WO2022086957A1

WO2022086957A1 - Peg-conjugated anti-mertk antibodies and methods of use

Info

Publication number: WO2022086957A1
Application number: PCT/US2021/055606
Authority: WO
Inventors: Wei-Ching Liang; Weiyu Lin; Yan Wu; Minhong Yan; Breanna Sachiko VOLLMAR; Mingjian FEI
Original assignee: Genentech Inc
Current assignee: Genentech Inc
Priority date: 2020-10-20
Filing date: 2021-10-19
Publication date: 2022-04-28
Anticipated expiration: 2023-04-20
Also published as: AR123855A1; TW202233671A

Abstract

The invention provides PEGylated anti-MerTK antibodies and methods of making and using the same.

Description

PEG-CONJUGATED ANTI-MERTK ANTIBODIES AND METHODS OF USE

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority to U.S. Provisional Application Serial No. 63/094,197, filed October 20, 2020, the disclosure of which is incorporated herein by reference in its entirety.

SUBMISSION OF SEQUENCE LISTING ON ASCII TEXT FILE

[0002] The content of the following submission on ASCII text file is incorporated herein by reference in its entirety: a computer readable form (CRF) of the Sequence Listing (file name: 146392051740SEQLIST.TXT, date recorded: October s, 2021, size: 60,047 bytes).

FIELD

[0003] The present invention relates to PEGylated anti-MeiTK. antibodies and methods of making and using the same.

BACKGROUND

[0004] Currently most cancer immuno-oncology (IO) therapies focus on modulating the activity of T cells, the adaptive arm of immune system, by blocking inhibitory pathways tliat serve as immunological checkpoints. However, long-lasting responses triggered by these therapies are limited to subpopulations of cancer patients. The relatively low response rate is caused by various immunosuppressive mechanisms in the tumor microenvironment. The innate immune system is an integral part of an effective immune response. Innate immune cells play a crucial part in initiating and subsequent direction of the adaptive immune response. Targeting the innate immune system may complement the adaptive immuno-oncology therapies (Midland, A., Nat. Rev. Drug Discov., 17: 3-5 (2018)).

[0005] Macrophages of the innate immune system are abundant in various types of solid tumors and may contribute to the relatively low response rate to T-cell based therapy. They are versatile cells capable of cartying out various functions, including phagocytosis. Macrophages are professional phagocytes highly specialized in removal of dying or dead cells, and cellular debris. It is estimated tltat billions of cells die every day in the human body. But it is rare to find apoptotic cells in tissues under normal physiological conditions thanks to the rapid and efficient clearance by phagocytes. In homeostasis, apoptotic cells are removed at the early stage of cell death before loss of plasma membrane integrity. Therefore, in general apoptosis is immunologically silent. In solid tumors, uncontrolled tumor growth is often accompanied bv increased cell death due to hypoxia and metabolic stress. To evade immune surveillance, tumors take advantage of the non-iininunogenic nature of

4- apoptosis. Tumor associated macrophages (TAMs) actively remove the dying tumor cells to avoid alerting the immune system.

[0006] MerTK has been shown to play a role in clearance of apoptotic ceils. Therefore, reduction of MerTK-mediated clearance of apoptotic ceils using MerTK inhibitors is an attractive therapeutic approach in treating cancer. Existing anti-MerTK antibodies have been described but may be unsuitable for therapeutic development. For example, White et al. (“MERTK-Specific Antibodies That Have Therapeutic Antitumor Activity in Mice Disrupt the Integrity of the Retinal Pigmented Epithelium in Cynomoigus Monkeys," presented at the American Association for Cancer Research Annual Meeting; March 31, 2019; Atlanta, GA) describe two anti-MerTK antibodies: one that binds to human MerTK with higher affinity (8.7xlO'ⁿM; SRF1), and one that binds to human MerTK with lower affinity (4.4xl0⁹) but cross-reacts with murine MerTK (SRF2). These antibodies were shown to inhibit various MerTK functions and inhibit tumor growth in combination with anti-PD-Ll antibody in a mouse model. However, both antibodies were found to promote retinal toxicity in cynomoigus monkey. As such, neither antibody would be acceptable as a therapeutic candidate.

These findings underscore the importance of examining multiple factors, not simply antibody affinity, in developing an efficacious therapeutic candidate with an acceptable safety profile.

[0007] Thus, there remains a need for an optimal therapy for treating, stabilizing, preventing, and/or delaying development of various cancers. In particular, anti-MerTK antibodies having optimal binding characteristics (e.g., on and off rates) as well as desired biological effects are needed. For example, the need exists for anti-MerTK antibodies with desired therapeutic effects but reduced or eliminated retinal toxicity .

[0008] All references cited herein, including patent applications, patent publications, and UniProtKB/Swiss-Prot Accession numbers are herein incorporated by reference in their entirety, as if each individual reference were specifically and individually indicated to be incorporated by reference.

SUMMARY

[0009] The present invention provides PEGylated anti-MerTK antibodies and methods of using the same.

[0010] In one aspect, the present disclosure provides a PEGylated antibody that binds to MerTK, wherein the antibody is conjugated to one or more polyethylene glycol (PEG) polymer(s); wherein each of the PEG polymer(s) is conjugated to a heavy chain or light chain of the antibody at an engineered cysteine residue; and wherein the antibody comprises: (1 ) a heavy chain variable domain (ATI) comprising (a) a CDR-H1 comprising the amino acid sequence of SY2AMG (SEQ ID NO: 1), (b) a CDR-H2 comprising the amino acid sequence of IINSYGNTYYANWAKG (SEQ ID NO: 2), and (c) a CDR-H3 comprising die amino acid sequence of DPGVSSNL (SEQ ID NO: 3), and a light chain variable domain (Vi.,) comprising (d) a CDR-L1 comprising the amino acid sequence of QASQNIYSGLA (SEQ ID NO: 4), (e) a CDR-L2 comprising the amino acid sequence of GASKLAS (SEQ ID NO: 5), and (f) a CDR-L3 comprising the amino acid sequence of QATYYSSNSVA (SEQ ID NO: 6); or (2) a heavy chain variable domain (VH) comprising (a) a CDR-H1 comprising the amino acid sequence of ANTMN (SEQ ID NO: 7), (b) a CDR-H2 comprising the amino acid sequence of IFTATGSTYYATWVNG (SEQ ID NO: 8), and (c) a CDR-H3 comprising the amino acid sequence of SGSGSSSGAFNI (SEQ ID NO: 9), and a light chain variable domain (VL) comprising (d) a CDR-L1 comprising the amino acid sequence of QASQSISSSLA (SEQ ID NO: 10), (e) a CDR-L2 comprising the amino acid sequence of AASILAS (SEQ ID NO: 11), and (f) a CDR-L3 comprising the amino acid sequence of QCTSYGSLFLGP (SEQ ID NO: 12). In one aspect, tire present disclosure provides a PEGylated antibody that binds to MerTK, wherein the antibody is conjugated to one or more polyethylene glycol (PEG) poiymer(s): wherein each of the PEG polvmer(s) is conjugated to a heavy chain or light chain of the antibody at an engineered cysteine residue; and wherein the antibody comprises: a heavy chain variable domain (VH) comprising (a) a CDR-H1 comprising the amino acid sequence of SYAMG (SEQ ID NO: 1), (b) a CDR-H2 comprising the amino acid sequence of IINSYGNTYYANWAKG (SEQ ID NO: 2), and (c) a CORES comprising the amino acid sequence of DPGVSSNL (SEQ ID NO: 3), and a light chain variable domain (VL) comprising (d) a CDR-L1 comprising tire amino acid sequence of QASQNIYSGLA (SEQ ID NO: 4), (e) a CDR-L2 comprising the amino acid sequence of GASKLAS (SEQ ID NO: 5), and (f) a CDR-L3 comprising the amino acid sequence of QATYYSSNSVA (SEQ ID NO: 6). In one aspect, the present disclosure provides a PEGylated antibody that binds to MerTK, wherein the antibody is conjugated to one or more polyethylene glycol (PEG) polymer(s); wherein each of the PEG polymer(s) is conjugated to a heavy chain or light chain of the antibody at an engineered cysteine residue; and wherein the antibody comprises: a heavy chain variable domain (VH) comprising (a) a CDR-H1 comprising the amino acid sequence of ANTMN (SEQ ID NO: 7), (b) a CDR-H2 comprising the amino acid sequence of IFTATGSTYYATWVNG (SEQ ID NO: 8), and (c) a CDR-H3 comprising the amino acid sequence of SGSGSSSGAFNI (SEQ ID NO: 9), and a light drain variable domain (VL) comprising (d) a CDR-L1 comprising the amino acid sequence of QASQSISSSLA (SEQ ID NO: 10), (e) a CDR-L2 comprising the amino acid sequence of AASILAS (SEQ ID NO: 11), and (f) a CDR-L3 comprising the amino acid sequence of QCTSYGSLFLGP (SEQ ID NO: 12).

[0011] In some embodiments, the VH domain comprises a sequence having at least 95% sequence identity to the amino acid sequence of EVQLVESGEGLVQPGGSLRLSCAASGFSLSSYAMGWVRQAPGKGLEYVGIINSYGNTYYAN WAKGRFTISRDNSKNTVYLQMGSLRAEDMAVYYCARDPGVSSNLWGRGTLVTVSS (SEQ ID NO: 13); and/or the VL domain comprises a sequence having at least 95% sequence identity to the amino acid sequence of DIQMTQSPSTLSASVGDRVTITCQASQNIYSGLAWYQQKPGKAPKLLIYGASKLASGVPSRFS GSGSGTEFTLTISSLQPDDFATYYCQATYYSSNSVAFGGGTKVEIK (SEQ ID NO: 14), In some embodiments, the VH domain comprises the amino acid sequence of

EVQLVESGEGLVQPGGSLRLSCAASGFSLSSYAMGWVRQAPGK.GLEYVGIINSYGNTYYAN WAKGRFTISRDNSKNTVYLQMGSLRAEDMAVYYCARDPGVSSNLWGRGTLVTVSS (SEQ ID NO: 13); and the VL domain comprises the amino acid sequence of

DIQMTQSPS’n.SASVGDRVTITCQASQNIYSGLAWYQQKPGKAPKLLIYGASKLASGVPSRFS GSGSGTEFTLTISSLQPDDFATYYCQATYYSSNSVAFGGGTKVEIK (SEQ ID NO: 14). In some embodiments, the VH domain comprises a sequence having at least 95% sequence identity to the amino acid sequence of

EQQLVESGEGLVQPGGSLRLSCAVSGFSLSSYAMGWVRQAPGKGLEWIGIINSYGNTYYAN WAKGRFTISRDSSKNTVYLQMGSLRAEDMAVYFCARDPGVSSNLWGPGTLVTVSS (SEQ ID NO: 15); and/or the VL domain comprises a sequence having at least 95% sequence identity to the amino acid sequence of

DVQMTQSPSTLSASVGDRVTITCQASQNIYSGLAWYQQKPGKPPKLLIYGASKLASGVPSRFS GSGSGTEFTETISSLQPDDFATYYCQATYYSSNSVAFGGGTKVEIK (SEQ ID NO: 16). In some embodiments, the VH domain comprises the amino acid sequence of

EQQLVESGEGLVQPGGSLRLSCAVSGFSLSSYAMGWVRQAPGKGLEWIGIINSYGNTYYAN WAKGRFTISRDSSKNTX^QMGSLRAEDMAVYFCARDPGVSSNLWGPGTLVTVSS (SEQ ID NO: 15); and the XT, domain comprises the amino acid sequence of

DVQMTQSPSTLSASVGDRVTITCQASQNIYSGLAWYQQKPGKPPKLLIYGASKLASGVPSRFS GSGSGTEFTLTISSLQPDDFATYYCQATYYSSNSVAFGGGTKVEIK (SEQ ID NO: 16). In some embodiments, the VH domain comprises a sequence having at least 95% sequence identity to the amino acid sequence of

QSVEESGGRLVTPGTPLTLTCTVSGFSLSSYAMGWVRQAPGKGLEWIGnNSYGNTYYANWA KGRFTISRTSTTVDLRMPSLTTEDTATYFCARDPGVSSNLWGPGTLVTVSS (SEQ ID NO: 17); and/or the XT, domain comprises a sequence having at least 95% sequence identity to the amino acid sequence of

DVty’NITQTPASVSEPVGGTVTIKCQASQNIYSGLAW'YQQKPGQPPKLLIYGASKLASGVSSRF KGSGSGTEFTLTISDLECADAATYYCQATYYSSNSVAFGGGTEVVVK (SEQ ID NO: 18). In some embodiments, the XT-I domai n comprises the amino acid sequence of

QSVEESGGRLVTPGTPLTLTCTVSGFSLSSYAMGWVRQAPGKGLEWIGIINSYGNTYYANWA KGRFTISRTSTTWLRMPSLTTEDTATYFCARDPGVSSNLWGPGTLVTVSS (SEQ ID NO: 17); and the VL domain comprises the amino acid sequence of

DWMTQTPASVSEPVGGTVTIKCQASQNrYSGLAWYQQKPGQPPKLLIYGASKLASGVSSRF KGSGSGTEFTLT1SDLECADAATYYCQATYYSSNSVAFGGGTEVVVK (SEQ ID NO: 18). In some embodiments, the VH domain comprises a sequence having at least 95% sequence identity to the amino acid sequence of QSVEESGGRLVTPGTPLTLTCWSGIDLSANTMNWVRQAPGKGLEWTGIFTATGSTA'YATWV NGRFTISKTSTTVDLKITSPTTEDTATYTCARSGSGSSSGAFNTWGPGTLVTVSL (SEQ ID NO: 19); and/or the VL domain comprises a sequence having at least 95% sequence identity to the amino acid sequence of

DPVLTQTPASVSEPVGGTVTIKCQASQSISSSLAWYQQKPGQPPKLLIYAASILASEISSRFKGS RSGTEFTLTISDLECADAATYYCQCTSYGSLFLGPFGGGTEVVVK (SEQ ID NO: 20). In some embodiments, the VH domain comprises die amino acid sequence of

QSVEESGGRLVTPGTPLTLTCTVSGIDLSANTMNWVRQAPGKGLEWIGIFTATGSTYYATWV NGRFTISKTSTTVDLKITSPTTEDTATYFCARSGSGSSSGAFNIWGPGTLVTVSL (SEQ ID NO: 19): and the VL domain comprises the amino acid sequence of

DPVLTQTPASVSEPVGGTVTIKCQASQSISSSLAWYQQKPGQPPKLLIYAASILASEISSRFKGS RSGTEFTLTISDLECADAATYYCQCTSYGSLFLGPFGGGTEVVVK (SEQ ID NO: 20).

[0012] In some embodiments, the heavy chain comprises the sequence

EVQLVESGEGLVQPGGSLRLSCAASGFSLSSYAMGWVRQAPGKGLEYVGIINSYGNTYYAN WAKGRFTISRDNSKNTVYLQMGSLRAEDMAVYYCARDPGVSSNLWGRGTLVTVSSASTKG PSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSV VTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEAAGGPSVFLFPPKPK DTL,MISRTPEVTCWA^/DVSHEDPE\TCFNWYVDGVEVHNAKTKPREEQYNSTYRWSVI.TVI., HQDWI.NGKEYKCKVSNKALGAPIEKTISKAKGQPREPQWn,PPSREEMTKNQVSLTCLVK GFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFELYSKLTVDKSRWQQGNVFSCSVMHEAL HNHYTQKSLSLSPG (SEQ ID NO: 21). In some embodiments, the heavy chain comprises the sequence

EVQLVESGEGLVQPGGSLRLSCAASGFSLSSYAMGWVRQAPGKGLEYVGIINSYGNTYYAN WAKGRFTISRDNSKNTVYLQMGSLRAEDMAVYYCARDPGVSSNLWGRGTLVTVSSASTKG PSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSV VTVPSSSLGTQTYICNVNHKPSNTKVDKKX⁷EPKSCDKTHTCPPCPAPELLGGPSV⁷FLFPPKPK DTLMISRTPEWCVV’VDVSHEDPEVKFNWYV'DGVEVHNAKTKPREEQYGSTYRVVSVLTVL HQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKG FYPSDIAVTWESNGQPENNYKTTPPVLDSDGSFFIATSKLTVDKSRWQQGNVFSCSVMHEALH NHYTQKSLSLSPG (SEQ ID NO: 22). In some embodiments, the light chain comprises the sequence DIQlvfTQSPS’n.SASVGDRVTITCQASQNIYSGLAWYQQKPGKAPKLLIYGASKLASGVPSRFS GSGSGTEFTLTISSLQPDDFATYYCQATYYSSNSVAFGGGTKVEIKRTVAAPSVFIFPPSDEQL KSGTASVVCLLNNFYPREAKVQWKWNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADY EKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 23).

[0013] In some embodiments, the antibody is conjugated to one or two PEG polymers. In some embodiments, the PEG polymer(s) are linear PEG polymers. In some embodiments, the PEG polvmer(s) are branched PEG polymers. In some embodiments, the branched PEG polymer(s) comprise 2 branches. In some embodiments, the PEGylated antibody lias a hydrodynamic radius of greater than about 6 ran. In some embodiments, the PEGylated antibody has a hydrodynamic radius of greater than or equal to about 10 sun. In some embodiments, the PEGylated antibody has a hydrodynamic radius of between about 6 ran and about 10 ran, or between about 9 nm and about 11 nm. In some embodiments, the PEG poly mer(s) each have a molecular weight of between about 10 kDa and about 40 kDa, between about 20 kDa and about 40 kDa, or between about 10 kDa and about 20 kDa, e.g., about 10 kDa, about 20 kDa, about 30 kDa, or about 40 kDa. In some embodiments, each of the PEG poly inert s) is conjugated to a heavy chain or light chain of the antibody at an engineered cysteine residue via maleirmde-cy steine conjugation. In some embodiments, each of the PEG polymer(s) is conjugated to a heavy chain or light chain of the antibody at an engineered cysteine residue via iodoacetamide-cy steine conjugation. In some embodiments, the engineered cysteine residue is selected from the group consisting of K149C of the light chain, K183C of the light chain, T186C of the heavy chain, and Y373C of the heavy drain, numbering of the light chain according to Rabat and numbering of the heavy chain according to EU index. In some embodiments, the antibody comprises two heavy chains, two light chains, and two PEG polymers; and wherein both light chains of the antibody comprise a K149C engineered cysteine conjugated to one of the two PEG polymers. In some embodiments, the antibody comprises two heavy drains, two light drains, and two PEG polymers; and wherein both light chains of the antibody comprise a K183C engineered cysteine conjugated to one of the two PEG polymers. In some embodiments, the antibody comprises two heavy chains, two light chains, and two PEG poly mers; and wherein both heavy chains of the antibody comprise a T186C engineered cysteine conjugated to one of the two PEG polymers. In some embodiments, the antibody comprises two heavy drains, two light chains, and two PEG polymers; and wherein both heavy chains of the antibody comprise a Y373C engineered cysteine conjugated to one of the two PEG polymers.

