WO2024206329A1

WO2024206329A1 - Nucleic acid molecules encoding bi-specific secreted engagers and uses thereof

Info

Publication number: WO2024206329A1
Application number: PCT/US2024/021507
Authority: WO
Inventors: Ailin Bai; Haley BRINJONES; Graham FARRINGTON; Joshua Frederick; Maija GARNAAS; Sushma GURUMURTHY; Ankita MISHRA; Maiko Obana; Rebecca RIDING; Zhen Zhang
Original assignee: ModernaTx Inc
Current assignee: ModernaTx Inc
Priority date: 2023-03-27
Filing date: 2024-03-26
Publication date: 2024-10-03
Anticipated expiration: 2025-09-27

Abstract

Described herein are bispecific NK cell engagers and nucleic acids encoding the same to treat various diseases, such as cancer (e.g., a B cell cancer).

Description

Attorney Docket No.: 45817-0158WO1 NUCLEIC ACID MOLECULES ENCODING BI-SPECIFIC SECRETED ENGAGERS AND USES THEREOF CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of priority of U.S. Provisional Application No.63/454,893 filed March 27, 2023, and U.S. Provisional Application No. 63/472,173 filed June 9, 2023, the contents of both of which are incorporated by reference in their entireties herein. SEQUENCE LISTING The instant application contains a Sequence Listing which has been submitted electronically in XML file format and is hereby incorporated by reference in its entirety. Said XML copy, created on March 26, 2024, is named 45817- 0158WO1_SL.xml and is 1,292,370 bytes in size. BACKGROUND Bispecific engagers are a type of bispecific binding molecule that can bind to two target molecules present on two cell types simultaneously to induce a particular function. Generally, a bispecific engager will bind one molecule on a cell or cells targeted for reduction or elimination (e.g., at least one molecule on a cell relating to a pathology, such as at least one tumor antigen) and a second molecule on an immune effector cell (e.g., a natural killer cell) to bring the effector cell into contact with or proximity to the cell targeted for reduction or elimination, such that killing of that cell can take place. Targeting a bispecific engager to a particular tissue or location (e.g., a tumor microenvironment), achieving an adequate half-life, and balancing efficacy with toxicity have all been challenging in the development of bispecific engager protein therapeutics. In addition, delivery of a bispecific engager which binds to a particular molecule on a tumor cell can result in selective expansion of tumor cells not bearing Attorney Docket No.: 45817-0158WO1 that molecule, and delivery of multiple protein therapeutics during a course of therapy is also problematic. Accordingly, there remains a need for therapeutic compositions and methods that address these current challenges. As set forth in more detail herein, this has been accomplished by the design of engager molecules that can be effectively delivered as nucleic acid molecules and expressed in vivo and that kill tumor cells. A multiplicity of such nucleic acid molecules encoding engager molecules which bind to different target molecules can also be delivered (either in the same or separate delivery vehicles) and expressed to maximize reduction or elimination of targeted cell populations. SUMMARY OF THE DISCLOSURE The present disclosure provides, among other things, bi-specific engager constructs that each specifically bind two different target molecules, which are efficiently expressed from nucleic acid molecules encoding them (e.g., mRNA), and therapeutic methods using these nucleic acid molecules to reduce or eliminate target cell populations. In one aspect, the present disclosure provides a composition comprising a nucleic acid encoding a bispecific engager, the nucleic acid comprising an mRNA polynucleotide comprising in order from the 5’ to 3’ end of the mRNA (i) an open reading frame encoding a VHH binding moiety that binds to a molecule expressed on the surface of a B cell, (ii) an open reading frame encoding an IgG4 PAA CH2 and CH3 domain, and (iii) an open reading frame encoding a VHH binding moiety that binds to a molecule expressed on the surface of an NK cell. In one instance, the open reading frame encoding an IgG4 PAA CH2 and CH3 domain encodes an amino acid sequence comprising a sequence that is at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to SEQ ID NO: 548. In some instances, an open reading frame encoding a GS (e.g., G4S (SEQ ID NO: 448)) linker follows (ii). The composition may further comprise one, two, or three other nucleic acids engaging a second, third, or fourth bispecific engager. These Attorney Docket No.: 45817-0158WO1 additional bispecific engagers each comprise an open reading frame encoding a VHH binding moiety that binds to a different molecule expressed on the surface of a B cell (e.g., CD38, BCMA, GPRC5D, and FcRH5). In one instance, these bispecific engagers comprise an open reading frame encoding a VHH binding moiety that binds to human CD16a. Compositions comprising the polypeptides encoded by the bispecific engager or bispecific engagers described above are also encompassed as part of the invention. In one aspect, the present disclosure provides a composition comprising a nucleic acid encoding a bispecific engager, the nucleic acid comprising an mRNA polynucleotide comprising in order from the 5’ to 3’ end of the mRNA (i) an open reading frame encoding a means for binding to a molecule expressed on the surface of a B cell (e.g., human FcRH5), (ii) an open reading frame encoding a hinge (e.g., IgG4 PAA CH2 and CH3 domain), and (iii) an open reading frame encoding a means for binding to a molecule expressed on the surface of an NK cell (e.g., human CD16a). In one instance, the open reading frame encoding an IgG4 PAA CH2 and CH3 domain encodes an amino acid sequence comprising a sequence that is at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to SEQ ID NO: 548. In some embodiments, the molecule expressed on the surface of a B cell is selected from CD38, BCMA, GPRC5D, and FcRH5. In some embodiments, the VHH binding moiety that binds to a molecule expressed on the surface of a B cell comprises (i) three complementarity-determining regions (CDRs) of any one VHH disclosed in Table 1 or Table A; (ii) three CDRs of any one VHH disclosed in Table 3 or Table B; (iii) three CDRs of any one VHH disclosed in Table 5 or Table C; or (iv) three CDRs of any one VHH disclosed in Table 7 or Table D. In some embodiments, the VHH binding moiety that binds to a molecule expressed on the surface of a B cell comprises (i) an amino acid sequence with at least 90%, at least 95%, or 100% identity to a heavy chain amino acid sequence disclosed Attorney Docket No.: 45817-0158WO1 in Table 2; (ii) an amino acid sequence with at least 90%, at least 95%, or 100% identity to a heavy chain amino acid sequence disclosed in Table 4; (iii) an amino acid sequence with at least 90%, at least 95%, or 100% identity to a heavy chain amino acid sequence disclosed in Table 6; or (iv) an amino acid sequence with at least 90%, at least 95%, or 100% identity to a heavy chain amino acid sequence disclosed in Table 8. In some embodiments, the molecule expressed on the surface of an NK cell is selected from B3GAT1 (CD57), CCR7 (CD197), CD16, CD16a, CD16b, CD2 CD226, CD244, CD27, CD3, CD300A, CD34, CD58, CD59, CD69, CSF2, CX3CR1, CXCR1 (CD128), CXCR3 (CD183), CXCR4, EOMES, GZMB, ICAM1 (CD54), IFNG, IL-15R, IL-1R, IL22, IL-2RB (CD122), IL-7R (CD127), ITGA1 (CD49a), ITGA2 (CD49b), ITGAL (CD11a), ITGAM (CD11b), ITGB2 (CD18), KIR, KIR2DL1, KIR2DL2, KIT (CD117), KLRB1C, KLRC1, KLRC2, KLRD1 (CD94), KLRF1, KLRG1, KLRK1, LILRB1, KLRA4, KLRA8, MICA/BNCAM1 (CD56), NK2D, NKP46 (NCR1, CD335), NCR2, NCR3 (CD337), PRF1, SELL (CD62L), SIGLEC7, SLAMF6, SPN, TBX21, and TNFa; or optionally, wherein the target molecule on the surface of a NK cell is selected from: CD16a, NKP46, NK2D, and MICA/B. In some embodiments, the molecule expressed on the surface of an NK cell is CD16a. In some instances, the VHH binding moiety that binds to CD16a comprises the three VHH CDRs of any one VHH set forth in SEQ ID NO: 1 or 5-24. In certain cases, the VHH binding moiety that binds to CD16a comprises the three VHH CDRs of any one CDR definition provided in Table I or Table II. In some embodiments, the VHH binding moiety that binds to a molecule expressed on the surface of an NK cell, wherein the molecule is CD16a, and wherein the VHH comprises: a CDR1 comprising the amino acid sequence GRTDSIYA (SEQ ID NO: 2), a CDR-2 comprising the amino acid sequence INSNTGRT (SEQ ID NO: 3), and a CDR-3 comprising the amino acid sequence AAGRGYGLLSISSNWYNY (SEQ ID NO: 4). Attorney Docket No.: 45817-0158WO1 In another instance, the VHH binding moiety that binds to CD16a comprises the three VHH CDRs according to any one CDR definition of Table I. In some embodiments, the VHH binding moiety that binds to a molecule expressed on the surface of an NK cell comprises an amino acid sequence with at least 90%, at least 95%, or 100% identity to any one of SEQ ID NOs: 1 or 5-24. In some embodiments, the bispecific engager comprises an amino acid sequence with at least 90%, at least 95%, or 100% identity to an amino acid sequence disclosed in Table 9. In some embodiments, the bispecific engager comprises an amino acid sequence comprising of SEQ ID NO: 433. In some embodiments, the bispecific engager comprises an amino acid sequence that is at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to an amino acid sequence of SEQ ID NO: 433, wherein the bispecific engager binds to both human FcRH5 and to human CD16a. In certain instances, the bispecific engager comprises an amino acid sequence that is at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to a sequence of SEQ ID NO: 392 linked directly or via a linker (e.g., glycine, glycine-serine linker) to an amino acid sequence that is at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to a sequence of SEQ ID NO:548 linked directly or via a linker (e.g., glycine, glycine- serine linker such as G4S (SEQ ID NO:448)) to an amino acid sequence that is at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to a sequence of SEQ ID NO:7. The bispecific engager binds to both human CD16a and human FcRH5. In some embodiments, the nucleic acid comprises a nucleotide sequence comprising any one of SEQ ID NO: 463 or encodes an amino acid sequence of SEQ ID NO: 464. In some instances, the nucleic acid comprises a nucleotide sequence of SEQ ID NO: 463 but lacking the nucleic acid sequence encoding the signal sequence. In some instances, the nucleic acid encodes an amino acid sequence of SEQ ID NO: 464 but lacking the signal sequence. Attorney Docket No.: 45817-0158WO1 In some embodiments, the composition may further comprise at least one additional nucleic acid encoding at least one additional bispecific engager, wherein the at least one additional bispecific engager is different from the bispecific engager of any one of the foregoing aspects or embodiments. In some embodiments, the at least one additional nucleic acid comprises an mRNA polynucleotide comprising in order from the 5’ to 3’ end of the mRNA (i) at least one additional open reading frame encoding at least one additional VHH binding moiety that binds to a molecule expressed on the surface of a B cell, (ii) at least one additional open reading frame encoding an IgG4 PAA CH2 and CH3 domain, and (iii) at least one additional open reading frame encoding a VHH binding moiety that binds to a molecule expressed on the surface of an NK cell. In some embodiments, the composition may further comprise a cytokine or at least one additional nucleic acid encoding a cytokine. In some embodiments, the cytokine is IL-15. In some instances, the cytokine is sushi-IL-15. sushi-IL-15 is composed of the NH2-terminal (amino acids 1–77, sushi+) domain of IL-15 receptor α coupled via a linker to IL-15 (see, e.g., Bouchaud et al., J Mol Biol 2008; 382:1–12; Huntington et al., J Exp Med 2009; 206:25–34; Bessard et al., Mol Cancer Ther (2009) 8 (9): 2736–2745). In some cases, the linker comprises or consists of the sequence: GGSGGGGSGGGSGGGGSLQ (SEQ ID NO:560). In some instances, the IL-15 is one described in U.S. Provisional Patent Application No.63/486,895, which is incorporated by reference in its entirety herein. In some embodiments, the composition further comprises a delivery vehicle. In some embodiments, the delivery vehicle comprises a lipid nanoparticle. In some embodiments, the lipid nanoparticle comprises: (a) an ionizable amino lipid of Formula (I): or its N-oxide, or a salt or isomer thereof, Attorney Docket No.: 45817-0158WO1 wherein R’^a is R’^branched; wherein R’^branched ;

wherein denotes a point of attachment;

wherein R^aβ, R^aγ, and R^aδ are each independently selected from the group consisting of H, C2-12 alkyl, and C2-12 alkenyl; R² and R³ are each independently selected from the group consisting of C_1-14 alkyl and C2-14 alkenyl; R⁴ is selected from the group consisting of -(CH2)nOH, wherein n is selected from the group , wherein

wherein R¹⁰ is N(R)2; each R is independently selected from the group consisting of C1-6 alkyl, C2-3 alkenyl, and H; wherein n2 is selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10; each R⁵ is independently selected from the group consisting of C1-3 alkyl, C2-3 alkenyl, and H; each R⁶ is independently selected from the group consisting of C1-3 alkyl, C2-3 alkenyl, and H; M and M’ are each independently selected from the group consisting of -C(O)O- and - OC(O)-; R’ is a C1-12 alkyl or C2-12 alkenyl; l is selected from the group consisting of 1, 2, 3, 4, and 5; and m is selected from the group consisting of 5, 6, 7, 8, 9, 10, 11, 12, and 13. In some embodiments, the ionizable amino lipid of Formula (I) comprises: R’^a is R’^branched; Attorney Docket No.: 45817-0158WO1 ; and

12 R^aδ are each H; R² and R³ are each C1-14 alkyl; ;

n2 is 2; each R⁵ is H; each R⁶ is H; M and M’ are each -C(O)O-; R’ is a C_1-12 alkyl; l is 5; and m is 7. In some embodiments, the ionizable amino lipid of Formula (I) comprises: R’a is R’branched; ;

R^aβ, R^aγ, and R^aδ are each H; R³ are each C1-14 alkyl; R⁴ is -(CH2)nOH; n is 2; each R5 is H; each R6 is H; M and M’ are each -C(O)O-; R’ is a C1-12 alkyl; Attorney Docket No.: 45817-0158WO1 l is 5; and m is 7. In some embodiments, the ionizable amino lipid of Formula (I) comprises: R’a is R’branched; ;

a R^aβ and R^aδ are each H; R^aγ is C_2-12 alkyl; R² and R³ are each C1-14 alkyl; R⁴ is -(CH₂)_nOH; n is 2; each R⁵ is H; each R⁶ is H; M and M’ are each -C(O)O-; R’ is a C1-12 alkyl; l is 5; and m is 7. In some embodiments, the lipid nanoparticle comprises an ionizable amino lipidselected from:

Attorney Docket No.: 45817-0158WO1 ,

or N-oxides, salts, or isomers thereof. In some embodiments, the lipid nanoparticle further comprises: a phospholipid, a structural lipid, and a PEG-lipid. In some embodiments, the lipid nanoparticle comprises: 40-50 mol% of an ionizable amino lipid, 30-45 mol% of a structural lipid, 5-15 mol% of a phospholipid, and 1-5 mol% of a PEG-lipid. In some embodiments, the lipid nanoparticle comprises: 45-50 mol% of the ionizable amino lipid, 35-40 mol% of the structural lipid, 8-12 mol% of the phospholipid, and 1.5-3.5 mol% of the PEG-lipid. In some embodiments, the phospholipid is selected from: 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), 1,2-dioleoyl-sn-glycero-3- phosphoethanolamine (DOPE), 1,2-dilinoleoyl-sn-glycero-3-phosphocholine (DLPC), 1,2-dimyristoyl-sn-gly cero-phosphocholine (DMPC), 1,2-dioleoyl-sn-glycero-3- phosphocholine (DOPC), l,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), 1,2- diundecanoyl-sn-glycero-phosphocholine (DUPC), 1-palmitoyl-2-oleoyl-sn-glycero- 3-phosphocholine (POPC), 1,2-di-O-octadecenyl-sn-glycero-3-phosphocholine (18:0 Diether PC), 1-oleoyl-2 cholesterylhemisuccinoyl-sn-glycero-3-phosphocholine (OChemsPC), 1-hexadecyl-sn-glycero-3-phosphocholine (C16 Lyso PC), 1,2- dilinolenoyl-sn-glycero-3-phosphocholine,1,2-diarachidonoyl-sn-glycero-3- Attorney Docket No.: 45817-0158WO1 phosphocholine, 1,2-didocosahexaenoyl-sn-glycero-3-phosphocholine, 1,2- diphytanoyl-sn-glycero-3-phosphoethanolamine (ME 16.0 PE), 1,2-distearoyl-sn- glycero-3-phosphoethanolamine, 1,2-dilinoleoyl-sn-glycero-3-phosphoethanolamine, 1,2-dilinolenoyl-sn-glycero-3-phosphoethanolamine, 1,2-diarachidonoyl-sn-glycero- 3-phosphoethanolamine, 1,2-didocosahexaenoyl-sn-glycero-3-phosphoethanolamine, 1,2-dioleoyl-sn-glycero-3-phospho-rac-(1-glycerol) sodium salt (DOPG), sphingomyelin, and mixtures thereof. In some embodiments, the phospholipid is DSPC. In some embodiments, the structural lipid is selected from: cholesterol, fecosterol, sitosterol, ergosterol, campesterol, stigmasterol, brassicasterol, tomatidine, tomatine, ursolic acid, alpha-tocopherol, hopanoids, phytosterols, steroids, derivatives thereof, and mixtures thereof. In some embodiments, the structural lipid is cholesterol or a derivative thereof. In some embodiments, the PEG-lipid is selected from: 1,2-dimyristoyl-sn-glycerol methoxypolyethylene glycol (PEG-DMG), 1,2-distearoyl- sn-glycero-3-phosphoethanolamine-N-[amino(polyethylene glycol)] (PEG-DSPE), PEG-disteryl glycerol (PEG-DSG), PEG-dipalmetoleyl, PEG-dioleyl, PEG-distearyl, PEG-diacylglycamide (PEG-DAG), PEG-dipalmitoyl phosphatidylethanolamine (PEG-DPPE), PEG-l,2-dimyristyloxlpropyl-3-amine (PEG-c-DMA), any mixtures thereof. In some embodiments, the PEG-lipid comprises a structure of: .

40-50 mol% of an ionizable amino lipid comprising a structure of: , Attorney Docket No.: 45817-0158WO1 30-45 mol% of cholesterol, 5-15 mol% of DSPC, and 1-5 mol% of a PEG-lipid. In some cases, the lipid nanoparticle comprises: about 47.5 mol % of ionizable amino lipid; about 39 mol % of cholesterol; about 10.5 mol % of DSPC; and about 3 mol % of PEG-lipid. In certain cases, the lipid nanoparticle comprises: 47.5 mol % of ionizable amino lipid; 39 mol % of cholesterol; 10.5 mol % of DSPC; and 3 mol % of PEG-lipid. In some cases, the ionizable amino acid lipid is heptadecan-9-yl 8-((2- hydroxyethyl)(8-(nonyloxy)-8-oxooctyl)amino)octanoate which has the formula (Compound 2) or a salt thereof. In

9-yl 8-((2- hydroxyethyl)(6-oxo-6-(undecyloxy)hexyl)amino)octanoate or a salt thereof. In some cases, the PEG lipid is 134-hydroxy- 3,6,9,12,15,18,21,24,27,30,33,36,39,42,45,48,51,54,57,60,63,66,69,72,75,78,81,84,87 ,90,93,96,99,102,105,108,111,114,117,120,123,126,129,132- tetratetracontaoxatetratriacontahectyl stearate which has the formula (Compound I) thereof.

In another aspect, the present disclosure provides a method of expressing a bispecific engager in a cell, comprising contacting a cell with a composition disclosed herein (e.g., a composition according to any one of the foregoing aspects or embodiments). Attorney Docket No.: 45817-0158WO1 another aspect, the present disclosure provides a method of reducing or eliminating a B cell population associated with a disease, comprising administering to a subject with a disease a composition disclosed herein (e.g., a composition according to any one of the foregoing aspects or embodiments). In some embodiments, the disease is a B cell cancer. In some embodiments, the disease is multiple myeloma. In some embodiments, the methods may further comprise administering a second composition comprising a second nucleic acid encoding a different bispecific engager, the second nucleic acid comprising a second mRNA comprising in order from the 5’ to 3’ end of the second mRNA (i) a second open reading frame encoding a second VHH binding moiety that binds to a molecule expressed on the surface of a B cell, (ii) a second open reading frame encoding an IgG4 PAA CH2 and CH3 domain, and (iii) a second open reading frame encoding a VHH binding moiety that binds to a molecule expressed on the surface of an NK cell. In some embodiments, the methods may further comprise administering a third composition comprising a third nucleic acid encoding a further different bispecific engager, the third nucleic acid comprising a third mRNA comprising in order from the 5’ to 3’ end of the third mRNA (i) a third open reading frame encoding a third VHH binding moiety that binds to a molecule expressed on the surface of a B cell, (ii) a third open reading frame encoding an IgG4 PAA CH2 and CH3 domain, and (iii) a third open reading frame encoding a VHH binding moiety that binds to a molecule expressed on the surface of an NK cell. In some embodiments, each of the composition, the second composition, and/or the third composition are present in the same delivery vehicle. In some embodiments, each of the composition, the second composition, and/or the third composition are present in different delivery vehicles. In some embodiments, each of the composition, the second composition, and/or the third composition are administered concurrently. In some embodiments, each of the composition, the second composition, and/or the third composition are administered sequentially. Attorney Docket No.: 45817-0158WO1 In some embodiments, the methods may further comprise administering to the subject a cytokine or an mRNA encoding the cytokine. In some embodiments, the cytokine is IL-15. In some instances, the cytokine is sushi-IL-15. sushi-IL-15 is composed of the NH2-terminal (amino acids 1–77, sushi+) domain of IL-15 receptor α coupled via a linker to IL-15. In some embodiments, the cytokine or mRNA encoding the cytokine and the composition are present in the same delivery vehicle. In some embodiments, the cytokine or mRNA encoding the cytokine and the composition are present in different delivery vehicles. Also provided are methods of treating relapsed or refractory multiple myeloma in a subject in need thereof, comprising administering to the subject a composition as described herein, comprising a nucleic acid encoding a bispecific engager as described herein, the nucleic acid comprising an mRNA polynucleotide comprising in order from the 5’ to 3’ end of the mRNA (i) an open reading frame encoding a VHH binding moiety that binds to a molecule expressed on the surface of a B cell, wherein the molecule expressed on the surface of a B cell is FcRH5 (ii) an open reading frame encoding an IgG4 PAA CH2 and CH3 domain, and (iii) an open reading frame encoding a VHH binding moiety that binds to a molecule expressed on the surface of an NK cell, wherein the molecule expressed on the surface of an NK cell is CD16a, wherein the composition further comprises a lipid nanoparticle delivery vehicle encapsulating the mRNA. In some aspects, prior to treatment, the subject has had prior exposure to one or more of all of a proteasome inhibitor, an immunomodulatory drug (IMiD), and an anti-cluster of differentiation (CD38) monoclonal antibody. In some aspects, prior to treatment, the subject has received at least three prior lines of prior therapy and/or is triple-class refractory. In some aspects, the subject is intolerant of one or more or all of a proteasome inhibitor, IMiD, or an anti-CD38 mAb. In any aspects, the mRNA may comprise a nucleotide sequence comprising any one of SEQ ID NO: 463 or encoding an amino acid sequence of SEQ ID NO: 464. In some instances, the mRNA comprises a nucleotide sequence of SEQ ID NO: 463 but without the signal sequence of SEQ ID NO:463. In some cases, that signal sequence is replaced by a different signal sequence. In some instances, the mRNA comprises a nucleotide sequence encoding an amino acid sequence of SEQ ID NO:433. Attorney Docket No.: 45817-0158WO1 In one aspect, the disclosure features a bispecific NK engager comprising in order from the N- to the C-terminal end of the polypeptide (i) a VHH binding moiety that binds to a molecule expressed on the surface of a B cell, (ii) a hinge region (e.g., a human IgG4 PAA hinge +CH2+ CH3 domain), and (iii) a VHH binding moiety that binds to a molecule expressed on the surface of an NK cell. In some instances, a linker links parts (ii) and (iii). The linker may be a Gly-Ser linker such as the linker set forth in SEQ ID NO: 448. In some cases, the mature bispecific NK engager polypeptide includes a signal peptide upstream of (i). In some cases, the signal peptide comprises the sequence of SEQ ID NO: 447. In some instances, the VHH binding moiety that binds to a molecule expressed on the surface of a B cell is a VHH that specifically binds one of CD38, BCMA, GPRC5D, or FcRH5. In one instance, the VHH binding moiety that binds to a molecule expressed on the surface of a B cell is a VHH that specifically binds BCMA. In another instance, the VHH binding moiety that binds to a molecule expressed on the surface of a B cell is a VHH that specifically binds FcRH5. In some cases, the VHH specifically binds CD38 and comprises the VHH-CDR1, VHH-CDR2, and VHH-CDR3 of any VHH set forth in SEQ ID NOs.: 238-252. In some cases, the VHH specifically binds CD38 and comprises the VHH-CDR1, VHH-CDR2, and VHH-CDR3 of any VHH set forth in SEQ ID NOs.: 238-252 and comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to an amino acid sequence of SEQ ID NOs.: 238-252. In some cases, the VHH specifically binds BCMA and comprises the VHH-CDR1, VHH- CDR2, and VHH-CDR3 of any VHH set forth in SEQ ID NOs.: 253-332 or 624-644. In some cases, the VHH specifically binds BCMA and comprises the VHH-CDR1, VHH-CDR2, and VHH-CDR3 of any VHH set forth in SEQ ID NOs.: 253-332 or 624-644 and comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to an amino acid sequence of SEQ ID NOs.: 253-332 or 624-644. In some cases, the VHH specifically binds GPRC5D and comprises the VHH-CDR1, VHH-CDR2, and VHH-CDR3 of any VHH set forth Attorney Docket No.: 45817-0158WO1 in SEQ ID NOs.: 333-376. In some cases, the VHH specifically binds GPRC5D and comprises the VHH-CDR1, VHH-CDR2, and VHH-CDR3 of any VHH set forth in SEQ ID NOs.: 333-376 and comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to an amino acid sequence of SEQ ID NOs.: 333-376. In some cases, the VHH specifically binds FcRH5 and comprises the VHH-CDR1, VHH-CDR2, and VHH-CDR3 of any VHH set forth in SEQ ID NOs.: 377-393 or 705-727. In certain cases, the VHH-CDR1, VHH-CDR2, and VHH-CDR3 comprise the sequences of SEQ ID NOs.: 213, 223, and 233, respectively. In some cases, the VHH specifically binds FcRH5 and comprises the VHH-CDR1, VHH-CDR2, and VHH-CDR3 of any VHH set forth in SEQ ID NOs.: 377-393 or 705-727 and comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to an amino acid sequence of SEQ ID NOs.: 377-393 or 705-727. In certain instances, the VHH binding moiety that binds to a molecule expressed on the surface of an NK cell, is a VHH that specifically binds CD16. In one instance, the VHH binds human CD16a. In another instance, the VHH binds cynomolgus CD16a. In another case, the VHH binds both human and cynomolgus CD16a. In some cases, the VHH specifically binds CD16 and comprises the VHH- CDR1, VHH-CDR2, and VHH-CDR3 of any VHH set forth in SEQ ID NOs.: 1 or 5- 24. In certain cases, the VHH-CDR1, VHH-CDR2, and VHH-CDR3 comprise the amino acid sequences of SEQ ID NOs.: 2, 3, and 4, respectively. In some cases, the VHH specifically binds CD16 and comprises the VHH-CDR1, VHH-CDR2, and VHH-CDR3 of any VHH set forth in SEQ ID NOs.: 1 or 5-24 and comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to an amino acid sequence of SEQ ID NOs.: 1 or 5-24. In some instances, the bispecific NK engager polypeptide binds both human FcRH5 and human CD16 (e.g., human CD16a) and comprises an amino sequence that Attorney Docket No.: 45817-0158WO1 is at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the sequence of SEQ ID NO:433. In another aspect, the disclosure provides nucleic acid encoding the bispecific NK engager polypeptide described herein. Also provided is a vector or vectors comprising the nucleic acid or nucleic acids encoding the bispecific NK engager polypeptide described herein. Also featured are host cells comprising the nucleic acid or nucleic acids or the vector and vectors described above. The disclosure also relates to methods of making the bispecific NK engager polypeptide described herein. The method involves culturing the host cell under conditions that facilitate the expression of the polypeptide and isolating the polypeptide. In some cases, the polypeptide can be formulated as a sterile pharmaceutical composition. In another aspect the disclosure features a polynucleotide comprising an mRNA comprising; (i) a 5’UTR; (ii) an ORF encoding any of the bispecific NK engager polypeptides described herein; (iii) a stop codon; and (iv) a 3’UTR. In some cases, the polynucleotide includes a 5’ terminal cap. In one case, the 5’ terminal cap is a m7GpppGmAG cap. In some cases, the polynucleotide comprises a poly A tail. In some cases, the poly A tail is 100 nt in length (SEQ ID NO: 730). In some cases, the mRNA comprises at least one chemically modified nucleobase, sugar, backbone, or any combination thereof. In some cases, the ORF comprises at least one modified uridine. In some cases, the modified uridine is 1-methylpseudourdine. In some cases, all uracils of the polynucleotide are N1-methylypseudouracils. In another aspect, the disclosure features a composition comprising a first polynucleotide comprising a first mRNA comprising; (i) a 5’UTR; (ii) an ORF encoding a first bispecific NK engager polypeptide; (iii) a stop codon; and (iv) a 3’UTR; and a second mRNA comprising; (i) a 5’UTR; (ii) an ORF encoding a second bispecific NK engager polypeptide; (iii) a stop codon; and (iv) a 3’UTR. In some instances, the composition comprises a third polynucleotide comprising a third mRNA comprising; (i) a 5’UTR; (ii) an ORF encoding a third bispecific NK engager polypeptide; (iii) a stop codon; and (iv) a 3’UTR. In certain instances, the Attorney Docket No.: 45817-0158WO1 composition comprises a fourth polynucleotide comprising a third mRNA comprising; (i) a 5’UTR; (ii) an ORF encoding a fourth bispecific NK engager polypeptide; (iii) a stop codon; and (iv) a 3’UTR. In some cases, the first bispecific NK engager polypeptide binds FcRH5 and CD16a. In some cases, the second bispecific NK engager polypeptide binds BCMA and CD16a. In some cases, the third bispecific NK engager polypeptide binds GPRC5D and CD16a. In some cases, the fourth bispecific NK engager polypeptide binds CD38 and CD16a. In some cases, the polynucleotide includes a 5’ terminal cap. In one case, the 5’ terminal cap is a m7GpppGmAG cap. In some cases, the polynucleotide comprises a poly A tail. In some cases, the poly A tail is 100 nt in length (SEQ ID NO: 730). In some cases, the mRNA comprises at least one chemically modified nucleobase, sugar, backbone, or any combination thereof. In some cases, the ORF or ORFs comprise(s) at least one modified uridine. In some cases, the modified uridine is 1-methylpseudourdine. In some cases, all uracils of the polynucleotide are N1-methylypseudouracils. In another aspect, the disclosure provides a pharmaceutical composition comprising the bispecific NK engager polypeptide or the polynucleotide(s) and a pharmaceutically acceptable carrier. In another aspect, the disclosure provides a pharmaceutical composition comprising the polynucleotide disclosed herein and a delivery agent. In some cases, the delivery agent is a lipid nanoparticle. In some cases, the lipid nanoparticle has mean particle size of from 80 nm to 160 nm. In certain cases, the lipid nanoparticle has a polydispersity index (PDI) of from 0.02 to 0.2 and/or a lipid:nucleic acid ratio of from 10 to 20. In some cases, the lipid nanoparticle comprises a neutral lipid, an ionizable amino lipid, a polyethyleneglycol (PEG) lipid, and/or a sterol. In certain cases, the neutral lipid is 1,2-distearoyl-sn-glycero-3-phosphocholine. In some cases, the PEG lipid is PEG 2000 dimyristoyl glycerol or OL56. In certain cases, the sterol is cholesterol. In some cases, multiple polynucleotides are encapsulated in the same lipid nanoparticle. Attorney Docket No.: 45817-0158WO1 Also provided herein are methods of treating a human subject in need thereof with a therapeutically effective amount of the bispecific NK engager polypeptide or the polynucleotide or pharmaceutical composition. In some cases, the human subject has multiple myeloma (MM). In some cases, the MM is relapsed MM. In some cases, the MM is refractory MM. In certain cases, the MM is relapsed refractory MM. In certain cases, the MM is double refractory MM. Also featured herein are methods of depleting or eliminating a B cell population associated with a disease in a human subject in need thereof, the method comprising administering to the human subject a therapeutically effective amount of the bispecific NK engager polypeptide or the polynucleotide or pharmaceutical composition. In some cases, the human subject has a B cell cancer. The disclosure also features a combination comprising a bispecific NK engager polypeptide, a polynucleotide or a pharmaceutical composition described herein and IL-15. In some cases, the IL-15 is sushi-IL-15 (see e.g., U.S. Prov. Appl. No.63/486,895). The fusion protein sushi-IL-15 is composed of the NH2-terminal (amino acids 1–77, sushi+) domain of IL-15 receptor α coupled via a linker (e.g., GGSGGGGSGGGSGGGGSLQ (SEQ ID NO:560) to IL-15. In certain cases, the sushi-IL-15 is linked to a serum albumin (e.g., HSA) or a VHH that specifically binds serum albumin (e.g., HSA). In certain cases, the serum albumin is human serum albumin. The disclosure also provides methods of using the combination described above. In some cases, provided herein are methods of treating a human subject in need thereof with a therapeutically effective amount of the combination. In some cases, the human subject has multiple myeloma (MM). In some cases, the MM is relapsed MM. In some cases, the MM is refractory MM. In certain cases the MM is relapsed refractory MM. In certain cases, the MM is double refractory MM. In other cases, this disclosure features methods of depleting or eliminating a B cell population associated with a disease in a human subject in need thereof. The method comprises Attorney Docket No.: 45817-0158WO1 administering to the human subject a therapeutically effective amount of the combination. In some cases, the human subject has a B cell cancer. In another aspect, the disclosure features a bispecific engager polypeptide comprising in order from the N to C terminus of the polypeptide (i) a VHH binding moiety that binds to a molecule expressed on the surface of a B cell, (ii) a hinge region (e.g., an IgG4 PAA CH2 and CH3 domain), and (iii) a VHH binding moiety that binds to a molecule expressed on the surface of an NK cell. In one instance, the IgG4 PAA CH2 and CH3 domain comprises an amino acid sequence that is at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to SEQ ID NO: 548. In some instances, a glycine serine linker (e.g., G4S (SEQ ID NO: 448)) follows (ii). In some instances, the disclosure provides a composition comprising two or more bispecific engager polypeptides. These additional bispecific engagers each comprise a VHH binding moiety that binds to a different molecule expressed on the surface of a B cell (e.g., CD38, BCMA, GPRC5D, and FcRH5). In one instance, these bispecific engagers each comprise a VHH binding moiety that binds to human CD16a. In another aspect, the disclosure features a bispecific engager polypeptide comprising in order from the N to C terminus of the polypeptide (i) a means for binding to a molecule expressed on the surface of a B cell (e.g., human FcRH5), (ii) a hinge region (e.g., an IgG4 PAA CH2 and CH3 domain), and (iii) a means for binding to a molecule expressed on the surface of an NK cell (e.g., human CD16a). In one instance, the hinge comprises a IgG4 PAA CH2 and CH3 domain which comprises an amino acid sequence that is at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to SEQ ID NO: 548. In some instances, a glycine serine linker (e.g., G4S (SEQ ID NO: 448)) follows (ii). In some instances, the disclosure provides a composition comprising two or more bispecific engager polypeptides. These additional bispecific engagers each comprise a VHH binding moiety that binds to a different molecule expressed on the surface of a B cell (e.g., CD38, BCMA, GPRC5D, and FcRH5). In one instance, these bispecific engagers each comprise a VHH binding moiety that binds to human CD16a. Attorney Docket No.: 45817-0158WO1 In some embodiments, the molecule expressed on the surface of a B cell is selected from CD38, BCMA, GPRC5D, and FcRH5. In some embodiments, the VHH binding moiety that binds to a molecule expressed on the surface of a B cell comprises (i) three complementarity-determining regions (CDRs) of any one VHH disclosed in Table 1 or Table A; (ii) three CDRs of any one VHH disclosed in Table 3 or Table B; (iii) three CDRs of any one VHH disclosed in Table 5 or Table C; or (iv) three CDRs of any one VHH disclosed in Table 7 or Table D. In some embodiments, the VHH binding moiety that binds to a molecule expressed on the surface of a B cell comprises (i) an amino acid sequence with at least 90%, at least 95%, or 100% identity to a heavy chain amino acid sequence disclosed in Table 2; (ii) an amino acid sequence with at least 90%, at least 95%, or 100% identity to a heavy chain amino acid sequence disclosed in Table 4; (iii) an amino acid sequence with at least 90%, at least 95%, or 100% identity to a heavy chain amino acid sequence disclosed in Table 6; or (iv) an amino acid sequence with at least 90%, at least 95%, or 100% identity to a heavy chain amino acid sequence disclosed in Table 8. In some embodiments, the molecule expressed on the surface of an NK cell is selected from B3GAT1 (CD57), CCR7 (CD197), CD16, CD16a, CD16b, CD2 CD226, CD244, CD27, CD3, CD300A, CD34, CD58, CD59, CD69, CSF2, CX3CR1, CXCR1 (CD128), CXCR3 (CD183), CXCR4, EOMES, GZMB, ICAM1 (CD54), IFNG, IL-15R, IL-1R, IL22, IL-2RB (CD122), IL-7R (CD127), ITGA1 (CD49a), ITGA2 (CD49b), ITGAL (CD11a), ITGAM (CD11b), ITGB2 (CD18), KIR, KIR2DL1, KIR2DL2, KIT (CD117), KLRB1C, KLRC1, KLRC2, KLRD1 (CD94), KLRF1, KLRG1, KLRK1, LILRB1, KLRA4, KLRA8, MICA/BNCAM1 (CD56), NK2D, NKP46 (NCR1, CD335), NCR2, NCR3 (CD337), PRF1, SELL (CD62L), SIGLEC7, SLAMF6, SPN, TBX21, and TNFa; or optionally, wherein the target molecule on the surface of a NK cell is selected from: CD16a, NKP46, NK2D, and MICA/B. Attorney Docket No.: 45817-0158WO1 In some embodiments, the molecule expressed on the surface of an NK cell is CD16a. In some instances, the VHH binding moiety that binds to CD16a comprises the three VHH CDRs of any one VHH set forth in SEQ ID NO: 1 or 5-24. In one aspect, the disclosure features the bispecific engager polypeptide comprises an amino acid sequence that is at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the sequence of SEQ ID NO:433. Also encompassed are mRNAs encoding such bispecific engager polypeptides. These mRNAs may include one or more or all of a 5’ terminal cap, a 5’UTR, a 3’UTR, and a poly A region described herein. In another aspect, the disclosure provides a pharmaceutical composition comprising the bispecific NK engager polypeptide described herein and a pharmaceutically acceptable carrier. The disclosure also features a combination comprising a bispecific NK engager polypeptide and the cytokine IL-15. In some cases, the IL-15 is sushi-IL-15 (see e.g., U.S. Prov. Appl. No.63/486,895). The fusion protein sushi-IL-15 is composed of the NH₂-terminal (amino acids 1–77, sushi+) domain of IL-15 receptor α coupled via a linker (e.g., GGSGGGGSGGGSGGGGSLQ (SEQ ID NO:560) to IL- 15. In certain cases, the sushi-IL-15 is linked to a serum albumin (e.g., HSA) or a VHH that specifically binds serum albumin (e.g., HSA). In certain cases, the serum albumin is human serum albumin. The disclosure also provides methods of using the polypeptide or combinations described above. In some cases, provided herein are methods of treating a human subject in need thereof with a therapeutically effective amount of the polypeptide or combination. In some cases, the human subject has multiple myeloma (MM). In some cases, the MM is relapsed MM. In some cases, the MM is refractory MM. In certain cases the MM is relapsed refractory MM. In certain cases, the MM is double refractory MM. In other cases, this disclosure features methods of depleting or eliminating a B cell population associated with a disease in a human subject in need Attorney Docket No.: 45817-0158WO1 thereof. The method comprises administering to the human subject a therapeutically effective amount of the combination. In some cases, the human subject has a B cell cancer. In one aspect, the disclosure features a lipid nanoparticle comprising a population of mRNAs encoding one, two, three, or four different bispecific engagers. In one instance, the lipid nanoparticle comprises a population of mRNAs that encode at one bispecific engager. In another instance, the lipid nanoparticle comprises a population of mRNAs that encode two bispecific engagers. In a different instance, the lipid nanoparticle comprises a population of mRNAs that encode three bispecific engagers. In a further instance, the lipid nanoparticle comprises a population of mRNAs that encode four bispecific engagers. These bispecific engagers comprise a VHH that binds to one or more molecules on the surface of a B cell (e.g., CD38, BCMA, GPRC5D, FcRH5) linked to a VHH that binds to a molecule on a NK cell (e.g., CD16). In some instances, any of the VHHs described herein, or variants thereof (e.g., sequences with at least 80% identity, at least 85% identity, at least 90% identity, or at least 95% identity to a VHH described herein but having all three VHH CDRs unaltered), can be used. In some cases the linker comprises an amino acid sequence that is at least 85%, at least 90%, at least 95%, or 100% identical to the sequence of SEQ ID NO:548. In some cases, the linker further comprises a sequence of SEQ ID NO:448. In one instance, a bispecific engager comprises an amino acid sequence that is at least 85%, at least 90%, at least 95%, or 100% identical to the sequence of SEQ ID NO:392 linked to an amino acid sequence that is at least 85%, at least 90%, at least 95%, or 100% identical to the sequence of SEQ ID NO:7, wherein the bispecific engager binds to both FCRH5 and CD16a. In a particular instance, bispecific engager comprises an amino acid sequence that is at least 85%, at least 90%, at least 95%, or 100% identical to the sequence of BE-40 (SEQ ID NO:433), wherein the bispecific engager binds to both FCRH5 and CD16a. In certain instances, the lipid nanoparticle further comprises an mRNA encoding a cytokine (e.g., human IL-15). In some cases, the mRNA encodes sushi- IL-15. In some cases, the mRNA encodes a polypeptide that is at least 85%, at least Attorney Docket No.: 45817-0158WO1 90%, at least 95%, or 100% identical to the sequence of SEQ ID NO:729. In other instances, a separate lipid nanoparticle is used to administer the cytokine (e.g., sushi- IL-15). In some cases, the lipid nanoparticle comprises an ionizable amino lipid, a PEG-lipid, a structural lipid, and a phospholipid. In certain cases the lipid nanoparticle comprises: about 47.5 mol % of ionizable amino lipid; about 39 mol % of cholesterol; about 10.5 mol % of DSPC; and about 3 mol % of PEG-lipid. In certain cases, the lipid nanoparticle comprises: 47.5 mol % of ionizable amino lipid; 39 mol % of cholesterol; 10.5 mol % of DSPC; and 3 mol % of PEG-lipid. In some cases, the ionizable amino acid lipid is heptadecan-9-yl 8-((2-hydroxyethyl)(8- (nonyloxy)-8-oxooctyl)amino)octanoate which has the formula (Compound 2), or a salt thereof. In

heptadecan-9-yl 8-((2- hydroxyethyl)(6-oxo-6-(undecyloxy)hexyl)amino)octanoate or a salt thereof. In some cases, the PEG lipid is 134-hydroxy- 3,6,9,12,15,18,21,24,27,30,33,36,39,42,45,48,51,54,57,60,63,66,69,72,75,78,81,84,87 ,90,93,96,99,102,105,108,111,114,117,120,123,126,129,132- tetratetracontaoxatetratriacontahectyl stearate which has the formula (Compound I) thereof.

The disclosure also provides methods of using the LNPs described above. In some cases, provided herein are methods of treating a human subject in need thereof with a therapeutically effective amount of the LNP or LNPs described above. In some cases, the human subject has multiple myeloma (MM). In some cases, the MM is Attorney Docket No.: 45817-0158WO1 relapsed MM. In some cases, the MM is refractory MM. In certain cases the MM is relapsed refractory MM. In certain cases, the MM is double refractory MM. In other cases, this disclosure features methods of depleting or eliminating a B cell population associated with a disease in a human subject in need thereof. The method comprises administering to the human subject a therapeutically effective amount of the LNP or LNPs. In some cases, the human subject has a B cell cancer. In some cases, administration is performed intravenously. In other cases, administration is performed subcutaneously. In certain cases, administration is performed intramuscularly. The following drawings and detailed description are examples and explanatory, but are not intended to be limiting. BRIEF DESCRIPTION OF THE DRAWINGS FIG.1 shows examples of different bispecific engager formats. On the left side of the figure the mRNA construct is shown and the proteins encoded are shown on the right. “Target 1 and 2” represent VHH moieties that bind to molecules on, for example, B cells associated with disease and “Effector” represents VHH moieties that bind to molecules on, for example, NK cells. Panel 1a represents the delivery of one mRNA and the protein dimer that is expressed. Panels 1b and 1c represent delivery of multiple, different mRNAs and the various protein dimers that are formed. The CH2- CH3 can be IgG1, IgG2, IgG4PAA, IgG1FEA, with optional linker sequences (shown as lines here) between the target binding moiety and CH2 domain and/or between the CH3 domain and the effector cell binding moiety. For example, the linker could be a hinge region from human IgG1, human IgG2, or human IgG4PAA. In one case the linker comprises or consists of the sequence of SEQ ID NO:548. FIG.2 shows FACS profiles of cell lines used in cytotoxicity assays demonstrating the expression levels of BCMA and FCRH5 on each cell lines. FIG.3A, 3B, and 3C show result from a cytotoxicity assay. Molm13 FCRH5 hi(BCMA-, FCRH5++; Fig.3A); RPMI 8226 (BCMA+, FCRH5-; Fig.3B); RPMI 8226-FCRH (BCMA+, FCRH5++; Fig.3C) were used to compare the potency of BE- Attorney Docket No.: 45817-0158WO1 40, BE-102, and BE-103 generated by mRNA transfection of Hela cells on each of the target cell lines. In each assay human NK cells (E) labeled with Calcein AM was mixed with target cells at an E:T ratio of 7.5:1. The percent cytotoxicity was determined based on the EC50 relationship between the fraction of cells killed versus dilution of the stock bispecific antibody. FIG.4 shows a comparison of expected depletion of target cells for two possible treatments, the outcome when a single therapeutic bispecific engager molecule is used versus a treatment with at least two molecules. FIG.5 shows anti-FCRH5 NK cell engager (NKE) formats evaluated for selection as a therapeutic. Both N and C terminal engager formats were evaluated. FIG.6 shows molecule characterization of bispecific molecules showing biophysical behavior in non-reducing (A) and reducing (B) SDS-PAGE. Affinity values are in in Table 18. FIG 7A and 7B show analytical gel filtration chromatography with anti-CD16 antibody in the N-terminal position following protein A purification. FIG 8A and 8B show analytical gel filtration with anti-CD16 antibody in the N-terminal position following preparative gel filtration. FIG.9 shows binding on FCRH5 transiently expressed on Expi293 cells. NK cell engager (NKE) constructed with parental anti-FCRH5 VHHs bind to human FCRH5 expressed by transiently transfected cells. FIG.10 shows evaluation of FACS binding of engager constructs with anti- CD16 in the N-terminal position and anti-TAA in the C-terminal position to full length FCRH5 transiently expressed on Expi293 cells. FIG.11 shows validation of FCRH5 on a lymphoma cell line SUDHL6 cells. FIG.12 shows cytotoxicity assays with anti-FCRH5 VHH antibodies oriented in the C-terminus of the engager format on RPMI. Attorney Docket No.: 45817-0158WO1 FIG.13A shows pharmacokinetic comparison of VHH antibodies C- and N- terminally fused to either two different FCs, IgG4PAA or IgG1FEA to evaluate expression and PK in vivo. FIG.13B shows that engager expression was detected (50-200 ng/ml) 120-168 hours after a single 0.25 mg/kg dose of RNA. FIG.14A shows analytical chromatography of humanized anti-FCRH5 bispecific engagers CD16-VHH4-IgG4PAA with c-terminally fused humanized forms of -FcRH5-VHH13, -FcRH5-VHH12, –FcRH5-VHH11 expressed in 293 cells purified on protein A and gel filtration. Analytical gel filtration of molecules post- Superdex 200 Polishing. FIG.14B shows the results for c-terminally fused –FcRH- VHH16, FcRH5-VHH17. FIG.15A shows analytical SEC comparison of IMAC purified [FcRH- VHH16]-IgG4PAA-[CD16-VHH4]-6xhis (6Xhis tag (SEQ ID NO: 731) for purification without pH 3 elution), and captured in totality from the supernatant, with 460best-IgG4-having (no his tag), purified with protein A and gel filtration (pH 3 elution from protein A). This shows the molecule has no aggregation upon expression. FIG.15B shows gel-filtration light scattering (GFLS) of BE-40 bispecific engager. Panel (a) shows the UV absorbance for the [FcRH-VHH16]-IgG4PAA- [CD16-VHH4]. FIG.16 shows cytotoxicity assays with humanized anti-FCRH5 VHH antibodies oriented in the C-terminus of the engager format using SUDHL6 cell line. FIG.17 shows N-terminal anti-FCRH5 NKEs demonstrate improved binding affinity. FIG.18 shows in a comparison of bidentate Fc engagers did not outperform N-terminal format of FcRH5-VHH11 engager in the non-humanized or humanized form. FIG.19 compares NK cell specific killing cells expressing FCRH5 (RPMI FCRH5 Hi) of C and N-terminal bispecific constructs using two different FcRH5 Attorney Docket No.: 45817-0158WO1 VHH (FcRH5-VHH11 and FcRH5-VHH15) in combination with CD16-VHH4. The results show the N-terminal FCR-VHH antibodies have improved killing. FIG.20 shows the serum concentrations of BE-40 ( FCR460h1-IgG4 PAA- 12C11h3) protein following a single IC infusion of an LNP-encapsulated mRNA encoding BE-40 engager protein. FIG.21 shows the percent of B cell subsets of CD20+ B cells in pre-dose cynomolgus monkeys. FIG.22 shows FcRH5 expression in B cell subpopulations in pre-dose cynomolgus monkeys. FIG.23 shows that an LNP-encapsulated mRNA encoding BE-40 engager protein induces specific depletion of CD21-CD27+ memory B cells at all dose levels. FIG.24 shows the percentage of memory B cell depletion after a single IV administration of an LNP-encapsulated mRNA encoding BE-40 engager protein. FIG.25 depicts the cytotoxicity against Molp-2 Target Cells in the presence of anti-FCRH5 NKE and HSA-sushi-IL-15 (HSA-sIL-15). FIG.26 shows the cytotoxicity against Molm13-LG-FCRH5 Target Cells in the presence of anti-FCRH5 NKE and HSA-sIL-15. DETAILED DESCRIPTION The constructs, compositions, and methods of the disclosure feature bispecific engager molecules for use in therapy, e.g., targeting cells for reduction or elimination in vivo. Specifically, the bispecific engager molecules of the disclosure specifically bind two different targets, a molecule on a B cell associated with disease (e.g., on a tumor cell or associated with an autoimmune disease) and an NK cell. These bispecific engagers bring the B cell targeted for destruction and the NK cell into proximity such that B cells expressing the target molecule in the population are reduced or eliminated. The bispecific engager molecules of the disclosure can be Attorney Docket No.: 45817-0158WO1 administered as mRNA molecules encoding the bispecific engager such that the bispecific engager molecules are made in vivo in a subject. Multiple mRNAs, each encoding a bispecific engager, but which comprise different binding moieties, can be administered in the same course of therapy, e.g., as shown in Fig.1. By so doing, the fraction of target cells depleted in a heterogeneous population can be significantly enhanced as shown in Fig.4. Alternatively, the bispecific engager molecules of the disclosure can be administered as a polypeptide(s). Design and Construction of Bispecific Engagers The efficacy of cancer treatments targeting specific B cell associated antigens may be hampered by challenges with delivery to the tumor microenvironment as well as tumor heterogeneity, which can result in selective expansion of non-targeted cells. The disclosed bispecific engagers, constructs, and methods advance the state of the art by addressing these challenges. For example, in one embodiment, a therapy employs a plurality of bispecific engagers by administering nucleic acids encoding them. Such therapies offer improved efficacy by permitting targeting of a plurality of B cell- associated molecules. For example, the expression of a plurality of bispecific engager molecules in vivo subsequent to administration of mRNA encoding different bispecific engagers permits improved targeting and depletion of inhomogeneous B cells targeted for reduction or elimination. As illustrated in the examples herein, transfecting cells with mRNA molecules (e.g., two different mRNA molecules each encoding a different bispecific engager) resulted in the production of a plurality of bispecific engagers targeting different antigens. The disclosed approaches can be used in vivo to express a plurality of bispecific engagers for effective targeting of cancer cells, more specifically B cells, e.g., cancerous B cells presenting different antigens, e.g., in the case of multiple myeloma, including relapsed or refractory multiple myeloma (RRMM). As discussed above, the present disclosure provides bispecific antibodies (“BsAbs”; also referred to herein as “engagers”) that have binding specificities for two different targets, e.g., a cell-surface protein, receptor, receptor subunit, or tissue- Attorney Docket No.: 45817-0158WO1 specific antigen, or other target, that is present on an effector cell (i.e., an NK cell) and a cell targeted for reduction or elimination (i.e., a B cell). Bispecific antibodies and engagers comprise at least two different variable domains, wherein each variable domain is capable of specifically binding to a separate target (i.e., two different molecules expressed on two different cell types). When multiple engagers are administered, they preferably comprise two different VHH that each bind different targets present on B cells. In general, the disclosed bispecific engagers will comprise a first antigen- binding domain (which binds to a molecule present on a cell targeted for reduction or elimination) and a second antigen binding domain (which targets a molecule present on an immune effector cell), and form a dimer (based on the presence of a CH2 and CH3 domain from an antibody molecule (which is typically altered to reduce effector function) when expressed in vivo from an mRNA molecule. If two mRNA molecules are administered with different binding moieties, some of the dimers produced will be heterodimers and some will be homodimers, thereby targeting two different antigens. Fig.1 also shows that three mRNAs can be administered, thereby targeting three different antigens. The subject therapeutics which can target a multiplicity of antigens result in more effective reduction or elimination of the target cell population. For example, in one embodiment, at least one binding moiety for a target antigen is present in a bispecific engager therapeutic when one mRNA is used in the therapy. In another embodiment, at least two binding moieties are present in a bispecific engager therapeutic when two mRNAs are used in the therapy. In another embodiment, at least three binding moieties are present in a bispecific engager therapeutic when three mRNAs are used in the therapy. In another embodiment, at least four binding moieties are present in a bispecific engager therapeutic when four mRNAs are used in the therapy. In another embodiment, at least five binding moieties are present in a bispecific engager therapeutic when five mRNAs are used in the therapy. In another embodiment, at least six binding moieties are present in a bispecific engager therapeutic when six mRNAs are used in the therapy. In another embodiment, at least seven binding moieties are present in a bispecific engager therapeutic when seven mRNAs are used in the therapy. In another embodiment, at least eight binding Attorney Docket No.: 45817-0158WO1 moieties are present in a bispecific engager therapeutic when eight mRNAs are used in the therapy, etc. The first antigen-binding domain and the second antigen-binding domain of the disclosed bispecific engagers are connected to one another in the engager construct via optional amino acid linkers and a CH2 and CH3 domain (typically modified to reduce effector function) to form a bispecific antibody. FIGs.1 and 5 show examples of bispecific engager formats. For example, FIG.5 shows two dimer formats in which two first antigen-binding domains (e.g., an anti-CD16 domains) and two second antigen-binding domains (e.g., an anti-tumor associate antigen or TAA domains) are bound to an Fc domain (the CH2 and CH3 domain of an antibody). The first and second antigen-binding domains may be connected at the N-terminus of the Fc domain, the C-terminus or the Fc domain, or both. In some embodiments, the antigen-binding domain targeting a B cell associated with disease (e.g., an antigen binding domain that binds to CD38, BCMA, GPRC5D, or FcRH5) is at the N- terminus of the bispecific engager and the binding domain targeting the immune effector cell (e.g., an antigen binding domain that binds to CD16a for binding to NK cells) is at the C-terminus of the bispecific engager. A signal sequence, which is not present in the mature peptide can be covalently linked to a VHH disclosed here. For example, the signal sequence can be from a human light chain (which is not present in the mature protein), and it can be covalently linked to a humanized VHH (e.g., a VHH that binds to a molecule present on the surface of a B cell targeted for reduction or elimination, the CH2 and CH3 domains from an IgG4 molecule (S228P/F234A/L235A), a G4S linker (SEQ ID NO: 448), and a humanized VHH binding to CD16a). This construct orientation was found to be efficiently expressed in cells, and minimize the formation of aggregates, and exhibit target cell killing. In addition, multiple engagers of this format were found to express in the same cell without a reduction in the potency of cell killing. Thus, one example of a suitable signal sequence is METPAQLLFLLLLWLPDTTG (SEQ ID NO: 447), and those of skill in the art will recognize that other leader sequences may be suitable as well. Attorney Docket No.: 45817-0158WO1 When expressed via an administered mRNA, the bispecific antibodies or engagers of the present disclosure are secreted (e.g., released from a cell, for example, into the extracellular milieu). For the purposes of such embodiments, the bispecific antibodies or engagers may be encoded by one or two or more mRNA, such that administration of the mRNA to a subject (e.g., a human subject) will result in expression and secretion of the encoded bispecific antibodies or engagers by cells in the subject’s body. mRNA molecules encoding the subject engagers are preferably administered via non-parenteral routes, such as intravenous (IV), intramuscular (IM), or subcutaneous (SC) routes. Bispecific antibodies or engagers of the present disclosure can comprise binding specificities that are directed towards (i) a molecule or target on a B cell associated with disease (e.g., CD38, BCMA, GPRC5D, or FcRH5) and (ii) a molecule or target on an NK cell. The molecule or target on the B cell may be associated with a disease involving B cells, such as certain types of cancer (e.g., multiple myeloma, including RRMM) or an immune disorder or autoimmune disease. In some embodiments, NK cell binding domain comprises binding specificity towards an NK antigen indicative of the state of the immune cells (e.g., an activated NK cell). Examples of NK cell targets or molecules to which the disclosed bispecific engagers may bind include, without limitation, B3GAT1 (CD57), CCR7 (CD197), CD16, CD16a, CD16b, CD2 CD226, CD244, CD27, CD3, CD300A, CD34, CD58, CD59, CD69, CSF2, CX3CR1, CXCR1 (CD128), CXCR3 (CD183), CXCR4, EOMES, GZMB, ICAM1 (CD54), IFNG, IL-15R, IL-1R, IL22, IL-2RB (CD122), IL- 7R (CD127), ITGA1 (CD49a), ITGA2 (CD49b), ITGAL (CD11a), ITGAM (CD11b), ITGB2 (CD18), KIR, KIR2DL1, KIR2DL2, KIT (CD117), KLRB1C, KLRC1, KLRC2, KLRD1 (CD94), KLRF1, KLRG1, KLRK1, LILRB1, KLRA4, KLRA8, MICA/BNCAM1 (CD56), NK2D, NKP46 (NCR1, CD335), NCR2, NCR3 (CD337), PRF1, SELL (CD62L), SIGLEC7, SLAMF6, SPN, TBX21, and TNFa. In some embodiments, bispecific antibodies of the present disclosure comprise a binding domain specific for CD16a, NKP46, NK2D, or MICA/B. In some embodiments, Attorney Docket No.: 45817-0158WO1 bispecific antibodies of the present disclosure comprise a binding domain specific for CD16a. Examples of B cell targets or molecules to which the discloses bispecific engagers may bind include, without limitation, BCMA, CADM1, CCR10, CD19, CD20, CD22, CD28, CD53, CD10, CD33, CD38, CD46, CD48/SLAMF2, CD56, CD138/SDC1, CD72, CD74/HLA-DR, CS-1, EVI2B, FcRH5, GGT1, GPRC5D, Integrin beta-7, LY9/CD229, SELPLG, SLAMF7, TACI, and TXNDC11. In some embodiments, bispecific antibodies of the present disclosure comprise a binding domain specific for FcRH5, GPRC5D, BCMA, or CD38. In some embodiments, bispecific antibodies of the present disclosure comprise a binding domain specific for FcRH5. In some embodiments, bispecific antibodies of the present disclosure comprise a binding domain specific for GPRC5D. In some embodiments, bispecific antibodies of the present disclosure comprise a binding domain specific for BCMA. In some embodiments, bispecific antibodies of the present disclosure comprise a binding domain specific for CD38. Single mRNA molecules encoding single bispecific engagers can be used to express single protein molecules, which illustrates bispecific antibodies targeting a single B cell target (e.g., FcRH5, GPRC5D, BCMA, or CD38) and a single NK binding domain (NK engager sequence). Co-expression of two different mRNA molecules each encoding bispecific engagers targeting a different B cell target, but the same NK binding domain or different NK binding domains that each bind the same target, may be used to engage NK cells with various cells for which is desirable to deplete (e.g., cancer cells. This strategy is useful for targeting heterogeneous cell populations that may express different markers, as is often the case in tumors (i.e., tumors often contain mixed populations of cancer cells). Co-expression of two different mRNA molecules each encoding bispecific engagers targeting a different B cell molecule and the same or a different NK molecule, may also be useful in treating cancers. Co-expression of three, four, or five or more different mRNA molecules each encoding bispecific engagers targeting a different B cell molecule (with the same or different NK binding domains) could also be used to treat diseases and reduce or Attorney Docket No.: 45817-0158WO1 deplete the number of B cells, which would be advantageous for heterologous tumors and cancers, as shown in FIG.5. Such engagers can be selected and formulated in response to the specific cancer cell population detected in a subject. When multiple mRNA encoding different bispecific engagers are administered to a subject, the mRNA may be formulated together in the same delivery composition or formulated separately in two or more different delivery compositions. For example, a first mRNA encoding a first bispecific engager and a second mRNA encoding a second bispecific engager (and, optionally, a third, fourth, fifth mRNA, etc., each encoding a further bispecific engager) can be formulated in a single pharmaceutical composition and administered to a subject concurrently. Alternatively, a first mRNA encoding a first bispecific engager and a second mRNA encoding a second bispecific engager (and, optionally, a third, fourth, fifth mRNA, each encoding a further bispecific engager) can be formulated into two different pharmaceutical compositions and administered concurrently, serially (i.e., back-to-back or one immediately following the other), or sequentially (i.e., with some predetermined amount of time between the administration of the first and second composition). The decision regarding how many different mRNAs should be administered, how many bispecific engagers should be expressed, and what B cell targets the bispecific engagers should bind may be determined based on a screening of a subject’s disease. The bispecific engagers described herein include an Fc domain. Examples of suitable Fc domains include the CH2 and CH3 domain from hu-IgG1, optionally containing one or several point mutations to abrogate Fc G1 functionality (e.g., L234F, L235E and D265A) (see Liu et al., Fc-Engineering for Modulated Effector Functions-Improving Antibodies for Cancer Treatment. Antibodies (Basel).2020 Nov 17; 9(4):64). Other modifications that can be incorporated in order to decrease or abolish Fc functionality in various IgG Fc domains include, but are not limited to, aglycosylation via N297 substitution (e.g., with A, Q, or G); L235A/G237A/E318A; L234A/L235A; S228P/L235E; G236R/L328R; S298G/T299A; L234F/L235E/P331S; H268Q/V309L/A330S/P331S; E233P/L234V/L235A/G236del/S267K; L234A/L235A/P329G; and V234A/G237A/P238S/H268A/V309L/ A330S/P331S. In Attorney Docket No.: 45817-0158WO1 some cases, the Fc domains include the hinge region, the CH2 and the CH3 domain from hu-IgG1, optionally containing one or several point mutations to abrogate Fc G1 functionality (e.g., L234F, L235E and D265A). Additionally or alternatively, if an IgG4 Fc domain is used, such an Fc domain may be stabilized with a PAA sequence (S228P/F234A/L235A) in the hinge region to prevent in vivo scrambling with other IgG4 antibodies that could be found in human sera (see Gillies et al., Cancer Res. (1999), 59: 2159–2166; Saunders, Conceptual Approaches to Modulating Antibody Effector Functions and Circulation Half-Life. Front Immunol.2019 Jun 7; 10:1296). In some cases, the IgG4 Fc comprises an IgG4 hinge region with a PAA mutation (S228P/F234A/L235A), along with an IgG4 CH2 and CH3 region. In one instance, the bispecific NK engagers of this disclosure comprises an amino acid sequence that is at least 80%, at least 85%, at least 90%, at least 95%, at least 86%, at least 97%, at least 98%, at least 99%, or 100% identical to the sequence set forth below: ESKYGPPCPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPE VQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCK VSNKGLPSSIEKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSD IAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVM HEALHNHYTQKSLSLSLGK (SEQ ID NO:548) As discussed in more detail below, also provided herein are nucleic acids, such as mRNA, that encode bispecific engagers, delivery vehicles, and methods of their use as disclosed herein. As an illustrative example, the following relates to just one example of a bispecific engager of this disclosure. Provided herein is an mRNA that encodes a tetravalent bispecific antibody (BsAb) as well as the encoded polypeptide that directs immune effector cells to, and induces killing of, FCRH5+ cell lines and patient samples. A NK Engager (NKE) composition of this disclosure comprises a lipid nanoparticle (LNP) encapsulating a single messenger RNA (mRNA) construct that encodes an N-terminal FCRH5 single domain antibody (sdAb) followed by an Attorney Docket No.: 45817-0158WO1 immunoglobulin G4 (IgG4) Fc domain with PAA mutations, and a C-terminal anti- CD16 sdAb (αFCRH5). The translated protein homodimerizes at the Fc domain resulting in a tetravalent construct. The expressed protein from the systemically delivered NKE composition is intended to target FCRH5+ malignant plasma cells in multiple myeloma patients by engaging NK cells and macrophages via CD16 to induce antibody-dependent cellular cytotoxicity (ADCC) and antibody-dependent cellular phagocytosis (ADCP), respectively. FCRH5, also known as FCRL5, is a member of the extended Fc receptor-like (FCRL) family. The human FCRL5 Ig8 extracellular domain (ECD) shares an 87 percent identity with cynomolgus monkey, demonstrating conserved domain structure and true orthology of cynomolgus FCRL5. Mouse FCRL5, also known as mouse FCRH3, has a shorter ECD with only 5 Ig-like domains compared to that of human FCRL5. The percent identities of amino acid sequences of human FCRL5 Ig8 to any mouse Ig- like domains is less than 60%, suggesting that mouse FCRL5 is not a true ortholog of human FCRL5 based on the domain structure and homology. The two human CD16a variants V176 and F176 share 92 percent identity with cynomolgus CD16a. Human CD16b variants share 89 and 90 percent identity (NA1 and NA2, respectively) with cynomolgus CD16a. There is no CD16b in cynomolgus. Using SPR, αFCRH5 NKE protein bound to full-length human FCRH5 ECD with an affinity of 6 nM and to human FCRH5-Ig8 with an affinity of 19 nM. The cross-binding affinity to cynomolgus FCRH5-Ig8 was 48 nM. The binding to the mouse homolog (FCRL5/FCRH3) could not be determined. The binding affinity of αFCHR5 NKE protein to human FCRL family members ranged from 389 nM (FCRH1) to no binding observed. αFCRH5 NKE protein bound to CD16a V176 with an affinity of 12 nM and to CD16a F176 with an affinity of 17 nM indicated that αFCRH5 NKE can bind to NK cells and macrophages similarly in individuals carrying different CD16a alleles. No binding was observed to CD16b NA1 while binding to CD16b NA2 had an affinity of 22 nM indicating αFCRH5 NKE only binds to neutrophils in individuals carrying the Attorney Docket No.: 45817-0158WO1 CD16b-NA2 allele. Binding of αFCRH5 NKE protein to cynomolgus CD16a had an affinity of 13 nM, while binding to rat CD16a had an affinity of 54nM. No binding of αFCRH5 NKE protein to mouse CD16 was observed. The target density of FCRH5 on human cancer cell lines was established. The transduced lines RPMI-8226-huFCRH5, RPMI-8226-cyFCRH5, and MOLM-13- huFCRH5 had target densities of 71787 molecules per cell at the cell surface, 5016 molecules per cell at the cell surface, and 15454 molecules per cell at the cell surface, respectively. In non-transduced SU-DHL-6 cells, the endogenous level of FCRH5 was 127 molecules per cell at the cell surface. The target density of FCRH5 on human B cells ranged from 235 to 580 FCHR5 molecules per cell at the cell surface. There was no significant FCRH5 expression on human T cells, NK cells, or monocytes. The target density of FCRH5 on patient samples ranged from 1300 to12954 cell surface molecules, with a range of FCRH5 positive cells from 32 to 96 percent positive. The binding affinity of αFCRH5 NKE protein to FCRH5 expressing cells were in the single digit nanomolar range for both human (6.7 nM) and cynomolgus (3.3 nM) FCRH5 transduced RPMI-8226 cells. The binding affinity of αFCRH5 NKE protein was 24.1 nM on endogenously FCRH5 expressing SU-DHL-6 cells. The αFCRH5 domain was required to bind human and cynomolgus B cells. αFCRH5 NKE protein bound to B cells with an affinity of 66.5 nM for human B cells and 1.5 nM for cynomolgus B cells. The CD16 domain was required to bind human and cynomolgus NK cell, NKT cell, CD14+ monocytes, and myeloid dendritic cells (mDCs). αFCRH5 NKE protein had a binding affinity of 3.8 nM for human NK cells and 2.4 nM for cynomolgus NK cells. αFCRH5 NKE protein had a binding affinity of 1.4 nM for human NKT cells and 2.4 nM for cynomolgus NKT cells. When gated on the anti-his+ only population (CD16+ and engager protein bound), αFCRH5 NKE protein had a binding affinity of 1.2 nM for human CD14+ monocytes and 1.5 nM for cynomolgus CD14+ monocytes. αFCRH5 NKE protein had a binding affinity of 1.2 nM for human CD11c+ CD14- mDCs and 2.6 nM for cynomolgus CD11c+ CD14- Attorney Docket No.: 45817-0158WO1 mDCs. Human and cynomolgus T cells and plasmacytoid DCs (pDCs) did not bind αFCRH5 NKE protein. The estimated binding affinity of αFCRH5 NKE protein to human neutrophils (from two donors) ranged from 10.6 to 11.9 nM, while for the NK Cells from the same donors ranged from 10.3 to 10.6 nM. Cell surface CD16 was progressively lost over time on NK cells upon incubation with αFCRH5 NKE protein. However, CD16 levels on neutrophils did not show significant reduction compared to cells in the absence of αFCRH5 NKE protein, suggesting that neutrophil binding of αFCRH5 NKE protein may play a minor role in target mediated drug disposition (TMDD). No dose dependence in CD16 loss was observed. For RPMI-8226-huFCRH5 cells, the EC50 of specific killing ranged from 7.1 pM (effector to target cell ratio of 10) to 8.6 pM (E:T ratio of 2.5). For RPMI-8226- cyFCRH5 cells, the EC50 of specific killing ranged from 11.3 pM (E:T ratio of 5) to 20.3 pM (E:T ratio of 2.5). For MOLM-13-huFCRH5, the EC50 was 7.8 nM at an E:T ratio of 8. For SU-DHL-6, the EC50 was 10.4 nM at an E:T ratio of 10. The potency of human NK cell cytotoxicity in the presence of human neutrophils induced by αFCRH5 NKE protein was established. The EC50 in the presence of neutrophils and αFCRH5 NKE protein was 16.4 pM and 11.2 pM for two donors tested, a 2- to 3-fold potency reduction compared to the no neutrophil control (EC50 = 6.4 pM). The potency of human NK cell cytotoxicity against MM patient derived tumor cells in the presence of αFCHR5 NKE protein was evaluated using an autologous system. Three of four donors evaluated showed specific killing of tumor cells with EC50 ranging from 18 to 316 pM. This illustrative example supports the utility of the several different bispecific engagers described further below. Binding Domains Among the molecular features of bispecific engagers described herein comprising target-binding domains described herein, it will be appreciated by one of Attorney Docket No.: 45817-0158WO1 skill in the art that the CDRs are those regions that predominantly dictate the target- binding properties of the molecule. Bispecific antibodies or engagers of the present disclosure can include single domain antibodies or VHH domains which bind to the target antigens described herein or known in the art. In general each binding domain will consist of a VHH. In some embodiments, the antigen binding domain portion comprises a mammalian VHH, such as a llama VHH or a humanized version thereof. The choice of target molecule binding domain may depend upon the type and number of antigens that are present on the surface of a target cell in any particular patient. In some aspects, alternate VHH domains binding to molecules on B cells or NK cells or tolerable variations in the disclosed binding domain will be known to those of skill in the art, while maintaining binding to the target antigen. Binding Domains Targeting Natural Killer (NK) Cells The disclosed bispecific engagers comprise at least one binding domain that binds to a target molecule on or associated with a NK cell. In general, the target of this binding domain will engage the NK cell and/or bring it into proximity with the B cell targeted by the other binding domain of the bispecific engager. The target molecule on the NK cell can be selected from B3GAT1 (CD57), CCR7 (CD197), CD16, CD16a, CD16b, CD2 CD226, CD244, CD27, CD3, CD300A, CD34, CD58, CD59, CD69, CSF2, CX3CR1, CXCR1 (CD128), CXCR3 (CD183), CXCR4, EOMES, GZMB, ICAM1 (CD54), IFNG, IL-15R, IL-1R, IL22, IL-2RB (CD122), IL- 7R (CD127), ITGA1 (CD49a), ITGA2 (CD49b), ITGAL (CD11a), ITGAM (CD11b), ITGB2 (CD18), KIR, KIR2DL1, KIR2DL2, KIT (CD117), KLRB1C, KLRC1, KLRC2, KLRD1 (CD94), KLRF1, KLRG1, KLRK1, LILRB1, KLRA4, KLRA8, MICA/BNCAM1 (CD56), NK2D, NKP46 (NCR1, CD335), NCR2, NCR3 (CD337), PRF1, SELL (CD62L), SIGLEC7, SLAMF6, SPN, TBX21, and TNFa. In some embodiments, the target molecule on the NK cell can be selected from CD16a, NKP46, NK2D, and MICA/B. In some embodiments, the target molecule on the NK cell is CD16a. Attorney Docket No.: 45817-0158WO1 In some embodiments, the binding domain that binds to an NK cell (i.e., a non-tumor cell or immune cell) binds to CD16, or more specifically, CD16a. In certain instances, the binding domain can bind to both human CD16a variants, namely V176 and F176. In some instances, the binding domain can also bind to both cynomolgus CD16a. In some instances, the binding domain can also bind to CD16b NA2. In some instances, the binding domain can does not bind to CD16b NA1. Binding to CD16 allows bispecific engagers to recruit or engage NK cells. An anti- CD16 antibody or binding domain suitable for the disclosed bispecific engagers may comprise one or more of the following CDRs: a CDR1 having the amino acid sequence GRTDSIYA (SEQ ID NO: 2) or an amino acid sequence having up to two amino acid substitutions (e.g., conservative amino acid substitutions) relative to SEQ ID NO: 2; a CDR2 having the amino acid sequence INSNTGRT (SEQ ID NO: 3), or an amino acid sequence having up to two amino acid substitutions (e.g., conservative amino acid substitutions) relative to SEQ ID NO: 3; and a CDR3 having the amino acid sequence AAGRGYGLLSISSNWYNY(SEQ ID NO: 4), or an amino acid sequence having up to two amino acid substitutions (e.g., conservative amino acid substitutions) relative to SEQ ID NO: 4. In some instances, the binding domain that binds to an NK cell is a VHH that specifically binds CD16 (e.g., human and cynomolgus CD16a) and comprises the VHH CDR1 comprising the sequence of SEQ ID NO:2, the VHH CDR2 comprising the sequence of SEQ ID NO:3, and the VHH CDR3 comprising the sequence of SEQ ID NO:4. In some instances, the binding domain that binds to an NK cell is a VHH that specifically binds CD16 (e.g., human and cynomolgus CD16a) and comprises a VHH CDR1, a VHH CDR2, and a VHH CDR3 of the VHH with the sequence of SEQ ID NO:7. Table I: Exemplary CDR Definitions for CD16 VHH CDR VHH-CDR1 VHH-CDR2 VHH-CDR3 Definition IMGT

