WO2025245176A1 - Multispecific antibody constructs - Google Patents

Abstract

Multispecific antibodies described herein comprise paired mutations in the heavy chain constant domain (CH1) and light chain constant domain (CL) that increase the correct pairing of heavy and light chains.

Description

Attorney Docket No.14681-WO-PCT MULTISPECIFIC ANTIBODY CONSTRUCTS Cross-Reference To Related Applications [0001] This application claims the benefit of US Provisional Application No. 63/650,594, filed May 22, 2024, the entire contents of which are hereby incorporated herein by reference. Field [0002] Embodiments described herein generally relate to multispecific antibody designs that facilitate correct pairing. Background [0003] Generation of multispecific antibodies is challenging. A bispecific antibody, for example, has two different heavy chains, two different light chains, and at least two different variable regions. Expressing the heavy and light chains of two antibodies expressed a cell to generate the bispecific antibody theoretically results in 16 different combinations (10 different molecules). Only one of the possible theoretical combinations is bispecific and the remaining pairings are non-functional or monospecific. Consequently, the yield and purity of the bispecific antibody is unfavorable especially for large scale production of therapeutic bispecific antibodies. Summary [0004] Multispecific antibodies described herein comprise paired mutations in the heavy chain constant domain (CH1) and light chain constant domain (CL) that increase the correct pairing of heavy and light chains. In certain embodiments, multispecific antibodies comprise paired mutations in the CH1 and CL domains further comprise Fc region mutations that modulate effector function and/or facilitate heavy chain pairing. Brief Description of the Drawings [0005] FIG.1 depicts a schematic identifying the CH1, CL, and CH3 amino acid substitutions in Antibody 1 (Ab1) and Antibody 2 (Ab2). [0006] FIG.2 depicts a protein gel of Antibody 1 (Ab1) and Antibody 2 (Ab2). Attorney Docket No.14681-WO-PCT [0007] FIG.3 depicts Mono S polishing chromatograms of Antibody 1 (Ab1) and Antibody 2 (Ab2). [0008] FIG.4 depicts a protein gel of Antibody 1 (Ab1) and Antibody 2 (Ab2) after Mono S polishing. [0009] FIG.5 depicts tumor killing activity of Antibody 1 (Ab1) and Antibody 2 in a A549 lung cancer cell line (target) cocultured with human T cells (effector) in a 1:4 T:E ratio. The graph plots % cell death (y-axis) as a function of drug concentration (log, nM) (x-axis). [0010] FIG.6 depicts Protein A yields from antibodies having matched VH/VL pairs but having different mutation sets, either Mutation Set 1 or Mutation Set 2. [0011] FIG.7 depicts mass spectrometry data from antibodies having matched VH/VL pairs but having different mutation sets, either Mutation Set 1 or Mutation Set 2. [0012] FIG.8 depicts Analytical Size Exclusion Chromatography (AnSEC) data from antibodies having matched VH/VL pairs but having different mutation sets, either Mutation Set 1 or Mutation Set 2. [0013] FIG.9 depicts viability of CHO1 cells that were stably transfected with antibodies having matched VH/VL pairs but having different mutation sets, either Mutation Set 1 or Mutation Set 2. [0014] FIG.10 depicts protein gels of a panel of bispecific antibodies in a 1+1 Fab format or 2+1 Fab format, and having CH1/CL Pairing Mutation Set 1 or Pairing Mutation Set 2, that were purified by a PrismA step and Jetted SP35 IEX polishing step. Definitions [0015] It will be understood that amino acid substitutions offset by slash “/” within a grouping means each residue is included (e.g., L234A/L235A/D265S means L234A, L235A and D265S). [0016] In various embodiments, amino acid substitution residues are numbering according to EU or Kabat numbering convention. If a numbering convention is not indicated, a person of skill will readily be able to determine whether the residue number is according to EU or Kabat based on the wildtype amino acid residue at the indicated residue number. Similarly, a person of skill will readily be able to determine whether an amino acid has been incorrectly associated with EU instead of Kabat or vice versa, based on the wildtype amino acid residue at the indicated residue number. Attorney Docket No.14681-WO-PCT Detailed Description I. Heavy Chain Constant Region 1 and Light Chain Constant Region Paired Mutations [0017] In several embodiments, multispecific antibodies described herein comprise paired mutations in the heavy chain constant domain (CH1) and light chain constant domain (CL). In certain embodiments, residues in the CH1 and CL domains are substituted with charged amino acids to increase the correct pairing of heavy and light chains. Modified CH1 and CL domains provided herein improve manufacturing of multispecific antibodies, which may include bivalent, trivalent and tetravalent antibodies, by increasing the purity of the correct multispecific chain pairing. [0018] In certain embodiments, a multispecific antibody comprises a first CL domain comprising a S176K and N137K or N137R mutation; a first CH1 domain comprising a S188D and T192D mutation; a second CL domain comprising a S176D and N137D mutation; and a second CH1 domain comprising a S188K and T192K mutation; wherein the residues are numbered according to Kabat (Kabat, E.A. et al., Sequences of proteins of immunological interest.5th Edition - US Department of Health and Human Services, NIH publication 91-3242, pp 662,680,689 (1991)). In certain aspects, the first CL domain is linked to the first CH1 domain and the second CL domain is linked to the second CH1 domain. In certain aspects, the first CL domain is linked to the first CH1 domain by a disulfide bond and the second CL domain is linked to the second CH1 domain by a disulfide bond. [0019] In certain embodiments, a multispecific antibody comprises a first CL domain comprising a S176K and N137K mutation; a first CH1 domain comprising a S188D and T192D mutation; a second CL domain comprising a S176D and N137D mutation; and a second CH1 domain comprising a S188K and T192K mutation; wherein the residues are numbered according to Kabat (Kabat, E.A. et al., Sequences of proteins of immunological interest.5th Edition - US Department of Health and Human Services, NIH publication 91-3242, pp 662,680,689 (1991)). [0020] In certain embodiments, a multispecific antibody comprises a first CL domain comprising a S176K and N137R mutation; a first CH1 domain comprising a S188D and T192D mutation; a second CL domain comprising a S176D and N137D mutation; and a second CH1 domain comprising a S188K and T192K mutation; wherein the residues are numbered according to Kabat (Kabat, E.A. et al., Sequences of proteins of immunological interest.5th Edition - US Department of Health and Human Services, NIH publication 91-3242, pp 662,680,689 (1991)). Attorney Docket No.14681-WO-PCT [0021] In certain embodiments, a multispecific antibody comprises a first Fab region comprising a VH domain, a VL domain, a CL domain, and a CH1 domain, wherein the CL domain comprises a S176K and N137K or N137R mutation, and the CH1 domain comprises a S188D and T192D mutation; a second Fab region comprising a VH domain, a VL domain, a CL domain, and a CH1 domain, wherein the CL domain comprises a S176D and N137D mutation and the CH1 domain comprises a S188K and T192K mutation; and a Fc region comprising a first CH2 domain, a second CH2 domain, a first CH3 domain, and a second CH3 domain, wherein the Fc region comprises one or more mutations for enhancing or reducing Fc effector function, wherein the residues are numbered according to Kabat (Kabat, E.A. et al., Sequences of proteins of immunological interest.5th Edition - US Department of Health and Human Services, NIH publication 91-3242, pp 662,680,689 (1991)). [0022] In certain embodiments, a multispecific antibody comprises a first Fab region comprising a VH domain, a VL domain, a CL domain, and a CH1 domain, wherein the CL domain comprises a S176K and N137K mutation, and the CH1 domain comprises a S188D and T192D mutation; a second Fab region comprising a VH domain, a VL domain, a CL domain, and a CH1 domain, wherein the CL domain comprises a S176D and N137D mutation and the CH1 domain comprises a S188K and T192K mutation; and a Fc region comprising a first CH2 domain, a second CH2 domain, a first CH3 domain, and a second CH3 domain, wherein the Fc region comprises one or more mutations for enhancing or reducing Fc effector function, wherein the residues are numbered according to Kabat (Kabat, E.A. et al., Sequences of proteins of immunological interest.5th Edition - US Department of Health and Human Services, NIH publication 91-3242, pp 662,680,689 (1991)). [0023] In certain embodiments, a multispecific antibody comprises a first Fab region comprising a VH domain, a VL domain, a CL domain, and a CH1 domain, wherein the CL domain comprises a S176K and N137R mutation, and the CH1 domain comprises a S188D and T192D mutation; a second Fab region comprising a VH domain, a VL domain, a CL domain, and a CH1 domain, wherein the CL domain comprises a S176D and N137D mutation and the CH1 domain comprises a S188K and T192K mutation; and a Fc region comprising a first CH2 domain, a second CH2 domain, a first CH3 domain, and a second CH3 domain, wherein the Fc region comprises one or more mutations for enhancing or reducing Fc effector function, wherein the residues are numbered according to Kabat (Kabat, E.A. et al., Sequences of proteins of immunological interest.5th Edition - US Department of Health and Human Services, NIH publication 91-3242, pp 662,680,689 (1991)). Attorney Docket No.14681-WO-PCT [0024] In certain embodiments, a multispecific antibody comprises a first CL domain comprising a S176K and N137K or N137R mutation; a first CH1 domain comprising a S183D and T187D mutation; a second CL domain comprising a S176D and N137D mutation; and a second CH1 domain comprising a S183K and T187K mutation; wherein the residues are numbered according to EU numbering (Edelman, G.M. et al., Proc. Natl. Acad. USA, 63, 78-85 (1969)). [0025] In certain embodiments, a multispecific antibody comprises a first CL domain comprising a S176K and N137K mutation; a first CH1 domain comprising a S183D and T187D mutation; a second CL domain comprising a S176D and N137D mutation; and a second CH1 domain comprising a S183K and T187K mutation; wherein the residues are numbered according to EU numbering (Edelman, G.M. et al., Proc. Natl. Acad. USA, 63, 78-85 (1969)). [0026] In certain embodiments, a multispecific antibody comprises a first CL domain comprising a S176K and N137R mutation; a first CH1 domain comprising a S183D and T187D mutation; a second CL domain comprising a S176D and N137D mutation; and a second CH1 domain comprising a S183K and T187K mutation; wherein the residues are numbered according to EU numbering (Edelman, G.M. et al., Proc. Natl. Acad. USA, 63, 78-85 (1969)). [0027] In certain embodiments, a multispecific antibody comprises a first Fab region comprising a VH domain, a VL domain, a CL domain, and a CH1 domain, wherein the CL domain comprises a S176K and N137K or N137R mutation, and the CH1 domain comprises a S183D and T187D mutation; a second Fab region comprising a VH domain, a VL domain, a CL domain, and a CH1 domain, wherein the CL domain comprises a S176D and N137D mutation and the CH1 domain comprises a S183K and T187K mutation; and a Fc region comprising a first CH2 domain, a second CH2 domain, a first CH3 domain, and a second CH3 domain, wherein the Fc region comprises one or more mutations for enhancing or reducing Fc effector function, wherein the residues are numbered according to EU numbering (Edelman, G.M. et al., Proc. Natl. Acad. USA, 63, 78-85 (1969)). [0028] In certain embodiments, a multispecific antibody comprises a first Fab region comprising a VH domain, a VL domain, a CL domain, and a CH1 domain, wherein the CL domain comprises a S176K and N137K mutation, and the CH1 domain comprises a S183D and T187D mutation; a second Fab region comprising a VH domain, a VL domain, a CL domain, and a CH1 domain, wherein the CL domain comprises a S176D and N137D mutation and the CH1 domain comprises a S183K and T187K mutation; and a Fc region comprising a first CH2 domain, a second CH2 domain, a first CH3 domain, and a second CH3 domain, wherein the Fc Attorney Docket No.14681-WO-PCT region comprises one or more mutations for enhancing or reducing Fc effector function, wherein the residues are numbered according to EU numbering (Edelman, G.M. et al., Proc. Natl. Acad. USA, 63, 78-85 (1969)). [0029] In certain embodiments, a multispecific antibody comprises a first Fab region comprising a VH domain, a VL domain, a CL domain, and a CH1 domain, wherein the CL domain comprises a S176K and N137R mutation, and the CH1 domain comprises a S183D and T187D mutation; a second Fab region comprising a VH domain, a VL domain, a CL domain, and a CH1 domain, wherein the CL domain comprises a S176D and N137D mutation and the CH1 domain comprises a S183K and T187K