[0014] In one aspect, tire present disclosure provides a method of producing a PEGylated antibody that binds to MeiTK, comprising contacting (i.e., convalently attaching) an antibody comprising one or more free engineered cysteine residues with one or more polyethylene glycol (PEG) polymer(s) comprising a maleimide moiety under conditions suitable for each of the PEG polymer(s) to be conjugated to an engineered cysteine residue of the antibody via thioether linkage. In some embodiments, the one or more free engineered cysteine residues of the antibody are contacted (i.e., chemically reacted) with the maleimide moiety of the one or more PEG polymer(s) at a pH between 7.0 and 7.5. In one aspect, the present disclosure provides a method of producing a PEGylated antibody that binds to MerTK, comprising contacting (i.e., convalently attaching) an antibody comprising one or more free engineered cysteine residues with one or more polyethylene glycol (PEG) polymer(s) comprising an iodoacetamide moiety under conditions suitable for each of the PEG polymer(s) to be conjugated to an engineered cysteine residue of the antibody via thioether linkage. In some embodiments, the antibody comprises a heavy chain variable domain (VH) comprising (a) a CDR-H1 comprising the amino acid sequence of SYAMG (SEQ ID NO: i), (b) a CDR-H2 comprising the amino acid sequence of IINSYGNTYYANWAKG (SEQ ID NO: 2), and (c) a CDR-H3 comprising the amino acid sequence of DPGVSSNL (SEQ ID NO: 3), and a light chain variable domain (VL) comprising (d) a CDR-LJ comprising the amino acid sequence of QASQNIYSGLA (SEQ ID NO: 4), (e) a CDR-L2 comprising the amino acid sequence of GASKLAS (SEQ ID NO: 5), and (f) a CDR-L3 comprising the amino acid sequence of QATYYSSNSVA (SEQ ID NO: 6). In some embodiments, the antibody comprises heavy chain variable domain ( VH) comprising (a) a CDR-H1 comprising the amino acid sequence of ANTMN (SEQ ID NO: 7), (b) a CDR-H2 comprising the amino acid sequence of IFTATGSTYYATWVNG (SEQ ID NO: 8), and (c) a CDR-H3 comprising the amino acid sequence of SGSGSSSGAFNI (SEQ ID NO: 9), and a light chain variable domain (VL) comprising (d) a CDR-L1 comprising the amino acid sequence of QASQSISSSLA (SEQ ID NO: 10), (e) a CDR-L2 comprising the amino acid sequence of AASILAS (SEQ ID NO: 11), and (f) a CDR-L3 comprising the amino acid sequence of QCTSYGSLFLGP (SEQ ID NO: 12).

[0015] In some embodiments, the PEG polymer(s) are conjugated to the antibody at a ratio of 2.0 polymers per antibody. In some embodiments, prior to contacting (i.e., convalently attaching) the antibody with the PEG polymer(s), each of the free engineered cysteine residues is blocked with a cysteine or glutathione moiety; and the method further comprises deblocking the engineered cysteine residue. In some embodiments, the method further comprises, after contacting (i.e., convalently attaching) the antibody with the PEG polymer(s), purifying tire PEGylated antibody from unconjugated antibody and PEG polymer. In some embodiments, the PEGylated antibody is purified by hydrophobic interaction chromatography .

[0016] In some embodiments, the VH domain comprises a sequence having at least 95% sequence identity to the amino acid sequence of EVQLVESGEGLVQPGGSLRLSCAASGFSLSSYAMGWVRQAPGKGLEYVGIINSYGNTYYAN WAKGRFTTSRDNSKNTVYLQMGSLRAEDMAVYYCARDPGVSSNLWGRGTLVTVSS (SEQ ID NO: 13); and/or the VL domain comprises a sequence having at least 95% sequence identity to the amino acid sequence of DIQMTQSPSTLSASVGDRVnTCQASQNIYSGLAWYQQKPGKAPKLLIYGASKLASGVPSRFS GSGSGTEFTLTISSIXJPDDFATYYCQATYYSSNSVAFGGGTKVEIK (SEQ ID NO: 14). In some embodiments, the VH domain comprises the amino acid sequence of EVQLVESGEGLVQPGGSLRLSCAASGFSLSSYAMGWVRQAPGKGLEYVGIINSYGNTYYAN WAKGRFTISRDNSKNTVYLQMGSLRAEDMAVYYCARDPGVSSNLWGRGTLVTVSS (SEQ ID NO: 13); and the VL domain comprises tlie amino acid sequence of DIQMTQSPSTLSASVGDRVTITCQASQNIYSGLAWYQQKPGKAPKLLIYGASKLASGVPSRFS GSGSGTEFTLTISSLQPDDFATYYCQATYYSSNSVAFGGGTKVEIK (SEQ ID NO: 14). In some embodiments, the VH domain comprises a sequence having at least 95% sequence identity to the amino acid sequence of EQQLVESGEGLVQPGGSLRLSCAVSGFSLSSYAMGWVRQAPGKGLEW1GIINSYGNTYYAN WAKGRFTISRDSSKNTVYLQMGSLRAEDMAVYFCARDPGVSSNLWGPGTLVWSS (SEQ ID NO: 15); and/or the VL domain comprises a sequence having at least 95% sequence identity to the amino acid sequence of DVQMTQSPSTLSASVGDRVTITCQASQNIYSGLAWYQQKPGKPPKLLIYGASKLASGVPSRFS GSGSGTEFTLTISSLQPDDFATYYCQATYYSSNSVAFGGGTKVEIK (SEQ ID NO: 16). In some embodiments, the VH domain comprises the amino acid sequence of EQQLVESGEGLVQPGGSLRLSCAVSGFSLSSYAMGWVRQAPGKGLEWIGIINSYGNTYYAN WAKGRFTISRDSSKNTVYLQMGSLRAEDMAVYFCARDPGVSSNLWGPGTLVTVSS (SEQ ID NO: 15); and the VL domain comprises the amino acid sequence of

DVQMTQSPSTLSASVGDRVTITCQASQNIYSGLAWYQQKPGKPPKLLIYGASKLASGVPSRFS GSGSGTEFTLTISSLQPDDFATYY'CQA’TY'YSSNSVAFGGGTKVEIK (SEQ ID NO: 16). In some embodiments, the VH domain comprises a sequence having at least 95% sequence identity to the amino acid sequence of

QSVEESGGRLVTPGTPLTLTCTVSGFSLSSYAMGWVRQAPGKGLEWIGIINSYGNTYYANWA KGRFTISRTSTTVDLRMPSLTTEDTATYFCARDPGVSSNLWGPGTLVTVSS (SEQ ID NO: 17); and/or the VL domain comprises a sequence having at least 95% sequence identity to the amino acid sequence of

DWMTQTPASVSEPVGGTVTIKCQASQNIYSGLAWYQQKPGQPPKLLIYGASKLASGVSSRF KGSGSGTEFTLTISDLECADAATYYCQATYYSSNSVAF'GGGTEVVVK (SEQ ID NO: 18). In some embodiments, the VH domain comprises the amino acid sequence of QSVEESGGRLVTPGTPLTLTCTVSGFSLSSYAMGWVRQAPGKGLEWIGIINSYGNTYYANWA KGRFTISRTSTTVDLRMPSLTTEDTATYFCARDPGVSSNLWGPGTLVTVSS (SEQ ID NO: 17); and tiie VL domain comprises the amino acid sequence of

DVVMTQTPASVSEPVGGTVTIKCQASQNIYSGLAWYQQKPGQPPKLLIYGASKLASGVSSRF KGSGSGTEFTLTISDLECADAATYYCQATYYSSNSVAFGGGTEVWK (SEQ ID NO: 18). In some embodiments, the VH domain comprises a sequence having at least 95% sequence identity to the amino acid sequence of

QSVEESGGRLVTPGTPLTL,TCTVSGIDLSANTMNWVR.QAPGKGLEWIGIFTATGSTYYATWV NGRFTISKTSTTVDLKITSPTTEDTATYFCARSGSGSSSGAFNIWGPGTLVTVSL (SEQ ID NO: 19); and/or the VL domain comprises a sequence having at least 95% sequence identity to the amino acid sequence of

DPVLTQTPASVSEPVGGTVIIKCQASQSISSSLAWYQQKPGQPPKLLIYAASILASE1SSRFKGS RSGTEFTLTISDLECADAATYYCQCTSYGSLFLGPFGGGTEVVVK (SEQ ID NO: 20). In some embodiments, the VH domain comprises the amino acid sequence of QSVEESGGRLVTPGTPLTLTCTVSGIDLSANTMNWVRQAPGKGLEW1GIFTATGSTYYATWV NGRFTISKTSTTVDLKITSPTTEDTATYTCARSGSGSSSGAFNTWGPGTLVTVSL (SEQ ID NO: 19); and the VL domain comprises the amino acid sequence of

DPVLTQTPASVSEPVGGTVTIKCQASQSISSSLAWYQQKPGQPPKLLIYAASILASEISSRFKGS RSGTEFTLTISDLECADAATYYCQCTSYGSLFLGPFGGGTEVWK (SEQ ID NO: 20).

[0017] In some embodiments, the heavy chain comprises the sequence

EVQIATiSGEGLVQPGGSLRLSCAASGFSLSSYAMGWVRQAPGKGLEYVGTINSYGNTYYAN WAKGRFTISRDNSKNTVYLQMGSLRAEDMAVYYCARDPGVSSNLWGRGTLVTVSSASTKG PSVTPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSV VTVPSSSLGTQTYICNVNIdKPSNTKVDIvKVEPKSCDKTHTCPPCP/kPEAAGGPSVFLFPPKPK DTLMISRTPEVTCWVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVL HQDWLNGKEYKCKVSNKALGAPIEKTISKAKGQPREPQVYTLPPSREEMIKNQVSLTCLVK GFYPSD1AVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEAL HNHYTQKSLSLSPG (SEQ ID NO: 21). In some embodiments, the heavy chain comprises the sequence

EVQLVESGEGLVQPGGSLRLSCAASGFSLSSYAMGWVRQAPGKGLEYVGIINSYGNTY'YAN WAKGRFTISRDNSKNTVYLQMGSLRAEDMAWYCARDPGVSSNLWGRGTLVTVSSASTKG PSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSV VTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPK DTT,MISRTPEVTCWA^/DVSHEDPE\TCFNWYVDGVEVHNAKTKPREEQYGSTYRWSVI.TVI., I-IQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKG FYPSDIAVEWESNGQPENNYKTTPP\⁷LDSDGSFFLYSKLT\⁷DKSRWQQGNVFSCSVMIIEALH NHYTQKSLSLSPG (SEQ ID NO:22). In some embodiments, the light chain comprises the sequence DIQMTQSPSTLSASVGDRVT1TCQASQNIYSGLAWYQQKPGKAPKLL1YGASKLASGVPSRFS GSGSGTEFTLTISSLQPDDFATYYCQATYYSSNSVAFGGGTKVEIKRTVAAPSVFIFPPSDEQL KSGTASWCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADY EKHKWACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 23).

[0018] In some embodiments, the antibody is conjugated to one or two PEG polymers. In some embodiments, the PEG polymer(s) are linear PEG polymers. In some embodiments, the PEG polymer(s) are branched PEG polymers. In some embodiments, the branched PEG poly mer(s) comprise 2. branches. In some embodiments, the PEGylated antibody lias a hydrodynamic radius of greater than about 6 am. In some embodiments, the PEGylated antibody has a hydrodynamic radius of greater than or equal to about 10 nm. In some embodiments, the PEGylated antibody lias a hydrodynamic radius of between about 6 mn and about 10 mu, or between about 9 nm and about 11 nm. In some embodiments, the PEG poly mer(s) each have a molecular weight of between about 10 kDa and about 40 kDa, between about 20 kDa and about 40 kDa, or between about 10 kDa and about 20 kDa, e.g., about 10 kDa, about 20 kDa, about 30 kDa, or about 40 kDa. In some embodiments, each of the PEG polymer(s) is conjugated to a heavy chain or light chain of the antibody at an engineered cysteine residue via maleimide-cvsteine conjugation. In some embodiments, each of the PEG polymer(s) is conjugated to a heavy chain or light chain of the antibody at an engineered cysteine residue via iodoacetamide-cvsteine conjugation. In some embodiments, the engineered cysteine residue is selected from the group consisting of K149C of the light chain, K183C of the light chain, T186C of the heavy chain, and Y373C of the heavy chain, numbering of the light chain according to Kabat and numbering of the heavy chain according to EU index. In some embodiments, the antibody comprises two heavy drains, two light chains, and two PEG polymers; and wherein both light chains of the antibody comprise a K149C engineered cysteine conjugated to one of die two PEG polymers. In some embodiments, die antibody comprises two heavy chains, two light chains, and two PEG polymers; and wherein both light chains of the antibody comprise a K183C engineered cysteine conjugated to one of the two PEG polymers. In some embodiments, the antibody comprises two heavy chains, two light chains, and two PEG polymers; and wherein both heavy chains of the antibody comprise a Ti 86C engineered cysteine conjugated to one of the two PEG polymers. In some embodiments, the antibody comprises two heavy chains, two light chains, and two PEG polymers; and wherein both heavy chains of the antibody comprise a Y373C engineered cysteine conjugated to one of the two PEG polymers.

[0019] In one aspect, the present disclosure provides a PEGylated antibody tlrat binds to MerTK produced by the method of any one of the above embodiments. In one aspect, the present disclosure provides a pharmaceutical composition comprising the PEGy lated anti-MerTK antibody of any one of tiie above embodiments and a pharmaceutically acceptable carrier.

[0028] In one aspect, the present disclosure provides a method of treating an individual having cancer comprising administering to the individual an effective amount of the PEGylated antibody or composition of any one of the above embodiments. In one aspect, the present disclosure provides a method of reducing MerTK-mediated clearance of apoptotic ceils in an individual comprising administering to the individual an effective amount of the PEGylated antibody or composition of any one of the above embodiments. In one aspect, the present disclosure provides the PEGylated anti- MerTK antibody or composition of any one of the above embodiments for use as a medicament. In one aspect, the present disclosure provides the PEGylated anti-MerTK antibody or composition of any one of the above embodiments for use in treating cancer. In one aspect, the present disclosure provides the PEGylated anti-MerTK antibody or composition of any one of the above embodiments for use in the any of the methods disclosed herein, e.g., in methods of treating an individual having cancer and/or reducing MerTK-mediated clearance of apoptotic cells in an individual. In one aspect, the present disclosure provides the PEGylated anti-MerTK antibody or composition of any one of the above embodiments for use in the manufacture of a medicament, e.g., for treating an individual having cancer and/or reducing MerTK-mediated clearance of apoptotic ceils in an individual.

[0021] In some embodiments, administration of the PEGylated antibody causes reduced retinal toxicity in the individual, as compared to administration of an antibody that binds to MerTK tha t is not PEGylated. In some embodiments, the method further comprises administering an additional therapeutic agent to the individual. In some embodiments, the additional therapeutic agent is selected from one or more of tamoxifen, letrozole, exemestane, anastrozole, irinotecan, cetuximab, fulvestrant, vinorelbine, erlotinib, bevacizumab, vincristine, imatinib mesylate, sorafenib, lapatinib, trastuzumab, cisplatin, gemcitabine, methotrexate, vinblastine, carboplatin, paclitaxel, 5-fIuorouracil, doxorubicin, bortezomib, melpbalan, prednisone, and docetaxel. In some embodiments, the additional therapeutic agent is an immune checkpoint inhibitor. In some embodiments, the immune checkpoint inhibitor is selected from the group consisting of a cytotoxic T-lymphocyte-associated protein 4 (CTLA4) inhibitor, a programmed ceil death protein 1 (PD-1) binding antagonist, and a programmed deathligand 1 (PDL1) binding antagonist. In some embodiments, tire immune checkpoint inhibitor is a PDL1 binding antagonist. In some embodiments, the PDL1 binding antagonist is an anti-PDLl antibody, e.g., atezolizumab. In some embodiments, the method further comprises administering an effective amount of an additional chemotherapeutic agent to the individual. In some embodiments, the cancer is colon cancer. In some embodiments, the cancer is selected from urothelial carcinoma, non-small cell lung cancer, triple negative breast cancer, small cell lung cancer, hepatocellular carcinoma, and melanoma. In some embodiments, clearance of apoptotic cells is reduced by about 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3,

3.4, 3.5, 3.6, 3.7, 3.8, 3.9, 4.0, 4.1, 4.2, 4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6,

5.7, 5.8, 5.9, 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7,3, 7.4, 7.5, 7.6, 7.7, 7.8, 7,9, or 8.0 fold.

[0022] It is to be understood that one, some, or all of the properties of the various embodiments described herein may be combined to form other embodiments of the present disclosure. These and other aspects of the disclosure will become apparent to one of skill in the art. These and other embodiments of the disclosure are further described by the detailed description that follows.

BRIEF DESCRIPTION OF THE DR WINGS

[0023] FIGS. 1A-1C depict on-target ocular toxicity using an anti-MerTK antibody. FIG. 1A is a schematic of the retinal pigment epithelium (RPE) depicting the expression of MerTK on the apical membrane of the RPE. Blockage of MerTK by anti-MerTK antibody in the RPE results in failure to phagocytize shedding photoreceptor outer segments (POS), leading to accumulation of debris and ultimate degeneration and loss of photoreceptors. FIG. IB shows the retinal macrophage infiltration in outer photoreceptor (PR) layer in cynomolgus monkey following administration of 30 mg kg I3B4 anti-MerTK mAb once every 3 weeks for 6 weeks (total 3 doses). The outer nuclear layer (ONE) is labelled in the image. Hematoxylin & Eosin (HE), 200X, FIG. 1C shows the retinal toxicity in Balb/c mice following administration of 30 mg/kg 14C9 anti-MerTK mAb twice weekly for 4 weeks (8 total doses). Compared with the control group, animals receiving 30 mg/kg 14C9 had marked outer retinal degeneration, characterized by PR vacuolization, increased cellularity of the PR layer, and decreased cellularity', degeneration and necrosis of the ONL. Hematoxylin & Eosin (HE), 200X.

[0024] FIG. 2 is a schematic representation of anti-MerTK cysteine-engineered antibody -PEG polymer conjugates, comprising either 40 kDa linear PEG (left) and 40 kDa 2-arm branched PEG (right).

[0025] FIGs. 3 A & 3B show the analysis of 14C9 cysteine-engineered antibody -PEG conjugates binding by Biacore SPR. A total of 6 THIOMAB conjugates on 14C9.C90S, including 2 PEGylated moieties (linear PEG40K and branched PEG40K) on 3 sites (LC_K149C, HC_T 182C, LC_K183C), were evaluated their IgG binding affinity against mouse MerTK along with either untreated or control-treated 14C9.C90S by Biacore SPR. FIG. 3A shows the results for the binding analysis of linear PEG40K~conjugated I4C9 THIOMAB, while FIG. 3B shows the results for branched PEG40K-conjugated 14C9 THIOMAB. The results indicated that all PEG-conjugated 14C9.C90S IgG variants appeared to have a moderate improvement (3-5 fold) in binding affinity to mouse MerTK, with no difference between the conjugation sites.

[0026] FIG. 4 shows the results of binding analysis of 14C9 Fab conjugates. The 14C9.C90S EC K149C series of THIOMAB conjugates (linear PEG40K and branched PEG40K) in Fabs were assessed for their binding affinity against the surface bound mouse MerTK by Biacore SPR. All variants exhibited about a 3-fold drop in binding affinity against mouse MerTK, with no difference in between the conjugation sites.

[0027] FIGS. 5A-5B show a comparison of the inhibitory activity of the PEG40 conjugated 14C9 anti-MerTK antibodies and parental mAb in mouse peritoneal macrophage mediated efferocytosis. FIG. 5A shows the results of the efferocytosis assay with PEG-conjugated 14C9 antibodies. Each panel represents the analysis for antibodies with a different conjugation site: KI49C on the light chain (top left), K183C on the light chain (top right), or T182C on the heavy chain (bottom). FIG. SB summarizes the potency and efficacy of 14C9 antibody and conjugates in the efferocytosis assay.

[0028] FIGS. 6A-6B shoiv a comparison of the inhibitory activity of the PEG40 conjugated 14C9 LC K149C antibodies and parental mAb in mouse peritoneal roacrophage-mediated efferocytosis. FIG. 6A shows the results of the efferocytosis assay with PEG-conjugated 14C9 LC K149C antibodies using commercial (left) and in-house isolated mouse peritoneal macrophages (right). FIG. 6B summarizes the potency and efficacy of 14C9 antibody and 14C9 LC K149C conjugates in the efferocytosis assay.

[0029] FIG. 7 show's the binding of anti-murine MerTK parental antibody and THIOMAB antibody -PEG conjugates to primary murine peritoneal macrophages as measured by FACS.