Attorney Docket No.: 45817-0158WO1 K

Table II: Exemplary CDR Definitions for Another CD16 VHH CDR VHH-CDR1 VHH-CDR2 VHH-CDR3 D fi i i

In some instances, the binding domain that binds to an NK cell is a VHH that specifically binds CD16 (e.g., human and cynomolgus CD16a) and comprises a VHH CDR1, a VHH CDR2, and a VHH CDR3 according to any one CDR definition provided in Table I or Table II. In some cases, the VHH that specifically binds CD16 is humanized. In some cases, one or more of positions 42, 49, 50, and 52 (IMGT numbering) of an anti- CD16 VHH are humanized. In some cases, one or more of positions 42, 49, 50, and Attorney Docket No.: 45817-0158WO1 52 (IMGT numbering) of an anti-CD16 VHH are not humanized. In certain cases positions 42 and/or 52 (IMGT numbering) of an anti-CD16 VHH are not humanized. In certain cases positions 49 and/or 50 IMGT numbering) of an anti-CD16 VHH are humanized In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of: QVQLVESGGGLVQAGGSLRLSCAASGRTDSIYAMGWFRQAPGKERDFVAAI NSNTGRTYHADSVKGRFTISRDNAKNMVYLQMNSLKPEDTAVYYCAAGRG YGLLSISSNWYNYWGQGTQVTVSS (VH1, SEQ ID NO: 1; CD16-VHH1), wherein the three VHH CDRs are not altered. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 1. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 1. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise the amino acid sequence of SEQ ID NO: 1. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of: EVQLVESGGGLVQPGGSLRLSCAASGRTDSIYAMGWFRQAPGKELDFVSAIN SNTGRTYHADSVKGRFTISRDNAKNTVYLQMNSLRAEDTAVYYCAAGRGYG LLSISSNWYNYWGQGTLVTVSS (SEQ ID NO: 5; CD16-VHH2), wherein the three VHH CDRs are not altered. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, Attorney Docket No.: 45817-0158WO1 or 100% identical) to the amino acid sequence of SEQ ID NO: 5. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 5. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise the amino acid sequence of SEQ ID NO: 5. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of: QVQLVESGGGLVQPGGSLRLSCAASGRTDSIYAMGWFRQAPGKELEFVAAIN SNTGRTYHADSVKGRFTISRDNAKNTLYLQMNSLKAEDTAVYYCAAGRGYG LLSISSNWYNYWGQGTLVTVSS (SEQ ID NO: 6; CD16-VHH3), wherein the three VHH CDRs are not altered. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 6. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 6. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise the amino acid sequence of SEQ ID NO: 6. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of: EVQLVESGGGLVQPGGSLRLSCAASGRTDSIYAMGWFRQAPGKEREFVSAIN SNTGRTYHADSVKGRFTISRDNAKNMVYLQMNSLRAEDTAVYYCAAGRGY GLLSISSNWYNYWGQGTLVTVSS (SEQ ID NO: 7; CD16-VHH4), wherein the three VHH CDRs are not altered. In some embodiments, the anti-CD16 antibody or Attorney Docket No.: 45817-0158WO1 binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 7. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 7. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise the amino acid sequence of SEQ ID NO: 7. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of: EVQLVESGGGLVQPGGSLRLSCAASGRTDSIYAMGWFRQAPGKELEFVSAIN SNTGRTYHADSVKGRFTISRDNAKNMVYLQMNSLKAEDTAVYYCAAGRGY GLLSISSNWYNYWGQGTLVTVSS (SEQ ID NO: 8; CD16-VHH5), wherein the three VHH CDRs are not altered. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 8. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 8. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise the amino acid sequence of SEQ ID NO: 8. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of: QVQLVESGGGLVQPGGSLRLSCAASGRTDSIYAMGWFRQAPGKELEFVSAIN SNTGRTYHADSVKGRFTISRDNAKNTLYLQMNSLKPEDTAVYYCAAGRGYG Attorney Docket No.: 45817-0158WO1 LLSISSNWYNYWGQGTLVTVSS (SEQ ID NO: 9; CD16-VHH6), wherein the three VHH CDRs are not altered. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 9. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 9. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise the amino acid sequence of SEQ ID NO: 9. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of: QVQLVESGGGLVQPGGSLRLSCAASGRTDSIYAMGWFRQAPGKEREFVSAIN SNTGRTYHADSVKGRFTISRDNAKNTLYLQMNSLRPEDTAVYYCAAGRGYG LLSISSNWYNYWGQGTLVTVSS (SEQ ID NO: 10; CD16-VHH7), wherein the three VHH CDRs are not altered. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 10. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 10. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise the amino acid sequence of SEQ ID NO: 10. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of: Attorney Docket No.: 45817-0158WO1 EVQLVESGGGLVQPGGSLRLSCAASGRTDSIYAMGWFRQAPGKEREFVSAIN SNTGRTYHADSVKGRFTISRDNAKNTVYLQMNSLRPEDTAVYYCAAGRGYG LLSISSNWYNYWGQGTLVTVSS (SEQ ID NO: 11; CD16-VHH8), wherein the three VHH CDRs are not altered. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 11. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 11. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise the amino acid sequence of SEQ ID NO: 11. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of: QVQLVESGGGLVQPGGSLRLSCAASGRTDSIYAMGWFRQAPGKELEFVSAIN SNTGRTYHADSVKGRFTISRDNAKNTVYLQMNSLKAEDTAVYYCAAGRGYG LLSISSNWYNYWGQGTLVTVSS (SEQ ID NO: 12; CD16-VHH9), wherein the three VHH CDRs are not altered. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 12. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 12. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise the amino acid sequence of SEQ ID NO: 12. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, Attorney Docket No.: 45817-0158WO1 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of: QVQLVESGGGLVQPGGSLRLSCAASGRTDSIYAMGWFRQAPGKERDFVSAIN SNTGRTYHADSVKGRFTISRDNAKNTLYLQMNSLRPEDTAVYYCAAGRGYG LLSISSNWYNYWGQGTLVTVSS (SEQ ID NO: 13; CD16-VHH10), wherein the three VHH CDRs are not altered. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 13. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 13. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise the amino acid sequence of SEQ ID NO: 13. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of: EVQLVESGGGLVQPGGSLRLSCAASGRTDSIYAMGWFRQAPGKERDFVSAIN SNTGRTYHADSVKGRFTISRDNAKNTVYLQMNSLKAEDTAVYYCAAGRGYG LLSISSNWYNYWGQGTLVTVSS (SEQ ID NO: 14; CD16-VHH11), wherein the three VHH CDRs are not altered. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 14. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 14. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise the amino acid sequence of SEQ ID NO: 14. Attorney Docket No.: 45817-0158WO1 In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of: EVQLVESGGGLVQPGGSLRLSCAASGRTDSIYAMGWFRQAPGKELDFVSAIN SNTGRTYHADSVKGRFTISRDNAKNMVYLQMNSLRPEDTAVYYCAAGRGY GLLSISSNWYNYWGQGTLVTVSS (SEQ ID NO: 15; CD16-VHH12), wherein the three VHH CDRs are not altered. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 15. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 15. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise the amino acid sequence of SEQ ID NO: 15. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of: QVQLVESGGGLVQPGGSLRLSCAASGRTDSIYAMGWFRQAPGKERDFVSAIN SNTGRTYHADSVKGRFTISRDNAKNTVYLQMNSLRAEDTAVYYCAAGRGYG LLSISSNWYNYWGQGTLVTVSS (SEQ ID NO: 16; CD16-VHH13), wherein the three VHH CDRs are not altered. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 16. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 16. In Attorney Docket No.: 45817-0158WO1 some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise the amino acid sequence of SEQ ID NO: 16. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of: QVQLVESGGGLVQPGGSLRLSCAASGRTDSIYAMGWFRQAPGKERDFVAAIN SNTGRTYHADSVKGRFTISRDNAKNTLYLQMNSLRAEDTAVYYCAAGRGYG LLSISSNWYNYWGQGTLVTVSS (SEQ ID NO: 17; CD16-VHH14), wherein the three VHH CDRs are not altered. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 17. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 17. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise the amino acid sequence of SEQ ID NO: 17. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of: QVQLVESGGGLVQPGGSLRLSCAASGRTDSIYAMGWFRQAPGKEREFVSAIN SNTGRTYHADSVKGRFTISRDNAKNMLYLQMNSLKAEDTAVYYCAAGRGY GLLSISSNWYNYWGQGTLVTVSS (SEQ ID NO: 18; CD16-VHH15), wherein the three VHH CDRs are not altered. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 18. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise Attorney Docket No.: 45817-0158WO1 an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 18. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise the amino acid sequence of SEQ ID NO: 18. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of: QVQLVESGGGLVQPGGSLRLSCAASGRTDSIYAMGWFRQAPGKEREFVAAIN SNTGRTYHADSVKGRFTISRDNAKNTLYLQMNSLKAEDTAVYYCAAGRGYG LLSISSNWYNYWGQGTLVTVSS (SEQ ID NO: 19; CD16-VHH16), wherein the three VHH CDRs are not altered. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 19. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 19. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise the amino acid sequence of SEQ ID NO: 19. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of: QVQLVESGGGLVQPGGSLRLSCAASGRTDSIYAMGWFRQAPGKERDFVSAIN SNTGRTYHADSVKGRFTISRDNAKNTLYLQMNSLKAEDTAVYYCAAGRGYG LLSISSNWYNYWGQGTLVTVSS (SEQ ID NO: 20; CD16-VHH17), wherein the three VHH CDRs are not altered. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, Attorney Docket No.: 45817-0158WO1 or 100% identical) to the amino acid sequence of SEQ ID NO: 20. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 20. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise the amino acid sequence of SEQ ID NO: 20. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of: QVQLVESGGGLVQPGGSLRLSCAASGRTDSIYAMGWFRQAPGKELDFVAAIN SNTGRTYHADSVKGRFTISRDNAKNMLYLQMNSLRAEDTAVYYCAAGRGY GLLSISSNWYNYWGQGTLVTVSS (SEQ ID NO: 21; CD16-VHH18), wherein the three VHH CDRs are not altered. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 21. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 21. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise the amino acid sequence of SEQ ID NO: 21. In some embodiments, the anti-CD16 antibody or binding domain (e.g., scFv or VHH) may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of: QVQLVESGGGLVQPGGSLRLSCAASGRTDSIYAMGWFRQAPGKERDFVSAIN SNTGRTYHADSVKGRFTISRDNAKNMLYLQMNSLKAEDTAVYYCAAGRGY GLLSISSNWYNYWGQGTLVTVSS (SEQ ID NO: 22; CD16-VHH19), wherein the three VHH CDRs are not altered. In some embodiments, the anti-CD16 antibody or Attorney Docket No.: 45817-0158WO1 binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 22. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 22. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise the amino acid sequence of SEQ ID NO: 22. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of: EVQLVESGGGLVQPGGSLRLSCAASGRTDSIYAMGWFRQAPGKERDFVAAIN SNTGRTYHADSVKGRFTISRDNAKNTVYLQMNSLKPEDTAVYYCAAGRGYG LLSISSNWYNYWGQGTLVTVSS (SEQ ID NO: 23; CD16-VHH20), wherein the three VHH CDRs are not altered. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 23. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 23. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise the amino acid sequence of SEQ ID NO: 23. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of: QVQLVESGGGLVQPGGSLRLSCAASGRTDSIYAMGWFRQAPGKERDFVAAIN SNTGRTYHADSVKGRFTISRDNAKNTVYLQMNSLKAEDTAVYYCAAGRGYG Attorney Docket No.: 45817-0158WO1 LLSISSNWYNYWGQGTLVTVSS (SEQ ID NO: 24; CD16-VHH21), wherein the three VHH CDRs are not altered. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 24. In some embodiments, the anti-CD16 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 24. In some embodiments, anti-CD16 the antibody or binding domain (e.g., VHH) may comprise the amino acid sequence of SEQ ID NO: 24. Binding Domains Targeting B cell Associated Molecules The B cell binding domain of a disclosed bispecific engager may bind to any molecule associated with a B cell targeted for reduction or elimination. For example, the molecule targeted on a B cell can be a tumor associated antigen (TAA) or another molecule associated with or expressed on a B cell. Non-limiting examples of molecules that may serve as targets on a B cell include BCMA, CADM1, CCR10, CD19, CD20, CD22, CD28, CD53, CD10, CD33, CD38, CD46, CD48/SLAMF2, CD56, CD138/SDC1, CD72, CD74/HLA-DR, CS-1, EVI2B, FcRH5, GGT1, GPRC5D, Integrin beta-7, LY9/CD229, SELPLG, SLAMF7, TACI, and TXNDC11. In some embodiments, the molecule targeted on the B cell is selected from FcRH5, GPRC5D, BCMA, and CD38. Examples of CDRs for anti-CD38 antibodies or binding domains include: (a) a CDR1 comprising the amino acid sequence selected from the group consisting of GFILDTYS (SEQ ID NO: 25); GFIFSDKV (SEQ ID NO: 26); RSIFEINTMT (SEQ ID NO: 27); GFSLDYYH (SEQ ID NO: 28); GAIVSAES (SEQ ID NO: 29); GTFSSINL (SEQ ID NO: 30); GSISGLNT (SEQ ID NO: 31); GSSVSMNS (SEQ ID NO: 32); GFIYSIST (SEQ ID NO: 33); GRYFRINA (SEQ ID NO: 34); GTFSSIAL (SEQ ID NO: 35); and GIIFRIFS (SEQ ID NO: 36); or an amino acid sequence having up to two amino acid substitutions (e.g., conservative Attorney Docket No.: 45817-0158WO1 amino acid substitutions) relative to any one of SEQ ID NOs: 25-36; and (b) a CDR2 comprising the amino acid sequence selected from the group consisting of: ISSRDGNT (SEQ ID NO: 37); ITPGGTAT (SEQ ID NO: 38); SRGATT (SEQ ID NO: 39); ISSSDGYT (SEQ ID NO: 40); IISGSKS (SEQ ID NO: 41); DYTEGTT (SEQ ID NO: 42); IISGTMT (SEQ ID NO: 43); ITPGDRI (SEQ ID NO: 44); ITSGGNT (SEQ ID NO: 45); ISNDGST (SEQ ID NO: 46); VSIGGVT (SEQ ID NO: 47); and ITSGGNT (SEQ ID NO: 48); or an amino acid sequence having up to two amino acid substitutions (e.g., conservative amino acid substitutions) relative to any one of SEQ ID NOs: 37-48; and (c) a CDR3 comprising the amino acid sequence selected from the group consisting of: AAGAQAHCTIFTSYFNSDYYRRYNY (SEQ ID NO: 49); RIGGPGGRYDN (SEQ ID NO: 50); SADRYGFGYGDNDY (SEQ ID NO: 51); AASPRRLACAGSLYPPLSADFSS (SEQ ID NO: 52); KRTERIWTNNPQVY (SEQ ID NO: 53); WLMVRAGDVY (SEQ ID NO: 54); TFKEITRDSRSY (SEQ ID NO: 55); NIGATRPPFGA (SEQ ID NO: 56); NTAWRETIVSRV (SEQ ID NO: 57); NVKALPFLSSNELSY (SEQ ID NO: 58); WRMEGAGDVY (SEQ ID NO: 59); and NVAIPSGIVDRSA (SEQ ID NO: 60); or an amino acid sequence having up to two amino acid substitutions (e.g., conservative amino acid substitutions) relative to any one of SEQ ID NOs: 49-60. Examples of anti-CD38 CDRs from VHH domains suitable for incorporation in the disclosed bispecific engagers are shown in Table 1 below. Table 1 – Examples of anti-CD38 CDRs Antibody CDR1 CDR2 CDR3 VHH C^D38-VHH1 _GFILDTYS ISSRDGNT AAGAQAHCTIFTSYFNSDYYRRY

Attorney Docket No.: 45817-0158WO1 C^{D38 HH5} _{AI AE} II K KRTERI T P Y

e ea pes o a - s o o a s su a e o incorporation in the disclosed bispecific engagers are shown in Table A below. Table A. Exemplary CDR definitions for anti-CD38 VHHs IMGT CDR Definition Clone Name VHH-CDR1 VHH-CDR2 VHH-CDR3 GFILDTYS ISSRDGNT AAGAQAHCTIFTSYFNSDYYRRYNY CD38-VHH1 (SEQ ID NO:25) (SEQ ID NO:37) (SEQ ID NO:49) GFIFSDKV ITPGGTAT RIGGPGGRYDN CD38-VHH2 (SEQ ID NO:26) (SEQ ID NO:38) (SEQ ID NO:50) RSIFEINTMT SRGATT SADRYGFGYGDNDY CD38-VHH3 (SEQ ID NO:27) (SEQ ID NO:39) (SEQ ID NO:51) Kabat CDR Definition Clone Name VHH-CDR1 VHH-CDR2 VHH-CDR3 CD38-VHH1 TYSVA CISSRDGNTFYSDSVKG GAQAHCTIFTSYFNSDYYRRYNY (SEQ ID NO: 561) (SEQ ID NO:562) (^{SEQ ID NO:563)} CD38-VHH2 DKVMS TITPGGTATSYTESVKG GGPGGRYDN Attorney Docket No.: 45817-0158WO1 (SEQ ID NO: 564) (SEQ ID NO: 565) (SEQ ID NO:566 ) CD38-VHH3 INTMTMG ASRGATTNYADSVKG DRYGFGYGDNDY (SEQ ID NO:567) (SEQ ID NO:568) (SEQ ID NO:569 ) Enhanced Chothia CDR Definition Clone Name VHH-CDR1 VHH-CDR2 VHH-CDR3 CD38-VHH1 GFILDTYSVA CISSRDGNTF GAQAHCTIFTSYFNSDYYRRYNY (SEQ ID NO:570 ) (SEQ ID NO:571) (SEQ ID NO:572 ) CD38-VHH2 GFIFSDKVMS TITPGGTATS GGPGGRYDN (SEQ ID NO:573 ) (SEQ ID NO:574 ) (SEQ ID NO:575 ) CD38-VHH3 RSIFEINTMTMG ASRGATTN DRYGFGYGDNDY (SEQ ID NO:576 ) (SEQ ID NO:577 ) (^{SEQ ID NO:578 )} Contact CDR Definition Clone Name VHH-CDR1 VHH-CDR2 VHH-CDR3 DTYSVA GVSCISSRDGNTF AAGAQAHCTIFTSYFNSDYYRRYN CD38-VHH1 (SEQ ID NO:579 ) (SEQ ID NO:580 ) (SEQ ID NO:581) SDKVMS WVSTITPGGTATS RIGGPGGRYD CD38-VHH2 (SEQ ID NO: 582) (SEQ ID NO: 583) (SEQ ID NO: 584) EINTMTMG LISASRGATTN SADRYGFGYGDND CD38-VHH3 (SEQ ID NO: 585 ) (SEQ ID NO:586 ) (SEQ ID NO: 587) Chothia CDR Definition Clone Name VHH-CDR1 VHH-CDR2 VHH-CDR3 GFILDTY SSRDGN GAQAHCTIFTSYFNSDYYRRYNY CD38-VHH1 (SEQ ID NO: 588) (SEQ ID NO: 589) (SEQ ID NO: 590) GFIFSDK TPGGTA GGPGGRYDN CD38-VHH2 (SEQ ID NO:591) (SEQ ID NO: 592) (SEQ ID NO: 593) RSIFEIN RGAT DRYGFGYGDNDY CD38-VHH3 (SEQ ID NO: 594) (SEQ ID NO:595) (SEQ ID NO: 596) In some instances, a bispecific engager of this disclosure comprises the three CDRs of any one VHH disclosed in Table 1 or Table A. Attorney Docket No.: 45817-0158WO1 Examples of anti-CD38 VHH domains suitable for incorporation in the disclosed bispecific engagers are shown in Table 2 below. Table 2 – Examples of anti-CD38 VHH domains Antibody Amino acid sequence SEQ ID

Attorney Docket No.: 45817-0158WO1 CD38 VHH12 V LVESGGGLV AGGSLRLSCVASGIIFRIFSMGWYR APGK R 249

In some embodiments, the anti-CD38 antibody or binding domain (e.g., VHH) comprises an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of any one of SEQ ID NOs: 238- 252, wherein the three VHH CDRs are not altered. In some embodiments, the anti- CD38 antibody or binding domain (e.g., VHH) comprises an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of any one of SEQ ID NOs: 238-252. In some embodiments, the anti-CD38 antibody or binding domain (e.g., VHH) comprises an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of any one of SEQ ID NOs: 238-252. In some embodiments, the anti-CD38 antibody or binding domain (e.g., VHH) comprises an amino acid sequence that has 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 amino acid substitutions within the amino acid sequence of any one of SEQ ID NOs: 238-252. In some embodiments, the anti-CD38 antibody or binding domain (e.g., VHH) comprises the amino acid sequence of any one of SEQ ID NOs: 238-252. Examples of CDRs for anti-BCMA antibodies or binding domains include: (a) a CDR1 comprising the amino acid sequence selected from the group consisting of GRTLSPYT (SEQ ID NO: 61); GRTLNNYV (SEQ ID NO: 62); GSIFAYHV (SEQ ID NO: 63); GRTFSDYT (SEQ ID NO: 64); GRPLRMYN (SEQ Attorney Docket No.: 45817-0158WO1 ID NO: 65); GSTFSRYA (SEQ ID NO: 66); GFTLSSYW (SEQ ID NO: 67); GFTYSSYW (SEQ ID NO: 68); GGTLEYYA (SEQ ID NO: 69); GFTFSSYW (SEQ ID NO: 70); GRIDSGYT (SEQ ID NO: 71); GFTFGSYW (SEQ ID NO: 72); GHTLNSYA (SEQ ID NO: 73); GFTFSTYS (SEQ ID NO: 74); GHTFSNSA (SEQ ID NO: 75); GRTFSGYT (SEQ ID NO: 76); GRTFSSYA (SEQ ID NO: 77); GRTIGSFV (SEQ ID NO: 78); GFTFSNYA (SEQ ID NO: 79); VRYFSTYA (SEQ ID NO: 80); GFTFDDYA (SEQ ID NO: 81); GFNLTSDA (SEQ ID NO: 82); GRTFASFA (SEQ ID NO: 83); GFTLGYYA (SEQ ID NO: 84); GFTFEDYA (SEQ ID NO: 85); GFTFEDYA (SEQ ID NO: 86); GNFLRFNA (SEQ ID NO: 87); GSFSSIDT (SEQ ID NO: 88); EQNFSADY (SEQ ID NO: 89); EQNFSTDD (SEQ ID NO: 90); RSIFSINA (SEQ ID NO: 91) GSIFSINP (SEQ ID NO: 92); GRPIDTYA (SEQ ID NO: 93); GRTISTYA (SEQ ID NO: 94); GRPLECYA (SEQ ID NO: 95); GRRLEGYT (SEQ ID NO: 96); GPRLEGYT (SEQ ID NO: 97); GRTFNLYA (SEQ ID NO: 98); GVTFSNYV (SEQ ID NO: 99); GRTVSNYA (SEQ ID NO: 100); GRTFSNYA (SEQ ID NO:101); GRTFSRYA (SEQ ID NO: 102); and GFTFTAYD (SEQ ID NO: 544); or an amino acid sequence having up to two amino acid substitutions (e.g., conservative amino acid substitutions) relative to any one of SEQ ID NOs: 61-102 or 544; and (b) a CDR2 comprising the amino acid sequence selected from the group consisting of: ITARGDWT (SEQ ID NO: 103); MWWSGGSP (SEQ ID NO: 104); ITSGGST (SEQ ID NO: 105); MWWSGGSP (SEQ ID NO: 106); SVWTDGKP (SEQ ID NO: 107); ISWSGKTT (SEQ ID NO: 108); ISWSGGST (SEQ ID NO: 109); IKPESGIT (SEQ ID NO: 110); ISTGGDTTDT (SEQ ID NO: 111); ITWSGGST (SEQ ID NO: 112); INTDGDST (SEQ ID NO: 113); VVGSDGRD (SEQ ID NO: 114); IDTSGGHV (SEQ ID NO: 115); ISRSGEKT (SEQ ID NO: 116); IDARGVNT (SEQ ID NO: 117); ISLNGGNT (SEQ ID NO: 118); AVGSDGST (SEQ ID NO: 119); VSWTGDNT (SEQ ID NO: 120); VNWRGSST (SEQ ID NO: 121); INANSDTT (SEQ ID NO: 122); ISWNGGST (SEQ ID NO: 123); ISRT (SEQ ID NO: 124); ISWSGGFT (SEQ ID NO: 125); IASSDGST (SEQ ID NO: 126); ISGRDGST (SEQ ID NO: 127); TSKNDRMP (SEQ ID NO: 128); ISSGGRT (SEQ ID NO: 129); INRGGDT (SEQ ID NO: 130); ISNSGRT (SEQ ID NO: 131); Attorney Docket No.: 45817-0158WO1 ITNSGTT (SEQ ID NO: 132); ITNGGTT (SEQ ID NO: 133); FTSGGTT (SEQ ID NO: 134); VSWAGVYT (SEQ ID NO: 135); VSWAGSYT (SEQ ID NO: 136); ISGNGVNT (SEQ ID NO: 137); ITRNRVNT (SEQ ID NO: 138); ISRNRVNT (SEQ ID NO: 139); INRSGTP (SEQ ID NO: 140); ITRSGTST (SEQ ID NO: 141); and ISSDSGDT (SEQ ID NO: 545); or an amino acid sequence having up to two amino acid substitutions (e.g., conservative amino acid substitutions) relative to any one of SEQ ID NOs: 103-141 or 545; and (c) a CDR3 comprising the amino acid sequence selected from the group consisting of: VRDLLGRDDY (SEQ ID NO: 142); AATWVGTSEYRH (SEQ ID NO: 143); AIGYYSGSYYWERSGDY (SEQ ID NO: 144); AATWVGTSEYQH (SEQ ID NO: 145); QALTGGSWILDY (SEQ ID NO: 146); FMNVWADTSDSSADAY (SEQ ID NO: 147); FGDIIRSGERSDYEY (SEQ ID NO: 148); VREDYDSAYVGDY (SEQ ID NO: 149); VALNLWGTDLEHDY (SEQ ID NO: 150); AAQFVEVEILVRSYEY (SEQ ID NO: 151); AAIVTRSDGHQYDY (SEQ ID NO: 152); AATNYYSDYLDHLSRGY (SEQ ID NO: 153); ARVENTWESIY (SEQ ID NO: 154); GAWNFVKNDAY (SEQ ID NO: 155); GGWEFAT (SEQ ID NO: 156); ATLGFAS (SEQ ID NO: 157); VKLDSGAWSLAE (SEQ ID NO: 158); AAWNWGHHEYTY (SEQ ID NO: 159); ARWSWDIGADFGS (SEQ ID NO: 160); VSDSYIGGLYATYVY (SEQ ID NO: 161); AAPYGSSQNLEYDY (SEQ ID NO: 162); VPVSHSDSVCGSPYMDY (SEQ ID NO: 163); AADNAADRCSLSIYNYNL (SEQ ID NO: 164); GEWGGFSL (SEQ ID NO: 165); VAPCFWFDTVIAGTDPRYDY (SEQ ID NO: 166); AAVRGPIVSMDPDLCRPVEFDY (SEQ ID NO: 167); AATNGPAITLFPCHINYWLYDN (SEQ ID NO: 168); ASADWRSPTPFPCGVSRSLYDH (SEQ ID NO: 169); WSAPDY (SEQ ID NO: 170); AGSFTLATGDDFGS (SEQ ID NO: 171); VAEPFSFRRRA (SEQ ID NO: 172); GESTTGWAECDFGC (SEQ ID NO: 173); NANSRYGVGWYNY (SEQ ID NO: 174); NVRGGHCDPRYWREY (SEQ ID NO: 175); NVRGGHYDPRYWREY (SEQ ID NO: 176); AATKLPWNTIVMVQRSYCDY (SEQ ID NO: 177); AATKLPWNTSVMVKRSVYDY (SEQ ID NO: 178); AATTVPVINLEISHMTY (SEQ ID NO: 179); AATNLPGITLLMSHMNYCDY (SEQ ID NO: 180); Attorney Docket No.: 45817-0158WO1 AATTVPVINLQVSHINY (SEQ ID NO: 181); AAKVFPMATLDDDVYDY (SEQ ID NO: 182); AAESHRRNTIVIVTTPDEYDY (SEQ ID NO: 183); AADPSSGYNLFARTVVAFARYDY (SEQ ID NO: 184); AAHPAGAHGGLIYKN (SEQ ID NO: 185); AADPTGAHVGAIYKN (SEQ ID NO: 186); AADLRGSSWYFDGVDY (SEQ ID NO: 187); AADLRGSSWYFDGMDY (SEQ ID NO: 188); AAHEAQYSSRWSGTEKGYDY (SEQ ID NO: 189); and ARLGAYSTTYDY (SEQ ID NO: 546); or an amino acid sequence having up to two amino acid substitutions (e.g., conservative amino acid substitutions) relative to any one of SEQ ID NOs: 142-188 or 546. Examples of anti-BCMA CDRs from VHH domains suitable for incorporation in the disclosed bispecific engagers are shown in Table 3 below. Table 3 – Examples of anti-BCMA CDRs Antibody CDR1 CDR2 CDR3 VHH D

Attorney Docket No.: 45817-0158WO1 BCMA RID YT D RD AAT YY DYLDHL R Y EQ Q Y N H

Attorney Docket No.: 45817-0158WO1 BCMA E F ADY I RT AEPF FRRRA

Attorney Docket No.: 45817-0158WO1 BCMA RTF RYA ITR T T AAHEA Y R TEK YDY

Other examples of anti-BCMA CDRs from VHH domains suitable for incorporation in the disclosed bispecific engagers are shown in Table B below. Table B. Exemplary CDR definitions for anti-BCMA VHHs IMGT CDR Definition Clone Name VHH-CDR1 VHH-CDR2 VHH-CDR3 GSIFAYHV ITSGGST AIGYYSGSYYWERSGDY BCMA-VHH3 (SEQ ID NO:63) (SEQ ID NO:105) (SEQ ID NO:144) GRTLNNYV MWWSGGSP AATWVGTSEYRH BCMA-VHH2 (SEQ ID NO:62) (SEQ ID NO:104) (SEQ ID NO:143) Kabat CDR Definition Clone Name VHH-CDR1 VHH-CDR2 VHH-CDR3 YHVMG LITSGGSTNYTDSVKG GYYSGSYYWERSGDY BCMA-VHH3 (SEQ ID NO:600) (SEQ ID NO:601) (SEQ ID NO:602) NYVVA TWVGTSEYRH TMWWSGGSPWYSDNVKG BCMA-VHH2 (SEQ ID NO:603) (SEQ ID NO:604) (SEQ ID NO:605) Enhanced Chothia CDR Definition Clone Name VHH-CDR1 VHH-CDR2 VHH-CDR3 GSIFAYHVMG LITSGGSTN GYYSGSYYWERSGDY BCMA-VHH3 (SEQ ID NO:606) (SEQ ID NO:607) (SEQ ID NO:608) Attorney Docket No.: 45817-0158WO1 GRTLNNYVVA TMWWSGGSPW TWVGTSEYRH BCMA-VHH2 (SEQ ID NO:609) (SEQ ID NO:610) (SEQ ID NO:611) Contact CDR Definition Clone Name VHH-CDR1 VHH-CDR2 VHH-CDR3 AYHVMG LVALITSGGSTN AIGYYSGSYYWERSGD BCMA-VHH3 (SEQ ID NO:612) (SEQ ID NO:613) (SEQ ID NO:614) NNYVVA FVATMWWSGGSPW AATWVGTSEYR BCMA-VHH2 (SEQ ID NO:615) (SEQ ID NO:616) (SEQ ID NO:617) Chothia CDR Definition Clone Name VHH-CDR1 VHH-CDR2 VHH-CDR3 GSIFAYH TSGGS GYYSGSYYWERSGDY BCMA-VHH3 (SEQ ID NO:618) (SEQ ID NO: 619) (SEQ ID NO:620) GRTLNNY WWSGGS TWVGTSEYRH BCMA-VHH2 (SEQ ID NO:621) (SEQ ID NO:622) (SEQ ID NO:623) In some instances, a bispecific engager of this disclosure comprises the three CDRs of any one VHH disclosed in Table 3 or Table B. Examples of anti-BCMA VHH domains suitable for incorporation in the disclosed bispecific engagers are shown in Table 4 below. Table 4 – Examples of anti-BCMA VHH domains Antibody Amino acid sequence SEQ ID VHH NO BCMA VHH1 AV LVDSGGGLVTAGDSLTLSCVASGRTLSPYTAGWFR APGRE 253

Attorney Docket No.: 45817-0158WO1 BCMAVHH2 HV LVESGGGLV TGGSLRLSCAASGRTLNNYVVAWFR APGKE 254

Attorney Docket No.: 45817-0158WO1 BCMA VKLEESGGGLV AGGSLRLSCAASGHTFSNSAMGWIR APGKE 268

Attorney Docket No.: 45817-0158WO1 BCMA V LVESGGGLV AGGSLRLSCVVSGSFSSIDTVDWYR APGK 282

Attorney Docket No.: 45817-0158WO1 BCMA V LVESGGGLV AGATLRLSCAASGVTFSNYVMGWFR APGK 296

Attorney Docket No.: 45817-0158WO1 BCMA V LVESGGGLV PGGSLRLSCAASGSIFAYHVMGWYR APGK 309

Attorney Docket No.: 45817-0158WO1 BCMA EV LVESGGGVV PGGSLRLSCAASGSIFAYHVMGWYR APGK 323

Attorney Docket No.: 45817-0158WO1 BCMA V LVESGGGLV PGGSLRLSCAASGRTLNNYVVAWFR APGKE 629

Attorney Docket No.: 45817-0158WO1 BCMA HV LVESGGGLV PGGSLRLSCAASGRTLNNYVVAWFR APGKE 643

An additional exemplary anti-BCMA (“BCMA-VHH102”) may comprise a CDR1 comprising an amino acid sequence of GFTFTAYD (SEQ ID NO: 544), a CDR2, comprising an amino acid sequence of ISSDSGDT (SEQ ID NO: 545), and a CDR3 comprising an amino acid sequence of ARLGAYSTTYDY (SEQ ID NO: 546). The variable heavy sequence of BCMA-VHH102 comprises an amino acid sequence of EVQLVESGGGLVQPGGSLRLSCAASGFTFTAYDMGWVRQAPGKGPEWVSLI SSDSGDTWYDDSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCARLGAY STTYDYWGQGTLVTVSS (SEQ ID NO: 547). In some embodiments, the anti-BCMA antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of any one of SEQ ID NOs: 253- 332, 547 or 624-644, wherein the three VHH CDRs are not altered. In some embodiments, the anti-BCMA antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of any one of SEQ ID NOs: 253-332, 547 or 624-644. In some embodiments, the anti-BCMA antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of any one of SEQ ID NOs: 253-332, 547 or 624-644. In some embodiments, the anti-BCMA antibody or binding domain (e.g., VHH) may comprise the amino acid sequence of any one of SEQ ID NOs: 253-332, 547 or 624-644. Attorney Docket No.: 45817-0158WO1 Example of CDRs for anti-GPRC5D antibodies or binding domains include: (a) a CDR1 comprising the amino acid sequence selected from the group consisting of GRTVSSYA (SEQ ID NO: 190); GRTASAYV (SEQ ID NO: 191); GIIFSASN (SEQ ID NO: 192); GGFGMMYS (SEQ ID NO: 193); RIRFSINV (SEQ ID NO: 194); and SERTFRSYT (SEQ ID NO: 195); or an amino acid sequence having up to two amino acid substitutions (e.g., conservative amino acid substitutions) relative to any one of SEQ ID NOs: 190-195; and (b) a CDR2 comprising the amino acid sequence selected from the group consisting of: ISWSGRST (SEQ ID NO: 196); ISGGA (SEQ ID NO: 197); VTGGGSI (SEQ ID NO: 198); RTIDGST (SEQ ID NO: 199); IAAGGTT (SEQ ID NO: 200); and ISWSHSST (SEQ ID NO: 201); or an amino acid sequence having up to two amino acid substitutions (e.g., conservative amino acid substitutions) relative to any one of SEQ ID NOs: 196-201; and (c) a CDR3 comprising the amino acid sequence selected from the group consisting of: ATSRAVIPGRDWNYYEY (SEQ ID NO: 202); AAERGMRRLTESYQYDY (SEQ ID NO: 203); NARRSYSH (SEQ ID NO: 204); NAKPLNGRLTQY (SEQ ID NO: 205); NAVLSTLVLPSTY (SEQ ID NO: 206); and AADLRLLPEEYDY (SEQ ID NO: 207); or an amino acid sequence having up to two amino acid substitutions (e.g., conservative amino acid substitutions) relative to any one of SEQ ID NOs: 202-207. Examples of anti-GPRC5D CDRs from VHH domains suitable for incorporation in the disclosed bispecific engagers are shown in Table 5 below. Table 5 – Examples of Anti-GPRc5D CDRs Antibody CDR1 CDR2 CDR3 VHH GPRC5D- GRTVSSYA ISWSGRST ATSRAVIPGRDWNYYEY

Attorney Docket No.: 45817-0158WO1 GPRC5D F MMY RTID T AKPL RLT Y

Other examples of anti-GPRC5D CDRs from VHH domains suitable for incorporation in the disclosed bispecific engagers are shown in Table C below. Table C. Exemplary CDR definitions for anti-GPRC5D VHHs IMGT CDR Definition Clone Name VHH-CDR1 VHH-CDR2 VHH-CDR3 GRTVSSYA ISWSGRST ATSRAVIPGRDWNYYEY GPRC5D-VHH1 (SEQ ID NO:190) (SEQ ID NO:196) (SEQ ID NO: 202) GIIFSASN VTGGGSI NARRSYSH GPRC5D-VHH3 (SEQ ID NO: 192) (SEQ ID NO: 198) (SEQ ID NO: 204) GRTASAYV ISGGA AAERGMRRLTESYQYDY GPRC5D-VHH2 (SEQ ID NO: 191) (SEQ ID NO: 197) (SEQ ID NO: 203) GGFGMMYS RTIDGST NAKPLNGRLTQY GPRC5D-VHH4 (SEQ ID NO: 193) (SEQ ID NO: 199) (SEQ ID NO: 205) Kabat CDR Definition Clone Name VHH-CDR1 VHH-CDR2 VHH-CDR3 SYAMG AISWSGRSTYYADSMKG SRAVIPGRDWNYYEY GPRC5D-VHH1 (SEQ ID NO:645) (SEQ ID NO:646) (SEQ ID NO:647) ASNLA GVTGGGSINYADSVKG RRSYSH GPRC5D-VHH3 (SEQ ID NO:648) (SEQ ID NO:649) (SEQ ID NO:650) AYVMG GISGGAYYADSVKG ERGMRRLTESYQYDY GPRC5D-VHH2 (SEQ ID NO:651) (SEQ ID NO:652) (SEQ ID NO:653) Attorney Docket No.: 45817-0158WO1 MYSMG ARTIDGSTNYADSVKD KPLNGRLTQY GPRC5D-VHH4 (SEQ ID NO:654) (SEQ ID NO:655) (SEQ ID NO:656) Enhanced Chothia CDR Definition Clone Name VHH-CDR1 VHH-CDR2 VHH-CDR3 GRTVSSYAMG AISWSGRSTY SRAVIPGRDWNYYEY GPRC5D-VHH1 (SEQ ID NO:657) (SEQ ID NO:658) (SEQ ID NO:659) GIIFSASNLA GVTGGGSIN RRSYSH GPRC5D-VHH3 (SEQ ID NO:660) (SEQ ID NO:661) (SEQ ID NO:662) GRTASAYVMG GISGGAY ERGMRRLTESYQYDY GPRC5D-VHH2 (SEQ ID NO:663) (SEQ ID NO:664) (SEQ ID NO:665) GGFGMMYSMG ARTIDGSTN KPLNGRLTQY GPRC5D-VHH4 (SEQ ID NO:666) (SEQ ID NO:667) (SEQ ID NO:668) Contact CDR Definition Clone Name VHH-CDR1 VHH-CDR2 VHH-CDR3 SSYAMG FVSAISWSGRSTY ATSRAVIPGRDWNYYE GPRC5D-VHH1 (SEQ ID NO: 669) (SEQ ID NO: 670) (SEQ ID NO:671) SASNLA LVAGVTGGGSIN NARRSYS GPRC5D-VHH3 (SEQ ID NO: 672) (SEQ ID NO: 673) (SEQ ID NO: 674) SAYVMG FVAGISGGAY AAERGMRRLTESYQYD GPRC5D-VHH2 (SEQ ID NO: 675) (SEQ ID NO: 676) (SEQ ID NO:677) MMYSMG LVAARTIDGSTN NAKPLNGRLTQ GPRC5D-VHH4 (SEQ ID NO: 678) (SEQ ID NO: 679) (SEQ ID NO: 680) Chothia CDR Definition Attorney Docket No.: 45817-0158WO1 Clone Name VHH-CDR1 VHH-CDR2 VHH-CDR3 GRTVSSY SWSGRS SRAVIPGRDWNYYEY GPRC5D-VHH1 (SEQ ID NO: 681) (SEQ ID NO: 682) (SEQ ID NO: 683) GIIFSAS TGGGS RRSYSH GPRC5D-VHH3 (SEQ ID NO: 684) (SEQ ID NO: 685) (SEQ ID NO: 686) GRTASAY ERGMRRLTESYQYDY ISGG GPRC5D-VHH2 (SEQ ID NO: 687) (SEQ ID NO: 688) (SEQ ID NO:689) GGFGMMY TIDGS KPLNGRLTQY GPRC5D-VHH4 (SEQ ID NO: 690) (SEQ ID NO: 691) (SEQ ID NO: 692) In some instances, a bispecific engager of this disclosure comprises the three CDRs of any one VHH disclosed in Table 5 or Table C. Examples of anti-GPRC5D VHH domains suitable for incorporation in the disclosed bispecific engagers are shown in Table 6 below. Table 6 – Examples of Anti-GPRc5D VHH Antibody VHH Amino acid sequence SEQ ID NO:

Attorney Docket No.: 45817-0158WO1 YL MNSLKPDDTAVYYCNAVLSTLVLPSTYWG GT VT

Attorney Docket No.: 45817-0158WO1 GPRC5DVHH17 VKLVESGGGLV PGGSLRLSCVGSGIIFSASNLAWYR A 349

Attorney Docket No.: 45817-0158WO1 L MNSLRAEDTAVYYCAAERGMRRLTESY YDYWG G

Attorney Docket No.: 45817-0158WO1 VYL MDSLRAEDTAVYYCNAKPLNGRLT YWG GTLVT

n some em o ments, t e ant - ant o y or n ng oma n (e.g., VHH) may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of any one of SEQ ID NOs: 333- 376, wherein the three VHH CDRs are not altered. In some embodiments, the anti- GPRC5D antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of any one of SEQ ID NOs: 333-376. In some embodiments, the anti-GPRC5D antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of any one of SEQ ID NOs: 333-376. In some embodiments, the anti- GPRC5D antibody or binding domain (e.g., VHH) may comprise the amino acid sequence of any one of SEQ ID NOs: 333-376. Examples of CDRs for anti-FcRH5 antibodies or binding domains include: (a) a CDR1 comprising the amino acid sequence selected from the group consisting of GITVSRND (SEQ ID NO: 208); VHIISHYS (SEQ ID NO: 209); GHTLSTYA (SEQ ID NO: 210); GRTFSTYA (SEQ ID NO: 211); GSHFSIAT (SEQ ID NO: 212); GRTYNNYA (SEQ ID NO: 213); GRTFSTYG (SEQ ID NO: 214); RSSFSNNA (SEQ ID NO: 215); GRTSSRAA (SEQ ID NO: 216); and GSIFSINA Attorney Docket No.: 45817-0158WO1 (SEQ ID NO: 217)or an amino acid sequence having up to two amino acid substitutions (e.g., conservative amino acid substitutions) relative to any one of SEQ ID NOs: 208-217; and (b) a CDR2 comprising the amino acid sequence selected from the group consisting of: IMNRVGST (SEQ ID NO: 218); IPVSGRVP (SEQ ID NO: 219); IARDGGAT (SEQ ID NO: 220); IDTTGSAS (SEQ ID NO: 221); LSSSGRP (SEQ ID NO: 222); ISRSGGMT (SEQ ID NO: 223); ISRSGGAT (SEQ ID NO: 224); ITKGGVT (SEQ ID NO: 225); ISWSGGTT (SEQ ID NO: 226); and ITSGGST (SEQ ID NO: 227); or an amino acid sequence having up to two amino acid substitutions (e.g., conservative amino acid substitutions) relative to any one of SEQ ID NOs: 218- 227; and (c) a CDR3 comprising the amino acid sequence selected from the group consisting of: NALNTVITWP (SEQ ID NO: 228); AAYPRKGLEGNEYEY (SEQ ID NO: 229); AASSMFSTAKRDYSY (SEQ ID NO: 230); AAARRYSTAPGDYDY (SEQ ID NO: 231); KANLKRFFIEERYRDY (SEQ ID NO: 232); AAYVGGFSTARRDYSY (SEQ ID NO: 233); AGTRRAFSTGLRDYDY (SEQ ID NO: 234); NTIPFRSA (SEQ ID NO: 235); AAARIFTTARNDYDH (SEQ ID NO: 236); and NALGGFVPSYG (SEQ ID NO: 237), or an amino acid sequence having up to two amino acid substitutions (e.g., conservative amino acid substitutions) relative to any one of SEQ ID NOs: 228-237. Examples of anti-FcRH5 CDRs from VHH domains suitable for incorporation in the disclosed bispecific engagers are shown in Table 7 below. Table 7 – Examples of Anti-FcRH5 CDRs Antibody VHH CDR1 CDR2 CDR3 F^cRH5-VHH1 _GITVSRND IMNRVGST NALNTVITWP (SEQ ID NO: 208) (SEQ ID NO: 218) (SEQ ID NO: 228) Y Y Y