mutation; and a Fc region comprising a first CH2 domain, a second CH2 domain, a first CH3 domain, and a second CH3 domain, wherein the Fc region comprises one or more mutations for enhancing or reducing Fc effector function, wherein the residues are numbered according to EU numbering (Edelman, G.M. et al., Proc. Natl. Acad. USA, 63, 78-85 (1969)). II. Fc Region Modifications A. Effector Function Modifications [0030] In certain embodiments, multispecific antibodies described herein comprising paired mutations in the heavy chain constant domain (CH1) and light chain constant domain (CL) can further comprise an Fc region having one or more amino acid substitutions. In certain embodiments, a multispecific antibody comprises a first CL domain comprising a S176K and N137K or N137R mutation (numbering according to Kabat); a first CH1 domain comprising a S188D and T192D mutation (numbering according to Kabat); a second CL domain comprising a S176D and N137D mutation (numbering according to Kabat); a second CH1 domain comprising a S188K and T192K mutation (numbering according to Kabat); and a Fc region comprising one or more amino acid substitutions. [0031] In certain embodiments, a multispecific antibody comprises a first CL domain comprising a S176K and N137K or N137R mutation (numbering according to EU); a first CH1 domain comprising a S183D and T187D mutation (numbering according to EU); a second CL domain comprising a S176D and N137D mutation (numbering according to EU); a second CH1 domain comprising a S183K and T187K mutation (numbering according to EU); and a Fc region comprising one or more amino acid substitutions. [0032] Any of the Fc amino acid substitutions described herein may be combined with the paired mutations in the heavy chain constant domain (CH1) and light chain constant domain Attorney Docket No.14681-WO-PCT (CL) described herein. Although certain Fc amino acid substitutions may be described in the context of IgG1, IgG2, IgG3, or IgG4, it will be understood that such amino acid substitutions may be used in any immunologlobulin G (IgG) Fc region. In other words, description of certain Fc amino acid substitutions in the context of IgG1, IgG2, IgG3, or IgG4 is not limited to that particular subclass and may pertain to any immunologlobulin G (IgG). A person of skill in the art will understand amino acid numbering differences among the isotypes and be able to renumber accordingly. [0033] The multispecific antibodies described herein may comprise an Fc comprising one or more amino acid modifications, typically to alter one or more functional properties of the antibody, such as serum half-life, complement fixation, Fc receptor binding, and/or antigen- dependent cellular cytotoxicity. For example, one may make modifications in the Fc region in order to generate an Fc variant with (a) increased or decreased antibody- dependent cell- mediated cytotoxicity (ADCC), (b) increased or decreased complement mediated cytotoxicity (CDC), (c) increased or decreased affinity for Clq and/or (d) increased or decreased affinity for a Fc receptor relative to the parent Fc. Such Fc region variants will generally comprise at least one amino acid modification in the Fc region. Combining amino acid modifications is thought to be particularly desirable. For example, the variant Fc region may include two, three, four, five, etc substitutions therein, e.g. of the specific Fc region positions identified herein. Exemplary Fc sequence variants are disclosed herein, and are also provided at U.S. Pat. Nos.5,624,821; 6,277,375; 6,737,056; 6,194,551; 7,317,091; 8,101,720; PCT Patent Publications WO 00/42072; WO 01/58957; WO 04/016750; WO 04/029207; WO 04/035752; WO 04/074455; WO 04/099249; WO 04/063351 ; WO 05/070963; WO 05/040217, WO 05/092925 and WO 06/020114. [0034] ADCC activity may be reduced by modifying the Fc region. In certain embodiments, sites that affect binding to Fc receptors may be removed (e.g., by mutation), preferably sites other than salvage receptor binding sites. In other embodiments, an Fc region may be modified to remove an ADCC site. ADCC sites are known in the art; see, for example, Sarmay et al. (1992) Molec. Immunol.29 (5): 633-9 with regard to ADCC sites in IgG1. In one embodiment, _{the G236R and L328R variant of human IgG1 effectively eliminates Fc R binding. Horton et al.} (2011) J. Immunol.186:4223 and Chu et al. (2008) Mol. Immunol.45:3926. In other _{embodiments, the Fc having reduced binding to Fc Rs comprised the amino acid substitutions} L234A, L235E and G237A. Gross et al. (2001) Immunity 15:289. In some embodiments, the Fc comprises the mutations L234A and L235E. Attorney Docket No.14681-WO-PCT [0035] CDC activity may also be reduced by modifying the Fc region. Mutations at IgG1 positions D270, K322, P329 and P331, specifically alanine mutations D270A, K322A, P329A and P331A, significantly reduce the ability of the corresponding antibody to bind Clq and activate complement. Idusogie et al. (2000) Immunol.164:4178; WO 99/51642. Modification of position 331 of IgG1 (e.g. P331S) has been shown to reduce complement binding. Tao et al. (1993 ) J. Exp. Med.178:661 and Canfield & Morrison (1991) J. Exp. Med.173:1483. In another example, one or more amino acid residues within amino acid positions 231 to 239 are altered to thereby reduce the ability of the antibody to fix complement. WO 94/29351. In some embodiments, the Fc with reduced complement fixation has the amino acid substitutions A330S and P331S. Gross et al. (2001) Immunity 15:289. [0036] For uses where effector function is to be avoided altogether, e.g. when antigen binding alone is sufficient to generate the desired therapeutic benefit, and effector function only leads to (or increases the risk of) undesired side effects, IgG4 antibodies may be used, or antibodies or fragments lacking the Fc region or a substantial portion thereof can be devised, or the Fc may be mutated to eliminate glycosylation altogether (e.g. N297A). Alternatively, a hybrid construct of human IgG2 (CRI domain and hinge region) and human IgG4 (CR2 and CR3 domains) has been generated that is devoid of effector function, lacking the ability to bind _{the Fc Rs (like IgG2) and unable to activate complement (like IgG4). Rother et al. (2007) Nat.} Biotechnol.25 : 1256. See also Mueller et al. (1997) Mol. Immunol.34:441; Labrijn et al. (2008) Curr. Op. Immunol.20:479 (discussing Fc modifications to reduce effector function generally). [0037] In other embodiments, the Fc region is altered by replacing at least one amino acid residue with a different amino acid residue to reduce all effector function(s) of the antibody. For example, one or more amino acids selected from amino acid residues 234, 235, 236, 237, 297, 318, 320 and 322 can be replaced with a different amino acid residue such that the antibody has decreased affinity for an effector ligand but retains the antigen-binding ability of the parent antibody. The effector ligand to which affinity is altered can be, for example, an Fc receptor (residues 234, 235, 236, 237, 297) or the CI component of complement (residues 297, 318, 320, 322). U.S. Patent Nos.5,624,821 and 5,648,260, both by Winter et al. [0038] WO 88/007089 proposed modifications in the IgG Fc region to decrease binding to _{Fc RI to decrease ADCC (234A; 235E; 236A; G237A) or block binding to complement} component Cl q to eliminate CDC (E318A or V/K320A and K322A/Q). See also Duncan & Winter (1988) Nature 332 :563; Chappel et al. (1991) Pro Nat'l Acad. Sci. (USA) 88:9036; and Attorney Docket No.14681-WO-PCT _{Sondermann et al. (2000) Nature 406:267 (discussing the effects of these mutations on Fc RIII} binding). [0039] Fc modifications reducing effector function also include substitutions, insertions, and deletions at positions 234, 235, 236, 237, 267, 269, 325, and 328, such as 234G, 235G, 236R, 237K, 267R, 269R, 325L, and 328R. An Fc variant may comprise 236R/328R. Other _{modifications for reducing Fc R and complement interactions include substitutions 297A, 234A,} 235A, 237A, 318A, 228P, 236E, 268Q, 309L, 330S , 331 S , 220S, 226S, 229S, 238S, 233P, and 234V. These and other modifications are reviewed in Strohl (2009) Current Opinion in Biotechnology 20:685-691. Effector functions (both ADCC and complement activation) can be reduced, while maintaining neonatal FcR binding (maintaining half-life), by mutating IgG residues at one or more of positions 233-236 and 327-331, such as E233P, L234V, L235A, optionally G236A, A327G, A330S and P331S in IgG1; E233P, F234V, L235A, optionally G236A in IgG4; and A330S and P331 S in IgG2. See Armour et al. (1999) Eur. J. Immunol. 29:2613; WO 99/58572. Other mutations that reduce effector function include L234A and L235A in IgG1 (Alegre et al. (1994) Transplantation 57: 1537); V234A and G237A in IgG2 (Cole et al. (1997) J. Immunol.159:3613; see also U.S . Pat. No.5,834,597); and S228P and L235E for IgG4 (Reddy et al. (2000) J. Immunol.164: 1925). Another combination of mutations for reducing effector function in a human IgG1 include L234F, L235E and P331S. Oganesyan et al. (2008) Acta Crystallogr. D. Biol. Crystallogr.64:700. See generally Labrijn et gal. (2008) Curr. Op. Immunol.20:479. Additional mutations found to decrease effector function in the context of an Fc (IgG1) fusion protein (abatacept) are C226S, C229S and P238S (EU residue numbering). Davis et al. (2007) J. Immunol.34:2204. [0040] Other Fc variants having reduced ADCC and/or CDC are disclosed at Glaesner et al. (2010) Diabetes Metab. Res. Rev.26:287 (F234A and L235A to decrease ADCC and ADCP in an IgG4); Hutchins et al. (1995) Proc. Nat'l Acad. Set. (USA) 92: 11980 (F234A, G237A and E318A in an IgG4); An et al. (2009) MAbs 1 :572 and U.S. Pat. App. Pub.2007/0148167 (H268Q, V309L, A330S and P331S in an IgG2); McEarchern et al. (2007) Blood 109: 1185-92 (C226S, C229S, E233P, L234V, L235A in an IgG1); Vafa et al. (2014) Methods 65(1): 114-26 (V234V, G237A, P238S, H268A, V309L, A330S, P331S in an IgG2). [0041] In certain embodiments, an Fc is chosen that has essentially no effector function, i.e., _{it has reduced binding to Fc Rs and reduced complement fixation. An exemplary Fc, e.g., IgG1} Fc, that is effectorless comprises the following five mutations: L234A, L235E, G237A, A330S and P331S. Gross et al. (2001) Immunity 15:289. These five substitutions may be combined Attorney Docket No.14681-WO-PCT with N297A to eliminate glycosylation as well. In certain embodiments, the Fc region comprises the mutations L234A, L235A, and G237A. As used herein "IgG1.3" refers to an IgG1 heavy chain comprising L234A, L235E and G237A. IgG1 constant regions comprising these three mutations may also comprise additional mutations, such as those described herein. An IgG1.3 Fc provides an antibody with significantly reduced effector function, such as ADCC and CDC. In certain embodiments, an Fc comprises the mutations of IgG1.3 and additional mutations, e.g., P238K. In certain embodiments, an antibody comprises an IgG1.3 heavy chain constant region, which constant region does not comprise any other than mutation that modulates effector function, in addition to L234A, L235E and G237A. In certain embodiments, an antibody comprises an IgG1.3 heavy chain constant region, which constant region does not comprise any other than mutation, in addition to L234A, L235E and G237A. In certain embodiments, the Fc region comprises the mutations L23A, L235A, and D265S (AAS). In certain embodiments, the Fc region comprises the mutations L23A, L235A, and P329G (LALAPG). In certain embodiments, the Fc region comprises the mutations L234A, L235A, G237A, P238S, H268A, A330S, and P331S. [0042] Alternatively, ADCC activity may be increased by modifying the Fc region. With regard to ADCC activity, an IgG1 constant domain, rather than an IgG2 or IgG4, might be chosen for use in a drug where ADCC is desired. Alternatively, the Fc region may be modified to increase antibody dependent cellular cytotoxicity (ADCC) and/or to increase the affinity for an Fc receptor by modifying one or more amino acids at the following positions: 234, 235, 236, 238, 239, 240, 241, 243, 244, 245, 247, 248, 249, 252, 254, 255, 256, 258, 262, 263, 264, 265, 267, 268, 269, 270, 272, 276, 278, 280, 283, 285, 286, 289, 290, 292, 293, 294, 295, 296, 298, 299, 301, 303, 305, 307, 309, 312, 313, 315, 320, 322, 324, 325, 326, 327, 329, 330, 331 , 332, 333, 334, 335, 337, 338, 340, 360, 373, 376, 378, 382, 388, 389, 398, 414, 416, 419, 430, 433, 434, 435, 436, 437, 438 or 439. See WO 2012/142515; see also WO 00/42072. Exemplary substitutions include 236A, 239D, 239E, 268D, 267E, 268E, 268F, 324T, 332D, and 332E. Exemplary variants include 239D/332E, 236A/332E, 236A/239D/332E, 268F/324T, 267E/268F, 267E/324T, and 267E/268F/324T. For example, human IgG1 Fcs comprising the G236A _{variant, which can optionally be combined with I332E, have been shown to increase the Fc IIA / Fc IIB binding affinity ratio approximately 15-fold. Richards et al. (2008) Mol. Cancer Therap. 