[0030] FIGS. 8A & 8B show a comparison of the inhibitory’ activity of the PEG40 conjugated

14C9 Fabs and parental mAb and Fab in mouse peritoneal macrophage mediated efferocytosis. FIG. 8A shows the results of the efferocytosis assay with PEG-conjugated 14C9 Fabs. FIG. 88 summarizes the potency and efficacy of 14C9 mAb, Fab and Fab PEG conjugates.

[0031] FIG. 9 shows the results of examining MerTK occupancy and type I IFN signaling following a single high dose (30 mg/kg) of PEG-conjugated 14C9 antibodies or parental mAb (n = 8/group). Analysis of IFNb and interferon-stimulated genes (ISGs) induction in tumors is shown. [0032] FIGS. K1A-10C show the results of analysis MerTK occupancy in tumor-associated macrophages (TAMs) and retinal pigment epithelium cells (RPEs) by and-MerTK antibody conjugates after a single high dose (30 mg/kg) of antibody. FIG. 10A shows the results of the occupancy analysis on TAMs and RPEs (n = 8/group), while FIG. 108 summarizes the occupancy results. FIG. 10C shows the levels of unoccupied MerTK on free-MerTK+ RPE cells. The MFI (mean fluorescence intensity) analysis suggested partial occupancy of MerTK.

[0033] FIGS. 11A-11D show the MerTK occupancy on RPEs after a repeated high dose treatment (45 mg/kg on Day 1 and Day 5) with anti-MerTK antibody conjugates. FIG. 11A shows levels of occupancy of MerTK, while FIG. 118 summarizes the observed occupancy. FIG. 11C shows the levels of unoccupied MerTK on free-MerTK+ RPE cells after incubation with anti-MerTK antibody conjugates. FIG. 1 ID summarizes the molecular weight of the anti-MerTK antibody conjugates.

[0034] FIGS. 12A-12C show the analysis of MerTK occupancy in tumor-associated macroplrages (TAMs) after a low repeated dose treatment (2.5 mg/kg on Day 1 and Day 5) with anti- MerTK antibody conjugates. FIG. 12A shows the results of the occupancy analysis on TAMs. FIG. 128 summarizes the occupancy results. FIG. 12C shows the levels of unoccupied MerTK on free- MerTK+ TAMs.

[0035] FIG. 13 shows the induction of IFNb and interferon-stimulated genes (ISGs) in tumors after low repeated dose (2.5 mg/kg) with PEG-conjugated anti-MerTK antibodies.

DET 'All..ED DESCRIPTION

I. DEFINITIONS

[0036] It is to be understood that this disclosure is not limited to particular compositions or biological systems, which can, of course, vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting. [0037] As used in this specification and the appended claims, the singular forms “a”, “an” and “the” include plural referents unless the content clearly dictates otherwise. Thus, for example, reference to “a molecule” optionally includes a combination of two or more such molecules, and the like. [0038] The term “about” as used herein refers to the usual error range for the respective value readily known to the skilled person in this technical field. Reference to “about” a value or parameter herein includes (and describes) embodiments that are directed to that value or parameter per se.

[0039] It is understood that aspects and embodiments of the present disclosure include “comprising,” “consisting,” and “consisting essentially of’ aspects and embodiments.

[0040] An “acceptor human framework” for the purposes herein is a framework comprising the amino acid sequence of a light chain variable domain (VL) framework or a heavy chain variable domain ( VH) framework derived from a human immunoglobulin framework or a human consensus framework, as defined below. An acceptor human framework “derived from” a human immunoglobulin framework or a human consensus framework may comprise the same amino acid sequence thereof, or it may contain amino acid sequence changes. In some aspects, the number of amino acid changes are 10 or less, 9 or less, 8 or less, 7 or less, 6 or less, 5 or less, 4 or less, 3 or less, or 2 or less. In some aspects, the VL acceptor human framework is identical in sequence to the VL human immunoglobulin framework sequence or human consensus framework sequence.

[0041] “Affinity" refers to the strength of the sum total of noncovalent interactions between a single binding site of a molecule (e.g., an antibody) and its binding partner (e.g., an antigen). Unless indicated otherwise, as used herein, “binding affinity” refers to intrinsic binding affinity which reflects a 1:1 interaction between members of a binding pair (e.g,, antibody and antigen). The affinity of a molecule X for its partner ¥ can generally be represented by the dissociation constant (K_D). Affinity can be measured by common methods known in the art, including those described herein. Specific illustrative and exemplary methods for measuring binding affinity are described in the following.

[0042] An “affinity matured” antibody refers to an antibody with one or more alterations in one or more complementary' determining regions (CDRs), compared to a parent antibody which does not possess such alterations, such alterations resulting in an improvement in the affinity of the antibody for antigen.

[0043] The terms “anti-MeiTK antibody" and “an antibody that binds to MerTK” refer to an antibody that is capable of binding MerTK with sufficient affinity such that the antibody is useful as a diagnostic and/or therapeutic agent in targeting MerTK. In one aspect, the extent of binding of an anti-MerTK antibody to an unrelated, non-MerTK protein is less than about 10% of the binding of the antibody to MerTK as measured, e.g., by surface plasmon resonance (SPR). In certain aspects, an antibody that binds to MerTK lias a dissociation constant (KD) of < IpM, < 100 nM, < 10 nM, < 1 nM, < 0.1 nM, < 0.01 nM, or < 0.001 nM (e.g., 10'⁸ M or less, e.g., from 10“⁸ M to 10’^i:i M, e.g., from IO’⁹ M to 10'^1J M). An antibody is said to “specifically bind” to MerTK when the antibody lias a KD of IpM or less. In certain aspects, an anti-MerTK antibody binds to an epitope of MerTK that is conserved among MerTK from different species. [0044] The term “antibody” herein is used in the broadest sense and encompasses various antibody structures, including but not limited to monoclonal antibodies, polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies), and antibody fragments so long as they exhibit the desired antigen-binding activity.

[0045] An “antibody fragment” refers to a molecule other than an intact antibody that comprises a portion of an intact antibody that binds the antigen to which the intact antibody binds. Examples of antibody fragments include but are not limited to Fv, Fab, Fab', Fab'-SH, F(ab')2; diabodies; linear antibodies; single-chain antibody molecules (e.g., scFv, and scFab); single domain antibodies (dAbs); and multispecific antibodies formed from antibody fragments. For a review of certain antibody fragments, see Holliger and Hudson, Nature Biotechnology 23 : 1126-1136 (2005).

[0046] The term “chimeric” antibody refers to an antibody in which a portion of the heavy and/or light chain is derived from a particular source or species, while the remainder of the heavy and/or light chain is derived from a different source or species.

[0047] The “class” of an antibody' refers to the ty pe of constant domain or constant region possessed by its heavy drain. There are five major classes of antibodies: IgA, IgD, IgE, IgG, and IgM, and several of these may be further divided into subclasses (isotypes), e.g., IgGi , IgG2, IgG3, IgG4, IgAl, and IgA2. In certain aspects, the antibody is of the IgGi isotype. In certain aspects, the antibody is of the IgGi isotype with the P329G, L234A and L235A mutation to reduce Fc-region effector function. In other aspects, the antibody is of the IgG2 isotype. In certain aspects, the antibody is of the IgG4 isotype with the S228P mutation in the hinge region to improve stability of IgG4 antibody . The heavy chain constant domains that correspond to tire different classes of immunoglobulins are called a, 8, s, v, and p, respectively. The light chain of an antibody may be assigned to one of two types, called kappa (K) and lambda (X), based on the amino acid sequence of its constant domain.

[0048] The terms “constant region derived from human origin” or “human constant region" as used in the current application denotes a constant heavy chain region of a human antibody of the subclass IgG i , IgG2, IgG3, or IgG4 and/or a constant light chain kappa or lambda region. Such constant regions are well known in the state of the art and e.g, described by Kabat, E, A., et al,, Sequences of Proteins of Immunological Interest, 5th ed., Public Health Service, National Institutes of Health Bethesda, AID (1991) (see also e.g. Johnson, G., and Wu, T.T., Nucleic Acids Res. 28 (2000) 214-218; Kabat, E.A., et al., Proc. Natl. Acad, Sci. USA 72 (1975) 2785-2788). Unless otherwise specified herein, numbering of amino acid residues in the constant region is according to the EU numbering system, also called the EU index of Kabat, as described in Kabat, E.A. et al.. Sequences of Proteins of Immunological Interest, 5th ed., Public Health Service, National Institutes of Health.. Bethesda, MD (1991), NIH Publication 91-3242.

[0049] “Effector functions” refer to those biological activities attributable to the Fc region of an antibody, which vary with the antibody isotype. Examples of antibody effector functions include: Clq binding and complement dependent cytotoxicity (CDC); Fc receptor binding; antibodydependent cell-mediated cytotoxicity (ADCC); phagocytosis; down regulation of cell surface receptors (e.g., B cell receptor); and B cell activation.

[0050] An “effective amount" of an agent, e.g,, a pharmaceutical composition, refers to an amount effective, at dosages and for periods of time necessary, to achieve the desired therapeutic or prophylactic result.

[0051] The term “Fc region” herein is used to define a C-terminal region of an immunoglobulin heavy chain that contains at least a portion of the constant region. The term includes native sequence Fc regions and variant Fc regions. In one aspect, a human IgG heavy chain Fc region extends from Cys226, or from Pro230, to the caiboxyl-terminus of the heavy chain. However, antibodies produced by host cells may undergo post-translational cleavage of one or more, particularly one or two, amino acids from the C-terminus of the heavy chain. Therefore an antibody produced by a host cell by expression of a specific nucleic acid molecule encoding a full-length heavy chain may include the full-length heavy chain, or it may include a cleaved variant of the full-length heavy chain. This may be the case where the final two C-terminal amino acids of the heavy chain are glycine (G446) and lysine (K447, EU numbering system). Therefore, the C-terminal lysine (Lys447), or the C-terminal glycine (Gly 446) and lysine (Lys447), of the Fc region may or may not be present. Amino acid sequences of heavy chains including an Fc region are denoted herein without C-terminal glycinelysine dipeptide if not indicated otherwise. In one aspect, a heavy chain including anFc region as specified herein, comprised m an antibody according to the invention, comprises an additional C- terminal gly cine -lysine dipeptide (G446 and K447, EU numbering system). In one aspect, a heavy chain including an Fc region as specified herein, comprised in an antibody according to the invention, comprises an additional C-terminal glycine residue (G446, numbering according to EU index). Unless otherwise specified herein, numbering of amino acid residues in the Fc region or constant region is according to the EU numbering system, also called the EU index, as described in Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD, 1991.

[0052] “Framework” or “FR” refers to variable domain residues other than complementary' determining regions (CDRs). The FR of a variable domain generally consists of four FR domains: FR I , FR2, FR3, and FR4, Accordingly, the CDR and FR sequences generally appear in the following sequence in VH (or VL): FR1-CDR-H1(CDR-L1)-FR2- CDR-H2(CDR-L2)-FR3- CDR-H3(CDR- L3)-FR4.

[0053] The terms “full length antibody”, “intact antibody”, and “whole antibody” are used herein interchangeably to refer to an antibody having a structure substantially similar to a native antibody structure or having heavy chains that contain an Fc region as defined herein.

[0054] The terms “host cell”, “host ceil line”, and “host cell culture” are used interchangeably and refer to cells into which exogenous nucleic acid has been introduced, including the progeny of such cells. Host cells include “transformants" and “transformed cells”, which include the primary transformed cell and progeny derived therefrom without regard to the number of passages. Progeny may not be completely identical in nucleic acid content to a parent cell, but may contain mutations. Mutant progeny that have the same function or biological activity as screened or selected for in the originally transformed cell are included herein.

[0055] A “human antibody” is one which possesses an amino acid sequence which corresponds to that of an antibody produced by a human or a human cell or derived from a non-human source that utilizes human antibody repertoires or other human antibody -encoding sequences. This definition of a human antibody specifically excludes a humanized antibody comprising non-human antigen-binding residues.

[0056] A “human consensus framework” is a framework which represents the most commonly occurring amino acid residues in a selection of human immunoglobulin VL or VH framework sequences. Generally, the selection of human immunoglobulin VL or VH sequences is from a subgroup of variable domain sequences. Generally, the subgroup of sequences is a subgroup as in Kabat et al, Sequences of Proteins of Immunological Interest, Fifth Edition, NTH Publication 91- 3242, Bethesda MD (1991), vols. 1-3. In one aspect, for the VL, the subgroup is subgroup kappa I as in Kabat et al., supra. In one aspect, for the VH, the subgroup is subgroup III as in Kabat et al., supra.

[0057] A “humanized” antibody refers to a chimeric antibody comprising amino acid residues from non-human CDRs and amino acid residues from human FRs, In certain aspects, a humanized antibody will comprise substantially all of at least one, and typically two, variable domains, in winch all or substantially all of the CDRs correspond to those of a non-human antibody, and all or substantially all of the FRs correspond to those of a human antibody. A humanized antibody optionally may comprise at least a portion of an antibody constant region derived from a human antibody. A “humanized form” of an antibody, e.g., a non-human antibody, refers to an antibody that has undergone humanization.

[0058] The term “hypervariable region” or “HVR” as used herein refers to each of the regions of an antibody variable domain which are hypeivariable in sequence and which determine antigen binding specificity, for example “complementarity determining regions” (“CDRs”). Generally, antibodies comprise six CDRs: three in the VH (CDR-H1, CDR-H2, CDR-H3), and three in the VL (CDR-L1, CDR-L2, CDR-L3). Exemplary CDRs herein include:

(a) hypeivariable loops occurring at amino acid residues 26-32. (LI), 50-52 (L2), 91-96 (L3), 26-32 (Hl), 53-55 (H2), and 96-101 ( II 3 ) (Chothia and Lesk, J. Mol. Biol. 196:901-917 (1987));

(b) CDRs occurring at amino acid residues 24-34 (LI), 50-56 (L2), 89-97 (L3), 31-35b (Hl), 50-65 (H2), and 95-102 (H3) (Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Sendee, National Institutes of Health, Bethesda, MD (1991)); and (c) antigen contacts occurring at amino acid residues 27c-36 (LI), 46-55 (L2), 89-96 (L3), 30- 35b (Hl), 47-58 (H2), and 93-101 (H3) (MacCallum et al. J. Mol. Biol. 262: 732-745 (1996)). Unless otherwise indicated, the CDRs are determined according to Rabat et al., supra. One of skill in the art will understand that the CDR designations can also be determined according io Chothia, supra, McCallum, supra, or any other scientifically accepted nomenclature system.

[0059] In one aspect, CDR residues comprise those identified in section II. A or elsewhere in the specification.

[0060] An “immunoconjugate” is an antibody conjugated to one or more heterologous molecule(s), including but not limited to a cytotoxic agent.

[0061] An “individual” or “subject” is a mammal. Mammals include, but are not limited to, domesticated animals (e.g., cows, sheep, cats, dogs, and horses), primates (e.g., humans and nonhuman primates such as monkeys), rabbits, and rodents (e.g., mice and rats). In certain aspects, the individual or subject is a human.

[0062] An “isolated” antibody is one which has been separated from a component of its natural environment. In some aspects, an antibody is purified to greater than 95% or 99% purity as determined by, for example, electrophoretic (e.g., SDS-PAGE, isoelectric focusing (IEF), capillary electrophoresis) or chromatographic (e.g., ion exchange or reverse phase HPLC) methods. For a review of methods for assessment of antibody purity, see, e.g., Flatman et al., J. Chromatogr. B 848:79-87 (2007).

[0063] The term “nucleic acid molecule” or “polynucleotide” includes any compound and/or substance that comprises a polymer of nucleotides. Each nucleotide is composed of a base, specifically a purine- or pyrimidine base (i.e. cytosine (C), guanine (G), adenine (A), thymine (T) or uracil (U)), a sugar (i.e. deoxyribose or ribose), and a phosphate group. Often, die nucleic acid molecule is described by the sequence of bases, whereby said bases represent the primary structure (linear structure) of a nucleic acid molecule. The sequence of bases is typically represented from 5’ to 3’. Herein, the term nucleic acid molecule encompasses deoxyribonucleic acid (DNA) including e.g., complementary’ DNA (cDN A) and genomic DNA, ribonucleic acid (RNA), in particular messenger RNA (mRNA), synthetic forms of DNA or RNA, and mixed polymers comprising two or more of these molecules. The nucleic acid molecule may be linear or circular. In addition, the term nucleic acid molecule includes both, sense and antisense strands, as well as single stranded and double stranded forms. Moreover, the herein described nucleic acid molecule can contain naturally occurring or non-naturally occurring nucleotides. Examples of non-naturally occurring nucleotides include modified nucleotide bases with derivatized sugars or phosphate backbone linkages or chemically modified residues. Nucleic acid molecules also encompass DNA and RNA molecules which are suitable as a vector for direct expression of an antibody of the invention in vitro and/or in vivo, e.g., in a host or patient. Such DNA (e.g., cDNA) or RNA (e.g., mRNA) vectors, can be unmodified or modified. For example, mRNA can be chemically modified to enhance the stability' of the RNA vector and/or expression of the encoded molecule so that mRNA can be injected into a subject to generate the antibody in vivo (see e.g., Stadler ert al, Nature Medicine 2017, published online 12 June 2017, doi:10.1038/nm.4356 or EP 2 101 823 Bl).

[0064] An “isolated” nucleic acid refers to a nucleic acid molecule that has been separated from a component of its natural environment. An isolated nucleic acid includes a nucleic acid molecule contained in cells that ordinarily contain the nucleic acid molecule, but the nucleic acid molecule is present extrachromosomally or at a chromosomal location that is different from its natural chromosomal location.

[0065] “Isolated nucleic acid encoding an anti-MerTK antibody” refers to one or more nucleic acid molecules encoding anti-MerTK antibody heavy and light chains (or fragments thereof), including such nucleic acid molecule(s) in a single vector or separate vectors, and such nucleic acid molecule(s) present at one or more locations in a host cell.

[0066] The term “monoclonal antibody" as used herein refers to an antibody obtained from a population of substantially homogeneous antibodies, i.e., the individual antibodies comprising the population are identical and/or bind the same epitope, except for possible variant antibodies, e.g., containing naturally occurring mutations or arising during production of a monoclonal antibody preparation, such variants generally being present in minor amounts. In contrast to polyclonal antibody preparations, which typically include different antibodies directed against different determinants (epitopes), each monoclonal antibody of a monoclonal antibody preparation is directed against a single determinant on an antigen. Thus, the modifier “monoclonal” indicates the character of the antibody as being obtained from a substantially homogeneous population of antibodies, and is not to be construed as requiring production of the antibody by any particular method. For example, tlie monoclonal antibodies in accordance with the present invention may be made by a variety of techniques, including but not limited to the hybridoma method, recombinant DNA methods, phagedisplay methods, and methods utilizing transgenic animals containing all or part of the human immunoglobulin loci, such methods and other exemplary⁷ methods for making monoclonal antibodies being described herein.

[0067] A “naked antibody" refers to an antibody that is not conjugated to a heterologous moiety’ (e.g., a cytotoxic moiety ) or radiolabel. The naked antibody may be present in a pharmaceutical composition,

[0068] “Native antibodies” refer to naturally occurring immunoglobulin molecules w ith vary ing structures. For example, native IgG antibodies are heterotetrameric glycoproteins of about 150,000 daltons, composed of two identical light chains and two identical heavy chains that are disulfide- bonded. From N- to C-terminus, each heavy chain has a variable domain ( VI-I), also called a vanable heavy domain or a heavy chain variable region, followed by three constant heavy domains (CHI, CH2, and CH3). Similarly, from N- to C-terminus, each light chain has a variable domain (VL), also called a variable light domain or a light drain variable region, followed by a constant light (CL) domain.

[0069] The term “package insert” is used to refer to instructions customarily included in commercial packages of therapeutic products, that contain information about the indications, usage, dosage, administration, combination therapy, contraindications and/or warnings concerning the use of such therapeutic products.