Attorney Docket No.: 45817-0158WO1 F^{RH5 HH5} _{HF IAT} L RP KA LKRFFIEERYRDY S D H

Other examples of anti-FcRH5 CDRs from VHH domains suitable for incorporation in the disclosed bispecific engagers are shown in Table D below. Table D. Exemplary CDR definitions for anti-FcRH5 VHHs IMGT CDR Definition Clone Name VHH-CDR1 VHH-CDR2 VHH-CDR3 GITVSRND IMNRVGST NALNTVITWP FcRH5-VHH1 (SEQ ID NO:208) (SEQ ID NO:218) (SEQ ID NO:228) Kabat CDR Definition Clone Name VHH-CDR1 VHH-CDR2 VHH-CDR3 RNDMG IMNRVGSTDTADSVKG LNTVITWP FcRH5-VHH1 (SEQ ID NO: 693) (SEQ ID NO:694) (SEQ ID NO:695) Enhanced Chothia CDR Definition Clone Name VHH-CDR1 VHH-CDR2 VHH-CDR3 GITVSRNDMG LNTVITWP IMNRVGSTD FcRH5-VHH1 (SEQ ID NO:696) (SEQ ID NO:697) (SEQ ID NO:698) Attorney Docket No.: 45817-0158WO1 Contact CDR Definition Clone Name VHH-CDR1 VHH-CDR2 VHH-CDR3 SRNDMG LVAIMNRVGSTD NALNTVITW FcRH5-VHH1 (SEQ ID NO:699) (SEQ ID NO:700) (SEQ ID NO:701) Chothia CDR Definition Clone Name VHH-CDR1 VHH-CDR2 VHH-CDR3 GITVSRN NRVGS LNTVITWP FcRH5-VHH1 (SEQ ID NO:702) (SEQ ID NO:703) (SEQ ID NO:704)

Atty. Docket No.45817-0158WO1 In some instances, a bispecific engager of this disclosure comprises the three CDRs of any one VHH disclosed in Table 7 or Table D. Examples of anti-FcRH5 VHH domains suitable for incorporation in the disclosed bispecific engagers are shown in Table 8 below. Table 8 – Examples of Anti-FcRH5 VHH Antibody VHH Amino acid sequence SEQ ID

Atty. Docket No.45817-0158WO1 F^{RH5 HH11} _{L E L A LRL AA RTY YAM FR} 387

Atty. Docket No.45817-0158WO1 F^{RH5 HH22} _{L E L P LRL AA RTF TY M FR A} 709

Atty. Docket No.45817-0158WO1 M LRAEDTA YY AAARIFTTAR DYDH TL T

VHH) may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of any one of SEQ ID NOs: 377- 393 or 705-727, wherein the three VHH CDRs are not altered. In some embodiments, the anti-FcRH5 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of any one of SEQ ID NOs: 377-393 or 705-727. In some embodiments, the anti-FcRH5 antibody or binding domain (e.g., VHH) may comprise an amino acid sequence that is at least Atty. Docket No.45817-0158WO1 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of any one of SEQ ID NOs: 377-393 or 705-727. In some embodiments, the anti-FcRH5 antibody or binding domain (e.g., VHH) may comprise the amino acid sequence of any one of SEQ ID NOs: 377-393 or 705-727. In some embodiments, a VHH incorporated into a bispecific engager disclosed herein may be a murine-specific VHH, e.g., the VHH specifically binds a murine antigen. In some embodiments, a VHH incorporated into a bispecific engager disclosed herein may be a rat-specific VHH, e.g., the VHH specifically binds a rat antigen. In some embodiments, a VHH incorporated into a bispecific engager disclosed herein may be a llama-specific VHH, e.g., the VHH specifically binds a llama antigen. In some embodiments, a VHH incorporated into a bispecific engager disclosed herein may be a human-specific VHH, e.g., the VHH specifically binds the human antigen. In some embodiments, a VHH incorporated into a bispecific engager disclosed herein may be human-specific even if the VHH is not human or humanized. In some aspects, tolerable variations in the binding domains will be known to those of skill in the art, while maintaining binding to the target antigen. Given the disclosure provides exemplary bispecific engager therapeutics and describes the delivery of such therapeutics (e.g., via mRNA), those of ordinary skill in the art will understand that other VHH domains which bind to B cells or NK cells can be included in the disclosed bispecific engager constructs. Indeed, there are numerous binding domains of which those of ordinary skill would be aware that can be adapted into the disclosed bispecific engager format, such as the binding doamins disclosed in Han et al., Leukemia, 2021, 10:3002-3006; Zhoa et al., J. Hematol. Oncol., 2018;11:141; US 2023/0058669; WO 2022/246004; US 11242376; US 2022/0265710; WO 2010/104949; EP3689908; WO 2023/034740; WO 2023/034741; and WO 2021/113853, all of which are herein incorporated by reference. Linkers Atty. Docket No.45817-0158WO1 The disclosed bispecific engagers may contain a linker, which connects two domains of the bispecific engager. In some embodiments, a linker may directly or indirectly connect the two binding domains of a bispecific engager. In some embodiments, the linker may connect a binding domain with an Fc domain. In some embodiments, a linker may connect the C-terminus of an Fc domain with a binding domain. In some embodiments, a linker may connect the N-terminus of an Fc domain with a binding domain. In some embodiments, a bispecific engager may comprise two separate linkers, each of which connect different domains of the bispecific engager. The linkers used in the disclosed bispecific engagers can be encoded in vivo via expression of an mRNA. The linker may be flexible. The linker may comprise 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 or more peptides. A link may comprise guanine (G), serine (S), or various repeats or combinations thereof. Suitable linkers can include, but are not limited to GGGGS (SEQ ID NO: 448; i.e., a “G4S linker”), GSGG (SEQ ID NO: 449), GSGGSG (SEQ ID NO: 450), GSGGGG (SEQ ID NO: 451), GSG, GSGGSGGSGGSGGG (SEQ ID NO: 453), GGGSGGGSGG (SEQ ID NO: 454), GGGG (SEQ ID NO: 455), GGG, SGG, GGGSGGSG (SEQ ID NO: 458), GGSGGSGGGS (SEQ ID NO: 459), and GGGGGS (SEQ ID NO: 460). In some embodiments, the linker comprises GGGGS (SEQ ID NO: 448). Signal Sequences A signal sequence (sometimes referred to as signal peptide, targeting signal, localization signal, localization sequence, leader sequence, or leader peptide) is a short polypeptide sequence (usually 10-30 amino acids long) that is present at the N- terminus of a newly synthesized protein. The signal sequence can help direct a newly synthesized protein toward a secretory pathway. Thus, because the disclosed bispecific engagers are excreted from a cell upon expression in vivo, the disclosed bispecific engagers will generally comprise a signal sequence, which is ultimately cleaved off the mature protein during protein processing and translocation. Atty. Docket No.45817-0158WO1 Various signal sequences that can guide a protein toward a secretion pathway are known in the art. For example, the signal peptide of a human light chain can be used to direct the disclosed bispecific engagers for secretion upon translation in vivo. Thus, in some embodiments, a human light chain signal sequence can be covalently linked to a humanized VHH (e.g., a VHH that binds to a molecule present on the surface of a B cell targeted for reduction or elimination, the CH2 and CH3 domains from an IgG4 molecule (S228P/F234A/L235A), a G4S linker (SEQ ID NO: 448), and a humanized VHH binding to CD16a). This construct orientation was found to be efficiently expressed in cells, and minimize the formation of aggregates, and exhibit target cell killing. In addition, multiple engagers of this format were found to express in the same cell without a reduction in the potency of cell killing. In some embodiments, the signal sequence is METPAQLLFLLLLWLPDTTG (SEQ ID NO: 447). Those of skill in the art will recognize that other leader sequences may be suitable as well. Examples of Bispecific Engager Amino Acid Sequences The foregoing binding domains can be combined in numerous possible ways to prepare a bispecific engager, and the format of a bispecific engager of the present disclosure is not particularly limited. Moreover, binding domains that specifically bind to B cell associated molecules other than CD38, BCMA, GPRc5D, and FcRh5 can be used for or incorporated into bispecific engagers for the purposes of the present disclosure, and binding domains that specifically bind to NK cell molecules other than CD16a can be used for or incorporated into bispecific engagers for the purposes of the present disclosure (e.g., binding domains that bind to B3GAT1 (CD57), CCR7 (CD197), CD16, CD16a, CD16b, CD2 CD226, CD244, CD27, CD3, CD300A, CD34, CD58, CD59, CD69, CSF2, CX3CR1, CXCR1 (CD128), CXCR3 (CD183), CXCR4, EOMES, GZMB, ICAM1 (CD54), IFNG, IL-15R, IL-1R, IL22, IL-2RB (CD122), IL- 7R (CD127), ITGA1 (CD49a), ITGA2 (CD49b), ITGAL (CD11a), ITGAM (CD11b), ITGB2 (CD18), KIR, KIR2DL1, KIR2DL2, KIT (CD117), KLRB1C, KLRC1, KLRC2, KLRD1 (CD94), KLRF1, KLRG1, KLRK1, LILRB1, KLRA4, KLRA8, Atty. Docket No.45817-0158WO1 MICA/BNCAM1 (CD56), NK2D, NKP46 (NCR1, CD335), NCR2, NCR3 (CD337), PRF1, SELL (CD62L), SIGLEC7, SLAMF6, SPN, TBX21, or TNFa). Accordingly, the present disclosure encompasses a variety of bispecific engagers of various formats and that bind to various targets. Nevertheless, specific examples of bispecific engagers that can be used alone or in combination are disclosed in Table 9. These bispecific engagers (as well as any other bispecific engagers within the scope of the disclosure) can be administered to a subject alone or in combination. Additionally, these bispecific engagers (as well as any other bispecific engagers within the scope of the disclosure) can be used by administering to a subject an mRNA that encodes the bispecific engager. For example, bispecific engager (including, but not limited to those disclosed in Table 9) can be used by administering to a subject an mRNA encoding the bispecific engager. Two different bispecific engagers (including, but not limited to those disclosed in Table 9) can be used by administering to a subject (i) a single mRNA that encodes both bispecific engagers, or (ii) two mRNA, each one encoding a different bispecific engager. The same arrangement could be used for 3, 4, or 5 or more different bispecific engagers. Table 9 – Examples of Bispecific Engagers Bispecific Amino acid sequence SEQ ID Engager NO

Atty. Docket No.45817-0158WO1 VSLTCLVKGFYPSDIAVEWESNG PENNYKTTPPVLDSDGSFFL

Atty. Docket No.45817-0158WO1 APGKEREFVAGISRSGGMTGYAESVKGRFTISRDNAKNMVFL M

Atty. Docket No.45817-0158WO1 EDTAVYYCAAGRGYGLLSISSNWYNYWG GTLVTVSSESKYGPP

Atty. Docket No.45817-0158WO1 KN VSLTCLVKGFYPSDIAVEWESNG PENNYKTTPPVLDSDGSF

Atty. Docket No.45817-0158WO1 APGKEREFVAAISRSGGATAYAASVKGRFTISRDNSKNTLYL MN

Atty. Docket No.45817-0158WO1 EDTAVYYCAAGRGYGLLSISSNWYNYWG GTLVTVSSESKYGPP

Atty. Docket No.45817-0158WO1 KN VSLTCLVKGFYPSDIAVEWESNG PENNYKTTPPVLDSDGSF

Atty. Docket No.45817-0158WO1 GK RELVAFITSGGSTNYADSVKGRFTISRDNSKNTLYL MNSLR

Atty. Docket No.45817-0158WO1 BE 38 ESKYGPPCPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVVV 431

Atty. Docket No.45817-0158WO1 LNGKEYKCKVSNKGLPSSIEKTISKAKG PREP VYTLPPS EEMT

Atty. Docket No.45817-0158WO1 BE46 EV LVESGGGLV PGGSLRLSCAASGRTDSIYAMGWFR APGKE 439

Atty. Docket No.45817-0158WO1 FLYSRLTVDKSRW EGNVFSCSVMHEALHNHYT KSLSLSLGKG

Atty. Docket No.45817-0158WO1 EDTAVYYCARLGAYSTTYDYWG GTLVTVSSDKTHTCPPCPAPE

In some embodiments, the bispecific engager may comprise an amino acid sequence that is at least 85% identical (e.g., at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of any one of SEQ ID NOs: 394-446. In some embodiments, the bispecific engager may comprise an amino acid sequence that is at least 90% identical (e.g., at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of any one of SEQ ID NOs: 394-446. In some embodiments, the bispecific engager may comprise an amino acid sequence that is at least 95% identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) to the amino acid sequence of any one of SEQ ID NOs: 394-446. In some embodiments, the bispecific engager may comprise the amino acid sequence of any one of SEQ ID NOs: 394-446. When a bispecific engager is expressed, it may initially include a signal sequence that is ultimately cleaved off to yield a mature protein. Accordingly, the signal sequence may be encoded as part of an open reading frame (ORF) encoding any bispecific engager disclosed here. As noted above, the signal sequence can be from a human light chain. One example of a suitable signal sequence is METPAQLLFLLLLWLPDTTG (SEQ ID NO: 447), and those of skill in the art will recognize that other leader sequences may be suitable as well. In some aspects, tolerable variations in the binding domain will be known to those of skill in the art, while maintaining binding to the target antigen. Atty. Docket No.45817-0158WO1 Affinity of VHH binding moieties and bispecific engagers of the disclosure VHH domains and bispecific engagers of the disclosure may have an affinity for a target B cell molecule or NK cell target molecule of, for example, from 1 nM to 100 nM (e.g., from 10 nM to 90 nM, from 20 nM to 80 nM, from 30 nM to 70 nM, from 40 nM to 60 nM, or about 50 nM). In some embodiments, VHH domains and bispecific engagers of the disclosure have an affinity for a target B cell molecule or NK cell target molecule of from about 1 nM to about 100 nM. In some embodiments, VHH domains and bispecific engagers of the disclosure have an affinity for a target B cell molecule or NK cell target molecule of from about 1 nM to about 90 nM. In some embodiments, VHH domains and bispecific engagers of the disclosure have an affinity for a target B cell molecule or NK cell target molecule of from about 1 nM to about 80 nM. In some embodiments, VHH domains and bispecific engagers of the disclosure have an affinity for a target B cell molecule or NK cell target molecule of from about 1 nM to 60 nM. In some embodiments, VHH domains and bispecific engagers of the disclosure have an affinity for a target B cell molecule or NK cell target molecule of from about 1 nM to 40 nM. In some embodiments, VHH domains and bispecific engagers of the disclosure have an affinity for a target B cell molecule or NK cell target molecule of from about 1 nM to 20 nM. The specific binding of VHH domains and bispecific engagers described herein to a target B cell molecule or NK cell target molecule can be determined by any of a variety of established methods. The affinity can be represented quantitatively by various measurements, including the equilibrium constant (K_D) of the VHH domain- and bispecific engager-antigen complex dissociation. The equilibrium constant, K_D, which describes the interaction of the antigen with a binding domain described herein is the chemical equilibrium constant for the dissociation reaction of a bound complex into solvent-separated antigen and binding domain molecules that do not interact with one another. VHH domains and bispecific engagers described herein include those that specifically bind to target B cell molecules or NK cell target molecules with a KD Atty. Docket No.45817-0158WO1 value of less than 100 nM (e.g., less than 95 nM, 90 nM, 85 nM, 80 nM, 75 nM, 70 nM, 65 nM, 60 nM, 55 nM, 50 nM, 45 nM, 40 nM, 35 nM, 30 nM, 25 nM, 20 nM, 15 nM, 10 nM, 5 nM, 4 nM, 3 nM, 2 nM, or 1 nM). In some embodiments, the VHH domains and bispecific engagers described herein specifically bind to a target B cell molecule or NK cell target molecules with a KD value of less than 10 nM (e.g., less than 9 nM, 8 nM, 7 nM, 6 nM, 5 nM, 4 nM, 3 nM, 2 nM, or 1 nM). VHH domains and bispecific engagers described herein can also be characterized by a variety of in vitro binding assays. Examples of experiments that can be used to determine the KD a binding domain include, e.g., surface plasmon resonance, isothermal titration calorimetry, fluorescence anisotropy, ELISA-based assays, gene expression assays, and protein expression assays, among others. ELISA represents a particularly useful method for analyzing binding domain activity, as such assays typically require minimal concentrations of binding domains. A common signal that is analyzed in a typical ELISA assay is luminescence, which is typically the result of the activity of a peroxidase conjugated to a secondary antibody that specifically binds a primary antibody. In a direct ELISA experiment, this binding can be quantified, e.g., by analyzing the luminescence that occurs upon incubation of an HRP substrate (e.g., 2,2’-azino-di-3- ethylbenzthiazoline sulfonate) with an antigen- antibody, antigen-antigen-binding fragment, or antigen- binding domain complex bound to a HRP-conjugated secondary antibody. Kinetic properties of antibodies or binding domains It is also possible to quantitatively characterize the kinetic association and dissociation of VHH domains and bispecific engagers described herein with a target B cell molecule or NK cell target molecule. This can be done, e.g., by monitoring the rate of binding domain-antigen complex formation according to established procedures. For example, one can use surface plasmon resonance (SPR) to determine the rate constants for the formation (kon) and dissociation (koff) of a binding domain- antigen complex. These data also enable calculation of the equilibrium constant of (KD) of the binding domain-antigen complex dissociation since the equilibrium Atty. Docket No.45817-0158WO1 constant of this unimolecular dissociation can be expressed as the ratio of the koff to k_on values. SPR is a technique that is particularly advantageous for determining kinetic and thermodynamic parameters of antigen-binding domain interactions since the experiment does not require that one component be modified by attachment of a chemical label. Rather, the antigen is typically immobilized on a solid metallic surface which is treated in pulses with solutions of increasing concentrations of binding domain. Binding domain-antigen binding induces distortion in the angle of reflection of incident light at the metallic surface, and this change in refractive index over time as binding domain is introduced to the system can be fit to established regression models in order to calculate the association and dissociation rate constants of a binding domain-antigen interaction. VHH domains and bispecific engagers described herein may exhibit high k_on and low koff values upon interaction with the target B cell molecule or NK cell target molecule. For example, VHH domains and bispecific engagers described herein may exhibit kon values in the presence of the target B cell molecule or NK cell target molecule of greater than 10⁴ M^-1s^-1 (e.g., 1.0 x 10⁴ M^-1s^-1, 1.5 x 10⁴ M^-1s^-1, 2.0 x 10⁴ M^-1s^-1, 2.5 x 10⁴ M^-1s^-1, 3.0 x 10⁴ M^-1s^-1, 3.5 x 10⁴ M^-1s^-1, 4.0 x 10⁴ M^-1s^-1, 4.5 x 10⁴ M- ¹s^-1, 5.0 x 10⁴ M^-1s^-1, 5.5 x 10⁴ M^-1s^-1, 6.0 x 10⁴ M^-1s^-1, 6.5 x 10⁴ M^-1s^-1, 7.0 x 10⁴ M^-1s- ¹, 7.5 x 10⁴ M^-1s^-1, 8.0 x 10⁴ M^-1s^-1, 8.5 x 10⁴ M^-1s^-1, 9.0 x 10⁴ M^-1s^-1, 9.5 x 10⁴ M^-1s^-1, 1.0 x 10⁵ M^-1s^-1, 1.5 x 10⁵ M^-1s^-1, 2.0 x 10⁵ M^-1s^-1, 2.5 x 10⁵ M^-1s^-1, 3.0 x 10⁵ M^-1s^-1, 3.5 x 10⁵ M^-1s^-1, 4.0 x 10⁵ M^-1s^-1, 4.5 x 10⁵ M^-1s^-1, 5.0 x 10⁵ M^-1s^-1, 5.5 x 10⁵ M^-1s^-1, 6.0 x 10⁵ M^-1s^-1, 6.5 x 10⁵ M^-1s^-1, 7.0 x 10⁵ M^-1s^-1, 7.5 x 10⁵ M^-1s^-1, 8.0 x 10⁵ M^-1s^-1, 8.5 x 10⁵ M^-1s^-1, 9.0 x 10⁵ M^-1s^-1, 9.5 x 10⁵ M^-1s^-1, or 1.0 x 10⁶ M^-1s^-1). VHH domains and bispecific engagers described herein may exhibit low koff values when bound to the target B cell molecule or NK cell target molecule. For instance, VHH domains and bispecific engagers described herein may exhibit koff values of less than 10^-3 s^-1 when complexed to the target B cell molecule or NK cell target molecule (e.g., 1.0 x 10^-3 s- ¹, 9.5 x 10^-4 s^-1, 9.0 x 10^-4 s^-1, 8.5 x 10^-4 s^-1, 8.0 x 10 ^-4 s^-1, 7.5 x 10^-4 s^-1, 7.0 x 10^-4 s^-1, 6.5 x 10^-4 s^-1, 6.0 x 10^-4 s^-1, 5.5 x 10^-4 s^-1, 5.0 x 10^-4 s^-1, 4.5 x 10^-4 s^-1, 4.0 x 10^-4 s^-1, 3.5 x 10^-4 s^-1, 3.0 x 10^-4 s^-1, 2.5 x 10^-4 s^-1, 2.0 x 10^-4 s^-1, 1.5 x 10^-4 s^-1, 1.0 x 10^-4 s^-1, 9.5 x Atty. Docket No.45817-0158WO1 10^-5 s^-1, 9.0 x 10^-5 s^-1, 8.5 x 10^-5 s^-1, 8.0 x 10^-5 s^-1, 7.5 x 10^-5 s^-1, 7.0 x 10^-5 s^-1, 6.5 x 10^-5 s^-1, 6.0 x 10^-5 s^-1, 5.5 x 10^-5 s^-1, 5.0 x 10^-5 s^-1, 4.5 x 10^-5 s^-1, 4.0 x 10^-5 s^-1, 3.5 x 10^-5 s^-1, 3.0 x 10^-5 s^-1, 2.5 x 10^-5 s^-1, 2.0 x 10^-5 s^-1, 1.5 x 10^-5 s^-1, or 1.0 x 10^-5 s^-1). Methods for Humanization VHH domains described herein can be derived from, e.g., Llamas by immunization using methods known in the art. Such VHH preferably are humanized. As an example, one strategy that can be used to design humanized VHH domains described herein is set forth in Hanf et al. (Methods.65(1):68-76 (2014)). Briefly sequences of the CDRs of VH1 are annotated using the IMGT numbering scheme. Each VHH nucleotide sequence is generated and used to identify the nearest human germline VH sequences by searching for similar sequences with the NCBI IgBLAST program. Common J and D gene sequences were attached to the VH as the acceptor. Next the most similar human VH sequences are identified using BLASTp and used to choose the nearest framework sequences into which the VH1 CDR sequences are grafted replacing the human CDRs. As an example, Rosetta/Alpha fold are used to create the structural 3D homology model the of the appropriate CDRs that were grafted into the acceptor framework. The framework residues that were critical for huVH/VL interactions are back mutated to llama sequence canonical llama residues, also potentially structural defects due to mismatches at the graft interface can be fixed by mutating some framework residues to llama, or by mutating some residues on the CDRs’ backside to human or to a de novo designed sequence. CDR stabilizing or overall fold stabilizing sequences were then back-mutated to the corresponding llama sequence to maintain the biophysical properties and target binding affinity. Similarly, this strategy can also be used to produce primatized VHH domains, as one can substitute, for example, one or more, or all, of the CDRs of a primate antibody consensus sequence with, for example, one or more, or all, of the CDRs of a VHH domain of the disclosure. Consensus primate antibody sequences known in the art (see, e.g., U.S. Patent Nos. Atty. Docket No.45817-0158WO1 5,658,570; 5,681,722; and 5,693,780; the disclosures of each of which are incorporated herein by reference). Combination of Bispecific Engagers and Additional Therapeutics The disclosed bispecific engagers can be used or expressed in vivo as individual constructs (e.g., one nucleic acid encoding one bispecific engager is administered to a subject) or in mixtures of bispecific engagers (e.g., more than one nucleic acid, each expressing a different bispecific engager are administered to a subject), as discussed herein. Additionally, the nucleic acids expressing bispecific engagers may be combined with one or more additional agent known in the art to be useful in the treatment of a disorder associated with B cell dysfunction (e.g., cancer), such as a cytokine. Suitable cytokines include, for example, various interleukins (IL), (e.g., IL-15), interferons, and/or checkpoint inhibitors. Such additional art recognized therapeutics may be administered in protein form or as nucleic acid molecules (either in the same or separate delivery vehicles as the engagers). Cytokines are a category of small proteins important in cell signaling. Cytokines cannot cross the lipid bilayer of cells, but they have been shown to be involved in autocrine, paracrine and endocrine signaling as immunomodulating agents. Cytokines have also been used in certain cancer therapy regimen. For the purposes of the present disclosure, in some embodiments, a nucleic acid (e.g., mRNA) can be administered to a subject along with a cytokine (e.g., IL-15) or a nucleic acid (e.g., mRNA) that encodes the cytokine. When a cytokine or nucleic acid encoding a cytokine is administered in combination with one or more mRNA encoding one of more bispecific engagers, the cytokine or nucleic acid encoding the cytokine may be formulated together with the mRNA encoding the bispecific engager as part of the same composition or formulated separately in two or more different compositions. For example, the cytokine or nucleic acid encoding the cytokine and the mRNA encoding a bispecific engager (and, optionally, a second, third, fourth, fifth mRNA, each encoding a different bispecific engager) can be formulated in a single pharmaceutical composition and administered to a subject concurrently. Atty. Docket No.45817-0158WO1 Alternatively, the cytokine or nucleic acid encoding the cytokine and a mRNA encoding a bispecific engager (and, optionally, a second, third, fourth, fifth mRNA, each encoding a different bispecific engager) can be formulated into two different pharmaceutical compositions and administered concurrently, serially (i.e., back-to- back or one immediately following the other), or sequentially (i.e., with some predetermined amount of time between the administration of the first and second composition). The decision regarding how many different mRNAs should be administered, how many bispecific engagers should be expressed, what cytokines should be used, and what B cell targets the bispecific engagers should bind may be determined based on a screening and analysis of a subject’s disease. Examples of cytokines that may be combined with or co-expressed with the disclosed bispecific engagers include, but are not limited to, IL-15, granulocyte- macrophage colony-stimulating factor (GM-CSF), IFN gamma (IFNγ), IL-2, IL-7, IL- 12, and IL-21. An mRNA encoding a cytokine may comprise the same components discussed herein with respect to expression of the disclosed bispecific engagers. Nucleic Acids Encoding Bispecific Engagers The compositions of the disclosure can be administered in protein form, but preferably in the form of nucleic acids which are expressed in vivo. The examples of nucleic acids described herein may be used to deliver any of the disclosed bispecific engagers to a subject. These nucleic acids (e.g., RNAs, such as mRNAs) may be used as therapeutic agents to express the bispecific antibodies and engagers of the disclosure as a therapy to treat a target disease. An example of a specific mRNA construct that can be used in therapy is provided in Table 21 at the end of Example 3. The present disclosure provides nucleic acid, such as mRNA, encoding the disclosed bispecific antibodies or engagers. The nucleic acids, such as mRNA, can be used as therapeutics, which can be administered to a subject in order to treat a disease (e.g., cancer). Atty. Docket No.45817-0158WO1 In particular, the present disclosure provides methods of expressing at least one or multiple (e.g., 2, 3, 4, or 5 or more) of the disclosed bispecific engagers in vivo in a subject by administering to the subject one or more nucleic acids (e.g., mRNA) that encode the various bispecific engagers. For example, two different bispecific engagers of the present disclosure can be concurrently expressed in a subject by administering to the subject a first mRNA encoding a first bispecific engager and a second mRNA encoding a second bispecific engager, wherein the first and the second bispecific engagers each independently bind to different TAAs or the same TAA, and wherein the immune cell target molecule- binding domains may bind to the same type of immune cell (e.g., NK cell) or different types of immune cells. By way of illustration, the first mRNA may encode a bispecific engager with an anti-BCMA binding domain and an anti-CD16 binding domain and the second mRNA may encode a bispecific engager with an anti-FcRH5 binding domain and an anti-CD16 binding domain. By way of further illustration, the first mRNA may encode a bispecific engager with an anti-BCMA binding domain and an anti-CD16 binding domain and the second mRNA may encode a bispecific engager with an anti-FcRH5 binding domain and a different anti-NK cell molecule. As these illustrations show, the bispecific engagers can be combined in various ways, as needed, to successfully target heterologous populations of tumor cells. Similarly, two different bispecific engagers of the present disclosure can be concurrently expressed in a subject by administering to the subject a single mRNA encoding a first bispecific engager and a second bispecific engager. As with the embodiments in which two mRNA are administered, the first and second bispecific engagers may bind to different or the same TAAs and different or the same immune cell target molecules. This section provides examples of nucleic acids that may be used to encode bispecific antibodies or engagers of the disclosure. Using the genetic code, various nucleotide sequences encoding the engagers of the invention can be designed using methods known in the art. In some embodiments, the polynucleotide of the present Atty. Docket No.45817-0158WO1 disclosure (e.g., a RNA, e.g., an mRNA) comprises: a 5’ terminal cap (e.g., m⁷Gp- ppGm-A, e.g., Cap1, or an analog thereof); a 5’ UTR; a nucleotide sequence (e.g., an ORF) encoding a secreted engager; a 3′-UTR ; and a polyA tail (e.g., about 100 nucleotides in length. In some embodiments, the poly A tail is protected (e.g., with an inverted deoxy-thymidine). In some instances, the poly A tail comprises or consists of A100-UCUAG-A20-inverted deoxy-thymidine (SEQ ID NO: 530). The nucleic acid molecules of the disclosure may include one or more alterations. Herein, in a nucleotide, nucleoside, or polynucleotide (such as the nucleic acids of the disclosure (e.g., an mRNA or an oligonucleotide)), the terms “alteration” or, as appropriate, “alternative” refer to alteration with respect to A, G, U or C ribonucleotides. In one embodiment, the polynucleotide comprises at least one chemically modified nucleobase, e.g., N1-methylpseudouracil or 5-methoxyuracil. In certain embodiments, all uracils in the polynucleotide are N1-methylpseudouracils. The alterations may be various distinct alterations. In some embodiments, where the nucleic acid is an mRNA, the coding region, the flanking regions, and/or the terminal regions may contain one, two, or more (optionally different) nucleoside or nucleotide alterations. In some embodiments, an alternative polynucleotide introduced to a cell may exhibit reduced degradation in the cell, as compared to an unaltered polynucleotide. Modified Nucleotide Sequences Encoding Bispecific Engagers In some embodiments, the polynucleotide (e.g., a RNA, e.g., an mRNA) of the present disclosure comprises a chemically modified nucleobase, for example, a chemically modified uracil, e.g., pseudouracil, N1-methylpseudouracil, 5- methoxyuracil, or the like. In some embodiments, the mRNA is a uracil-modified sequence comprising an ORF encoding an Engager polypeptide, wherein the mRNA comprises a chemically modified nucleobase, for example, a chemically modified uracil, e.g., pseudouracil, N1-methylpseudouracil, or 5-methoxyuracil. Atty. Docket No.45817-0158WO1 In certain aspects of the present disclosure, when the modified uracil base is connected to a ribose sugar, as it is in polynucleotides, the resulting modified nucleoside or nucleotide is referred to as modified uridine. In some embodiments, uracil in the polynucleotide is at least about 25%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least 90%, at least 95%, at least 99%, or about 100% modified uracil. In some embodiments, uracil in the polynucleotide is at least 95% modified uracil. In some embodiments, uracil in the polynucleotide is 100% modified uracil. In embodiments where uracil in the polynucleotide is at least 95% modified uracil overall uracil content can be adjusted such that an mRNA provides suitable protein expression levels while inducing little to no immune response. In some embodiments, the uracil content of the ORF is between about 100% and about 150%, between about 100% and about 110%, between about 105% and about 115%, between about 110% and about 120%, between about 115% and about 125%, between about 120% and about 130%, between about 125% and about 135%, between about 130% and about 140%, between about 135% and about 145%, between about 140% and about 150% of the theoretical minimum uracil content in the corresponding wild-type ORF (%UTM). In other embodiments, the uracil content of the ORF is between about 121% and about 136% or between 123% and 134% of the %U_TM. In some embodiments, the uracil content of the ORF encoding an Engager polypeptide is about 115%, about 120%, about 125%, about 130%, about 135%, about 140%, about 145%, or about 150% of the %UTM. In this context, the term "uracil" can refer to modified uracil and/or naturally occurring uracil. In some embodiments, the uracil content in the ORF of the mRNA encoding an Engager polypeptide of the present disclosure is less than about 30%, about 25%, about 20%, about 15%, or about 10% of the total nucleobase content in the ORF. In some embodiments, the uracil content in the ORF is between about 10% and about 20% of the total nucleobase content in the ORF. In other embodiments, the uracil content in the ORF is between about 10% and about 25% of the total nucleobase content in the ORF. In some embodiments, the uracil content in the ORF of the Atty. Docket No.45817-0158WO1 mRNA encoding an Engager polypeptide is less than about 20% of the total nucleobase content in the open reading frame. In this context, the term "uracil" can refer to modified uracil and/or naturally occurring uracil. In further embodiments, the ORF of the mRNA encoding an Engager polypeptide having modified uracil and adjusted uracil content has increased Cytosine (C), Guanine (G), or Guanine/Cytosine (G/C) content (absolute or relative). In some embodiments, the overall increase in C, G, or G/C content (absolute or relative) of the ORF is at least about 2%, at least about 3%, at least about 4%, at least about 5%, at least about 6%, at least about 7%, at least about 10%, at least about 15%, at least about 20%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, at least about 95%, or at least about 100% relative to the G/C content (absolute or relative) of the wild-type ORF. In some embodiments, the G, the C, or the G/C content in the ORF is less than about 100%, less than about 90%, less than about 85%, or less than about 80% of the theoretical maximum G, C, or G/C content of the corresponding wild type nucleotide sequence encoding the Engager polypeptide (%GTMX; %CTMX, or %G/CTMX). In some embodiments, the increases in G and/or C content (absolute or relative) described herein can be conducted by replacing synonymous codons with low G, C, or G/C content with synonymous codons having higher G, C, or G/C content. In other embodiments, the increase in G and/or C content (absolute or relative) is conducted by replacing a codon ending with U with a synonymous codon ending with G or C. In further embodiments, the ORF of the mRNA encoding an Engager polypeptide of the present disclosure comprises modified uracil and has an adjusted uracil content containing less uracil pairs (UU) and/or uracil triplets (UUU) and/or uracil quadruplets (UUUU) than the corresponding wild-type nucleotide sequence encoding the Engager polypeptide. In some embodiments, the ORF of the mRNA encoding an Engager polypeptide of the present disclosure contains no uracil pairs and/or uracil triplets and/or uracil quadruplets. In some embodiments, uracil pairs and/or uracil triplets and/or uracil quadruplets are reduced below a certain threshold, e.g., no more than 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 Atty. Docket No.45817-0158WO1 occurrences in the ORF of the mRNA encoding the Engager polypeptide. In a particular embodiment, the ORF of the mRNA encoding the Engager polypeptide of the present disclosure contains less than 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 non-phenylalanine uracil pairs and/or triplets. In some embodiments, the ORF of the mRNA encoding the Engager polypeptide contains no non-phenylalanine uracil pairs and/or triplets. In further embodiments, the ORF of the mRNA encoding an Engager polypeptide of the present disclosure comprises modified uracil and has an adjusted uracil content containing fewer uracil-rich clusters than the corresponding wild-type nucleotide sequence encoding the Engager polypeptide. In some embodiments, the ORF of the mRNA encoding the Engager polypeptide of the present disclosure contains uracil-rich clusters that are shorter in length than corresponding uracil-rich clusters in the corresponding wild-type nucleotide sequence encoding the Engager polypeptide. In further embodiments, alternative lower frequency codons are employed. At least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 99%, or 100% of the codons in the Engager polypeptide–encoding ORF of the modified uracil-comprising mRNA are substituted with alternative codons, each alternative codon having a codon frequency lower than the codon frequency of the substituted codon in the synonymous codon set. The ORF also has adjusted uracil content, as described above. In some embodiments, at least one codon in the ORF of the mRNA encoding the Engager polypeptide is substituted with an alternative codon having a codon frequency lower than the codon frequency of the substituted codon in the synonymous codon set. Methods for Modifying Polynucleotides Atty. Docket No.45817-0158WO1 The disclosure includes modified polynucleotides comprising a polynucleotide described herein (e.g., a polynucleotide, e.g. mRNA, comprising a nucleotide sequence encoding an Engager polypeptide). The modified polynucleotides can be chemically modified and/or structurally modified. When the polynucleotides of the present disclosure are chemically and/or structurally modified the polynucleotides can be referred to as "modified polynucleotides." The present disclosure provides for modified nucleosides and nucleotides of a polynucleotide (e.g., RNA polynucleotides, such as mRNA polynucleotides) encoding an Engager polypeptide. A "nucleoside" refers to a compound containing a sugar molecule (e.g., a pentose or ribose) or a derivative thereof in combination with an organic base (e.g., a purine or pyrimidine) or a derivative thereof (also referred to herein as "nucleobase"). A “nucleotide" refers to a nucleoside including a phosphate group. Modified nucleotides can be synthesized by any useful method, such as, for example, chemically, enzymatically, or recombinantly, to include one or more modified or non-natural nucleosides. Polynucleotides can comprise a region or regions of linked nucleosides. Such regions can have variable backbone linkages. The linkages can be standard phosphodiester linkages, in which case the polynucleotides would comprise regions of nucleotides. The modified polynucleotides disclosed herein can comprise various distinct modifications. In some embodiments, the modified polynucleotides contain one, two, or more (optionally different) nucleoside or nucleotide modifications. In some embodiments, a modified polynucleotide, introduced to a cell can exhibit one or more desirable properties, e.g., improved protein expression, reduced immunogenicity, or reduced degradation in the cell, as compared to an unmodified polynucleotide. In some embodiments, a polynucleotide of the present disclosure (e.g., a polynucleotide comprising a nucleotide sequence encoding an Engager polypeptide) is structurally modified. As used herein, a "structural" modification is one in which two or more linked nucleosides are inserted, deleted, duplicated, inverted or randomized in a polynucleotide without significant chemical modification to the Atty. Docket No.45817-0158WO1 nucleotides themselves. Because chemical bonds will necessarily be broken and reformed to effect a structural modification, structural modifications are of a chemical nature and hence are chemical modifications. However, structural modifications will result in a different sequence of nucleotides. For example, the polynucleotide "ATCG" can be chemically modified to "AT-5meC-G". The same polynucleotide can be structurally modified from "ATCG" to "ATCCCG". Here, the dinucleotide "CC" has been inserted, resulting in a structural modification to the polynucleotide. Therapeutic compositions of the present disclosure comprise, in some embodiments, at least one nucleic acid (e.g., RNA) having an open reading frame encoding a secreted engager wherein the nucleic acid comprises nucleotides and/or nucleosides that can be standard (unmodified) or modified as is known in the art. In some embodiments, nucleotides and nucleosides of the present disclosure comprise modified nucleotides or nucleosides. Such modified nucleotides and nucleosides can be naturally-occurring modified nucleotides and nucleosides or non-naturally occurring modified nucleotides and nucleosides. Such modifications can include those at the sugar, backbone, or nucleobase portion of the nucleotide and/or nucleoside as are recognized in the art. In some embodiments, a naturally-occurring modified nucleotide or nucleotide of the disclosure is one as is generally known or recognized in the art. Non-limiting examples of such naturally occurring modified nucleotides and nucleotides can be found, inter alia, in the widely recognized MODOMICS database. In some embodiments, a non-naturally occurring modified nucleotide or nucleoside of the disclosure is one as is generally known or recognized in the art. Non-limiting examples of such non-naturally occurring modified nucleotides and nucleosides can be found, inter alia, in published US Application Nos. PCT/US2012/058519; PCT/US2013/075177; PCT/US2014/058897; PCT/US2014/058891; PCT/US2014/070413; PCT/US2015/36773; PCT/US2015/36759; PCT/US2015/36771; or PCT/IB2017/051367 all of which are incorporated by reference herein. Atty. Docket No.45817-0158WO1 In some embodiments, at least one RNA (e.g., mRNA) of the present disclosure is not chemically modified and comprises the standard ribonucleotides consisting of adenosine, guanosine, cytosine and uridine. In some embodiments, nucleotides and nucleosides of the present disclosure comprise standard nucleoside residues such as those present in transcribed RNA (e.g. A, G, C, or U). In some embodiments, nucleotides and nucleosides of the present disclosure comprise standard deoxyribonucleosides such as those present in DNA (e.g. dA, dG, dC, or dT). Hence, nucleic acids of the disclosure (e.g., DNA nucleic acids and RNA nucleic acids, such as mRNA nucleic acids) can comprise standard nucleotides and nucleosides, naturally-occurring nucleotides and nucleosides, non-naturally-occurring nucleotides and nucleosides, or any combination thereof. Nucleic acids of the disclosure (e.g., RNA nucleic acids, such as mRNA nucleic acids), in some embodiments, comprise various (more than one) different types of standard and/or modified nucleotides and nucleosides. In some embodiments, a particular region of a nucleic acid contains one, two or more (optionally different) types of standard and/or modified nucleotides and nucleosides. In some embodiments, a modified RNA nucleic acid (e.g., a modified mRNA nucleic acid), introduced to a cell or organism, exhibits reduced degradation in the cell or organism, respectively, relative to an unmodified nucleic acid comprising standard nucleotides and nucleosides. In some embodiments, a modified RNA nucleic acid (e.g., a modified mRNA nucleic acid), introduced into a cell or organism, may exhibit reduced immunogenicity in the cell or organism, respectively (e.g., a reduced innate response) relative to an unmodified nucleic acid comprising standard nucleotides and nucleosides. Nucleic acids (e.g., RNA nucleic acids, such as mRNA nucleic acids), in some embodiments, comprise non-natural modified nucleotides that are introduced during synthesis or post-synthesis of the nucleic acids to achieve desired functions or Atty. Docket No.45817-0158WO1 properties. The modifications may be present on internucleotide linkages, purine or pyrimidine bases, or sugars. The modification may be introduced with chemical synthesis or with a polymerase enzyme at the terminal of a chain or anywhere else in the chain. Any of the regions of a nucleic acid may be chemically modified. The present disclosure provides for modified nucleosides and nucleotides of a nucleic acid (e.g., RNA nucleic acids, such as mRNA nucleic acids). A “nucleoside” refers to a compound containing a sugar molecule (e.g., a pentose or ribose) or a derivative thereof in combination with an organic base (e.g., a purine or pyrimidine) or a derivative thereof (also referred to herein as “nucleobase”). A “nucleotide” refers to a nucleoside, including a phosphate group. Modified nucleotides may by synthesized by any useful method, such as, for example, chemically, enzymatically, or recombinantly, to include one or more modified or non-natural nucleosides. Nucleic acids can comprise a region or regions of linked nucleosides. Such regions may have variable backbone linkages. The linkages can be standard phosphodiester linkages, in which case the nucleic acids would comprise regions of nucleotides. Modified nucleotide base pairing encompasses not only the standard adenosine-thymine, adenosine-uracil, or guanosine-cytosine base pairs, but also base pairs formed between nucleotides and/or modified nucleotides comprising non- standard or modified bases, wherein the arrangement of hydrogen bond donors and hydrogen bond acceptors permits hydrogen bonding between a non-standard base and a standard base or between two complementary non-standard base structures, such as, for example, in those nucleic acids having at least one chemical modification. One example of such non-standard base pairing is the base pairing between the modified nucleotide inosine and adenine, cytosine or uracil. Any combination of base/sugar or linker may be incorporated into nucleic acids of the present disclosure. In some embodiments, modified nucleobases in nucleic acids (e.g., RNA nucleic acids, such as mRNA nucleic acids) comprise N1-methyl-pseudouridine (m1ψ), 1-ethyl-pseudouridine (e1ψ), 5-methoxy-uridine (mo5U), 5-methyl-cytidine (m5C), and/or pseudouridine (ψ). In some embodiments, modified nucleobases in Atty. Docket No.45817-0158WO1 nucleic acids (e.g., RNA nucleic acids, such as mRNA nucleic acids) comprise 5- methoxymethyl uridine, 5-methylthio uridine, 1-methoxymethyl pseudouridine, 5- methyl cytidine, and/or 5-methoxy cytidine. In some embodiments, the polyribonucleotide includes a combination of at least two (e.g., 2, 3, 4 or more) of any of the aforementioned modified nucleobases, including but not limited to chemical modifications. In some embodiments, a RNA nucleic acid of the disclosure comprises N1- methyl-pseudouridine (m1ψ) substitutions at one or more or all uridine positions of the nucleic acid. In some embodiments, a RNA nucleic acid of the disclosure comprises N1- methyl-pseudouridine (m1ψ) substitutions at one or more or all uridine positions of the nucleic acid and 5-methyl cytidine substitutions at one or more or all cytidine positions of the nucleic acid. In some embodiments, a RNA nucleic acid of the disclosure comprises pseudouridine (ψ) substitutions at one or more or all uridine positions of the nucleic acid. In some embodiments, a RNA nucleic acid of the disclosure comprises pseudouridine (ψ) substitutions at one or more or all uridine positions of the nucleic acid and 5-methyl cytidine substitutions at one or more or all cytidine positions of the nucleic acid. In some embodiments, a RNA nucleic acid of the disclosure comprises uridine at one or more or all uridine positions of the nucleic acid. In some embodiments, nucleic acids (e.g., RNA nucleic acids, such as mRNA nucleic acids) are uniformly modified (e.g., fully modified, modified throughout the entire sequence) for a particular modification. For example, a nucleic acid can be uniformly modified with N1-methyl-pseudouridine, meaning that all uridine residues in the mRNA sequence are replaced with N1-methyl-pseudouridine. Similarly, a Atty. Docket No.45817-0158WO1 nucleic acid can be uniformly modified for any type of nucleoside residue present in the sequence by replacement with a modified residue such as those set forth above. The nucleic acids of the present disclosure may be partially or fully modified along the entire length of the molecule. For example, one or more or all or a given type of nucleotide (e.g., purine or pyrimidine, or any one or more or all of A, G, U, C) may be uniformly modified in a nucleic acid of the disclosure, or in a predetermined sequence region thereof (e.g., in the mRNA including or excluding the polyA tail). In some embodiments, all nucleotides X in a nucleic acid of the present disclosure (or in a sequence region thereof) are modified nucleotides, wherein X may be any one of nucleotides A, G, U, C, or any one of the combinations A+G, A+U, A+C, G+U, G+C, U+C, A+G+U, A+G+C, G+U+C or A+G+C. The nucleic acid may contain from about 1% to about 100% modified nucleotides (either in relation to overall nucleotide content, or in relation to one or more types of nucleotide, i.e., any one or more of A, G, U or C) or any intervening percentage (e.g., from 1% to 20%, from 1% to 25%, from 1% to 50%, from 1% to 60%, from 1% to 70%, from 1% to 80%, from 1% to 90%, from 1% to 95%, from 10% to 20%, from 10% to 25%, from 10% to 50%, from 10% to 60%, from 10% to 70%, from 10% to 80%, from 10% to 90%, from 10% to 95%, from 10% to 100%, from 20% to 25%, from 20% to 50%, from 20% to 60%, from 20% to 70%, from 20% to 80%, from 20% to 90%, from 20% to 95%, from 20% to 100%, from 50% to 60%, from 50% to 70%, from 50% to 80%, from 50% to 90%, from 50% to 95%, from 50% to 100%, from 70% to 80%, from 70% to 90%, from 70% to 95%, from 70% to 100%, from 80% to 90%, from 80% to 95%, from 80% to 100%, from 90% to 95%, from 90% to 100%, and from 95% to 100%). It will be understood that any remaining percentage is accounted for by the presence of unmodified A, G, U, or C. The nucleic acids may contain at a minimum 1% and at maximum 100% modified nucleotides, or any intervening percentage, such as at least 5% modified nucleotides, at least 10% modified nucleotides, at least 25% modified nucleotides, at least 50% modified nucleotides, at least 80% modified nucleotides, or at least 90% Atty. Docket No.45817-0158WO1 modified nucleotides. For example, the nucleic acids may contain a modified pyrimidine such as a modified uracil or cytosine. In some embodiments, at least 5%, at least 10%, at least 25%, at least 50%, at least 80%, at least 90% or 100% of the uracil in the nucleic acid is replaced with a modified uracil (e.g., a 5-substituted uracil). The modified uracil can be replaced by a compound having a single unique structure, or can be replaced by a plurality of compounds having different structures (e.g., 2, 3, 4 or more unique structures). In some embodiments, at least 5%, at least 10%, at least 25%, at least 50%, at least 80%, at least 90% or 100% of the cytosine in the nucleic acid is replaced with a modified cytosine (e.g., a 5-substituted cytosine). The modified cytosine can be replaced by a compound having a single unique structure, or can be replaced by a plurality of compounds having different structures (e.g., 2, 3, 4 or more unique structures). Messenger RNA The present disclosure features compositions including one or more mRNAs, where each mRNA encodes at least one (e.g., 1, 2, 3, 4, or 5 or more) bispecific engagers as described herein. mRNAs of the disclosure may each include (i) a 5’-cap structure; (ii) a 5’-UTR; (iii) an open reading frame encoding the polypeptide; (iv) a 3’-untranslated region (3’-UTR); and (v) a poly-A region. In some embodiments, the mRNA includes from about 30 to about 3,000 (e.g., from 30 to 50, from 30 to 100, from 30 to 250, from 30 to 500, from 30 to 750, from 30 to 1,000, from 30 to 1,500, from 30 to 2,000, from 30 to 2,500, from 50 to 100, from 50 to 250, from 50 to 500, from 50 to 750, from 50 to 1,000, from 50 to 1,500, from 50 to 2,000, from 50 to 2,500, from 50 to 3,000, from 100 to 500, from 100 to 750, from 100 to 1,000, from 100 to 1,500, from 100 to 2,000, from 100 to 2,500, from 100 to 3,000, from 500 to 750, from 500 to 1,000, from 500 to 1,500, from 500 to 2,000, from 500 to 2,500, from 500 to 3,000, from 1,000 to 1,500, from 1,000 to 2,000, from 1,000 to 2,500, from 1,000 to 3,000, from 1,500 to 2,000, from 1,500 to 2,500, from 1,500 to 3,000, from 2,000 to 3,000, from 2,000 to 2,500, or from 2,500 to 3,000) nucleotides. Atty. Docket No.45817-0158WO1 More specifically, provided herein are nucleic acids that encode VHH domains and bispecific engagers of the disclosure. Any of the proteins disclosed herein may be encoded by an open reading frame (ORF) of an mRNA. In some embodiments, the ORF of the mRNA comprises the nucleic acid sequence (wherein all uracils are N1-methylpseudouracils) : AUGGAGACCCCUGCCCAGCUGCUGUUCCUGCUGCUGCUGUGGCUGCCCG ACACCACCGGCCAGGUGCAGUUGCAGGAGUCCGGCGGAGGCCUGGUCCA ACCCGGCGGCUCACUGCGGCUUAGCUGCGCCGCAAGCGGCCGAACCUAC AACAACUACGCCAUGGGGUGGUUUCGGCAAGCCCCAGGCAAGGAACGG GAGUUCGUGGCCGGCAUCAGCCGGAGCGGCGGCAUGACCGGCUACGCCG AGAGCGUGAAGGGUCGGUUCACCAUUAGCCGGGAUAACAGCAAGAAUA CCGUCUACCUUCAGAUGAACAGCCUUAGAGCCGAGGACACCGCUGUUUA CUACUGUGCCGCCUACGUGGGCGGCUUCAGCACCGCCCGGCGGGACUAC AGCUACUGGGGACAGGGGACCCAAGUGACAGUGAGCAGCGAGAGCAAG UACGGACCUCCUUGCCCUCCCUGUCCAGCCCCAGAGGCCGCUGGAGGCC CAAGCGUGUUCCUGUUCCCACCCAAGCCCAAGGACACCCUGAUGAUCAG CCGGACCCCAGAGGUGACCUGCGUGGUGGUGGACGUGAGCCAGGAGGA CCCCGAGGUGCAGUUCAACUGGUACGUGGACGGCGUGGAGGUGCACAA CGCCAAGACCAAGCCCCGGGAGGAGCAGUUCAACAGCACCUACCGGGUG GUGAGCGUGCUGACCGUGCUGCACCAGGACUGGCUGAACGGCAAGGAG UACAAGUGCAAGGUGAGCAACAAGGGCCUGCCCAGCAGCAUCGAGAAG ACCAUCAGCAAGGCAAAGGGCCAACCUCGGGAGCCACAGGUGUACACCC UGCCUCCCAGCCAGGAGGAGAUGACCAAGAACCAGGUGAGCCUGACCUG CCUGGUGAAGGGCUUCUACCCCAGCGACAUCGCCGUGGAGUGGGAGAGC AACGGCCAGCCCGAGAACAACUACAAGACCACUCCACCAGUGCUGGACA GCGACGGCAGCUUCUUCCUGUACAGCCGGCUGACCGUGGACAAGAGCCG GUGGCAGGAGGGCAACGUGUUCAGCUGCAGCGUGAUGCACGAGGCCCU GCACAACCACUACACCCAGAAGAGCCUGAGCCUCAGCCUGGGCAAGGGU GGAGGCGGGAGCGAGGUGCAGCUGGUGGAAAGCGGCGGCGGCCUGGUG CAACCCGGCGGCAGCCUGCGACUGUCCUGUGCCGCCUCUGGCCGGACCG Atty. Docket No.45817-0158WO1 ACAGCAUCUACGCCAUGGGCUGGUUCCGGCAGGCACCCGGCAAGGAGCG GGAGUUCGUGAGCGCCAUCAACAGCAACACCGGCCGGACCUACCACGCC GACAGCGUGAAGGGCCGGUUCACCAUCAGCCGGGACAACGCCAAGAACA UGGUGUACCUGCAGAUGAACAGCCUGCGGGCCGAGGAUACCGCCGUGU ACUAUUGCGCCGCCGGACGGGGUUACGGCCUGCUGAGCAUCAGCAGCAA CUGGUACAACUACUGGGGCCAGGGCACCCUGGUGACCGUGAGCAGC (SEQ ID NO: 463), as disclosed in Table 21 at the end of Example 3. It should be noted that the ORF may include a different signal sequence than the one listed in SEQ ID NO:463. Similar such open reading frames can be designed from the protein sequences disclosed herein or with alternate VHH binding domains or FC domains known in the art using the genetic code and the codon optimization strategies known to one of ordinary skill in the art. Untranslated Regions (UTRs) Untranslated regions (UTRs) are nucleic acid sections of a polynucleotide before a start codon (5′ UTR) and after a stop codon (3′ UTR) that are not translated. In some embodiments, a polynucleotide (e.g., a ribonucleic acid (RNA), e.g., a messenger RNA (mRNA)) of the present disclosure comprising an open reading frame (ORF) encoding an Engager polypeptide further comprises a UTR (e.g., a 5′ UTR or functional fragment thereof, a 3′ UTR or functional fragment thereof, or a combination thereof). A UTR (e.g., 5′ UTR or 3′ UTR) can be homologous or heterologous to the coding region in a polynucleotide. In some embodiments, the UTR is homologous to the ORF encoding the Engager polypeptide. In some embodiments, the UTR is heterologous to the ORF encoding the Engager polypeptide. In some embodiments, the polynucleotide comprises two or more 5′ UTRs or functional fragments thereof, each of which has the same or different nucleotide sequences. In some embodiments, the polynucleotide comprises two or more 3′ UTRs Atty. Docket No.45817-0158WO1 or functional fragments thereof, each of which has the same or different nucleotide sequences. In some embodiments, the 5′ UTR or functional fragment thereof, 3′ UTR or functional fragment thereof, or any combination thereof is sequence optimized. In some embodiments, the 5′UTR or functional fragment thereof, 3′ UTR or functional fragment thereof, or any combination thereof comprises at least one chemically modified nucleobase, e.g., N1-methylpseudouracil or 5-methoxyuracil. UTRs can have features that provide a regulatory role, e.g., increased or decreased stability, localization and/or translation efficiency. A polynucleotide comprising a UTR can be administered to a cell, tissue, or organism, and one or more regulatory features can be measured using routine methods. In some embodiments, a functional fragment of a 5′ UTR or 3′ UTR comprises one or more regulatory features of a full length 5′ or 3′ UTR, respectively. Natural 5′UTRs bear features that play roles in translation initiation. They harbor signatures like Kozak sequences that are commonly known to be involved in the process by which the ribosome initiates translation of many genes. Kozak sequences have the consensus CCR(A/G)CCAUGG, where R is a purine (adenine or guanine) three bases upstream of the start codon (AUG), which is followed by another ‘G’.5′ UTRs also have been known to form secondary structures that are involved in elongation factor binding. By engineering the features typically found in abundantly expressed genes of specific target organs, one can enhance the stability and protein production of a polynucleotide. For example, introduction of 5′ UTR of liver-expressed mRNA, such as albumin, serum amyloid A, Apolipoprotein A/B/E, transferrin, alpha fetoprotein, erythropoietin, or Factor VIII, can enhance expression of polynucleotides in hepatic cell lines or liver. Likewise, use of 5′UTR from other tissue-specific mRNA to improve expression in that tissue is possible for muscle (e.g., MyoD, Myosin, Myoglobin, Myogenin, Herculin), for endothelial cells (e.g., Tie-1, CD36), for Atty. Docket No.45817-0158WO1 myeloid cells (e.g., C/EBP, AML1, G-CSF, GM-CSF, CD11b, MSR, Fr-1, i-NOS), for leukocytes (e.g., CD45, CD18), for adipose tissue (e.g., CD36, GLUT4, ACRP30, adiponectin) and for lung epithelial cells (e.g., SP-A/B/C/D). In some embodiments, UTRs are selected from a family of transcripts whose proteins share a common function, structure, feature or property. For example, an encoded polypeptide can belong to a family of proteins (i.e., that share at least one function, structure, feature, localization, origin, or expression pattern), which are expressed in a particular cell, tissue or at some time during development. The UTRs from any of the genes or mRNA can be swapped for any other UTR of the same or different family of proteins to create a new polynucleotide. In some embodiments, the 5′ UTR and the 3′ UTR can be heterologous. In some embodiments, the 5′ UTR can be derived from a different species than the 3′ UTR. In some embodiments, the 3′ UTR can be derived from a different species than the 5′ UTR. Co-owned International Patent Application No. PCT/US2014/021522 (Publ. No. WO/2014/164253, incorporated herein by reference in its entirety) provides a listing of exemplary UTRs that can be utilized in the polynucleotide of the present disclosure as flanking regions to an ORF. Additional exemplary UTRs of the application include, but are not limited to, one or more 5′UTR and/or 3′UTR derived from the nucleic acid sequence of: a globin, such as an α- or β-globin (e.g., a Xenopus, mouse, rabbit, or human globin); a strong Kozak translational initiation signal; a CYBA (e.g., human cytochrome b-245 α polypeptide); an albumin (e.g., human albumin7); a HSD17B4 (hydroxysteroid (17-β) dehydrogenase); a virus (e.g., a tobacco etch virus (TEV), a Venezuelan equine encephalitis virus (VEEV), a Dengue virus, a cytomegalovirus (CMV) (e.g., CMV immediate early 1 (IE1)), a hepatitis virus (e.g., hepatitis B virus), a sindbis virus, or a PAV barley yellow dwarf virus); a heat shock protein (e.g., hsp70); a translation initiation factor (e.g., elF4G); a glucose transporter (e.g., hGLUT1 (human glucose transporter 1)); an actin (e.g., human α or β actin); a GAPDH; a tubulin; a histone; a Atty. Docket No.45817-0158WO1 citric acid cycle enzyme; a topoisomerase (e.g., a 5′UTR of a TOP gene lacking the 5′ TOP motif (the oligopyrimidine tract)); a ribosomal protein Large 32 (L32); a ribosomal protein (e.g., human or mouse ribosomal protein, such as, for example, rps9); an ATP synthase (e.g., ATP5A1 or the β subunit of mitochondrial H⁺-ATP synthase); a growth hormone (e.g., bovine (bGH) or human (hGH)); an elongation factor (e.g., elongation factor 1 α1 (EEF1A1)); a manganese superoxide dismutase (MnSOD); a myocyte enhancer factor 2A (MEF2A); a β-F1-ATPase, a creatine kinase, a myoglobin, a granulocyte-colony stimulating factor (G-CSF); a collagen (e.g., collagen type I, alpha 2 (Col1A2), collagen type I, alpha 1 (Col1A1), collagen type VI, alpha 2 (Col6A2), collagen type VI, alpha 1 (Col6A1)); a ribophorin (e.g., ribophorin I (RPNI)); a low density lipoprotein receptor-related protein (e.g., LRP1); a cardiotrophin-like cytokine factor (e.g., Nnt1); calreticulin (Calr); a procollagen- lysine, 2-oxoglutarate 5-dioxygenase 1 (Plod1); and a nucleobindin (e.g., Nucb1). In some embodiments, the 5′ UTR is selected from the group consisting of a βglobin 5′ UTR; a 5′UTR containing a strong Kozak translational initiation signal; a cytochrome b-245 α polypeptide (CYBA) 5′ UTR; a hydroxysteroid (17-β) dehydrogenase (HSD17B4) 5′ UTR; a Tobacco etch virus (TEV) 5′ UTR; a Venezuelen equine encephalitis virus (TEEV) 5′ UTR; a 5′ proximal open reading frame of rubella virus (RV) RNA encoding nonstructural proteins; a Dengue virus (DEN) 5′ UTR; a heat shock protein 70 (Hsp70) 5′ UTR; a eIF4G 5′ UTR; a GLUT1 5′ UTR; functional fragments thereof and any combination thereof. In some embodiment, the mRNA comprises a 5’-UTR comprising a nucleic acid sequence of AGGAAAUAAGAGAGAAAAGAAGAGUAAGAAGAAAUAUAAGACCCCGGC GCCGCCACC (SEQ ID NO: 467), as showing in Table 21 at the end of Example 3. In some embodiment, the mRNA comprises a 3’-UTR comprising a nucleic acid sequence of UGAUAAUAGGCUGGAGCCUCGGUGGCCUAGCUUCUUGCCCCUUGGGCCU CCCCCCAGCCCCUCCUCCCCUUCCUGCACCCGUACCCCCGUGGUCUUUGA Atty. Docket No.45817-0158WO1 AUAAAGUCUGAGUGGGCGGC (SEQ ID NO: 468), as showing in Table 21 at the end of Example 3. In some embodiments, the 3′ UTR is selected from the group consisting of a βglobin 3′ UTR; a CYBA 3′ UTR; an albumin 3′ UTR; a growth hormone (GH) 3′ UTR; a VEEV 3′ UTR; a hepatitis B virus (HBV) 3′ UTR; α-globin 3′UTR; a DEN 3′ UTR; a PAV barley yellow dwarf virus (BYDV-PAV) 3′ UTR; an elongation factor 1 α1 (EEF1A1) 3′ UTR; a manganese superoxide dismutase (MnSOD) 3′ UTR; a β subunit of mitochondrial H(+)-ATP synthase (β-mRNA) 3′ UTR; a GLUT13′ UTR; a MEF2A 3′ UTR; a β-F1-ATPase 3′ UTR; functional fragments thereof, and combinations thereof. Wild-type UTRs derived from any gene or mRNA can be incorporated into the polynucleotides of the present disclosure. In some embodiments, a UTR can be altered relative to a wild type or native UTR to produce a variant UTR, e.g., by changing the orientation or location of the UTR relative to the ORF; or by inclusion of additional nucleotides, deletion of nucleotides, swapping or transposition of nucleotides. In some embodiments, variants of 5′ or 3′ UTRs can be utilized, for example, mutants of wild type UTRs, or variants wherein one or more nucleotides are added to or removed from a terminus of the UTR. Additionally, one or more synthetic UTRs can be used in combination with one or more non-synthetic UTRs. See, e.g., Mandal and Rossi, Nat. Protoc.2013 8(3):568-82, the contents of which are incorporated herein by reference in their entirety. UTRs or portions thereof can be placed in the same orientation as in the transcript from which they were selected or can be altered in orientation or location. Hence, a 5′ and/or 3′ UTR can be inverted, shortened, lengthened, or combined with one or more other 5′ UTRs or 3′ UTRs. In some embodiments, the polynucleotide comprises multiple UTRs, e.g., a double, a triple or a quadruple 5′ UTR or 3′ UTR. For example, a double UTR Atty. Docket No.45817-0158WO1 comprises two copies of the same UTR either in series or substantially in series. For example, a double beta-globin 3′UTR can be used (see US2010/0129877, the contents of which are incorporated herein by reference in its entirety). The polynucleotides of the present disclosure can comprise combinations of features. For example, the ORF can be flanked by a 5′UTR that comprises a strong Kozak translational initiation signal and/or a 3′UTR comprising an oligo(dT) sequence for templated addition of a poly-A tail. A 5′UTR can comprise a first polynucleotide fragment and a second polynucleotide fragment from the same and/or different UTRs (see, e.g., US2010/0293625, herein incorporated by reference in its entirety). Other non-UTR sequences can be used as regions or subregions within the polynucleotides of the present disclosure. For example, introns or portions of intron sequences can be incorporated into the polynucleotides of the present disclosure. Incorporation of intronic sequences can increase protein production as well as polynucleotide expression levels. In some embodiments, the polynucleotide of the present disclosure comprises an internal ribosome entry site (IRES) instead of or in addition to a UTR (see, e.g., Yakubov et al., Biochem. Biophys. Res. Commun.2010 394(1):189-193, the contents of which are incorporated herein by reference in their entirety). In some embodiments, the polynucleotide comprises an IRES instead of a 5′ UTR sequence. In some embodiments, the polynucleotide comprises an ORF and a viral capsid sequence. In some embodiments, the polynucleotide comprises a synthetic 5′ UTR in combination with a non-synthetic 3′ UTR. In some embodiments, the UTR can also include at least one translation enhancer polynucleotide, translation enhancer element, or translational enhancer elements (collectively, "TEE," which refers to nucleic acid sequences that increase the amount of polypeptide or protein produced from a polynucleotide. As a non-limiting example, the TEE can be located between the transcription promoter and the start codon. In some embodiments, the 5′ UTR comprises a TEE. Atty. Docket No.45817-0158WO1 In one aspect, a TEE is a conserved element in a UTR that can promote translational activity of a nucleic acid such as, but not limited to, cap-dependent or cap-independent translation. 5′ UTR sequences 5′ UTR sequences are important for ribosome recruitment to the mRNA and have been reported to play a role in translation (Hinnebusch A, et al., (2016) Science, 352:6292: 1413-6). Disclosed herein, inter alia, is a polynucleotide, e.g., mRNA, comprising an open reading frame (e.g., any one of SEQ ID NOs: 461, 463, or 465) encoding an Engager polypeptide (e.g., any one of SEQ ID NO: 394-446), which polynucleotide has a 5′ UTR that confers an increased half-life, increased expression and/or increased activity of the polypeptide encoded by said polynucleotide, or of the polynucleotide itself. In some embodiments, a polynucleotide disclosed herein comprises: (a) a 5′- UTR (e.g., as provided in Table 10 or a variant or fragment thereof); (b) a coding region comprising a stop element (e.g., as described herein); and (c) a 3′-UTR (e.g., as described herein), and LNP compositions comprising the same. In some embodiments, the polynucleotide comprises a 5′-UTR comprising a sequence provided in Table 10 or a variant or fragment thereof (e.g., a functional variant or fragment thereof). In some embodiments, the polynucleotide having a 5′ UTR sequence provided in Table 10 or a variant or fragment thereof, has an increase in the half-life of the polynucleotide, e.g., about 1.5-20-fold increase in half-life of the polynucleotide. In some embodiments, the increase in half-life is about 1.5-, 2-, 3-, 4-, 5-, 6-, 7-, 8-, 9-, 10-, 11-, 12-, 13-, 14-, 15-, 16-, 17-, 18-, 19- or 20-fold, or more. In some embodiments, the increase in half life is about 1.5-fold or more. In some embodiments, the increase in half life is about 2-fold or more. In some embodiments, the increase in half life is about 3-fold or more. In some embodiments, the increase in half life is about 4-fold or more. In some embodiments, the increase in half life is about 5-fold or more. Atty. Docket No.45817-0158WO1 In some embodiments, the polynucleotide having a 5′ UTR sequence provided in Table 10 or a variant or fragment thereof, results in an increased level and/or activity, e.g., output, of the polypeptide encoded by the polynucleotide. In some embodiments, the 5′UTR results in about 1.5-20-fold increase in level and/or activity, e.g., output, of the polypeptide encoded by the polynucleotide. In some embodiments, the increase in level and/or activity is about 1.5-, 2-, 3-, 4-, 5-, 6-, 7-, 8-, 9-, 10-, 11-, 12-, 13-, 14-, 15-, 16-, 17-, 18-, 19- or 20-fold, or more. In some embodiments, the increase in level and/or activity is about 1.5-fold or more. In some embodiments, the increase in level and/or activity is about 2-fold or more. In some embodiments, the increase in level and/or activity is about 3-fold or more. In some embodiments, the increase in level and/or activity is about 4-fold or more. In some embodiments, the increase in level and/or activity is about 5-fold or more. In some embodiments, the increase is compared to an otherwise similar polynucleotide which does not have a 5′ UTR, has a different 5′ UTR, or does not have a 5′ UTR described in Table 10 or a variant or fragment thereof. In some embodiments, the increase in half-life of the polynucleotide is measured according to an assay that measures the half-life of a polynucleotide. In some embodiments, the increase in level and/or activity, e.g., output, of the polypeptide encoded by the polynucleotide is measured according to an assay that measures the level and/or activity of a polypeptide. In some embodiments, the 5′ UTR comprises a sequence provided in Table 10 or a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to a 5′ UTR sequence provided in Table 10, or a variant or a fragment thereof. In some embodiments, the 5′ UTR comprises a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 470, SEQ ID NO: 471, SEQ ID NO: 472, SEQ ID NO: 473, SEQ ID NO: 474, SEQ ID NO: 475, SEQ ID NO: 476, SEQ ID NO: 477, SEQ ID NO: 478, or SEQ ID NO: 498. Atty. Docket No.45817-0158WO1 In some embodiments, the 5′ UTR comprises a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 470. In some embodiments, the 5′ UTR comprises a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 471. In some embodiments, the 5′ UTR comprises a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 472. In some embodiments, the 5′ UTR comprises a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 473. In some embodiments, the 5′ UTR comprises a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 474. In some embodiments, the 5′ UTR comprises a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 475. In some embodiments, the 5′ UTR comprises a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 476. In some embodiments, the 5′ UTR comprises a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 477. In some embodiments, the 5′ UTR comprises a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 478. In some embodiments, the 5′ UTR comprises a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 498. In some embodiments, the 5′ UTR comprises the sequence of SEQ ID NO: 470. In some embodiments, the 5′ UTR consists of the sequence of SEQ ID NO: 470. In some embodiments, the 5′ UTR comprises the sequence of SEQ ID NO: 475. In some embodiments, the 5′ UTR consists of the sequence of SEQ ID NO: 475. In some embodiments, the 5′ UTR comprises the sequence of SEQ ID NO: 476. In some embodiments, the 5′ UTR consists of the sequence of SEQ ID NO: 476. In some embodiments, the 5′ UTR comprises the sequence of SEQ ID NO: 498. In some embodiments, the 5′ UTR consists of the sequence of SEQ ID NO: 498. In some embodiments, a 5′ UTR sequence provided in Table 10 has a first nucleotide (not shown) which is an A. In some embodiments, a 5′ UTR sequence provided in Table 10 has a first nucleotide (not shown) which is a G. Atty. Docket No.45817-0158WO1 Table 10 – 5’ UTR Sequences SEQ ID Sequence Sequence U G U G C C A C U U G U G G U or U C C