7:2517; Moore et al. ( 2010) mAbs 2: 181. Other modifications for enhancing Fc R and} complement interactions include but are not limited to substitutions 298A, 333A, 334A, 326A, 2471, 339D, 339Q, 280H, 290S, 298D, 298V, 243L, 292P, 300L, 396L, 3051, and 396L. These Attorney Docket No.14681-WO-PCT and other modifications are reviewed in Strohl (2009) Current Opinion in Biotechnology 20:685-691. Specifically, both ADCC and CDC may be enhanced by changes at position E333 of IgG1, e.g. E333A. Shields et al. (2001) /. Biol. Chem.276:6591. The use of P247I and A339D/Q mutations to enhance effector function in an IgG1 is disclosed at WO 2006/020114, and D280H, K290S + S298D/V is disclosed at WO 2004/074455. The K326A W and E333A/S variants have been shown to increase effector function in human IgG1, and E333S in IgG2. Idusogie et al. (2001) J. Immunol.166:2571. Specifically, the binding sites on human IgG1 for _{Fc Rl , Fc RII, Fc RIII and FcRn have been mapped, and variants with improved binding have} been described. Shields et al. (2001) J. Biol. Chem.276:6591-6604. Specific mutations at _{positions 256, 290, 298, 333, 334 and 339 were shown to improve binding to Fc RIII, including} the combination mutants T256A/S298A, S298A/E333A, S298A/K224A and _{S298A/E333A/K334A (AAA) (having enhanced Fc RIIIa binding and ADCC activity). Other IgG1 variants with strongly enhanced binding to Fc RIIIa have been identified, including} variants with S239D/I332E (DE) and S239D/I332E/A330L mutations which showed the greatest _{increase in affinity for Fc RIIIa, a decrease in Fc RIIb binding, and strong cytotoxic activity in} cynomolgus monkeys. Lazar et a/. (2006) Proc. Nat'lAcad Sci. (USA) 103:4005; Awan et al. (2010) Blood 115: 1204; Desjarlais & Lazar (2011) Exp. Cell Res.317: 1278. In addition, IgG1 mutants containing L235V, F243L, R292P, Y300L, V305I and P396L mutations which _{exhibited enhanced binding to Fc RIIIa and concomitantly enhanced ADCC activity in transgenic mice expressing human Fc RIIIa in models of B cell malignancies and breast cancer} have been identified. Stavenhagen et al. (2007) Cancer Res.67:8882; U.S. Pat. No.8,652,466; Nordstrom et al. (2011) Breast Cancer Res.13:R123. Different IgG isotypes also exhibit differential CDC activity (IgG3>IgG1>IgG2~IgG4). Dangl et al. (1988) EMBO J.7: 1989. For uses in which enhanced CDC is desired, it is also possible to introduce mutations that increase binding to Clq. The ability to recruit complement (CDC) may be enhanced by mutations at K326 and/or E333 in an IgG2, such as K326W (which reduces ADCC activity) and E333S, to increase binding to Clq, the first component of the complement cascade. Idusogie et al. (2001) J. Immunol.166:2571. Introduction of S267E / H268F / S324T (alone or in any combination) into human IgG1 enhances Clq binding. Moore et al. (2010) mAbs 2: 181. The Fc region of the IgG1/IgG3 hybrid isotype antibody "113F" of Natsume et al. (2008) Cancer Res.68:3863 (figure 1 therein) also confers enhanced CDC. See also Michaelsen et al. (2009) Scand. J. Immunol.70:553 and Redpath et al. (1998) Immunology 93:595. Additional mutations that can increase or decrease effector function are disclosed at Dall'Acqua et al. (2006) J. Immunol.177: Attorney Docket No.14681-WO-PCT 1129. See also Carter (2006) Nat. Rev. Immunol.6:343; Presta (2008) Curr. Op. Immunol. 20:460. [_{0043] Fc variants that enhance affinity for the inhibitory receptor Fc RIIb may also be} used, e.g. to enhance apoptosis-inducing or adjuvant activity. Li & Ravetch (2011) Science 333: 1030; Li & Ravetch (2012) Proc. Nat'l Acad. Sci (USA) 109: 10966; U.S. Pat. App. Pub. 2014/0010812. Such variants may provide an antibody with immunomodulatory activities _{related to Fc RIIb+ cells, including for example B cells and monocytes. In one embodiment, the Fc variants provide selectively enhanced affinity to Fc RIIb relative to one or more activating receptors. Modifications for altering binding to Fc RIIb include one or more modifications at a} position selected from the group consisting of 234, 235, 236, 237, 239, 266, 267, 268, 325, 326, _{327, 328, and 332, according to the EU index. Exemplary substitutions for enhancing Fc RIIb} affinity include but are not limited to 234D, 234E, 234F, 234W, 235D, 235F, 235R, 235Y, 236D, 236N, 237D, 237N, 239D, 239E, 266M, 267D, 267E, 268D, 268E, 327D, 327E, 328F, 328W, 328Y, and 332E. Exemplary substitutions include 235Y, 236D, 239D, 266M, 267E, _{268D, 268E, 328F, 328W, and 328Y. Other Fc variants for enhancing binding to Fc RIIb} include 235Y/267E, 236D/267E, 239D/268D, 239D/267E, 267E/268D, 267E/268E, and 267E/328F. Specifically, the S267E, G236D, S239D, L328F and I332E variants, including the S267E + L328F double variant, of human IgG1 are of particular value in specifically enhancing _{affinity for the inhibitory Fc RIIb receptor. Chu et al. (2008) Mol. Immunol. 45 :3926; U.S. Pat. App. Pub. 2006/024298; WO 2012/087928. Enhanced specificity for Fc RIIb (as distinguished from Fc RIIaR) may be obtained by adding the P238D substitution. Mimoto et al. (2013)} Protein. Eng. Des. & Selection 26:589; WO 2012/115241. [0044] In some embodiments of any of the IgG modified Fc, the Fc comprises N297A mutation according to EU numbering. In some embodiments of any of the IgG modified Fc, the Fc comprises N297K mutation according to EU numbering. In some embodiments of any of the IgG modified Fc, the Fc comprises D265A and N297A mutations according to EU numbering. In some embodiments of any of the IgG modified Fc, the Fc comprises D270A mutations according to EU numbering. In some embodiments, the IgG modified Fc comprises L234A and L235A mutations according to EU numbering. In some embodiments of any of the IgG modified Fc, the Fc comprises L234A and G237A mutations according to EU numbering. In some embodiments of any of the IgG modified Fc, the Fc comprises L234A, L235A and G237A mutations according to EU numbering. In some embodiments of any of the IgG modified Fc, the Fc comprises one or more (including all) of E233D, G237D, P238D, H268D, P271G, L328E, Attorney Docket No.14681-WO-PCT and A330R mutations according to EU numbering. In some embodiments of any of the IgG modified Fc, the Fc comprises one or more of S267E/L328F mutations according to EU numbering. In some embodiments of any of the IgG modified Fc, the Fc comprises P238D, L328E, E233D, G237D, H268D, P271G and A330R mutations according to EU numbering. In some embodiments of any of the IgG modified Fc, the Fc comprises G237D, P238D, H268D, P271G, L328E, and A330R mutations according to EU numbering. In some embodiments of any of the IgG modified Fc, the Fc comprises G237D, P238D, H268D, P271G, and A330R mutations according to EU numbering. In some embodiments of any of the IgG modified Fc, the Fc comprises P238D, S267E, L328E, E233D, G237D, H268D, P271G and A330R mutations according to EU numbering. In some embodiments of any of the IgG modified Fc, the Fc comprises P238D, S267E, L328E, G237D, H268D, P271G and A330R mutations according to EU numbering. In some embodiments of any of the IgG modified Fc, the Fc comprises C226S, C229S, E233P, L234V, and L235A mutations according to EU numbering. In some embodiments of any of the IgG modified Fc, the Fc comprises L234F, L235E, and P331S (FES) mutations according to EU numbering. In some embodiments of any of the IgG modified Fc, the Fc comprises L234A, L235A, and P331S (LALAPS) mutations according to EU numbering. In some embodiments of any of the IgG modified Fc, the Fc comprises S267E and L328F mutations according to EU numbering. In some embodiments of any of the IgG modified Fc, the Fc comprises N325S and L328F (NSLF) mutations according to EU numbering. In some embodiments of any of the IgG modified Fc, the Fc comprises S267E mutations according to EU numbering. In some embodiments of any of the IgG modified Fc, the Fc comprises a substitute of the constant heavy 1 (CH1) and hinge region of IgG1 with CH1 and hinge region of IgG2 (amino acids 118-230 of IgG2 according to EU numbering) with a Kappa light chain. In some embodiments, the Fc region comprises the mutations L234F, L235Q, and K322Q (FQQ). In some embodiments, the IgG is IgG1. [0045] In certain embodiments, an antibody comprises a heavy chain constant region comprising an IgG1 constant domain comprising one or more of L234A, L235E and G237A. As used herein "IgG1.3" refers to an IgG1 heavy chain comprising L234A, L235E and G237A. IgG1 constant regions comprising these three mutations may also comprise additional mutations, such as those described herein. An IgG1.3 Fc provides an antibody with significantly reduced effector function, such as ADCC and CDC. In certain embodiments, an Fc comprises the mutations of IgG1.3 and additional mutations, e.g., P238K. In certain embodiments, an antibody comprises an IgG1.3 heavy chain constant region, which constant region does not comprise any Attorney Docket No.14681-WO-PCT other than mutation that modulates effector function, in addition to L234A, L235E and G237A. In certain embodiments, an antibody comprises an IgG1.3 heavy chain constant region, which constant region does not comprise any other than mutation, in addition to L234A, L235E and G237A. [0046] In certain embodiments, an IgG1 Fc comprising a P238K mutation comprises no other mutations relative to a wild type IgG1 Fc, e.g., those described herein. In certain embodiments, an IgG1 Fc comprising a P238K mutation comprises 1-5 amino acid changes in addition to P238K relative to the wild type human IgG1 Fc, provided that the IgG1 Fc has reduced effector function. [0047] In certain embodiments, an IgG1 Fc comprising a P238K mutation does not comprise any other mutation that reduces effector function. In certain embodiments, an IgG1 Fc comprising a P238K mutation comprises 1-5 mutations that reduces effector function. [0048] In certain embodiments, an IgG Fc comprising a P238K mutation also comprises an L235E mutation and/or a K322A mutation, and may, in certain embodiments not contain any additional Fc mutation that modulates Fc effector function, e.g., it does not include a mutation at P330, P331, or a mutation in the lower hinge, e.g., at amino acids 234 and 236- 237. The IgG may be an IgG1 or IgG2. [0049] In certain embodiments, an IgG Fc comprises a P238K mutation. In certain embodiments, an IgG Fc comprises the mutations L235E and P238K. In certain embodiments, an IgG Fc comprises the mutations L235E, P238K, and K322A. [0050] In certain embodiments, an antibody comprises a heavy chain constant region comprising an IgG2 constant domain, or at least the hinge thereof, wherein the IgG2 constant domain or hinge thereof comprises a mutation selected from the group consisting of P238A, P238K, L235A, K322A, and optionally a mutation at C219 and/or C220, e.g., C219S and/or C220S. [0051] In some embodiments of any of the antibodies provided herein, the modified antibody Fc is an IgG2 modified Fc. In some embodiments, the IgG2 modified Fc comprises one or more modifications. For example, in some embodiments, the IgG2 modified Fc comprises one or more amino acid substitutions (e.g relative to a wild-type Fc region of the same isotype). In some embodiments of any of the IgG2 modified Fc, the one or more amino acid substitutions are selected from V234A (Alegre et al. Transplantation 57:1537-1543 (1994); Xu et al. Cell Immunol , 200:16-26 (2000)); G237A (Cole et al. Transplantation , 68:563-571 (1999)); H268Q, V309L, A330S, P331S (US 2007/0148167; Armour et al. Eur J Immunol 29: 2613-2624 (1999); Attorney Docket No.14681-WO-PCT Armour et al. The Haematology Journal (Suppl.1):27 (2000); Armour et al. The Haematology Journal (Suppl. l):27 (2000)), C131S, C219S, and/or C220S (White et al. Cancer Cell 27, 138- 148 (2015)); S267E, L328F (Chu et al. Mol Immunol, 45:3926-3933 (2008)); and M252Y, S254T, and/or T256E (YTE) according to the EU numbering convention. In some embodiments of any of the IgG2 modified Fc, the Fc comprises an amino acid substitution at positions V234A and G237A according to EU numbering. In some embodiments of any of the IgG2 modified Fc, the Fc comprises an amino acid substitution at positions C219S or C220S according to EU numbering. [0052] In some embodiments of any of the IgG2 modified Fc, the Fc comprises an amino acid substitution at positions A330S and P331S according to EU numbering. In some embodiments of any of the IgG2 modified Fc, the Fc comprises an amino acid substitution at positions S267E and L328F according to EU numbering. [0053] In some embodiments of any of the IgG2 modified Fc, the Fc comprises a C127S amino acid substitution according to the EU numbering convention (White et al., (2015) Cancer Cell 27, 138-148; Lightle et al. Protein Sci.19:753-762 (2010); and WO 2008/079246). In some embodiments of any of the IgG2 modified Fc, the antibody has an IgG2 isotype with a Kappa light chain constant domain that comprises a C214S amino acid substitution according to the EU numbering convention (White et al. Cancer Cell 27:138-148 (2015); Lightle et al. Protein Sci. 19:753-762 (2010); and WO 2008/079246). In some embodiments of any of the IgG2 modified Fc, the Fc comprises a C220S amino acid