[0070] “Percent (%) amino acid sequence identity” with respect to a reference polypeptide sequence is defined as the percentage of amino acid residues in a candidate sequence that are identical with the amino acid residues in the reference polypeptide sequence, after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent sequence identity, and not considering any conservative substitutions as part of the sequence identity for the purposes of the alignment. Alignment for purposes of determining percent amino acid sequence identity can be achieved in various ways that are within the skill in the art, for instance, using publicly available computer software such as BLAST, BLAST-2, Clustal W, Megalign (DNASTAR) software or the FASTA program package. Those skilled in the art can determine appropriate parameters for aligning sequences, including any algorithms needed to achieve maximal alignment over the full length of the sequences being compared. Alternatively, the percent identity values can be generated using the sequence comparison computer program ALIGN-2. The ALIGN-2 sequence comparison computer program was authored by Genentech, Inc,, and the source code has been filed with user documentation in the U.S. Copyright Office, Washington D.C., 20559, where it is registered under U.S. Copyright Registration No. TXU510087 and is described in WO 2001/007611.

[0071] Unless otherwise indicated, for purposes herein, percent amino acid sequence identity values are generated using the ggsearch program of the FASTA package version 36.3.8c or later with a BLOSUM50 comparison matrix. The FASTA program package was authored by W. R. Pearson and D. J. Lipman (1988), “Improved Tools for Biological Sequence Analysis”, PNAS 85:2444-2448; W. R. Pearson (1996) “Effective protein sequence comparison” Meth. Enzvmol. 266:227- 258; and Pearson et. al. (1997) Genomics 46:24-36 and is publicly available from www.fasta.bioch.virginia.edu/fasta www2/fasta down. shtml or www. ebi.ac.uk/Tools/sss/fasta. Alternatively, a public server accessible at fasta.bioch.virginia.edu/fasta_www2/index.cgi can be used to compare the sequences, using the ggsearch (global proteimprotein) program and default options (BLOSUM50; open: -10; ext: -2; Ktup = 2) to ensure a global, rather than local, alignment is performed. Percent amino acid identity is given in the output alignment header.

[0072] The term “pharmaceutical composition” or “pharmaceutical formulation” refers to a preparation which is in such form as to permit the biological activity of an active ingredient contained therein to be effective, and which contains no additional components which are unacceptably toxic to a subject to which the pharmaceutical composition would be administered. [0073] A “pharmaceutically acceptable earner” refers to an ingredient in a pharmaceutical composition or formulation, other than an active ingredient, which is nontoxic to a subject. A pharmaceutically acceptable earner includes, but is not limited to, a buffer, excipient, stabilizer, or preservative.

[0074] The term “MerTK,” as used herein, refers to any native MerTK from any vertebrate source, including mammals such as primates (e.g. humans) and rodents (e.g., mice and rats), unless otherwise indicated. The term encompasses “full-length,” unprocessed MerTK as well as any form of MerTK that results from processing in the cell. The term also encompasses naturally occurring variants of MerTK, e.g., splice variants or allelic variants. The amino acid sequence of an exemplary human MerTK is described in US 2006/0121562.

[0075] As used herein, “treatment” (and grammatical variations thereof such as “treat” or “treating”) refers to clinical intervention in an attempt to alter the natural course of a disease in the individual being treated, and can be performed either for prophylaxis or during the course of clinical pathology. Desirable effects of treatment include, but are not limited to, preventing occurrence or recurrence of disease, alleviation of symptoms, diminishment of any direct or indirect pathological consequences of the disease, preventing metastasis, decreasing the rate of disease progression, amelioration or palliation of the disease state, and remission or improved prognosis. In some aspects, antibodies of the invention are used to delay development of a disease or to slow the progression of a disease.

[0076] The term “vanable region” or “variable domain” refers to the domain of an antibody heavy or light chain that is involved in binding the antibody to antigen. The variable domains of the heavy chain and light chain (VH and VL, respectively) of a native antibody generally have similar structures, with each domain comprising four conserved framework regions (FRs) and three complementary determining regions (CDRs). (See, e.g., Kindt et al. Kuby Immunology, 6^th ed., W.H. Freeman and Co., page 91 (2007).) A single VH or VL domain may be sufficient to confer antigenbinding specificity. Furthermore, antibodies that bind a particular antigen may be isolated using a VH or VL domain from an antibody' that binds the antigen to screen a library' of complementary VL. or VH domains, respectively. See, e.g., Portolano et al., J. Immunol. 150:880-887 (1993); Clarkson et al.. Nature 352:624-628 ( 1991),

[0077] The term “vector”, as used herein, refers to a nucleic acid molecule capable of propagating another nucleic acid to which it is linked. The term includes the vector as a selfreplicaring nucleic acid structure as well as the vector incorporated into the genome of a host cell into which it lias been introduced. Certain vectors are capable of directing the expression of nucleic acids to which they are operatively linked. Such vectors are referred to herein as “expression vectors”. II. COMPOSITIONS AND METHODS

[0078] In one aspect, the invention is based, in part, on the observation that MeiTK antibody treatment can lead to on-target retinal toxicity (in particular, cells in the photoreceptor and outer nuclear layers) in preclinical primate and mouse models. As disclosed herein, increasing the hydrodynamic radius of anti-MerTK antibodies (e.g., by conjugating one or more polyethylene glycol, PEG, poly inerts) to the antibody) can reduce ocular distribution following systemic administration, thereby reducing distribution to RPE cells and resultant retinal toxicity. In certain aspects, PEGylated antibodies that bind to MerTK are provided. PEGylated antibodies of the invention are useful, e.g., for the treatment of cancer.

[0079] C-Mer proto-oncogene tyrosine kinase (MerTK) is a receptor tyrosine kinase which transduces extracellular signals upon binding to various ligands, such as galectin-3, Protein S, and Gas6, thus activating expression of effector genes. The MerTK pathway regulates essential cellular processes, including cell survival, cytokine production, migration, differentiation, and phagocytosis (Cabernoy N., et al. J Cell Physio. 22' 1 (2012): 401-407; Wu, G., et al. Cell Death & Disease 8 (2017): e2700). Expression of MerTK is found in a variety of hematopoeietic cell types, such as macropliages, dendritic cells, natural killer (NK) cells. Importantly, the MerTK receptor pathway is active in several solid and hematological cancers, including colon cancer (Wu, G., et al. Cell Death & Disease 8 (2017): e2700).

[0080] However, MerTK is also expressed in the retinal pigmented epithelium and is required for turnover of the photoreceptor outer segments (POS) critical for vision. Indeed, mutations in MerTK have been found in patients with retinal dystrophies.

[0081] The MerTK receptor is composed of an extracellular component, a transmembrane (TM) domain, and an intracellular component. As shown in the diagram below, the extracellular or ligandbinding region of MerTK contains two immunoglobulin (Ig)-like domains and two fibronectin (FN) type Ill-like domains.

[0082] In human MerTK, for example, the two Ig-like domains are defined by amino acid residues 76-195 and amino acid residues 199-283, respectively. Additionally, the two fibronectin-Eke domains of human MerTK are defined by amino acid residues 286-384 and amino acid residues 388- 480, respectively. The intracellular region of MerTK contains a tyrosine kinase (TK) domain, which autophosphorylates specific ty rosine residues following ligand binding to the extracellular region and facilitates MerTK receptor dimerization, thus activating downstream effector gene expression (Toledo, R. A, et al. Clin Can. Res. 22 (2016): 2301-2.312). Human MerTK comprises the amino acid sequence: MGPAPLPLLLGLFLPALWRRAITEAREEAKPYPLFPGPFPGSLQTDHTPLLSLPHASGYQPAL MFSPTQPGRPHTGNVAIPQVTSVESKPLPPLAFKHTVGHIILSEHKGVKFNCSISVPNIYQDTTI SWWKDGKELLGAHHAITQFYPDDEVTAITASFSITSVQRSDNGSYICKMKINNEEIVSDPIYTEV QGLPHFTKQPESMNVTRNTAFNIYCQAVGPPEPVNIFWVQNSSRVNEQPEKSPSVT.TVPGLTE MAVFSCEAHNDKGLTVSKGVQINIKAIPSPPTEVSIRNSTAHSILISWVPGFDGYSPFRNCSIQV KE ADPLSNG SVMIFNTS ALPHLYQIKQLQ AL ANYSIGVSCMNEIGWS A VSPWIL A STTEG APS V /WLNVTVFLNESSDNVDIRWMKPPT 'KQQDGELVGYRISHVWQSAGISKELLEEVGQNGSR ARISVQ\aiNATCT\aUAAVTRGGVGPFSDP\⁷KIFIPAI-IGWVDYzAPSSTPAPGNADPVLIIFGCF CGFILIGLILYISLAIRKRVQETKFGNAFTEEDSELVVNYIAKKSFCRRAIELTLHSLGVSEELQN KLEDVVIDRNLLILGKILGEGEFGSVMEGNLKQEDGTSLKVAVKTMKLDNSSQREIEEFLSEA ACMKDFSHPNVIRLLGVCIEMSSQGIPKPMVILPFMKYGDLHTYLLYSRLETGPKH1PLQTLL KFMVDIALGMEYLSNRNFLHRDLAARNCMLRDDMTVCVADFGLSKKIYSGDYYRQGRIAK MPVKWIAIESLADRVYTSKSDVWAFGVTMWEIATRGMTPYPGVQNHEMYDYLLHGHRLKQ PEDCLDELYEIMYSCWRTDPLDRPTFSVLRLQLEKLLESLPDVRNQADVIYVNTQLLESSEGL AQGSTLAPLDLNIDPDSTIASCTPRAAISWTAEVHDSKPHEGRYILNGGSEEWEDLTSAPSAA VTAEKNSVLPGERLVRNGVSWSHSSMLPLGSSLPDELLFADDSSEGSEVLM (SEQ ID NO: 42).

A. Exemplary Anti-MerTK Antibodies

[0083] In one aspect, the invention provides antibodies (e.g., PEGylated antibodies) that bind to

MerTK. In one aspect, provided are isolated antibodies that bind to MerTK. In one aspect, the invention provides antibodies that specifically bind to MerTK. In certain aspects, the anti-MerTK antibodies are PEGylated, i.e., are conjugated to one or more (e.g., i or 2) PEG polymers.

[0084] In one aspect, the invention provides an anti-MerTK antibody comprising at least one, at least two, at least three, at least four, at least five, or all six CDRs selected from (a) CDR-H1 comprising the amino acid sequence of SYAMG (SEQ ID NO: 1); (b) CDR-H2 comprising the amino acid sequence of IINSYGNTYYANWAKG (SEQ ID NO: 2); (c) CDR-H3 comprising the amino acid sequence of DPGVSSNL (SEQ ID NO: 3); (d) CDR-L1 comprising the amino acid sequence of QASQNIYSGLA (SEQ ID NO: 4); (e) CDR-L2 comprising the amino acid sequence of GASKLAS (SEQ ID NO: 5); and (f) CDR-L3 comprising the amino acid sequence of QATYYSSNSVA (SEQ ID NO: 6).

[0085] In one aspect, the invention provides an antibody comprising at least one, at least two, or all three VH CDR sequences selected from (a) CDR-HI comprising the amino acid sequence of SEQ ID NO: I; (b) CDR-H2 comprising the amino acid sequence of SEQ ID NO:2; and (c) CDR-H3 comprising the amino acid sequence of SEQ ID NOG. In one aspect, the antibody comprises CDR- H3 comprising the amino acid sequence of SEQ ID NO:3. In another aspect, the antibody comprises CDR-H3 comprising the amino acid sequence of SEQ ID NOG and CDR-L3 comprising the amino acid sequence of SEQ ID NOE. In a further aspect, the antibody comprises CDR-H3 comprising the amino acid sequence of SEQ ID NOG, CDR-L3 comprising the amino acid sequence of SEQ ID NOE, and CDR-H2 comprising the amino acid sequence of SEQ ID NO:2. In a further aspect, the antibody comprises (a) CDR-H1 comprising the amino acid sequence of SEQ ID NO: 1; (b) CDR-H2 comprising the amino acid sequence of SEQ ID NO:2; and (c) CDR-H3 comprising tire amino acid sequence of SEQ ID NOG.

[0086] In another aspect, the invention provides an antibody comprising at least one, at least two, or all three VL CDR sequences selected from (a) CDR-L1 comprising the amino acid sequence of SEQ ID NO:4; (b) CDR-L2 comprising the amino acid sequence of SEQ ID NO:5; and (c) CDR-L3 comprising the amino acid sequence of SEQ ID NOE. In one aspect, the antibody comprises (a) CDR-L1 comprising the amino acid sequence of SEQ ID NO:4; (b) CDR-L2 comprising the amino acid sequence of SEQ ID NO:5; and (c) CDR-L3 comprising the amino acid sequence of SEQ ID NOE.

[0087] In another aspect, the invention provides an anti-MerTK antibody' comprising at least one, at least two, at least three, at least four, at least five, or ah six CDRs selected from (a) CDR-HI comprising the amino acid sequence of ANTMN (SEQ ID NO: 7); (b) CDR-H2 comprising the amino acid sequence of IFTATGSTYYATWVNG (SEQ ID NO: 8); (c) CDR-I-I3 comprising the amino acid sequence of SGSGSSSGAFNI (SEQ ID NO: 9); (d) CDR-L1 comprising the amino acid sequence of QASQSISSSLA (SEQ ID NO: 10); (e) CDR-L2 comprising the amino acid sequence of AASILAS (SEQ ID NO: 11); and (f) CDR-L3 comprising the amino acid sequence of QCTSYGSLFLGP (SEQ ID NO: 12).

[0088] In one aspect, the invention provides an antibody comprising at least one, at least two, or all three VH CDR sequences selected from (a) CDR-HI comprising the amino acid sequence of SEQ ID NO:7; (b) CDR-H2 comprising the amino acid sequence of SEQ ID NO:8; and (c) CDR-H3 comprising the amino acid sequence of SEQ ID NON. In one aspect, the antibody comprises CDR- H3 comprising the amino acid sequence of SEQ ID NON. In another aspect, the antibody' comprises CDR-H3 comprising the amino acid sequence of SEQ ID NO:9 and CDR-L3 comprising the amino acid sequence of SEQ ID NO: 12. In a further aspect, the antibody' comprises CDR-H3 comprising the amino acid sequence of SEQ ID NON, CDR-L3 comprising the amino acid sequence of SEQ ID NO: 12, and CDR-H2 comprising the amino acid sequence of SEQ ID NOG. In a further aspect, the antibody comprises (a) CDR-HI comprising the amino acid sequence of SEQ ID NO:7; (b) CDR-H2 comprising the amino acid sequence of SEQ ID NOG; and (c) CDR-H3 comprising the amino acid sequence of SEQ ID NON.

[0089] In another aspect, the invention provides an antibody comprising at least one, at least two, or ail three VL CDR sequences selected from (a) CDR-LI comprising the amino acid sequence of SEQ ID NO: 10; (b) CDR-L2 comprising the amino acid sequence of SEQ ID NO:I 1; and (c) CDR-L3 comprising the amino acid sequence of SEQ ID NO: 12. In one aspect, the antibody comprises (a) CDR-L1 comprising the amino acid sequence of SEQ ID NO: 10; (b) CDR-L2 comprising the amino acid sequence of SEQ ID NO: i 1; and (c) CDR-L3 comprising the amino acid sequence of SEQ ID NO: 12.

[0090] In any of the aspects provided herein, an anti-MerTK antibody is humanized. In one aspect, an anti-MerTK antibody further comprises an acceptor human framework, e.g. a human immunoglobulin framework or a human consensus framework,

[0091] In another aspect, an anti-MerTK antibody further comprises a VII or VL comprising an FR1, FR2, FR3, or FR4 sequence of SEQ ID NOs:13 or 14, respectively.

EVQLVESGEGLVQPGGSLRLSCAASGFSLSSYAMGWVRQAPGKGLEYVGIINSYGNTYYAN WAKGRFTISRDNSKNTVYLQMGSLRAEDMAVYYCARDPGVSSNLWGRGTLVTVSS (SEQ ID NO: 13)

DIQMTQSPSTLSASVGDRVTITCQASQNIYSGLAWYQQKPGKAPKLLIYGASKLASGVPSRFS GSGSGTEFTLTISSLQPDDFATYYCQATYYSSNSVAFGGGTKVEIK (SEQ ID NO: 14)

[0092] In another aspect, an anti-MerTK antibody comprises one or more of the CDR sequences of the VII of SEQ ID NO: 13. In another embodiment, an anti-MerTK antibody comprises one or more of tire CDR sequences of the VL of SEQ ID NO: 14. In another embodiment, an anti-MerTK antibody comprises the CDR sequences of the VH of SEQ ID NO: 13 and the CDR sequences of the VL of SEQ ID NO: 14.

[0093] In a further aspect, an anti-MerTK antibody comprises the CDR-H1, CDR-H2 and CDR-

H3 amino acid sequences of the VH domain of SEQ ID NO: 13 and the CDR-L1 , CDR-L2 and CDR- L3 amino acid sequences of the VL domain of SEQ ID NO: 14.

[0094] In one aspect, an anti-MerTK antibody comprises one or more of the heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO: 13 and a framework of at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% sequence identity to the framework amino acid sequence of the VH domain of SEQ ID NO: 13. In one aspect, the anti-MerTK antibody comprises the three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO: 13 and a framework of at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% sequence identity to the framework ammo acid sequence of the VH domain of SEQ ID NO: 13. In one aspect, the anti-MerTK antibody comprises the three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO: 13 and a framework of at least 95% sequence identity to the framework amino acid sequence of the VH domain of SEQ ID NO: 13. In another aspect, the anti-MerTK antibody comprises the three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO: 13 and a framework of at least of at least 98% sequence identity’ to the framework amino acid sequence of the VH domain of SEQ ID NO: 13.

[0095] In one aspect, an anti-MerTK antibody comprises one or more of the light drain CDR amino acid sequences of the VL domain of SEQ ID NO: 14 and a framework of at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% sequence identity to the framework amino acid sequence of the VL domain of SEQ ID NO: 14. In one aspect, the anti-MerTK antibody comprises the three light chain CDR amino acid sequences of the VL domain of SEQ ID NO: 14 and a framework of at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% sequence identity to the framework amino acid sequence of the VL domain of SEQ ID NO: 14. In one aspect, the anti-MerTK antibody comprises the three light chain CDR amino acid sequences of the VL domain of SEQ ID NO: 14 and a framework of at least 95% sequence identity to the framework amino acid sequence of the VL domain of SEQ ID NO: 14. In another aspect, the anti-MerTK antibody comprises the three light chain CDR amino acid sequences of the VL domain of SEQ ID NO: 14 and a framework of at least particularly of at least 98% sequence identity to the framework amino acid sequence of the VH domain of SEQ ID NO: 13.

[0096] In one aspect, the anti-MerTK antibody comprises (a) CDR-H1 comprising the amino acid sequence of SEQ ID NO: 1; (b) CDR-H2 comprising the amino acid sequence of SEQ ID NO:2;

(c) CDR-H3 comprising the amino acid sequence of SEQ ID NO:3; (d) CDR -LI comprising the amino acid sequence of SEQ ID NO:4; (e) CDR-L2 comprising the amino acid sequence of SEQ ID NO:5; and (f) CDR-L3 comprising the amino acid sequence of SEQ ID NO:6, and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 13, and a VL domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 14. In one aspect, the VII domain lias at least 95% sequence identity to the amino acid sequence of SEQ ID NO:13. In one aspect, the VL domain has at least 95% sequence identity to the amino acid sequence of SEQ ID NO: 14.

[0097] In one aspect, the anti-MerTK antibody comprises (a) CDR-H1 comprising the amino acid sequence of SEQ ID NO: 1; (b) CDR-H2 comprising the amino acid sequence of SEQ ID NO:2: (c) CDR-H3 comprising the amino acid sequence of SEQ ID NO: 3: (d) CDR-L1 comprising the amino acid sequence of SEQ ID NO:4; (e) CDR-L2 comprising the amino acid sequence of SEQ ID NO:5; and (f) CDR-L3 comprising the amino acid sequence of SEQ ID NO:6, and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 13, and a VL domain having at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 14; wherein the antibody specifically binds to MerTK. In one aspect, the VH domain lias at least 95% sequence identity to the amino acid sequence of SEQ ID NO: 13. In one aspect, the VL domain lias at least 95% sequence identity to the amino acid sequence of SEQ ID NO: 14.