Atty. Docket No.45817-0158WO1 A13 GGAAAAUCUGUAUUAGGUUGGCGUGUUCUUUGGUCGGU C U C U A U G A G A A G G U G C C U U A C G C

Atty. Docket No.45817-0158WO1 In some embodiments, the 5′ UTR comprises a variant of SEQ ID NO:50. In some embodiments, the variant of SEQ ID NO: 470 comprises a nucleic acid sequence of Formula A: G G A A A U C G C A A A A (N2)X (N3)X C U (N4)X (N5)X C G C G U U A G A U U U C U U U U A G U U U U C U N6 N7 C A A C U A G C A A G C U U U U U G U U C U C G C C (N8 C C)x (SEQ ID NO: 479), wherein: (N2)x is a uracil and x is an integer from 0 to 5, e.g., wherein x =3 or 4; (N₃)_x is a guanine and x is an integer from 0 to 1; (N4)x is a cytosine and x is an integer from 0 to 1; (N₅)_x is a uracil and x is an integer from 0 to 5, e.g., wherein x =2 or 3; N6 is a uracil or cytosine; N₇ is a uracil or guanine; and N8 is adenine or guanine and x is an integer from 0 to 1. In some embodiments (N2)x is a uracil and x is 0. In some embodiments (N2)x is a uracil and x is 1. In some embodiments (N2)x is a uracil and x is 2. In some embodiments (N₂)_x is a uracil and x is 3. In some embodiments, (N₂)_x is a uracil and x is 4. In some embodiments (N2)x is a uracil and x is 5. In some embodiments, (N3)x is a guanine and x is 0. In some embodiments, (N₃)_x is a guanine and x is 1. In some embodiments, (N4)x is a cytosine and x is 0. In some embodiments, (N₄)_x is a cytosine and x is 1. In some embodiments, (N₅)_x is a uracil and x is 0. In some embodiments, (N₅)_x is a uracil and x is 1. In some embodiments, (N5)x is a uracil and x is 2. In some embodiments, (N₅)_x is a uracil and x is 3. In some embodiments, (N₅)_x is a uracil and x is 4. In some embodiments (N5)x is a uracil and x is 5. In some embodiments, N6 is a uracil. In some embodiments, N6 is a cytosine. In some embodiments, N7 is a uracil. In some embodiments, N7 is a guanine. Atty. Docket No.45817-0158WO1 In some embodiments, N8 is an adenine and x is 0. In some embodiments, N8 is an adenine and x is 1. In some embodiments, N8 is a guanine and x is 0. In some embodiments, N8 is a guanine and x is 1. In some embodiments, the 5′ UTR comprises a variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498. In some embodiments, the variant of SEQ ID NO:50, SEQ ID NO:55, SEQ ID NO:56, or SEQ ID NO:78 comprises a sequence with at least 50%, 60%, 70%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity to SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises a sequence with at least 50% identity to SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises a sequence with at least 60% identity to SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises a sequence with at least 70% identity to SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises a sequence with at least 80% identity to SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises a sequence with at least 90% identity to SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises a sequence with at least 95% identity to SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises a sequence with at least 96% identity to SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID Atty. Docket No.45817-0158WO1 NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises a sequence with at least 97% identity to SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises a sequence with at least 98% identity to SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises a sequence with at least 99% identity to SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises a uridine content of at least 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, or 80%. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises a uridine content of at least 5%. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises a uridine content of at least 10%. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises a uridine content of at least 20%. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises a uridine content of at least 30%. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises a uridine content of at least 40%. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises a uridine content of at least 50%. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises a uridine content of at least 60%. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises a uridine content of at least 70%. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises a uridine content of at least 80%. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises at least 2, 3, 4, 5, 6 or 7 consecutive Atty. Docket No.45817-0158WO1 uridines (e.g., a polyuridine tract). In some embodiments, the polyuridine tract in the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises at least 1-7, 2-7, 3-7, 4-7, 5-7, 6-7, 1-6, 1-5, 1-4, 1-3, 1-2, 2-6, or 3-5 consecutive uridines. In some embodiments, the polyuridine tract in the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises 4 consecutive uridines. In some embodiments, the polyuridine tract in the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises 5 consecutive uridines. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 polyuridine tracts. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises 3 polyuridine tracts. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises 4 polyuridine tracts. In some embodiments, the variant of SEQ ID NO: 470, SEQ ID NO: 475, SEQ ID NO: 476, or SEQ ID NO: 498 comprises 5 polyuridine tracts. In some embodiments, one or more of the polyuridine tracts are adjacent to a different polyuridine tract. In some embodiments, each of, e.g., all, the polyuridine tracts are adjacent to each other, e.g., all of the polyuridine tracts are contiguous. In some embodiments, one or more of the polyuridine tracts are separated by 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 2, 13, 14, 15, 16, 17, 18.19, 20, 30, 40, 50 or 60 nucleotides. In some embodiments, each of, e.g., all of, the polyuridine tracts are separated by 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 2, 13, 14, 15, 16, 17, 18.19, 20, 30, 40, 50 or 60 nucleotides. In some embodiments, a first polyuridine tract and a second polyuridine tract are adjacent to each other. In some embodiments, a subsequent, e.g., third, fourth, fifth, sixth or seventh, eighth, ninth, or tenth, polyuridine tract is separated by 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, Atty. Docket No.45817-0158WO1 2, 13, 14, 15, 16, 17, 18.19, 20, 30, 40, 50 or 60 nucleotides from the first polyuridine tract, the second polyuridine tract, or any one of the subsequent polyuridine tracts. In some embodiments, a first polyuridine tract is separated by 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 2, 13, 14, 15, 16, 17, 18.19, 20, 30, 40, 50 or 60 nucleotides from a subsequent polyuridine tract, e.g., a second, third, fourth, fifth, sixth or seventh, eighth, ninth, or tenth polyuridine tract. In some embodiments, one or more of the subsequent polyuridine tracts are adjacent to a different polyuridine tract. In some embodiments, the 5′ UTR comprises a Kozak sequence, e.g., a GCCRCC nucleotide sequence wherein R is an adenine or guanine. In some embodiments, the Kozak sequence is disposed at the 3′ end of the 5′UTR sequence. In an aspect, the polynucleotide (e.g., mRNA) comprising an open reading frame (e.g., SEQ ID NO: 463) encoding an Engager polypeptide (e.g., SEQ ID NO: 464 or 433) and comprising a 5′ UTR sequence disclosed herein is formulated as an LNP. In some embodiments, the LNP composition comprises: (i) an ionizable amino lipid, e.g., an amino lipid; (ii) a sterol or other structural lipid; (iii) a non-cationic helper lipid or phospholipid; and (iv) a PEG-lipid. In another aspect, the LNP compositions of the disclosure are used in a method of treating argininosuccinic aciduria in a subject. In another aspect, an LNP composition comprising a polynucleotide disclosed herein encoding an Engager polypeptide, e.g., as described herein, can be administered with an additional agent, e.g., as described herein. 3′ UTR sequences 3′UTR sequences have been shown to influence translation, half-life, and subcellular localization of mRNAs (Mayr C., Cold Spring Harb. Persp. Biol.2019 Oct 1;11(10):a034728). Atty. Docket No.45817-0158WO1 Disclosed herein, inter alia, is a polynucleotide, e.g., mRNA, comprising an open reading frame (e.g., any one of SEQ ID NOs: 461, 463, or 465) encoding an Engager polypeptide (e.g., any one of SEQ ID NO: 394-446), which polynucleotide has a 3′ UTR that confers an increased half-life, increased expression and/or increased activity of the polypeptide encoded by said polynucleotide, or of the polynucleotide itself., a polynucleotide disclosed herein comprises: (a) a 5′-UTR (e.g., as described herein); (b) a coding region comprising a stop element (e.g., as described herein); and (c) a 3′-UTR (e.g., as provided in Table 11 or a variant or fragment thereof), and LNP compositions comprising the same. In some embodiments, the polynucleotide comprises a 3′-UTR comprising a sequence provided in Table 11 or a variant or fragment thereof. In some embodiments, the polynucleotide having a 3′ UTR sequence provided in Table 11 or a variant or fragment thereof, results in an increased half-life of the polynucleotide, e.g., about 1.5-10-fold increase in half-life of the polynucleotide. In some embodiments, the increase in half-life is about 1.5-, 2-, 3-, 4-, 5-, 6-, 7-, 8-, 9-, or 10-fold, or more. In some embodiments, the increase in half-life is about 1.5-fold or more. In some embodiments, the increase in half-life is about 2-fold or more. In some embodiments, the increase in half-life is about 3-fold or more. In some embodiments, the increase in half-life is about 4-fold or more. In some embodiments, the increase in half-life is about 5-fold or more. In some embodiments, the increase in half-life is about 6-fold or more. In some embodiments, the increase in half-life is about 7-fold or more. In some embodiments, the increase in half-life is about 8-fold. In some embodiments, the increase in half-life is about 9-fold or more. In some embodiments, the increase in half-life is about 10-fold or more. In some embodiments, the polynucleotide having a 3′ UTR sequence provided in Table 11 or a variant or fragment thereof, results in a polynucleotide with a mean half-life score of greater than 10. Atty. Docket No.45817-0158WO1 In some embodiments, the polynucleotide having a 3′ UTR sequence provided in Table 11 or a variant or fragment thereof, results in an increased level and/or activity, e.g., output, of the polypeptide encoded by the polynucleotide. In some embodiments, the increase is compared to an otherwise similar polynucleotide which does not have a 3′ UTR, has a different 3′ UTR, or does not have a 3′ UTR of Table 11 or a variant or fragment thereof. In some embodiments, the polynucleotide comprises a 3′ UTR sequence provided in Table 11 or a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to a 3′ UTR sequence provided in Table 11, or a fragment thereof. In some embodiments, the 3′ UTR comprises a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 500, SEQ ID NO: 501, SEQ ID NO: 502, SEQ ID NO: 503, SEQ ID NO: 504, SEQ ID NO: 505, SEQ ID NO: 506, SEQ ID NO: 507, SEQ ID NO: 508, SEQ ID NO: 509, SEQ ID NO: 510, SEQ ID NO: 511, SEQ ID NO: 512, SEQ ID NO: 513, SEQ ID NO: 514, SEQ ID NO: 515, SEQ ID NO: 525, SEQ ID NO: 526, and SEQ ID NO: 527. In some embodiments, the 3′ UTR comprises the sequence of SEQ ID NO: 500, or a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 500. In some embodiments, the 3′ UTR comprises the sequence of SEQ ID NO: 501, or a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 501. In some embodiments, the 3′ UTR comprises the sequence of SEQ ID NO: 502, or a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 502. In some embodiments, the 3′ UTR comprises the sequence of SEQ ID NO: 503, or a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 503. In some embodiments, the 3′ UTR comprises the sequence of SEQ ID NO: 504, or a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO :504. In some embodiments, the 3′ UTR comprises the sequence of SEQ ID NO: 505, or a sequence with at least 80%, 85%, Atty. Docket No.45817-0158WO1 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 505. In some embodiments, the 3′ UTR comprises the sequence of SEQ ID NO: 506, or a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 506. In some embodiments, the 3′ UTR comprises the sequence of SEQ ID NO: 507, or a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 507. In some embodiments, the 3′ UTR comprises the sequence of SEQ ID NO: 508, or a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 508. In some embodiments, the 3′ UTR comprises the sequence of SEQ ID NO: 509, or a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 509. In some embodiments, the 3′ UTR comprises the sequence of SEQ ID NO: 510, or a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 510. In some embodiments, the 3′ UTR comprises the sequence of SEQ ID NO: 511, or a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 511. In some embodiments, the 3′ UTR comprises the sequence of SEQ ID NO: 512, or a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 512. In some embodiments, the 3′ UTR comprises the sequence of SEQ ID NO: 513, or a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 513. In some embodiments, the 3′ UTR comprises the sequence of SEQ ID NO: 514, or a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 514. In some embodiments, the 3′ UTR comprises the sequence of SEQ ID NO: 515, or a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 515. In some embodiments, the 3′ UTR comprises the sequence of SEQ ID NO: 525, or a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 525. In some embodiments, the 3′ UTR comprises the sequence of SEQ ID NO: 526, or a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 526. In some embodiments, the 3′ UTR comprises the sequence of SEQ ID NO: 527, or a sequence with at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identity to SEQ ID NO: 527. Atty. Docket No.45817-0158WO1 Table 11 – 3’ UTR Sequences SEQ Sequence Sequence G C A CC U C C C U C U CC A C G G G G G C G G C G G U G C A G C A G C C

Atty. Docket No.45817-0158WO1 CUCCAUAAAGUAGGAAACACUACAGUGGUCUUUGAAUAAAGUCU U C C G G G A G G U G U U U G G U U U G G U C A C A U A G A U G G U U A G G U G C

Atty. Docket No.45817-0158WO1 527 B28 UAAAGCUAAACCUCACUCACGGCCACAUUGAGUGCCAGGCUCCG G U

Regions having a 5′ Cap The disclosure also includes a polynucleotide that comprises both a 5′ cap and a polynucleotide of the present disclosure (e.g., a polynucleotide comprising a nucleotide sequence encoding an Engager polypeptide to be expressed). The 5′ cap structure of a natural mRNA is involved in nuclear export, increasing mRNA stability and binds the mRNA Cap Binding Protein (CBP), which is responsible for mRNA stability in the cell and translation competency through the association of CBP with poly(A) binding protein to form the mature cyclic mRNA species. The cap further assists the removal of 5′ proximal introns during mRNA splicing. Endogenous mRNA molecules can be 5′-end capped generating a 5′-ppp-5′- triphosphate linkage between a terminal guanosine cap residue and the 5′-terminal transcribed sense nucleotide of the mRNA molecule. This 5′-guanylate cap can then be methylated to generate an N7-methyl-guanylate residue. The ribose sugars of the terminal and/or anteterminal transcribed nucleotides of the 5′ end of the mRNA can optionally also be 2′-O-methylated.5′-decapping through hydrolysis and cleavage of the guanylate cap structure can target a nucleic acid molecule, such as an mRNA molecule, for degradation. In some embodiments, the polynucleotides of the present disclosure (e.g., a polynucleotide comprising a nucleotide sequence encoding an Engager polypeptide) incorporate a cap moiety. In any of the embodiments disclosed herein, a 5’ terminal cap may terminate at the 3’ end with an A or G, even if not shown in the disclosure below. In some embodiments, polynucleotides of the present disclosure comprise a non-hydrolyzable cap structure preventing decapping and thus increasing mRNA half- Atty. Docket No.45817-0158WO1 life. Because cap structure hydrolysis requires cleavage of 5′-ppp-5′ phosphorodiester linkages, modified nucleotides can be used during the capping reaction. For example, a Vaccinia Capping Enzyme from New England Biolabs (Ipswich, MA) can be used with α-thio-guanosine nucleotides according to the manufacturer's instructions to create a phosphorothioate linkage in the 5′-ppp-5′ cap. Additional modified guanosine nucleotides can be used such as α-methyl-phosphonate and seleno-phosphate nucleotides. Additional modifications include, but are not limited to, 2′-O-methylation of the ribose sugars of 5′-terminal and/or 5′-anteterminal nucleotides of the polynucleotide (as mentioned above) on the 2′-hydroxyl group of the sugar ring. Multiple distinct 5′-cap structures can be used to generate the 5′-cap of a nucleic acid molecule, such as a polynucleotide that functions as an mRNA molecule. Cap analogs, which herein are also referred to as synthetic cap analogs, chemical caps, chemical cap analogs, or structural or functional cap analogs, differ from natural (i.e., endogenous, wild-type or physiological) 5′-caps in their chemical structure, while retaining cap function. Cap analogs can be chemically (i.e., non-enzymatically) or enzymatically synthesized and/or linked to the polynucleotides of the present disclosure. For example, the Anti-Reverse Cap Analog (ARCA) cap contains two guanines linked by a 5′-5′-triphosphate group, wherein one guanine contains an N7 methyl group as well as a 3′-O-methyl group (i.e., N7,3′-O-dimethyl-guanosine-5′- triphosphate-5′-guanosine (m⁷G-3′mppp-G; which can equivalently be designated 3′ O-Me-m⁷G(5′)ppp(5′)G). The 3′-O atom of the other, unmodified, guanine becomes linked to the 5′-terminal nucleotide of the capped polynucleotide. The N7- and 3′-O- methlyated guanine provides the terminal moiety of the capped polynucleotide. Another exemplary cap is mCAP, which is similar to ARCA but has a 2′-O- methyl group on guanosine (i.e., N7,2′-O-dimethyl-guanosine-5′-triphosphate-5′- guanosine, m⁷Gm-ppp-G). Atty. Docket No.45817-0158WO1 Another exemplary cap is m⁷G-ppp-Gm-A (i.e., N7,guanosine-5′-triphosphate- 2′-O-dimethyl-guanosine-adenosine). In some embodiments, the cap is a dinucleotide cap analog. As a non-limiting example, the dinucleotide cap analog can be modified at different phosphate positions with a boranophosphate group or a phosphoroselenoate group such as the dinucleotide cap analogs described in U.S. Patent No. US 8,519,110, the contents of which are herein incorporated by reference in its entirety. In some embodiments, the cap is a cap analog is a N7-(4-chlorophenoxyethyl) substituted dinucleotide form of a cap analog known in the art and/or described herein. Non-limiting examples of a N7-(4-chlorophenoxyethyl) substituted dinucleotide form of a cap analog include a N7-(4-chlorophenoxyethyl)- G(5′)ppp(5′)G and a N7-(4-chlorophenoxyethyl)-m^3′-OG(5′)ppp(5′)G cap analog (See, e.g., the various cap analogs and the methods of synthesizing cap analogs described in Kore et al. Bioorganic & Medicinal Chemistry 201321:4570-4574; the contents of which are herein incorporated by reference in its entirety). In some embodiments, a cap analog of the present disclosure is a 4-chloro/bromophenoxyethyl analog. Polynucleotides of the present disclosure can also be capped post-manufacture (whether IVT or chemical synthesis), using enzymes, in order to generate more authentic 5′-cap structures. As used herein, the phrase "more authentic" refers to a feature that closely mirrors or mimics, either structurally or functionally, an endogenous or wild type feature. That is, a "more authentic" feature is better representative of an endogenous, wild-type, natural or physiological cellular function and/or structure as compared to synthetic features or analogs, etc., of the prior art, or which outperforms the corresponding endogenous, wild-type, natural or physiological feature in one or more respects. Non-limiting examples of more authentic 5′cap structures of the present disclosure are those that, among other things, have enhanced binding of cap binding proteins, increased half-life, reduced susceptibility to 5′ endonucleases and/or reduced 5′decapping, as compared to synthetic 5′cap structures known in the art (or to a wild-type, natural or physiological 5′cap structure). For Atty. Docket No.45817-0158WO1 example, recombinant Vaccinia Virus Capping Enzyme and recombinant 2′-O- methyltransferase enzyme can create a canonical 5′-5′-triphosphate linkage between the 5′-terminal nucleotide of a polynucleotide and a guanine cap nucleotide wherein the cap guanine contains an N7 methylation and the 5′-terminal nucleotide of the mRNA contains a 2′-O-methyl. Such a structure is termed the Cap1 structure. This cap results in a higher translational-competency and cellular stability and a reduced activation of cellular pro-inflammatory cytokines, as compared, e.g., to other 5′cap analog structures known in the art. Cap structures include, but are not limited to, 7mG(5′)ppp(5′)N1pN2p (cap 0), 7mG(5′)ppp(5′)N1mpNp (cap 1), and 7mG(5′)- ppp(5′)N1mpN2mp (cap 2). As a non-limiting example, capping chimeric polynucleotides post- manufacture can be more efficient as nearly 100% of the chimeric polynucleotides can be capped. This is in contrast to ~80% when a cap analog is linked to a chimeric polynucleotide in the course of an in vitro transcription reaction. According to the present disclosure, 5′ terminal caps can include endogenous caps or cap analogs. According to the present disclosure, a 5′ terminal cap can comprise a guanine analog. Useful guanine analogs include, but are not limited to, inosine, N1-methyl-guanosine, 2′fluoro-guanosine, 7-deaza-guanosine, 8-oxo- guanosine, 2-amino-guanosine, LNA-guanosine, and 2-azido-guanosine. Also provided herein are exemplary caps including those that can be used in co-transcriptional capping methods for ribonucleic acid (RNA) synthesis, using RNA polymerase, e.g., wild type RNA polymerase or variants thereof, e.g., such as those variants described herein. In some embodiments, caps can be added when RNA is produced in a “one-pot” reaction, without the need for a separate capping reaction. Thus, the methods, in some embodiments, comprise reacting a polynucleotide template with an RNA polymerase variant, nucleoside triphosphates, and a cap analog under in vitro transcription reaction conditions to produce RNA transcript. As used here the term “cap” includes the inverted G nucleotide and can comprise one or more additional nucleotides 3’ of the inverted G nucleotide, e.g., 1, 2, Atty. Docket No.45817-0158WO1 3, or more nucleotides 3’ of the inverted G nucleotide and 5’ to the 5’ UTR, e.g., a 5’ UTR described herein. Exemplary caps comprise a sequence of GG, GA, or GGA, wherein the underlined, italicized G is an in inverted G nucleotide followed by a 5’-5’- triphosphate group. In some embodiments, a cap comprises a compound of Formula (C-I) a

;

a modified nucleobase; X2 is O, S(O)p, NR24 or CR25R26 in which p is 0, 1, or 2; Y0 is O or CR6R7; Y1 is O, S(O)_n, CR₆R₇, or NR₈, in which n is 0, 1 , or 2; each --- is a single bond or absent, wherein when each --- is a single bond, Yi is O, S(O)_n, CR₆R₇, or NR₈; and when each --- is absent, Y₁ is void; Y2 is (OP(O)R4)m in which m is 0, 1, or 2, or -O-(CR40R41)u-Q0-(CR42R43)v-, in which Q₀ is a bond, O, S(O)_r, NR₄₄, or CR₄₅R₄₆, r is 0, 1 , or 2, and each of u and v Atty. Docket No.45817-0158WO1 independently is 1, 2, 3 or 4; each R₂ and R₂' independently is halo, LNA, or OR₃; each R3 independently is H, C1-C6 alkyl, C2-C6 alkenyl, or C2-C6 alkynyl and R3, when being C1-C6 alkyl, C2-C6 alkenyl, or C2-C6 alkynyl, is optionally substituted with one or more of halo, OH and C1-C6 alkoxyl that is optionally substituted with one or more OH or OC(O)-C1-C6 alkyl; each R₄ and R₄' independently is H, halo, C₁-C₆ alkyl, OH, SH, SeH, or BH₃-; each of R6, R7, and R8, independently, is -Q1-T1, in which Q1 is a bond or C1-C3 alkyl linker optionally substituted with one or more of halo, cyano, OH and C₁-C₆ alkoxy, and T1 is H, halo, OH, COOH, cyano, or Rs1, in which Rs1 is C1-C3 alkyl, C2-C6 alkenyl, C₂-C₆ alkynyl, C₁- C₆ alkoxyl, C(O)O-C₁-C₆ alkyl, C₃-C₈ cycloalkyl, C₆-C₁₀ aryl, NR31R32, (NR31R32R33)⁺, 4 to 12- membered heterocycloalkyl, or 5- or 6- membered heteroaryl, and R_s1 is optionally substituted with one or more substituents selected from the group consisting of halo, OH, oxo, C1-C6 alkyl, COOH, C(O)O-C1- C₆ alkyl, cyano, C₁-C₆ alkoxyl, NR₃₁R₃₂, (NR₃₁R₃₂R₃₃)⁺, C₃-C₈ cycloalkyl, C₆- C10 aryl, 4 to 12-membered heterocycloalkyl, and 5- or 6-membered heteroaryl; each of R₁₀, R₁₁, R₁₂, R₁₃ R₁₄, and R₁₅, independently, is -Q₂-T₂, in which Q₂ is a bond or C1-C3 alkyl linker optionally substituted with one or more of halo, cyano, OH and C₁-C₆ alkoxy, and T₂ is H, halo, OH, NH₂, cyano, NO₂, N₃, R_s2, or OR_s2, in which R_s2 is C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C8 cycloalkyl, C6-C10 aryl, NHC(O)- C₁-C₆ alkyl, NR₃₁R₃₂, (NR₃₁R₃₂R₃₃)⁺, 4 to 12-membered heterocycloalkyl, or 5- or 6- membered heteroaryl, and Rs2 is optionally substituted with one or more substituents selected from the group consisting of halo, OH, oxo, C₁-C₆ alkyl, COOH, C(O)O-C₁- C6 alkyl, cyano, C1 - C6 alkoxyl, NR31R32, (NR31R32R33)⁺, C3-C8 cycloalkyl, C6- C10 aryl, 4 to 12-membered heterocycloalkyl, and 5- or 6- membered heteroaryl; or alternatively R₁₂ together with R₁₄ is oxo, or R₁₃ together with R₁₅ is oxo, each of R20, R21, R22, and R23 independently is -Q3-T3, in which Q3 is a bond or C1-C3 alkyl linker optionally substituted with one or more of halo, cyano, OH and C₁-C₆ alkoxy, and T3 is H, halo, OH, NH2, cyano, NO2, N3, RS3, or ORS3, in which RS3 is C1- C₆ alkyl, C₂-C₆ alkenyl, C₂-C₆ alkynyl, C₃-C₈ cycloalkyl, C₆-C₁₀ aryl, NHC(O)-C₁- C6 alkyl, mono-C1-C6 alkylamino, di-C1-C6 alkylamino, 4 to 12-membered Atty. Docket No.45817-0158WO1 heterocycloalkyl, or 5- or 6-membered heteroaryl, and Rs3 is optionally substituted with one or more substituents selected from the group consisting of halo, OH, oxo, C1-C6 alkyl, COOH, C(O)O-C1-C6 alkyl, cyano, C1-C6 alkoxyl, amino, mono-C1-C6 alkylamino, di-C1-C6 alkylamino, C3-C8 cycloalkyl, C6-C10 aryl, 4 to 12-membered heterocycloalkyl, and 5- or 6-membered heteroaryl; each of R24, R25, and R26 independently is H or C1-C6 alkyl; each of R₂₇ and R₂₈ independently is H or OR₂₉; or R₂₇ and R₂₈ together form O-R₃₀- O; each R29 independently is H, C1-C6 alkyl, C2-C6 alkenyl, or C2-C6 alkynyl and R29, when being C₁-C₆ alkyl, C₂-C₆ alkenyl, or C₂-C₆ alkynyl, is optionally substituted with one or more of halo, OH and C1-C6 alkoxyl that is optionally substituted with one or more OH or OC(O)-C₁-C₆ alkyl; R30 is C1-C6 alkylene optionally substituted with one or more of halo, OH and C1-C6 alkoxyl; each of R31, R32, and R33, independently is H, C1-C6 alkyl, C3-C8 cycloalkyl, C6-C10 aryl, 4 to 12-membered heterocycloalkyl, or 5- or 6-membered heteroaryl; each of R40, R41, R42, and R43 independently is H, halo, OH, cyano, N3, OP(O)R47R48, or C₁-C₆ alkyl optionally substituted with one or more OP(O)R₄₇R₄₈, or one R₄₁ and one R43, together with the carbon atoms to which they are attached and Q0, form C4- C₁₀ cycloalkyl, 4- to 14-membered heterocycloalkyl, C₆-C₁₀ aryl, or 5- to 14- membered heteroaryl, and each of the cycloalkyl, heterocycloalkyl, phenyl, or 5- to 6- membered heteroaryl is optionally substituted with one or more of OH, halo, cyano, N3, oxo, OP(O)R47R48, C1-C6 alkyl, C1-C6 haloalkyl, COOH, C(O)O-C1-C6 alkyl, C1- C₆ alkoxyl, C₁-C₆ haloalkoxyl, amino, mono-C₁-C₆ alkylamino, and di-C₁-C₆ alkylamino; R44 is H, C1-C6 alkyl, or an amine protecting group; each of R₄₅ and R₄₆ independently is H, OP(O)R₄₇R₄₈, or C₁-C₆ alkyl optionally substituted with one or more OP(O)R47R48, and each of R₄₇ and R₄₈, independently is H, halo, C₁-C₆ alkyl, OH, SH, SeH, or BH₃. Atty. Docket No.45817-0158WO1 It should be understood that a cap analog, as provided herein, may include any of the cap analogs described in international publication WO 2017/066797, published on 20 April 2017, incorporated by reference herein in its entirety. In some embodiments, the B₂ middle position can be a non-ribose molecule, such as arabinose. In some embodiments R2 is ethyl-based. Thus, in some embodiments, a cap comprises the following structure: (C-II).

Atty. Docket No.45817-0158WO1 In other embodiments, a cap comprises the following structure:

a cap (C-

Atty. Docket No.45817-0158WO1 In still other embodiments, a cap comprises the following structure:

. R is a methyl group (e.g., C₁ alkyl). In some embodiments, R is an ethyl group (e.g., C2 alkyl). In some embodiments, a cap comprises a sequence selected from the following sequences: GAA, GAC, GAG, GAU, GCA, GCC, GCG, GCU, GGA , GGC, GGG, GGU, GUA, GUC, GUG, and GUU. In some embodiments, a cap comprises GAA. In some embodiments, a cap comprises GAC. In some embodiments, a cap comprises GAG. In some embodiments, a cap comprises GAU. In some embodiments, a cap comprises GCA. In some embodiments, a cap comprises GCC. In some embodiments, a cap comprises GCG. In some embodiments, a cap comprises GCU. In some embodiments, a cap comprises GGA. In some embodiments, a cap comprises GGC. In some embodiments, a cap comprises GGG. In some embodiments, a cap comprises GGU. In some embodiments, a cap comprises GUA. In some embodiments, a cap comprises GUC. In some embodiments, a cap comprises GUG. In some embodiments, a cap comprises GUU. Atty. Docket No.45817-0158WO1 In some embodiments, a cap comprises a sequence selected from the following sequences: m⁷GpppApA, m⁷GpppApC, m⁷GpppApG, m⁷GpppApU, m⁷GpppCpA, m⁷GpppCpC, m⁷GpppCpG, m⁷GpppCpU, m⁷GpppGpA, m⁷GpppGpC, m⁷GpppGpG, m⁷GpppGpU, m⁷GpppUpA, m⁷GpppUpC, m⁷GpppUpG, and m⁷GpppUpU. In some embodiments, a cap comprises m⁷GpppApA. In some embodiments, a cap comprises m⁷GpppApC. In some embodiments, a cap comprises m⁷GpppApG. In some embodiments, a cap comprises m⁷GpppApU. In some embodiments, a cap comprises m⁷GpppCpA. In some embodiments, a cap comprises m⁷GpppCpC. In some embodiments, a cap comprises m⁷GpppCpG. In some embodiments, a cap comprises m⁷GpppCpU. In some embodiments, a cap comprises m⁷GpppGpA. In some embodiments, a cap comprises m⁷GpppGpC. In some embodiments, a cap comprises m⁷GpppGpG. In some embodiments, a cap comprises m⁷GpppGpU. In some embodiments, a cap comprises m⁷GpppUpA. In some embodiments, a cap comprises m⁷GpppUpC. In some embodiments, a cap comprises m⁷GpppUpG. In some embodiments, a cap comprises m⁷GpppUpU. A cap, in some embodiments, comprises a sequence selected from the following sequences: m⁷G3 _^OMepppApA, m⁷G3 _^OMepppApC, m⁷G3 _^OMepppApG, m⁷G_{3 ^OMe}pppApU, m⁷G_{3 ^OMe}pppCpA, m⁷G_{3 ^OMe}pppCpC, m⁷G_{3 ^OMe}pppCpG, m⁷G_{3 ^OMe}pppCpU, m⁷G_{3 ^OMe}pppGpA, m⁷G_{3 ^OMe}pppGpC, m⁷G_{3 ^OMe}pppGpG, m⁷G3 _^OMepppGpU, m⁷G3 _^OMepppUpA, m⁷G3 _^OMepppUpC, m⁷G3 _^OMepppUpG, and m⁷G3 ^OMepppUpU. In some embodiments, a cap comprises m⁷G3 _^OMepppApA. In some embodiments, a cap comprises m⁷G_{3 ^OMe}pppApC. In some embodiments, a cap comprises m⁷G_{3 ^OMe}pppApG. In some embodiments, a cap comprises m⁷G3 _^OMepppApU. In some embodiments, a cap comprises m⁷G3 _^OMepppCpA. In some embodiments, a cap comprises m⁷G3 _^OMepppCpC. In some embodiments, a cap comprises m⁷G_{3 ^OMe}pppCpG. In some embodiments, a cap comprises m⁷G_{3 ^OMe}pppCpU. In some embodiments, a cap comprises m⁷G_{3 ^OMe}pppGpA. In some Atty. Docket No.45817-0158WO1 embodiments, a cap comprises m⁷G_{3 ^OMe}pppGpC. In some embodiments, a cap comprises m⁷G_{3 ^OMe}pppGpG. In some embodiments, a cap comprises m⁷G3 _^OMepppGpU. In some embodiments, a cap comprises m⁷G3 _^OMepppUpA. In some embodiments, a cap comprises m⁷G3 _^OMepppUpC. In some embodiments, a cap comprises m⁷G_{3 ^OMe}pppUpG. In some embodiments, a cap comprises m⁷G_{3 ^OMe}pppUpU. In some embodiments, a cap comprises a sequence selected from the following sequences: m⁷G3 _^OMepppA2 _^OMepA, m⁷G3 _^OMepppA2 _^OMepC, m⁷G3 _^OMepppA2 _^OMepG, m⁷G_{3 ^OMe}pppA_{2 ^OMe}pU, m⁷G_{3 ^OMe}pppC_{2 ^OMe}pA, m⁷G_{3 ^OMe}pppC_{2 ^OMe}pC, m⁷G_{3 ^OMe}pppC_{2 ^OMe}pG, m⁷G_{3 ^OMe}pppC_{2 ^OMe}pU, m⁷G_{3 ^OMe}pppG_{2 ^OMe}pA, m⁷G3 _^OMepppG2 _^OMepC, m⁷G3 _^OMepppG2 _^OMepG, m⁷G3 _^OMepppG2 _^OMepU, m⁷G3 _^OMepppU2 _^OMepA, m⁷G3 _^OMepppU2 _^OMepC, m⁷G3 _^OMepppU2 _^OMepG, and m⁷G_{3 ^OMe}pppU_{2 ^OMe}pU. In some embodiments, a cap comprises m⁷G_{3 ^OMe}pppA_{2 ^OMe}pA. In some embodiments, a cap comprises m⁷G_{3 ^OMe}pppA_{2 ^OMe}pC. In some embodiments, a cap comprises m⁷G3 _^OMepppA2 _^OMepG. In some embodiments, a cap comprises m⁷G3 _^OMepppA2 _^OMepU. In some embodiments, a cap comprises m⁷G_{3 ^OMe}pppC_{2 ^OMe}pA. In some embodiments, a cap comprises m⁷G_{3 ^OMe}pppC_{2 ^OMe}pC. In some embodiments, a cap comprises m⁷G_{3 ^OMe}pppC_{2 ^OMe}pG. In some embodiments, a cap comprises m⁷G3 _^OMepppC2 _^OMepU. In some embodiments, a cap comprises m⁷G3 _^OMepppG2 _^OMepA. In some embodiments, a cap comprises m⁷G_{3 ^OMe}pppG_{2 ^OMe}pC. In some embodiments, a cap comprises m⁷G_{3 ^OMe}pppG_{2 ^OMe}pG. In some embodiments, a cap comprises m⁷G3 _^OMepppG2 _^OMepU. In some embodiments, a cap comprises m⁷G3 _^OMepppU2 _^OMepA. In some embodiments, a cap comprises m⁷G_{3 ^OMe}pppU_{2 ^OMe}pC. In some embodiments, a cap comprises m⁷G_{3 ^OMe}pppU_{2 ^OMe}pG. In some embodiments, a cap comprises m⁷G3 _^OMepppU2 _^OMepU. Atty. Docket No.45817-0158WO1 A cap, in still other embodiments, comprises a sequence selected from the following sequences: m⁷GpppA2 _^OMepA, m⁷GpppA2 _^OMepC, m⁷GpppA2 _^OMepG, m⁷GpppA2 _^OMepU, m⁷GpppC2 _^OMepA, m⁷GpppC2 _^OMepC, m⁷GpppC2 _^OMepG, m⁷GpppC_{2 ^OMe}pU, m⁷GpppG_{2 ^OMe}pA, m⁷GpppG_{2 ^OMe}pC, m⁷GpppG_{2 ^OMe}pG, m⁷GpppG_{2 ^OMe}pU, m⁷GpppU_{2 ^OMe}pA, m⁷GpppU_{2 ^OMe}pC, m⁷GpppU_{2 ^OMe}pG, and m⁷GpppU2 _^OMepU. In some embodiments, a cap comprises m⁷GpppA2 _^OMepA. In some embodiments, a cap comprises m⁷GpppA2 _^OMepC. In some embodiments, a cap comprises m⁷GpppA_{2 ^OMe}pG. In some embodiments, a cap comprises m⁷GpppA_{2 ^OMe}pU. In some embodiments, a cap comprises m⁷GpppC_{2 ^OMe}pA. In some embodiments, a cap comprises m⁷GpppC2 _^OMepC. In some embodiments, a cap comprises m⁷GpppC2 _^OMepG. In some embodiments, a trinucleotide cap comprises m⁷GpppC_{2 ^OMe}pU. In some embodiments, a cap comprises m⁷GpppG_{2 ^OMe}pA. In some embodiments, a cap comprises m⁷GpppG_{2 ^OMe}pC. In some embodiments, a cap comprises m⁷GpppG2 _^OMepG. In some embodiments, a cap comprises m⁷GpppG2 _^OMepU. In some embodiments, a cap comprises m⁷GpppU2 _^OMepA. In some embodiments, a cap comprises m⁷GpppU_{2 ^OMe}pC. In some embodiments, a cap comprises m⁷GpppU_{2 ^OMe}pG. In some embodiments, a cap comprises m⁷GpppU2 _^OMepU. In some embodiments, a cap comprises m⁷Gpppm⁶A_2’OmepG. In some embodiments, a cap comprises m⁷Gpppe⁶A2’OmepG. In some embodiments, a cap comprises GAG. In some embodiments, a cap comprises GCG. In some embodiments, a cap comprises GUG. In some embodiments, a cap comprises GGG. In some embodiments, a cap comprises any one of the following structures: Atty. Docket No.45817-0158WO1 ; or .

In some embodiments, the cap comprises ^m7GpppN1N2N3, where N1, N2, and N₃ are optional (i.e., can be absent or one or more can be present) and are independently a natural, a modified, or an unnatural nucleoside base. In some embodiments, ^m7G is further methylated, e.g., at the 3’ position. In some embodiments, the ^m7G comprises an O-methyl at the 3’ position. In some embodiments N₁, N₂, and N₃ if present, optionally, are independently an adenine, a uracil, a guanidine, a thymine, or a cytosine. In some embodiments, one or more (or all) of N₁, N₂, and N₃, if present, are methylated, e.g., at the 2’ position. In some Atty. Docket No.45817-0158WO1 embodiments, one or more (or all) of N1, N2, and N3, if present have an O-methyl at structure:

(C-IX) wherein B1, B2, and B3 are independently a natural, a modified, or an unnatural nucleoside based; and R1, R2, R3, and R4 are independently OH or O-methyl. In some embodiments, R3 is O-methyl and R4 is OH. In some embodiments, R3 and R4 are O- methyl. In some embodiments, R4 is O-methyl. In some embodiments, R1 is OH, R2 is OH, R₃ is O-methyl, and R₄ is OH. In some embodiments, R₁ is OH, R₂ is OH, R₃ is O-methyl, and R4 is O-methyl. In some embodiments, at least one of R1 and R2 is O-methyl, R₃ is O-methyl, and R₄ is OH. In some embodiments, at least one of R₁ and R is O-methy

2 l, R3 is O-methyl, and R4 is O-methyl. In some embodiments, B1, B3, and B3 are natural nucleoside bases. In some embodiments, at least one of B1, B2, and B3 is a modified or unnatural base. In some embodiments, at least one of B₁, B₂, and B₃ is N6-methyladenine. In some embodiments, B1 is adenine, cytosine, thymine, or uracil. In some embodiments, B1 is adenine, B₂ is uracil, and B₃ is adenine. In some embodiments, R₁ and R₂ are OH, R3 and R4 are O-methyl, B1 is adenine, B2 is uracil, and B3 is adenine. In some embodiments the cap comprises a sequence selected from the following sequences: GAAA, GACA, GAGA, GAUA, GCAA, GCCA, GCGA, GCUA, GGAA, GGCA, GGGA, GGUA, GUCA, and GUUA. In some embodiments Atty. Docket No.45817-0158WO1 the cap comprises a sequence selected from the following sequences: GAAG, GACG, GAGG, GAUG, GCAG, GCCG, GCGG, GCUG, GGAG, GGCG, GGGG, GGUG, a sequence GCAU, GUCU, GUGU, from the GCGC, GUUC. from the

m⁷G_{3 ^OMe}pppCpCpN, m⁷G_{3 ^OMe}pppCpGpN, m⁷G_{3 ^OMe}pppCpUpN, m⁷G_{3 ^OMe}pppGpApN, m⁷G_{3 ^OMe}pppGpCpN, m⁷G_{3 ^OMe}pppGpGpN, m⁷G3 _^OMepppGpUpN, m⁷G3 _^OMepppUpApN, m⁷G3 _^OMepppUpCpN, m⁷G3 _^OMepppUpGpN, and m⁷G3 _^OMepppUpUpN, where N is a natural, a modified, or an unnatural nucleoside base. A cap, in some embodiments, comprises a sequence selected from the following sequences: m⁷G3 _^OMepppA2 _^OMepApN, m⁷G3 _^OMepppA2 _^OMepCpN, m⁷G_{3 ^OMe}pppA_{2 ^OMe}pGpN, m⁷G_{3 ^OMe}pppA_{2 ^OMe}pUpN, m⁷G₃

_^OMepApN, m⁷G_{3 ^OMe}pppC_{2 ^OMe}pCpN, m⁷G_{3 ^OMe}pppC_{2 ^OMe}pGpN, m⁷G_{3 ^OMe}pppC_{2 ^OMe}pUpN, m⁷G3 _^OMepppG2 _^OMepApN, m⁷G3 _^OMepppG2 _^OMepCpN, m⁷G3 _^OMepppG2 _^OMepGpN, m⁷G3 _^OMepppG2 _^OMepUpN, m⁷G3 _^OMepppU2 _^OMepApN, m⁷G3 _^OMepppU2 _^OMepCpN, m⁷G_{3 ^OMe}pppU_{2 ^OMe}pGpN, and m⁷G_{3 ^OMe}pppU_{2 ^OMe}pUpN, where N is a natural, a modified, or an unnatural nucleoside base. A cap, in some embodiments, comprises a sequence selected from the following sequences: m⁷GpppA_{2 ^OMe}pApN, m⁷GpppA_{2 ^OMe}pCpN, m⁷GpppA_{2 ^OMe}pGpN, m⁷GpppA_{2 ^OMe}pUpN, m⁷GpppC_{2 ^OMe}pApN, m⁷GpppC2 _^OMepCpN, m⁷GpppC2 _^OMepGpN, m⁷GpppC2 _^OMepUpN, Atty. Docket No.45817-0158WO1 m⁷GpppG_{2 ^OMe}pApN, m⁷GpppG_{2 ^OMe}pCpN, m⁷GpppG_{2 ^OMe}pGpN, m⁷GpppG_{2 ^OMe}pUpN, m⁷GpppU_{2 ^OMe}pApN, m⁷GpppU_{2 ^OMe}pCpN, m⁷GpppU2 _^OMepGpN, and m⁷GpppU2 _^OMepUpN, where N is a natural, a modified, or an unnatural nucleoside base. A cap, in some embodiments, comprises a sequence selected from the following sequences: m⁷G3 _^OMepppA2 _^OMepA2 _^OMepN, m⁷G3 _^OMepppA2 _^OMepC2 _^OMepN, m⁷G3 _^OMepppA2 _^OMepG2 _^OMepN, m⁷G3 _^OMepppA2 _^OMepU2 _^OMepN, m⁷G_{3 ^OMe}pppC_{2 ^OMe}pA_{2 ^OMe}pN, m⁷G_{3 ^OMe}pppC_{2 ^OMe}pC_{2 ^OMe}pN, m⁷G_{3 ^OMe}pppC_{2 ^OMe}pG_{2 ^OMe}pN, m⁷G_{3 ^OMe}pppC_{2 ^OMe}pU_{2 ^OMe}pN, m⁷G3 _^OMepppG2 _^OMepA2 _^OMepN, m⁷G3 _^OMepppG2 _^OMepC2 _^OMepN, m⁷G3 _^OMepppG2 _^OMepG2 _^OMepN, m⁷G3 _^OMepppG2 _^OMepU2 _^OMepN, m⁷G_{3 ^OMe}pppU_{2 ^OMe}pA_{2 ^OMe}pN, m⁷G_{3 ^OMe}pppU_{2 ^OMe}pC_{2 ^OMe}pN, m⁷G_{3 ^OMe}pppU_{2 ^OMe}pG_{2 ^OMe}pN, and m⁷G_{3 ^OMe}pppU_{2 ^OMe}pU_{2 ^OMe}pN, where N is a natural, a modified, or an unnatural nucleoside base. A cap, in some embodiments, comprises a sequence selected from the following sequences: m⁷GpppA_{2 ^OMe}pA_{2 ^OMe}pN, m⁷GpppA_{2 ^OMe}pC_{2 ^OMe}pN, m⁷GpppA2 _^OMepG2 _^OMepN, m⁷GpppA2 _^OMepU2 _^OMepN, m⁷GpppC2 _^OMepA2 _^OMepN, m⁷GpppC2 _^OMepC2 _^OMepN, m⁷GpppC2 _^OMepG2 _^OMepN, m⁷GpppC2 _^OMepU2 _^OMepN, m⁷GpppG_{2 ^OMe}pA_{2 ^OMe}pN, m⁷GpppG_{2 ^OMe}pC_{2 ^OMe}pN, m⁷GpppG_{2 ^OMe}pG_{2 ^OMe}pN, m⁷GpppG_{2 ^OMe}pU_{2 ^OMe}pN, m⁷GpppU_{2 ^OMe}pA_{2 ^OMe}pN, m⁷GpppU_{2 ^OMe}pC_{2 ^OMe}pN, m⁷GpppU2 _^OMepG2 _^OMepN, and m⁷GpppU2 _^OMepU2 _^OMepN, where N is a natural, a modified, or an unnatural nucleoside base. In some embodiments, a cap comprises GGAG. In some embodiments, a cap comprises the following structure: Atty. Docket No.45817-0158WO1 (C‐X). In some embodiments, the mRNA comprises a m7GpppGmAG cap, as disclosed in Table 21 at the end of Example 3. Poly-A Tails In some embodiments, the polynucleotides of the present disclosure (e.g., a polynucleotide comprising a nucleotide sequence encoding an Engager polypeptide) further comprise a poly-A tail. In some embodiments, terminal groups on the poly-A tail can be incorporated for stabilization. In some embodiments, a poly-A tail comprises des-3′ hydroxyl tails. During RNA processing, a long chain of adenine nucleotides (poly-A tail) can be added to a polynucleotide (e.g., an mRNA molecule) in order to increase stability. Immediately after transcription, the 3′ end of the transcript can be cleaved to free a 3′ hydroxyl. Then poly-A polymerase adds a chain of adenine nucleotides to the RNA. The process, called polyadenylation, adds a poly-A tail that can be between, for example, approximately 80 to approximately 250 residues long, including approximately 80, 90, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240 or 250 residues long. In some embodiments, the poly-A tail is 100 nucleotides in length (SEQ ID NO: 528). PolyA tails can also be added after the construct is exported from the nucleus. Atty. Docket No.45817-0158WO1 According to the present disclosure, terminal groups on the poly A tail can be incorporated for stabilization. Polynucleotides of the present disclosure can include des-3′ hydroxyl tails. They can also include structural moieties or 2'-O-methyl modifications as taught by Junjie Li, et al. (Current Biology, vol.15, 1501–1507, August 23, 2005), the contents of which are incorporated herein by reference in its entirety). The polynucleotides of the present disclosure can be designed to encode transcripts with alternative polyA tail structures including histone mRNA. According to Norbury, "Terminal uridylation has also been detected on human replication- dependent histone mRNAs. The turnover of these mRNAs is thought to be important for the prevention of potentially toxic histone accumulation following the completion or inhibition of chromosomal DNA replication. These mRNAs are distinguished by their lack of a 3ʹ poly(A) tail, the function of which is instead assumed by a stable stem–loop structure and its cognate stem–loop binding protein (SLBP); the latter carries out the same functions as those of PABP on polyadenylated mRNAs" (Norbury, "Cytoplasmic RNA: a case of the tail wagging the dog," Nature Reviews Molecular Cell Biology; AOP, published online 29 August 2013; doi:10.1038/nrm3645), the contents of which are incorporated herein by reference in its entirety. Unique poly-A tail lengths provide certain advantages to the polynucleotides of the present disclosure. Generally, the length of a poly-A tail, when present, is greater than 30 nucleotides in length. In some embodiments, the poly-A tail is greater than 35 nucleotides in length (SEQ ID NO: 732) (e.g., at least or greater than about 35, 40, 45, 50, 55, 60, 70, 80, 90, 100, 120, 140, 160, 180, 200, 250, 300, 350, 400, 450, 500, 600, 700, 800, 900, 1,000, 1,100, 1,200, 1,300, 1,400, 1,500, 1,600, 1,700, 1,800, 1,900, 2,000, 2,500, and 3,000 nucleotides). In some embodiments, the polynucleotide or region thereof includes from about 30 to about 3,000 nucleotides (e.g., from 30 to 50, from 30 to 100, from 30 to 250, from 30 to 500, from 30 to 750, from 30 to 1,000, from 30 to 1,500, from 30 to Atty. Docket No.45817-0158WO1 2,000, from 30 to 2,500, from 50 to 100, from 50 to 250, from 50 to 500, from 50 to 750, from 50 to 1,000, from 50 to 1,500, from 50 to 2,000, from 50 to 2,500, from 50 to 3,000, from 100 to 500, from 100 to 750, from 100 to 1,000, from 100 to 1,500, 500 3,000, of the This feature

polynucleotides. In this context, the poly-A tail can be 10, 20, 30, 40, 50, 60, 70, 80, 90, or 100% greater in length than the polynucleotide or feature thereof. The poly-A tail can also be designed as a fraction of the polynucleotides to which it belongs. In this context, the poly-A tail can be 10, 20, 30, 40, 50, 60, 70, 80, or 90% or more of the total length of the construct, a construct region or the total length of the construct minus the poly-A tail. Further, engineered binding sites and conjugation of polynucleotides for Poly-A binding protein can enhance expression. Additionally, multiple distinct polynucleotides can be linked together via the PABP (Poly-A binding protein) through the 3′-end using modified nucleotides at the 3′-terminus of the poly-A tail. Transfection experiments can be conducted in relevant cell lines at and protein production can be assayed by ELISA at 12hr, 24hr, 48hr, 72hr and day 7 post-transfection. In some embodiments, the polynucleotides of the present disclosure are designed to include a polyA-G Quartet region. The G-quartet is a cyclic hydrogen bonded array of four guanine nucleotides that can be formed by G-rich sequences in both DNA and RNA. In some embodiments, the G-quartet is incorporated at the end Atty. Docket No.45817-0158WO1 of the poly-A tail. The resultant polynucleotide is assayed for stability, protein production and other parameters including half-life at various time points. It has been discovered that the polyA-G quartet results in protein production from an mRNA equivalent to at least 75% of that seen using a poly-A tail of 120 nucleotides alone (SEQ ID NO: 529). In some embodiments, the polyA tail comprises an alternative nucleoside, e.g., inverted thymidine. PolyA tails comprising an alternative nucleoside, e.g., inverted thymidine, may be generated as described herein. For instance, mRNA constructs may be modified by ligation to stabilize the poly(A) tail. Ligation may be performed using 0.5-1.5 mg/mL mRNA (5′ Cap1, 3′ A100), 50 mM Tris-HCl pH 7.5, 10 mM MgCl2, 1 mM TCEP, 1000 units/mL T4 RNA Ligase 1, 1 mM ATP, 20% w/v polyethylene glycol 8000, and 5:1 molar ratio of modifying oligo to mRNA. Modifying oligo has a sequence of 5’-phosphate-AAAAAAAAAAAAAAAAAAAA- (inverted deoxythymidine (idT) (SEQ ID NO:531)) (see below). Ligation reactions are mixed and incubated at room temperature (~22°C) for, e.g., 4 hours. Stable tail mRNA are purified by, e.g., dT purification, reverse phase purification, hydroxyapatite purification, ultrafiltration into water, and sterile filtration. The resulting stable tail-containing mRNAs contain the following structure at the 3’end, starting with the polyA region: A₁₀₀-UCUAGAAAAAAAAAAAAAAAAAAAA- inverted deoxythymidine (SEQ ID NO: 530). Modifying oligo to stabilize tail (5’-phosphate- AAAAAAAAAAAAAAAAAAAA-(inverted deoxythymidine)(SEQ ID NO: 531)):