substitution according to the EU numbering convention. In some embodiments of any of the IgG2 modified Fc, the antibody has an IgG2 isotype with a Kappa light chain constant domain that comprises a C214S amino acid substitution according to the EU numbering convention. In some embodiments of any of the IgG2 modified Fc, the Fc comprises a C219S amino acid substitution according to the EU numbering convention. In some embodiments of any of the IgG2 modified Fc, the antibody has an IgG2 isotype with a Kappa light chain constant domain that comprises a C214S amino acid substitution according to the EU numbering convention. In some embodiments of any of the IgG2 modified Fc, the Fc includes an IgG2 isotype heavy chain constant domain 1(CH1) and hinge region (White et al. Cancer Cell 27:138-148 (2015)). In certain embodiments of any of the IgG2 modified Fc, the IgG2 isotype CHI and hinge region comprise the amino acid sequence of 118-230 according to EU numbering. In some embodiments of any of the IgG2 modified Fc, the antibody Fc region comprises a S267E amino acid substitution, a L328F amino acid substitution, Attorney Docket No.14681-WO-PCT or both, and/or a N297A or N297Q amino acid substitution according to the EU numbering convention. [0054] In some embodiments of any of the IgG2 modified Fc, the Fc further comprises one or more amino acid substitution at positions E430G, E430S, E430F, E430T, E345K, E345Q, E345R, E345Y, S440Y, and S440W according to EU numbering. In some embodiments of any of the IgG2 modified Fc, the Fc may further comprise one or more mutations to enhance the antibody half-life in human serum (e.g., one or more (including all) of M252Y, S254T, and T256E (YTE) mutations according to the EU numbering convention). In some embodiments of any of the IgG2 modified Fc, the Fc may further comprise A330S and P331S. [0055] In some embodiments of any of the IgG2 modified Fc, the Fc is an IgG2/4 hybrid Fc. In some embodiments, the IgG2/4 hybrid Fc comprises IgG2 aa 118 to 260 and IgG4 aa 261 to 447. In some embodiments of any IgG2 modified Fc, the Fc comprises one or more amino acid substitutions at positions H268Q, V309L, A330S, and P331S according to EU numbering. [0056] In some embodiments of any of the IgG1 and/or IgG2 modified Fc, the Fc comprises one or more additional amino acid substitutions selected from A330L, L234F; L235E, or P331 S according to EU numbering; and any combination thereof. [0057] In certain embodiments of any of the IgG1 and/or IgG2 modified Fc, the Fc comprises one or more amino acid substitutions at a residue position selected from C127S, L234A, L234F, L235A, L235E, S267E, K322A, L328F, A330S, P331S, E345R, E430G, S440Y, and any combination thereof according to EU numbering. In certain embodiments, the Fc comprises the amino acid substitutions L234A, L235A, and K322A. In some embodiments of any of the IgG1 and/or IgG2 modified Fc, the Fc comprises an amino acid substitution at positions E430G, L243A, L235A, and P331S according to EU numbering. In some embodiments of any of the IgG1 and/or IgG2 modified Fc, the Fc comprises an amino acid substitution at positions E430G and P331 S according to EU numbering. In some embodiments of any of the IgG1 and/or IgG2 modified Fc, the Fc comprises an amino acid substitution at positions E430G and K322A according to EU numbering. In some embodiments of any of the IgG1 and/or IgG2 modified Fc, the Fc comprises an amino acid substitution at positions E430G, A330S, and P331S according to EU numbering. In some embodiments of any of the IgG1 and/or IgG2 modified Fc, the Fc comprises an amino acid substitution at positions E430G, K322A, A330S, and P331S according to EU numbering. In some embodiments of any of the IgG1 and/or IgG2 modified Fc, the Fc comprises an amino acid substitution at positions E430G, K322A, and A330S according to EU numbering. In some embodiments of any of the IgG1 and/or IgG2 Attorney Docket No.14681-WO-PCT modified Fc, the Fc comprises an amino acid substitution at positions E430G, K322A, and P331S according to EU numbering. [0058] In some embodiments of any of the IgG1 and/or IgG2 modified Fc, the Fc comprises an amino acid substitution at positions S267E and L328F according to EU numbering. In some embodiments of any of the IgG1 and/or IgG2 modified Fc, the Fc comprises an amino acid substitution at position C127S according to EU numbering. In some embodiments of any of the IgG1 and/or IgG2 modified Fc, the Fc comprises an amino acid substitution at positions E345R, E430G and S440Y according to EU numbering. In some embodiments, the Fc comprises one or more of the following mutations: SE (S267E), SELF (S267E/L328F), SDIE (S239D/I332E), SEFF, GASDALIE (G236A/S239D/A330L/I332E), and/or one or more mutations at the following amino acids: E233, L235, G237, P238, H268, P271, L328, A330 and K322. In some embodiments, the Fc comprises the mutations G236A, S239D, and I332E (ADE). In some embodiments, the Fc comprises the mutations G236A, A330L, and I332E (GAALIE). In some embodiments, the Fc comprises the mutations F243L, R292P, Y300L, V305I, and P396L (LPLIL). In some embodiments, the Fc comprises the mutations L235V, F243L, R292P, Y300L, and P396L (VLPLL). [0059] In some embodiments of any of the antibodies provided herein, the modified antibody Fc is an IgG4 modified Fc. In some embodiments, the IgG4 modified Fc comprises one or more modifications. For example, in some embodiments, the IgG4 modified Fc comprises one or more amino acid substitutions (e.g., relative to a wild-type Fc region of the same isotype). In some embodiments of any of the IgG4 modified Fc, the one or more amino acid substitutions are selected from L235A, G237A, S229P, L236E (Reddy et al. J Immunol 164:1925- 1933(2000)), S267E, E318A, L328F, M252Y, S254T, and/or T256E according to the EU numbering convention. In some embodiments of any of the IgG4 modified Fc, the Fc may further comprise L235A, G237A, and E318A according to the EU numbering convention. In some embodiments of any of the IgG4 modified Fc, the Fc may further comprise S228P and L235E according to the EU numbering convention. In some embodiments of any of the IgG4 modified Fc, the IgG4 modified Fc may further comprise S267E and L328F according to the EU numbering convention. [0060] In some embodiments of any of the IgG4 modified Fc, the IgG4 modified Fc comprises may be combined with an S228P mutation according to the EU numbering convention (Angal et al. Mol Immunol.30: 105-108 (1993)) and/or with one or more mutations Attorney Docket No.14681-WO-PCT described in (Peters et al. J Biol Chem.287(29):24525-33 (2012)) to enhance antibody stabilization. [0061] In some embodiments of any of the IgG4 modified Fc, the IgG4 modified Fc may further comprise one or more mutations to enhance the antibody half-life in human serum ( e.g ., one or more (including all) of M252Y, S254T, and T256E mutations according to the EU numbering convention). [0062] In some embodiments of any of the IgG4 modified Fc, the Fc comprises L235E according to EU numbering. In certain embodiments of any of the IgG4 modified Fc, the Fc comprises one or more amino acid substitutions at a residue position selected from C127S, F234A, L235A, L235E, S267E, K322A, L328F, E345R, E430G, S440Y, and any combination thereof, according to EU numbering. In some embodiments of any of the IgG4 modified Fc, the Fc comprises an amino acid substitution at positions E430G, L243A, L235A, and P331S according to EU numbering. In some embodiments of any of the IgG4 modified Fc, the Fc comprises an amino acid substitution at positions E430G and P331S according to EU numbering. In some embodiments of any of the IgG4 modified Fc, the Fc comprises an amino acid substitution at positions E430G and K322A according to EU numbering. In some embodiments of any of the IgG4 modified Fc, the Fc comprises an amino acid substitution at position E430 according to EU numbering. In some embodiments of any of the IgG4 modified Fc, the Fc region comprises an amino acid substitution at positions E430G and K322A according to EU numbering. In some embodiments of any of the IgG4 modified Fc, the Fc comprises an amino acid substitution at positions S267E and L328F according to EU numbering. In some embodiments of any of the IgG4 modified Fc, the Fc comprises an amino acid substitution at position C127S according to EU numbering. In some embodiments of any of the IgG4 modified Fc, the Fc comprises an amino acid substitution at positions E345R, E430G and S440Y according to EU numbering. [0063] In some embodiments, an Fc region comprises mutations that increase FcRn binding. In certain embodiments, an Fc region comprises the mutations M428L/N434S (LS). The M428L/N434S (LS) mutation increases half-life and FcRn binding at pH 6.0 (Zalevsky et al., Nat Biotechnol 2010 28:157–159). [0064] It will be understood that a multispecific antibody may comprise any of the Fc region mutations described herein in one or both of the CH2 domains or in one or both of the CH3 domains of the Fc region. Attorney Docket No.14681-WO-PCT B. Knob-Into-Hole Mutations [0065] In certain embodiments, multispecific antibodies described herein comprising paired mutations in the heavy chain constant domain (CH1) and light chain constant domain (CL) can further comprise an Fc region comprising a knob-into-hole modification. In certain embodiments, multispecific antibodies described herein comprising paired mutations in the heavy chain constant domain (CH1) and light chain constant domain (CL) can further comprise an Fc region comprising a knob-into-hole modification and one or more amino acid substitutions described herein that increase or decrease Fc effector function. [0066] Knob-into-hole technology is described in e.g. U.S. Pat. Nos.5,731,168; 7,695,936; 8,216,805; 8,765,412; Ridgway et al., Prot Eng 9, 617-621 (1996); and Carter, J Immunol Meth 248, 7-15 (2001). Generally, the method involves introducing a protuberance (“knob”) at the interface of a first polypeptide and a corresponding cavity (“hole”) in the interface of a second polypeptide, such that the protuberance can be positioned in the cavity so as to promote heterodimer formation and hinder homodimer formation. Protuberances are constructed by replacing small amino acid side chains from the interface of the first polypeptide with larger side chains (e.g. tyrosine or tryptophan). Compensatory cavities of identical or similar size to the protuberances are created in the interface of the second polypeptide by replacing large amino acid side chains with smaller ones (e.g. alanine or threonine). The protuberance and cavity can be made by altering the nucleic acid encoding the polypeptides, e.g. by site-specific mutagenesis, or by peptide synthesis. In a specific embodiment a knob modification comprises the amino acid substitution T366W in one of the two subunits of the Fc domain, and the hole modification comprises the amino acid substitutions T366S, L368A and Y407V in the other one of the two subunits of the Fc domain. In a further specific embodiment, the subunit of the Fc domain comprising the knob modification additionally comprises the amino acid substitution S354C, and the subunit of the Fc domain comprising the hole modification additionally comprises the amino acid substitution Y349C. Introduction of these two cysteine residues results in the formation of a disulfide bridge between the two subunits of the Fc region, thus further stabilizing the dimer (Carter, J Immunol Methods 248, 7-15 (2001)). Thus, in such configurations, a first Fc region polypeptide comprises amino acid modifications to form the “knob” and a second Fc region polypeptide comprises amino acid modifications to form the “hole” thus forming an Fc heterodimer comprising complementary Fc polypeptides. [0067] In some aspects, the bispecific antibody comprises an Fc region comprising a first polypeptide and a second polypeptide. In some aspects, the first polypeptide comprises the Attorney Docket No.14681-WO-PCT amino acid substitution T366Y, and the second polypeptide comprises the amino acid substation Y407T. In some aspects, the first polypeptide comprises the amino acid substitution T366W, and the second polypeptide comprises the amino acid substitutions T366S, L368W, and Y407V. In some aspects, the first polypeptide comprises the amino acid substitution T366W, and the second polypeptide comprises the amino acid substitutions T366S, L368A, and Y407V. In some aspects, the first polypeptide comprises the amino acid substitutions T366W, and S354C and the second polypeptide comprises the amino acid substitutions T366S, L368A, Y407V, and Y349C. In some aspects, the first polypeptide comprises Y349C, T366S, L368A, Y407V disulfide- stabilized Hole mutations (EU numbering, and the second polypeptide comprises S354C and T366W disulfide-stabilized Knob mutations (EU numbering). In some aspects, the first polypeptide comprises the amino acid substitutions T350V, L351Y, F405A, Y407V, and the second polypeptide comprises the amino acid substitutions T350V, T366L, K392L, and T394W. In some aspects, the