[0098] In another aspect, an anti-MerTK antibody comprises a heavy chain variable domain (VH) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 13. In one aspect, an anti-MerTK antibody comprises a heavy chain variable domain (VH) sequence having at least 95%, sequence identity to the amino acid sequence of SEQ ID NO: 13. In certain aspects, a VH sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, but an anti- MerTK antibody comprising that sequence retains the ability to bind to MeiTK. In certain aspects, a total of 1 to 10 amino acids have been substituted, inserted and/or deleted in SEQ ID NO: 13. In certain aspects, substitutions, insertions, or deletions occur in regions outside the CDRs (i.e. , in the FRs). Optionally, the anti-MerTK antibody comprises the VH sequence in SEQ ID NO: 13, including post-translational modifications of that sequence. In a particular aspect, the VH comprises one, two or three CDRs selected from: (a) CDR-H1, comprising die amino acid sequence of SEQ ID NO: 1, (b) CDR-H2, comprising the amino acid sequence of SEQ ID NO:2, and (c) CDR-H3, comprising the amino acid sequence of SEQ ID NO:3. In another aspect, an anti-MerTK antibody is provided, wherein the antibody comprises a light chain variable domain (VL) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 14. In one aspect, an anti-MerTK antibody comprises a light chain variable domain (VL) sequence having at least 95% sequence identity’ to the amino acid sequence of SEQ ID NO: 14. In certain aspects, a VL sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, but an anti-MerTK antibody comprising that sequence retains the ability to bind to MerTK. In certain aspects, a total of 1 to 10 amino acids have been substituted, inserted and/or deleted in SEQ ID NO: 14. In certain aspects, the substitutions, insertions, or deletions occur in regions outside the CDRs (i.e., in the FRs). Optionally , the anti-MerTK antibody comprises the VL sequence in SEQ ID NO: 14, including post-translational modifications of that sequence. In a particular aspect, the VL comprises one, two or three CDRs selected from: (a) CDR- Ll, comprising the amino acid sequence of SEQ ID NO:4, (b) CDR-L2, comprising the amino acid sequence of SEQ ID NO:5, and (c) CDR-L3, comprising the amino acid sequence of SEQ ID NO:6. [0099] In another aspect, an anti-MerTK antibody is provided, wherein the antibody comprises a

VH sequence as in any of the aspects provided above, and a VL sequence as in any of the aspects provided above. In one aspect, the antibody comprises the VH and VL sequences in SEQ ID NO: 13 and SEQ ID NO: 14, respectively, including post-translational modifications of those sequences. [0100] In another aspect, an anti-MerTK antibody further comprises a VH or VL comprising an

FR1, FR2, FR3, or FR4 sequence of SEQ ID NOs:15 or 16, respectively.

EQQLVESGEGLVQPGGSLRLSCAVSGFSLSSYAMGWVRQAPGKGLEWIGIINSYGNTYYAN WAKGRFTISRDSSKNTVYLQMGSLRAEDMAVYFCARDPGVSSNLWGPGTLVTVSS (SEQ ID NO: 15)

DVQMTQSPSTLSASVGDRVTITCQASQNIYSGLAWYQQKPGKPPKLLIYGASKLASGVPSRFS GSGSGTEFTLTISSLQPDDFATYYCQATYYSSNSVAFGGGTKVEIK (SEQ ID NO: 16) [0101] In another aspect, an anti-MerTK antibody comprises one or more of the CDR sequences of the VH of SEQ ID NO: 15. In another embodiment, an anti-MerTK antibody comprises one or more of the CDR sequences of the VL of SEQ ID NO: 16. In another embodiment, an anti-MerTK antibody comprises the CDR sequences of the VH of SEQ ID NO: 15 and the CDR sequences of the VL of SEQ ID NO: 16.

[0102] In a further aspect, an anti-MerTK antibody comprises the CDR-H1 , CDR-H2 and CDR- H3 amino acid sequences of the VH domain of SEQ ID NO: 15 and the CDR-L1, CDR-L2 and CDR- L3 amino acid sequences of the VL domain of SEQ ID NO: 16.

[0103] In one aspect, an anti-MerTK antibody comprises one or more of the heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO: 15 and a framework of at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% sequence identity to the framework amino acid sequence of the VH domain of SEQ ID NO: 15. In one aspect, the anti-MerTK antibody comprises the three heavy drain CDR amino acid sequences of the VH domain of SEQ ID NO: 15 and a framework of at least 85%, 86%, 87%, 88%, 89%, 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%,, 98%,, 99% sequence identity to the framework amino acid sequence of the VH domain of SEQ ID NO: 15. In one aspect, the anti-MerTK antibody comprises the three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO: 15 and a framework of at least 95%> sequence identity to the framework amino acid sequence of the VH domain of SEQ ID NO: 15. In another aspect, the anti-MerTK antibody comprises the three heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO: 15 and a framework of at least of at least 98% sequence identity to the framework amino acid sequence of die VII domain of SEQ ID NO: 15.

[0104] In one aspect, an anti-MerTK antibody comprises one or more of the light chain CDR amino acid sequences of the VL domain of SEQ ID NO: 16 and a framework of at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93% 94%, 95%, 96%, 97%, 98%, 99% sequence identity to the framework amino acid sequence of the VL domain of SEQ ID NO: 16. In one aspect, the anti-MerTK antibody comprises the three light chain CDR amino acid sequences of the VL domain of SEQ ID NO: 16 and a framework of at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% sequence identity to the framework amino acid sequence of the VL domain of SEQ ID NO: 16. In one aspect, the anti-MerTK antibody comprises the three light chain CDR amino acid sequences of the VL domain of SEQ ID NO: 16 and a framework of at least 95% sequence identity to the framework amino acid sequence of the VL domain of SEQ ID NO: 16. In another aspect, the anti-MerTK antibody comprises the three light chain CDR amino acid sequences of the VL domain of SEQ ID NO: 16 and a framework of at least particularly of at least 98% sequence identity to the framework amino acid sequence of the VH domain of SEQ ID NO: 15.

[0105] In one aspect, the anti-MerTK antibody comprises (a) CDR-H1 comprising the amino acid sequence of SEQ ID NO: 1: (b) CDR-H2 comprising the amino acid sequence of SEQ ID NO:2; (c) CDR-H3 comprising the amino acid sequence of SEQ ID NO:3; (d) CDR-L1 comprising the amino acid sequence of SEQ ID NO:4; (e) CDR-L2 comprising the amino acid sequence of SEQ ID NO:5; and (f) CDR-L3 comprising the amino acid sequence of SEQ ID NO:6, and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 15, and a VL domain having at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 16. In one aspect, the VH domain has at least 95% sequence identity to the amino acid sequence of SEQ ID NO: 15. In one aspect, the VL domain Isas at least 95% sequence identity to the amino acid sequence of SEQ ID NO: 16.

[0106] In one aspect, the anti-MeiTK antibody comprises (a) CDR-H1 comprising the amino acid sequence of SEQ ID NO:1; (b) CDR-H2 comprising the amino acid sequence of SEQ ID NO:2; (c) CDR-H3 comprising the amino acid sequence of SEQ ID NO:3; (d) CDR-L1 comprising the amino acid sequence of SEQ ID NO:4; (e) CDR-L2 comprising the amino acid sequence of SEQ ID NO:5; and (f) CDR-L3 comprising the amino acid sequence of SEQ ID NO:6, and a VH domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 15, and a VL. domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 16; wherein the antibody specifically binds to MerTK. In one aspect, the VH domain has at least 95% sequence identity to the amino acid sequence of SEQ ID NO: 15. In one aspect, the VI. domain has at least 95% sequence identity to the amino acid sequence of SEQ ID NO: 16.

[0107] In another aspect, an anti-MerTK antibody comprises a heavy chain variable domain (VH) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 15. In one aspect, an anti-MerTK antibody comprises a heavy chain variable domain (VH) sequence having at least 95%, sequence identity to the amino acid sequence of SEQ ID NO: 15. In certain aspects, a VH sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, but an anti- MerTK antibody comprising that sequence retains the ability to bind to MerTK. In certain aspects, a total of I to 10 amino acids have been substituted, inserted and/or deleted in SEQ ID NO: 15. In certain aspects, substitutions, insertions, or deletions occur in regions outside the CDRs (i.e., in the FRs). Optionally, the anti-MerTK antibody comprises the VH sequence in SEQ ID NO: 13, including post-translational modifications of that sequence. In a particular aspect, the VH comprises one, two or three CDRs selected from: (a) CDR-H1, comprising the amino acid sequence of SEQ ID NO:1, (b) CDR-H2, comprising the amino acid sequence of SEQ ID NOV, and (c) CDR-H3, comprising the amino acid sequence of SEQ ID NO:3. In another aspect, an anti-MerTK antibody is provided, wherein the antibody comprises a light chain variable domain (VL) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequence of SEQ ID NO: 16. In one aspect, an anti-MerTK antibody comprises a light chain variable domain (VL) sequence having at least 95% sequence identity to the amino acid sequence of SEQ ID NO: 16. In certain aspects, a VL sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, but an anti-MerTK antibody comprising that sequence retains the ability to bind to MerTK. In certain aspects, a total of 1 to 10 amino acids have been substituted, inserted and/or deleted in SEQ ID NO: 16. In certain aspects, the substitutions, insertions, or deletions occur in regions outside the CDRs (i.e., in the FRs). Optionally, the anti-MerTK antibody comprises the VL sequence in SEQ ID NO: 16, including post-translational modifications of that sequence. In a particular aspect, the VL comprises one, two or three CDRs selected from: (a) CDR- Ll, comprising the amino acid sequence of SEQ ID NO:4, (b) CDR-L2, comprising the amino acid sequence of SEQ ID NO:5, and (c) CDR-L3, comprising the amino acid sequence of SEQ ID NO:6. [0108] In another aspect, an anti-MerTK antibody is provided, wherein the antibody comprises a VH sequence as in any of the aspects provided above, and a VL. sequence as in any of the aspects provided above. In one aspect, the antibody comprises the VH and VL sequences in SEQ ID NO: 15 and SEQ ID NO: 16, respectively, including post-translational modifications of those sequences.

[0109] In another aspect, an anti-MerTK antibody comprises one or more of the CDR sequences of the VH of SEQ ID NO: 17. In another embodiment, an anti-MerTK antibody comprises one or more of the CDR sequences of the VL of SEQ ID NO: 18. In another embodiment, an anti-MerTK antibody comprises the CDR sequences of the VH of SEQ ID NO: 17 and the CDR sequences of the VL of SEQ ID NO: 18.

QSVEESGGRLVTPGTPLTLTCTVSGFSLSSYAMGWVRQAPGKGLEWIGIINSYGNTYYANWA KGRFTISRTSTTVDLRMPSLTTEDTATYFCARDPGVSSNLWGPGTLVTVSS (SEQ ID NO: 17)

DWMTQTPASVSEPVGGTVTIKCQASQNIYSGLAWYQQKPGQPPKLLIYGASKLASGVSSRF KGSGSGTEFTLTISDLECADAATYYCQATYYSSNSVAF'GGGTEVVVK (SEQ ID NO: 18)

[0110] In a further aspect, an anti-MerTK antibody comprises the CDR-H1, CDR-H2 and CDR- H3 amino acid sequences of the VH domain of SEQ ID NO: 17 and the CDR-I., 1 , CDR-L2 and CDR- L3 amino acid sequences of the VL domain of SEQ ID NO: 18.

[0111] In one aspect, an anti-MerTK antibody comprises one or more of the heavy chain CDR amino acid sequences of the VH domain of SEQ ID NO: 17 and one, two, three, or four human framework sequences (e.g., HC-FR1, HC-FR2, HC-FR3, and/or HC-FR4). In one aspect, an anti- MerTK antibody comprises one or more of the light chain CDR amino acid sequences of the VL domain of SEQ ID NO: 18 and one, two, three, or four human framework sequences (e.g., LC-FRI, LC-FR2, LC-FR3, and/or LC-FR4).

[0112] In another aspect, an anti-MerTK antibody is provided, wherein the antibody comprises a VH sequence as in any of the aspects provided above, and a VL sequence as in any of the aspects provided above. In one aspect, the antibody comprises the VH and VL sequences in SEQ ID NO: 17 and SEQ ID NO: 18, respectively, including post-translational modifications of those sequences.

[0113] In another aspect, an anti-MerTK antibody comprises one or more of the CDR sequences of the VH of SEQ ID NO: 19. In another embodiment, an anti-MerTK antibody comprises one or more of the CDR sequences of the VL of SEQ ID NO:20, In another embodiment, an anti-MerTK antibody comprises the CDR sequences of the VH of SEQ ID NO: 19 and the CDR sequences of the VL of SEQ ID NO:20.

QSVEESGGRLVTPGTPLTLTCTVSGIDLSANTMNWVRQAPGKGLEWIGIFTATGSTYYATWV NGRFTISKTSTTVDLKITSPTTEDTATYFCARSGSGSSSGAFNIWGPGTLVTVSL (SEQ ID NO: 19)

DPVLTQTPASVSEPVGGTVTTKCQASQSISSSLAWYQQKPGQPPKLLIYAASILASEISSRFKGS RSGTEFTLTISDLECADAATYYCQCTSYGSLFLGPFGGGTEVWK (SEQ ID NO: 20)

[0114] In a further aspect, an anti-MerTK antibody comprises the CDR-H1, CDR-H2 and CDR- H3 amino acid sequences of the VH domain of SEQ ID NO: 19 and the CDR-L1, CDR-L2 and CDR- L3 amino acid sequences of the VL domain of SEQ ID NO: 20.

[0115] In one aspect, an anti-MerTK antibody comprises one or more of the heavy drain CDR amino acid sequences of the VH domain of SEQ ID NO: 19 and one, two, three, or four human framework sequences (e.g., HC-FR1, HC-FR2, HC-FR3, and/or HC-FR4). In one aspect, an anti- MerTK antibody comprises one or more of the light chain CDR amino acid sequences of the VL domain of SEQ ID NQ:20 and one, two, three, or four human framework sequences (e.g., LC-FRl, LC-FR2, LC-FR3, and/or LC-FR4).

[0116] In another aspect, an anti-MerTK antibody is provided, wherein the antibody comprises a VH sequence as in any of the aspects provided above, and a VL sequence as in any of the aspects provided above. In one aspect, the antibody comprises the VH and VL sequences in SEQ ID NO: 19 and SEQ ID NO:20, respectively, including post-translational modifications of those sequences.

[0117] In a further aspect of the invention, an anti-MerTK antibody according to any of the above aspects is a monoclonal antibody, including a chimeric, humanized or human antibody. In one aspect, an anti-MerTK antibody is an antibody fragment, e.g., a Fv, Fab, Fab’, scFv, diabody, or F(ab’)2 fragment.

[0118] In another aspect, the antibody is a full length antibody, e.g., an intact IgGl antibody or other antibody class or isotype as defined herein.

[0119] In one aspect, additionally the C-terminal glycine (Gly446) is present. In one aspect, additional!}' the C-terminal lysine (Lys447) is present. In one aspect, additionally the C -terminal glycine (Gly446) and the C-terminal lysine (Lys447) is present.

[0120] In certain embodiments, the antibody comprises at least one mutation in the Fc region that reduces binding to Fc receptors and/or complement. In one embodiment, the antibody comprises a set of mutations in the Fc region referred to as LALAPG (L234A, L235 A, and P329G, numbering according to EU index). LALAPG and other Fc mutations are described further below and, for example, in U.S. Patent No. 8,969,526.

[0121] In one aspect, the anti-MerTK antibody comprises a heavy chain comprising the sequence EVQIAT’SGEGLVQPGGSLRLSCAASGFSLSSYAMGWVRQAPGKGLEYVGnNSYGNTYYAN WAKGRFTISRDNSKNrVYLQMGSLRAEDMAVYYCARDPGVSSNLWGRGTLVTVSSASTKG PSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSV VTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEAAGGPSVFLFPPKPK DTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVL HQDWLNGKEYKCKVSNKALGAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVK GFYPSD1AVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEAL HNHYTQKSLSLSPG (SEQ ID NO: 21).

[0122] In one aspect, the anti-MerTK antibody comprises an Fc region (e.g., a human IgGl Fc region) comprising an N297G mutation, numbering according to EU index.

[0123] In one aspect, the anti-MerTK antibody comprises a heavy chain comprising the sequence EVQLVESGEGLVQPGGSLRLSCAASGFSLSSYAMGWVRQAPGKGLEYVGIINSYGNTYYAN WAKGRFTISRDNSKNTVAG,QMGSLRAEDMAVYYCARDPGVSSNLWGRGTLVTVSSASTKG PSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSV VTWSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPK DTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYGSTYRVVSVLTVL I-IQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKG FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALH NH YTQKSLSLSPG (SEQ ID NO:22).

[0124] In one aspect, the anti-MerTK antibody comprises a light chain comprising the sequence DIQMTQSPSTLSASVGDRVTITCQASQNIYSGLAWYQQKPGKAPKLLIYGASKLASGVPSRFS GSGSGTEFTLTISSLQPDDFATYYCQATYYSSNSVAFGGGTKVEIKRTVAAPSVFIFPPSDEQL KSGTASWCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADY EKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 23).

[0125] In one aspect, the anti-MerTK antibody comprises a heavy chain comprising the sequence of SEQ ID NO:21 and a light chain comprising the sequence of SEQ ID NO:23. In one aspect, the anti-MerTK antibody comprises a heavy chain comprising the sequence of SEQ ID NO:22 and a light chain comprising the sequence of SEQ ID NO: 23.

B. PEG Polymers [0126] Certain aspects of the present disclosure relate to PEGylated antibodies that bind to MerTK. Any of the anti-MerTK antibodies of the present disclosure (e.g, as described supra} may be PEGylated.

[0127] As is known in the art, PEGylation of an antibody refers to conjugation (e.g. , chemical coupling) of a polymer of PEG monomers to the antibody. The structure of a PEG monomer is provided below, and PEG polymers are typically expressed as (O-CH2-CH2)„-OCH3, with /? referring to the number of PEG monomers. A variety of PEG polymers suitable for use herein are known in the art. See, e.g., Jevsevar, S. el al. (2010) Biolechnol. J. 5:113-128.

[0128] In some embodiments, the PEG polymer(s) are linear PEG polymers. In some embodiments, the PEG poiymer(s) are branched PEG polymers. In some embodiments, the branched PEG polymer(s) comprise 2 or more branches. In some embodiments, the branched PEG poly mer(s) comprise 2 branches.

[0129] In some embodiments, the PEGylated antibody lias a hydrodynamic radius of greater than about 6 nm. In some embodiments, the PEGylated antibody lias a hydrodynamic radius of greater than or equal to about 10 mu. In some embodiments, the PEGylated antibody has a hydrodynamic radius of between about 9 mu and about 11 nm, between about 6 nm and about 11 nm, or between about 6 nm and about 10 nm.

[0130] In some embodiments, an anti-MerTK antibody of the present disclosure is conjugated to one or more PEG polymer(s). In some embodiments, an anti-MerTK antibody of the present disclosure is conjugated to one or two PEG polymer] s).

[0131] In some embodiments, the PEG polymer(s) each have a molecular weight of between about 10 kDa and about 40 kDa. In some embodiments, the PEG polymer(s) each have a molecular weight of between about 20 kDa and about 40 kDa. In some embodiments, the PEG polymer(s) each have a molecular weight of between about 10 kDa and about 20 kDa. In some embodiments, the PEG poly mer(s) each have a molecular weight of about 10 kDa, about 20 kDa, about 30 kDa, or about 40 kDa.