Atty. Docket No.45817-0158WO1 In some instances, the polyA tail comprises A100-UCUAG-A20-inverted deoxy-thymidine (SEQ ID NO: 530). In some instances, the polyA tail consists of A100-UCUAG-A20-inverted deoxy-thymidine (SEQ ID NO: 530). Start codon region The invention also includes a polynucleotide that comprises both a start codon region and the polynucleotide described herein (e.g., a polynucleotide comprising a nucleotide sequence encoding an Engager polypeptide). In some embodiments, the polynucleotides of the present disclosure can have regions that are analogous to or function like a start codon region. In some embodiments, the translation of a polynucleotide can initiate on a codon that is not the start codon AUG. Translation of the polynucleotide can initiate on an alternative start codon such as, but not limited to, ACG, AGG, AAG, CTG/CUG, GTG/GUG, ATA/AUA, ATT/AUU, TTG/UUG (see Touriol et al. Biology of the Cell 95 (2003) 169-178 and Matsuda and Mauro PLoS ONE, 2010 5:11; the contents of each of which are herein incorporated by reference in its entirety). As a non-limiting example, the translation of a polynucleotide begins on the alternative start codon ACG. As another non-limiting example, polynucleotide translation begins on the alternative start codon CTG or CUG. As yet another non- Atty. Docket No.45817-0158WO1 limiting example, the translation of a polynucleotide begins on the alternative start codon GTG or GUG. Nucleotides flanking a codon that initiates translation such as, but not limited to, a start codon or an alternative start codon, are known to affect the translation efficiency, the length and/or the structure of the polynucleotide. (See, e.g., Matsuda and Mauro PLoS ONE, 20105:11; the contents of which are herein incorporated by reference in its entirety). Masking any of the nucleotides flanking a codon that initiates translation can be used to alter the position of translation initiation, translation efficiency, length and/or structure of a polynucleotide. In some embodiments, a masking agent can be used near the start codon or alternative start codon in order to mask or hide the codon to reduce the probability of translation initiation at the masked start codon or alternative start codon. Non-limiting examples of masking agents include antisense locked nucleic acids (LNA) polynucleotides and exon-junction complexes (EJCs) (See, e.g., Matsuda and Mauro describing masking agents LNA polynucleotides and EJCs (PLoS ONE, 20105:11); the contents of which are herein incorporated by reference in its entirety). In some embodiments, a masking agent can be used to mask a start codon of a polynucleotide in order to increase the likelihood that translation will initiate on an alternative start codon. In some embodiments, a masking agent can be used to mask a first start codon or alternative start codon in order to increase the chance that translation will initiate on a start codon or alternative start codon downstream to the masked start codon or alternative start codon. In some embodiments, a start codon or alternative start codon can be located within a perfect complement for a miRNA binding site. The perfect complement of a miRNA binding site can help control the translation, length and/or structure of the polynucleotide similar to a masking agent. As a non-limiting example, the start codon or alternative start codon can be located in the middle of a perfect complement for a miRNA binding site. The start codon or alternative start codon can be located after the first nucleotide, second nucleotide, third nucleotide, fourth nucleotide, fifth Atty. Docket No.45817-0158WO1 nucleotide, sixth nucleotide, seventh nucleotide, eighth nucleotide, ninth nucleotide, tenth nucleotide, eleventh nucleotide, twelfth nucleotide, thirteenth nucleotide, fourteenth nucleotide, fifteenth nucleotide, sixteenth nucleotide, seventeenth nucleotide, eighteenth nucleotide, nineteenth nucleotide, twentieth nucleotide or twenty-first nucleotide. In some embodiments, the start codon of a polynucleotide can be removed from the polynucleotide sequence in order to have the translation of the polynucleotide begin on a codon that is not the start codon. Translation of the polynucleotide can begin on the codon following the removed start codon or on a downstream start codon or an alternative start codon. In a non-limiting example, the start codon ATG or AUG is removed as the first 3 nucleotides of the polynucleotide sequence in order to have translation initiate on a downstream start codon or alternative start codon. The polynucleotide sequence where the start codon was removed can further comprise at least one masking agent for the downstream start codon and/or alternative start codons in order to control or attempt to control the initiation of translation, the length of the polynucleotide and/or the structure of the polynucleotide. In some embodiments, the poly(A) tail may be 80 nucleotides, 120 nucleotides, or 160 nucleotides in length. In some embodiments, the poly(A) tail may be 20, 40, 80, 100, 120, 140 or 160 nucleotides in length. In some embodiments, the mRNA comprises a 100 nucleotide poly(A) tail (SEQ ID NO: 730), as disclosed in Table 21 at the end of Example 3. Stop Codon Region The present disclosure also includes a polynucleotide that comprises both a stop codon region and the polynucleotide described herein (e.g., a polynucleotide comprising a nucleotide sequence encoding an Engager polypeptide). In some embodiments, the polynucleotides of the present disclosure can include at least two stop codons before the 3′ untranslated region (UTR). The stop codon can be selected from TGA, TAA and TAG in the case of DNA, or from UGA, UAA and UAG in the epent r n. nide y A nide y A Atty. Docket No.45817-0158WO1 tail, e.g., as described herein. In some embodiments, the polynucleotide further comprises a 3’ stabilizing region, e.g., as described herein. In some embodiments, a polynucleotide of the disclosure comprises (a) a 5’ UTR described in Table 10 or a variant or fragment thereof; (b) a coding region comprising a stop element provided herein; and (c) a 3’ UTR described in Table 11 or a variant or fragment thereof. In some embodiments, the polynucleotide further comprises a cap structure, e.g., as described herein, or a poly A tail, e.g., as described herein. In some embodiments, the polynucleotide further comprises a 3’ stabilizing region, e.g., as described herein. Table 12 – Exemplary 3’ UTR and stop element sequences SEQ ID Sequence information Sequence NO 532 3’ UTR with stop C11 UAGGGUUAAGCUGGAGCCUCGGUGGCCUAGCUUCUUGCCCC (underlined) UUGGGCCUCCCCCCAGCCCCUCCUCCCCUUCCUGCACCCGUA CCCCCGUGGUCUUUGAAUAAAGUCUGAGUGGGCGGC 533 3’ UTR with stop C10 UAAAGCUCCGCUGGAGCCUCGGUGGCCUAGCUUCUUGCCCC (underlined) UUGGGCCUCCCCCCAGCCCCUCCUCCCCUUCCUGCACCCGUA CCCCCGUGGUCUUUGAAUAAAGUCUGAGUGGGCGGC 534 3’ UTR with stop C9 UAAGCCCCUGCUGGAGCCUCGGUGGCCUAGCUUCUUGCCCC (underlined) UUGGGCCUCCCCCCAGCCCCUCCUCCCCUUCCUGCACCCGUA CCCCCGUGGUCUUUGAAUAAAGUCUGAGUGGGCGGC 535 3’ UTR with stop C8 UAAGCACCCGCUGGAGCCUCGGUGGCCUAGCUUCUUGCCCC (underlined) UUGGGCCUCCCCCCAGCCCCUCCUCCCCUUCCUGCACCCGUA CCCCCGUGGUCUUUGAAUAAAGUCUGAGUGGGCGGC 536 3’ UTR with stop C7 UAAGCCCCUCCGGGGGCCUCGGUGGCCUAGCUUCUUGCCCC (underlined) UUGGGCCUCCCCCCAGCCCCUCCUCCCCUUCCUGCACCCGUA CCCCCGUGGUCUUUGAAUAAAGUCUGAGUGGGCGGC 537 3’ UTR with stop C6 UAAGGCUAAGCUGGAGCCUCGGUGGCCUAGCUUCUUGCCCC (underlined) UUGGGCCUCCCCCCAGCCCCUCCUCCCCUUCCUGCACCCGUA CCCCCGUGGUCUUUGAAUAAAGUCUGAGUGGGCGGC 538 3’ UTR with stop C5 UAAGUCUCCGCUGGAGCCUCGGUGGCCUAGCUUCUUGCCCC (underlined) UUGGGCCUCCCCCCAGCCCCUCCUCCCCUUCCUGCACCCGUA CCCCCGUGGUCUUUGAAUAAAGUCUGAGUGGGCGGC 539 3’ UTR with stop C4 UAAAGCUAAGCUGGAGCCUCGGUGGCCUAGCUUCUUGCCCC (underlined) UUGGGCCUCCCCCCAGCCCCUCCUCCCCUUCCUGCACCCGUA CCCCCGUGGUCUUUGAAUAAAGUCUGAGUGGGCGGC 540 3’ UTR with stop C3 UAAGUCUAAGCUGGAGCCUCGGUGGCCUAGCUUCUUGCCCC (underlined) UUGGGCCUCCCCCCAGCCCCUCCUCCCCUUCCUGCACCCGUA CCCCCGUGGUCUUUGAAUAAAGUCUGAGUGGGCGGC 541 3’ UTR with C10 stop UAAAGCUCCAUAAAGUAGGAAACACUACAGCUGGAGCCUCG (underlined) GUGGCCUAGCUUCUUGCCCCUUGGGCCUCCAUAAAGUAGGA AACACUACAUCCCCCCAGCCCCUCCUCCCCUUCCUGCACCCG Atty. Docket No.45817-0158WO1 UACCCCCUCCAUAAAGUAGGAAACACUACAGUGGUCUUUGA AUAAAGUCUGAGUGGGCGGC 542 3’ UTR with C7 stop UAAGCCCCUCCGGGGUCCAUAAAGUAGGAAACACUACAGCC (underlined) UCGGUGGCCUAGCUUCUUGCCCCUUGGGCCUCCAUAAAGUA GGAAACACUACAUCCCCCCAGCCCCUCCUCCCCUUCCUGCAC CCGUACCCCCUCCAUAAAGUAGGAAACACUACAGUGGUCUU UGAAUAAAGUCUGAGUGGGCGGC 543 3’ UTR with C8 stop UAAAGCUCCCCGGGGUCCAUAAAGUAGGAAACACUACAGCC (underlined) UCGGUGGCCUAGCUUCUUGCCCCUUGGGCCUCCAUAAAGUA GGAAACACUACAUCCCCCCAGCCCCUCCUCCCCUUCCUGCAC CCGUACCCCCUCCAUAAAGUAGGAAACACUACAGUGGUCUU UGAAUAAAGUCUGAGUGGGCGGC Polynucleotide Comprising an mRNA Encoding an Engager polypeptide In some embodiments, a polynucleotide of the present disclosure, for example a polynucleotide comprising an mRNA nucleotide sequence encoding an Engager polypeptide, comprises from 5′ to 3′ end: (i) a 5′ cap such as provided above; (ii) a 5′ UTR, such as the sequences provided above; (iii) an ORF encoding a human Engager polypeptide; (iv) at least one stop codon; (v) a 3′ UTR, such as the sequences provided above; and (vi) a poly-A tail provided above. Methods of Making Polynucleotides The present disclosure also provides methods for making a polynucleotide of the present disclosure (e.g., a polynucleotide comprising a nucleotide sequence encoding an Engager polypeptide) or a complement thereof. In some aspects, a polynucleotide (e.g., a RNA, e.g., an mRNA) disclosed herein, and encoding an Engager polypeptide, can be constructed using in vitro transcription (IVT). In other aspects, a polynucleotide (e.g., a RNA, e.g., an mRNA) disclosed herein, and encoding an Engager polypeptide, can be constructed by chemical synthesis using an oligonucleotide synthesizer. Atty. Docket No.45817-0158WO1 In other aspects, a polynucleotide (e.g., a RNA, e.g., an mRNA) disclosed herein, and encoding an Engager polypeptide is made by using a host cell. In certain aspects, a polynucleotide (e.g., a RNA, e.g., an mRNA) disclosed herein, and encoding an Engager polypeptide is made by one or more combination of the IVT, chemical synthesis, host cell expression, or any other methods known in the art. Naturally occurring nucleosides, non-naturally occurring nucleosides, or combinations thereof, can totally or partially naturally replace occurring nucleosides present in the candidate nucleotide sequence and can be incorporated into a sequence- optimized nucleotide sequence (e.g., a RNA, e.g., an mRNA) encoding an Engager polypeptide. The resultant polynucleotides, e.g., mRNAs, can then be examined for their ability to produce protein and/or produce a therapeutic outcome. In Vitro Transcription / Enzymatic Synthesis The present disclosure also provides methods for making a polynucleotide disclosed herein or a complement thereof. In some aspects, a polynucleotide (e.g., an mRNA) disclosed herein can be constructed using in vitro transcription. In other aspects, a polynucleotide (e.g., an mRNA) disclosed herein can be constructed by chemical synthesis using an oligonucleotide synthesizer. In other aspects, a polynucleotide (e.g., an mRNA) disclosed herein is made by using a host cell. In certain aspects, a polynucleotide (e.g., an mRNA) disclosed herein is made by one or more combination of the IVT, chemical synthesis, host cell expression, or any other methods known in the art. Naturally occurring nucleosides, non-naturally occurring nucleosides, or combinations thereof, can totally or partially naturally replace occurring nucleosides present in the candidate nucleotide sequence and can be incorporated into a sequence- optimized nucleotide sequence (e.g., an mRNA) encoding an engager polypeptide. The resultant mRNAs can then be examined for their ability to produce a therapeutic outcome. Atty. Docket No.45817-0158WO1 While RNA can be made synthetically using methods well known in the art, In some embodiments an RNA transcript (e.g., mRNA transcript) is synthesized by contacting a DNA template with a RNA polymerase (e.g., a T7 RNA polymerase or a T7 RNA polymerase variant) under conditions that result in the production of RNA transcript. In some aspects, the present disclosure provides methods of performing an IVT (in vitro transcription) reaction, comprising contacting a DNA template with the RNA polymerase (e.g., a T7 RNA polymerase, such as a T7 RNA polymerase variant) in the presence of nucleoside triphosphates and buffer under conditions that result in the production of RNA transcripts. Other aspects of the present disclosure provide capping methods, e.g., co- transcriptional capping methods or other methods known in the art. In some embodiments, a capping method comprises reacting a polynucleotide template with a T7 RNA polymerase variant, nucleoside triphosphates, and a cap analog under in vitro transcription reaction conditions to produce RNA transcript. IVT conditions typically require a purified linear DNA template containing a promoter, nucleoside triphosphates, a buffer system that includes dithiothreitol (DTT) and magnesium ions, and a RNA polymerase. The exact conditions used in the transcription reaction depend on the amount of RNA needed for a specific application. Typical IVT reactions are performed by incubating a DNA template with a RNA polymerase and nucleoside triphosphates, including GTP, ATP, CTP, and UTP (or nucleotide analogs) in a transcription buffer. A RNA transcript having a 5 ^ terminal guanosine triphosphate is produced from this reaction. A deoxyribonucleic acid (DNA) is simply a nucleic acid template for RNA polymerase. A DNA template may include a polynucleotide encoding an Engager polypeptide. A DNA template, in some embodiments, includes a RNA polymerase promoter (e.g., a T7 RNA polymerase promoter) located 5' from and operably linked to polynucleotide encoding an Engager polypeptide. A DNA template may also Atty. Docket No.45817-0158WO1 include a nucleotide sequence encoding a polyadenylation (polyA) tail located at the 3' end of the gene of interest.

Atty. Docket No.45817-0158WO1 It should be understood that the term “nucleotide” includes naturally-occurring nucleotides, synthetic nucleotides and modified nucleotides, unless indicated otherwise. Examples of naturally-occurring nucleotides used for the production of RNA, e.g., in an IVT reaction, as provided herein include adenosine triphosphate (ATP), guanosine triphosphate (GTP), cytidine triphosphate (CTP), uridine triphosphate (UTP), and 5-methyluridine triphosphate (m⁵UTP). In some embodiments, adenosine diphosphate (ADP), guanosine diphosphate (GDP), cytidine diphosphate (CDP), and/or uridine diphosphate (UDP) are used. Examples of nucleotide analogs include, but are not limited to, antiviral nucleotide analogs, phosphate analogs (soluble or immobilized, hydrolyzable or non- hydrolyzable), dinucleotide, trinucleotide, e.g., a cap analog, or a precursor/substrate for enzymatic capping (vaccinia or ligase), a nucleotide labeled with a functional group to facilitate ligation/conjugation of cap or 5 ^ moiety (IRES), a nucleotide labeled with a 5 ^ PO₄ to facilitate ligation of cap or 5 ^ moiety, or a nucleotide labeled with a functional group/protecting group that can be chemically or enzymatically cleaved. Examples of antiviral nucleotide/nucleoside analogs include, but are not limited, to Ganciclovir, Entecavir, Telbivudine, Vidarabine and Cidofovir. Modified nucleotides may include modified nucleobases. For example, a RNA transcript (e.g., mRNA transcript) of the present disclosure may include a modified nucleobase selected from pseudouridine (ψ), 1-methylpseudouridine (m1ψ), 1- ethylpseudouridine, 2-thiouridine, 4’-thiouridine, 2-thio-1-methyl-1-deaza- pseudouridine, 2-thio-1-methyl-pseudouridine, 2-thio-5-aza-uridine , 2-thio- dihydropseudouridine, 2-thio-dihydrouridine, 2-thio-pseudouridine, 4-methoxy-2- thio-pseudouridine, 4-methoxy-pseudouridine, 4-thio-1-methyl-pseudouridine, 4-thio- pseudouridine, 5-aza-uridine, dihydropseudouridine, 5-methyluridine, 5- methoxyuridine (mo5U) and 2’-O-methyl uridine. In some embodiments, a RNA transcript (e.g., mRNA transcript) includes a combination of at least two (e.g., 2, 3, 4 or more) of the foregoing modified nucleobases. Atty. Docket No.45817-0158WO1 The nucleoside triphosphates (NTPs) as provided herein may comprise unmodified or modified ATP, modified or unmodified UTP, modified or unmodified GTP, and/or modified or unmodified CTP. In some embodiments, NTPs of an IVT reaction comprise unmodified ATP. In some embodiments, NTPs of an IVT reaction comprise modified ATP. In some embodiments, NTPs of an IVT reaction comprise unmodified UTP. In some embodiments, NTPs of an IVT reaction comprise modified UTP. In some embodiments, NTPs of an IVT reaction comprise unmodified GTP. In some embodiments, NTPs of an IVT reaction comprise modified GTP. In some embodiments, NTPs of an IVT reaction comprise unmodified CTP. In some embodiments, NTPs of an IVT reaction comprise modified CTP. The concentration of nucleoside triphosphates and cap analog present in an IVT reaction may vary. In some embodiments, NTPs and cap analog are present in the reaction at equimolar concentrations. In some embodiments, the molar ratio of cap analog (e.g., trinucleotide cap) to nucleoside triphosphates in the reaction is greater than 1:1. For example, the molar ratio of cap analog to nucleoside triphosphates in the reaction may be 2:1, 3:1, 4:1, 5:1, 6:1, 7:1, 8:1, 9:1, 10:1, 15:1, 20:1, 25:1, 50:1, or 100:1. In some embodiments, the molar ratio of cap analog (e.g., trinucleotide cap) to nucleoside triphosphates in the reaction is less than 1:1. For example, the molar ratio of cap analog (e.g., trinucleotide cap) to nucleoside triphosphates in the reaction may be 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:9, 1:10, 1:15, 1:20, 1:25, 1:50, or 1:100. The composition of NTPs in an IVT reaction may also vary. For example, ATP may be used in excess of GTP, CTP and UTP. As a non-limiting example, an IVT reaction may include 7.5 millimolar GTP, 7.5 millimolar CTP, 7.5 millimolar UTP, and 3.75 millimolar ATP. The same IVT reaction may include 3.75 millimolar cap analog (e.g., trinucleotide cap). In some embodiments, the molar ratio of G:C:U:A:cap is 1:1:1:0.5:0.5. In some embodiments, the molar ratio of G:C:U:A:cap is 1:1:0.5:1:0.5. In some embodiments, the molar ratio of G:C:U:A:cap is 1:0.5:1:1:0.5. In some embodiments, the molar ratio of G:C:U:A:cap is 0.5:1:1:1:0.5. Atty. Docket No.45817-0158WO1 In some embodiments, a RNA transcript (e.g., mRNA transcript) includes a modified nucleobase selected from pseudouridine (ψ), 1-methylpseudouridine (m¹ψ), 5-methoxyuridine (mo⁵U), 5-methylcytidine (m⁵C), α-thio-guanosine and α-thio- adenosine. In some embodiments, a RNA transcript (e.g., mRNA transcript) includes a combination of at least two (e.g., 2, 3, 4 or more) of the foregoing modified nucleobases. In some embodiments, a RNA transcript (e.g., mRNA transcript) includes pseudouridine (ψ). In some embodiments, a RNA transcript (e.g., mRNA transcript) includes 1-methylpseudouridine (m¹ψ). In some embodiments, a RNA transcript (e.g., mRNA transcript) includes 5-methoxyuridine (mo⁵U). In some embodiments, a RNA transcript (e.g., mRNA transcript) includes 5-methylcytidine (m⁵C). In some embodiments, a RNA transcript (e.g., mRNA transcript) includes α-thio-guanosine. In some embodiments, a RNA transcript (e.g., mRNA transcript) includes α-thio- adenosine. In some embodiments, the polynucleotide (e.g., RNA polynucleotide, such as mRNA polynucleotide) is uniformly modified (e.g., fully modified, modified throughout the entire sequence) for a particular modification. For example, a polynucleotide can be uniformly modified with 1-methylpseudouridine (m¹ψ), meaning that all uridine residues in the mRNA sequence are replaced with 1- methylpseudouridine (m¹ψ). Similarly, a polynucleotide can be uniformly modified for any type of nucleoside residue present in the sequence by replacement with a modified residue such as any of those set forth above. Alternatively, the polynucleotide (e.g., RNA polynucleotide, such as mRNA polynucleotide) may not be uniformly modified (e.g., partially modified, part of the sequence is modified). Each possibility represents a separate embodiment of the present disclosure. In some embodiments, the buffer system contains tris. The concentration of tris used in an IVT reaction, for example, may be at least 10 mM, at least 20 mM, at least 30 mM, at least 40 mM, at least 50 mM, at least 60 mM, at least 70 mM, at least Atty. Docket No.45817-0158WO1 80 mM, at least 90 mM, at least 100 mM or at least 110 mM phosphate. In some embodiments, the concentration of phosphate is 20-60 mM or 10-100 mM. In some embodiments, the buffer system contains dithiothreitol (DTT). The concentration of DTT used in an IVT reaction, for example, may be at least 1 mM, at least 5 mM, or at least 50 mM. In some embodiments, the concentration of DTT used in an IVT reaction is 1-50 mM or 5-50 mM. In some embodiments, the concentration of DTT used in an IVT reaction is 5 mM. In some embodiments, the buffer system contains magnesium. In some embodiments, the molar ratio of NTP to magnesium ions (Mg²⁺; e.g., MgCl2) present in an IVT reaction is 1:1 to 1:5. For example, the molar ratio of NTP to magnesium ions may be 1:1, 1:2, 1:3, 1:4 or 1:5. In some embodiments, the molar ratio of NTP plus cap analog (e.g., trinucleotide cap, such as GAG) to magnesium ions (Mg²⁺; e.g., MgCl₂) present in an IVT reaction is 1:1 to 1:5. For example, the molar ratio of NTP+trinucleotide cap (e.g., GAG) to magnesium ions may be 1:1, 1:2, 1:3, 1:4 or 1:5. In some embodiments, the buffer system contains Tris-HCl, spermidine (e.g., at a concentration of 1-30 mM), TRITON^® X-100 (polyethylene glycol p-(1,1,3,3- tetramethylbutyl)-phenyl ether) and/or polyethylene glycol (PEG). The addition of nucleoside triphosphates (NTPs) to the 3 ^ end of a growing RNA strand is catalyzed by a polymerase, such as T7 RNA polymerase, for example, any one or more of the T7 RNA polymerase variants (e.g., G47A) of the present disclosure. In some embodiments, the RNA polymerase (e.g., T7 RNA polymerase variant) is present in a reaction (e.g., an IVT reaction) at a concentration of 0.01 mg/ml to 1 mg/ml. For example, the RNA polymerase may be present in a reaction at a concentration of 0.01 mg/mL, 0.05 mg/ml, 0.1 mg/ml, 0.5 mg/ml or 1.0 mg/ml. In some embodiments, the polynucleotide of the present disclosure is an IVT polynucleotide. Traditionally, the basic components of an mRNA molecule include at least a coding region, a 5′UTR, a 3′UTR, a 5′ cap and a poly-A tail. The IVT Atty. Docket No.45817-0158WO1 polynucleotides of the present disclosure can function as mRNA but are distinguished from wild-type mRNA in their functional and/or structural design features which serve, e.g., to overcome existing problems of effective polypeptide production using nucleic-acid based therapeutics. The primary construct of an IVT polynucleotide comprises a first region of linked nucleotides that is flanked by a first flanking region and a second flaking region. This first region can include, but is not limited to, the encoded Engager polypeptide. The first flanking region can include a sequence of linked nucleosides which function as a 5’ untranslated region (UTR) such as the 5’ UTR of any of the nucleic acids encoding the native 5’ UTR of the polypeptide or a non-native 5’UTR such as, but not limited to, a heterologous 5’ UTR or a synthetic 5’ UTR. The IVT encoding an Engager polypeptide can comprise at its 5 terminus a signal sequence region encoding one or more signal sequences. The flanking region can comprise a region of linked nucleotides comprising one or more complete or incomplete 5′ UTRs sequences. The flanking region can also comprise a 5′ terminal cap. The second flanking region can comprise a region of linked nucleotides comprising one or more complete or incomplete 3′ UTRs which can encode the native 3’ UTR of an Engager polypeptide, or a non-native 3’ UTR such as, but not limited to, a heterologous 3’ UTR or a synthetic 3’ UTR. The flanking region can also comprise a 3′ tailing sequence. The 3’ tailing sequence can be, but is not limited to, a polyA tail, a polyA- G quartet and/or a stem loop sequence. Additional and exemplary features of IVT polynucleotide architecture and methods of making a polynucleotide are disclosed in PCT International application WO 2017/201325, filed on 18 May 2017, the entire contents of which are hereby incorporated by reference. Chemical synthesis Standard methods can be applied to synthesize an isolated polynucleotide sequence encoding an isolated polypeptide of interest, such as a polynucleotide of the present disclosure (e.g., a polynucleotide comprising a nucleotide sequence encoding Atty. Docket No.45817-0158WO1 an Engager polypeptide). For example, a single DNA or RNA oligomer containing a codon-optimized nucleotide sequence coding for the particular isolated polypeptide can be synthesized. In other aspects, several small oligonucleotides coding for portions of the desired polypeptide can be synthesized and then ligated. In some aspects, the individual oligonucleotides typically contain 5′ or 3′ overhangs for complementary assembly. A polynucleotide disclosed herein (e.g., a RNA, e.g., an mRNA) can be chemically synthesized using chemical synthesis methods and potential nucleobase substitutions known in the art. See, for example, International Publication Nos. WO2014093924, WO2013052523; WO2013039857, WO2012135805, WO2013151671; U.S. Publ. No. US20130115272; or U.S. Pat. Nos. US8999380 or US8710200, all of which are herein incorporated by reference in their entireties. Pharmaceutical Compositions Pharmaceutical compositions containing a bispecific engager as described herein or nucleic acid encoding the same can be prepared using methods known in the art. Pharmaceutical compositions described herein may contain a bispecific engager or nucleic acid encoding the same in combination with one or more pharmaceutically acceptable excipients. For instance, pharmaceutical compositions described herein can be prepared using physiologically acceptable carriers, excipients, or stabilizers (Remington's Pharmaceutical Sciences (19th ed., 1995), incorporated herein by reference), and in a desired form, e.g., in the form of lyophilized formulations or aqueous solutions. The compositions can also be prepared so as to contain the active agent (e.g., bispecific engager or nucleic acid encoding the same) at a desired concentration. For example, a pharmaceutical composition described herein may contain at least 10% (e.g., 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, 97%, 98%, 99%, 99.5%, 99.9%, or 100%) active agent by weight (w/w). Additionally, an active agent that can be incorporated into a pharmaceutical formulation can itself have a desired level of purity. For example, a polypeptide or Atty. Docket No.45817-0158WO1 nucleic acid described herein may be characterized by a certain degree of purity after isolating it from cell culture media or after chemical synthesis Pharmaceutical compositions can be prepared for storage as lyophilized formulations or aqueous solutions by mixing the active agent having

degree of purity with optional pharmaceutically acceptable carriers, excipients or stabilizers typically employed in the art, e.g., buffering agents, stabilizing agents, preservatives, isotonifiers, non-ionic detergents, antioxidants, and other miscellaneous additives. See, e.g., Remington's Pharmaceutical Sciences (19th ed., 1995), incorporated herein by reference). Such additives must be nontoxic to the recipients at the dosages and concentrations employed. Buffering agents Buffering agents help to maintain the pH in the range which approximates physiological conditions. Suitable buffering agents for use with the pharmaceutical compositions of the disclosure include both organic and inorganic acids and salts thereof, such as citrate buffers (e.g., monosodium citrate-disodium citrate mixture, citric acid-trisodium citrate mixture, citric acid-monosodium citrate mixture, etc.), succinate buffers (e.g., succinic acid- monosodium succinate mixture, succinic acid- sodium hydroxide mixture, succinic acid- disodium succinate mixture, etc.), tartrate buffers (e.g., tartaric acid-sodium tartrate mixture, tartaric acid-potassium tartrate mixture, tartaric acid-sodium hydroxide mixture, etc.), fumarate buffers (e.g., fumaric acid-monosodium fumarate mixture, fumaric acid- disodium fumarate mixture, monosodium fumarate-disodium fumarate mixture, etc.), gluconate buffers (e.g., gluconic acid-sodium gluconate mixture, gluconic acid-sodium hydroxide mixture, gluconic acid-potassium gluconate mixture, etc.), oxalate buffer (e.g., oxalic acid- sodium oxalate mixture, oxalic acid-sodium hydroxide mixture, oxalic acid-potassium oxalate mixture, etc.), lactate buffers (e.g., lactic acid-sodium lactate mixture, lactic acid-sodium hydroxide mixture, lactic acid-potassium lactate mixture, etc.), and acetate buffers (e.g., acetic acid-sodium acetate mixture, acetic acid-sodium Atty. Docket No.45817-0158WO1 hydroxide mixture, etc.). Additionally, phosphate buffers, histidine buffers, and trimethylamine salts such as Tris can be used. Preservatives Preservatives can be added to a composition described herein, for example, to inhibit microbial growth. Suitable preservatives for use with the pharmaceutical compositions of the disclosure include phenol, benzyl alcohol, meta-cresol, methyl paraben, propyl paraben, octadecyldimethylbenzyl ammonium chloride, benzalconium halides (e.g., chloride, bromide, and iodide), hexamethonium chloride, and alkyl parabens such as methyl or propyl paraben, catechol, resorcinol, cyclohexanol, and 3-pentanol. Isotonifiers, also known as “stabilizers,” can be added to ensure isotonicity of liquid compositions described herein and include polhydric sugar alcohols, for example trihydric or higher sugar alcohols, such as glycerin, arabitol, xylitol, sorbitol, and mannitol. Stabilizers refer to a broad category of excipients which can range in function from a bulking agent to an additive which solubilizes the therapeutic agent or helps to prevent denaturation or adherence to the container wall. Typical stabilizers can be polyhydric sugar alcohols; amino acids such as arginine, lysine, glycine, glutamine, asparagine, histidine, alanine, ornithine, L- leucine, 2-phenylalanine, glutamic acid, threonine, etc.; organic sugars or sugar alcohols, such as lactose, trehalose, stachyose, mannitol, sorbitol, xylitol, ribitol, myoinisitol, galactitol, glycerol and the like, including cyclitols such as inositol; polyethylene glycol; amino acid polymers; sulfur containing reducing agents, such as urea, glutathione, thioctic acid, sodium thioglycolate, thioglycerol, a- monothioglycerol and sodium thio sulfate; low molecular weight polypeptides (e.g., peptides of 10 residues or fewer); proteins such as HAS, BSA, MSA, gelatin, or immunoglobulins; hydrophilic polymers, such as polyvinylpyrrolidone monosaccharides, such as xylose, mannose, fructose, glucose; disaccharides such as lactose, maltose, sucrose and trisaccharides such as raffinose; and polysaccharides such as dextran. Detergents Atty. Docket No.45817-0158WO1 In some embodiments, non-ionic surfactants or detergents (also known as “wetting agents”) are added to the pharmaceutical composition, for example, to help solubilize the therapeutic agent as well as to protect the therapeutic agent against agitation-induced aggregation, which also permits the formulation to be exposed to shear surface stressed without causing denaturation of the protein. Suitable non-ionic surfactants include, for example and without limitation, polysorbates (20, 80, etc.), polyoxamers (184, 188 etc.), Pluronic polyols, polyoxyethylene sorbitan monoethers (TWEEN®-20, TWEEN®-80, etc.). Other pharmaceutical carriers Alternative pharmaceutically acceptable carriers that can be incorporated into a pharmaceutical composition described herein may include dextrose, sucrose, sorbitol, mannitol, starch, rubber arable, potassium phosphate, arginate, gelatin, potassium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrups, methyl cellulose, methylhydroxy benzoate, propylhydroxy benzoate, talc, magnesium stearate, and mineral oils, but not limited to. A pharmaceutical composition described herein may further include a lubricant, a humectant, a sweetener, a flavoring agent, an emulsifier, a suspending agent, and a preservative. Details of suitable pharmaceutically acceptable carriers and formulations can be found in Remington's Pharmaceutical Sciences (19th ed., 1995), which is incorporated herein by reference. Lipid Nanoparticle (LNP) Compositions The present disclosure provides LNP compositions with advantageous properties. The lipid nanoparticle compositions described herein may be used for the delivery of therapeutic and/or prophylactic agents, e.g., mRNAs, to mammalian cells or organs. For example, the lipid nanoparticles described herein have little or no immunogenicity. For example, the lipid compounds disclosed herein have a lower immunogenicity as compared to a reference lipid (e.g., MC3, KC2, or DLinDMA). For example, a formulation comprising a lipid disclosed herein and a therapeutic or prophylactic agent, e.g., mRNA, has an increased therapeutic index as compared to a Atty. Docket No.45817-0158WO1 corresponding formulation which comprises a reference lipid (e.g., MC3, KC2, or DLinDMA) and the same therapeutic or prophylactic agent. In some embodiments, the present application provides pharmaceutical compositions comprising: (a) a delivery agent comprising a lipid nanoparticle; and (b) a polynucleotide comprising a nucleotide sequence encoding a bispecific engager or VHH domain of the disclosure. Lipid Nanoparticles In some embodiments, polynucleotides of the present disclosure (e.g., mRNA) are included in a lipid nanoparticle (LNP). Lipid nanoparticles according to the present disclosure may comprise: (i) an ionizable amino lipid(e.g., an ionizable amino lipid); (ii) a sterol or other structural lipid; (iii) a non-cationic helper lipid or phospholipid; and (iv) a PEG-modified lipid. In some embodiments, lipid nanoparticles according to the present disclosure further comprise one or more polynucleotides of the present disclosure (e.g., mRNA). The lipid nanoparticles according to the present disclosure can be generated using components, compositions, and methods as are generally known in the art, see, for example PCT/US2016/052352; PCT/US2016/068300; PCT/US2017/037551; PCT/US2015/027400; PCT/US2016/047406; PCT/US2016000129; PCT/US2016/014280; PCT/US2016/014280; PCT/US2017/038426; PCT/US2014/027077; PCT/US2014/055394; PCT/US2016/52117; PCT/US2012/069610; PCT/US2017/027492; PCT/US2016/059575 and PCT/US2016/069491 all of which are incorporated by reference herein in their entirety. In some embodiments, the lipid nanoparticle comprises an ioniziable cationic lipid (e.g., an ionizable amino lipid) at a content of 20-60 mol.%, 25-60 mol.%, 30-60 mol.%, 35-60 mol.%, 40-60 mol.%, 45-60 mol.%, 20-55 mol.%, 25-55 mol.%, 30-55 mol.%, 35-55 mol.%, 40-55 mol.%, 45-55 mol.%, 20-50 mol.%, 25-50 mol.%, 30-50 mol.%, 35-50 mol.%, or 40-50 mol.%. For example, the lipid nanoparticle may Atty. Docket No.45817-0158WO1 comprise an ionizable cationic lipid (e.g., an ionizable amino lipid) at a content of 40- 50 mol.%, 45-50 mol.%, 45-46 mol.%, 46-47 mol.%, 47-48 mol.%, 48-49 mol.%, or 49-50 mol.%, for example about 45 mol.%, about 45.5 mol.%, about 46 mol.%, about 46.5 mol.%, about 47 mol.%, about 47.5 mol.%, about 48 mol.%, about 48.5 mol.%, about 49 mol.%, or about 49.5 mol.% ionizable cationic lipid (e.g., an ionizable amino lipid). In some embodiments, the lipid nanoparticle comprises a non-cationic helper lipid or phospholipid at a content of 5-25 mol.%. For example, the lipid nanoparticle may comprise a non-cationic helper lipid or phospholipid at a content of molar ratio of 5-25 mol.%, 5-20 mol.%, 5-15 mol.%, 10-25 mol.%, 10-20 mol.%, 10-15 mol.%, 5-6 mol.%, 6-7 mol.%, 7-8 mol.%, 8-9 mol.%, 9-10 mol.%, 10-11 mol.%, 11-12 mol.%, 12-13 mol.%, 13-14 mol.%, 14-15 mol.%, 10-14 mol.%, 10-13 mol.%, 10-12 mol.%, 10-11 mol.%, 9-15 mol.%, 9-14 mol.%, 9-13 mol.%, 9-12 mol.%, or 9-11 mol.% non-cationic lipid. In some embodiments, the lipid nanoparticle comprises a sterol or other structural lipid at a content molar ratio of 25-55 mol.%, 25-50 mol.%, 25-45 mol.%, 25-40 mol.%, 25-35 mol.%, 30-55 mol.%, 30-50 mol.%, 30-45 mol.%, 30-40 mol.%, 30-35 mol.%, 35-55 mol.%, 35-50 mol.%, 35-45 mol.%, 35-40 mol.%, 25-30 mol.%, 30-35 mol.%, 25-28 mol.%, 28-30 mol.%, 30-33 mol.%, 35-38 mol.%, 38-40 mol.%, 36-40 mol.%, 37-40 mol.%, 38-40 mol.%, 38-39 mol.%, 36-40 mol.%, 37-40 mol.%, 36-39 mol.%, or 37-39 mol.%. For example, the lipid nanoparticle may comprise a sterol or other structural lipid at a content of about 30 mol.%, about 30.5 mol.%, about 31.0 mol.%, about 31.5 mol.%, about 32.0 mol.%, about 32.5 mol.%, about 33.0 mol.%, about 33.5 mol.%, about 34.0 mol.%, about 34.5 mol.%, about 35.0 mol.%, about 35.5 mol.%, about 36.0 mol.%, about 36.5 mol.%, about 37.0 mol.%, about 37.5 mol.%, about 38.0 mol.%, about 38.5 mol.%, about 39.0 mol.%, about 39.5 mol.%, about 40.0 mol.%, about 40.5 mol.%, about 41.0 mol.%, about 41.5 mol.%, about 42.0 mol.%, about 42.5 mol.%, about 43.0 mol.%, about 43.5 mol.%, about 44.0 mol.%, about 44.5 mol.%, or about 45.0 mol.%. Atty. Docket No.45817-0158WO1 In some embodiments, the lipid nanoparticle comprises a PEG-modified lipid at a content of 0.5-15 mol.%, 1.0-15 mol.%, 1.5-15 mol.%, 2.0-15 mol.%, 2.5-15 mol.%, 3.0-15 mol.%, 3.5-15 mol.%, 4.0-15 mol.%, 4.5-15 mol.%, 5.0-15 mol.%, 10- 15 mol.%, 0.5-10 mol.%, 0.5-5 mol.%, 0.5-4.5 mol.%, 0.5-4.0 mol.%, 0.5-3.5 mol.%, 0.5-3.0 mol.%, 0.5-2.5 mol.%, 0.5-2.0 mol.%, 0.5-1.5 mol.%, 0.5-1.0 mol.%, 1.0-10 mol.%, 1.0-5 mol.%, 1.0-4.5 mol.%, 1.0-4.0 mol.%, 1.0-3.5 mol.%, 1.0-3.0 mol.%, 1.0-2.5 mol.%, 1.0-2.0 mol.%, 1.0-1.5 mol.%, 1.5-5.0 mol.%, 1.5-4.5 mol.%, 1.5-4.0 mol.%, 1.5-3.5 mol.%, 1.5-3.0 mol.%, 1.5-2.5 mol.%, 1.5-2.0 mol.%, 2.0-5.0 mol.%, 2.0-4.5 mol.%, 2.0-4.0 mol.%, 2.0-3.5 mol.%, 2.0-3.0 mol.%, or 2.0-2.5 mol.%. For example, the lipid nanoparticle may comprise a PEG-modified lipid at a content of a about 0.5 mol.%, about 1.0 mol.%, about 1.5 mol.%, about 2.0 mol.%, about 2.5 mol.%, about 3.0 mol.%, about 3.5 mol.%, about 4.0 mol.%, about 4.5 mol.%, about 5.0 mol.%, about 6.0 mol.%, about 7.0 mol.%, about 8.0 mol.%, about 9.0 mol.%, about 10.0 mol.%, or about 15.0 mol.%. In some embodiments, the lipid nanoparticle comprises: (i) 20 to 60 mol.% ionizable cationic lipid (e.g., ionizable amino lipid), (ii) 25 to 55 mol.% sterol or other structural lipid, (iii) 5 to 25 mol.% non-cationic lipid (e.g., phospholipid), and (iv) 0.5 to 15 mol.% PEG-modified lipid. In some embodiments, the lipid nanoparticle comprises: (i) 40 to 50 mol.% ionizable cationic lipid (e.g., ionizable amino lipid), (ii) 30 to 45 mol.% sterol or other structural lipid, (iii) 5 to 15 mol.% non-cationic lipid (e.g., phospholipid), and (iv) 1 to 5 mol.% PEG-modified lipid. In some embodiments, the lipid nanoparticle comprises: (i) 45 to 50 mol.% ionizable cationic lipid (e.g., ionizable amino lipid), (ii) 35 to 45 mol.% sterol or other structural lipid, (iii) 8 to 12 mol.% non-cationic lipid (e.g., phospholipid), and (iv) 1.5 to 3.5 mol.% PEG-modified lipid. Ionizable Amino Lipids Atty. Docket No.45817-0158WO1 In some embodiments, the lipid nanoparticle of the present disclosure comprises an ionizable cationic lipid (e.g., an ionizable amino lipid) that is a compound of Formula (I): (I) or its N-oxide, or a salt or isomer thereof, wherein R’^a is R’^branched; wherein R’^branched is: ; wherein denotes a point of attachment; wherein R^aα, R^aβ, R^aγ, and R^aδ are each independently selected from the group consisting of H, C_2-12 alkyl, and C_2-12 alkenyl; R² and R³ are each independently selected from the group consisting of C_1-14 alkyl and C_2-14 alkenyl; R⁴ is selected from the group consisting of -(CH₂)_nOH, wherein n is selected from the group consisting of 1, 2, 3, 4, and 5, and , wherein denotes a point of attachment; wherein R¹⁰ is N(R)₂; each R is independently selected from the group consisting of C_1-6 alkyl, C_2-3 alkenyl, and H; and n2 is selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10; each R⁵ is independently selected from the group consisting of C_1-3 alkyl, C_2-3 alkenyl, and H; each R⁶ is independently selected from the group consisting of C_1-3 alkyl, C_2-3 alkenyl, and H; Atty. Docket No.45817-0158WO1 M and M’ are each independently selected from the group consisting of -C(O)O- and -OC(O)-; R’ is a C_1-12 alkyl or C_2-12 alkenyl; l is selected from the group consisting of 1, 2, 3, 4, and 5; and m is selected from the group consisting of 5, 6, 7, 8, 9, 10, 11, 12, and 13. In some embodiments, in Formula (I), R’^a is R’^branched; R’^branched is ; denotes a point of attachment; R^aα, R^aβ, R^aγ, and R^aδ are each H; R² and R³ are each C_1-14 alkyl; R⁴ is -(CH₂)_nOH; n is 2; each R⁵ is H; each R⁶ is H; M and M’ are each -C(O)O-; R’ is a C_1-12 alkyl; l is 5; and m is 7. In some embodiments, in Formula (I), R’^a is R’^branched; R’^branched is ; denotes a point of attachment; R^aα, R^aβ, R^aγ, and R^aδ are each H; R² and R³ are each C_1-14 alkyl; R⁴ is -(CH₂)_nOH; n is 2; each R⁵ is H; each R⁶ is H; M and M’ are each -C(O)O-; R’ is a C_1-12 alkyl; l is 3; and m is 7. In some embodiments of the compounds of Formula (I), R’^a is R’^branched; R’^branched is ; denotes a point of attachment; R^aα is C2-12 alkyl; R^aβ, R^aγ, and R^aδ are each H; R² and R³ are each C_1-14 alkyl; R⁴ is ; R¹⁰ is NH(C1-6 alkyl); n2 is 2; R⁵ is H; each R⁶ is H; M and M’ are each -C(O)O-; R’ is a C_1-12 alkyl; l is 5; and m is 7. In some embodiments of the compounds of Formula (I), R’^a is R’^branched; Atty. Docket No.45817-0158WO1 R’^branched is ; denotes a point of attachment; R^aα, R^aβ, and R^aδ are each H; R^aγ is C_2-12 alkyl; R² and R³ are each C_1-14 alkyl; R⁴ is -(CH₂)_nOH; n is 2; each R⁵ is H; each R⁶ is H; M and M’ are each -C(O)O-; R’ is a C_1-12 alkyl; l is 5; and m is 7. In some embodiments, the compound of Formula (I) is selected from: , , and . In some embodiments, the compound of Formula (I) is: (Compound 2). In some embodiments, the compound of Formula (I) is: (Compound 3). In some embodiments, the compound of Formula (I) is: Atty. Docket No.45817-0158WO1 (Compound 4). In some aspects, the disclosure relates to a compound of Formula (Ia): (Ia) or its N-oxide, or a salt or isomer thereof, wherein R’^a is R’^branched; wherein R’^branched is: ; wherein denotes a point of attachment; wherein R^aβ, R^aγ, and R^aδ are each independently selected from the group consisting of H, C2-12 alkyl, and C2-12 alkenyl; R² and R³ are each independently selected from the group consisting of C_1-14 alkyl and C2-14 alkenyl; R⁴ is selected from the group consisting of -(CH2)nOH wherein n is selected from the group consisting of 1, 2, 3, 4, and 5, and , wherein denotes a point of attachment; wherein R¹⁰ is N(R)2; each R is independently selected from the group consisting of C1-6 alkyl, C2-3 alkenyl, and H; and n2 is selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10; each R⁵ is independently selected from the group consisting of C1-3 alkyl, C2-3 alkenyl, and H; each R⁶ is independently selected from the group consisting of C_1-3 alkyl, C2-3 alkenyl, and H; M and M’ are each independently selected from the group consisting of -C(O)O- and Atty. Docket No.45817-0158WO1 -OC(O)-; R’ is a C_1-12 alkyl or C_2-12 alkenyl; l is selected from the group consisting of 1, 2, 3, 4, and 5; and m is selected from the group consisting of 5, 6, 7, 8, 9, 10, 11, 12, and 13. the disclosure relates to a compound of Formula (Ib):

(Ib) or its N-oxide, or a salt or isomer thereof,

R’^branched is: ; wherein denotes a point of attachment; wherein R^aα, R^aβ, R^aγ, and R^aδ are each independently selected from the group consisting of H, C2-12 alkyl, and C2-12 alkenyl; R² and R³ are each independently selected from the group consisting of C_1-14 alkyl and C2-14 alkenyl; R⁴ is -(CH₂)_nOH, wherein n is selected from the group consisting of 1, 2, 3, 4, and 5;

each R⁵ is independently selected from the group consisting of C1-3 alkyl, C2-3 alkenyl, and H;

each R⁶ is independently selected from the group consisting of C1-3 alkyl, C2-3 alkenyl, and H; M and M’ are each independently selected from the group consisting of -C(O)O- and - OC(O)-; R’ is a C1-12 alkyl or C2-12 alkenyl; l is selected from the group consisting of 1, 2, 3, 4, and 5; and m is selected from the group consisting of 5, 6, 7, 8, 9, 10, 11, 12, and 13. Atty. Docket No.45817-0158WO1 In some embodiments of Formula (I) or (Ib), R’^a is R’^branched; R’^branched is

; denotes a point of attachment; R^aβ, R^aγ, and R^aδ are each H; R² and R³ are each C1-14 alkyl; R⁴ is -(CH2)nOH; n is 2; each R⁵ is H; each R⁶ is H; M and M’ are each -C(O)O-; R’ is a C1-12 alkyl; l is 5; and m is 7. In some embodiments of Formula (I) or (Ib), R’^a is R’^branched; R’^branched is ; denotes a point of attachment; R^aβ and R^aδ are each H; R^aγ is C_2-12 alkyl; R² and R³ are each C_1-14 alkyl; R⁴ is -(CH₂)_nOH; n is 2; each R⁵ is H; each R⁶ is H; M and M’ are each -C(O)O-; R’ is a C1-12 alkyl; l is 5; and m is 7. In some embodiments, the disclosure relates to a compound of Formula (Ic): (Ic) or its N-oxide, or a salt or isomer thereof, wherein R’^a is R’^branched; wherein

R’^branched is: ; wherein denotes a point of attachment; wherein R^aα, R^aβ, R^aγ, and R^aδ are each independently selected from the group consisting of H, C_2-12 alkyl, and C_2-12 alkenyl; R² and R³ are each independently selected from the group consisting of C1-14 alkyl and C_2-14 alkenyl; R⁴ is , wherein denotes a point of attachment; wherein R¹⁰ is N(R)2; each R is independently selected from the group consisting of Atty. Docket No.45817-0158WO1 C1-6 alkyl, C2-3 alkenyl, and H; n2 is selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10; each from the group consisting of C1-3 alkyl, C2-3 alkenyl, and H; each from the group consisting of C1-3 alkyl, C2-3

alkenyl, and H; M and M’ are each independently selected from the group consisting of -C(O)O- and - OC(O)-; R’ is a C_1-12 alkyl or C_2-12 alkenyl; l is 2, 3, 4, and 5; and m is 5, 6, 7, 8, 9, 10, 11, 12, and 13.

R’^branched; R’^branched is ; R^aγ, and R^aδ are each H; R^aα is C2-12 alkyl; R² and

R³ are each C1-14 alkyl; R⁴ is ; denotes a point of attachment; R¹⁰ is NH(C_1-6 alkyl); n2 is 2; each R⁵ is H; each R⁶ is H; M and M’ are each -C O-; R’ is a C1-12 l is 5; and m is 7. compound of Formula (Ic) is:

(Compound 3).

Atty. Docket No.45817-0158WO1 In some (II): thereof, wherein R’^a is

R’^branched is: and R’^cyclic is: ; and R’^b is:

wherein denotes a point of attachment; R^aγ and R^aδ are each independently selected from the group consisting of H, C1-12 alkyl, and is selected from the group consisting R^bγ and consisting of H, C_1-12 alkyl, and

is selected from the group consisting of C_1-12 alkyl and C_2-12 alkenyl; R² and R³ are each independently selected from the group consisting of C1-14 alkyl and C_2-14 R⁴ is from the

group consisting of 1, 2, 3, 4, and 5, and , of attachment; is independently selected from the group consisting of and n2 is selected from the group consisting of 1, 2, 3,

Atty. Docket No.45817-0158WO1 each Y^a is R*”^a and C2-15 alkenyl; and s is 2

m is selected from 1, 2, 3, 4, 5, 6, 7, 8, and 9; l is selected from 1, 2, 3, 4, 5, 6, 7, 8, and 9. In some aspects, the disclosure relates to a compound of Formula (II-a):

isomer thereof, wherein R’^a is R’^branched or R’^cyclic; wherein R’^branched is: and R’^b is: or ; wherein denotes a point of attachment; R^aγ and R^aδ are each independently selected from the group consisting of H, C1-12 alkyl, and C_2-12 alkenyl, wherein at least one of R^aγ and R^aδ is selected from the group consisting of C1-12 alkyl and C2-12 alkenyl; R^bγ and R^bδ are each independently selected from the group consisting of H, C1-12 alkyl, and C_2-12 alkenyl, wherein at least one of R^bγ and R^bδ is selected from the group consisting of C1-12 alkyl and C2-12 alkenyl; R² and R³ are each independently selected from the group consisting of C_1-14 alkyl and C2-14 alkenyl; R⁴ is selected from the group consisting of -(CH₂)_nOH wherein n is selected from the 5, and , of attachment; Atty. Docket No.45817-0158WO1 from the group consisting of C_1-6 the group consisting of 1, 2, 3, 4, 5,

each a 12 or 12 m is selected from 1, 2, 3, 4, 5, 6, 7, 8, and 9; l is selected from 1, 2, 3, 4, 5, 6, 7, 8, and 9. In some aspects, the disclosure relates to a compound of Formula (II-b): (II-b) or its N-oxide, or a salt or isomer thereof, wherein R’^a is R’^branched or R’^cyclic; wherein R’^branched is: and R’^b is: or ; wherein denotes a point of attachment; R^aγ and R^bγ are each independently selected from the group consisting of C_1-12 alkyl and C2-12 alkenyl; R² and R³ are each independently selected from the group consisting of C_1-14 alkyl and of -(CH₂)_nOH wherein n is selected from the

group consisting of 1, 2, 3, 4, and 5, and , wherein denotes a point of attachment; wherein R¹⁰ is N each R is selected from the group consisting of selected from the group consisting of 1, 2, 3,

C_2-12 alkenyl; Atty. Docket No.45817-0158WO1 m is selected from 1, 2, 3, 4, 5, 6, 7, 8, and 9; 6, 7, 8, and 9. relates to a compound of Formula (II-c):

a salt or isomer thereof,

wherein R’^branched is: and R’^b is: ; wherein denotes a point of attachment; wherein R^aγ is selected from the group consisting of C_1-12 alkyl and C_2-12 alkenyl; selected from the group consisting of C1-14 alkyl and consisting of -(CH2)nOH wherein n is selected from the

group consisting of 1, 2, 3, 4, and 5, and , wherein denotes a point of attachment; wherein R¹⁰ is N(R)₂; each R is independently selected from the group consisting of C_1-6 alkyl, C2-3 alkenyl, and H; and n2 is selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10; R’ is a C1-12 alkyl or C2-12 alkenyl; m is selected from 1, 2, 3, 4, 5, 6, 7, 8, and 9; l is selected from 1, 2, 3, 4, 5, 6, 7, 8, and 9. Atty. Docket No.45817-0158WO1 In some aspects, the

or isomer thereof, wherein R’^a is

wherein R’^branched is: and R’^b is: ; wherein denotes a point of attachment; wherein R^aγ and R^bγ are each independently selected from the group consisting of C_1- nOH wherein n is selected from the

group consisting of 1, 2, 3, 4, and 5, and , wherein denotes a point of attachment; selected from the group consisting of n2 is selected from the group consisting of 1, 2, 3, or C_2-12 alkenyl;

7, 8, and 9; l is selected from 1, 2, 3, 4, 5, 6, 7, 8, and 9. Formula (II-e): thereof,

Atty. Docket No.45817-0158WO1 wherein R’^branched is: and R’^b is: ; wherein denotes a point of attachment; wherein R^aγ is selected from the group consisting of C1-12 alkyl and C2-12 alkenyl; R² and R³ are each independently selected from the group consisting of C_1-14 alkyl and C2-14 alkenyl; R⁴ is -(CH2)nOH wherein n is selected from the group consisting of 1, 2, 3, 4, and 5; R’ is a C_1-12 alkyl or C_2-12 alkenyl; m is selected l is selected In some

, (II-a), (II-b), (II-c), (II-d), or (II-e), m and l are each independently selected from 4, 5, and 6. In some embodiments of the compound of Formula (II), (II-a), (II-b), (II-c), (II-d), or (II-e), m and l are each 5. In some embodiments of the compound of Formula (II), (II-a), (II-b), (II-c), (II-d), or (II-e), each R’ independently is a C1-12 alkyl. In some embodiments of the compound of Formula (II), (II-a), (II-b), (II-c), (II-d), or (II-e), each R’ independently is a C2-5 alkyl. In some embodiments of the compound of Formula (II), (II-a), (II-b), (II-c), (II-d), or (II-e), R’^b is: and R² and R³ are each independently a C1-14 alkyl. In some embodiments of the compound of Formula (II), (II-a), (II-b), (II-c), (II-d), or are each independently a C_6-10 alkyl. In some (II), (II-a), (II-b), (II-c), (II-d), or (II-e), each a C8 alkyl.