first polypeptide comprises the amino acid substitutions K360D, D399M, and Y407A, and the second polypeptide comprises the amino acid substitutions E345R, Q347R, T366V, and K409V. In some aspects, the first polypeptide comprises the amino acid substitutions K409D and K392D, and the second polypeptide comprises the amino acid substitutions D399K and E356K. In some aspects, the first polypeptide comprises the amino acid substitutions K360E and K409W, and the second polypeptide comprises the amino acid substitutions Q347R, D399V, and F405T. In some aspects, the first polypeptide comprises the amino acid substitutions L360E, K409W, and Y349C, and the second polypeptide comprises the amino acid substitutions Q347R, D399V, F405T, and S354C. In some aspects, the first polypeptide comprises the amino acid substitutions K370E and K409W, and the second polypeptide comprises the amino acid substitutions E357N, D399V, and F405T. In some aspects, the substitution is according to EU numbering. [0068] In some aspects, the knob mutation comprises the amino acid substitution T366W according to EU numbering. [0069] In some aspects, the hole mutation comprises the amino acids substitutions T366S, L368A, and Y407V according to EU numbering. [0070] In a specific embodiment, in (the CH3 domain of) the first subunit of the Fc domain (the “knobs” subunit) the threonine residue at position 366 is replaced with a tryptophan residue (T366W), and in (the CH3 domain of) the second subunit of the Fc domain (the “hole” subunit) the tyrosine residue at position 407 is replaced with a valine residue (Y407V). In one embodiment, in the second subunit of the Fc domain additionally the threonine residue at Attorney Docket No.14681-WO-PCT position 366 is replaced with a serine residue (T366S) and the leucine residue at position 368 is replaced with an alanine residue (L368A) (numberings according to EU index). [0071] In yet a further embodiment, in the first subunit of the Fc domain additionally the serine residue at position 354 is replaced with a cysteine residue (S354C) or the glutamic acid residue at position 356 is replaced with a cysteine residue (E356C) (particularly the serine residue at position 354 is replaced with a cysteine residue), and in the second subunit of the Fc domain additionally the tyrosine residue at position 349 is replaced by a cysteine residue (Y349C) (numberings according to EU index). Introduction of these two cysteine residues results in formation of a disulfide bridge between the two subunits of the Fc domain, further stabilizing the dimer (Carter, J Immunol Methods 248, 7-15 (2001)). [0072] In a particular embodiment, the first subunit of the Fc domain comprises the amino acid substitutions S354C and T366W, and the second subunit of the Fc domain comprises the amino acid substitutions Y349C, T366S, L368A and Y407V (numbering according to EU index). C. Fc Heterodimerization Modifications [0073] In certain embodiments, multispecific antibodies described herein comprising paired mutations in the heavy chain constant domain (CH1) and light chain constant domain (CL) can further comprise an Fc region comprising a heterodimerization modifications that enhance pairing of heavy chains. In certain embodiments, multispecific antibodies described herein comprising paired mutations in the heavy chain constant domain (CH1) and light chain constant domain (CL) can further comprise an Fc region comprising heterodimerization modifications that enhance pairing of heavy chains and one or more amino acid substitutions described herein that increase or decrease Fc effector function. [0074] In certain embodiments, an IgG comprises two heavy chains, each chain comprising a set of mutations that enhance pairing of the two heavy chains. In some embodiments, an IgG comprises (i) a first heavy chain comprising the Fc mutations T371V, L372Y, F436A, and Y438V; and (ii) a second heavy chain comprising the Fc mutations T371V, T389L, K420L, and T422W, wherein the numbering is according to Kabat). D. Glycosylation Modifications [0075] In some embodiments, the glycosylation of a multispecific antibody described herein may be modified. For example, an aglycoslated antibody can be made (i.e., the antibody lacks glycosylation). Glycosylation can be altered to, for example, increase the affinity of the antibody Attorney Docket No.14681-WO-PCT for antigen. Such carbohydrate modifications can be accomplished by, for example, altering one or more sites of glycosylation within the antibody sequence. For example, one or more amino acid substitutions can be made that result in elimination of one or more variable region framework glycosylation sites to thereby eliminate glycosylation at that site. Such aglycosylation can increase the affinity of the antibody for antigen. Such an approach is described in further detail in U.S. Patent Nos.5,714,350 and 6,350,861 by Co et al. [0076] Glycosylation of the constant region on N297 (EU numbering) can be prevented by mutating the N297 residue to another residue, e.g., N297A, and/or by mutating an adjacent amino acid, e.g., 298 to thereby reduce glycosylation on N297. [_{0077] In some embodiments, the interaction of antibodies with Fc Rs can be enhanced by} modifying the glycan moiety attached to each Fc fragment at the N297 residue (EU numbering). In particular, the absence of core fucose residues strongly enhances ADCC via improved _{binding of IgG to activating Fc RIIIA without altering antigen binding or CDC. Natsume et al.} (2009) Drug Des. Devel. Ther.3 :7. There is convincing evidence that afucosylated tumor- specific antibodies translate into enhanced therapeutic activity in mouse models in vivo. Nimmerjahn & Ravetch (2005) Science 310: 1510; Mossner et a/. (2010) Blood 115:4393. [0078] Modification of antibody glycosylation can be accomplished by, for example, expressing the antibody in a host cell with altered glycosylation machinery. Cells with altered glycosylation machinery have been described in the art and can be used as host cells in which to express recombinant antibodies of this disclosure to thereby produce an antibody with altered glycosylation. For example, the cell lines Ms704, Ms705, and Ms709 lack the fucosyltransferase gene, FUT8 (a-(l,6) fucosyltransferase) (see U.S. Pat. App. Publication No.20040110704; Yamane-Ohnuki et al. (2004) Biotechnol. Bioeng.87: 614), such that antibodies expressed in these cell lines lack fucose on their carbohydrates. As another example, EP 1176195 also describes a cell line with a functionally disrupted FUT8 gene as well as cell lines that have little or no activity for adding fucose to the N-acetylglucosamine that binds to the Fc region of the antibody, for example, the rat myeloma cell line YB2/0 (ATCC CRL 1662). PCT Publication WO 03/035835 describes a variant CHO cell line, Lecl3, with reduced ability to attach fucose to Asn (297)-linked carbohydrates, also resulting in hypofucosylation of antibodies expressed in that host cell. See also Shields et al. (2002) J. Biol. Chem.277:26733. Antibodies with a modified glycosylation profile can also be produced in chicken eggs, as described in PCT Publication No. WO 2006/089231. Alternatively, antibodies with a modified glycosylation profile can be produced in plant cells, such as Lemna. See e.g. U.S. Publication No. Attorney Docket No.14681-WO-PCT 2012/0276086. PCT Publication No. WO 99/54342 describes cell lines engineered to express glycoprotein-modifying glycosyl transferases (e.g., beta(l,4)- N-acetylglucosaminyltransferase III (GnTIII)) such that antibodies expressed in the engineered cell lines exhibit increased bisecting GlcNac structures which results in increased ADCC activity of the antibodies. See also Umana et al. (1999) Nat. Biotech.17: 176. Alternatively, the fucose residues of the antibody may be cleaved off using a fucosidase enzyme. For example, the enzyme alpha-L-fucosidase removes fucosyl residues from antibodies. Tarentino et al. (1975) Biochem.14:5516. Core fucosylation may also be reduced by culturing antibody-producing cells in the presence of small molecule fucose analogs, such as those described at EP2282773B1, or in the presence of castanospermine, as described at WO 08/052030. [0079] Because nonfucosylated antibodies exhibit greatly enhanced ADCC and/or ADCP compared with fucosylated antibodies, antibody preparations need not be completely free of fucosylated heavy chains to be useful in the present invention. Residual levels of fucosylated heavy chains will not significantly interfere with the ADCC and/or ADCP activity of a preparation substantially of nonfucosylated heavy chains. Antibodies produced in conventional CHO cells, which are fully competent to add core fucose to N-glycans, may nevertheless comprise from a few percent up to 15% nonfucosylated antibodies. Nonfucosylated antibodies may exhibit ten-fold higher affinity, and up to 30- to 100-fold enhancement of ADCC and/or ADCP activity, so even a small increase in the proportion of nonfucosylated antibodies may drastically increase the ADCC and/or ADCP activity of a preparation. Any preparation comprising more nonfucosylated antibodies than would be produced in normal CHO cells in culture may exhibit some level of enhanced ADCC and/or ADCP. Such antibody preparations are referred to herein as preparations having reduced fucosylation. Depending on the original level of nonfucosylation obtained from normal CHO cells, reduced fucosylation preparations may comprise 80%, 70%, 60% 50%, 30%, 20%, 10% and even 5% nonfucosylated antibodies. Reduced fucosylation may be functionally defined as preparations exhibiting 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, two-fold, three-fold or greater enhancement of ADCC and/or ADCP compared with antibodies prepared in normal CHO cells, and not with reference to any fixed percentage of nonfucosylated species. [0080] The level of nonfucosylation may be structurally defined. As used herein, “nonfucosylated” or “afucosylated” (terms used synonymously) refers to antibody preparations in which over 95% of heavy chains lack fucose, such as over 96%, over 97%, over 98%, over 99%, or 100%. The term “hypofucosylated” refers to antibody preparations in which more than Attorney Docket No.14681-WO-PCT 80% and less than or equal to 95% heavy chains lack fucose, e.g. antibody preparations in which between 85 and 95%, between 80 and 85%, between 80 and 90%, between 85 and 90%, or between 90 and 95% of heavy chains lack fucose. The term “hypofucosylated or nonfucosylated” refers to antibody preparations in which 80% or more of heavy chains lack fucose. The term “reduced fucosylation” refers to antibody preparations in which between 10 and 80% of heavy chains lack fucose, such as 20-80%, 30-80%, 40-80%, 50-80%, 60-80%, 70- 80%, 20-70%, 30-70%, 40-70%, 50-70%, 60-70%, 20-60%, 30-60%, 40-60%, 50-60%, 20-50%, 30- 50%, 40-50%, 20-40%, 30-40%, 10-20%, 10-30%, or 20-30%. E. Multispecific Formats [0081] Multispecific antibodies provided herein can have various formats. In certain embodiments, a multispecific antibody is bispecific and capable of specifically binding to at least two distinct antigenic determinants. [0082] In certain embodiments, a multispecific antibody provided herein is in a 1+1 bispecific format comprising two binding sites each formed by a pair of an antibody heavy chain variable domain (VH) and an antibody light chain variable domain (VL) binding to different antigens or to different epitopes on the same antigen. In certain aspects, the 1+1 bispecific format comprises two Fab regions that bind to different antigens or to different epitopes on the same antigen. [0083] In certain embodiments, a multispecific antibody provided herein is in a 2+1 bispecific format comprising two binding sites for a first antigen or epitope and one binding site for a second antigen or epitope. In certain aspects, the 2+1 bispecific format comprises three Fab regions, wherein two Fab regions bind to the same first antigen and the third Fab region binds to a second antigen that is different from the first antigen. In certain aspects, the two Fab regions that bind to the same first antigen bind to the same epitope. In certain aspects, the two Fab regions that bind to the same first antigen bind to different epitopes. [0084] In certain embodiments, a multispecific antibody provided herein is in a 2+2 bispecific format comprising two binding sites for a first antigen or epitope and two binding sites for a second antigen or epitope). In certain aspects, the 2+2 bispecific format comprises four Fab regions. In certain aspects, two Fab regions bind to the same first antigen and the other two Fab regions both bind to the same second antigen wherein the first and second antigen are different. In certain aspects, two Fab regions that bind to the same first antigen bind to the same Attorney Docket No.14681-WO-PCT epitopes of the first antigen. In certain aspects, two Fab regions that bind to the same first antigen bind to different epitopes of the first antigen. [0085] In certain embodiments, a multispecific anitbody, such as a bispecific