[0132] In some embodiments, the PEG polymer(s) (e.g. , one or two PEG polymers) are conjugated to an anti-MerTK antibody of the present disclosure at the heavy and/or light chain(s). In some embodiments, the PEG polymer(s) (e.g., one or two PEG polymers) are conjugated to an anti- MerTK antibody of the present disclosure at an engineered cysteine residue of the heavy and/or light chain(s). Engineered cysteine residues are described in greater detail infra.

[0133] In some embodiments, the engineered cysteine residue is selected from the group consisting of K149C of the light chain, K I83C of the light chain, T186C of the heavy chain, and Y373C of the heavy chain (numbering of the light chain according to Kabat and numbering of the heavy chain according to EU index). For example, in some embodiments, the antibody comprises two heavy chains, two light chains, and two PEG polymers; and both light chains of the antibody comprise a K149C engineered cysteine conjugated to one of the two PEG polymers. In some embodiments, the antibody comprises two heavy chains, two light chains, and two PEG poly mers; and both light chains of the antibody comprise a K183C engineered cysteine conjugated to one of the two PEG poly mers. In some embodiments, the antibody comprises two heavy chains, two light chains, and two PEG polymers; and both heavy chains of the antibody comprise a T186C engineered cysteine conjugated to one of the two PEG polymers. In some embodiments, the antibody comprises two heavy chains, two light chains, and two PEG polymers; and both heavy chains of the antibody comprise a Y373C engineered cysteine conjugated to one of the two PEG polymers.

[0134 ] Certain aspects of the present disclosure rela te to methods of producing PEGylated antibodies that bind to MerTK. Any of the anti-MerTK antibodies of the present disclosure (e.g. , as described supra) may find use in the methods disclosed herein. In some embodiments, PEG polvmer(s) are conjugated to a heavy chain or light chain of an anti-MerTK antibody of the present disclosure at an engineered cysteine residue via maleimide-cysteine conjugation. In some embodiments, PEG polymer(s) are conjugated to a heavy chain or light chain of an anti-MerTK antibody of the present disclosure at an engineered cysteine residue via iodoacetamide-cysteine conjugation.

[0135] In some embodiments, the methods comprise contacting (i.e., convalently attaching) an antibody comprising one or more free engineered cysteine residues with one or more polyethylene glycol (PEG) polymer(s) comprising a maleimide moiety under conditions suitable for each of the PEG polymer(s) to be conjugated to an engineered cysteine residue of the antibody via thioether linkage. In some embodiments, the conjugation is performed at a pH suitable to avoid conjugation to lysines and/or opening of the maleimide ring. In some embodiments, the conjugation is performed at pH 7 to pH 7.5. Non-limiting reaction conditions are exemplified infra. In some embodiments, the conjugation reaction is monitored, e.g., using HPLC and size exclusion chromatography (SEC) to resolve antibody species with different polymer ratios.

[0136] In some embodiments, the methods comprise contacting (i.e., convalently attaching) an antibody comprising one or more free engineered cysteine residues with one or more polyethylene glycol (PEG) polymer(s) comprising an iodoacetamide moiety under conditions suitable for each of the PEG polymer(s) to be conjugated to an engineered cysteine residue of the antibody via thioether linkage.

[0137] In some embodiments, PEG polymers are conjugated to the antibody at a ratio of 2.0 polymers per antibody. Ratio of PEGtantibody can be determined, e.g., using SEC and HPLC.

[0138] In some embodiments, each of the free engineered cysteine residues is blocked with a cysteine or glutathione moiety prior to the conjugation. In some embodiments, the methods further comprise, prior to conjugation, deblocking the free engineered cysteine residues. In some embodiments, deblocking the free engineered cysteine residues is performed by reduction at pH 8.5, e.g., particularly for sites with a higher thiol pKa (including but not limited to K149C of the light chain). In some embodiments, deblocking the free engineered cysteine residues is performed with more reductant for sites with a higher thiol pKa (including but not limited to K149C of the light drain). In some embodiments, the reductant is DTT. Non-limiting reaction conditions are exemplified infra. In some embodiments, after deblocking, the antibody is re-oxidized. In some embodiments, after deblocking, the antibody is purified, e.g. , using cationic exchange chromatography to remove the reductant and the reduced cysteine & glutathione.

[0139] In some embodiments, the methods further comprise purifying the PEGylated antibody from unconjugated antibody and PEG polymer after conjugation. Non-limiting purification methods are exemplified infra. For example, in some embodiments, the PEGylated antibody can be purified hydrophobic interaction chromatography (HIC).

1. MerTK Biological Activity

[0140] In some embodiments, the antibodies (e.g., PEGylated anti-MerTK antibodies) reduce MerTK mediated clearance of apoptotic cells by phagocytes, e.g., the clearance of apoptotic cells is reduced by 1-10 fold, 1-8 foid, 1-5 fold, 1-4 fold, 1-3 fold, 1-2 fold, 2-10 fold, 2-8 fold, 2-5 fold, 2-4 fold, 2-3 fold, 3-10 fold, 3-8 fold, 3-5 fold, 3-4 fold, or by about 1.1 fold, 1.2 fold, 1.3 fold, 1.4 fold,

I.5 fold, 1.6 fold, 1.7 fold, 1.8 fold, 1.9 fold, 2.0 fold, 2.1 fold, 2.2 fold, 2.3 fold, 2.4 fold, 2.5 fold, 2.6 fold, 2.7 fold, 2.8 fold, 2.9 fold, 3.0 fold, 3.1 fold, 3.2 fold, 3.3 fold, 3.4 fold, 3.5 fold, 3.6 fold, 3.7 fold, 3.8 fold, 3.9 fold, 4.0 fold, 4.1 fold, 4.2 fold, 4.3 fold, 4.4 fold, 4.5 fold, 4.6 fold, 4.7 fold, 4.8 fold, 4.9 fold, 5.0 fold, 5.1 fold, 5.2 fold, 5.3 fold, 5.4 fold, 5.5 fold, 5.6 fold, 5.7 fold, 5.8 fold, 5.9 fold, 6.0 fold, 6.1 fold, 6.2 fold, 6.3 fold, 6.4 fold, 6.5 fold, 6.6 fold, 6.7 fold, 6.8 fold, 6.9 fold, 7.0 fold, 7. 1 fold, 7.2 fold, 7.3 fold, 7.4 fold, 7.5 fold, 7.6 fold, 7.7 fold, 7.8 fold, 7.9 fold, or 8.0 fold. In some embodiments, the phagocytes are macrophages. In some such embodiments, the macrophages are tumor-associated macrophages (TAMs). In humans, TAMs may be identified based on expression of various cell-surface markers, including CD 14, HLA-DR (MHC class II), CD312, CD 1 I 5, CD 16, CD163, CD2.04, CD206, and C ID 301. Furthermore, the production of specific functional biomarkers, such as matrix metalloproteinases, IL- 10, inducible nitric oxide synthase (iNOS), TNF -alpha, or IL-12 may be combined with cell-surface biomarkers to accurately identify TAM populations (Quatromoni,

J., et al., Am J Transl Res. 4 (2012): 376-389.) The clearance of apoptotic cells may be measured by any assay known to one of skill in the art for such purpose. For example, for in vitro apoptotic cell clearance assays, phagocytes such as mouse peritoneal macrophages or human monocyte derived macrophages are used. Apoptotic cells are generated by treatment with dexamethasone and labeled with a detection probe. Phagocytosis can be analyzed by microscopy or flow cytometry after incubation apoptotic cells with phagocytes. In some embodiments, the clearance of apoptotic cells is reduced as measured in such an apoptotic cell clearance assay at room temperature. For example, for in vivo apoptotic clearance assays, mice are injected with dexamethasone to induce thymocyte death. Resident macrophages in the thymus recognize and engulf the dying/dead cells (Seitz, H. M. J Immunol. 178(9) 5635-5642 (2007). In some embodiments, the clearance of apoptotic cells is reduced as measured in such an apoptotic cell clearance assay in vivo. In some embodiments, the antibodies reduce ligand-mediated MerTK signaling. In some embodiments, the ligand is hGAS6-Fc (EC50 = ~ 84 pM). In some embodiments, the antibodies induce a pro-inflammatory response. In some embodiments, lite antibodies induce a type I IFN response.

[0141] In some embodiments, an anti-MerTK antibody of the present disclosure (e.g., a PEGylated anti-MerTK antibody) reduces phagocytic activity of apoptotic cells by about 10-100%, 20-100%, 30-100%, 40-100%, 50-100%, 60-100?% 70-100%, 75-100%, 80-100%, 85-100%, 90- 100%, 95-100%, 10-95%, 20-95%, 30-95%, 40-95%, 50-95%, 60-95%, 70-95%, 75-95%, 80-95%, 85-95%, 90-95%, 10-90%, 20-90%, 30-90%, 40-90%, 50-90%, 60-90%, 70-90%, 75-90%, 80-90%, 85-90%, 10-85%, 20-85%, 30-85%, 40-85%, 50-85%, 60-85%, 70-85%, 75-85%, 80-85%, 10-80%, 20-80%, 30-80%, 40-80%, 50-80%, 60-80%, 70-80%, 75-80%, 10-75%, 20-75%, 30-75%, 40-75%, 50-75%, 60-75%, 70-75%, 10-70%, 20-70%, 30-70%, 40-70%, 50-70%, 60-70%, 10-65%, 20-65%, 30-65%, 40-65%, 50-65%, 60-65%, 10-60%, 2.0-60%, 30-60%, 40-60%, 50-60%, 10-55%, 2.0-55%, 30-55%, 40-55%, 50-55%, 10-40%, 20-40%, or 30-40%, or by at least about 10%, 20%, 30%, 40%, 50%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98% or 99%. In some embodiments, tire anti-MerTK antibody has a half maximal inhibitory concentration (IC50) for reducing phagocytic activity of apoptotic cells of about 1 pM - 50 pM, 1 pM - 100 pM, 1 pM - 500 pM, 1 pM - 1 nM, 1 pM - 1.5 nM, 5 pM - 50 pM, 5 pM - 100 pM, 5 pM - 500 pM, 5 pM - 1 nM, 5 pM - 1.5 nM, 10 pM - 50 pM, 10 pM - 100 pM, 10 pM - 500 pM, 10 pM - 1 nM, 10 pM - 1.5 nM, 50 pM - 100 pM, 50 pM - 500 pM, 50 pM - 1 nM, 50 pM - 1.5 nM, 100 pM - 500 pM, 100 pM - 1 nM, or 100 pM - 1.5 nM. Exemplary methods for determining phagocytic activity and IC50 are described in the Examples herein below.

[0142] In some embodiments, an anti-MerTK antibody of the present disclosure (e.g. a PEGylated anti-MerTK antibody) enhances the activity of a checkpoint inhibitor by about 1-2 fold, 1- 5 fold, 1-10 fold, 1-15 fold, 1-20 fold, 1-25 fold, 1-30 fold, 1-50 fold, 1-75 fold, 1-100 fold, 1-150 fold, 1-200 fold, 1-250 fold, 1.5-2. fold, 1.5-5 fold, 1.5-10 fold, 1.5-15 fold, 1.5-20 fold, 1.5-25 fold, 1.5-30 fold, 1.5-50 fold, 1.5-75 fold, 1.5-100 fold, 1.5-150 fold, 1.5-200 fold, 1.5-250 fold, 2-5 fold, 2-10 fold, 2-15 fold, 2-20 fold, 2-25 fold, 2-30 fold, 2-50 fold, 2-75 fold, 2-100 fold, 2-150 fold, 2- 200 fold, 2-250 fold, 2.5-5 fold, 2.5-10 fold, 2.5-15 fold, 2.5-20 fold, 2.5-25 fold, 2.5-30 fold, 2.5-50 fold, 2.5-75 fold, 2.5-100 fold, 2.5-150 fold, 2.5-200 fold, 2.5-250 fold, 5-10 fold, 5-15 fold, 5-20 fold, 5-25 fold, 5-30 fold, 5-50 fold, 5-75 fold, 5-100 fold, 5-150 fold, 5-200 fold, 5-250 fold, 10-15 fold, 10-20 fold, 10-25 fold, 10-30 fold, 10-50 fold, 10-75 fold, 10-100 fold, 10-150 fold, 10-200 fold, 10-250 fold, 20-25 fold, 20-30 fold, 20-50 fold, 20-75 fold, 20-100 fold, 20-150 fold, 20-200 fold, 20- 250 fold, 25-30 fold, 25-50 fold, 25-75 fold, 25-100 fold, 25-150 fold, 25-200 fold, or 25-250 fold or by at least about 1 fold, 2 fold, 5 fold, 10 fold, 15 fold 20 fold 25 fold, 30 fold, 40 fold, 50 fold 60 fold, 70 fold, 75 fold, 80 fold, 90 fold, 100 fold, 125 fold, 150 fold, 200 fold, 225 fold or 250 fold. In certain embodiments, an anti-MerTK antibody of the present disclosure enhances the activity of a checkpoint inhibitor as determined using an assay as described in the Examples herein below, such as, for example, by determining a reduction in tumor volume in a mouse tumor model using a combmation of an anti-MerTK antibody plus a checkpoint inhibitor as compared to the reduction in tumor volume using the checkpoint inhibitor alone. In certain embodiments, the reduction in tumor volume is determined after at least 10 days, 14 days, 20 days, 21 days or 30 days after treatment with the therapeutic agents. In certain embodiments, the checkpoint inhibitor is a anti-PDl axis antagonist. In one exemplary embodiment, the checkpoint inhibitor is an anti-PD-Ll antibody. In another embodiment, the checkpoint inhibitor is an anti-PDl antibody.

[0143] In some embodiments, an anti-MerTK antibody of the present disclosure (e.g., a PEGylated anti-MerTK antibody) increases cell-free DNA (ciDNA) and/or circulating tumor DNA (ctDNA), e.g. , in a blood or plasma sample, by about 1-2 fold, 1 -3 fold, 1-4 fold, 1-5 fold, 1-10 fold, 1.5-2 fold, 1.5-3 fold, 1.5-4 fold, 1.5-5 fold, 1.5-10 fold, 2-3 fold, 2-4 fold, 2-5 fold, 2-10 fold, 3-5 fold, 3-10 fold, 4-5 fold, 4-10 fold, 5-10 fold, or by at least about 1 fold, 2 fold, 3 fold, 4 fold, 5 fold, or 10 fold. In certain embodiments, an anti-MerTK antibody of the present disclosure increases cell- free DNA (cfDNA) and/or circulating tumor DNA (ctDNA) as determined using an assay as described in the Examples herein below, such as, for example, by isolating cfDNA and/or ctDNA from a blood or plasma sample and detecting levels of cfDNA and/or ctDNA using PCR and quantitative DNA electrophoresis.

2. Antibody Affinity

[0144] In certain aspects, an antibody provided herein has a dissociation constant (K_D) of < IpM, < 100 nM, < 10 nM, < 1 nM, < 0.1 nM, < 0.01 nM, or < 0.001 nM (e.g., 10'⁸M or less, e.g., from 10“⁸ M to 10'¹³ M, e.g., from 10'⁹M to 10’ⁱ³ M).

[0145] In one aspect, K_D is measured using a BIACORE® surface plasmon resonance assay. For example, an assay using a BIACORE®-2000 or a BIACORE ®-3000 (BIAcore, Inc., Piscataway, N.I) is performed at 25°C with immobilized antigen CM5 chips at -10 response units (RU). In one aspect, carboxy methylated dextran biosensor chips (CM5, BIACORE, Inc.) are activated with AAnhyl-T ’- (3- dimethylaminopropyl)-carbodiimide hydrochloride (EDC) and 2v-hydro xysuccinimide (NTIS) according to the supplier’s instructions. Antigen is diluted with 10 mM sodium acetate, pH 4.8, to 5 pg/ ml (—0.2 uM) before injection at a flow rate of 5 pl/minute to achieve approximately 10 response units (RU) of coupled protein. Following the injection of antigen, 1 M ethanolamine is injected to block unreacted groups. For kinetics measurements, two-fold serial dilutions of Fab (0.78 nM to 500 nM) are injected in PBS with 0.05% polysorbate 20 (TWEEN-20™) surfactant (PBST) at 25°C at a flow rate of approximately 25 pl/min. Association rates (k_on) and dissociation rates (k_on) are calculated using a simple one-to-one Langmuir binding model (BI ACORE® Evaluation Software version 3.2) by simultaneously fitting the association and dissociation sensorgrams. The equilibrium dissociation constant (K_D) is calculated as the ratio k_Off/k_On. See, e.g., Chen et al., J. Mol. Biol. 293:865-881 ( 1999). If the on-rate exceeds IO⁶ M"^s s'¹ by the surface plasmon resonance assay above, then the on-rate can be determined by using a fluorescent quenching technique that measures the increase or decrease in fluorescence emission intensity (excitation = 295 nm; emission = 340 ran, 16 ran band-pass) at 25°C of a 20 nM anti-antigen antibody (Fab form) in PBS, pH 7.2, in the presence of increasing concentrations of antigen as measured in a spectrometer, such as a stop-flow equipped spectrophometer (Aviv Instruments) or a 8000-series SLM-AMINCO ^IM spectrophotometer (Thermo Spectronic) with a stirred cuvette.

[0146] In an alternative method, KD is measured by a radiolabeled antigen binding assay (RIA). In one aspect, an RIA is performed with the Fab version of an antibody of interest and its antigen. For example, solution binding affinity’ of Fabs for antigen is measured by equilibrating Fab with a minimal concentration of (¹²⁵I)-labeled antigen in the presence of a titration series of unlabeled antigen, then capturing bound antigen with an anti-Fab antibody -coated plate (see, e.g., Chen et al,, J. Mol. Biol. 293:865-881(1999)). To establish conditions for the assay, MICROTITER® multi-well plates (Thermo Scientific) are coated overnight with 5 pg/ml of a capturing anti-Fab antibody (Cappel Labs) in 50 mM sodium carbonate (pH 9.6), and subsequently blocked with 2% (w/v) bovine serum albumin in PBS for two to five hours at room temperature (approximately 23°C). In a non-adsorbent plate (Nunc #269620), 100 pM or 26 pM [¹²⁵I] -antigen are mixed with serial dilutions of a Fab of interest (e.g., consistent with assessment of the anti-VEGF antibody, Fab-12, in Presta et ah, Cancer Res. 57:4593-4599 (1997)). The Fab of interest is then incubated overnight: however, the incubation may’ continue for a longer period (e.g., about 65 hours) to ensure that equilibrium is reached.

Thereafter, the mixtures are transferred to the capture plate for incubation at room temperature (e.g., for one hour). The solution is then removed and the plate washed eight times with 0.1% polysorbate 20 (TWEEN-20*) in PBS, When the plates have dried, 150 gl/well of scintillant (MICROSCINT-20 ^{1 M}; Packard) is added, and the plates are counted on a TOPCOUNT ^,M gamma counter (Packard) for ten minutes. Concentrations of each Fab that give less than or equal to 2.0% of maximal binding are chosen for use in competitive binding, assays,

3, Antibody Fragments

[0147] In certain aspects, an antibody provided herein is an antibody fragment.

[0148] In one aspect, the antibody fragment is a Fab, Fab’, Fab’-SH, or F(ab’)?. fragment, in particular a Fab fragment. Papain digestion of intact antibodies produces two identical antigenbinding fragments, called “Fab” fragments containing each the heavy- and light-chain variable domains ( VH and VL, respectively) and also the constant domain of the light chain (CL) and the first constant domain of the heavy chain (CHI). The term “Fab fragment” thus refers to an antibody fragment comprising a light chain comprising a VL domain and a CL domain, and a heavy chain fragment comprising a VH domain and a CHI domain. “Fab'¹ fragments” differ from Fab fragments by the addition of residues at the carboxy terminus of the CHI domain including one or more cysteines from the antibody hinge region. Fab’-SH are Fab’ fragments in which the cysteine residue(s) of the constant domains bear a free thiol group. Pepsin treatment yields an F(ab’ty fragment that has two antigen-binding sites (two Fab fragments) and a part of the Fc region. For discussion of Fab and F(ab’)2 fragments comprising salvage receptor binding epitope residues and having increased in vivo half-life, see U.S. Patent No. 5,869,046.