Atty. Docket No.45817-0158WO1 b), (II-c), is a C1-12

of the compound of Formula (II), (II-a), (II-b), (II-c), (II-d), or (II-e), R’^branched is: and R’^b is: , R^aγ is a C2-6 alkyl and R² and R³ are each independently a C6-10 alkyl. In some embodiments of the compound of Formula (II), (II-a), (II-b), (II-c), (II-d), or (II-e), R’^branched is: and R’^b is: , R^aγ is a C2-6 alkyl, and R² and R³ are each a C8 alkyl. In some embodiments of the compound of Formula (II), (II-a), (II-b), (II-c), (II-d), or (II-e), , and R^aγ and R^bγ are each a of Formula (II),

(II-a), , (II-c), (II-d), or (II-e), R’^branched is: , R’^b is: , and R^aγ and R^bγ are each a C_2-6 alkyl. In some embodiments of the compound of Formula (II), (II-a), (II-b), (II-c), (II-d), or (II-e), m and l are each independently selected from 4, 5, and 6 and each R’ independently is a C1-12 alkyl. In some embodiments of the compound of Formula (II), (II-a), (II-b), (II-c), (II-d), or (II-e), m and l are each 5 and each R’ independently is a C2-5 Atty. Docket No.45817-0158WO1 In some embodiments of the of ,

(II-d), or (II-e), is: , , m l are each independently selected from 4, 5, and 6, each R’ independently is a C1-12 alkyl, and R^aγ and R^bγ are each a C_1-12 alkyl. In some embodiments of the compound of Formula (II), (II-a), (II-b), (II-c), (II-d), or (II-e), R’^branched is: , R’^b is: , m and l are each 5, each R’ independently is a C2-5 alkyl, and R^aγ and R^bγ are In some

, (II-a), (II-b), (II-c), (II-d), or (II-e), R’^branched is: and R’^b is: , m and l are each independently selected from 4, 5, and 6, R’ is a C1-12 alkyl, R^aγ is a C1-12 alkyl and R² and R³ are each independently a C_6-10 alkyl. In some embodiments of the compound of Formula (II), (II-a), (II-b), (II-c), (II-d), or (II-e), R’^branched is: and R’^b is: , m and l are each 5, R’ is a C_2-5 alkyl, R^aγ is a C_2-6 alkyl, and R² and R³ are each a C8 alkyl. compound of Formula (II), (II-a), (II-b), (II-c), , wherein R¹⁰ is NH(C1-6 alkyl) and n2 is 2.

of Formula (II), (II-a), (II-b), (II-c), (II-d), or (II-e),

is

and n2 is 2. Atty. Docket No.45817-0158WO1 In some embodiments of the compound of Formula (II), (II-a), (II-b), (II-c), (II-d), or (II-e), R’^branched is: , R’^b is: , m and l are each is a C1-12 alkyl,

R^aγ and each a C_1-12 alkyl, and R⁴ is , wherein R¹⁰ is

NH(C1-6 , and n2 is 2. In some embodiments of the compound of Formula (II), (II-a), (II-b), (II-c), (II-d), or (II-e), R’^branched is: , R’^b is: , m and l are each 5, each R’ independently is a C2-5 alkyl, R^aγ and R^bγ are each a C2-6 alkyl, and R⁴ is , wherein R¹⁰ is NH(CH3) and , (II-c), l are each

, wherein R¹⁰ is NH(C1-6 alkyl) and n2 is 2. In some embodiments of the compound of Formula (II), (II-a), (II-b), (II-c), (II-d), or (II-e), R’^branched is: and R’^b is: m l R’ R^aγ a C_2-6 alkyl, R² and R³ Atty. Docket No.45817-0158WO1 are each a C8 alkyl, and R⁴ is , wherein R¹⁰ is NH(CH3) and n2 is 2. In some embodiments of the compound of Formula (II), (II-a), (II-b), (II-c), (II-d), or (II-e), R⁴ is -(CH2)nOH and n is 2, 3, or 4. In some embodiments of the compound of Formula (II), (II-a), (II-b), (II-c), (II-d), or (II-e), R⁴ is -(CH₂)_nOH and n is 2. In some embodiments of the compound of Formula (II), (II-a), (II-b), (II-c), , R’^b is: , m and l are 5, and 6, each R’ independently is a C_1-12 alkyl,

is -(CH2)nOH, and n is 2, 3, or 4. In some embodiments of the compound of Formula (II), (II-a), (II-b), (II-c), (II-d), or (II-e), R’^branched is:

m and l are each 5, each R’ is a C_2-5 alkyl, R^aγ and R^bγ are each a C_2-6 alkyl, R⁴ is -(CH₂)_nOH, and n

is 2. In some aspects, the disclosure relates to a compound of Formula (II-f): (II-f) or its N-oxide, or a salt or isomer thereof, wherein R’^a is R’^branched or R’^cyclic; wherein R’^branched is: and R’^b is: ; wherein denotes a point of attachment; Atty. Docket No.45817-0158WO1 R^aγ is a C1-12 alkyl; R² and R³ are each independently a C_1-14 alkyl; R⁴ is -(CH2)nOH wherein n is selected from the group consisting of 1, 2, 3, 4, and 5; R’ is a C1-12 alkyl; m is selected from 4, 5, and 6; and l is selected from 4, 5, and 6. In some the compound of Formula (II-f), m and l are each 5, and n is 2, 3, or 4.

In of the compound of Formula (II-f) R’ is a C2-5 alkyl, R^aγ is a C_2-6

and R³ are each a C_6-10 alkyl. In some embodiments of the compound of Formula (II-f), m and l are each 5, n is 2, R^aγ is a C2-6 alkyl, and R² and R³ are each a C6-10 alkyl.

In some aspects, the disclosure relates to a compound of Formula (II-g):

(II-g), wherein

group - n from the 4, and 5, and

a point of attachment, R¹⁰ is NH(C_1-6 alkyl), and n2 is selected

of 1, 2, and 3. Atty. Docket No.45817-0158WO1 In some aspects, the disclosure relates to a compound of Formula (II-h):

(II-h), wherein a C2-6 alkyl;

each R’ independently is a C2-5 alkyl; and R⁴ is selected from the group consisting of -(CH2)nOH wherein n is selected from the group consisting of 3, 4, and 5, and , wherein denotes a point of attachment, R¹⁰ is NH(C_1-6 alkyl), and n2 is selected from the group consisting of 1, 2, and 3. In some

, wherein R¹⁰ is NH(CH3) and n2 is 2. In some embodiments of the of Formula (CH2)

the disclosure relates to a compound having the Formula

:

, or a salt or isomer thereof, wherein Atty. Docket No.45817-0158WO1 R1, R2, R5 are independently selected from the group consisting of C5-20 alkyl, C_5-

-R”MR’, -R*YR”, -YR”, and -R*OR”; each M is independently selected from the group consisting of -C(O)O-, -OC(O)-, -OC(O)O-, -C(O)N(R’)-, -N(R’)C(O)-, -C(O)-, -C(S)-, -C(S)S-, -SC(S)-, group, and a heteroaryl group; from the group consisting of a bond, -CH₂-, , -OC(O)-, -C(O)-CH2-, -CH2

-C(O)-, -C(O)O-CH₂-, -OC(O)-CH₂-, -CH₂-C(O)O-, -CH₂-OC(O)-, -CH(OH)-, -C(S)-, from the group consisting of C1-12 alkyl and C2-12

alkenyl; each R is independently selected from the group consisting of C1-3 alkyl and a C3-6 carbocycle; each R’ is independently alkenyl, and H; and

each R” is independently selected from the group consisting of C3-12 alkyl and C3-12 alkenyl, and wherein: i) at least one of X¹, X², and X³ ii) at least one of R₁, R₂, R₃, R₄,

In some embodiments, R1, R2, R3, R4, X¹ is -CH2-; and X² and X³ are each -C(O)-.

In some embodiments, the compound of Formula (III) is:

(Compound VI), or a salt or isomer thereof.

Phospholipids Atty. Docket No.45817-0158WO1 The lipid composition of the lipid nanoparticle composition disclosed herein or

A phospholipid moiety can be group consisting of phosphatidyl choline, glycerol, phosphatidyl serine, phosphatidic and a

sphingomyelin.

acid, arachidic acid, arachidonic acid, eicosapentaenoic acid, behenic acid, a

to a membrane can allow one or more elements (e.g., a therapeutic agent) of a lipid- containing composition (e.g., LNPs) to pass through the membrane permitting, e.g., delivery of the one or more elements to a target tissue. Non-natural phospholipid species including natural species with modifications and substitutions including branching, oxidation, cyclization, and alkynes are also contemplated. For example, a phospholipid can be functionalized with or cross-linked to one or more alkynes replaced with a triple

can undergo a copper- upon exposure an reactions can be useful in functionalizing a lipid bilayer of a nanoparticle composition to facilitate membrane permeation or cellular recognition or in conjugating a nanoparticle composition to a useful component such as a targeting or imaging moiety Atty. Docket No.45817-0158WO1 Phospholipids include, but are not limited to, glycerophospholipids such as phosphatidylcholines, phosphatidylethanolamines, phosphatidylserines, phosphatidylinositols, phosphatidy glycerols, and phosphatidic acids. Phospholipids also include phosphosphingolipid, such as sphingomyelin. 1,2- 3- (DLPC), 3- , 1,2-

sn-glycero- 3- 3-phosphocholine (18:0 Diether

3-phosphocholine (OChemsPC), 1-hexadecyl-sn-glycero-3-phosphocholine (C16 Lyso PC), 1,2- dilinolenoyl-sn-glycero-3-phosphocholine,1,2-diarachidonoyl-sn-glycero-3- phosphocholine, 1,2-didocosahexaenoyl-sn-glycero-3-phosphocholine, 1,2- diphytanoyl-sn-glycero-3-phosphoethanolamine (ME 16.0 PE), 1,2-distearoyl-sn- glycero-3-phosphoethanolamine, 1,2-dilinoleoyl-sn-glycero-3-phosphoethanolamine, 1,2-dilinolenoyl-sn-glycero-3-phosphoethanolamine, 1,2-diarachidonoyl-sn-glycero- 3-phosphoethanolamine, 1,2-didocosahexaenoyl-sn-glycero-3-phosphoethanolamine, 1,2-dioleoyl-sn-glycero-3-phospho-rac-(1-glycerol) sodium salt (DOPG), sphingomyelin, and mixtures thereof. In certain embodiments, a phospholipid useful or potentially useful in the present disclosure is an analog or variant of DSPC. In certain embodiments, a phospholipid useful or potentially useful in the present disclosure is a compound of Formula (IV): (IV), or a salt thereof, wherein: each R¹ is independently optionally substituted alkyl; or optionally two R¹ are joined together with the intervening atoms to form optionally substituted monocyclic carbocyclyl or optionally substituted monocyclic heterocyclyl; or optionally three R¹ Atty. Docket No.45817-0158WO1 are joined together with the intervening atoms to form optionally substituted bicyclic

; each instance of L² is independently a bond or optionally substituted C1-6 alkylene, wherein one methylene unit of the optionally substituted C_1-6 alkylene is optionally replaced with O, N(R^N), S, C(O), C(O)N(R^N), NR^NC(O), C(O)O, OC(O), OC(O)O, OC(O)N(R^N), each instance of C_1-30 alkyl, optionally substituted C1-

alkynyl; optionally wherein one or more methylene units of R² are independently replaced with optionally substituted carbocyclylene, optionally substituted heterocyclylene, optionally substituted arylene, optionally substituted heteroarylene, N(R^N), O, S, C(O), - C(O)N(R^N), NR^NC(O), NR^NC(O)N(R^N), C(O)O, OC(O), OC(O)O, OC(O)N(R^N), - N(R^N), NR^NC(=NR^N), - (S), NR^NC(S)N(R^N), S(O), OS(O), - N(R^N)S(O), S(O)N(R^N), - S(O)2, N(R^N)S(O)2, S(O)2N(R^N), -

, , or (O)₂O; each instance of R^N is independently hydrogen, optionally substituted alkyl, or a nitrogen protecting group; Ring B is optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally and p is 1 or 2; provided that

Atty. Docket No.45817-0158WO1 , wherein ²

each instance of R is independently unsubstituted alkyl, unsubstituted alkenyl, or unsubstituted alkynyl. In some embodiments, the phospholipids may be one or more of the phospholipids described in U.S. Application No.62/520,530. Phospholipid head modifications In certain embodiments, a phospholipid useful or potentially useful in the (e.g., a modified choline head is DSPC, or in embodiments of at least one of compound of Formula (IV)

, , , ( )_v ( )_v ^N _O O O A ( )_v ( )_n P ( )_m , O , or a salt thereof, wherein: each t is independently 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10; each u is independently 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10; and each v is independently 1, 2, or 3. Atty. Docket No.45817-0158WO1 In certain embodiments, a compound of Formula (IV) is of Formula (IV-a): (IV-a), or a salt thereof. In certain embodiments, a phospholipid useful or potentially useful in the present disclosure comprises a cyclic moiety in place of the glyceride moiety. In certain embodiments, a phospholipid useful in the present disclosure is DSPC, or analog thereof, with a cyclic moiety in place of the glyceride moiety. In certain embodiments, the compound of Formula (IV) is of Formula (IV-b):

In certain embodiments, a phospholipid useful or potentially useful in the present disclosure comprises a modified tail. In certain embodiments, a phospholipid useful or potentially useful in the present disclosure is DSPC, or analog thereof, with a modified tail. As described herein, a “modified tail” may be a tail with shorter or longer aliphatic chains, aliphatic chains with branching introduced, aliphatic chains with substituents introduced, aliphatic chains wherein one or more methylenes are replaced by cyclic or heteroatom groups, or any combination thereof. For example, in certain embodiments, the compound of (IV) is of Formula (IV-a), or a salt thereof, wherein at least one instance of R² is each instance of R² is optionally substituted C_1- 30 alkyl, wherein one or more methylene units of R² are independently replaced with optionally substituted carbocyclylene, optionally substituted heterocyclylene, optionally substituted arylene, optionally substituted heteroarylene, N(R^N), O, S, - C(O), C(O)N(R^N), NR^NC(O), NR^NC(O)N(R^N), C(O)O, OC(O), OC(O)O, - OC(O)N(R^N), NR^NC(O)O, C(O)S, SC(O), C(=NR^N), C(=NR^N)N(R^N), NR^NC(=NR^N), NR^NC(=NR^N)N(R^N), C(S), C(S)N(R^N), NR^NC(S), NR^NC(S)N(R^N), S(O), OS(O), - Atty. Docket No.45817-0158WO1 S(O)O, OS(O)O, OS(O)2, S(O)2O, OS(O)2O, N(R^N)S(O), S(O)N(R^N), - N(R^N)S(O)N(R^N), OS(O)N(R^N), N(R^N)S(O)O, S(O)₂, N(R^N)S(O)₂, S(O)₂N(R^N), - N(R^N)S(O)2N(R^N), OS(O)2N(R^N), or N(R^N)S(O)2O. In certain embodiments, the compound of Formula (IV) is of Formula (IV-c): (IV-c), or a salt thereof, wherein: each x is independently an integer between 0-30, inclusive; and each instance is G is independently selected from the group consisting of optionally substituted carbocyclylene, optionally substituted heterocyclylene, optionally substituted arylene, optionally substituted heteroarylene, N(R^N), O, S, C(O), - C(O)N(R^N), NR^NC(O), NR^NC(O)N(R^N), C(O)O, OC(O), OC(O)O, OC(O)N(R^N), - NR^NC(O)O, C(O)S, SC(O), C(=NR^N), C(=NR^N)N(R^N), NR^NC(=NR^N), - NR^NC(=NR^N)N(R^N), C(S), C(S)N(R^N), NR^NC(S), NR^NC(S)N(R^N), S(O), OS(O), - S(O)O, OS(O)O, OS(O)₂, S(O)₂O, OS(O)₂O, N(R^N)S(O), S(O)N(R^N), - N(R^N)S(O)N(R^N), OS(O)N(R^N), N(R^N)S(O)O, S(O)2, N(R^N)S(O)2, S(O)2N(R^N), - N(R^N)S(O)₂N(R^N), OS(O)₂N(R^N), or N(R^N)S(O)₂O. Each possibility represents a separate embodiment of the present disclosure. In certain embodiments, a phospholipid useful or potentially useful in the present disclosure comprises a modified phosphocholine moiety, wherein the alkyl chain linking the quaternary amine to the phosphoryl group is not ethylene (e.g., n is not 2). Therefore, in certain embodiments, a phospholipid useful or potentially useful in the present disclosure is a compound of Formula (IV), wherein n is 1, 3, 4, 5, 6, 7, 8, 9, or 10. For example, in certain embodiments, a compound of Formula (IV) is of one of the following Formulae: 1 O R O O (⁾ A N P _m , ^R1 R¹ O , or a salt thereof. Atty. Docket No.45817-0158WO1 Alternative lipids In certain embodiments, a phospholipid useful or potentially useful in the present disclosure comprises a modified phosphocholine moiety, wherein the alkyl chain linking the quaternary amine to the phosphoryl group is not ethylene (e.g., n is not 2). Therefore, in certain embodiments, a phospholipid useful. In certain embodiments, an alternative lipid is used in place of a phospholipid of the present disclosure. In certain embodiments, an alternative lipid of the present disclosure is oleic acid. In certain embodiments, the alternative lipid is one of the following:

, ,

, , Atty. Docket No.45817-0158WO1 ,

, and . Structural Lipids of a pharmaceutical composition disclosed herein can lipids. As used herein, the term "structural lipid" containing sterol moieties.

lipids in the lipid nanoparticle may help mitigate aggregation of other lipids in the particle. Structural lipids can be selected from the group including but not limited to, cholesterol, fecosterol, sitosterol, ergosterol, campesterol, stigmasterol, brassicasterol, tomatidine, tomatine, ursolic acid, alpha- tocopherol, hopanoids, phytosterols, steroids, and mixtures thereof. In some embodiments, the structural lipid is a sterol. As defined herein, "sterols" are a subgroup of steroids consisting of steroid alcohols. In certain embodiments, the structural lipid is a steroid. In certain embodiments, the structural lipid is cholesterol. In certain embodiments, the structural lipid is an analog of cholesterol. In certain embodiments, the structural lipid is alpha-tocopherol. In some embodiments, the structural lipids may be one or more of the structural lipids described in U.S. Application No.62/520,530. Atty. Docket No.45817-0158WO1 can -

PEG-CerC14 or PEG-CerC20), PEG-modified dialkylamines and PEG-modified 1,2- diacyloxypropan-3-amines. Such lipids are also referred to as PEGylated lipids. For example, a PEG lipid can be PEG-c-DOMG, PEG-DMG, PEG-DLPE, PEG-DMPE, PEG-DPPC, or a PEG-DSPE lipid. In some embodiments, the PEG-lipid includes, but not limited to 1,2- sn- glycol (PEG-DMG), 1,2-distearoyl-sn- (polyethylene glycol)] (PEG-DSPE), PEG- PEG-dioleyl, PEG-distearyl, PEG-

, phosphatidylethanolamine (PEG- DPPE), or PEG-l,2-dimyristyloxlpropyl-3-amine (PEG-c-DMA). In some embodiments, the PEG-lipid is selected from the group consisting of a PEG-modified phosphatidylethanolamine, a PEG-modified phosphatidic acid, a PEG- modified ceramide, a PEG-modified dialkylamine, a PEG-modified diacylglycerol, a PEG-modified dialkylglycerol, and mixtures thereof. of the PEG-lipids includes those preferably from about C14 to about C16.

an mPEG-NH₂, has a size of about 1000, 2000, 5000, 10,000, 15,000 or 20,000 daltons. In some embodiments, the PEG- lipid is PEG_2k-DMG. In some embodiments, the lipid nanoparticles described herein can comprise a PEG lipid which is a non-diffusible PEG. Non-limiting examples of non-diffusible PEGs include PEG-DSG and PEG-DSPE. Atty. Docket No.45817-0158WO1 PEG-lipids are known in the art, such as those described in U.S. Patent No. 8,158,601 and International Publ. No. WO 2015/130584 A2, which are incorporated herein by reference in their entirety. In general, some of the other lipid components (e.g., PEG lipids) of various Formulae, described herein may be synthesized as described International Patent Application No. PCT/US2016/000129, filed December 10, 2016, entitled “Compositions and Methods for Delivery of Therapeutic Agents,” which is incorporated by reference in its entirety. The lipid component of a lipid nanoparticle composition may include one or more molecules comprising polyethylene glycol, such as PEG or PEG-modified lipids. Such species may be alternately referred to as PEGylated lipids. A PEG lipid is a lipid modified with polyethylene glycol. A PEG lipid may be selected from the non- limiting group including PEG-modified phosphatidylethanolamines, PEG-modified phosphatidic acids, PEG-modified ceramides, PEG-modified dialkylamines, PEG- modified diacylglycerols, PEG-modified dialkylglycerols, and mixtures thereof. For PEG-DMG, PEG-DLPE, PEG-DMPE, PEG-

lipids are a modified form of PEG DMG. PEG-DMG has the following structure: . In some embodiments, PEG lipids useful in the present disclosure can be PEGylated lipids described in International Publication No. WO2012099755, the contents of which is herein incorporated by reference in its entirety. Any of these exemplary PEG lipids described herein may be modified to comprise a hydroxyl group on the PEG chain. In certain embodiments, the PEG lipid is a PEG-OH lipid. As generally defined herein, a “PEG-OH lipid” (also referred to herein as “hydroxy- Atty. Docket No.45817-0158WO1 PEGylated lipid”) is a PEGylated lipid having one or more hydroxyl (–OH) groups on the lipid. In certain embodiments, the PEG-OH lipid includes one or more hydroxyl groups on the PEG chain. In certain embodiments, a PEG-OH or hydroxy-PEGylated lipid comprises an –OH group at the terminus of the PEG chain. Each possibility represents a separate embodiment of the present disclosure. In certain embodiments, a PEG lipid useful in the present disclosure is a compound of Formula (V). Provided herein are compounds of Formula (V):

group; r is an integer between 1 and 100, inclusive; L¹ is optionally substituted C1-10 alkylene, wherein at least one methylene of the optionally substituted C_1-10 alkylene is independently replaced with optionally substituted carbocyclylene, optionally substituted heterocyclylene, optionally substituted arylene, optionally substituted heteroarylene, O, N(R^N), S, C(O), - C(O)N(R^N), NR^NC(O), C(O)O, OC(O), OC(O)O, OC(O)N(R^N), NR^NC(O)O, or - NR^NC(O)N(R^N); D is a moiety obtained by click chemistry or a moiety cleavable under physiological conditions; m is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10; A is of the Formula: or ; each instance of L² is independently a bond or optionally substituted C1-6 alkylene, wherein one methylene unit of the optionally substituted C_1-6 alkylene is optionally (O)O, OC

one or more methylene units of R² are independently replaced with optionally Atty. Docket No.45817-0158WO1 - C , - (O), - S N , - N or a p (i.e., R³ Formula a R³ R^O r is

R⁵ is optionally substituted C10-40 alkyl, optionally substituted C10-40 alkenyl, or optionally substituted C_10-40 alkynyl; and optionally one or more methylene groups of R⁵ are replaced with optionally substituted carbocyclylene, optionally substituted Atty. Docket No.45817-0158WO1 heterocyclylene, optionally substituted arylene, optionally substituted heteroarylene, N(R^N), O, S, C(O), C(O)N(R^N), NR^NC(O), NR^NC(O)N(R^N), C(O)O, OC(O), - OC(O)O, OC(O)N(R^N), NR^NC(O)O, C(O)S, SC(O), C(=NR^N), C(=NR^N)N(R^N), - NR^NC(=NR^N), NR^NC(=NR^N)N(R^N), C(S), C(S)N(R^N), NR^NC(S), NR^NC(S)N(R^N), - S(O), OS(O), S(O)O, OS(O)O, OS(O)2, S(O)2O, OS(O)2O, N(R^N)S(O), S(O)N(R^N), N(R^N)S(O)N(R^N), OS(O)N(R^N), N(R^N)S(O)O, S(O)2, N(R^N)S(O)2, S(O)2N(R^N), - N(R^N)S(O)₂N(R^N), OS(O)₂N(R^N), or N(R^N)S(O)₂O; and each instance of R^N is independently hydrogen, optionally substituted alkyl, or a nitrogen protecting group. In certain embodiments, the compound of Formula (VI) is of Formula (VI- OH): (VI-OH), or a salt thereof. In some embodiments, r is 45. In yet other embodiments the compound of Formula (VI) is: or a salt thereof. In one embodiment, r is 40-50. In some embodiments, the compound of Formula (VI) is (Compound I). In some aspects, the lipid composition of the pharmaceutical compositions disclosed herein does not comprise a PEG-lipid. In some embodiments, the PEG-lipids may be one or more of the PEG lipids described in U.S. Application No.62/520,530. Atty. Docket No.45817-0158WO1 In some embodiments, a PEG lipid of the present disclosure comprises a PEG- modified phosphatidylethanolamine, a PEG-modified phosphatidic acid, a PEG- modified ceramide, a PEG-modified dialkylamine, a PEG-modified diacylglycerol, a PEG-modified dialkylglycerol, and mixtures thereof. In some embodiments, the PEG- modified lipid is PEG-DMG, PEG-c-DOMG (also referred to as PEG-DOMG), PEG- DSG and/or PEG-DPG. In some embodiments, a LNP of the present disclosure comprises an ionizable cationic lipid of any of Formula I, II, or III, a phospholipid comprising DSPC, a structural lipid, and a PEG lipid comprising PEG-DMG. In some embodiments, a LNP of the present disclosure comprises an ionizable cationic lipid of any of Formula I, II, or III, a phospholipid comprising DSPC, a structural lipid, and a PEG lipid comprising a compound having Formula VI. In some embodiments, a LNP of the present disclosure comprises an ionizable cationic lipid of Formula I, II or III, a phospholipid comprising a compound having Formula IV, a structural lipid, and the PEG lipid comprising a compound having Formula V or VI. In some embodiments, a LNP of the present disclosure comprises an ionizable cationic lipid of Formula I, II or III, a phospholipid comprising a compound having Formula IV, a structural lipid, and the PEG lipid comprising a compound having Formula V or VI. In some embodiments, a LNP of the present disclosure comprises an ionizable cationic lipid of Formula I, II or III, a phospholipid having Formula IV, a structural lipid, and a PEG lipid comprising a compound having Formula VI. In some embodiments, a LNP of the present disclosure comprises an ionizable cationic lipid of: Atty. Docket No.45817-0158WO1 , and a PEG lipid comprising Formula VI. comprises an ionizable

, and an alternative lipid comprising oleic acid. In some embodiments, a LNP of the present disclosure comprises an ionizable cationic lipid of: , an alternative lipid comprising oleic acid, a structural lipid comprising cholesterol, and a PEG lipid comprising a compound having Formula VI. In some embodiments, a LNP of the present disclosure comprises an ionizable

, a phospholipid comprising DOPE, a structural lipid comprising cholesterol, and a PEG lipid comprising a compound having Formula VI. Atty. Docket No.45817-0158WO1 In some embodiments, a LNP of the present disclosure comprises an ionizable cationic lipid of: , a phospholipid cholesterol, and a PEG lipid comprising a

In some embodiments, a LNP of the present disclosure comprises an N:P ratio of from about 2:1 to about 30:1. In some embodiments, a LNP of the present disclosure comprises an N:P ratio of about 6:1. In some embodiments, a LNP of the present disclosure comprises an N:P ratio of about 3:1. In some embodiments, a LNP of the present disclosure comprises a wt/wt ratio of the ionizable cationic lipid component to the RNA of from about 10:1 to about 100:1. In some embodiments, a LNP of the present disclosure comprises a wt/wt ratio of the ionizable cationic lipid component to the RNA of about 20:1. In some embodiments, a LNP of the present disclosure comprises a wt/wt ratio of the ionizable cationic lipid component to the RNA of about 10:1. In some embodiments, a LNP of the present disclosure has a mean diameter from about 50nm to about 150nm. In some embodiments, a LNP of the present disclosure has a mean diameter from about 70nm to about 120nm. Atty. Docket No.45817-0158WO1 As used herein, the term "alkyl", "alkyl group", or "alkylene" means a linear or saturated including one or more carbon atoms (e.g., one, two, three,

nine, ten, eleven, twelve, thirteen, fourteen, fifteen, sixteen, seventeen, eighteen, nineteen, twenty, or more carbon atoms), which is optionally substituted. The notation "C1-14 alkyl" means an optionally substituted linear or branched, saturated hydrocarbon including 1-14 carbon atoms. Unless otherwise specified, an alkyl group described herein refers to both unsubstituted and substituted alkyl groups. the term "alkenyl", "alkenyl group", or "alkenylene" means a linear

including two or more carbon atoms (e.g., two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, fifteen, sixteen, seventeen, eighteen, nineteen, twenty, or more carbon atoms) and at least one double bond, which is optionally substituted. The notation "C2-14 alkenyl" means an optionally substituted linear or branched hydrocarbon including 2-14 carbon atoms and at least one carbon-carbon double bond. An alkenyl group may include one, two, three, four, or more carbon-carbon double bonds. For example, C18 alkenyl may include one or more double bonds. A C₁₈ alkenyl group including two double bonds may be a linoleyl group. Unless otherwise specified, an alkenyl group described herein refers to both unsubstituted and substituted alkenyl groups. As used herein, the term "alkynyl", "alkynyl group", or "alkynylene" means a linear or branched hydrocarbon including two or more carbon atoms (e.g., two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, fifteen, sixteen, seventeen, eighteen, nineteen, twenty, or more carbon atoms) and at least one carbon-carbon triple bond, which is optionally substituted. The notation "C_2-14 alkynyl" means an optionally substituted linear or branched hydrocarbon including 2- 14 carbon atoms and at least one carbon-carbon triple bond. An alkynyl group may include one, two, three, four, or more carbon-carbon triple bonds. For example, C18 alkynyl may include one or more carbon-carbon triple bonds. Unless otherwise specified, an alkynyl group described herein refers to both unsubstituted and substituted alkynyl groups. Atty. Docket No.45817-0158WO1 As used herein, the term "carbocycle" or "carbocyclic group" means an rings of eleven,

or twenty membered rings. The notation "C3-6 carbocycle" means a carbocycle including a single ring having 3-6 carbon atoms. Carbocycles may include one or more carbon- carbon double or triple bonds and may be non-aromatic or aromatic (e.g., cycloalkyl

term "cycloalkyl" as used herein means a non-aromatic carbocycle and may or may not include any double or triple bond. Unless otherwise specified, carbocycles described herein refers to both unsubstituted and substituted carbocycle groups, i.e., optionally substituted carbocycles. As used herein, the term "heterocycle" or "heterocyclic group" means an optionally substituted mono- or multi-cyclic system including one or more rings, where at least one ring includes at least one heteroatom. Heteroatoms may be, for example, nitrogen, oxygen, or sulfur atoms. Rings may be three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, or fourteen membered rings. Heterocycles may include one or more double or triple bonds and may be non- aromatic or aromatic (e.g., heterocycloalkyl or heteroaryl groups). Examples of heterocycles include imidazolyl, imidazolidinyl, oxazolyl, oxazolidinyl, thiazolyl, thiazolidinyl, pyrazolidinyl, pyrazolyl, isoxazolidinyl, isoxazolyl, isothiazolidinyl, isothiazolyl, morpholinyl, pyrrolyl, pyrrolidinyl, furyl, tetrahydrofuryl, thiophenyl, pyridinyl, piperidinyl, quinolyl, and isoquinolyl groups. The term "heterocycloalkyl" as used herein means a non-aromatic heterocycle and may or may not include any double or triple bond. Unless otherwise specified, heterocycles described herein refers to both unsubstituted and substituted heterocycle groups, i.e., optionally substituted heterocycles. As used herein, the term "heteroalkyl", "heteroalkenyl", or "heteroalkynyl", refers respectively to an alkyl, alkenyl, alkynyl group, as defined herein, which further Atty. Docket No.45817-0158WO1 comprises one or more (e.g., 1, 2, 3, or 4) heteroatoms (e.g., oxygen, sulfur, nitrogen, boron, silicon, phosphorus) wherein the one or more heteroatoms is inserted between adjacent carbon atoms within the parent carbon chain and/or one or more heteroatoms is inserted between a carbon atom and the parent molecule, i.e., between the point of attachment. Unless otherwise specified, heteroalkyls, heteroalkenyls, or heteroalkynyls described herein refers to both unsubstituted and substituted heteroalkyls, heteroalkenyls, or heteroalkynyls, i.e., optionally substituted heteroalkyls, heteroalkenyls, or heteroalkynyls. As used herein, a "biodegradable group" is a group that may facilitate faster metabolism of a lipid in a mammalian entity. A biodegradable group may be selected from the group consisting of, but is not limited to, -C(O)O-, -OC(O)-, -C(O)N(R')-, -N(R')C(O)-, -C(O)-, -C(S)-, -C(S)S-, -SC(S)-, -CH(OH)-, -P(O)(OR')O-, -S(O)2-, an aryl group, and a heteroaryl group. As used herein, an "aryl group" is an optionally Examples of aryl group" is aromatic rings. imidazolyl,

optionally substituted. For example, M and M' can be selected from the non-limiting group consisting of optionally substituted phenyl, oxazole, and thiazole. In the Formulas herein, M and M' can be independently selected from the list of biodegradable groups above. Unless otherwise specified, aryl or heteroaryl groups described herein refers aryl or groups may be

may be selected from the group consisting of, but are not limited to, a halogen atom (e.g., a chloride, bromide, fluoride, or iodide group), a carboxylic acid (e.g., C(O)OH), an alcohol (e.g., a hydroxyl, OH), an ester (e.g., C(O)OR OC(O)R), an aldehyde (e.g., C(O)H), a carbonyl (e.g., C(O)R, alternatively represented by C=O), an acyl halide Atty. Docket No.45817-0158WO1 fluoride, chloride, and , an acetal (e.g., be the same or different (O)4^3-), a thiol (e.g., SH), a

sulfonic acid (e.g., S(O)2OH), a thial (e.g., C(S)H), a sulfate (e.g., S(O)4^2-), a sulfonyl (e.g., S(O)2 ), an amide (e.g., C(O)NR₂, or N(R)C(O)R), an azido (e.g., N₃), a nitro (e.g., NO₂), a cyano (e.g., CN), an isocyano (e.g., NC), an acyloxy (e.g., OC(O)R), an amino (e.g., NR₂, NRH, or NH₂), a carbamoyl (e.g., OC(O)NR₂, OC(O)NRH, or OC(O)NH₂), a sulfonamide (e.g., S(O)2NR2, S(O)2NRH, S(O)2NH2, N(R)S(O)2R, and a is an alkyl

five, or six substituents as defined herein. For example, a C_1-6 alkyl group may be further substituted with one, two, three, four, five, or six substituents as described herein. Compounds of the disclosure that contain nitrogens can be converted to N- oxides by treatment with an oxidizing agent (e.g., 3-chloroperoxybenzoic acid (mCPBA) and/or hydrogen peroxides) to afford other compounds of the disclosure.

hydroxy or N-alkoxy compounds. For example, N-hydroxy compounds can be prepared by oxidation of the parent amine by an oxidizing agent such as m CPBA. All shown and claimed nitrogen-containing compounds are also considered, when allowed by valency and structure, to cover both the compound as shown and its N- hydroxy (i.e., N-OH) and N-alkoxy (i.e., N-OR, wherein R is substituted or unsubstituted C₁-C₆ alkyl, C₁-C₆ alkenyl, C₁-C₆ alkynyl, 3-14-membered carbocycle or 3-14-membered heterocycle) derivatives. Atty. Docket No.45817-0158WO1 Other Lipid Composition Components The lipid composition of a pharmaceutical composition disclosed herein can include one or more components in addition to those described above. For example, the lipid composition can include one or more permeability enhancer molecules, carbohydrates, polymers, surface altering agents (e.g., surfactants), or other components. For example, a permeability enhancer molecule can be a molecule described by U.S. Patent Application Publication No.2005/0222064. Carbohydrates can include simple sugars (e.g., glucose) and polysaccharides (e.g., glycogen and derivatives and analogs thereof). A polymer can be included in and/or used to encapsulate or partially encapsulate a pharmaceutical composition disclosed herein (e.g., a pharmaceutical composition in lipid nanoparticle form). A polymer can be biodegradable and/or biocompatible. A polymer can be selected from, but is not limited to, polyamines, polyethers, polyamides, polyesters, polycarbamates, polyureas, polycarbonates, polystyrenes, polyimides, polysulfones, polyurethanes, polyacetylenes, polyethylenes, polyethyleneimines, polyisocyanates, polyacrylates, polymethacrylates, polyacrylonitriles, and polyarylates. The ratio between the lipid composition and the polynucleotide range can be from about 10:1 to about 60:1 (wt/wt). In some embodiments, the ratio between the lipid composition and the polynucleotide can be about 10:1, 11:1, 12:1, 13:1, 14:1, 15:1, 16:1, 17:1, 18:1, 19:1, 20:1, 21:1, 22:1, 23:1, 24:1, 25:1, 26:1, 27:1, 28:1, 29:1, 30:1, 31:1, 32:1, 33:1, 34:1, 35:1, 36:1, 37:1, 38:1, 39:1, 40:1, 41:1, 42:1, 43:1, 44:1, 45:1, 46:1, 47:1, 48:1, 49:1, 50:1, 51:1, 52:1, 53:1, 54:1, 55:1, 56:1, 57:1, 58:1, 59:1 or 60:1 (wt/wt). In some embodiments, the wt/wt ratio of the lipid composition to the polynucleotide encoding a therapeutic agent is about 20:1 or about 15:1. Atty. Docket No.45817-0158WO1 In some embodiments, the pharmaceutical composition disclosed herein can contain more than one polypeptides. For example, a pharmaceutical composition disclosed herein can contain two or more polynucleotides (e.g., RNA, e.g., mRNA). In some embodiments, the lipid nanoparticles described herein can comprise polynucleotides (e.g., mRNA) in a lipid:polynucleotide weight ratio of 5:1, 10:1, 15:1, 20:1, 25:1, 30:1, 35:1, 40:1, 45:1, 50:1, 55:1, 60:1 or 70:1, or a range or any of these ratios such as, but not limited to, 5:1 to about 10:1, from about 5:1 to about 15:1, from about 5:1 to about 20:1, from about 5:1 to about 25:1, from about 5:1 to about 30:1, from about 5:1 to about 35:1, from about 5:1 to about 40:1, from about 5:1 to about 45:1, from about 5:1 to about 50:1, from about 5:1 to about 55:1, from about 5:1 to about 60:1, from about 5:1 to about 70:1, from about 10:1 to about 15:1, from about 10:1 to about 20:1, from about 10:1 to about 25:1, from about 10:1 to about 30:1, from about 10:1 to about 35:1, from about 10:1 to about 40:1, from about 10:1 to about 45:1, from about 10:1 to about 50:1, from about 10:1 to about 55:1, from about 10:1 to about 60:1, from about 10:1 to about 70:1, from about 15:1 to about 20:1, from about 15:1 to about 25:1,from about 15:1 to about 30:1, from about 15:1 to about 35:1, from about 15:1 to about 40:1, from about 15:1 to about 45:1, from about 15:1 to about 50:1, from about 15:1 to about 55:1, from about 15:1 to about 60:1 or from about 15:1 to about 70:1. In some embodiments, the lipid nanoparticles described herein can comprise the polynucleotide in a concentration from approximately 0.1 mg/ml to 2 mg/ml such as, but not limited to, 0.1 mg/ml, 0.2 mg/ml, 0.3 mg/ml, 0.4 mg/ml, 0.5 mg/ml, 0.6 mg/ml, 0.7 mg/ml, 0.8 mg/ml, 0.9 mg/ml, 1.0 mg/ml, 1.1 mg/ml, 1.2 mg/ml, 1.3 mg/ml, 1.4 mg/ml, 1.5 mg/ml, 1.6 mg/ml, 1.7 mg/ml, 1.8 mg/ml, 1.9 mg/ml, 2.0 mg/ml or greater than 2.0 mg/ml. Nanoparticle Compositions In some embodiments, the pharmaceutical compositions disclosed herein are Formulated as lipid nanoparticles (LNP). Accordingly, the present disclosure also provides nanoparticle compositions comprising (i) a lipid composition comprising a Atty. Docket No.45817-0158WO1 delivery agent such as compound as described herein, and (ii) a polynucleotide encoding a polypeptide. In such nanoparticle composition, the lipid composition disclosed herein can encapsulate the polynucleotide encoding a polypeptide. Nanoparticle compositions are typically sized on the order of micrometers or smaller and can include a lipid bilayer. Nanoparticle compositions encompass lipid nanoparticles (LNPs), liposomes (e.g., lipid vesicles), and lipoplexes. For example, a nanoparticle composition can be a liposome having a lipid bilayer with a diameter of 500 nm or less. Nanoparticle compositions include, for example, lipid nanoparticles (LNPs), liposomes, and lipoplexes. In some embodiments, nanoparticle compositions are vesicles including one or more lipid bilayers. In certain embodiments, a nanoparticle composition includes two or more concentric bilayers separated by aqueous compartments. Lipid bilayers can be functionalized and/or crosslinked to one another. Lipid bilayers can include one or more ligands, proteins, or channels. In some embodiments, a lipid nanoparticle comprises an ionizable amino lipid, a structural lipid, a phospholipid, and mRNA. In some embodiments, the LNP comprises an ionizable amino lipid, a PEG-modified lipid, a sterol and a structural lipid. In some embodiments, the LNP has a molar ratio of about 40-50% ionizable amino lipid; about 5-15% structural lipid; about 30-45% sterol; and about 1-5% PEG- modified lipid. In some embodiments, the lipid nanoparticle comprises 47-49 mol.% ionizable cationic lipid (e.g. ionizable amino lipid, e.g., Compound I-1, Compound I-2, or Compound I-3), 10-12 mol.% non-cationic lipid (e.g., phospholipid, e.g., DSPC), 38- 40 mol.% sterol (e.g., cholesterol) or other structural lipid, and 1-3 mol.% PEG- modified lipid (e.g., PEG-DMG or Compound P-I). For instance, in some embodiments, the lipid nanoparticle (“LNP-1”) may comprise the following components at the following molar ratios: (i) 45-50 mol.% Compound I-1 Atty. Docket No.45817-0158WO1 (ii) 35-45 mol.% sterol (e.g., cholesterol); (iii) 8-12 mol.% phospholipid (e.g., DSPC or DOPE); and (iv) 1.5-3.5 mol.% PEG-lipid. For instance, in some embodiments, the lipid nanoparticle (“LNP-1A”) may comprise the following components at the following molar ratios: (i) 45-50 mol.% Compound I-1 (ii) 35-45 mol.% Cholesterol; (iii) 8-12 mol.% DSPC; and (iv) 1.5-3.5 mol.% PEG-lipid. For instance, in some embodiments, the lipid nanoparticle (“LNP-1B”) may comprise the following components at the following molar ratios: (i) 45-50 mol.% Compound I-1 (ii) 35-45 mol.% Cholesterol; (iii) 8-12 mol.% DSPC; and (iv) 1.5-3.5 mol.% PEG-lipid. In some embodiments, the lipid nanoparticle (“LNP-2”) may comprise the following: (i) 45-50 mol.% Compound I-2; (ii) 35-45 mol.% sterol (e.g., Cholesterol); (iii) 8-12 mol.% phospholipid (e.g., DSPC or DOPE); and (iv) 1.5-3.5 mol.% PEG-lipid. In some embodiments, the lipid nanoparticle (“LNP-2A”) may comprise the following: (i) 45-50 mol.% Compound I-2; (ii) 35-45 mol.% Cholesterol; (iii) 8-12 mol.% DSPC; and (iv) 1.5-3.5 mol.% PEG-lipid. Atty. Docket No.45817-0158WO1 For instance, in some embodiments, the lipid nanoparticle (“LNP-2B”) may comprise the following components at the following molar ratios: (i) 45-50 mol.% Compound I-2; (ii) 35-45 mol.% Cholesterol; (iii) 8-12 mol.% DSPC; and (iv) 1.5-3.5 mol.% PEG-lipid. In some embodiments, the lipid nanoparticle (“LNP-3”) may comprise the following: (i) 45-50 mol.% Compound I-3; (ii) 35-45 mol.% sterol (e.g., Cholesterol); (iii) 8-12 mol.% phospholipid (e.g., DSPC or DOPE); and (iv) 1.5-3.5 mol.% PEG-lipid. In some embodiments, the lipid nanoparticle (“LNP-3A”) may comprise the following: (i) 45-50 mol.% Compound I-3; (ii) 35-45 mol.% Cholesterol; (iii) 8-12 mol.% DSPC; and (iv) 1.5-3.5 mol.% PEG-lipid. In some embodiments, the lipid nanoparticle (“LNP-3B”) may comprise the following: (i) 45-50 mol.% Compound I-3; (ii) 35-45 mol.% Cholesterol; (iii) 8-12 mol.% DSPC; and (iv) 1.5-3.5 mol.% PEG-lipid. In some embodiments, the LNP has a polydispersity value of less than 0.4. In some embodiments, the LNP has a net neutral charge at a neutral pH. In some embodiments, the LNP has a mean diameter of 50-150 nm. In some embodiments, the LNP has a mean diameter of 80-100 nm. Atty. Docket No.45817-0158WO1 As generally defined herein, the term “lipid” refers to a small molecule that has hydrophobic or amphiphilic properties. Lipids may be naturally occurring or synthetic. Examples of classes of lipids include, but are not limited to, fats, waxes, sterol-containing metabolites, vitamins, fatty acids, glycerolipids, glycerophospholipids, sphingolipids, saccharolipids, and polyketides, and prenol lipids. In some instances, the amphiphilic properties of some lipids lead them to form liposomes, vesicles, or membranes in aqueous media. In some embodiments, a lipid nanoparticle (LNP) may comprise an ionizable amino lipid. As used herein, the term “ionizable amino lipid” has its ordinary meaning in the art and may refer to a lipid comprising one or more charged moieties. In some embodiments, an ionizable amino lipid may be positively charged or negatively charged. An ionizable amino lipid may be positively charged, in which case it can be referred to as “cationic lipid”. In certain embodiments, an ionizable amino lipid molecule may comprise an amine group, and can be referred to as an ionizable amino lipid. As used herein, a “charged moiety” is a chemical moiety that carries a formal electronic charge, e.g., monovalent (+1, or -1), divalent (+2, or -2), trivalent (+3, or - 3), etc. The charged moiety may be anionic (i.e., negatively charged) or cationic (i.e., positively charged). Examples of positively-charged moieties include amine groups (e.g., primary, secondary, and/or tertiary amines), ammonium groups, pyridinium group, guanidine groups, and imidizolium groups. In a particular embodiment, the charged moieties comprise amine groups. Examples of negatively- charged groups or precursors thereof, include carboxylate groups, sulfonate groups, sulfate groups, phosphonate groups, phosphate groups, hydroxyl groups, and the like. The charge of the charged moiety may vary, in some cases, with the environmental conditions, for example, changes in pH may alter the charge of the moiety, and/or cause the moiety to become charged or uncharged. In general, the charge density of the molecule may be selected as desired. It should be understood that the terms “charged” or “charged moiety” does not refer to a “partial negative charge" or “partial positive charge" on a molecule. The terms “partial negative charge" and “partial positive charge" are given their ordinary Atty. Docket No.45817-0158WO1 meaning in the art. A “partial negative charge" may result when a functional group comprises a bond that becomes polarized such that electron density is pulled toward one atom of the bond, creating a partial negative charge on the atom. Those of ordinary skill in the art will, in general, recognize bonds that can become polarized in this way. The ionizable amino lipid is sometimes referred to in the art as an “ionizable cationic lipid”. In some embodiments, the ionizable amino lipid may have a positively charged hydrophilic head and a hydrophobic tail that are connected via a linker structure. In addition to these, an ionizable amino lipid may also be a lipid including a cyclic amine group. In some embodiments, the ionizable amino lipid may be selected from, but not limited to, an ionizable amino lipid described in International Publication Nos. WO2013086354 and WO2013116126; the contents of each of which are herein incorporated by reference in their entirety. In yet another embodiment, the ionizable amino lipid may be selected from, but not limited to, Formula CLI-CLXXXXII of US Patent No.7,404,969; each of which is herein incorporated by reference in their entirety. In some embodiments, the lipid may be a cleavable lipid such as those described in International Publication No. WO2012170889, herein incorporated by reference in its entirety. In some embodiments, the lipid may be synthesized by methods known in the art and/or as described in International Publication Nos. WO2013086354; the contents of each of which are herein incorporated by reference in their entirety. Nanoparticle compositions can be characterized by a variety of methods. For example, microscopy (e.g., transmission electron microscopy or scanning electron microscopy) can be used to examine the morphology and size distribution of a nanoparticle composition. Dynamic light scattering or potentiometry (e.g., Atty. Docket No.45817-0158WO1 potentiometric titrations) can be used to measure zeta potentials. Dynamic light scattering can also be utilized to determine particle sizes. Instruments such as the Zetasizer Nano ZS (Malvern Instruments Ltd, Malvern, Worcestershire, UK) can also be used to measure multiple characteristics of a nanoparticle composition, such as particle size, polydispersity index, and zeta potential. The size of the nanoparticles can help counter biological reactions such as, but not limited to, inflammation, or can increase the biological effect of the polynucleotide. As used herein, “size” or “mean size” in the context of nanoparticle compositions refers to the mean diameter of a nanoparticle composition. In some embodiments, the polynucleotide encoding a polypeptide are formulated in lipid nanoparticles having a diameter from about 10 to about 100 nm such as, but not limited to, about 10 to about 20 nm, about 10 to about 30 nm, about 10 to about 40 nm, about 10 to about 50 nm, about 10 to about 60 nm, about 10 to about 70 nm, about 10 to about 80 nm, about 10 to about 90 nm, about 20 to about 30 nm, about 20 to about 40 nm, about 20 to about 50 nm, about 20 to about 60 nm, about 20 to about 70 nm, about 20 to about 80 nm, about 20 to about 90 nm, about 20 to about 100 nm, about 30 to about 40 nm, about 30 to about 50 nm, about 30 to about 60 nm, about 30 to about 70 nm, about 30 to about 80 nm, about 30 to about 90 nm, about 30 to about 100 nm, about 40 to about 50 nm, about 40 to about 60 nm, about 40 to about 70 nm, about 40 to about 80 nm, about 40 to about 90 nm, about 40 to about 100 nm, about 50 to about 60 nm, about 50 to about 70 nm, about 50 to about 80 nm, about 50 to about 90 nm, about 50 to about 100 nm, about 60 to about 70 nm, about 60 to about 80 nm, about 60 to about 90 nm, about 60 to about 100 nm, about 70 to about 80 nm, about 70 to about 90 nm, about 70 to about 100 nm, about 80 to about 90 nm, about 80 to about 100 nm and/or about 90 to about 100 nm. In some embodiments, the nanoparticles have a diameter from about 10 to 500 nm. In some embodiments, the nanoparticle has a diameter greater than 100 nm, greater than 150 nm, greater than 200 nm, greater than 250 nm, greater than 300 nm, Atty. Docket No.45817-0158WO1 greater than 350 nm, greater than 400 nm, greater than 450 nm, greater than 500 nm, greater than 550 nm, greater than 600 nm, greater than 650 nm, greater than 700 nm, greater than 750 nm, greater than 800 nm, greater than 850 nm, greater than 900 nm, greater than 950 nm or greater than 1000 nm. In some embodiments, the largest dimension of a nanoparticle composition is 1 µm or shorter (e.g., 1 µm, 900 nm, 800 nm, 700 nm, 600 nm, 500 nm, 400 nm, 300 nm, 200 nm, 175 nm, 150 nm, 125 nm, 100 nm, 75 nm, 50 nm, or shorter). A nanoparticle composition can be relatively homogenous. A polydispersity index can be used to indicate the homogeneity of a nanoparticle composition, e.g., the particle size distribution of the nanoparticle composition. A small (e.g., less than 0.3) polydispersity index generally indicates a narrow particle size distribution. A nanoparticle composition can have a polydispersity index from about 0 to about 0.25, such as 0.01, 0.02, 0.03, 0.04, 0.05, 0.06, 0.07, 0.08, 0.09, 0.10, 0.11, 0.12, 0.13, 0.14, 0.15, 0.16, 0.17, 0.18, 0.19, 0.20, 0.21, 0.22, 0.23, 0.24, or 0.25. In some embodiments, the polydispersity index of a nanoparticle composition disclosed herein can be from about 0.10 to about 0.20. The amount of a polynucleotide present in a pharmaceutical composition disclosed herein can depend on multiple factors such as the size of the polynucleotide, desired target and/or application, or other properties of the nanoparticle composition as well as on the properties of the polynucleotide. For example, the amount of an mRNA useful in a nanoparticle composition can depend on the size (expressed as length, or molecular mass), sequence, and other characteristics of the mRNA. The relative amounts of a polynucleotide in a nanoparticle composition can also vary. The relative amounts of the lipid composition and the polynucleotide present in a lipid nanoparticle composition of the present disclosure can be optimized according to considerations of efficacy and tolerability. For compositions including an mRNA as a polynucleotide, the N:P ratio can serve as a useful metric. Atty. Docket No.45817-0158WO1 As the N:P ratio of a nanoparticle composition controls both expression and tolerability, nanoparticle compositions with low N:P ratios and strong expression are desirable. N:P ratios vary according to the ratio of lipids to RNA in a nanoparticle composition. In general, a lower N:P ratio is preferred. The one or more RNA, lipids, and amounts thereof can be selected to provide an N:P ratio from about 2:1 to about 30:1, such as 2:1, 3:1, 4:1, 5:1, 6:1, 7:1, 8:1, 9:1, 10:1, 12:1, 14:1, 16:1, 18:1, 20:1, 22:1, 24:1, 26:1, 28:1, or 30:1. In certain embodiments, the N:P ratio can be from about 2:1 to about 8:1. In other embodiments, the N:P ratio is from about 5:1 to about 8:1. In certain embodiments, the N:P ratio is between 5:1 and 6:1. In one specific aspect, the N:P ratio is about is about 5.67:1. In some embodiments, the polynucleotides described herein can be Formulated for controlled release and/or targeted delivery. As used herein, "controlled release" refers to a pharmaceutical composition or compound release profile that conforms to a particular pattern of release to effect a therapeutic outcome. In some embodiments, the polynucleotides can be encapsulated into a delivery agent described herein and/or known in the art for controlled release and/or targeted delivery. As used herein, the term "encapsulate" means to enclose, surround or encase. As it relates to the formulation of the compounds of the present disclosure, encapsulation can be substantial, complete or partial. The term "substantially encapsulated" means that at least greater than 50, 60, 70, 80, 85, 90, 95, 96, 97, 98, 99, or greater than 99% of the pharmaceutical composition or compound of the present disclosure can be enclosed, surrounded or encased within the delivery agent. "Partial encapsulation" or “partially encapsulate” means that less than 10, 10, 20, 30, 4050 or less of the pharmaceutical composition or compound of the present disclosure can be enclosed, surrounded or encased within the delivery agent. In some embodiments, the therapeutic nanoparticle polynucleotide can be Formulated for sustained release. As used herein, "sustained release" refers to a pharmaceutical composition or compound that conforms to a release rate over a Atty. Docket No.45817-0158WO1 specific period of time. The period of time can include, but is not limited to, hours, days, weeks, months and years. As a non-limiting example, the sustained release nanoparticle of the polynucleotides described herein can be Formulated as disclosed in Intl. Pub. No. WO2010075072 and U.S. Pub. Nos. US20100216804, US20110217377, US20120201859 and US20130150295, each of which is herein incorporated by reference in their entirety. In some embodiments, the therapeutic nanoparticle polynucleotide can be Formulated to be target specific, such as those described in Intl. Pub. Nos. WO2008121949, WO2010005726, WO2010005725, WO2011084521 and WO2011084518; and U.S. Pub. Nos. US20100069426, US20120004293 and US20100104655, each of which is herein incorporated by reference in its entirety. The LNPs can be prepared using microfluidic mixers or micromixers. Exemplary microfluidic mixers can include, but are not limited to, a slit interdigital micromixer including, but not limited to those manufactured by Microinnova (Allerheiligen bei Wildon, Austria) and/or a staggered herringbone micromixer (SHM) (see, Zhigaltsev et al., Langmuir.28:3633-40 (2012); Belliveau et al., Molecular Therapy-Nucleic Acids.1:e37 (2012); Chen et al., J. Am. Chem. Soc. 134(16):6948-51 (2012); each of which is herein incorporated by reference in its entirety). Exemplary micromixers include Slit Interdigital Microstructured Mixer (SIMM-V2) or a Standard Slit Interdigital Micro Mixer (SSIMM) or Caterpillar (CPMM) or Impinging-jet (IJMM,) from the Institut für Mikrotechnik Mainz GmbH, Mainz Germany. In some embodiments, methods of making LNP using SHM further comprise mixing at least two input streams wherein mixing occurs by microstructure- induced chaotic advection (MICA). According to this method, fluid streams flow through channels present in a herringbone pattern causing rotational flow and folding the fluids around each other. This method can also comprise a surface for fluid mixing wherein the surface changes orientations during fluid cycling. Methods of generating LNPs using SHM include those disclosed in U.S. Pub. Nos. US20040262223 and US20120276209, each of which is incorporated herein by reference in their entirety. Atty. Docket No.45817-0158WO1 In some embodiments, the polynucleotides described herein can be Formulated in lipid nanoparticles using microfluidic technology (see, Whitesides, George M., Nature 442: 368-373 (2006); and Abraham et al., Science 295: 647-651 (2002); each of which is herein incorporated by reference in its entirety). In some embodiments, the polynucleotides can be Formulated in lipid nanoparticles using a micromixer chip such as, but not limited to, those from Harvard Apparatus (Holliston, MA) or Dolomite Microfluidics (Royston, UK). A micromixer chip can be used for rapid mixing of two or more fluid streams with a split and recombine mechanism. In some embodiment, the nanoparticles described herein are stealth nanoparticles or target-specific stealth nanoparticles such as, but not limited to, those described in U.S. Pub. No. US20130172406, herein incorporated by reference in its entirety. The stealth or target-specific stealth nanoparticles can comprise a polymeric matrix, which can comprise two or more polymers such as, but not limited to, polyethylenes, polycarbonates, polyanhydrides, polyhydroxyacids, polypropylfumerates, polycaprolactones, polyamides, polyacetals, polyethers, polyesters, poly(orthoesters), polycyanoacrylates, polyvinyl alcohols, polyurethanes, polyphosphazenes, polyacrylates, polymethacrylates, polycyanoacrylates, polyureas, polystyrenes, polyamines, polyesters, polyanhydrides, polyethers, polyurethanes, polymethacrylates, polyacrylates, polycyanoacrylates, or combinations thereof. Methods Using Bispecific Engagers In some aspects, bispecific antibodies or engagers of the present disclosure are administered to a subject in need thereof. Alternatively, mRNA (or other nucleic acids) encoding bispecific antibodies or engagers of the present disclosure are administered to a subject in need thereof. For example, an mRNA encoding a bispecific engager as disclosed herein may be administered to a subject. For another example, two mRNA, each encoding a different bispecific engager disclosed herein, may be administered to a subject. In general, the subject in need thereof is a subject with a disease, disorder, and/or condition that may be treated with technologies described herein. Atty. Docket No.45817-0158WO1 In some aspects, the subject in need thereof is a subject with cancer, such as a cancer associated with B cells. In some embodiments, the cancer may be multiple myeloma. In some embodiments, the multiple myeloma may be relapsed or refractory multiple myeloma (RRMM). In some embodiments, the cancer may be a B cell lymphoma, including a non-Hodgkin lymphoma such as Burkitt lymphoma, chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL), diffuse large B-cell lymphoma, follicular lymphoma, or mantle cell lymphoma. In embodiments where the subject has relapsed or refractory multiple myeloma, the subject may have previously been treated with one or more or all of a proteasome inhibitor, an immunomodulatory drug, and an anti-cluster of differentiation monoclonal antibody (e.g., CD38 mAb). The subject may have received at least one, at least two, or at least three prior lines of therapy. The subject may be triple-class refractory. Additionally or alternatively, the subject may be intolerant to one or more or all of a proteasome inhibitor, an immunomodulatory drug, and an anti-cluster of differentiation monoclonal antibody (e.g., CD38 mAb). In some aspects, the subject in need thereof is a subject with an immune condition associated with B cells, such as an autoimmune disease. The role of B cells in autoimmune diseases involves different cellular functions, including the well- established secretion of autoantibodies, autoantigen presentation and ensuing reciprocal interactions with T cells, secretion of inflammatory cytokines, and the generation of ectopic germinal centers. Indeed, B cells may be involved in the pathogenesis and/or progression of systemic lupus erythematosus, rheumatoid arthritis, and type 1 diabetes, among other autoimmune diseases, and therefore such diseases may be treated with the disclosed B cell engagers. In some aspects, the subject in need thereof is a mammal. In some embodiments, a mammal includes, for example and without limitation, a household pet (e.g., a dog, a cat, a rabbit, a ferret, a hamster), a livestock or farm animal (e.g., a cow, a pig, a sheep, a goat, a pig, a chicken or another poultry), a horse, a monkey, a laboratory animal (e.g., a mouse, a rat, a rabbit) and a human. In a preferred Atty. Docket No.45817-0158WO1 embodiment, the subject in need thereof is a human. Technologies of the present disclosure can be practiced in any subject in need thereof that is likely to benefit from administration of technologies of the present disclosure (e.g., a subject with cancer). In some embodiments, a subject in need thereof is a human. In some embodiments, the human is male. In some embodiments, the human is female. In some embodiments, the human is an adult (e.g., 18 or more years of age). In some embodiments, the adult is greater than 18 years old, greater than 25 years old, greater than 30 years old, greater than 40 years old, greater than 50 years old, greater than 55 years old, greater than 60 years old, greater than 65 years old, greater than 70 years old, greater than 75 years old, greater than 80 years old, greater than 85 years old, greater than 90 years old, greater than 95 years old, greater than 100 years old, or greater than 105 years old in age. In some embodiments, the human is a child. In some embodiments, the child is greater than 2 years old, greater than 3 years old, greater than 4 years old, greater than 5 years old, greater than 6 years old, greater than 7 years old, greater than 8 years old, greater than 9 years old, greater than 10 years old, greater than 11 years old, greater than 12 years old, greater than 13 years old, greater than 14 years old, greater than 15 years old, or greater than 16 years old in age. Routes of Administration and Dosing The disclosed bispecific engagers and nucleic acids (e.g., mRNA) encoding the same can be administered to a subject (e.g., a mammalian subject, such as a human) by a variety of routes. In some embodiments, the antibody or nucleic acid is administered to the subject intravenously, subcutaneously, intramuscularly, subdermally, parenterally, intrathecally, intracerebroventricularly, or transdermally. In some embodiments, mRNA encoding a bispecific engager as disclosed herein is administered intravenously. The most suitable route for administration in any given case will depend on the particular therapeutic agent administered, the patient, pharmaceutical formulation methods, and various patient-specific parameters, such as the patient's age, body Atty. Docket No.45817-0158WO1 weight, sex, severity of the diseases being treated, the patient’s diet, and the patient’s excretion rate. An appropriate dosage of the disclosed bispecific engagers and nucleic acids (e.g., mRNA) encoding the same will vary with the particular condition, disease and/or disease being treated, various subject-specific parameters (e.g., age, weight, physical condition of the subject), severity of the particular condition, disease, and/or disorder being treated, the nature of current or combination therapy (if any), the specific route of administration and other factors within the knowledge and expertise of a health practitioner. In some embodiments, a maximally tolerated dose of technologies described herein is to be used, e.g., the highest safe dose according to sound medical judgement. In some embodiments, technologies described herein are administered in an effective amount, e.g., a dose sufficient to provide one or more medically desirable results. A therapeutic regimen for use in accordance with treatments described herein may include administration of the disclosed bispecific engagers and nucleic acids (e.g., mRNA) encoding the same once a day, once every two days, once every three days, twice a week, once a week, once every two weeks, once every three weeks, once every month or four weeks, once every six weeks, once every two months or eight weeks, once every three months or twelve weeks. In some certain embodiments, a subject receives a single dose of bispecific engagers or nucleic acids (e.g., mRNA) encoding the same described herein. In certain embodiments, a subject receives a plurality of doses of disclosed bispecific engagers or nucleic acids (e.g., mRNA) encoding the same (e.g., at least two, at least three, at least four, at least five, at least six, at least eight, at least ten, or more doses). In some embodiments, technologies described herein are administered over a period of time, such as one week, two weeks, three weeks, four weeks, six weeks, two months, three months, four months, five months, six months, one year or more. Appropriate therapeutic regimens are readily understood by medical practitioners and such regimens may be designed by a medical practitioner for a particular patient (e.g., a patient-specific regimen). In some embodiments, a regimen may comprise a dose escalation study to evaluate safety and Atty. Docket No.45817-0158WO1 tolerability of escalating doses and/or to determine a maximum tolerated dose and/or to determine a recommended dose. Kits Also included herein are kits that contain disclosed bispecific engagers and nucleic acids (e.g., mRNA) encoding the same. In some embodiments, the kits provided herein contain one or more cells engineered to express and secrete one or more of the disclosed bispecific engagers and nucleic acids (e.g., mRNA) encoding the same. A kit described herein may include reagents that can be used to produce a pharmaceutical composition of the disclosure. Optionally, kits described herein may include reagents that can induce the expression of the disclosed bispecific engagers and nucleic acids (e.g., mRNA) encoding the same within cells (e.g., mammalian cells). Other kits described herein may include tools for engineering a prokaryotic or eukaryotic cell (e.g., a CHO cell or a BL21(DE3) E. Coli cell or an T cell) so as to express and secrete one or more of the disclosed bispecific engagers and nucleic acids (e.g., mRNA) encoding the same. For example, a kit may contain CHO cells stored in an appropriate media and optionally frozen according to methods known in the art. The kit may also contain one or more nucleic acids (e.g., mRNA) encoding one or more of the disclosed bispecific engagers. A kit described herein may also provide a package insert describing how the disclosed bispecific engagers and nucleic acids (e.g., mRNA) encoding the same may be administered to a subject for the treatment of a disease, disorder and/or condition (e.g., cancer). Definitions As used herein, the term “about” refers to a stated numerical term and a value that is no more than 10% above or below the value being described. For example, the Atty. Docket No.45817-0158WO1 term “about 5 nM” indicates disclosure of both the stated value of 5 nM and a range of from 4.5 nM to 5.5 nM. As used herein, the term “bispecific antibodies” refers to monoclonal, often human or humanized antibodies that have binding specificities for at least two different antigens. Bispecific antibodies of the disclosure may have binding specificities that are directed towards a tumor associated antigen and any other antigen, e.g., for a cell-surface protein, receptor, receptor subunit, or tissue-specific antigen. A bispecific antibody may also be an antibody or antigen-binding fragment thereof that includes two separate antigen-binding domains (e.g., VHH, optionally joined directly or indirectly by a linker). The binding domains may bind the same antigen or different antigens. A “bispecific engager” is a particular type of bispecific antibody that binds to an antigen associated with a disease state (e.g., a a target molecule on a B cell) and a molecule on an immune cell (e.g., a natural killer (NK) cell). The presently disclosed bispecific engagers generally comprise VHH as their binding domains. As used herein, the term “complementarity determining region” or “CDR” refers to a hypervariable region found in the light chain and/or the heavy chain variable domains of an antibody. The more highly conserved portions of variable domains are called the framework regions (FRs). As is appreciated in the art, the amino acid positions that delineate a hypervariable region of an antibody can vary, depending on the context and the various definitions known in the art. Some positions within a variable domain may be viewed as hybrid hypervariable positions in that these positions can be deemed to be within a hypervariable region under one set of criteria while being deemed to be outside a hypervariable region under a different set of criteria. One or more of these positions can also be found in extended hypervariable regions. The disclosure includes antibodies comprising modifications in these hybrid hypervariable positions. The variable domains of native heavy and light chains each comprise four framework regions that primarily adopt a β-sheet configuration, connected by three CDRs, which form loops that connect, and in some cases form part of, the β-sheet structure. The CDRs in each chain are held together in Atty. Docket No.45817-0158WO1 close proximity by the FR regions in the order FR1-CDR1-FR2-CDR2-FR3-CDR3- FR4 and, with the CDRs from the other antibody chains, contribute to the formation of the target binding site of antibodies (see, Kabat et al., Sequences of Proteins of Immunological Interest (National Institute of Health, Bethesda, Md. (1987); incorporated herein by reference). As used herein, numbering of immunoglobulin amino acid residues is performed according to the immunoglobulin amino acid residue numbering system of Kabat et al., unless otherwise indicated. As used herein, the terms “conservative mutation,” “conservative substitution,” “conservative amino acid substitution,” and the like refer to a substitution of one or more amino acids for one or more different amino acids that exhibit similar physicochemical properties, such as polarity, electrostatic charge, and/or steric volume. These properties are summarized for each of the twenty naturally-occurring amino acids in Table 13 below. Table 13 – Representative physicochemical properties of naturally-occurring amino acids Electrostatic 3 1 Side- Amino Acid Letter Letter chain character at Steric p ^† Code Code Polarity hysiological pH Volume (7.4) Alanine Ala A nonpolar neutral small Arginine Arg R polar cationic large Asparagine Asn N polar neutral intermediate Aspartic acid Asp D polar anionic intermediate Cysteine Cys C nonpolar neutral intermediate Glutamic acid Glu E polar anionic intermediate Glutamine Gln Q polar neutral intermediate Glycine Gly G nonpolar neutral small Both neutral and Histidine His H polar cationic forms in large equilibrium at pH 7.4 Atty. Docket No.45817-0158WO1 Isoleucine Ile I nonpolar neutral large Leucine Leu L nonpolar neutral large Lysine Lys K polar cationic large Methionine Met M nonpolar neutral large Phenylalanine Phe F nonpolar neutral large P^{roline Pro P non-} p_olar ^{neutral intermediate} Serine Ser S polar neutral small Threonine Thr T polar neutral intermediate Tryptophan Trp W nonpolar neutral bulky Tyrosine Tyr Y polar neutral large Valine Val V nonpolar neutral intermediate †based on volume in A³: 50-100 is small, 100-150 is intermediate, 150-200 is large, and >200 is bulky From this table it is appreciated that the conservative amino acid families include, e.g., (i) G, A, V, L, I, P, and M; (ii) D and E; (iii) C, S and T; (iv) H, K and R; (v) N and Q; and (vi) F, Y and W. A conservative mutation or substitution is therefore one that substitutes one amino acid for a member of the same amino acid family (e.g., a substitution of Ser for Thr or Lys for Arg). As used herein, the term “construct” refers to a fusion protein containing a first polypeptide domain bound to a second polypeptide domain. The polypeptide domains may each independently be binding domains (e.g., one that binds a B cell associated with a disease and one that binds to a NK cell), for instance, as described herein. The first polypeptide domain may be covalently bound to the second polypeptide domain, for instance, by way of a linker, such as a peptide linker or a disulfide bridge, among others. Exemplary linkers that may be used to join the polypeptide domains of a construct include, without limitation, those that are described in Leriche et al., Bioorg. Med. Chem., 20:571-582 (2012), the disclosure of which is incorporated herein by reference in its entirety. Atty. Docket No.45817-0158WO1 As used herein, the term “framework region” or “FW region” includes amino acid residues that are adjacent to the CDRs. FW region residues may be present in, for example, human antibodies, rodent-derived antibodies (e.g., murine antibodies), humanized antibodies, primatized antibodies, chimeric antibodies, antibody fragments (e.g., Fab fragments), single-chain antibody fragments (e.g., scFv fragments), antibody domains, and bispecific antibodies, among others. As used herein, the term “humanized” antibodies refers to forms of non- human (e.g., murine) antibodies that are chimeric immunoglobulins, or immunoglobulin chains or fragments thereof (such as Fv, Fab, Fab', F(ab')2 or other target-binding subdomains of antibodies), which contain minimal sequences derived from non-human immunoglobulin. In general, a humanized antibody will contain substantially all of at least one, and typically two, variable domains, in which all or substantially all of the CDRs correspond to those of a non-human immunoglobulin. All or substantially all of the FRs may also be those of a human immunoglobulin sequence. The humanized antibody may also contain at least a portion of an immunoglobulin constant region (Fc), typically that of a human immunoglobulin consensus sequence. Methods of antibody humanization are known in the art. See, e.g., Riechmann et al., Nature 332:323-7 (1988); U.S. Patent Nos: 5,530,101; 5,585,089; 5,693,761; 5,693,762; and 6,180,370 to Queen et al; EP239400; PCT publication WO 91/09967; U.S. Patent No.5,225,539; EP592106; and EP519596; the disclosure of each of which is incorporated herein by reference. As used herein, the term “lipid nanoparticle” refers to a transfer vehicle including one or more lipids (e.g., cationic lipids, non-cationic lipids, and PEG- modified lipids). Exemplary lipid nanoparticles are formulated to deliver one or more mRNA to one or more target cells. Examples of suitable lipids include, for example, the phosphatidyl compounds (e.g., phosphatidylglycerol, phosphatidylcholine, phosphatidylserine, phosphatidylethanolamine, sphingolipids, cerebrosides, and gangliosides). Lipid nanoparticles may contain a cationic lipid, or a lipid species with a net positive charge at a selected pH (e.g., physiological pH), to encapsulate and/or enhance the delivery of mRNA into the target cells. Atty. Docket No.45817-0158WO1 As used herein, the terms “messenger RNA” or “mRNA” refer to any polynucleotide which encodes a polypeptide of interest and which is capable of being translated to produce the encoded polypeptide of interest in vitro, in vivo, in situ, or ex vivo. Traditionally, the basic components of an mRNA molecule include a coding region, a 5’UTR, a 3’UTR, a 5’ cap, and a poly-A tail. As used herein, the terms “modified messenger RNA” or “modified mRNA” refer to mRNA polynucleotides that include naturally occurring and/or non-naturally occurring modifications, for example, of a sugar, a nucleobase, or an internucleoside linkage (e.g., to a linking phosphate, to a phosphodiester linkage, or to the phosphodiester backbone). Non-natural modified nucleotides may be introduced during synthesis of post-synthesis of the polynucleotides to achieve desired functions or properties. The modifications may be present on an internucleoside linkage, purine or pyrimidine base, or sugar. The modification may be introduced with chemical synthesis or with a polymerase enzyme at the terminal of a chain or anywhere else in the chain. Any of the regions of a polynucleotide may be chemically modified. As used herein, the term “nucleic acid” includes any compound containing a continuous segment of nucleosides joined by way of one or more internucleoside linkages (e.g., polymers of nucleosides linked by way of phosphodiester bonds). Exemplary nucleic acids include ribonucleic acids (RNA, in particular mRNA), deoxyribonucleic acids (DNA), threose nucleic acids (TNA), glycol nucleic acids (GNA), peptide nucleic acids (PNA), locked nucleic acids (LNA), or hybrids thereof. Nucleic acids also include RNAi inducers, RNAi agents, siRNAs, shRNAs, miRNAs, antisense RNAs, ribozymes, catalytic DNAs, tRNAs, RNAs that induce triple spiral formation, aptamers, vectors, and the like. In a preferred embodiment, the nucleic acid is one or more modified messenger RNAs (modified mRNAs). As used herein, the terms “percent (%) sequence identity,” “percent (%) identity,” and the like, with respect to a reference polynucleotide or polypeptide sequence, is defined as the percentage of nucleic acids or amino acids in a candidate sequence that are identical to the nucleic acids or amino acids in the reference Atty. Docket No.45817-0158WO1 polynucleotide or polypeptide sequence, after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent sequence identity. Alignment for purposes of determining percent nucleic acid or amino acid sequence identity can be achieved in various ways that are within the capabilities of one of skill in the art, for example, using publicly available computer software such as BLAST, BLAST-2, or Megalign software. Those skilled in the art can determine appropriate parameters for aligning sequences, including any algorithms needed to achieve maximal alignment over the full length of the sequences being compared. For example, percent sequence identity values may be generated using the sequence comparison computer program BLAST. As an illustration, the percent sequence identity of a given nucleic acid or amino acid sequence, A, to, with, or against a given nucleic acid or amino acid sequence, B, (which can alternatively be phrased as a given nucleic acid or amino acid sequence, A that has a certain percent sequence identity to, with, or against a given nucleic acid or amino acid sequence, B) is calculated as: 100 multiplied by (the fraction X/Y) where X is the number of nucleotides or amino acids scored as identical matches by a sequence alignment program (e.g., BLAST) in that program’s alignment of A and B, and where Y is the total number of nucleic acids in B. It will be appreciated that where the length of nucleic acid or amino acid sequence A is not equal to the length of nucleic acid or amino acid sequence B, the percent sequence identity of A to B will not equal the percent sequence identity of B to A. As used herein, the term “operatively linked” in the context of a polynucleotide fragment is intended to mean that the two polynucleotide fragments are joined such that the amino acid sequences encoded by the two polynucleotide fragments remain in-frame. As used herein, the term “VHH antibody” refers to a single-chain antibody that contains only a single heavy-chain variable domain. Unlike a traditional, full- length antibody, which includes heavy chains and light chains, each containing a corresponding variable domain (i.e., a heavy chain variable domain, VH, and a light chain variable domain, V_L) having three CDRs, a VHH antibody only includes one Atty. Docket No.45817-0158WO1 heavy-chain variable domain having a total of three CDRs (referred to herein as CDR- H1, CDR-H2, and CDR-H3). As used herein the phrase “specifically binds” refers to a binding reaction ,