antibody, can have a 1+1, 2+1, or 2+2 format described herein and comprise any of the CL, CH1, or CH3 mutations described herein. [0086] In certain embodiments, a multispecific antibody having a 1+1 format can comprise two Fab regions, wherein the first Fab region comprises (i) a CL domain comprising a S176K and N137K or N137R mutation and (ii) a CH1 domain comprising a S183D and T187D mutation; and wherein the second Fab region comprises (i) a CL domain comprising a S176D and N137D mutation and (ii) a CH1 domain comprising a S183K and T187K mutation; wherein the residues are numbered according to EU. In certain embodiments, the antibody having the 1+1 format comprises any Fc region mutation described herein. In certain aspects, the Fc region comprises a mutation in one or both CH2 domains wherein the mutation is L234A/L235A/D265S (AAS); L234A/L235A/P329G (LALAPG); S298A/E333A/K334A (AAA); N297K; M252Y/S254T/T256E (YTE); G236A/S239D/A330L/I332E (GASDALIE); G236A; S239D/I332E (DE); S239D/A330L/I332E (DLE); G236A/S239D/I332E (ADE); G236A/A330L/I332E (GAALIE); F243L/R292P/Y300L/V305I/P396L (LPLIL); L235V/F243L/R292P/Y300L/P396L (VLPLL); P238K/L235E/P238K; L235E/P238K/K322A; L234A/L235E/G237A; L234F/L235E/P331S (FES); L234F/L235Q/K322Q (FQQ); A330S/P331S; L234A/G237A; L234A/L235A/G237A; L234A/L235A/G237A/P238S/H268A/A330S/P331S; L234A/L235E; G236R/L328R; or L234A/L235A/K322A. In certain aspects, the Fc region comprises the mutations Y349C, T366S, L368A, Y407V (Hole mutations) in a first CH3 domain, and the mutations S354C and T366W (Knob mutations) in a second CH3 domain. In certain aspects, the Hole mutations and Knob mutations are disulfide stabilized. [0087] In certain embodiments, a multispecific antibody having a 2+1 format can comprise three Fab regions, wherein: the first Fab region is linked to the second Fab region in the N-terminal to C-terminal direction; the second Fab region is linked to a first CH2 domain in the N-terminal to C-terminal direction, which is linked to a first CH3 domain; the third Fab region is linked to a second CH2 domain in the N-terminal to C- terminal direction, which is linked to a second CH3 domain; Attorney Docket No.14681-WO-PCT the first Fab region and the third Fab region each comprises a variable region that binds to the same antigen, optionally wherein the variable regions are identical; the second Fab region comprises a variable region that binds to a different antigen than the first Fab region and the third Fab region; the first Fab region and the third Fab region each comprises a CH1 domain comprising the amino acid substitutions S188D and T192D; the first Fab region and the third Fab region each comprises a CL domain comprising the amino acid substitutions S176K and N137K; the second Fab region comprises a CH1 domain comprising the amino acid substitutions S188K and T192K; and the second Fab region comprises a CL domain comprising the amino acid substitutions S176D and N137D. [0088] In certain embodiments, the antibody having the 2+1 format comprises any Fc region mutation described herein. [0089] In certain embodiments, a multispecific antibody having a 2+1 format can comprise three Fab regions, wherein: the first Fab region is linked to the second Fab region in the N-terminal to C-terminal direction; the second Fab region is linked to a first CH2 domain in the N-terminal to C-terminal direction, which is linked to a first CH3 domain; the third Fab region is linked to a second CH2 domain in the N-terminal to C- terminal direction, which is linked to a second CH3 domain; the first Fab region and the third Fab region each comprises a variable region that binds to the same antigen, optionally wherein the variable regions are identical; the second Fab region comprises a variable region that binds to a different antigen than the first Fab region and the third Fab region; the first Fab region and the third Fab region each comprises a CH1 domain comprising the amino acid substitutions S188D and T192D; the first Fab region and the third Fab region each comprises a CL domain comprising the amino acid substitutions S176K and N137K; the second Fab region comprises a CH1 domain comprising the amino acid substitutions S188K and T192K; Attorney Docket No.14681-WO-PCT the second Fab region comprises a CL domain comprising the amino acid substitutions S176D and N137D; the first CH2 domain and/or the second CH2 domain each comprises the mutations L234A/L235A/D265S (AAS); L234A/L235A/P329G (LALAPG); S298A/E333A/K334A (AAA); N297K; M252Y/S254T/T256E (YTE); G236A/S239D/A330L/I332E (GASDALIE); G236A; S239D/I332E (DE); S239D/A330L/I332E (DLE); G236A/S239D/I332E (ADE); G236A/A330L/I332E (GAALIE); F243L/R292P/Y300L/V305I/P396L (LPLIL); L235V/F243L/R292P/Y300L/P396L (VLPLL); P238K/L235E/P238K; L235E/P238K/K322A; L234A/L235E/G237A; L234F/L235E/P331S (FES); L234F/L235Q/K322Q (FQQ); A330S/P331S; L234A/G237A; L234A/L235A/G237A; L234A/L235A/G237A/P238S/H268A/A330S/P331S; L234A/L235E; G236R/L328R; or L234A/L235A/K322A. [0090] In certain embodiments, a multispecific antibody having a 2+1 format can comprise three Fab regions, wherein: the first Fab region is linked to the second Fab region in the N-terminal to C-terminal direction; the second Fab region is linked to a first CH2 domain in the N-terminal to C-terminal direction, which is linked to a first CH3 domain; the third Fab region is linked to a second CH2 domain in the N-terminal to C- terminal direction, which is linked to a second CH3 domain; the first Fab region and the third Fab region each comprises a variable region that binds to the same antigen, optionally wherein the variable regions are identical; the second Fab region comprises a variable region that binds to a different antigen than the first Fab region and the third Fab region; the first Fab region and the third Fab region each comprises a CH1 domain comprising the amino acid substitutions S188D and T192D; the first Fab region and the third Fab region each comprises a CL domain comprising the amino acid substitutions S176K and N137K; the second Fab region comprises a CH1 domain comprising the amino acid substitutions S188K and T192K; the second Fab region comprises a CL domain comprising the amino acid substitutions S176D and N137D; Attorney Docket No.14681-WO-PCT the first CH2 domain and/or the second CH2 domain each comprises the mutations L234A/L235A/D265S (AAS); L234A/L235A/P329G (LALAPG); S298A/E333A/K334A (AAA); N297K; M252Y/S254T/T256E (YTE); G236A/S239D/A330L/I332E (GASDALIE); G236A; S239D/I332E (DE); S239D/A330L/I332E (DLE); G236A/S239D/I332E (ADE); G236A/A330L/I332E (GAALIE); F243L/R292P/Y300L/V305I/P396L (LPLIL); L235V/F243L/R292P/Y300L/P396L (VLPLL); P238K/L235E/P238K; L235E/P238K/K322A; L234A/L235E/G237A; L234F/L235E/P331S (FES); L234F/L235Q/K322Q (FQQ); A330S/P331S; L234A/G237A; L234A/L235A/G237A; L234A/L235A/G237A/P238S/H268A/A330S/P331S; L234A/L235E; G236R/L328R; or L234A/L235A/K322A; the first CH3 domain comprises S354C and T366W mutations (Knob mutations), optionally wherein the Knob mutations are disulfide stabilized; the second CH3 domain comprises Y349C, T366S, L368A, and Y407V mutations (Hole mutations), optionally wherein the Hole mutations are disulfide stabilized. [0091] In certain embodiments, a multispecific antibody having a 2+1 format can comprise a first heavy chain, a first light chain, a second heavy chain, and a second light chain, wherein: the first heavy chain comprises in the N-terminal to C-terminal direction a first VH domain, a first CH1 domain, a second VH domain, a second CH1 domain, a first CH2 domain, and a first CH3 domain; the first light chain comprises in the N-terminal to C-terminal direction a first VL domain, a first CL domain, a second VL domain, and a second CL domain; the second heavy chain comprises in the N-terminal to C-terminal direction a third VH domain, a third CH1 domain, a second CH2 domain, and a second CH3 domain; the second light chain comprises in the N-terminal to C-terminal direction a third VL domain and a third CL domain; the first CH1 domain and the third CH1 domain each comprises the amino acid substitutions S188D and T192D; the second CH1 domain comprises the amino acid substitutions S188K and T192K; the first CL domain and the third CL domain each comprises the amino acid substitutions S176K and N137K; and the second CH1 domain comprises the amino acid substitutions S176D and N137D. [0092] In certain embodiments, a multispecific antibody having a 2+1 format can comprise a first heavy chain, a first light chain, a second heavy chain, and a second light chain, wherein: Attorney Docket No.14681-WO-PCT the first heavy chain comprises in the N-terminal to C-terminal direction a first VH domain, a first CH1 domain, a second VH domain, a second CH1 domain, a first CH2 domain, and a first CH3 domain; the first light chain comprises in the N-terminal to C-terminal direction a first VL domain, a first CL domain, a second VL domain, and a second CL domain; the second heavy chain comprises in the N-terminal to C-terminal direction a third VH domain, a third CH1 domain, a second CH2 domain, and a second CH3 domain; the second light chain comprises in the N-terminal to C-terminal direction a third VL domain and a third CL domain; the first CH1 domain and the third CH1 domain each comprises the amino acid substitutions S188D and T192D; the second CH1 domain comprises the amino acid substitutions S188K and T192K; the first CL domain and the third CL domain each comprises the amino acid substitutions S176K and N137K; the second CH1 domain comprises the amino acid substitutions S176D and N137D; the first VH domain and the third VH domain are the same; the first VL domain and the third VL domain are the same; the second VH domain is different from the first VH domain and the third VH domain; and the second VL domain is different from the first VL domain and the third VL domain. [0093] In certain embodiments, a multispecific antibody having a 2+1 format can comprise a first heavy chain, a first light chain, a second heavy chain, and a second light chain, wherein: the first heavy chain comprises in the N-terminal to C-terminal direction a first VH domain, a first CH1 domain, a second VH domain, a second CH1 domain, a first CH2 domain, and a first CH3 domain; the first light chain comprises in the N-terminal to C-terminal direction a first VL domain, a first CL domain, a second VL domain, and a second CL domain; the second heavy chain comprises in the N-terminal to C-terminal direction a third VH domain, a third CH1 domain, a second CH2 domain, and a second CH3 domain; the second light chain comprises in the N-terminal to C-terminal direction a third VL domain and a third CL domain; Attorney Docket No.14681-WO-PCT the first CH1 domain and the third CH1 domain each comprises the amino acid substitutions S188D and T192D; the second CH1 domain comprises the amino acid substitutions S188K and T192K; the first CL domain and the third CL domain each comprises the amino acid substitutions S176K and N137K; the second CH1 domain comprises the amino acid substitutions S176D and N137D; the first VH domain and the third VH domain are the same; the first VL domain and the third VL domain are the same; the second VH domain is different from the first VH domain and the third VH domain; the second VL domain is different from the first VL domain and the third VL domain; and the first CH2 domain and/or the second CH2 domain each comprises the mutations L234A/L235A/D265S (AAS); L234A/L235A/P329G (LALAPG); S298A/E333A/K334A (AAA); N297K; M252Y/S254T/T256E (YTE); G236A/S239D/A330L/I332E (GASDALIE); G236A; S239D/I332E (DE); S239D/A330L/I332E (DLE); G236A/S239D/I332E (ADE); G236A/A330L/I332E (GAALIE); F243L/R292P/Y300L/V305I/P396L (LPLIL); L235V/F243L/R292P/Y300L/P396L (VLPLL); P238K/L235E/P238K; L235E/P238K/K322A; L234A/L235E/G237A; L234F/L235E/P331S (FES); L234F/L235Q/K322Q (FQQ); A330S/P331S; L234A/G237A; L234A/L235A/G237A; L234A/L235A/G237A/P238S/H268A/A330S/P331S; L234A/L235E; G236R/L328R; or L234A/L235A/K322A. [0094] In certain embodiments, a multispecific antibody having a 2+1 format can comprise a first heavy chain, a first light chain, a second heavy chain, and a second light chain, wherein: the first heavy chain comprises in the N-terminal to C-terminal direction a first VH domain, a first CH1 domain, a second VH domain, a second CH1 domain, a first CH2 domain, and a first CH3 domain; the first light chain comprises in the N-terminal to C-terminal direction a first VL domain, a first CL domain, a second VL domain, and a second CL domain; the second heavy chain comprises in the N-terminal to C-terminal direction a third VH domain, a third CH1 domain, a second CH2 domain, and a second CH3 domain; the second light chain comprises in the N-terminal to C-terminal direction a third VL domain and a third CL domain; Attorney Docket No.14681-WO-PCT the first CH1 domain and the third CH1 domain each comprises the amino acid substitutions S188D and T192D; the second CH1 domain comprises the amino acid substitutions S188K and T192K; the first CL domain and the third CL