[0149] In another aspect, the antibody fragment is a diabody, a triabody or a tetrabody. “Diabodies” are antibody fragments with two antigen-binding sites that may be bivalent or bispecific. See, for example, EP 404,097; WO 1993/01161; Hudson et al., Nat. Med. 9: 129-134 (2003); and Hollinger et al., Proc. Natl. Acad. Sci. USA 90: 6444-6448 (1993). Triabodies and tetrabodies are also described in Hudson et al., Nat. Med. 9:129-134 (2003).

[0150] In a further aspect, the antibody fragment is a single chain Fab fragment. A “single chain Fab fragment” or “scFab" is a polypeptide consisting of an antibody heavy chain variable domain (VH), an antibody heavy chain constant domain 1 (CHI), an antibody light chain variable domain (VL), an antibody light chain constant domain (CL) and a linker, wherein said antibody domains and said linker have one of the following orders in N-termriial to C -terminal direction: a) VH-CHl -linker- VL-CL, b) VL-CL-linker-VH-CHl, c) VH-CL-hnker-VL-CHl or d) VL-CHl-linlver-VFI-CL. In particular, said linker is a polypeptide of at least 30 ammo acids, preferably between 32 and 50 amino acids. Said single chain Fab fragments are stabilized via the natural disulfide bond between the CL domain and the CHI domain. In addition, these single chain Fab fragments might be further stabilized by generation of interchain disulfide bonds via insertion of cysteine residues (e.g., position 44 in the variable heavy chain and position 100 in the variable light chain according to Rabat numbering).

[(H 51] In another aspect, the antibody fragment is single-chain variable fragment (scFv). A “single-chain variable fragment” or “scFv” is a fusion protein of the variable domains of the heavy (VH) and light drains (VL) of an antibody, connected by a linker. In particular, the linker is a short polypeptide of 10 to 25 amino acids and is usually rich in glycine for flexibility, as well as serine or threonine for solubility, and can either connect the N-terminus of the VH with the C -terminus of the VL, or vice versa. This protein retains the specificity of the original antibody, despite removal of the constant regions and the introduction of the linker. For a review of scFv fragments, see, e.g., Pluckthun, in The Pharmacology of Monoclonal Antibodies, vol. 113, Rosenburg and Moore eds., (Springer- Verlag, New York), pp. 269-315 (1994); see also WO 93/16185; and U.S. Patent Nos. 5,571,894 and 5,587,458.

[0152] In another aspect, the antibody fragment is a single-domain antibody . “Single-domain antibodies” are antibody fragments comprising all or a portion of the heavy chain variable domain or all or a portion of the light chain variable domain of an antibody. In certain aspects, a single-domain antibody is a human single-domain antibody (Domantis, Inc., Waltham, MA; see, e.g., U.S. Patent No. 6,248,516 Bl).

[0153] Antibody fragments can be made by various techniques, including but not limited to proteolytic digestion of an intact antibody as well as recombinant production by recombinant host cells (e.g,, E. coli), as described herein.

4. Chimeric and humanized antibodies

[0154] In certain aspects, an antibody provided herein is a chimeric antibody. Certain chimeric antibodies are described, e.g., in U.S. Patent No. 4,816,567; and Morrison et al., Proc. Natl. Acad. Sci. USA, 81:6851-6855 (1984)). In one example, a chimeric antibody comprises a non-human variable region (e.g., a variable region derived from a mouse, rat, hamster, rabbit, or non-human primate, such as a monkey) and a human constant region. In a further example, a chimeric antibody is a “class switched” antibody in which the class or subclass has been changed from tlrat of the parent antibody. Chimeric antibodies include antigen-binding fragments thereof.

[0155] In certain aspects, a chimeric antibody is a humanized antibody. Typically, a non-human antibody is humanized to reduce immunogenicity to humans, while retaining die specificity and affinity of the parental non-human antibody. Generally, a humanized antibody comprises one or more variable domains in which the CDRs (or portions thereof) are derived from a non-human antibody, and FRs (or portions thereof) are derived from human antibody sequences. A humanized antibody optionally will also comprise at least a portion of a human constant region. In some aspects, some FR residues in a humanized antibody are substituted with corresponding residues from a non-human antibody (e.g., the antibody from which the CDR residues are derived), e.g., to restore or improve antibody specificity or affinity.

[0156] Humanized antibodies and methods of making them are reviewed, e.g., in Almagro and Fransson, Front. Biosci. 13 : 1619-1633 (2008), and are further described, e.g., in Riechmann et al., Nature 332:323-329 (1988); Queen et al., Proc. Nat ’! Acad. Sci. USA 86:10029-10033 (1989); US Patent Nos. 5, 821,337, 7,52.7,791, 6,982,321, and 7,087,409; Kashmiri et al.. Methods 36:25-34 (2005) (describing specificity determining region (SDR) grafting); Padlan, Mol. Immunol. 2.8:489-498 (1991) (describing “resurfacing”); Dall’Acqua et al., Methods 36:43-60 (2005) (describing “FR shuffling”); and Osbourn et al., Methods 36:61-68 (2005) and Klimka et al., Br. J. Cancer, 83:252- 260 (2000) (describing the “guided selection” approach to FR shuffling).

[0157] Human framework regions that may be used for humanization include but are not limited to: framework regions selected using the “best-fit” method (see, e.g., Sims et al. J. Immunol.

151 :2296 (1993)); framework regions derived from the consensus sequence of human antibodies of a particular subgroup of light or heavy chain variable regions (see, e.g., Carter et al. Proc. Natl. Acad. Sci. USA, 89:4285 (1992); and Presta et al. J. Immunol., 151:2623 (1993)); human mature (somatically mutated) framework regions or human germline framework regions (see, e.g., Almagro and Fransson, Front. Biosci. 13:1619-1633 (2008)); and framework regions derived from screening FR libraries (see, e.g., Baca et al., J. Biol. Chem. 272: 10678-10684 (1997) and Rosok et al., J. Biol. Chem. 271:22611 -22618 (1996)).

5. Multispecific anti bodies

[0158] In certain aspects, an antibody provided herein is a multispecific antibody, e.g., a bispecific antibody. “Multispecific antibodies” are monoclonal antibodies that have binding specificities for at least two different sites, i.e., different epitopes on different antigens or different epitopes on the same antigen. In certain aspects, the multispecific antibody has three or more binding specificities. In certain aspects, one of the binding specificities is for MerTK and the other specificity is for any other antigen. In certain aspects, bispecific antibodies may bind to two (or more) different epitopes of MerTK. Multispecific (e.g., bispecific) antibodies may also be used to localize cytotoxic agents or cells to cells which express MerTK. Multispecific antibodies may be prepared as full length antibodies or antibody fragments.

[0159] Techniques for making multispecific antibodies include, but are not limited to, recombinant co-expression of two immunoglobulin heavy chain-light chain pairs having different specificities (see Milstein and Cuello, Nature 305: 537 (1983)) and “knob-in-hole” engineering (see, e.g., U.S. Patent No. 5,731,168, and Atwell et al., J. Mol. Biol. 270:26 (1997)). Multi-specific antibodies may also be made by engineering electrostatic steering effects for making antibody Fc- heterodimeric molecules (see, e.g., WO 2009/089004); cross-linking two or more antibodies or fragments (see, e.g., US Patent No. 4,676,980, and Brennan et al., Science, 229: 81 (1985)); using leucine zippers to produce bi-specific antibodies (see, e.g., Kostelny et al., J. Immunol, 148(5): 1547- 1553 (1992) and WO 2011/034605); using the common light chain technology for circumventing the light chain mis-paiiing problem (see, e.g., WO 98/50431); using “diabody” technology for making bispecific antibody fragments (see, e.g., Hollinger et al., Proc. Natl. Acad. Sci. USA, 90:6444-6448 (1993)); and using single-chain Fv (sFv) dimers (see, e.g., Gruber et al., J. Immunol., 152:5368 (1994)); and preparing trispecific antibodies as described, e.g., in Tutt et al. J. Immunol. 147: 60 (1991).

[0160] Engineered antibodies witli three or more antigen binding sites, including for example, “Octopus antibodies”, orDVD-Ig are also included herein (see, e.g., WO 2001/77342 and WO 2008/024715). Other examples of multispecific antibodies with three or more antigen binding sites can be found in WO 2010/115589, WO 2010/112193, WO 2010/136172, WO 2010/145792, and WO 2013/026831. The bispecific antibody or antigen binding fragment thereof also includes a “Dual Acting FAb” or “DAF” comprising an antigen binding site that binds to MerTK as well as another different antigen, or two different epitopes of MerTK (see, e.g., US 2008/0069820 and WO 2015/095539).

[0161] Multi-specific antibodies may also be provided in an asymmetric form with a domain crossover in one or more binding arms of the same antigen specificity, i.e, by exchanging the VH/VL domains (see e.g., WO 2009/080252 and WO 2015/150447), the CH1/CL domains (see e.g., WO 2009/080253) or the complete Fab arms (see e.g., W^rO 2009/080251, WO 2016/016299, also see Schaefer et al, PNAS, 108 (2011) 1187-1191, and Klein at al., MAbs 8 (2016) 1010-20). In one aspect, the multispecific antibody comprises a cross-Fab fragment. The term “cross-Fab fragment” or “xFab fragment” or “crossover Fab fragment” refers to a Fab fragment, wherein either the vanable regions or the constant regions of the heavy and light chain are exchanged. A cross-Fab fragment comprises a polypeptide chain composed of the light chain variable region (VL) and the heavy chain constant region 1 (CH 1), and a polypeptide chain composed of the heavy chain variable region ( VH) and the light drain constant region (CL). Asymmetrical Fab arms can also be engineered by introducing charged or non-charged amino acid mutations into domain interfaces to direct comet Fab pairing. See e.g., WO 2016/172485.

[0162] Various further molecular formats for multispecific antibodies are known in the art and are included herein (see e.g,, Spiess et al.. Mol Immunol 67 (2015) 95-106).

6. Antibody variants

[0163] In certain aspects, ammo acid sequence variants of the antibodies provided herein are contemplated. For example, it may be desirable to alter the binding affinity and/or other biological properties of the antibody. Amino acid sequence variants of an antibody may be prepared by introducing appropriate modifications into the nucleotide sequence encoding the antibody, or by peptide synthesis. Such modifications include, for example, deletions from, and/or insertions into and/or substitutions of residues within the amino acid sequences of the antibody. Any combination of deletion, insertion, and substitution can be made to arrive at the final construct, provided that the final construct possesses the desired characteristics, e.g., antigen-binding. a) Substitution, Insertion, and Deletion Variants

[0164] In certain aspects, antibody variants having one or more amino acid substitutions are provided. Sites of interest for substitutional mutagenesis include the CDRs and FRs. Conservative substitutions are shown in Table 1 under the heading of “preferred substitutions”. More substantial changes are provided in Table 1 under the heading of “exemplary substitutions”, and as further described below in reference to amino acid side chain classes. Amino acid substitutions may be introduced into an antibody of interest and the products screened for a desired activity, e.g., retained/! mproved antigen binding, decreased immunogenicity, or improved ADCC or CDC. TABLE 1

[0165] Amino acids may be grouped according io common side-chain properties:

(1) hydrophobic: Norleucine. Mei, Ala, Vai, Leu, lie:

(2) neutral hydrophilic: Cys, Ser, Thr, Asn, Gin;

(3) acidic: Asp, Glu;

(4) basic: His, Lvs, Arg;

(5) residues that influence drain orientation: Gly, Pro;

(6) aromatic: Trp, Tyr, Phe.

[0166] Non-conseivative substitutions will entail exchanging a member of one of these classes for a member of another class. [0167] One type of substitutional variant involves substituting one or more hypervariable region residues of a parent antibody (e.g., a humanized or human antibody). Generally, the resulting variant(s) selected for further study will have modifications (e.g., improvements) in certain biological properties (e.g., increased affinity, reduced immunogenicity) relative to the parent antibody and/or will have substantially retained certain biological properties of the parent antibody. An exemplary substitutional variant is an affinity matured antibody, which may be conveniently generated, e.g., using phage display -based affinity maturation techniques such as those described herein. Briefly, one or more. CDR residues are mutated and the variant antibodies displayed on phage and screened for a particular biological activity (e.g., binding affinity).

[0168] Alterations (e.g., substitutions) may be made in CDRs, e.g., to improve antibody affinity.

Such alterations may be made in CDR “hotspots”, i.e., residues encoded by codons tliat undergo mutation at high frequency during the somatic maturation process (see, e.g., Chowdhury', Methods Mol. Biol. 207:179-196 (2008)), and/or residues that contact antigen, with the resulting variant VH or VL being tested for binding affinity. Affinity maturation by constructing and reselecting from secondary libraries has been described, e.g., in Hoogenboom et al. in Methods in Molecular Biology 178: 1-37 (O’Brien et al., ed., Human Press, Totowa, NJ, (2001).) In some aspects of affinity maturation, diversity is introduced into the variable genes chosen for maturation by any of a variety of methods (e.g., error-prone PCR, chain shuffling, or oligonucleotide-directed mutagenesis). A secondary library' is then created. The library' is then screened to identify any antibody variants with die desired affinity. Another method to introduce diversity involves CDR-directed approaches, in which several CDR residues (e.g., 4-6 residues at a time) are randomized. CDR residues involved in antigen binding may be specifically identified, e.g., using alanine scanning mutagenesis or modeling. CDR-H3 and CDR-L3 in particular are often targeted.

[0169] In certain aspects, substitutions, insertions, or deletions may occur within one or more CDRs so long as such alterations do not substantially reduce the ability of the antibody to bind antigen. For example, conservative alterations (e.g., conservative substitutions as provided herein) that do not substantially reduce binding affinity may be made in the CDRs. Such alterations may, for example, be outside of antigen contacting residues in the CDRs. In certain variant VH and VL sequences provided above, each CDR either is unaltered, or contains no more than one, two or three amino acid substitutions.

[0170] A useful method for identification of residues or regions of an antibody tliat may be targeted for mutagenesis is called “alanine scanning mutagenesis” as described by Cunningham and Wells (1989) Science, 244: 1081-1085. In this method, a residue or group of target residues (e.g., charged residues such as arg, asp, his, ly s, and glu) are identified and replaced by a neutral or negatively charged amino acid (e.g., alanine or poly alanine) to determine whether the interaction of the antibody with antigen is affected. Further substitutions may be introduced at the amino acid locations demonstrating functional sensitivity to the initial substitutions. Alternatively, or additionally, a crystal structure of an antigen-antibody complex may be used to identify contact points between the antibody and antigen. Such contact residues and neighboring residues may be targeted or eliminated as candidates for substitution. Variants may be screened to determine whether they contain the desired properties.

[0171] Amino acid sequence insertions include amino- and/or carboxyl-terminal fusions ranging in length from one residue to polypeptides containing a hundred or more residues, as well as intrasequence insertions of single or multiple amino acid residues. Examples of terminal insertions include an antibody with an N-temunal methionyl residue. Other insertional variants of the antibodymolecule include the fusion to the N- or C-terminus of the antibody to an enzyme (e.g., for ADEPT (antibody directed enzyme prodrug therapy)) or a polypeptide which increases the serum half-life of the antibody. b) GIvcosvSation variants

[0172] In certain aspects, an antibody provided herein is altered to increase or decrease the extent to which the antibody is glycosylated. Addition or deletion of glycosylation sites to an antibody may be conveniently accomplished by altering the amino acid sequence such that one or more glycosylation sites is created or removed.

[0173] Where the antibody comprises an Fc region, the oligosaccharide attached thereto may be altered. Native antibodies produced by mammalian ceils typically comprise a branched, biantennary oligosaccharide that is generally attached by an N-Iinkage to Asn297 of tire CH2 domain of the Fc region. See, e.g., Wright et al. TIBTECH 15:26-32 (1997). The oligosaccharide may include various carbohydrates, e.g., mannose, N-acetyl glucosamine (GlcNAc), galactose, and sialic acid, as well as a fucose attached to a GlcNAc in the “stem" of the biantennary oligosaccharide structure. In some aspects, modifications of the oligosaccharide in an antibody of the invention may be made in order to create antibody variants with certain improved properties.

[0174] In one aspect, antibody variants are provided having a non-fucosylated oligosaccharide, i.e. an oligosaccharide structure that lacks fucose attached (directly or indirectly) to an Fc region.

Such non-fucosylated oligosaccharide (also referred to as “afucosylated” oligosaccharide) particularly is an N-linked oligosaccharide which lacks a fucose residue attached to the first GlcNAc in the stem of the biantennary oligosaccharide structure. In one aspect, antibody variants are provided having an increased proportion of non-fucosylated oligosaccharides in the Fc region as compared to a native or parent antibody. For example, the proportion of non-fucosylated oligosaccharides may be at least about 20%, at least about 40%, at least about 60%, at least about 80%, or even about 100% (i.e. no fucosy lated oligosaccharides are present). The percentage of non-fucosylated oligosaccharides is the (average) amount of oligosaccharides lacking fucose residues, relative to the sum of all oligosaccharides attached to Asn 297 (e. g. complex, hybrid and high mannose structures) as measured by MALDI-TOF mass spectrometry, as described in WO 2006/082515, for example.

Asn297 refers to the asparagine residue located at about position 297 in the Fc region (EU numbering of Fc region residues); however, Asn297 may also be located about i 3 amino acids upstream or downstream of position 297, i.e., between positions 294 and 300, due to minor sequence variations in antibodies. Such antibodies having an increased proportion of non-fucosylated oligosaccharides in the Fc region may have improved FcyRIIIa receptor binding and/or improved effector function, in particular improved ADCC function. See, e.g., US 2003/0157108; US 2004/0093621.

[0175] Examples of cell lines capable of producing antibodies with reduced fucosy lation include Lee 13 CHO cells deficient in protein fucosy lation (Ripka et al. Arch. Biochem. Biophys. 249:533-545 (1986); US 2003/0157108; and WO 2004/056312, especially at Example 11), and knockout cell lines, such as alpha- 1, 6-fucosyltransferase gene, FUT8, knockout CHO cells (see, e.g., Yamane-Ohnuki et al. Biotech. Bioeng. 87:614-622 (2004): Kanda, Y. et al., Biotechnol. Bioeng., 94(4):680-688 (2006); and WO 2003/085107), or cells with reduced or abolished activity of a GDP -fucose synthesis or transporter protein (see, e.g., US2004259150, US2005031613, US2004132140, US2004110282).

[0176] In a further aspect, antibody variants are provided with bisected oligosaccharides, e.g., in which a biantennary oligosaccharide attached to the Fc region of the antibody is bisected by GlcNAc. Such antibody variants may have reduced fucosylation and/or improved ADCC function as described above. Examples of such antibody variants are described, e.g., in Umana et al., Nat Biotechnol 17, 176-180 (1999); Ferrara et al., Biotechn Bioeng 93, 851-861 (2006); W^rO 99/54342; WO 2004/065540, WO 2003/011878.

[0177] Antibody variants with at least one galactose residue in the oligosaccharide attached to tlie Fc region aie also provided. Such antibody variants may have improved CDC function. Such antibody variants are described, e.g., in WO 1997/30087; WO 1998/58964: and WO 1999/22764. c) Fc region variants

[0178] In certain aspects, one or more amino acid modifications may be introduced into the Fc region of an antibody provided herein, thereby generating an Fc region variant. The Fc region variant may comprise a human Fc region sequence (e.g., a human IgGl, IgG2, IgG3 or IgG4 Fc region) comprising an amino acid modification (e.g., a substitution) at one or more amino acid positions.