Atty. Docket No.45817-0158WO1 of days, weeks, months, or years). Alternatively, a patient may be symptomatic for a particular disease, but has yet to be diagnosed with the disease by a physician. Other patients that may be treated using the compositions and methods described herein include those that have been diagnosed as having a disease or disorder, and may or may not be showing symptoms of the disease as of yet. As used herein, the term “transfection” refers to any of a wide variety of techniques commonly used for the introduction of a nucleic acid molecule, e.g., exogenous DNA or RNA into a prokaryotic or eukaryotic host cell, e.g., electroporation, lipofection, calcium- phosphate precipitation, DEAE- dextran transfection and the like. As used herein, the terms “treat” or “treatment” refer to therapeutic treatment, in which the object is to inhibit or slow down (lessen) an undesired physiological change or disorder, such as a cancer or an immunological disorder (e.g., autoimmune disorders (and graft-versus-host disease, among others). Beneficial or desired clinical results of treatment include, without limitation, alleviation of symptoms, diminishment of extent of disease, stabilized (i.e., not worsening) state of disease, delay or slowing of disease progression, amelioration or palliation of the disease state, and remission (whether partial or total), whether detectable or undetectable. Those in need of treatment include those already having the condition or disorder, as well as those prone to have the condition or disorder or those in which the condition or disorder is to be inhibited. As used herein the term “variable region CDR” includes amino acids in a CDR or complementarity determining region as identified using sequence or structure-based methods. As used herein, the term “CDR” or “complementarity determining region” refers to the noncontiguous antigen-binding sites found within the variable regions of both heavy and light chain polypeptides. These particular regions have been described by Kabat et al., J. Biol. Chem.252:6609-6616 (1977) and Kabat, et al., Sequences of Proteins of Immunological Interest, Fifth Edition, U.S. Department of Health and Human Services, NIH Publication No.91 -3242 (1991); by Atty. Docket No.45817-0158WO1 Chothia et al., J. Mol. Biol.196:901-917 (1987), and by MacCallum et al., J. Mol. Biol.262:732-745 (1996) where the definitions include overlapping or subsets of amino acid residues when compared against each other. In certain embodiments, the term “CDR” is a CDR as defined by Kabat based on sequence comparisons. As used herein, the term “vector” includes a nucleic acid vector, e.g., a DNA vector, such as a plasmid, an RNA vector, virus or other suitable replicon (e.g., viral vector). A variety of vectors have been developed for the delivery of polynucleotides encoding exogenous proteins into a prokaryotic or eukaryotic cell. Examples of such expression vectors are disclosed in, e.g., WO 1994/11026; incorporated herein by reference. Expression vectors described herein contain a polynucleotide sequence as well as, e.g., additional sequence elements used for the expression of proteins and/or the integration of these polynucleotide sequences into the genome of a mammalian cell. Certain vectors that can be used for the expression of antibodies, antibody fragments, and/or binding proteins described herein include plasmids that contain regulatory sequences, such as promoter and enhancer regions, which direct gene transcription. Other useful vectors for expression of antibodies, antibody fragments, and/or binding proteins contain polynucleotide sequences that enhance the rate of translation of these genes or improve the stability or nuclear export of the mRNA that results from gene transcription. These sequence elements include, e.g., 5’ and 3’ untranslated regions, an internal ribosomal entry site (IRES), and polyadenylation signal site in order to direct efficient transcription of the gene carried on the expression vector. The expression vectors described herein may also contain a polynucleotide encoding a marker for selection of cells that contain such a vector. Examples of a suitable marker include genes that encode resistance to antibiotics, such as ampicillin, chloramphenicol, kanamycin, or nourseothricin. Combination of an NK Engager and IL-15 In some aspects, the disclosure features a composition comprising an NK engager polypeptide(s) of this disclosure and a cytokine (e.g., IL-15). In some instances the IL-15 is sushi-IL-15. Atty. Docket No.45817-0158WO1 In other aspects, the disclosure features a composition comprising an mRNA encoding an NK engager polypeptide(s) of this disclosure and an mRNA encoding a cytokine (e.g., IL-15). In some instances the IL-15 is sushi-IL-15. In some cases, the mRNA comprises a 5’UTR and a 3’UTR. In some cases, the mRNA further comprises a 5’cap and a poly A tail. In some instances, all uracils in the mRNA or mRNAs are N1-methylpseudouracils. Exemplary nucleic acid and amino acid sequences of an IL15 construct that can be used in combination with an NK engager (e.g., BE-40) are provided below. It is to be understood that the IL-15 construct instead of comprising HSA may comprise a VHH that specifically binds to HSA. Sequence SEQ ID NO: C^{ap m G-ppp-Gm-A} 5’UTR AGGAAAUAAGAGAGAAAAGAAGAGUAAGAAGAAAUAUAAGACCCCGGCGCCGCCA 467 CC (v1.1 5'UTR A-start) ORF AUGAAGUGGGUGACCUUCAUCAGCCUGCUGUUCCUGUUCAGCAGCGCCUACAGCA 728 GAGGCGUGUUCAGAAGAGACGCCCACAAGAGCGAGGUGGCCCACAGAUUCAAGGA (HSA- sushiL- CCUGGGCGAGGAGAACUUCAAGGCCCUGGUGCUGAUCGCCUUCGCCCAGUACCUG IL15) CAGCAGUGCCCUUUCGAGGACCACGUGAAGCUGGUGAACGAGGUGACCGAGUUCG CCAAGACCUGCGUGGCCGACGAGAGCGCCGAGAACUGCGACAAGAGCCUGCACAC CCUGUUCGGCGACAAGCUGUGCACCGUGGCCACCCUGAGAGAAACUUACGGCGAG AUGGCCGACUGCUGCGCCAAGCAGGAGCCAGAGCGGAACGAGUGCUUCCUCCAGC ACAAGGACGACAACCCUAACCUGCCUAGACUGGUAAGGCCUGAGGUGGACGUGAU GUGUACCGCCUUCCACGACAACGAGGAGACAUUCCUGAAGAAGUACCUGUACGAG AUCGCCAGAAGACACCCUUACUUCUACGCCCCUGAGUUGCUGUUCUUCGCGAAGA GAUACAAGGCCGCCUUCACCGAGUGCUGCCAGGCCGCCGAUAAGGCCGCGUGCCU GCUGCCUAAGCUGGACGAGCUGAGAGACGAGGGCAAGGCAUCCAGCGCUAAGCAG AGACUGAAGUGCGCCAGCCUGCAGAAGUUCGGAGAGAGAGCUUUCAAGGCGUGGG CAGUGGCUAGAUUGAGCCAAAGAUUCCCUAAGGCAGAAUUCGCUGAGGUGAGCAA GCUCGUGACUGACCUGACCAAGGUGCAUACAGAGUGCUGUCACGGCGACCUGCUG GAGUGCGCCGACGACAGAGCCGACCUGGCCAAGUACAUCUGCGAGAACCAGGACA GCAUCAGCAGCAAGCUGAAGGAGUGUUGUGAGAAGCCUCUAUUGGAGAAGAGUCA CUGCAUUGCCGAGGUGGAGAACGACGAGAUGCCUGCGGAUCUGCCAAGCUUGGCG GCCGACUUCGUGGAGAGCAAGGACGUGUGCAAGAACUACGCCGAGGCCAAGGACG Atty. Docket No.45817-0158WO1 UUUUCCUGGGCAUGUUCCUCUACGAGUACGCACGCAGACAUCCAGACUACAGCGU GGUGCUGCUGCUGAGACUGGCUAAGACAUACGAAACUACCCUGGAGAAGUGCUGC GCAGCGGCGGACCCUCACGAGUGUUACGCCAAGGUGUUCGACGAGUUCAAGCCUC UGGUGGAGGAGCCUCAGAACCUGAUCAAGCAGAACUGUGAGCUGUUCGAGCAGCU CGGCGAGUACAAGUUCCAGAACGCCCUGUUGGUCCGCUACACCAAGAAGGUGCCU CAAGUAAGUACCCCUACCCUGGUAGAGGUUAGUAGAAACCUGGGCAAGGUGGGCA GCAAGUGCUGUAAGCACCCAGAAGCUAAGAGGAUGCCUUGCGCCGAGGACUACCU GUCCGUUGUGCUGAACCAGCUGUGCGUGCUGCACGAGAAGACCCCUGUGAGCGAC AGAGUGACAAAGUGUUGCACCGAGAGCUUAGUGAAUAGAAGACCUUGCUUCAGCG CCCUGGAAGUUGACGAAACUUACGUGCCUAAGGAGUUCAACGCCGAGACUUUCAC AUUCCACGCCGACAUUUGCACUCUGAGCGAGAAGGAGAGACAGAUCAAGAAGCAG ACCGCCCUCGUAGAGUUGGUCAAGCACAAGCCGAAGGCAACCAAGGAACAGCUUA AGGCCGUGAUGGACGACUUCGCAGCCUUCGUCGAGAAGUGUUGUAAGGCCGACGA CAAGGAGACUUGCUUCGCAGAGGAAGGCAAGAAGUUGGUAGCCGCCUCUCAGGCU GCCUUGGGACUCGGUGGCGGCAGCAUCACCUGCCCGCCACCCAUGAGCGUGGAGC ACGCCGACAUCUGGGUGAAGAGCUACAGCCUGUACAGCCGGGAGCGGUACAUCUG CAACAGCGGCUUCAAGCGGAAGGCCGGCACCAGCAGCCUGACCGAGUGCGUGCUG AACAAGGCCACCAACGUGGCCCACUGGACCACCCCUAGCCUGAAGUGUAUACGGG AUCCCGCCCUGGUGCAUCAGCGACCCGCCCCACCAUCUGGCGGCAGUGGAGGCGG AGGAAGUGGCGGUGGUAGCGGUGGAGGUGGCAGCCUGCAGAACUGGGUGAACGUG AUCAGCGACCUGAAGAAGAUCGAGGACCUGAUCCAGAGCAUGCACAUCGACGCCA CCCUGUACACCGAGAGCGACGUGCACCCCAGCUGCAAGGUGACCGCCAUGAAGUG CUUCCUGCUGGAGCUGCAGGUGAUCAGCCUGGAGAGCGGCGACGCCAGCAUCCAC GACACCGUGGAGAACCUGAUCAUCCUGGCCAACAACAGCCUGAGCAGCAACGGCA ACGUGACCGAGAGCGGCUGCAAGGAGUGCGAGGAGCUGGAGGAGAAGAACAUCAA GGAGUUCCUGCAGAGCUUCGUGCACAUCGUGCAGAUGUUCAUCAACACCAGC Encoded MKWVTFISLLFLFSSAYSRGVFRRDAHKSEVAHRFKDLGEENFKALVLIAFAQYL 729 AA QQCPFEDHVKLVNEVTEFAKTCVADESAENCDKSLHTLFGDKLCTVATLRETYGE sequence MADCCAKQEPERNECFLQHKDDNPNLPRLVRPEVDVMCTAFHDNEETFLKKYLYE IARRHPYFYAPELLFFAKRYKAAFTECCQAADKAACLLPKLDELRDEGKASSAKQ RLKCASLQKFGERAFKAWAVARLSQRFPKAEFAEVSKLVTDLTKVHTECCHGDLL ECADDRADLAKYICENQDSISSKLKECCEKPLLEKSHCIAEVENDEMPADLPSLA ADFVESKDVCKNYAEAKDVFLGMFLYEYARRHPDYSVVLLLRLAKTYETTLEKCC AAADPHECYAKVFDEFKPLVEEPQNLIKQNCELFEQLGEYKFQNALLVRYTKKVP QVSTPTLVEVSRNLGKVGSKCCKHPEAKRMPCAEDYLSVVLNQLCVLHEKTPVSD RVTKCCTESLVNRRPCFSALEVDETYVPKEFNAETFTFHADICTLSEKERQIKKQ TALVELVKHKPKATKEQLKAVMDDFAAFVEKCCKADDKETCFAEEGKKLVAASQA ALGLGGGSITCPPPMSVEHADIWVKSYSLYSRERYICNSGFKRKAGTSSLTECVL NKATNVAHWTTPSLKCIRDPALVHQRPAPPSGGSGGGGSGGGSGGGGSLQNWVNV ISDLKKIEDLIQSMHIDATLYTESDVHPSCKVTAMKCFLLELQVISLESGDASIH DTVENLIILANNSLSSNGNVTESGCKECEELEEKNIKEFLQSFVHIVQMFINTS 3’UTR UGAUAAUAGGCUGGAGCCUCGGUGGCCUAGCUUCUUGCCCCUUGGGCCUCCCCCC 468 AGCCCCUCCUCCCCUUCCUGCACCCGUACCCCCGUGGUCUUUGAAUAAAGUCUGA v1.1 3'UTR GUGGGCGGC Atty. Docket No.45817-0158WO1 T^{ail Poly A (100 nt in length)} ₇₃₀ Note: All uracils in this table are N1-methyl pseudouracils. These combinations can be used to treat a human subject with a cancer or tumor. EXAMPLES The following examples provide those of ordinary skill in the art with a description of how the compositions and methods claimed herein can be performed, made, and evaluated, and are intended to be purely examples and are not intended to limit the scope of the present disclosure. As set forth in more detail herein, single mRNA molecules encoding bispecific engagers can be delivered to cells and expressed to target B cells for reduction or elimination and NK cells. The expression of multiple bispecific engager molecules in cells can improve the targeting and depletion of cell populations that are not homogenous, e.g., that have variable expression of tumor associated antigens (TAAs). In addition, these examples demonstrate the ability to produce multiple Fc bispecific engager molecules in cells by transfecting two or more mRNAs that encode for different bispecific engager molecules targeting different molecules on B cells. A potential loss of potency could occur when coexpressing two different Fc bispecific engager sequences due to recombination, which could generate a heteromeric bispecific engager molecule. Surprisingly, multiple Fc engagers can be expressed without losing potency. The following materials and methods were used in the examples: Flow cytometry assays: BCMA expressing cell line RPMI8226 and engineered cell lines expressing FcRH5 and both BCMA and FcRH5 such as Molm13-FcRH5 and RPMI8226-FcRH5 were resuspended in 2% H.I. FBS + PBS (FACS buffer) (Gibco [Waltham, MA], Catalog No.10010-023) at 2.5 x 10⁶ cells/mL, and 200 µL of the cell suspension was mixed in a 96-well round bottom Atty. Docket No.45817-0158WO1 tissue culture treated microplate (Corning [Corning, NY], Catalog No.3799). Cells were centrifuged at 1200 rpm for 5 minutes at 4⁰C and were washed 1x with PBS. Cells were resuspended in 200 µL of eBioscience™ Fixable Viability Dye eFluor™ 780 (ThermoFisher Scientific Catalog No.65-0865-18), diluted 1:500 in PBS, incubated for 20 minutes at 4⁰C. Cells were centrifuged at 1200 rpm for 5 minutes at 4⁰C, and incubated with 100 µL of Human TruStain FcX™ (Fc Receptor Blocking Solution) (BioLegend [San Diego, CA], Catalog No.422302) diluted 1:200 in FACS buffer for 15 minutes at 4⁰C, followed by staining with 100 µL of PE anti-human CD307e (FcRL5) Antibody (BioLegend [San Diego, CA], Catalog No.340304), and PE/Dazzle™ 594 anti-human CD269 (BCMA) Antibody (BioLegend [San Diego, CA], Catalog No.357512), diluted 1:50 in FACS buffer for 20 minutes at 4⁰C. Cells were centrifuged at 1200 rpm for 5 minutes at 4⁰C and washed 1X in PBS. Cellular fluorescence data was acquired on a BD LSRFortessa (Franklin Lakes, NJ) and analyzed using FlowJo software (FLOWJO [Ashland, OR]). In vitro cytotoxicity assay: Purified frozen human peripheral blood NK cells (Stemcell Technologies [Vancouver, Canada], Catalog No.70036) were thawed and rested in complete RPMI-1640 medium (ATCC Catalog No.30-2001) overnight. Target cells were stained with 50-100 µM Calcein, AM, cell-permeant dye (Thermo Fisher Scientific Catalog No. C3100MP) in serum free medium for 30 minutes at 37⁰C.15,000 washed target cells were combined with 112,500 NK cells to achieve desired effector (NK) to target (E:T) ratios of 7.5:1 and plated in a 96-well round bottom tissue culture treated microplate (Corning [Corning, NY], Catalog No.3799). Test articles prepared in serial dilutions were added to the co-culture and target cell lysis was measured after a 4h incubation at 37⁰C by fluorescence reading at 485/528 nM in the supernatants. Target cells alone wells were used as spontaneous calcein release control, and maximum killing was recorded from Triton X (1% final concentration) (Thermo Fisher Scientifc, Catalog No.85111) treated wells. Percent of specific killing was calculated as (F[sample]-F[spontaneous])/(F[maximum]- F[spontaneous])x100. Atty. Docket No.45817-0158WO1 MSD protein determination assays: mRNA-derived engager protein concentrations in tissue culture supernatant were quantified using a custom MSD assay. For the MSD assay, 96-well Multi-Array Sector plates (Meso Scale Diagnostics [Rockville, MD], Catalog No. L15XA-3) were coated with 40 μL per well of diluted capture antigen in 1X phosphate buffered saline (PBS) (Gibco [Waltham, MA], Catalog No.10010-023). Plates were sealed (VWR [Radnor, PA], Catalog No.60941-062) and incubated at 4^oC overnight (minimum of 8 to 24 hours) without shaking. Next day, plates were washed 3 times with 300 µL of 1X MSD Tris Wash Buffer (Meso Scale Diagnostics [Rockville, MD], Catalog No. R61TX-1). Thereafter, plates were blocked with 150 µL of MSD Blocker A solution (Meso Scale Diagnostics, Catalog No. R93AA-2). Plates were sealed and incubated for 1 hour at room temperature with shaking at 700-900 RPM. Plates were washed 3 times with 300 µL of 1X MSD Tris Wash Buffer and 25 µL of diluted standards, samples, blanks in Diluent 100 (Meso Scale Diagnostics, Catalog No. R50AA-3) were transferred to the plate. Plates were sealed and incubated at room temperature for 1 hour with shaking at 700-900 RPM. Plates were washed 3 times with 300 µL of 1X MSD Tris Wash Buffer and 25 µL of Detection solution diluted with Diluent 100 was transferred to the plate. Plates were sealed and incubated at room temperature for 1 hour with shaking at 700-900 RPM. Plates were washed 3 times with 300 µL of 1X MSD Tris Wash Buffer and 25 µL of Streptavidin Sulfo-Tag Labeled detection reagent (Meso Scale Diagnostics, Catalog No. R32AD-1) diluted with Diluent 100 was added to the plate and incubated for 1 hour at room temperature with shaking at 700-900 RPM. Plates were washed 3 times with 300 µL of 1X MSD Tris Wash Buffer and 150 µL of MSD Read Buffer T (4X with surfactant, diluted 1:2) (Meso Scale Diagnostics, Catalog No. R92TC-1) was added to the wells. Plates were immediately read on MESO SECTOR S 600 reader (Meso Scale Diagnostics, Catalog No. MESO SECTOR S 600) and analyzed using MSD Discovery Workbench software (Meso Scale Diagnostics) with raw data processed and exported directly from the software to Microsoft Excel to perform the final concentration calculations using dilution factor. Data were graphed in GraphPad Prism, v7.03 (GraphPad

Atty. Docket No.45817-0158WO1 Tbl 14 P ti dtil f i i diid l ti

Atty. Docket No.45817-0158WO1 F i l h k 250 d i b d 8% CO2 d

Initially each bispecific Fc engager was expressed recombinantly in 293 cells and the resulting proteins purified and characterized as shown in Table 15. Each purified homogenous bispecific protein was evaluated for a killing EC50 value against the target cell lines. Three different bispecific Fc engager molecules were produced using different anti-TAA. These included an engager comprising an anti- BCMA antibody (SEQ ID NO: 547) paired with the anti-CD16a mAb (SEQ ID NO: 7) using two different human Fc domains. The Fc domains used were either an hu- IgG1 containing several of point mutations (L234F, L235E and D265A, “FEA”)(Liu, 2020) to abrogate Fc G1 functionality (BE-103) or a stabilized IgG4 that contains a PAA sequence in the hinge region to prevent in vivo scrambling with other IgG4 Atty. Docket No.45817-0158WO1 antibodies that could be found in human sera (Giles, 1999, Saunders 2019,) (BE-102). The second bispecific was an anti-FCRH5 VHH antibody (SEQ ID NO: 392) paired with the anti-CD16a mAb (SEQ ID NO: 7), which was expressed only as the IgG4- PAA bispecific (BE-40). Table 15 – Concentrations of mRNA expressed molecules mRNA construct(s) mRNA Concentration transfected (µg/well) (ng/ml) of molecule(s) BE-40 1 3671 2 4237 BE-102 1 2054 2 3204 BE-103 1 2927 2 3688 BE-40 1 1704 BE-102 1 1470 Combination (2) (3174) BE-40 1 1649 BE-103 1 1836 Combination (2) (3485) To evaluate the ability of the bispecific antibodies to induce T-cell dependent depletion (killing) three different cell lines were selected for the bispecific antibody potency assays: Mol13-FCRH5 hi, which only expresses high levels of FCRH5, RPMI8226-parental which only expresses a low level of BCMA, and a cell line RPMI8226-FcRH5 hi that expresses both a low level BCMA level and high level of FCRH5. Fig.2 shows the level of expression of FCRH5 and BCMA for each of these cell lines. Next, the potency of each bispecific engager antibody to deplete target cells was determined using the protein generated from mRNA transfected HeLa cell supernatants. Each mRNA encoded bispecific was transfected into Hela cells and the EC50 and Emax of the supernatants based on depletion of the target cells was determined (Table 16). The concentration of each bispecific in the mRNA transfected Atty. Docket No.45817-0158WO1 HeLa cell supernatants was determined using the appropriate MSD assay as described in the methods. For the MSD assay each bispecific Fc engager molecule was captured in the MSD assay using the appropriate recombinant rhuFCRH5 or rhuBCMA. The bound bispecific antibody was then detected using biotinylated CD16a in combination with a streptavidin sulfo tagged detection molecule. Using the combination of the BCMA/CD16 or FCRH5/CD16 reagents insured that only a bispecific antibody containing the appropriate B cell binding arm, BCMA or FCRH5 was detected in the assay. The standard was a single bispecific engager, either anti-BCMA or anti- FCRH5. Because A control for the heteromeric bispecific engager containing both B cell binding moieties was lacking, this standard was used so that engager molecules containing both anti-FCRH5 and anti-BCMA VHH antibodies could be detected in both the BCMA and FCRH5 assay. Table 16 – Bispecific constructs expressed in 293 cells were purified and characterized. Each molecule was then titrated into killing assays and the commensurate EC50 and Emax determined in picomolar (pM) K_{illing assay potency EC50/Emax (pM)} α-B cell ^protein _{Molm13 FCRH5 hi} RPMI 8226 RPMI 8226 FCRH5 hi binding (BCMA-, FCRH5++) (BCMA+, FCRH5-) (BCMA+, FCRH5++) moiety FCRH5 BE-40 4.8/107.8 No depletion 5.2/47.5 BCMA BE-102 No depletion 7.2/46.1 15.7/45.4 BCMA BE-103 No depletion 6.4/48.2 9.3/47.3 The associated cellular depletion curves for each individual encoded molecule are shown in Fig.3A-3C. Each plot contains a particular cell line, Fig.3A (Molm13- FCRH5 hi) expressing FCRH only, Fig.3B (RPMI) expressing only BCMA, and Fig. 3C (RPMI8226-FCRH5 hi) expressing both BCMA and FCRH5. The expression results for single or combinations of BCMA and/or FCRH5 Fc engager bispecific antibodies is described in Table 16 above. The concentration of bispecific Fc engager molecules expressed was determined by the MSD. These determinations allowed for Atty. Docket No.45817-0158WO1 a comparison of the potency of the bispecific Fc engager molecules in the mRNA transfected Hela supernatants with the purified proteins previously tested. A negative control included in each of the single bispecific engager potency assays showed no killing of the cells lacking the appropriate antigen/target. In Table 17, the determined EC50 and Emax of cell depletion values for each single Fc bispecific engager were compared to the potency of a single mRNA or two mRNAs that encode a single and combination of bispecific Fc engagers, respectively. Table 17 – Bispecific constructs transfected into Hela cells and the corresponding potency of the molecules in killing of target cell lines Molm 13FCRH5 Hi cells, RPMI226 and RPMI 8226 expressing FCRH5 mRNA expressed bispecific Potency of bispecific killing of target cell (pm) Molm13 FCRH5 RPMI 8226 RPMI 8226- BE-40 BE-102 BE-103 hi (BCMA+,FCRH5-) FCRH hi (BCMA-, (BCMA+, FCRH5++) FCRH5++) µg mRNA) EC50 Emax EC50 Emax EC50 Emax 2 - - 9.5/125.7 No depletion 18.3/48.7 1 - - 5.5/136.0 No depletion 18.6/42.4 - 2 - No depletion 22.4/28.5 39.2/32.1 - 1 - No depletion 18.8/33.7 33.2/34.1 - - 2 No de letion 194/375 333/337

Atty. Docket No.45817-0158WO1 of a bispecific targeting a transfected cell line resulted in depletion of the target cell line. Conclusions: Combinations of mRNA encoding engager molecules transfected into and expressed by cells were able to produce nearly equivalent concentration levels of total bispecific antibodies compared to transfection of each of the bispecific antibodies separately. The activity of the individual bispecific antibodies was assessed on each of three cell lines, two of the cell lines Molm13-FCRH5 hi and RPMI8226 express a targeted antigen (FCRH5 or BCMA respectively) and one cell line RPMI8226- FCRH5 hi expresses both FCRH5 and BCMA. Transfection of mRNAs encoding two bispecific engagers targeting both FCRH5 and BCMA expressed bispecific Fc engager proteins that demonstrated comparable and, in many cases, superior in EC₅₀ and Emax, compared to single secreted Fc engagers for the cell line that expressed both TAAs. This demonstrates that the combination of mRNAs produced molecules that retained their potency and that the expression of bispecific molecules in combination produce highly potent molecules that can effectively target cells carrying more than a single target molecule. For treatment of diseases such as multiple myeloma, including relapsed or refractory multiple myeloma, a single formulation (e.g., in the same delivery vehicle or separate delivery vehicles) using this format can produce multiple, effective molecules. This gives a distinct advantage as the expression of target molecules on cancer cells is often diverse and agents targeting a single molecule are not able to achieve complete remission owing to depletion of only the fraction of the cancer cells. Using the subject mRNA constructs, a multitude of engagers can be produced in vivo that have the potential to target and deplete cancer cells that express either targeted TAA. The treatment with multiple engagers is expected to improve the treatment and achieve complete remission as shown in Fig.4. Sequences used for Example 1: Atty. Docket No.45817-0158WO1 BE-102 Nucleotide Sequence (note that all uracils in the sequence are N1- methylpseudouracils): AUGGAGACACCCGCCCAGCUGCUGUUCCUGCUACUGCUGUGGCUGCCCG ACACCACAGGCGAGGUGCAACUGGUGGAAUCAGGCGGCGGACUGGUGC AGCCCGGAGGAAGCCUGCGGCUGAGCUGUGCCGCCAGCGGCUUUACCUU UACCGCCUACGACAUGGGCUGGGUGCGGCAAGCACCAGGCAAGGGCCCC GAGUGGGUGAGCCUGAUCAGCAGCGACAGCGGCGACACCUGGUACGAC GACUCCGUGAAGGGCCGGUUCACCAUCAGCCGGGACAACAGCAAGAACA CGCUGUAUCUGCAGAUGAACAGCCUCAGGGCCGAGGACACAGCGGUGU ACUAUUGUGCCCGGCUGGGAGCCUACAGCACCACCUACGACUACUGGGG ACAGGGAACCCUAGUGACCGUGAGCAGCGAGAGCAAGUACGGACCUCCU UGCCCUCCCUGUCCAGCCCCAGAGGCCGCUGGAGGCCCAAGCGUGUUCC UGUUCCCACCCAAGCCCAAGGACACCCUGAUGAUCAGCCGGACCCCAGA GGUGACCUGCGUGGUGGUGGACGUGAGCCAGGAGGACCCCGAGGUGCA GUUCAACUGGUACGUGGACGGCGUGGAGGUGCACAACGCCAAGACCAA GCCCCGGGAGGAGCAGUUCAACAGCACCUACCGGGUGGUGAGCGUGCUG ACCGUGCUGCACCAGGACUGGCUGAACGGCAAGGAGUACAAGUGCAAG GUGAGCAACAAGGGCCUGCCCAGCAGCAUCGAGAAGACCAUCAGCAAGG CAAAGGGCCAACCUCGGGAGCCACAGGUGUACACCCUGCCUCCCAGCCA GGAGGAGAUGACCAAGAACCAGGUGAGCCUGACCUGCCUGGUGAAGGG CUUCUACCCCAGCGACAUCGCCGUGGAGUGGGAGAGCAACGGCCAGCCC GAGAACAACUACAAGACCACUCCACCAGUGCUGGACAGCGACGGCAGCU UCUUCCUGUACAGCCGGCUGACCGUGGACAAGAGCCGGUGGCAGGAGG GCAACGUGUUCAGCUGCAGCGUGAUGCACGAGGCCCUGCACAACCACUA CACCCAGAAGAGCCUGAGCCUCAGCCUGGGCAAGGGAGGUGGCGGAAGC GAGGUGCAGCUGGUGGAAAGCGGCGGCGGCCUGGUGCAACCCGGCGGC AGCCUGCGACUGUCCUGUGCCGCCUCUGGCCGGACCGACAGCAUCUACG CGGGAGUUCGUGA GCCGACAGCGUGAA ACAUGGUGUACCU UGUACUAUUGCGCC

Atty. Docket No.45817-0158WO1 GCCGGACGGGGUUACGGCCUGCUGAGCAUCAGCAGCAACUGGUACAACU (SEQ ID NO: 461). leader sequence): SLRLSCAASGFTFTA

YDMGWVRQAPGKGPEWVSLISSDSGDTWYDDSVKGRFTISRDNSKNTLYLQ MNSLRAEDTAVYYCARLGAYSTTYDYWGQGTLVTVSSESKYGPPCPPCPAPE AAGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVH NAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTIS KAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPEN NYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSL SLSLGKGGGGSEVQLVESGGGLVQPGGSLRLSCAASGRTDSIYAMGWFRQAP GKEREFVSAINSNTGRTYHADSVKGRFTISRDNAKNMVYLQMNSLRAEDTA VYYCAAGRGYGLLSISSNWYNYWGQGTLVTVSS (SEQ ID NO: 462). >BE-40 Nucleotide Sequence (note that all uracils in the sequence are N1- methylpseudouracils): AUGGAGACCCCUGCCCAGCUGCUGUUCCUGCUGCUGCUGUGGCUGCCCG ACACCACCGGCCAGGUGCAGUUGCAGGAGUCCGGCGGAGGCCUGGUCCA ACCCGGCGGCUCACUGCGGCUUAGCUGCGCCGCAAGCGGCCGAACCUAC AACAACUACGCCAUGGGGUGGUUUCGGCAAGCCCCAGGCAAGGAACGG GAGUUCGUGGCCGGCAUCAGCCGGAGCGGCGGCAUGACCGGCUACGCCG AGAGCGUGAAGGGUCGGUUCACCAUUAGCCGGGAUAACAGCAAGAAUA CCGUCUACCUUCAGAUGAACAGCCUUAGAGCCGAGGACACCGCUGUUUA CUACUGUGCCGCCUACGUGGGCGGCUUCAGCACCGCCCGGCGGGACUAC AGCUACUGGGGACAGGGGACCCAAGUGACAGUGAGCAGCGAGAGCAAG UACGGACCUCCUUGCCCUCCCUGUCCAGCCCCAGAGGCCGCUGGAGGCC CAAGCGUGUUCCUGUUCCCACCCAAGCCCAAGGACACCCUGAUGAUCAG CCGGACCCCAGAGGUGACCUGCGUGGUGGUGGACGUGAGCCAGGAGGA CCCCGAGGUGCAGUUCAACUGGUACGUGGACGGCGUGGAGGUGCACAA CGCCAAGACCAAGCCCCGGGAGGAGCAGUUCAACAGCACCUACCGGGUG GUGAGCGUGCUGACCGUGCUGCACCAGGACUGGCUGAACGGCAAGGAG Atty. Docket No.45817-0158WO1 UACAAGUGCAAGGUGAGCAACAAGGGCCUGCCCAGCAGCAUCGAGAAG ACCAUCAGCAAGGCAAAGGGCCAACCUCGGGAGCCACAGGUGUACACCC UGCCUCCCAGCCAGGAGGAGAUGACCAAGAACCAGGUGAGCCUGACCUG CCUGGUGAAGGGCUUCUACCCCAGCGACAUCGCCGUGGAGUGGGAGAGC

ACAGCAUCUACGCCAUGGGCUGGUUCCGGCAGGCACCCGGCAAGGAGCG GGAGUUCGUGAGCGCCAUCAACAGCAACACCGGCCGGACCUACCACGCC GACAGCGUGAAGGGCCGGUUCACCAUCAGCCGGGACAACGCCAAGAACA UGGUGUACCUGCAGAUGAACAGCCUGCGGGCCGAGGAUACCGCCGUGU ACUAUUGCGCCGCCGGACGGGGUUACGGCCUGCUGAGCAUCAGCAGCAA CUGGUACAACUACUGGGGCCAGGGCACCCUGGUGACCGUGAGCAGC (SEQ ID NO: 463). BE-40 Amino Acid Sequence (before processing of leader sequence): METPAQLLFLLLLWLPDTTGQVQLQESGGGLVQPGGSLRLSCAASGRTYN NYAMGWFRQAPGKEREFVAGISRSGGMTGYAESVKGRFTISRDNSKNTVYL QMNSLRAEDTAVYYCAAYVGGFSTARRDYSYWGQGTQVTVSSESKYGPPCP PCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVD GVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSS IEKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESN GQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNH YTQKSLSLSLGKGGGGSEVQLVESGGGLVQPGGSLRLSCAASGRTDSIYAMG WFRQAPGKEREFVSAINSNTGRTYHADSVKGRFTISRDNAKNMVYLQMNSL RAEDTAVYYCAAGRGYGLLSISSNWYNYWGQGTLVTVSS (SEQ ID NO: 464). BE-103 Nucleotide Sequence (note that all uracils in the sequence are N1- methylpseudouracils): Atty. Docket No.45817-0158WO1 AUGGAGACCCCUGCCCAGCUGCUGUUCCUGCUGCUGCUGUGGCUGCCCG ACACCACCGGCGAGGUGCAGCUGGUCGAAAGCGGCGGCGGAUUGGUCCA ACCCGGCGGCUCGUUACGCCUGAGUUGUGCCGCGAGCGGCUUCACCUUC ACCGCCUACGACAUGGGGUGGGUCAGACAGGCUCCUGGCAAGGGCCCUG