domain each comprises the amino acid substitutions S176K and N137K; the second CH1 domain comprises the amino acid substitutions S176D and N137D; the first VH domain and the third VH domain are the same; the first VL domain and the third VL domain are the same; the second VH domain is different from the first VH domain and the third VH domain; the second VL domain is different from the first VL domain and the third VL domain; the first CH2 domain and/or the second CH2 domain each comprises the mutations L234A/L235A/D265S (AAS); L234A/L235A/P329G (LALAPG); S298A/E333A/K334A (AAA); N297K; M252Y/S254T/T256E (YTE); G236A/S239D/A330L/I332E (GASDALIE); G236A; S239D/I332E (DE); S239D/A330L/I332E (DLE); G236A/S239D/I332E (ADE); G236A/A330L/I332E (GAALIE); F243L/R292P/Y300L/V305I/P396L (LPLIL); L235V/F243L/R292P/Y300L/P396L (VLPLL); P238K/L235E/P238K; L235E/P238K/K322A; L234A/L235E/G237A; L234F/L235E/P331S (FES); L234F/L235Q/K322Q (FQQ); A330S/P331S; L234A/G237A; L234A/L235A/G237A; L234A/L235A/G237A/P238S/H268A/A330S/P331S; L234A/L235E; G236R/L328R; or L234A/L235A/K322A; the first CH3 domain comprises S354C and T366W mutations (Knob mutations), optionally wherein the Knob mutations are disulfide stabilized; and the second CH3 domain comprises Y349C, T366S, L368A, and Y407V mutations (Hole mutations), optionally wherein the Hole mutations are disulfide stabilized. [0095] In certain embodiments, a multispecific antibody has a 2+2 format. It will be understood that a fourth Fab region can be added to any of the 2+1 format embodiments described herein to arrive at a 2+2 format. It will be understood that any of the Fab regions described herein can be combined together in various multispecific formats (e.g., 1+1+1, 1+1+1+1, 3+1, etc.). It will be understood that an scFv or a single domain antibody fragment Attorney Docket No.14681-WO-PCT can be used in combination with a Fab described herein for any format (e.g., 1+1+1, 1+1+1+1, 3+1, etc.). III. EXAMPLES [0096] The following examples are included for illustrative purposes only and are not intended to limit the scope of the invention. Example 1 Design of Bispecific IgG Antibodies have CH1, CL, and Fc Mutations [0097] Two bispecific IgG antibodies (Antibody 1 and Antibody 2) were designed having mutations in the CL domains, CH1 domains, and CH3 domains as depicted in FIG.1. Both bispecific antibodies had a 1+1 format with one Fab targeting CD3 and the other Fab targeting ROR1. The VH and VL sequences were the same in Antibody 1 and Antibody 2. [0098] CD3 Fab: the CL domain of the CD3 Fab in Antibody 1 had S176K and N137K mutations, whereas the CL domain of the CD3 Fab in Antibody 2 had S176K and N137R mutations. The CH1 domain of the CD3 Fab in Antibody 1 and Antibody 2 had S183D and T187D mutations. Numbering is according to EU. [0099] ROR1 Fab: Antibody 1 and Antibody 2 had the same mutations in the CL domain and CH1 domain of the ROR1 Fab. The CL mutations were S176D and N137D. The CH1 mutations were S183K and T187K. Numbering is according to EU. [0100] Fc region: Antibody 1 and Antibody 2 had the same mutations in the CH3 domains of the Fc region. The mutations were T371V/L372Y/F436A/Y438V in the CH3 domain of the CD3 heavy chain and T371V/T389L/K420L/T422W in the CH3 domain of the ROR1 heavy chain. Numbering is according to Kabat. Example 2 Expression and Purification of Bispecific IgG Antibodies have CH1, CL, and Fc Mutations [0101] Antibody 1 and Antibody 2 were each expressed by transient transfection of Expi- CHO cells at a 1:1:1:1 (LC1:HC1:HC2:LC2) DNA ratio in 1L volume. The titer range for each antibody was approximately 100-200 mg/L. [0102] The antibodies were purified by Protein A chromatography followed by Mono S cation exchange chromatography for polishing. FIG.2 shows a protein gel of the two antibodies after Protein A chromatography. FIG.3 shows the chromatograms of the Mono S polishing. FIG.4 shows a protein gel of the two antibodies after Mono S polishing. The purification results are summarized in Table E1 below. Attorney Docket No.14681-WO-PCT Final Volume % Monomer Antibody Final mg/mL Final Total mg (mL) Intact MS Ab 1 1.53 9 13.77 95 Ab 2 2.09 9 18.81 95 [0103] The mass of both Antibody 1 and Antibody 2 were confirmed as full length antibody species deglycosylated. Approximately 5% of Heavy Chain 2/Light Chain 2 observed. Both antibodies were similar in expression and purification. Highly pure bispecific antibodies with % monomer of ~95% were obtained. Light chain mispairing was not detected with Antibody 1 or Antibody 2. Example 3 T-cell Killing Activity of ROR1xCD3 Bispecific Antibodies [0104] Antibody 1 and Antibody 2 were tested for functional cell killing activity in a human T cell killing assay. A549 cancer cells (target) and human T cells (effector) were co-cultured at a 1:4 target to effector ratio for 4 days in the presence of Antibody 1, Antibody 2, a positive control bispecific antibody that does not have the same CH1 and CL mutations as Antibody 1 or Antibody 2, negative control antibodies, or tumor plus T cells without antibody. As shown in FIG.5, Antibody 1 and Antibody 2 demonstrated similar tumor killing activity as the positive control antibody. Example 4 Comparison of Bispecific IgG Antibodies in 1+1 format having different sets of CH1 and CL pairing mutations and Fc Mutations [0105] Eight bispecific IgG antibodies (Antibody 3 to 10) were designed having mutations in the CL domains, CH1 domains, and CH3 domains. All bispecific antibodies had a 1+1 format with one Fab targeting Tumor Antigen 1 (TAA1) and the other Fab targeting Tumor Antigen 2 (TAA2). The eight antibodies represented four pairs with each pair having a distinct Variable Region (VR) relative to the other pairs, and each antibody of a pair having either Mutation Set 1 or Mutation Set 2. This grouping allowed comparison of Mutation Set 1 and Mutation Set 2 in the context of four different variable regions. [0106] Mutation Set 1 (Antibodies 3, 5, 7, & 9) CL chain 1 (human kappa) having the following mutations (EU numbering): F116A, Q124R, L135V, T178R Attorney Docket No.14681-WO-PCT CL chain 2 (human kappa) having the following mutations (EU numbering): Q124E, L135W, Q160E, T180E IgG1 heavy chain 1 having the following mutations: (EU= A141W, L145E, K147T, Q175E) + L234A + L235A + D265S + KEMA mixed allotype IgG1 heavy chain 2 having the following mutations: (EU: Q124E, L135W, Q160E, T180E) [0107] Mutation Set 2 (knob-in-hole) (Antibodies 4, 6, 8, & 10) CL chain 1 (human kappa) having S176K and N137K mutations; CL chain 2 (human kappa) having S176D and N137D mutations; IgG1 heavy chain 1 having the following mutations: S188D + T192D in CH1; KEMA mixed allotype (K214, E356, M358, A431) with L234A, L235A, D265S mutations for low effector function; and Y349C, T366S, L368A, Y407V disulfide-stabilized Hole mutations; IgG1 heavy chain 2 having the following mutations: S188K + T192K in CH1; KEMA mixed allotype (K214, E356, M358, A431) with L234A, L235A, D265S mutations for low effector function; S354C, T366W disulfide-stabilized Knob mutations [0108] The bispecific antibodies were transiently transfected in CHO1 cells and purified by Protein A chromatography (PrismA) and characterized by mass spectrometry. In the matched VH/VL pairs comparing the two different mutation sets, Mutation Set 1 exhibited greater Protein A yield in 3 of 4 pairs (FIG.6). However, mass spec data demonstrated greater purity for Mutation Set 2 (FIG.7). 3 of 4 pairs having Mutation Set 2 were 100% heterodimer, whereas 0 of 4 pairs having Mutation Set 1 were 100% heterodimer. Antibody 5 (Mutation Set 1) had 56% LC1-HC2 mispairing. The antibodies were further evaluated by Analytical Size Exclusion Chromatography (AnSEC). Atypical high molecular species were observed in (2/4) antibodies having Mutation Set 1 compared to (1/4) antibodies having Mutation Set 2 (FIG.8). Pairwise, antibodies having Mutation Set 1 had greater typical high molecular weight species than antibodies having Mutation Set 2. Antibodies having Mutation Set 2 had superior quality by AnSEC relative to antibodies having Mutation Set 1 in all four variable region pairs. [0109] The bispecific antibodies were also stably transfected into CHO1 cells and evaluated for recovery. As shown in FIG.9, CHO1 cells stably transfected to express antibodies having Mutation Set 2 showed a tendency to recover faster than CHO1 cells stably transfected to express antibodies having Mutation Set 1. Two pools of cells stably expressing antibodies having Mutation Set 1 did not recover. Attorney Docket No.14681-WO-PCT Example 5 Expression and Purification of 1+1 and 2+1 format Bispecific Antibodies having CH1 and CL pairing mutations [0110] Bispecific antibodies in the 1+1 Fab format or 2+1 Fab format having CH1/CL Pairing Mutation Set 1 or Pairing Mutation Set 2 were expressed and purified. Pairing Mutation Set 1: CL chain 1 (human kappa) having S176K and N137K mutations; CL chain 2 (human kappa) having S176D and N137D mutations; CH1 chain 1 having S188D and T192D mutations; and CH1 chain 2 having S188K and T192K mutations [0112] Pairing Mutation Set 2: CL chain 1 (human kappa) having S176K and N137R mutations; CL chain 2 (human kappa) having S176D and N137D mutations; CH1 chain 1 having S188D and T192D mutations; and CH1 chain 2 having S188K and T192K mutations [0113] The antibodies were purified by Protein A chromatography (PrismA) and a 5 ml/min isocratic elution. The PrismA column was equilibrated in PBS (5 column volumes), sample was injected at a rate of 5 mL/min, washed in PBS + 0.1% Triton-X (5 column volumnes), washed in PBS (5 column volumes), and eluted with 50mM citrate pH 3.0 isocratic (6 column volumes). 2mL fractions were collected and neutralized with 1:101M sodium acetate pH5.2. The yields obtained are reported in Table E2. Table E2 Pairing % Yie C^{hain Tit} Volume Expected Total ld Antibody Targets Mutation ^er (_mg/mL) (L) Total m from Set g mg PrismA 11 ROR1xROR1 1 1+1 71.7 0.5 35.8 29.12 81 12 ROR1xROR1 2 1+1 114.5 0.5 57.3 48.8 85 13 FCRH5xCD3 1 1+1 33.1 0.5 16.6 9.04 55 14 FCRH5xCD3 2 1+1 32.3 0.5 16.2 8.24 51 15 ROR1xCD3 1 1+2 48.7 0.5 24.3 19.44 80 16 ROR1xCD3 2 1+2 68.4 0.5 34.2 30.4 89 [0114] Antibodies were polished by Jetted SP35 IEX chromatography. A 5mL column was loaded with the PrismA pool diluted 1:10 with CX-1 Buffer A, equilibrated with 5 column volumes of Buffer A, washed with 5 column volumes of Buffer A, and eluted with 35 column Attorney Docket No.14681-WO-PCT volumes of Buffer B gradient 1-100 then 5 column volumes of 100% isocratic Buffer B. 2mL fractions were collected. Protein Gels from the PrismA step and Jetted SP35 IEX polishing step are shown in FIG.10. Next, the antibodies were further polished by Hydrophobic interaction chromatography (HIC). Sample was diluted 1:10 with HIC buffer A (50mM sodium phosphate pH 5.8, 1.5M ammonium Sulfate). A 5 mL HIC phenyl column was equilibrated with 5 column volumes (CV) of HIC buffer A. Sample was run through the column, washed with 5 CV of HIC Buffer A, then eluted with a 0-100% gradient of HIC Buffer B (50mM sodium phosphate pH 5.8) in 35 CV, then 5 CV of HIC Buffer B.2mL fractions were collected. The yields obtained after Protein A, Jetted SP35 IEX polishing, and HIC polishing are reported in Table E3. Table E3 Pairing on _Chain ^Fi Final Final Final Antibody Targets Mutati ^nal A₂₈₀ ( Volume Total Set mg/mL) (mL) mg 11 ROR1xROR1 1 1+1 2.22 1.38 5.7 7.88 12 ROR1xROR1 2 1+1 5.77 3.59 6.7 24 13 FCRH5xCD3 1 1+1 0.59 0.38 4.7 1.77 14 FCRH5xCD3 2 1+1 0.38 0.242 10.8 2.62 15 ROR1xCD3 1 1+2 1.15 0.72 9.8 7.04 16 ROR1xCD3 2 1+2 2.93 1.73 4 8.974 [0115] The antibodies were analyzed by mass spectrometry. The mass was confirmed as full length deglycosylated target antibody. No other impurity was observed. Example 6 Comparison of Bispecific IgG Antibodies in 2+1 format having different sets of CH1 and CL pairing mutations and Fc Mutations [0116] Antibody 17 Light Chain 1 (Target A) o CL mutations: S176K and N137K Light chain 2 (Target A and Target B-VR1) o CL mutations (Target A Fab region): S176K and N137K o CL mutations (Target B-VR1 Fab region): S176D and N137D Heavy Chain 1 (Target A) o CH1 mutations: S188D and T192D o Fc mutations KEMA mixed allotype (K214, E356, M358, A431) Attorney Docket No.14681-WO-PCT L234A, L235A, D265S Y349C, T366S, L368A, Y407V disulfide-stabilized Hole mutations Heavy Chain 2 (Target A and Target B-VR1) o CH1 mutations (Target A Fab region) S188D and T192D o CH1 mutations (Target B-VR1 Fab region) S188K and T192K o Fc mutations KEMA mixed allotype (K214, E356, M358, A431) L234A, L235A, D265S S354C, T366W disulfide-stabilized Knob mutations [0117] Antibody 18 Light Chain 1 (Target A) o CL mutations: S176K and N137K Light chain 2 (Target A and Target B-VR2) o CL mutations (Target A Fab region): S176K and N137K o CL mutations (Target B-VR2 Fab region): S176D and N137D Heavy Chain 1 (Target A) o CH1 mutations: S188D and T192D o Fc mutations KEMA mixed allotype (K214, E356, M358, A431) L234A, L235A, D265S Y349C, T366S, L368A, Y407V disulfide-stabilized Hole mutations Heavy Chain 2 (Target A and Target B-VR2) o CH1 mutations (Target A Fab region) S188D and T192D o CH1 mutations (Target B-VR2 Fab region) S188K and T192K o Fc mutations KEMA mixed allotype (K214, E356, M358, A431) Attorney Docket No.14681-WO-PCT L234A, L235A, D265S S354C, T366W disulfide-stabilized Knob mutations [0118] Antibody 19 Light Chain 1 (Target A) o CL mutations: F116A, Q124R, L135V, T178R (EU numbering) Light chain 2 (Target A and Target B-VR1) o CL mutations (Target A Fab region): F116A, Q124R, L135V, T178R (EU numbering) o CL mutations (Target B-VR1 Fab region): Q124E, L135W, Q160E, T180E Heavy Chain 1 (Target A) o CH1 mutations: A141W, L145E, K147T, Q175E o Fc mutations KEMA mixed allotype (K214, E356, M358, A431) L234A, L235A, D265S T371V, L372Y, F436A, Y438V Heavy Chain 2 (Target A and Target B-VR1) o CH1 mutations (Target A Fab region): A141W, L145E, K147T, Q175E (EU numbering) o CH1 mutations (Target B-VR1 Fab region): Q175K (EU numbering) o Fc mutations KEMA mixed allotype (K214, E356, M358, A431) L234A, L235A, D265S T371V, T389L, K420L, T422W [0119] Antibody 20 Light Chain 1 (Target A) o CL mutations: F116A, Q124R, L135V, T178R (EU numbering) Light chain 2 (Target A and Target B-VR3) o CL mutations (Target A Fab region): F116A, Q124R, L135V, T178R (EU numbering) Attorney Docket No.14681-WO-PCT o CL mutations (Target B-VR3 Fab region): Q124E, L135W, Q160E, T180E Heavy Chain 1 (Target A) o CH1 mutations: A141W, L145E, K147T, Q175E o Fc mutations KEMA mixed allotype (K214, E356, M358, A431) L234A, L235A, D265S T371V, L372Y, F436A, Y438V Heavy Chain 2 (Target A and Target B-VR3) o CH1 mutations (Target A Fab region): A141W, L145E, K147T, Q175E (EU numbering) o CH1 mutations (Target B-VR3 Fab region): Q175K (EU numbering) o Fc mutations KEMA mixed allotype (K214, E356, M358, A431) L234A, L235A, D265S T371V, T389L, K420L, T422W