CAACCCCGAGAGCCCCAGGUGUAUACACUGCCUCCGAGCCGGGACGAGC UGACCAAGAACCAGGUGAGCCUGACCUGCCUGGUGAAGGGCUUCUACCC CAGCGACAUCGCCGUGGAGUGGGAGAGCAACGGCCAGCCCGAGAACAAC UACAAGACCACCCCGCCUGUGCUGGACAGCGACGGCAGCUUCUUCCUGU ACAGCAAGCUGACCGUGGACAAGAGCCGGUGGCAGCAGGGCAACGUGU UCAGCUGCAGCGUGAUGCACGAGGCCCUGCACAACCACUACACCCAGAA GAGCCUGUCUCUGUCACCAGGCGGAGGCGGCGGAAGCCAGGUGCAGCUG GUGGAGAGCGGCGGAGGUCUGGUGCAGGCCGGCGGUAGCCUGAGGCUG AGUUGCGCCGCUAGCGGCAGAACCGACAGCAUCUACGCCAUGGGCUGGU UCCGGCAGGCACCCGGCAAGGAGCGGGACUUCGUGGCCGCCAUCAACAG CAACACCGGCCGGACCUACCACGCCGACAGCGUGAAGGGCCGGUUCACC AUCAGCCGGGACAACGCCAAGAACAUGGUGUACCUGCAGAUGAACAGCC UGAAGCCCGAGGACACCGCCGUGUACUAUUGCGCUGCUGGCCGGGGAUA CGGCCUGCUGAGCAUCAGCAGCAACUGGUACAACUACUGGGGCCAGGGU ACCCAGGUGACCGUGAGCAGC (SEQ ID NO: 465). Atty. Docket No.45817-0158WO1 BE-103 Amino Acid Sequence (before processing of leader sequence): METPAQLLFLLLLWLPDTTGEVQLVESGGGLVQPGGSLRLSCAASGFTFTA YDMGWVRQAPGKGPEWVSLISSDSGDTWYDDSVKGRFTISRDNSKNTLYLQ MNSLRAEDTAVYYCARLGAYSTTYDYWGQGTLVTVSSDKTHTCPPCPAPEF EGGPSVFLFPPKPKDTLMISRTPEVTCVVVAVSHEDPEVKFNWYVDGVEVHN AKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISK AKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENN YKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLS LSPGGGGGSQVQLVESGGGLVQAGGSLRLSCAASGRTDSIYAMGWFRQAPG KERDFVAAINSNTGRTYHADSVKGRFTISRDNAKNMVYLQMNSLKPEDTAV YYCAAGRGYGLLSISSNWYNYWGQGTQVTVSS (SEQ ID NO: 466). Example 2: Evaluation of NK Binding Moieties Methods: Selection of Exemplary NK cell binding moieties: Immunization and anti- CD16a VHH antibody selection: Llamas were immunized with either formulated protein that encoded huCD16a then cynoCD16a and immunized subcutaneously on an alternating basis every two weeks. The first dose of huCD16a was in complete Freund’s adjuvant, all following immunizations were with incomplete Freund’s adjuvant. After four immunizations sera from animals were tested for binding in ELISA assays to hu and cyno CD16a. At this point a sufficient titer was reached animals were given an IV dose of huCD16a protein in PBS and PBMCs were harvested from a bleed three days post the IV dose. B-cell sorting and antibody selection: B-cells were sorted at a concentration of 1 x 10⁶ cells/mL on a Sony FACS-Sorter while gating for double positive VhH and CD16a positive antibodies using biotinylated CD16a, streptavidin-APC and Rabbit anti-Vhh-FITC (Rabbit anti-camelid VHH AF-647, Genescript). Cells were sorted three to a well and cultured using the methods of J Immunol 2016; 197:4163-4176. The sorted B-cells were then cultured with CD154-expressing stromal cells expressing huCD154. B cells were cultured in R5 medium (RPMI 1640 with 5% Atty. Docket No.45817-0158WO1 human serum [Sigma], 55 mM 2-ME, 2 mM L-glutamine, 100 U/ml penicillin, 100 mg/ml streptomycin, 10 mM HEPES, 1 mM sodium pyruvate, and 1% MEM nonessential amino acids [Invitrogen]. Cells were then cultured for 7-10 days and screened for binding to huCD16a and cyno CD16 in using ELISA binding assays. The individual CD16 molecules were coated at 2 µg/mL on a plate and 3-fold diluted supernatants were evaluated for anti-body binding using anti-llama Fc-HRP detection. Cells that were positive were subject to RT-TAP (Clargo et al.2014) to produce VHH antibodies. Cell binding of VHH-Fc fusions, FACS Assays and Analysis: CHO cell lines that stably express CD16a and CD16b were resuspended in 1% H.I. FCS 1x PBS (FACS buffer) at 500,000 cells/ml, and 100 µl of the cell suspension was mixed in a 96-well V bottom polypropylene plate (Nunc) with 100 µl of VHH-Fc fusion molecules in the same buffer. After one-hour incubation at 4°C, pelleted cells were washed 3x and then resuspended in 100 µl Goat anti-human IgG Fc – PE (Jackson ImmunoResearch Labs, West Grove, PA), diluted 1/300 in FACS buffer, incubated 1h at 4°C, centrifuged at 1200 rpm for 3 min; cell pellets were washed, fixed with 150 µl/well 1% paraformaldehyde for 10 min at room temperature, centrifuged, the fix solution was decanted and cell pellets were resuspended in 175 µl FACS buffer for analysis. Cellular fluorescence was determined on FACScan equipped with FloJo software (Becton-Dickinson, Franklin Lakes, NJ). The data were plotted as a function of mean channel fluorescence versus the concentration; the receptor EC50 values were determined by fitting the data with a single site total binding model using GraphPad Prism 6.0 (La Jolla, CA). FACS binding analysis of CD16-VHH4 to neutrophils and NK cells was carried out in a similar manner. Transient protein expression: Proteins were transiently expressed by co- transfecting separate plasmids into ExpiCHO-S™ Cells (Thermofisher) that express the appropriate heavy and light chain genes for heavy and light chain antibodies. Single plasmids encoding anti-TAA or anti-CD16 sdAbs fused to the IgG4-PAA format or in line were built into single plasmids. The anti-TAA/CD16 molecules were transiently expressed in ExpiCHO cells using the EpxiFectamine CHO Atty. Docket No.45817-0158WO1 (Thermofisher) transfection reagents. The ExpiCHO-S cells were cultured in ExpiCHO expression medium (ThermoFisher Scientific) in a shaker incubator set at 125 rpm, 37 ^ C and 8% CO2. The day prior to transfection, ExpiCHO-S cells were seeded at 3 x 10⁶ cells per ml in 30 ml of ExpiCHO Expression medium. On the day of transfection cells were split using pre-warmed (37 ^ C) ExpiCHO expression media to a density of 6 x 10⁶ cells per ml. Using the manufacturer’s recommended protocol, a total 20 µg of a single plasmid and 72 µL of ExpiFectamine CHO reagent were mixed in 2.4 ml of cold Opti-PRO SFM (Thermo Fisher Scientific), after incubating the mixture for 2 minutes it was then slowly added to the cells. One day (~20h) post- transfection, 7.2 ml of ExpiCHO Feed and 180 µl of ExpiCHO Enhancer were added to the ExpiFectamine-transfected cultures. Culture supernatants were harvested when cell viability dropped below 60% (~day 8), clarified by centrifugation at 3,000 rpm for 30 min and filtered using a 0.2 μm filter (Thermo Fisher Scientific). Purification of single domain antibodies: Expressed single domain antibodies with a 6xHis C-terminal tag (SEQ ID NO: 731) were purified from ExpiCHO media by capture on TALON (TAKRA) cobalt immobilized metal affinity chromatography (IMAC) resin. Resin was prewashed in 50 mM sodium phosphate buffer pH 7.4 (wash buffer) with 15x the volume of resin, then centrifuged for 2 min @ 700 x g to pellet the resin. The supernatant discarded and the resin wash repeated. To the filtered media containing the expressed protein was added 3 mL of IMAC resin, then allowed to bind overnight with gentle shaking. The media resin mixture was gravity loaded into a 25 ml column. Columns containing IMAC resin were then washed with 10 column volumes (~30ml) of wash buffer (PBS pH 7.4 (Life Tech cat# 10010- 023)., 2 mM imidazole) then 2x with 10ml of wash buffer. Protein was then eluted using a total of 7.5 mL 150 mM imidazole /PBS pH 7 in three 2.5 mL aliquots. Proteins were then dialyzed exhaustively using Slide-A-Lyzer® 10 or 3K as appropriate, (Dialysis cassette, Pierce) versus 1 x PBS (100mM NaPO4 pH 6.8, 200mM NaCl). Alternatively, antibodies were purified using Protein A. Protein A purification: The Protein A column was prewashed and equilibrated with 20 times of column volume (CV) of PBS buffer pH 7.4 (wash Atty. Docket No.45817-0158WO1 buffer) at 1ml/min. The media containing the expressed protein was filtered and loaded into the column at 1 ml/min. The column was then washed with 20 times CV of wash buffer at 1ml/min. Protein was then eluted using 20 times CV of elution buffer (citrate buffer pH3.4) and collected at 500 µl per vial. The eluted proteins were detected by NanoDrop at 280 nm. Fractions with protein were pooled and then dialyzed exhaustively into 1 x PBS. The proteins then were concentrated into desired concentration using spin concentrator. Protein characterization: SDS PAGE was run on each sample using gradient gels NuPAGE Bis-Tris 4-12% gradient gels using a MES running buffer (Thermo Fisher Scientific). Samples were prepared with either reducing or nonreducing sample buffer and briefly heated to 95 ^ C. N-ethyl maleimide was added to samples electrophoresed in nonreducing buffer to cap any free thiols and prevent unwanted disulfide scrambling as the samples cooled. Molecular weight standards (Blue Plus protein, Thermofisher) were included on the SDS-PAGE. Non-denaturing protein electrophoresis was performed running 1 μg of each purified protein sample; reducing conditions were performed mixing each purified sample with 10 μl of Sample Reducing Agent (Invitrogen, Carlsbad, CA) and heating at 70°C for 10 min before electrophoresis on NuPAGE 4-12% Bis-Tris Mini Gels 1.0 mm (Invitrogen, Carlsbad, CA). The bands were visualized by SimplyBlue™ SafeStain (Invitrogen, Carlsbad, CA) staining, and the gel was dried using DryEase Mini-Gel Drying System (Invitrogen, Carlsbad, CA). All procedures were performed according to the manufacturer’s instructions. Analysis of native molecule homogeneity and determination of molecular weight was done using Size Exclusion Chromatography with Light Scattering (SEC- LS) when indicated. Size exclusion chromatography (SEC) was carried out on a Zenix SEC 3004.6 x 300 mm (Sepax Technologies) in 20 mM sodium phosphate pH 7.2, 150 mM NaCl (PBS), 0.05% NaAzide at a flow rate of 0.35ml/min using an Agilent 1260 UPLC. In addition to UV detection, the eluent was monitored with a refractive index detector (Waters, Milford, MA). Light scattering was monitored using a Wyatt Atty. Docket No.45817-0158WO1 Dawn 18 angle, coupled with an Optrex refractometer. Intact mass of molecules was determined mass spectrometry (Merrigen, Lowell MA). SPR Binding assay determinations of VHH-his antibodies: Briefly, binding of an anti-CD16 VHH antibody molecule (SEQ ID NO: 1; “CD16-VH1”) to various CD16 molecules was analyzed on a Biacore T200 (Cytiva) at 25°C. The CD16- VHH1-His was covalently coated to CM5 sensor chip by amine-coupling using the coupling kit (Cytiva). Various Fc receptor molecules were injected over each flow cell at the flow rate of 30 µl /min in HBS-EP+ buffer at concentrations ranging from 3 nM to 200 nM. A buffer injection served as a negative control. Channel 1 was blocked by ethanolamide and served as a reference channel. Upon completion of each association and dissociation cycle, surfaces were regenerated with 10 nM glycine pH 2.5 solution. The association rates (ka), dissociation rate constants (kd), and affinity constants (KD) were calculated using Biacore T200 evaluation software. Each fit was evaluated by the agreement between experimental data and the calculated fits, where the Chi2 values were below 10% of Rmax. Surface densities of the molecule were optimized to minimize mass transfer while maintaining enough response. All ka, kd, KD reported here represent the means and standard errors of at least two experiments. SPR binding on an OCTET RED96 was used to analyze the affinity CD16- VHH1 humanized antibodies for target CD16a. Biotinylated CD16a V176 was loaded to Streptavidin sensor to 0.5. A concentration series of 50 nM to 1.56 nM along with a 0 nM control was used to measure the kinetics with association phase for 240 seconds and dissociation phase for 320 seconds. Running buffer was 1X kinetic buffer from OCTET. The supernatant was then injected into a BSA pre-loaded CM5 Chip in a BiaCore 8K.200 nM CD16a was injected as analyte for 120 seconds and dissociation was carried out in 360 seconds. All the data were processed using the Biacore 8K Evaluation software version 1.1. Flow cell 1 and blank injection of buffer in each cycle were used as double reference for Response Units subtraction. Three negative Atty. Docket No.45817-0158WO1 controls were included in the assay: 3H01 is an irrelevant VHH, NC is an irrelevant antibody, and blank is buffer. Humanization: The VHH antibodies were humanized. Briefly, sequences of the complementarity-determining regions (CDRs) of CD16-VHH1 were annotated using the IMGT numbering scheme. The nucleotide sequence was generated and used to identify the nearest human germline VH sequences by searching for similar sequences with the NCBI IgBLAST program. Common J and D gene sequences were attached to the VH as the acceptor. Next the most similar human VH sequences are identified using BLASTp and used to choose the nearest framework sequences into which the CDR sequences are grafted replacing the human CDRs. Rosetta was used to create the structural 3D homology model the of the CD16- VHH1 CDRs that were grafted into the acceptor framework. The framework residues that were critical for huVH/VL interactions are back mutated to llama sequence canonical llama residues, also potentially structural defects due to mismatches at the graft interface can be fixed by mutating some framework residues to llama, or by mutating some residues on the CDRs’ backside to human or to a de novo designed sequence. CDR stabilizing or overall fold stabilizing sequences were then back- mutated to the corresponding llama sequence to maintain the biophysical properties and target binding affinity. This typically generates about 20 humanized variants, each of the CD16- VHH1 variants was expressed as SASA fusion (anti-BSA VHH, Genscript) in 293 cells and the binding affinity validated, if binding is and stability is maintained with several variants than the most humanized is selected. Example 3: Evaluation of NK binding moieties in exemplary bispecific constructs targeting CD16a and FCRH5 In vitro cytotoxicity assays: Bispecific molecules were evaluated for the ability to engage NK cells and kill target cells expressing FCRH5, the procedure used to assess in vitro cell cytotoxicity is a calcein release assay. This assay is specifically Atty. Docket No.45817-0158WO1 used to assess the potency of NK cell cytotoxicity against FcRH5 expressing target cells. Purified frozen human NK cells (IQ Biosciences) were thawed and rested in complete RPMI-1640 medium overnight. Target cells were stained with 100 µM Calcein in serum free medium for 30 minutes at 37C.15,000 washed target cells were combined with different numbers of NK cells to achieve desired effector (NK) to target (E:T) ratios and plated in each well of a 96 well round-bottom plate. Test articles prepared in serial dilutions were added to the co-culture and target cell lysis was measured after a 4h incubation at 37⁰C by fluorescence reading at 485/528 nM in the supernatants. Target cells alone wells were used as spontaneous calcein release control, and maximum killing was recorded from Triton X (1% final concentration) treated wells. Percent of specific killing was calculated as (F[sample]- F[spontaneous])/(F[maximum]-F[spontaneous])x100. Results: Fc Engager constructs: Various bispecific formats including varied orientations of the anti-CD16 and anti-FCRH5 VHH antibody domains were evaluated as Fc-bispecific engager molecules. These Fc-bispecific engagers are designed bind and activate NK immune cells expressing CD16a and bind target tumor cells through Vhh antibody binding to the tumor associated antigen (TAA). These engager molecules serve two roles, cross linking driven by anti-CD16 binding activates NK cells to become cytotoxic, while also specifically targeting these activated NK cells to tumor cells. The initial bispecific design contained an Fc framework in which single domain antibodies were linked N- and C-terminally to the Fc domain, sdAb-(IgG-Fc)- sdAb. The immune engagers form homodimers utilizing an Fc scaffold to create a molecule that could bivalently binds to both the targeted tumor associated antigen, which may be a B cell antigen associated with a disease (i.e., cancer) such as FcRH5, and anti-CD16a on NK cells (Fig.5). The anti-CD16a and anti-VHH have two potential orientations for the anti-TAA and anti-CD16a Vhh domains (i) an anti-CD16 VHH at the N-terminal sdAb position and an anti-FcRH5 VHH at the C-terminal Atty. Docket No.45817-0158WO1 sdAb position or (ii) an anti-FcRH5 VHH at the N-terminal sdAb position and an anti- CD16 VHH at the C-terminal sdAb position (Fig.5). The potency of the Fc engager is dependent on multiple factors including retention of both the TAA and CD16a target affinity in these bidentate constructs, appropriate orientation of the bispecific molecule bound to the respective cellular surfaces, in vivo expression levels and expression homogeneity of the final molecule. Fc Engager Construct design: The antigen binding domains incorporated were humanized llama VHH anti-CD16 antibodies, as use of VHH formats offers a particular flexibility for therapeutic protein design allowing evaluation of multiple molecular formats to optimize molecule efficacy from the multitude of tested designs (Brinkmann and Kontermann, MAbs.2017 Feb/Mar;9(2):182-212. doi: 10.1080/19420862.2016.1268307). Evaluation and characterization of a-CD16a/a-FCRH5 Fc Engager molecules: From an initial number of VHH antibodies that showed binding affinity to FCRH5 protein, five were deemed of interest to be built into a series of the various Fc bispecific engager molecules. The five VHH antibodies were incorporated into Fc engager bispecific engager molecules (Fig.5) with N-terminus fused to an anti-CD16 VHH antibodies and C-termini fused to an anti-FCRH5 VHH antibody. Each Fc engager bispecific antibodies was transiently expressed using 293 cells and showed homogenous expression of the desired molecules by SDS PAGE (Fig.6). The purified bispecific engager molecules showed varied amounts of aggregated protein following protein A purification using a low pH elution Figs.7A and 7B to elute the protein from the column, as described in the methods. Each of the molecules was therefore further purified to near homogeneity using preparative gel filtration, before for evaluation of binding affinities to the various forms of FCRH5. The binding affinities for each of the bispecific NK-engager was initially determined using the purified proteins in SPR binding assays. The binding affinity for each bivalent Fc engager construct was determined to huFCRH5, huFCRH5-IgG (domain 8 of FCRH5-Fc) and the cross reactive cynoFCRH5-IgG, results are Atty. Docket No.45817-0158WO1 tabulated in Table 18. In addition, the FACS binding affinity for each of these constructs was also evaluated for binding to both the full extracellular domains of huFCRH5 using RPMI cells which have high expression of FCRH5 (Fig.9). We also constructed two bispecific engagers using FcRH5-VHH15 and FcRH5-VHH14 using a G1-Fc and containing FEA mutation to reduce the Fc interaction with Fcg receptors (McCarthy, 2015). These molecules had been purified and characterized in a similar manner to the G4-(PAA) Fc bispecific constructs. The affinity of the molecules containing the G1-FEA Fc showed virtually identical binding in FACS, Fig.9. The Fc bispecific engager labeled BMK3 contained an irrelevant C-terminal VHH in the bispecific G4-(PAA) engager format and was included in the FACS assay as negative control for nonspecific binding of these constructs to the cell line Fig.9. The anti-FCRh5 bispecific engager molecules in Table 18 were also evaluated for cross binding to FCRH5 homologs. Fig.10 shows the FACS binding of each the NK bispecific engagers to 293 cells that transiently express FCRH3, only the bispecific engagers with anti-TAA domains FcRH5-VHH13 and FcRH5-VHH15 showed any significant FACS binding to FCRH3. In each of these assays, BMK3 contains a VHH antibody derived to a different target; this molecule serves as the negative control for background FACS binding. Since FCRH3 is expressed on NK cells it would be undesirable to bind this target as it might drive autolysis of NK cells in vivo, thus both FcRH5-VHH13 and FcRH5-VHH15 may cause NK autolysis. Li et al. (2017) showed the bispecific antibody binding to immunoglobulin domain 8 of FCRH5 was important for efficient anti-FCRH5/CD3 synapse formation and lead to efficient cytotoxicity of multiple myeloma cells. Under the premise that the NK anti-CD16 synapse would be similarly distance sensitive the Vhhs binding affinity to domain 8 was evaluated using Biacore-SPR binding with only FcRH5- VHH14 showing poor affinity, interestingly FcRH5-VHH14 retained a much tighter affinity for the intact FCRH5. It is not possible to determine if the lower affinity binding of FcRH5-VHH14 is due to misfolding of the domain 8 FCRH5 alone or if the epitope is partially missing in domain 8 FCRH5 construct, therefore this molecule was not eliminated at this point. Atty. Docket No.45817-0158WO1 To facilitate therapeutic development affinity to cyno FCRH5 was considered necessary to support preclinical toxicity evaluation. Each anti-FCRH5 VHH was evaluated in SPR binding to cyno FCRH5. FcRH5-VHH14 showed poor affinity to cyno FCRH5 with >100 nM Kd as determined by SPR. Therefore, FcRH5-VHH14 was considered to have a low rank as development candidate. We then evaluated each bispecific engager construct for potency in cytotoxicity assays. The cytotoxicity assays were done with two cell lines SUDHL6 and RPMI8226-hFCRH5. The SUDHL6 has a very low level of FCRH5 expressed on the cell surface as evaluated by staining shift shown in Fig.11 with a known positive control f(ab) which binds FCRH5. The RPMI8226-hFCRH5 has much greater expression. Table 18 – Measurement of affinity (nM) of a CD16- IgG4(PAA)-aFCRH5 to various forms of FCRH5 VHH FACS binding FACS binding SPR SPR SPR affinity FCRH5 to full length to full D8 affinity affinity to to cyno-D8 huFCRH5 huFCRH5 intact hu-D8 FCRH5 transfected into transfected into FCRH5 FCRH5 Expi-293 cells Expi-293 cells FcRH5- 62 5.3 0.5 0.5 20 VHH13 FcRH5- 74 17 1.5 1.3 34 VHH12 FcRH5- 17 43 4.6 4.6 11 VHH11 FcRH5- 50500 140 14.1 400 1900 VHH14 FcRH5- 18 34 0.5 0.5 30 VHH15 The initial ranking criteria for these bispecific engager molecules included assessing protein binding of anti-FCRH5 dimeric binding affinities of <10 nM to intact huFCRH5 in SPR binding assays, 2) positive binding to D8 of huFCRH5, 3) binding to cyno FCRH5, 4) and a lack of cross-reactive binding to FCRH3 a homologous FCRH5 family member. The binding results are summarized in Table 18. Assessing the binding data in total, affinity to FCRH5 and FCRH3, the binding Atty. Docket No.45817-0158WO1 results indicate that the lead molecules based on these criteria are FcRH5-VHH11, FcRH5-VHH15, and FcRH5-VHH12. Next each of the bispecific engager molecules was then evaluated for cytotoxic potency in assays using the RPMI cells as the target cell, these cells express high levels of FCRH5. The relative cytotoxicity of each of each molecule is shown in Fig.11, based on relative EC50 of cytotoxicity and percent relative specific killing for these molecules we ranked the molecules for potency as follows: FcRH5- VHH11>FcRH5-VHH15>FcRH5-VHH12> FcRH5-VHH14 and FcRH5-VHH13. Comparison of two Fc domains in the bispecific engager format for in vivo and in vitro expression levels and pharmacokinetics. NK cells bound to bispecific engagers that retain immunologically functional Fc domain have the potential to drive NK cell autolysis, we therefore compared the expression and pharmacokinetics of bispecific engagers with different Fc domains. The bispecific NK engager molecules were designed with an Fc that has little or no Fc receptor binding activity. Two Fc sequences IgG1FEA (L234F, L235E and D265A) and IgG4 PAA (S228P/Phe234Ala/Leu235Ala) were evaluated which are known to have suppressed CD16a binding affinity. Both Fc sequences include mutations that seek to abolish FcgRIII binding, while retaining Fc/FcRn interactions (G1 FEA (Engelberts, et al., EBioMedicine.2020;52:102625. doi: 10.1016/j.ebiom.2019.102625; and G4 PAA, Saunders 2019 Front. Immunol., 07 June 2019 Sec. Comparative Immunology, Volume 10 - 2019 | doi.org/10.3389/fimmu.2019.01296) to retain molecular half-life. To evaluate these two options, we first determined if there was a difference in expression levels of two different bispecific engager designs incorporating two different Fc sequences IgG4(PAA) and IgG1(FEA) using two VHH molecules, CD16-VHH4 (SEQ ID NO:7) and BCMA-VHH81 (SEQ ID NO: 547). Therefore, two different formats for the bispecific engagers described in (Fig.5) were evaluated. Four bispecific engagers were expressed containing an Fc with either IgG4(PAA) or IgG1(FEA), with the anti-TAA/anti-CD16 VHH domains ordered in both N and C Atty. Docket No.45817-0158WO1 orientations (Fig.5). The four bispecific engagers were initially evaluated for in vitro expression level. Table 19 compares the peak expression levels achieved at 24 h for each molecule when HEK293 cells are transfected with mRNA and found in vivo with mice dosed IV. As shown in Table 19 the expression of bispecific molecules containing IgG4(PAA) Fc domain was superior to molecules containing the IgG1(FEA) Fc domain. Independent of the Fc domain used, expression of bispecific engagers having CD16-VHH4 fused to the C-termini was superior to having CD16- VHH4 on the N-terminus of the Fc, indicating that the preferred position for CD16- VHH4 is the C-terminus for superior expression. Table 19 – In vitro expression of bispecific engager molecules when mRNA is transfected into Expi293 cells. Pharmacokinetic parameters of in vivo expressed bispecific engagers in mice, following 0.25 mg/Kg dosing of formulated mRNA into mice _{In vitro In vivo} _Group ^{Avg conc.} AUC Cmax (_ug/mL) (0-168 h) _(ng/mL) ^{Tmax (h)} [CD16-VHH4]-IgG4 PAA- ^{10762 180.3} [_{BCMA-VHH81]-His} ^3.22 ₂₄ [BCMA-VHH81]-IgG4 PAA- ^{145535 2522} [_{CD16-VHH4]-His} ^8.97 ₂₄ [CD16-VHH4]-IgG1 FEA- ^{7122 91.78} [_BCMA-VHH81] ^1.30 ₄₈ [BCMA-VHH81]-IgG1 FEA- ⁷⁷³⁰ [_CD16-VHH4] ^{3.29 0 1037} ₄₈ We then evaluated in vivo expression levels to determine if the superior expression parameters observed in the in vitro transfection experiments were maintained in vivo. The pharmacokinetics of each of the bispecific engagers in mice were evaluated for each of the mRNA expressed bispecific molecules G4(PAA) and G1(FEA) with the appropriate N and C-terminal VHH BCMA-VHH81and CD16- VHH4 (Table 19). Each molecule was formulated with a lipid nanoparticle (LNP) Atty. Docket No.45817-0158WO1 comprising an ionizable amino lipid(shown as Formula I above), DSPC, cholesterol, and a PEG-lipid (shown as Compound I above), and then dosed intravenously (IV) into mice. The expression levels of the bispecific engagers align with that which was seen with the in vitro HEK293 expression, higher expression with the IgG4(PAA) Fc molecule and higher expression with the N-terminal BCMA-VHH81construct Fig. 13A. Thus, a clear selection preference for the use of IgG4(PAA) over IgG1(FEA) was evident in this case. Additionally, the orientation of having CD16-VHH4 on the C-termini was also clearly preferred. Interestingly, the pharmacokinetic half-lives of the molecules are unaffected by the choice of the two Fc functionalities IgG4(PAA) versus IgG1(FEA) or the N- or C- orientation of the CD16-VHH4 or BCMA-VHH81 antibodies as shown in Fig.13B results expression levels in vivo versus time are virtually identical, thus main difference is the higher expression when CD16-VHH4 is in the C-terminus. Thus, a bispecific engager design in which CD16-VHH4 was oriented on the C-terminus of the bispecific engager Fc domain was considered favorable and the activity of the molecules with the CD16-VHH4 on the N-termini and the anti-B cell moiety on the C-terminus were further evaluated. Humanization Expression and characterization the bispecific engagers with CD16-VHH4 in the C-terminal position. All of our anti-FCRH5 antibodies were humanized and each of these molecules was expressed in vitro using 293 cells and purified as described in the methods. Each of the prospective anti-FCHR5 molecules was humanized, resulting in the design of at least 5 different humanized molecule designs for each anti-FCRH5 Vhh antibody. For each of these humanized molecules a highly human construct was transiently expressed in 293 cells Table 20 below. Each of these molecules was purified using protein A, Table 20 shows the characteristics for selected humanized molecules and the transient expression yields for each of these molecules. The analytical SEC for each of the purified bispecific engagers is shown in Figs.14A and 14B. Atty. Docket No.45817-0158WO1 Table 20 – Characteristics of humanized bispecific NK engager molecules Molecule Native Sequence PI Yield [FCRH5 VHH] (Daltons) length (mg) BE-11 [VHH27] 103870.48 473 8.64 0.65 BE-16 [VHH22] 105570.12 482 8.65 2.28 BE-26 [VHH32] 105524.06 481 8.54 2.07 BE-21 [VHH16] 105784.28 482 8.53 1.29 BE-31 [VHH17] 103980.50 476 8.24 0.9 The affinity for each binding domain of the bispecific molecules were then determined using Biacore and FACS binding. It had been previously determined that the NK Fc-bispecific molecules with an N-terminal CD16-VHH4 showed aggregation when purified by protein A. It was suspected that the constructs were being disrupted by the low pH elution of the molecules from protein A. Therefore, the Fc engager BE-9 was also expressed with a C-terminal His Tag allowing for the use of an IMAC matrix (Immobilized Metal Affinity Column) for purification and avoiding the harsh low pH 3.0 or less elution from protein A. A comparison of the analytical SEC showed the IMAC purified protein BE-9 (with a HIS tag), showed no aggregate in the total protein captured from the supernatant and had a virtually equivalent elution time (Fig.15A). The 6x-his tag (SEQ ID NO: 731) shows a slight interaction with silica SEC column backbone leading to some slight tailing of the peak, the molecule is of homogenous molecular weight as shown by GFLS (gel-filtration light scattering) indicting the tailing is not due to degradation of the molecule (Fig.15B). These results upon initial expression the molecule is well folded and does not show detectable aggregates. Based on binding affinity further efforts were focused on the top three candidate molecules containing FcRH5-VHH11, FcRH5-VHH15, and FcRH5- VHH12. Each of these molecules in which the anti-TAA was in the C-terminal position had been previously evaluated using a cytotoxicity assay (Fig.16). The most potent molecules were identified as containing one of these three engagers, however we eliminated engagers containing FcRH5-VHH12 as not a suitable candidate as previously mentioned due to the strong risk NK cell killing. For the final selection, Atty. Docket No.45817-0158WO1 we then undertook a direct comparison of FcRH5-VHH11 and FcRH5-VHH15 in the N-terminal engager format design and demonstrated that the N-terminal engager format had superior binding affinity (Fig.17). A comparison of the humanized FcRH5-VHH11 (SEQ ID NO: 392) and FcRH5-VHH15(SEQ ID NO: 393) in killing assays showed that the BE-40 (the construct comprising SEQ ID NO:392 + IgG4PAA hinge, CH2, CH3 domains + G₄S (SEQ ID NO:448)+ SEQ ID NO:7) had the best cytotoxicity based in EC50 value. These results lead us to select this molecule as a therapeutic monovalent candidate molecule. GAAAUA 7) CUGCUGC AGUUGCA CGGCUCA ACCUACA

U U U UUU CCCCAG GCAAGGAACGGGAGUUCGUGGCCGGCAUCAGCCGGA GCGGCGGCAUGACCGGCUACGCCGAGAGCGUGAAGG GUCGGUUCACCAUUAGCCGGGAUAACAGCAAGAAUA CCGUCUACCUUCAGAUGAACAGCCUUAGAGCCGAGG ACACCGCUGUUUACUACUGUGCCGCCUACGUGGGCG GCUUCAGCACCGCCCGGCGGGACUACAGCUACUGGGG ACAGGGGACCCAAGUGACAGUGAGCAGCGAGAGCAA GUACGGACCUCCUUGCCCUCCCUGUCCAGCCCCAGAG GCCGCUGGAGGCCCAAGCGUGUUCCUGUUCCCACCCA AGCCCAAGGACACCCUGAUGAUCAGCCGGACCCCAGA GGUGACCUGCGUGGUGGUGGACGUGAGCCAGGAGGA CCCCGAGGUGCAGUUCAACUGGUACGUGGACGGCGU GGAGGUGCACAACGCCAAGACCAAGCCCCGGGAGGA GCAGUUCAACAGCACCUACCGGGUGGUGAGCGUGCU GACCGUGCUGCACCAGGACUGGCUGAACGGCAAGGA GUACAAGUGCAAGGUGAGCAACAAGGGCCUGCCCAG Atty. Docket No.45817-0158WO1 CAGCAUCGAGAAGACCAUCAGCAAGGCAAAGGGCCA ACCUCGGGAGCCACAGGUGUACACCCUGCCUCCCAGC CAGGAGGAGAUGACCAAGAACCAGGUGAGCCUGACC UGCCUGGUGAAGGGCUUCUACCCCAGCGACAUCGCCG UGGAGUGGGAGAGCAACGGCCAGCCCGAGAACAACU ACAAGACCACUCCACCAGUGCUGGACAGCGACGGCAG CUUCUUCCUGUACAGCCGGCUGACCGUGGACAAGAG CCGGUGGCAGGAGGGCAACGUGUUCAGCUGCAGCGU GAUGCACGAGGCCCUGCACAACCACUACACCCAGAAG AGCCUGAGCCUCAGCCUGGGCAAGGGUGGAGGCGGG AGCGAGGUGCAGCUGGUGGAAAGCGGCGGCGGCCUG GUGCAACCCGGCGGCAGCCUGCGACUGUCCUGUGCCG CCUCUGGCCGGACCGACAGCAUCUACGCCAUGGGCUG GUUCCGGCAGGCACCCGGCAAGGAGCGGGAGUUCGU GAGCGCCAUCAACAGCAACACCGGCCGGACCUACCAC GCCGACAGCGUGAAGGGCCGGUUCACCAUCAGCCGG GACAACGCCAAGAACAUGGUGUACCUGCAGAUGAAC AGCCUGCGGGCCGAGGAUACCGCCGUGUACUAUUGC GCCGCCGGACGGGGUUACGGCCUGCUGAGCAUCAGC AGCAACUGGUACAACUACUGGGGCCAGGGCACCCUG GUGACCGUGAGCAGC (SEQ ID NO: 463) 3’ UTR UGAUAAUAGGCUGGAGCCUCGGUGGCCUAGCUUCUU GCCCCUUGGGCCUCCCCCCAGCCCCUCCUCCCCUUCC UGCACCCGUACCCCCGUGGUCUUUGAAUAAAGUCUG AGUGGGCGGC (SEQ ID NO: 468) Corresponding amino acid sequence METPAQLLFLLLLWLPDTTGQVQLQESGGGLVQPGGSLR LSCAASGRTYNNYAMGWFRQAPGKEREFVAGISRSGGM TGYAESVKGRFTISRDNSKNTVYLQMNSLRAEDTAVYYC AAYVGGFSTARRDYSYWGQGTQVTVSSESKYGPPCPPC PAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVS QEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVV SVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISKAK GQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDI AVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDK SRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGKGGG GSEVQLVESGGGLVQPGGSLRLSCAASGRTDSIYAMGWF RQAPGKEREFVSAINSNTGRTYHADSVKGRFTISRDNAK NMVYLQMNSLRAEDTAVYYCAAGRGYGLLSISSNWYN YWGQGTLVTVSS (SEQ ID NO: 464) PolyA tail 100 nt (SEQ ID NO: 730) Example 4: In vivo animal studies Bioanalysis of pharmacokinetic samples from cynomolgus monkeys Atty. Docket No.45817-0158WO1 To determine the pharmacokinetic parameters for the mRNA-derived NKE protein, BE-40 (FCR460h1-IgG4 PAA-12C11h3) protein concentrations in serum were quantified using a custom MSD assay. Briefly, 96-well Multi-Array Sector plates (Meso Scale Diagnostics [Rockville, MD], Catalog No. L15XA) were coated with capture antigen at 2 μg/mL (Recombinant Human FCRL5/FcRH5 Protein; R&D Systems [Minneapolis, MN], Catalog No.2078-FC-050) in PBS overnight at 4^oC. Cynomolgus monkey serum samples were diluted 1:40 with Diluent 100 (Meso Scale Diagnostics, Catalog No. R50AA-3) and incubated in the coated plates for 1 hour at ambient room temperature with shaking at 700-900 RPM.0.5 μg/mL Detection Antigen solution (Biotinylated Human Fc gamma RIIIA/CD16a (F176) Protein [Acro

in the peripheral blood were profiled by flow cytometry using markers CD45, CD3, CD19, CD20, CD16, CD8, CD14, CD11b, CD11c, and CD86. Subsets of B cells were further gated by naïve/memory markers CD21 and CD27. For pre-dose blood samples, FcRH5 levels in B cell subsets were assessed using a biotinylated recombinant protein FCR460h1-IgG4 PAA and PE-streptavidin (panel 1, Table 22). Frequency and median fluorescence intensity (MFI) data from flow panel 2 (Table 23) were collected for pharmacodynamic evaluation. Table 22: Flow Cytometry Staining Reagents for Panel 1 Parameter Antibody Fluorophore Clone Isotype Vendor Catalog No. Dilution Atty. Docket No.45817-0158WO1 AF700-A CD11c AF700 3.9 IgG1 k BioLegend 301648 50 APC-Cy7-A CD11b APC-Cy7 ICRF44 IgG1 k BioLegend 301342 50 BV421-A CD27 BV421 O323 IgG1 k BioLegend 302824 50 BV605-A CD86 BV605 2331 IgG1 k BD 562999 50 (FUN- Biosciences 1) PE- CD3 PE- SP34-2 IgG1 λ BD 562406 50 Dazzle594 - Dazzle594 Biosciences A PE-A Streptavidin PE N/A N/A BioLegend 405204 400 PerCP CD20 PerCP 2H7 IgG2b BioLegend 302326 50 Cy5.5-A Cy5.5 BV785-A CD45 BV785 D058- IgG1 k BD 563861 50 1283 Biosciences FITC-A CD21 AF488 Bly4 IgG1 k BD 561372 50 BIosciences BV711-A CD8 BV711 SK1 IgG1 k BioLegend 344734 50 BV650-A CD14 BV650 M5E2 IgG2a BioLegend 301836 50 k PE-Cy7-A CD19 PE-Cy7 J3-119 IgG1 Beckman IM3628U 50 Coulter BV510-A Fixable BV510 N/A N/A ThermoFisher 65-0866-14 1000 Viability Dye eFluor506 Table 23: Flow Cytometry Staining Reagents for Panel 2 (Pre and Post-dose Samples) Parameter Antibody Fluorophore Clone Isotype Vendor Catalog No. Dilution AF700-A CD11c AF700 3.9 IgG1 k BioLegend 301648 50 APC-A Camelid iFluor647 N/A IgG Genscript A01994 100 VHH APC-Cy7- CD11b APC-Cy7 ICRF44 IgG1 k BioLegend 301342 50 A BV421-A CD27 BV421 O323 IgG1 k BioLegend 302824 50 BV605-A CD86 BV605 2331 IgG1 k BD 562999 50 (FUN- Biosciences 1) PE- CD3 PE- SP34-2 IgG1 L BD 562406 50 Dazzle5 -A PE-A 50

Atty. Docket No.45817-0158WO1 PerCP CD20 PerCP Cy5.5 2H7 IgG2b BioLegend 302326 50 Cy5.5-A BV785-A CD45 BV785 D058- IgG1 k BD 563861 50 1283 Biosciences FITC-A CD21 AF488 Bly4 IgG1 k BD 561372 50 BIosciences BV711-A CD8 BV711 SK1 IgG1 k BioLegend 344734 50 BV650-A CD14 BV650 M5E2 IgG2a BioLegend 301836 50 k PE-Cy7-A CD19 PE-Cy7 J3-119 IgG1 Beckman IM3628U 50 Coulter BV510-A Fixable BV510 N/A N/A ThermoFisher 65-0866-14 1000 Viability Dye eFluor506 Single dose pharmacokinetics of the mRNA-derived NKE protein in cynomolgus monkeys An LNP-encapsulated mRNA that encodes BE-40 (FCR460h1-IgG4 PAA- 12C11h3) engager protein was manufactured in a fed-batch in vitro transcription process using a T7 polymerase variant and a co-transcriptional capping analog. Purified mRNA was formulated in an LNP that consisted of SM-86, cholesterol, DSPC, and OL- 56. Nine female cynomolgus monkeys were randomized and assigned to study groups according to Table 24. The LNP-encapsulated mRNA was administered through a 1-hr intravenous infusion using an infusion pump connected to a syringe with an infusion line and a temporary indwelling catheter. Serum samples were collected for protein PK analysis, and single cell suspensions from RBC lysed whole blood were prepared for flow cytometry-based PD analysis. Following a single IV infusion, serum concentrations of FCR460h1-IgG4 PAA-12C11h3 protein were quantifiable up to 169-217 hours post start of infusion (SOI), and the TEmax of the NKE protein was 13 or 25 hours post SOI (FIG.20, Table 25). Mean effective t_1/2 values were 16.4, 21.6, and 29.5 hours at 0.5, 1.5, and 3 mg/kg/dose, respectively. The systemic exposure to the NKE protein, as defined by mean Emax and AUEC0-673hr, increased with increasing dose level across the dose range Atty. Docket No.45817-0158WO1 following a single IV infusion of the LNP-encapsulated mRNA but displayed non- linear pharmacokinetics. Table 24: Single dose NHP study design Group Dose Dose Dose Route No. of No. Level Volume Concentration Females (mg/kg) (mL/kg) (mg/mL) 1 0.5 5 0.1 IV 3 infusion 2 1.5 5 0.3 IV 3 infusion

Atty. Docket No.45817-0158WO1 CD21 CD27 i b d CD21 CD27 b h d f

exposure to a proteasome inhibitor, an immunomodulatory drug (IMiD), and an anti- cluster of differentiation (CD38) monoclonal antibody. Subjects will have received at least 3 prior lines of therapy or be triple-class refractory. Subjects that are intolerant of a proteasome inhibitor, IMiD, or anti-CD38 mAb also will be eligible. Subjects may have measurable disease, defined as at least one of the following: (i) serum M-protein ≥ 0.5 g/dL; urine M-protein ≥ 200 mg/24 hours; (iii) involved free light chain (FLC) ≥ 100 mg/L and an abnormal FLC ratio; (iv) Atty. Docket No.45817-0158WO1 plasmacytoma with a single diameter ≥ 2 cm; and/or (v) bone marrow plasma cells > 30%. Females of childbearing potential must not be pregnant or breastfeeding, must be using a contraceptive that is highly effective, and must have a negative highly sensitive pregnancy test (urine or blood as required by local regulations) within 14 days before the first dose of the study treatment. Key exclusion criteria may include any clinically relevant concurrent medical disease or condition that would be judged to compromise subject safety or interfere with the evaluation of the safety of the study treatment; cardiopulmonary disease requiring supplemental oxygen to maintain adequate oxygenation; a history of confirmed progressive multifocal leukoencephalopathy; antibody-based immunotherapy (monoclonal antibody, bispecific antibody, antibody drug conjugate, radioimmunoconjugate) within 21 days prior to cycle 1, day 1 of treatment; immunomodulatory agent therapy within 7 days of cycle 1, day 1 of treatment; corticosteroid therapy, ≥ 140 mg of prednisone or equivalent cumulative dose, within 14 days prior to cycle 1, day 1; planning to receive a live attenuated vaccine during the study or having received a live vaccine within 30 days before the first dose of the study drug. Interventions: The LNP-encapsulated mRNA will be administered by intravenous infusion over 1 hour. A starting dose of 0.3 mg/kg mRNA may be used. Dosing may be on Days 1, 8, and 15 of a 28-day cycle for up to 12 cycles; alternative dosing schedules may be explored. Escalating dose levels will be explored. Results: Participants will be monitored for safety and efficacy. Blood and bone marrow samples may be taken to assess pharmacokinetics and pharmacodynamics. Participants may be followed up for safety for 90 days after their last dose of the study mRNA or after the start of next anti-cancer therapy, whichever is earlier. Participants may be followed for up to 2 years or longer from their first study dose to assess progression-free survival and overall survival. Maximum tolerated dose and/or a recommended dose for Phase 2 studies may be determined. The pharmacokinetics (e.g., Cmax, AUC), pharmacodynamics (e.g., Emax, AUEC) Atty. Docket No.45817-0158WO1 and preliminary efficacy (e.g., Overall Response Rate, Clinical Benefit Rate, Duration of Response, Progression-Free Survival, Overall Survival) of the mRNA treatment

fixed, and analyzed using flow cytometry. Target cells were identified as GFP+ or CFSE+ cells, and percent killing calculated for each well as

between the target cell count for the treated well and the average target cell count of untreated wells, divided by the average target cell count of the untreated wells. Co-culture experiments demonstrated an increase in NK cell-mediated cytotoxicity in the presence of NKE (BE-40) and HSA-sIL-15 relative to NKE alone as demonstrated by increased cell killing by the NKE in the presence of increasing amounts of HSA-sIL-15. This correlated with a reduction in the EC₅₀ of cytotoxicity, demonstrated with both Molp-2 (FIG.25) and Molm13-LG-FCRH5 (FIG.26) target cells. Atty. Docket No. 45817-0158WO1

Table 23: Flow Cytometry Staining Reagents for Panel 2 (Pre and Post-dose Samples)

[0716] Single dose pharmacokinetics of the mRNA-derived NKE protein in cynomolgus monkeys

|0717| An LNP-encapsulated mRNA that encodes BE-40 (FCR460hl-IgG4 PAA- 12Cl lh3) engager protein was manufactured in a fed-batch in vitro transcription process using a T7 polymerase variant and a co-transcriptional capping analog. Purified mRNA was formulated in an LNP that consisted of SM-86, cholesterol, DSPC, and OL- 56. Nine female cynomolgus monkeys were randomized and assigned to study groups

292

SUBSTITUTE SHEET (RULE 26) according to Table 24. The LNP-encapsulated mRNA was administered through a 1-hr intravenous infusion using an infusion pump connected to a syringe with an infusion line and a temporary indwelling catheter. Serum samples were collected for protein PK analysis, and single cell suspensions from RBC lysed whole blood were prepared for flow cytometry-based PD analysis.

[0718] Following a single IV infusion, serum concentrations of FCR460hl-IgG4 PAA-12Cllh3 protein were quantifiable up to 169-217 hours post start of infusion (SOI), and the TEmax of the NKE protein was 13 or 25 hours post SOI (FIG. 20, Table 25). Mean effective ti/2 values were 16.4, 21.6, and 29.5 hours at 0.5, 1.5, and 3 mg/kg/dose, respectively. The systemic exposure to the NKE protein, as defined by mean Emax and AUECo-673hr, increased with increasing dose level across the dose range following a single IV infusion of the LNP-encapsulated mRNA but displayed nonlinear pharmacokinetics.

Table 24: Single dose NHP study design

293

SUBSTITUTE SHEET (RULE 26) Atty. DocketNo. 45817-0158WO1

Table 25: Individual Serum Protein Pharmacokinetics Parameters in Female Monkeys Following a Single IV Infusion of LNP- encapsulated mRNA (Note that mRNA-2736 encodes BE-40)

[0719] Pharmacodynamics of single dose anti-FcRH5 NKE mRNA in cynomolgus monkeys

[0720] FcRH5 is exclusively expressed in the B cell lineage in the peripheral blood. The expression pattern among B cell subsets was further defined using the naive and memory B cell markers CD21 and CD27. In the 9 animals enrolled in the study, the CD21+CD27- naive subset and CD21-CD27+ memory subset together accounted for the vast majority (>90%) of total CD20+ B cells (FIG. 21). The percentage of FCRH5+ cells ranged from 56% - 93% in CD21-CD27+ memory B cells, 10 - 50% in CD21-CD27- B cells, and 5 - 14% in CD21+CD27- naive B cells (FIG. 22, upper panel). The MFI of FCRH5 was highest on CD21-CD27+ memory B cells, followed by CD21- CD27- subset (Fig. 22, lower panel). Following a single dose IV infusion of the LNP-encapsulated mRNA discussed above, a 56-93% reduction of CD21-CD27+ memory B cells was observed at all dose levels (0.5 mg/kg, 1.5 mg/kg, and 3 mg/kg; FIGs. 23 and 24). Statistical analysis of the reduction of CD21-CD27+ memory B cells did not reach significance due to the small sample size and large variability of B cell subsets between animals. Consistent with the lack of FCRH5 expression, there was no consistent change in frequency in the CD21+ CD27- naive B cell subset. Example 5: Phase I safety and tolerability study of mRNA encoding a bispecific engager targeting CD16a and FcRH5 in subjects with relapsed or refractory multiple myeloma.

|07211 An open-label, Phase I, dose escalation, first-in-human clinical study will be performed in subjects with relapsed or refractory multiple myeloma (RRMM) to evaluate safety and tolerability of escalating doses of LNP-encapsulated mRNA encoding a bispecific engager as disclosed herein targeting CD16a and FcRH5 (the BE-40 candidate described in Example 3 above). The LNP-encapsulated mRNA will be administered intravenously.

[0722] Subjects: Subjects will be 18 years old or older, and have RRMM and prior exposure to a proteasome inhibitor, an immunomodulatory drug (IMiD), and an anticluster of differentiation (CD38) monoclonal antibody. Subjects will have received at least 3 prior lines of therapy or be triple-class refractory. Subjects that are intolerant of a proteasome inhibitor, IMiD, or anti-CD38 mAb also will be eligible.

295

SUBSTITUTE SHEET (RULE 26) [0723] Subjects may have measurable disease, defined as at least one of the following: (i) serum M-protein > 0.5 g/dL; urine M-protein > 200 mg/24 hours; (iii) involved free light chain (FLC) > 100 mg/L and an abnormal FLC ratio; (iv) plasmacytoma with a single diameter > 2 cm; and/or (v) bone marrow plasma cells > 30%. Females of childbearing potential must not be pregnant or breastfeeding, must be using a contraceptive that is highly effective, and must have a negative highly sensitive pregnancy test (urine or blood as required by local regulations) within 14 days before the first dose of the study treatment.

[0724] Key exclusion criteria may include any clinically relevant concurrent medical disease or condition that would be judged to compromise subject safety or interfere with the evaluation of the safety of the study treatment; cardiopulmonary disease requiring supplemental oxygen to maintain adequate oxygenation; a history of confirmed progressive multifocal leukoencephalopathy; antibody -based immunotherapy (monoclonal antibody, bispecific antibody, antibody drug conjugate, radioimmunoconjugate) within 21 days prior to cycle 1, day 1 of treatment; immunomodulatory agent therapy within 7 days of cycle 1, day 1 of treatment; corticosteroid therapy, > 140 mg of prednisone or equivalent cumulative dose, within 14 days prior to cycle 1, day 1; planning to receive a live attenuated vaccine during the study or having received a live vaccine within 30 days before the first dose of the study drug.

[0725] Interventions: The LNP-encapsulated mRNA will be administered by intravenous infusion over 1 hour. A starting dose of 0.3 mg/kg mRNA may be used. Dosing may be on Days 1, 8, and 15 of a 28-day cycle for up to 12 cycles; alternative dosing schedules may be explored. Escalating dose levels will be explored.

[0726] Results: Participants will be monitored for safety and efficacy. Blood and bone marrow samples may be taken to assess pharmacokinetics and pharmacodynamics. Participants may be followed up for safety for 90 days after their last dose of the study mRNA or after the start of next anti-cancer therapy, whichever is earlier. Participants may be followed for up to 2 years or longer from their first

296

SUBSTITUTE SHEET (RULE 26) study dose to assess progression-free survival and overall survival. Maximum tolerated dose and/or a recommended dose for Phase 2 studies may be determined. The pharmacokinetics (e.g., Cmax, AUG), pharmacodynamics (e.g., Emax, AUEC) and preliminary efficacy (e.g., Overall Response Rate, Clinical Benefit Rate, Duration of Response, Progression-Free Survival, Overall Survival) of the mRNA treatment may be assessed. Clinical benefit rate may be defined as the proportion of participants whose best overall response is a minimal response or better, as determined based on International Myeloma Working Group criteria, between the first dose of the study treatment and the date of progression or the date of subsequent anticancer therapy, whichever occurs first.

Example 6: HSA-sIL-15 Synergistically Improves NKE Cytotoxicity In Vitro

J07271 Cytotoxicity mediated by an anti-FCRH5 NKE (BE-40) and HSA-sIL- 15 proteins was tested in an in vitro co-culture of healthy donor PBMCs and NK cells with FCRH5-expressing target cells. Overnight-rested PBMCs and isolated NK cells were mixed at a 1 :2 ratio, stained with CellTrace Violet, and used as effector cells. GFP-expressing Molml3-LG-FCRH5 cells or carboxyfluorescein succinimidyl ester (CFSE)-stained Molp-2 cells served as target cells. Cells were seeded in a 96-well plate at a ratio of 100,000 effector cells: 1000 target cells per well. Varying concentrations of NKE and/or HSA-sIL-15 protein were added to each well followed by incubating the cocultures for 24 hours at 37°C and 5% CO2. After incubation, cells were stained with antibodies against human lymphocyte and activation markers, fixed, and analyzed using flow cytometry. Target cells were identified as GFP+ or CFSE+ cells, and percent killing calculated for each well as the difference between the target cell count for the treated well and the average target cell count of untreated wells, divided by the average target cell count of the untreated wells.

[07281 Co-culture experiments demonstrated an increase in NK cell-mediated cytotoxicity in the presence of NKE (BE-40) and HSA-sIL-15 relative to NKE alone as demonstrated by increased cell killing by the NKE in the presence of increasing amounts of HSA-sIL-15. This correlated with a reduction in the EC50 of cytotoxicity.

297

SUBSTITUTE SHEET (RULE 26) demonstrated with both Molp-2 (FIG.25) and Molml3-LG-FCRH5 (FIG. 26) target cells.

298

SUBSTITUTE SHEET (RULE 26)

Claims

Atty. Docket No.45817-0158WO1 WHAT IS CLAIMED IS: 1. A composition comprising a nucleic acid encoding a bispecific engager, the nucleic acid comprising an mRNA polynucleotide comprising in order from the 5’ to 3’ end of the mRNA (i) an open reading frame encoding a VHH binding moiety that binds to a molecule expressed on the surface of a B cell, (ii) an open reading frame encoding an IgG4 PAA CH2 and CH3 domain, and (iii) an open reading frame encoding a VHH binding moiety that binds to a molecule expressed on the surface of an NK cell. 2. The composition of claim 1, wherein the molecule expressed on the surface of a B cell is selected from CD38, BCMA, GPRC5D, and FcRH5. 3. The composition of claim 1 or 2, wherein the VHH binding moiety that binds to a molecule expressed on the surface of a B cell comprises (i) a set of complementarity-determining regions (CDRs) disclosed in Table 1; (ii) a set of CDRs disclosed in Table 3; (iii) a set of CDRs disclosed in Table 5; or (iv) a set of CDRs disclosed in Table 7, optionally wherein the VHH binding moiety specifically binds to FcRH5 and comprises the amino acid sequences of the VHH CDR1, VHH CDR2 and VHH CDR3 set forth in SEQ ID NOs.: 213, 223, and 233, respectively. 4. The composition of any one of claims 1-3, wherein the VHH binding moiety that binds to a molecule expressed on the surface of a B cell comprises (i) an amino acid sequence with at least 90%, at least 95%, or 100% identity to a heavy chain amino acid sequence disclosed in Table 2; (ii) an amino acid sequence with at least 90%, at least 95%, or 100% identity to a heavy chain amino acid sequence disclosed in Table 4; (iii) an amino acid sequence with at least 90%, at least 95%, or 100% identity to a heavy chain amino acid sequence disclosed in Table 6; or (iv) an amino acid sequence with at least 90%, at least 95%, or 100% identity to a heavy chain amino acid sequence disclosed in Table 8, optionally wherein the VHH binding moiety that binds to a molecule expressed on the surface of a B cell comprises an amino acid 291 Atty. Docket No.45817-0158WO1 sequence with at least 85%, at least 90%, at least 95%, or 100% identity to the amino acid sequence of SEQ ID NO:392. 5. The composition of any one of claims 1-4, wherein the molecule expressed on the surface of an NK cell is selected from B3GAT1 (CD57), CCR7 (CD197), CD16, CD16a, CD16b, CD2 CD226, CD244, CD27, CD3, CD300A, CD34, CD58, CD59, CD69, CSF2, CX3CR1, CXCR1 (CD128), CXCR3 (CD183), CXCR4, EOMES, GZMB, ICAM1 (CD54), IFNG, IL-15R, IL-1R, IL22, IL- 2RB (CD122), IL-7R (CD127), ITGA1 (CD49a), ITGA2 (CD49b), ITGAL (CD11a), ITGAM (CD11b), ITGB2 (CD18), KIR, KIR2DL1, KIR2DL2, KIT (CD117), KLRB1C, KLRC1, KLRC2, KLRD1 (CD94), KLRF1, KLRG1, KLRK1, LILRB1, KLRA4, KLRA8, MICA/BNCAM1 (CD56), NK2D, NKP46 (NCR1, CD335), NCR2, NCR3 (CD337), PRF1, SELL (CD62L), SIGLEC7, SLAMF6, SPN, TBX21, and TNFa; or optionally, wherein the target molecule on the surface of a NK cell is selected from: CD16a, NKP46, NK2D, and MICA/B. 6. The composition of any one of claims 1-5, wherein the molecule expressed on the surface of an NK cell is CD16a. 7. The composition of any one of claims 1-6, wherein the VHH binding moiety that binds to a molecule expressed on the surface of an NK cell comprises: (i) a VHH CDR1, a VHH CDR2, and a VHH CDR3 according to any one CDR definition set out in Table I; (ii) a VHH CDR1, a VHH CDR2, and a VHH CDR3 according to any one CDR definition set out in Table II; or (iii) a CDR1 comprising the amino acid sequence GRTDSIYA (SEQ ID NO: 2), a CDR-2 comprising the amino acid sequence INSNTGRT (SEQ ID NO: 3), and a CDR-3 comprising the amino acid sequence AAGRGYGLLSISSNWYNY(SEQ ID NO: 4). 8. The composition of any one of claims 1-7, wherein the VHH binding moiety that binds to a molecule expressed on the surface of an NK cell comprises an amino acid sequence with at least 90%, at least 95%, or 100% identity to any 292 Atty. Docket No.45817-0158WO1 one of SEQ ID NOs: 1 or 5-24, optionally wherein the VHH binding moiety that binds to a molecule expressed on the surface of an NK cell comprises an amino acid sequence with at least 85%, at least 90%, at least 95%, or 100% identity to the amino acid sequence set forth in SEQ ID NO:7. 9. The composition of any one of claims 1-8, wherein the bispecific engager comprises an amino acid sequence with at least 90%, at least 95%, or 100% identity to an amino acid sequence disclosed in Table 9, optionally wherein the bispecific engager comprises an amino acid sequence with at least 85%, at least 90%, at least 95%, or 100% identity to the amino acid sequence set forth in SEQ ID NO:433. 10. The composition of any one of claims 1-9, wherein the bispecific engager comprises the amino acid sequence comprising of SEQ ID NO: 433. 11. The composition of any one of claims 1-10, wherein the nucleic acid comprises the nucleotide sequence of SEQ ID NO: 463 along with a stop codon at the 3’end, or encodes the amino acid sequence of SEQ ID NO: 464, or encodes the amino acid sequence of amino acids 21 to 502 of SEQ ID NO: 464. 12. The composition of any one of claims 1-11, further comprising at least one additional nucleic acid encoding at least one additional bispecific engager, wherein the at least one additional bispecific engager is different from the bispecific engager of any one of claims 1-11. 13. The composition of claim 12, wherein the at least one additional nucleic acid comprises an mRNA polynucleotide comprising in order from the 5’ to 3’ end of the mRNA (i) at least one additional open reading frame encoding at least one additional VHH binding moiety that binds to a molecule expressed on the surface of a B cell, (ii) at least one additional open reading frame encoding an IgG4 PAA CH2 and CH3 domain (SEQ ID NO:548), and (iii) at least one 293 Atty. Docket No.45817-0158WO1 additional open reading frame encoding a VHH binding moiety that binds to a molecule expressed on the surface of an NK cell. 14. The composition of any one of claims 1-13, further comprising a cytokine or at least one additional nucleic acid encoding a cytokine. 15. The composition of claim 14, wherein the cytokine is IL-15, optionally wherein the cytokine is HSA-sIL-15 and comprises an amino acid sequence that is at least 85%, 90%, 95%, or 100% identical to SEQ ID NO: 729. 16. The composition of any one of claims 1-15, further comprising a delivery vehicle. 17. The composition of claim 16, wherein the delivery vehicle comprises a lipid nanoparticle. 18. The composition of claim 17, wherein the lipid nanoparticle comprises: (a) an ionizable amino lipid of Formula (I): or its N-oxide, or a salt or isomer thereof,

;

wherein R^aβ, ^{aγ aδ}

R , and R are each independently selected from the group consisting of H, C2-12 alkyl, and C2-12 alkenyl; R² and R³ are each independently selected from the group consisting of C_1-14 alkyl and C2-14 alkenyl; R⁴ is selected from the group consisting of -(CH₂)_nOH, wherein n is selected from 294 Atty. Docket No.45817-0158WO1 the group ,

wherein denotes a point of attachment;

wherein is N(R)2; each R is independently selected from the group consisting of C_1-6 alkyl, C_2-3 alkenyl, and H; wherein n2 is selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10; each R⁵ is independently selected from the group consisting of C_1-3 alkyl, C_2-3 alkenyl, and H; each R⁶ is independently selected from the group consisting of C1-3 alkyl, C2-3 alkenyl, and H; M and M’ are each independently selected from the group consisting of -C(O)O- and -OC(O)-; R’ is a C1-12 alkyl or C2-12 alkenyl; l is selected from the group consisting of 1, 2, 3, 4, and 5; and m is selected from the group consisting of 5, 6, 7, 8, 9, 10, 11, 12, and 13. 19. The composition of claim 18, wherein the ionizable amino lipid of Formula (I) comprises: R’^a is R’^branched; ; and R^aδ are each H; R² and R³ are each C_1-14 alkyl;

;

each R⁵ is H; 295 Atty. Docket No.45817-0158WO1 each R⁶ is H; M and M’ are each -C(O)O-; R’ is a C1-12 alkyl; l is 5; and m is 7. 20. The composition of claims 18, wherein the ionizable amino lipid of Formula (I) comprises: R’a is R’branched; ;

R^aβ, R^aγ, and R^aδ are each H;

R⁴ is -(CH2)nOH; n is 2; each R5 is H; each R6 is H; M and M’ are each -C(O)O-; R’ is a C1-12 alkyl; l is 5; and m is 7. 21. The composition of claims 18, wherein the ionizable amino lipid of Formula (I) comprises: R’a is R’branched; ;

296 Atty. Docket No.45817-0158WO1 R^aγ is C2-12 alkyl; R² and R³ are each C_1-14 alkyl; R⁴ is -(CH2)nOH; n is 2; each R⁵ is H; each R⁶ is H; M and M’ are each -C(O)O-; R’ is a C_1-12 alkyl; l is 5; and m is 7. 22. The composition of claim 17, wherein the lipid nanoparticle comprises an ionizable amino lipidselected from:

23. The composition of any one of claims 17-22, wherein the lipid nanoparticle further comprises: a phospholipid, a structural lipid, and a PEG-lipid. 297 Atty. Docket No.45817-0158WO1 24. The composition of any one of claims 17-23, wherein the lipid nanoparticle comprises: 40-50 mol% of an ionizable amino lipid, 30-45 mol% of a structural lipid, 5-15 mol% of a phospholipid, and 1-5 mol% of a PEG-lipid. 25. The composition of claim 24, wherein the lipid nanoparticle comprises: 45-50 mol% of the ionizable amino lipid, 35-40 mol% of the structural lipid, 8-12 mol% of the phospholipid, and 1.5-3.5 mol% of the PEG-lipid. 26. The composition of any one of claims 23-25, wherein the phospholipid is selected from: 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), 1,2-dioleoyl-sn-glycero- 3-phosphoethanolamine (DOPE), 1,2-dilinoleoyl-sn-glycero-3-phosphocholine (DLPC), 1,2-dimyristoyl-sn-gly cero-phosphocholine (DMPC), 1,2-dioleoyl- sn-glycero-3-phosphocholine (DOPC), l,2-dipalmitoyl-sn-glycero-3- phosphocholine (DPPC), 1,2-diundecanoyl-sn-glycero-phosphocholine (DUPC), 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), 1,2-di- O-octadecenyl-sn-glycero-3-phosphocholine (18:0 Diether PC), 1-oleoyl-2 cholesterylhemisuccinoyl-sn-glycero-3-phosphocholine (OChemsPC), 1- hexadecyl-sn-glycero-3-phosphocholine (C16 Lyso PC), 1,2-dilinolenoyl-sn- glycero-3-phosphocholine,1,2-diarachidonoyl-sn-glycero-3-phosphocholine, 1,2-didocosahexaenoyl-sn-glycero-3-phosphocholine, 1,2-diphytanoyl-sn- glycero-3-phosphoethanolamine (ME 16.0 PE), 1,2-distearoyl-sn-glycero-3- phosphoethanolamine, 1,2-dilinoleoyl-sn-glycero-3-phosphoethanolamine, 1,2-dilinolenoyl-sn-glycero-3-phosphoethanolamine, 1,2-diarachidonoyl-sn- glycero-3-phosphoethanolamine, 1,2-didocosahexaenoyl-sn-glycero-3- phosphoethanolamine, 1,2-dioleoyl-sn-glycero-3-phospho-rac-(1-glycerol) sodium salt (DOPG), sphingomyelin, and mixtures thereof. 298 Atty. Docket No.45817-0158WO1 27. The composition of any one of claims 23-26, where the phospholipid is DSPC. 28. The composition of any one of claims 23-27, wherein the structural lipid is selected from: cholesterol, fecosterol, sitosterol, ergosterol, campesterol, stigmasterol, brassicasterol, tomatidine, tomatine, ursolic acid, alpha-tocopherol, hopanoids, phytosterols, steroids, derivatives thereof, and mixtures thereof. 29. The composition of any one of claims 23-28, wherein the structural lipid is cholesterol or a derivative thereof. 30. The composition of any one of claims 23-29, wherein the PEG-lipid is selected from: 1,2-dimyristoyl-sn-glycerol methoxypolyethylene glycol (PEG-DMG), 1,2- distearoyl-sn-glycero-3-phosphoethanolamine-N-[amino(polyethylene glycol)] (PEG-DSPE), PEG-disteryl glycerol (PEG-DSG), PEG-dipalmetoleyl, PEG- dioleyl, PEG-distearyl, PEG-diacylglycamide (PEG-DAG), PEG-dipalmitoyl phosphatidylethanolamine (PEG-DPPE), PEG-l,2-dimyristyloxlpropyl-3- amine (PEG-c-DMA), any mixtures thereof. 31. The composition of any one of claims 23-30, wherein the PEG-lipid comprises a structure of: . 32.

comprises: 40-50 mol% of an ionizable amino lipidcomprising a structure of: ,

Atty. Docket No.45817-0158WO1 5-15 mol% of DSPC, and 1-5 mol% of a PEG-lipid. 33. A method of expressing a bispecific engager in a cell, comprising contacting a cell with a composition according to any one of claims 1-32. 34. A method of reducing or eliminating a B cell population associated with a disease, comprising administering to a subject with a disease a composition according to any one of claims 1-32. 35. The method of claim 34, wherein the disease is a B cell cancer. 36. The method of 34 or 35, wherein the disease is multiple myeloma, optionally wherein the disease is relapsed or refractory multiple myeloma. 37. The method of any one of claims 34-36, further comprising administering a second composition comprising a second nucleic acid encoding a different bispecific engager, the second nucleic acid comprising a second mRNA comprising in order from the 5’ to 3’ end of the second mRNA (i) a second open reading frame encoding a second VHH binding moiety that binds to a molecule expressed on the surface of a B cell, (ii) a second open reading frame encoding an IgG4 PAA CH2 and CH3 domain, and (iii) a second open reading frame encoding a VHH binding moiety that binds to a molecule expressed on the surface of an NK cell. 38. The method of claim 37, further comprising administering a third composition comprising a third nucleic acid encoding a further different bispecific engager, the third nucleic acid comprising a third mRNA comprising in order from the 5’ to 3’ end of the third mRNA (i) a third open reading frame encoding a third VHH binding moiety that binds to a molecule expressed on the surface of a B cell, (ii) a third open reading frame encoding an IgG4 PAA CH2 and CH3 domain, and (iii) a third open reading frame encoding a VHH binding moiety that binds to a molecule expressed on the surface of an NK cell. 300 Atty. Docket No.45817-0158WO1 39. The method of claim 37 or 38, wherein each of the composition, the second composition, and/or the third composition are present in the same delivery vehicle. 40. The method of claim 37 or 38, wherein each of the composition, the second composition, and/or the third composition are present in different delivery vehicles. 41. The method of claim 37 or 38, wherein each of the composition, the second composition, and/or the third composition are administered concurrently. 42. The method of claim 37 or 38, wherein each of the composition, the second composition, and/or the third composition are administered sequentially. 43. The method of any one of claims 34-42, further comprising administering to the subject a cytokine or an mRNA encoding the cytokine. 44. The method of claim 43, wherein the cytokine is IL-15, optionally wherein the cytokine is HSA-sIL-15, and comprises an amino acid sequence that is at least 85%, 90%, 95%, or 100% identical to SEQ ID NO: 729. 45. The method of claim 43 or 44, wherein the cytokine or mRNA encoding the cytokine and the composition are present in the same delivery vehicle. 46. The method of claim 43 or 44, wherein the cytokine or mRNA encoding the cytokine and the composition are present in different delivery vehicles. 47. A method of treating relapsed or refractory multiple myeloma in a subject in need thereof, comprising administering to the subject a composition according to claim 1, wherein the molecule expressed on the surface of a B cell is FcRH5 and the molecule expressed on the surface of an NK cell is CD16a, wherein the composition further comprises a lipid nanoparticle delivery vehicle encapsulating the mRNA. 301 Atty. Docket No.45817-0158WO1 48. The method of claim 47, wherein prior to treatment the subject has had prior exposure to one or more of all of a proteasome inhibitor, an immunomodulatory drug (IMiD), and an anti-cluster of differentiation (CD38) monoclonal antibody. 49. The method of claim 47 or claim 48, wherein prior to treatment the subject has received at least three prior lines of prior therapy and/or is triple-class refractory. 50. The method of any one of claims 47-49, wherein the subject is intolerant of one or more or all of a proteasome inhibitor, IMiD, or an anti-CD38 mAb. 51. The method of any of claims 47-50, wherein the mRNA comprises a nucleotide sequence comprising any one of SEQ ID NO: 463 or that encodes an amino acid sequence of SEQ ID NO: 464 or 433. 52. A composition comprising (1) a nucleic acid encoding a bispecific engager, the nucleic acid comprising an mRNA polynucleotide comprising in order from the 5’ to 3’ end of the mRNA (i) an open reading frame encoding a VHH binding moiety that binds to a molecule expressed on the surface of a B cell, optionally wherein the molecule expressed on the surface of a B cell is FcRH5, (ii) an open reading frame encoding a hinge region, optionally wherein the hinge is an IgG4 PAA CH2 and CH3 domain, and (iii) an open reading frame encoding a VHH binding moiety that binds to a molecule expressed on the surface of an NK cell, optionally wherein the molecule expressed on the surface of an NK cell is human CD16a, and (2) a nucleic acid encoding a polypeptide comprising a human interleukin 15 (IL-15) cytokine, optionally wherein the IL-15 cytokine is HSA-sIL-15. 53. The composition of claim 52, wherein the polypeptide comprising a human interleukin 15 (IL-15) cytokine comprises an amino acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, 302 Atty. Docket No.45817-0158WO1 at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the sequence set forth in SEQ ID NO:729. 54. The composition of claim 52, wherein the nucleic acid encoding the polypeptide comprises a nucleic acid sequence that is at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the sequence set forth in SEQ ID NO:728. 55. A lipid nanoparticle comprising a composition according to any one of claims 52-54. 56. A pharmaceutical composition comprising the composition according to any one of claims 52-54, and a pharmaceutically acceptable carrier. 57. A pharmaceutical composition comprising the lipid nanoparticle according to claim 55, and a pharmaceutically acceptable carrier. 58. A method of reducing or eliminating a B cell population associated with a disease, comprising administering to a subject with the disease a composition according to any one of claims 52-54, a lipid nanoparticle according to claim 55, or a pharmaceutical composition according to any one of claims 56 or 57, optionally wherein the subject is human. 59. The method of claim 58, wherein the disease is a B cell cancer. 60. The method of any one of claims 58 or 59, wherein the disease is multiple myeloma, optionally wherein the disease is relapsed multiple myeloma or refractory multiple myeloma. 303