Claims

Attorney Docket No.14681-WO-PCT CLAIMS WHAT IS CLAIMED: 1. A multispecific antibody comprising: a first Fab region comprising a VH domain, a VL domain, a CL domain, and a CH1 domain, wherein the CL domain comprises a S176K and N137K or N137R mutation, and the CH1 domain comprises a S188D and T192D mutation; a second Fab region comprising a VH domain, a VL domain, a CL domain, and a CH1 domain, wherein the CL domain comprises a S176D and N137D mutation and the CH1 domain comprises a S188K and T192K mutation; and a Fc region comprising a first CH2 domain, a second CH2 domain, a first CH3 domain, and a second CH3 domain, wherein the Fc region comprises one or more mutations for enhancing or reducing Fc effector function, wherein the residues are numbered according to Kabat. 2. The multispecific antibody of claim 1, wherein the multispecific antibody comprises: a first heavy chain comprising from N-terminus to C-terminus: the VH domain and the CH1 domain of the first Fab region, the first CH2 domain, and the first CH3 domain; a second heavy chain comprising from N to C terminus: the VH domain and the CH1 domain of the second Fab region, the second CH2 domain, and the second CH3 domain; a first light chain comprising from N to C terminus: the VL domain and the CL domain of the first Fab region; and a second light chain comprising from N to C terminus: the VL domain and the CL domain of the second Fab region. 3. The multispecific antibody of claim 1 or 2, wherein the first Fab region and second Fab region bind to different antigens. 4. The multispecific antibody of claims 1 or 2, wherein the first Fab region and second Fab region bind to different epitopes of the same antigen. 5. The multispecific antibody of any of claims 1-4, wherein the multispecific antibody further comprises a third Fab region comprising a VH domain, a VL domain, a CL Attorney Docket No.14681-WO-PCT domain, and a CH1 domain, and wherein the CL domain and CH1 domain has the same mutations as the CL domain and CH1 domain of the first Fab region or the same mutations as the CL domain and CH1 domain of the second Fab region. 6. The multispecific antibody of claim 5, wherein the third Fab region binds to the same antigen as the first Fab region. 7. The multispecific antibody of claim 6, wherein the third Fab region binds to the same epitope as the first Fab region. 8. The multispecific antibody of claim 7, wherein the sequences of the VH domains of the third Fab region and the first Fab region are identical and the sequences of the VL domains of the third Fab region and the first Fab region are identical. 9. The multispecific antibody of claim 6, wherein the third Fab region binds to a different epitope of the same antigen as the first Fab region. 10. The multispecific antibody of claim 5, wherein the third Fab region binds to the same antigen as the second Fab region. 11. The multispecific antibody of claim 10, wherein the third Fab region binds to the same epitope as the second Fab region. 12. The multispecific antibody of claim 11, wherein the sequences of the VH domains of the third Fab region and the second Fab region are identical and the sequences of the VL domains of the third Fab region and the second Fab region are identical. 13. The multispecific antibody of claim 10, wherein the third Fab region binds to a different epitope of the same antigen as the second Fab region. 14. The multispecific antibody of any of claims 5-13, wherein the third Fab region is covalently linked to the first Fab region. Attorney Docket No.14681-WO-PCT 15. The multispecific antibody of claim 14, wherein the CH1 domain of the third Fab region is covalently linked to the VH domain of the first Fab region. 16. The multispecific antibody of any of claims 5-13, wherein the third Fab region is covalently linked to the second Fab region. 17. The multispecific antibody of claim 16, wherein the CH1 domain of the third Fab region is covalently linked to the VH domain of the second Fab region. 18. The multispecific antibody of any of claims 14-16, wherein the third Fab region is covalently linked to the VH domain by a peptide linker comprising (G3S)x wherein x is 1-5, (G4S)x wherein x is 1-5, GGGGSGGGGS, GGGS or GGGGS. 19. The multispecific antibody of any of claims, wherein the Fc region comprises knob-in-hole mutations. 20. The multispecific antibody of claim 19, wherein the knob-in-hole mutations comprise T366Y in one CH3 domain (knob domain) and Y407T in the other CH3 domain (hole domain). 21. The multispecific antibody of claim 19, wherein the knob-in-hole mutations comprise T366W in one CH3 domain (knob domain) and T366S, L368A, and Y407V in the other CH3 domain (hole domain). 22. The multispecific antibody of any of claims 19-21, wherein the Fc region further comprises an engineered disulfide bond between the CH3 domains near the knob-in-hole mutations. 23. The multispecific antibody of claim 22, wherein the disulfide bond is between S354C in one CH3 domain (knob domain) and Y349C in the other CH3 domain (hole domain). 24. The multispecific antibody of any of claims 1-23, wherein the mutations for enhancing or reducing Fc effector function comprise one or more of the following: L234A, L235A, and D265S (AAS); Attorney Docket No.14681-WO-PCT L234A, L235A, and P329G (LALAPG); S298A, E333A, and K334A (AAA); N297K; M428L and N434S (LS); M252Y, S254T, and T256E (YTE); G236A, S239D, A330L, and I332E (GASDALIE); G236A; S239D and I332E (DE); S239D, A330L, and I332E (DLE); G236A, S239D, and I332E (ADE); G236A, A330L, and I332E (GAALIE); F243L, R292P, Y300L, V305I, P396L (LPLIL); L235V, F243L, R292P, Y300L, P396L (VLPLL); P238K; L235E and P238K; L235E, P238K, and K322A; L234A, L235E and G237A; L234F, L235E, and P331S (FES); L234F, L235Q, and K322Q (FQQ); A330S, and P331S; L234A and G237A; L234A, L235A, and G237A; L234A, L235A, G237A, P238S, H268A, A330S, and P331S; L234A and L235E; G236R and L328R; or L234A, L235A, and K322A 25. The multispecific antibody of any of claims 1-24, wherein the mutation for reducing Fc effector function reduces antibody-dependent cellular cytotoxicity (ADCC), antibody-dependent cellular phagocytosis (ADCP), Fc receptor binding, complement-dependent cytotoxicity (CDC), or C1q binding. Attorney Docket No.14681-WO-PCT 2_{6. The multispecific antibody of claim 25, wherein the Fc receptor is a Fc receptor} or neonatal Fc receptor (FcRn). 2_{7. The multispecific antibody of claim 26, wherein the Fc receptor is Fc RI (CD64), Fc RIIa (CD32a), Fc RIIb (CD32b), Fc RIIc (CD32c), Fc RIIIa (CD16a), or Fc RIIIb} (CD16b). 28. The multispecific antibody of any of claims 1-27, wherein the Fc region is afucosylated. 29. The multispecific antibody of any of claims 1-28, wherein the antibody is an immunoglobulin G (IgG). 30. The multispecific antibody of claim 29, wherein the IgG is IgG1, IgG2, IgG3, or IgG4. 31. The multispecific antibody of any of claims 1-30, wherein the antibody comprises a 1+1 format. 32. The multispecific antibody of claim 31, wherein the 1+1 format comprises two Fab regions, wherein the first Fab region comprises (i) a CL domain comprising a S176K and N137K or N137R mutation and (ii) a CH1 domain comprising a S183D and T187D mutation; and wherein the second Fab region comprises (i) a CL domain comprising a S176D and N137D mutation and (ii) a CH1 domain comprising a S183K and T187K mutation; wherein the residues are numbered according to EU. 33. The multispecific antibody of claim 32, wherein the Fc region of the antibody having the 1+1 format comprises a mutation in one or both CH2 domains, wherein the mutation is L234A/L235A/D265S (AAS); L234A/L235A/P329G (LALAPG); S298A/E333A/K334A (AAA); N297K; M252Y/S254T/T256E (YTE); G236A/S239D/A330L/I332E (GASDALIE); G236A; S239D/I332E (DE); S239D/A330L/I332E (DLE); G236A/S239D/I332E (ADE); G236A/A330L/I332E (GAALIE); F243L/R292P/Y300L/V305I/P396L (LPLIL); L235V/F243L/R292P/Y300L/P396L (VLPLL); P238K/L235E/P238K; L235E/P238K/K322A; L234A/L235E/G237A; L234F/L235E/P331S (FES); L234F/L235Q/K322Q (FQQ); Attorney Docket No.14681-WO-PCT A330S/P331S; L234A/G237A; L234A/L235A/G237A; L234A/L235A/G237A/P238S/H268A/A330S/P331S; L234A/L235E; G236R/L328R; or L234A/L235A/K322A. 34. The multispecific antibody of claim 32 or 33, wherein the Fc region comprises the mutations Y349C, T366S, L368A, Y407V (Hole mutations) in a first CH3 domain, and the mutations S354C and T366W (Knob mutations) in a second CH3 domain. 35. The multispecific antibody of claim 34, wherein the Hole mutations and Knob mutations are disulfide stabilized. 36. The multispecific antibody of any of claims 5-35, wherein the antibody comprises a 1+1+1 or 2+1 format. 37. The multispecific antibody of claim 36, wherein the antibody has a 2+1 format comprising three Fab regions, wherein: the first Fab region is linked to the second Fab region in the N-terminal to C-terminal direction; the second Fab region is linked to a first CH2 domain in the N-terminal to C-terminal direction, which is linked to a first CH3 domain; the third Fab region is linked to a second CH2 domain in the N-terminal to C- terminal direction, which is linked to a second CH3 domain; the first Fab region and the third Fab region each comprises a variable region that binds to the same antigen, optionally wherein the variable regions are identical; the second Fab region comprises a variable region that binds to a different antigen than the first Fab region and the third Fab region; the first Fab region and the third Fab region each comprises a CH1 domain comprising the amino acid substitutions S188D and T192D; the first Fab region and the third Fab region each comprises a CL domain comprising the amino acid substitutions S176K and N137K; the second Fab region comprises a CH1 domain comprising the amino acid substitutions S188K and T192K; and Attorney Docket No.14681-WO-PCT the second Fab region comprises a CL domain comprising the amino acid substitutions S176D and N137D. 38. The multispecific antibody of claim 37, wherein the Fc region of the antibody having the 2+1 format comprises a mutation in one or both CH2 domains, wherein the mutation is L234A/L235A/D265S (AAS); L234A/L235A/P329G (LALAPG); S298A/E333A/K334A (AAA); N297K; M252Y/S254T/T256E (YTE); G236A/S239D/A330L/I332E (GASDALIE); G236A; S239D/I332E (DE); S239D/A330L/I332E (DLE); G236A/S239D/I332E (ADE); G236A/A330L/I332E (GAALIE); F243L/R292P/Y300L/V305I/P396L (LPLIL); L235V/F243L/R292P/Y300L/P396L (VLPLL); P238K/L235E/P238K; L235E/P238K/K322A; L234A/L235E/G237A; L234F/L235E/P331S (FES); L234F/L235Q/K322Q (FQQ); A330S/P331S; L234A/G237A; L234A/L235A/G237A; L234A/L235A/G237A/P238S/H268A/A330S/P331S; L234A/L235E; G236R/L328R; or L234A/L235A/K322A. 39. The multispecific antibody of claim 37 or 38, wherein the Fc region comprises the mutations Y349C, T366S, L368A, Y407V (Hole mutations) in a first CH3 domain, and the mutations S354C and T366W (Knob mutations) in a second CH3 domain. 40. The multispecific antibody of claim 39, wherein the Hole mutations and Knob mutations are disulfide stabilized. 41. The multispecific antibody of any of claims 36-40, wherein the antibody further comprises a fourth Fab region. 42. The multispecific antibody of claim 33, wherein the antibody comprises a 1+1+1+1, 3+1, or 2+2 format.