[0179] In certain aspects, the invention contemplates an antibody vanant that possesses some but not all effector functions, which make it a desirable candidate for applications in which the half life of tlie antibody in vivo is important yet certain effector functions (such as complement-dependent cytotoxicity (CDC) and antibody -dependent cell-mediated cytotoxicity (ADCC)) are unnecessary or deleterious. In vitro and/or in vivo cytotoxicity assays can be conducted to confirm the reduction/depletion of CDC and/or ADCC activities. For example, Fc receptor (FcR) binding assays can be conducted to ensure that the antibody lacks FcvR binding (hence likely lacking ADCC activity), but retains FcRn binding ability’. The primary’ cells for mediating ADCC, NK cells, express FcyRIII only, whereas monocytes express FcyRI, FcyRU and FcyRIII. FcR expression on hematopoietic cells is summarized in Table 3 on page 464 of Ravetch and Kinet,z1nww. Rev. Immunol. 9:457-492 (1991). Non-limiting examples of in vitro assays to assess ADCC activity of a molecule of interest is described in U.S. Patent No. 5,500,362 (see, e.g., Hellstrom, I. et al. Proc. Nat ’I Acad. Sci. USA 83:7059-7063 (1986)) and Hellstrom, I et al.. Proc. Natl Acad. Sci. USA 82:1499-1502 (1985); 5,821,337 (see Bniggemann, M. et al., J. Exp. Med. 166:1351-1361 (1987)). Alternatively, nonradioactive assays methods may be employed (see, for example, ACT!'™ non-radioactive cytotoxicity assay for flow cytometry (CellTechnology, Inc. Mountain View, CA; and CytoTox 96® nonradioactive cytotoxicity assay (Promega, Madison, Wl). Useful effector cells for such assays include peripheral blood mononuclear cells (PBMC) and Natural Killer (NK) cells. Alternatively, or additionally, ADCC activity’ of the molecule of interest may be assessed in vivo, e.g., in a animal model such as that, disclosed in Clynes et al. Proc. Nat ’i Acad. Sci. USA 95:652-656 (1998). Clq binding assays may also be carried out to confirm that the antibody is unable to bind Clq and hence lacks CDC activity. See, e.g., Clq and C3c binding ELISA in WO 2006/029879 and

WO 2005/100402. To assess complement activation, a CDC assay may be performed (see, for example, Gazzano-Santoro et al., J. Immunol. Methods 202: 163 (1996); Cragg, M.S. et al., Blood 101: 1045-1052 (2003); and Cragg, M.S. and MJ. Glennie, Blood 103:2738-2743 (2004)). FcRn binding and in vivo clearance/italf life determinations can also be performed using methods known in the art (see, e.g., Petkova, S.B. et al., for 7. Immunol. 18(12): 1759-1769 (2006); WO 2013/120929 Al). [0180] Antibodies with reduced effector function include those with substitution of one or more of Fc region residues 238, 265, 269, 270, 297, 327 and 329 (U.S. Patent No. 6,737,056). Such Fc mutants include Fc mutants with substitutions at two or more of amino acid positions 265, 269, 270, 297 and 327, including the so-called “DANA” Fc mutant with substitution of residues 265 and 297 to alanine (US Patent No. 7,332,581).

[0181] Certain antibody variants with improved or diminished binding to FcRs are described. (See, e.g., U.S. Patent No. 6,737,056; WO 2004/056312, and Shields et al., J. Biol. Chem. 9(2): 6591- 6604 (2001).)

[0182] In certain aspects, an antibody variant comprises an Fc region with one or more amino acid substitutions which improve ADCC, e.g., substitutions at positions 298, 333, and/or 334 of the Fc region (EU numbering of residues).

[0183] In certain aspects, an antibody variant comprises an Fc region with one or more amino acid substitutions which diminish FcyR binding, e.g., substitutions at positions 234 and 235 of the Fc region (EU numbering of residues). In one aspect, the substitutions are L234A and L235A (LALA). In certain aspects, the antibody variant further comprises D265A and/or P329G in an Fc region derived from a human IgGl Fc region. In one aspect, the substitutions are L234A, L235A and P329G (LALA-PG) in an Fc region derived from a human IgGl Fc region. (See, e.g., WO 2012/130831). In another aspect, the substitutions are L234A, L235A and D265 A (LALA-DA) in an Fc region derived from a human IgGl Fc region.

[0184] In some aspects, alterations are made in the Fc region that result in altered (i.e., either improved or diminished) Clq binding and/or Complement Dependent Cytotoxicity (CDC), e.g., as described in US Patent No. 6,194,551 , WO 99/51642, and Idusogie et al. J. Immunol. 164: 4178-4184 (2000).

[0185] Antibodies with increased half lives and improved binding to the neonatal Fc receptor (FcRn), which is responsible for the transfer of maternal IgGs to the fetus (Guyer et al., J. Immunol.

117:587 (1976) and Kim et al., J. Immunol. 24:249 (1994)), are described in US2005/0014934 (Hinton et al.). Those antibodies comprise an Fc region with one or more substitutions therein which improve binding of the Fc region to FcRn. Such Fc variants include those with substitutions at one or more of Fc region residues: 238, 252, 254, 256, 265, 272, 286, 303, 305, 307, 311, 312, 317, 340, 356, 360, 362, 376, 378, 380, 382, 413, 424 or 434, e.g., substitution of Fc region residue 434 (See, e.g., US Patent No. 7,371,826; Dall'Acqua, W.F., et al. J. Biol. Chem. 281 (2006) 23514-23524).

[0186] Fc region residues critical to the mouse Fc-monse FcRn interaction have been identified by site-directed mutagenesis (see e.g. Dall’Acqua, W.F., et al. J. Immunol 169 (2002) 5171-5180). Residues 12.53, H310, H433, N434, and H435 (EU numbering of residues) are involved in the interaction (Medesan, C,, et al., Eur. J, Immunol. 26 ( 1996) 2533; Firan, M., et al., Int. Immunol. 13 (2001) 993; Kim, J.K., et al., Eur. J. Immunol. 24 (1994) 542). Residues 1253, H310, and H435 were found to be critical for the interaction of human Fc with murine FcRn (Kim, J.K., et al., Eur. J. Immunol. 29 (1999) 2819). Studies of the human Fc-human FcRn complex have shown that residues 1253, S254, H435, and Y436 are crucial for the interaction (Firan, M., et al., Int. Immunol. 13 (2001) 993; Shields, R.L., et al., J. Biol. Chem. 276 (2001) 6591-6604). In Yeung, Y.A., et al. (J. Immunol.

182 (2009) 7667-7671) various mutants of residues 248 to 259 and 301 to 317 and 376 to 382 and 424 to 437 have been reported and examined.

[0187] In certain aspects, an antibody variant comprises an Fc region with one or more amino acid substitutions, which reduce FcRn binding, e.g., substitutions at positions 253, and/or 310, and/or

435 of the Fc-region (EU numbering of residues). In certain aspects, the antibody variant comprises an Fc region with the amino acid substitutions at positions 253, 310 and 435. In one aspect, the substitutions are 1253 A, H310A and H435A in an Fc region derived from a human IgGl Fc-region, See, e.g., Grevys, A,, et al., J. Immunol. 194 (2015) 5497-5508.

[0188] In certain aspects, an antibody variant comprises an Fc region with one or more amino acid substitutions, which reduce FcRn binding, e.g., substitutions at positions 310, and/or 433, and/or

436 of the Fc region (EU numbering of residues). In certain aspects, the antibody variant comprises an Fc region with the amino acid substitutions at positions 310, 433 and 436. In one aspect, the substitutions are H310A, H433A and Y436A in an Fc region derived from a human IgGl Fc-region. (See, e.g., WO 2014/177460 Al). [0189] In certain aspects, an antibody variant comprises an Fc region with one or more amino acid substitutions which increase FcRn binding, e.g., substitutions at positions 252, and/or 254, and/or 256 of the Fc region (EU numbering of residues). In certain aspects, the antibody variant comprises an Fc region with amino acid substitutions at positions 2.52, 254, and 256. In one aspect, the substitutions are M252Y, S2.54T and T256E in an Fc region derived from a human IgGl Fc-region. See also Duncan & Winter, Nature 32.2:738-40 (1988); U.S. Patent No. 5,648,2.60; U.S. Patent No. 5,62.4,821 ; and WO 94/29351 concerning oilier examples of Fc region variants.

[0190] The C-terminus of the heavy chain can be a shortened C-terminus in which one or two of the C terminal amino acid residues have been removed. In one preferred aspect, the C-terminus of the heavy chain is a shortened C-terminus ending PG. In one aspect of all aspects as reported herein, an antibody comprising a heavy chain including a C-terminal CH3 domain as specified herein, comprises the C-terminal glycine-lysine dipeptide (G446 and K447, EU index numbering of amino acid positions). In one aspect of all aspects as reported herein, an antibody comprising a heavy drain including a C-terminal CH3 domain, as specified herein, comprises a C-terminal glycine residue (G446, EU index numbering of amino acid positions). d) Cysteine engineered antibody variants

[0191] In certain aspects, it may be desirable to create cysteine engineered antibodies, e.g., TH1OMAB™ antibodies, in which one or more residues of an antibody are substituted with cysteine residues, in particular aspects, the substituted residues occur at accessible sites of the antibody. By substituting those residues with cysteine, reactive thiol groups are thereby positioned at accessible sites of the antibody and may be used to conjugate the antibody to other moieties, such as PEG poiymer(s), drug moieties, or linker-drug moieties, to create an immunoconjugate or PEGylated antibody, as described further herein. Cysteine engineered antibodies may be generated as described, e.g., in U.S. Patent No. 7,521,541, 8,30,930, 7,855,275, 9,000,130, WO 2016040856, or WO 201 1/156328. e) Antibody derivatives

[0192] In certain aspects, an antibody provided herein may be further modified to contain additional nonproteinaceous moieties that are known in the art and readily available. The moieties suitable for derivatization of die antibody include but are not limited to water soluble polymers. Nonlimiting examples of water soluble polymers include, but are not limited to, polyethylene glycol (PEG), copolymers of ethylene glycol/propylene glycol, carboxymethylcellulose, dextran, polyvinyl alcohol, polyvinyl pyrrolidone, poly-1, 3 -dioxolane, poly- 1,3, 6-trioxane, ethylene/maleic anhydride copolymer, polyaminoacids (either homopolymers or random copolymers), and dextran or polypvinyl pyrrolidone)poly ethylene glycol, propropylene glycol homopolymers, proly propylene oxide/etbylene oxide co-polymers, polyoxy ethylated polyols (e.g., glycerol), polyvinyl alcohol, and mixtures thereof. Polyethylene glycol propionaldehyde may have advantages in manufacturing due to its stability in water. The polymer may be of any molecular weight, and may be branched or unbranched. The number of polymers attached to the antibody may vary’, and if more than one polymer are attached, they can be the same or different molecules. In general, the number and/or type of polymers used for derivatization can be determined based on considerations including, but not limited to, the particular properties or functions of the antibody to be improved, whether the antibody derivative will be used in a therapy under defined conditions, etc.

7. Immunoconjugates

[0193] The invention also provides immunoconjugates comprising an anti-MerTK antibody herein conjugated (chemically bonded) to one or more therapeutic agents such as cytotoxic agents, chemotherapeutic agents, drags, growth inhibitory agents, toxins (e.g., protein toxins, enzymatically' active toxins of bacterial, fungal, plant, or animal origin, or fragments thereof), or radioactive isotopes.

[0194] In one aspect, an immunoconjugate is an antibody -drug conjugate (ADC) in which an antibody is conjugated to one or more of the therapeutic agents mentioned above. The antibody is typically connected to one or more of the therapeutic agents using linkers. An overview of ADC technology including examples of therapeutic agents and drugs and linkers is set forth in Pharmacol Review 68:3-19 (2016).

[0195] In another aspect, an immunoconjugate comprises an antibody as described herein conjugated to an enzymatically active toxin or fragment thereof, including but not limited to diphtheria A drain, nonbinding active fragments of diphtheria toxin, exotoxin A chain (from Pseudomonas aemginosa), ricin A chain, abrin A drain, modeccin A chain, alpha-sarcin, Aleurites fordii proteins, dianthin proteins, Phytolaca americana proteins (PAPI, PAPII, and P AP-S), momordica charantia inhibitor, curcin, crotin, sapaonaria officinalis inhibitor, gelonin, mitogellin, restrictocin, phenomycin, enomycin, and the tricothecenes.

[0196] In another aspect, an immunoconjugate comprises an antibody as described herein conjugated to a radioactive atom to form a radioconjugate. A variety of radioactive isotopes are available for the production of radioconjugates. Examples include At²¹ ‘, I ¹³, 1¹²⁵, Y⁹⁰, Re¹⁸⁶, Re¹⁸⁸, Sm¹⁵’, Bi²¹², P ^!?, Pb^{2 12} and radioactive isotopes of Lu. When the radioconjugate is used for detection, it may comprise a radioactive atom for scintigraphic studies, for example tc99m or 1123, or a spin label for nuclear magnetic resonance (NMR) imaging (also known as magnetic resonance imaging, mri), such as iodine-123 again, iodine-131, indium-i l l, fluorine-19, carbon-13, nitrogen-15, oxy gen- 17, gadolinium, manganese or iron.

[0197] Conjugates of an antibody and cytotoxic agent may be made using a variety of bifunctional protein coupling agents such as N-succinimidyl-3-(2-pyridyldithio) propionate (SPDP), succinimidyl-4-(N-maleimidomethyl) cyclohexane- 1 -carboxylate (SMCC), iminothiolane (IT), bifunctional derivatives of imidoesters (such as dimethyl adipimidate HC1), active esters (such as disuccinimidvl suberate), aldehydes (such as glutaraldehyde), bis-azido compounds (such as bis (p- azidobenzoyl) hexanediamine), bis-diazonium derivatives (such as bis-(p-diazomumbenzoyl)- ethylenediamine), diisocyanates (such as toluene 2,6-diisocyanate), and bis-active fluorine compounds (such as I,5-difluoro-2,4-dimtrobenzene). For example, a ricin immunotoxin can be prepared as described in Vitetta et al.. Science 238: 1098 (1987). Carbon-14-labeled 1- isotluocyanatobenzyl-3-methyldiethylene triaminepentaacetic acid (MX-DTPA) is an exemplary chelating agent for conjugation of radionucleotide to die antibody. See WO 94/11026. The linker may be a “cleavable linker" facilitating release of a cytotoxic drug in the ceil. For example, an acid- labile linker, peptidase-sensitive linker, photolabile linker, dimethyl linker or disulfide-containing linker (Chari et al., Cancer Res. 52:127-131 (1992); U.S. Patent No. 5,208,020) may be used.

[0198] The immunuoconjugates or ADCs herein expressly contemplate, but are not limited to such conjugates prepared with cross-linker reagents including, but not limited to, BMPS, EMCS, GMBS, HBVS, LC-SMCC, MBS, MPBH, SBAP, SIA, STAB, SMCC, SMPB, SMPH, sulfo-EMCS, sulfo-GMBS, sulfo-KMUS, sulfo-MBS, sulfo-SIAB, sulfo-SMCC, and sulfo-SMPB, and SVSB (succinimidyl-(4-vinylsulfone)benzoate) which are commercially available (e.g., from Pierce Biotechnology, Inc., Rockford, IL,, U.S.A).

C. Recombinant Methods and Compositions

[0199] Antibodies may be produced using recombinant methods and compositions, e.g., as described in US 4,816,567. For these methods one or more isolated nucleic acid(s) encoding an antibody are provided.

[0200] In case of a native antibody or native antibody fragment two nucleic acids are required, one for the light chain or a fragment thereof and one for the heavy chain or a fragment thereof. Such nucleic acid(s) encode an amino acid sequence comprising the VL and/or an amino acid sequence comprising the VH of the antibody (e.g., the light and/or heavy chain(s) of the antibody). These nucleic acids can be on the same expression vector or on different expression vectors.

[0201] In case of a bispecific antibody with heterodimeric heavy chains four nucleic acids are required, one for the first light chain, one for the first heavy chain comprising the first heteromonomeric Fc -region polypeptide, one for the second light chain, and one for the second heavy chain comprising the second heteromonomeric Fc -region polypeptide. The four nucleic acids can be comprised in one or more nucleic acid molecules or expression vectors. Such nucleic acid(s) encode an amino acid sequence comprising the first VL and/or an amino acid sequence comprising the first VH including the first heteromonomeric Fc-region and/or an amino acid sequence comprising the second VL and/or an amino acid sequence comprising the second VH including the second heteromonomeric Fc-reglon of the antibody (e.g., the first and/or second light and/or the first and/or second heavy chains of the antibody). These nucleic acids can be on the same expression vector or on different expression vectors, normally these nucleic acids are located on two or three expression vectors, i.e. one vector can comprise more than one of these nucleic acids. Examples of these bispecific antibodies are CrossMabs (see, e.g., Schaefer, W. ei al, PNAS, 108 (2011) 11187-1191), For example, one of the heteromonomeric heavy chain compri ses the so-called “knob mutations” (T366W and optionally one of S354C or Y349C) and the other comprises the so-called “hole mutations” (T366S, L368A and Y407V and optionally Y349C or S354C) (see, e.g.. Carter, P. et al,, Iimnunotechnol. 2 (1996) 73) according to EU index numbering.

[0202] In one aspect, isolated nucleic acids encoding an antibody as used in the methods as reported herein are provided.

[0203] In one aspect, a method of making an anti-MerTK antibody is provided, wherein the method comprises culturing a host ceil comprising nucleic acld(s) encoding the antibody, as provided above, under conditions suitable for expression of the antibody, and optionally recovering the antibody from the host cell (or host cell culture medium).

[0284] For recombinant production of an anti-MerTK antibody, nucleic acids encoding the antibody, e.g., as described above, are isolated and inserted into one or more vectors for further cloning and/or expression in a host cell. Such nucleic acids may be readily isolated and sequenced using conventional procedures (e.g., by using oligonucleotide probes that are capable of binding specifically to genes encoding the heavy and light chains of the antibody ) or produced by recombinant methods or obtained by chemical sy nthesis.

[0205] Suitable host cells for cloning or expression of antibody -encoding vectors include prokaryotic or eukaryotic cells described herein. For example, antibodies may be produced in bacteria, in particular when glycosylation and Fc effector function are not needed. For expression of antibody fragments and polypeptides in bacteria, see, e.g., US 5,648,237, US 5,789,199, and US 5,840,523. (See also Chariton, K.A., In: Methods in Molecular Biology, Vol. 248, Lo, B.K.C. (ed.), Humana Press, Totowa, NJ (2003), pp. 245-254, describing expression of antibody fragments in E. coli.) After expression, the antibody may be isolated from the bacterial cell paste in a soluble fraction and can be further purified.

[0206] In addition to prokaryotes, eukaryotic microbes such as filamentous fungi or yeast are suitable cloning or expression hosts for antibody -encoding vectors, including fungi and yeast strains whose glycosylation pathway s have been “humanized”, resulting in the production of an antibody with a partially or fully human glycosylation pattern. See Gerngross, T.U., Nat. Biotech. 22 (2004) 1409-1414: and Li, H. et al., Nat. Biotech. 24 (2006) 210-215.

[0207] Suitable host cells for the expression of (glycosylated) antibody are also derived from multicellular organisms (invertebrates and vertebrates). Examples of invertebrate cells include plant and insect cells. Numerous bacuioviral strains have been identified which may be used in conjunction with insect cells, particularly for transfection of Spodoptera frugiperda cells.

[0208] Plant cell cultures can also be utilized as hosts. See, e.g., US 5,959,177, US 6,040,498, US 6,420,548, US 7, 125,978, and US 6,417,42.9 (describing PLANTIBODIESTM technology for producing antibodies in transgenic plants),

[0209] Vertebrate cells may also be used as hosts. For example, mammalian cell lines that are adapted to grow in suspension may be useful. Other examples of useful mammalian host cell lines are monkey kidney CV1 line transformed by SV40 (COS-7); human embryonic kidney line (293 or 293T cells as described, e.g., in Graham, F.L. et al., J. Gen Virol. 36 (1977) 59-74); baby hamster kidney cells (BHK); mouse sertoli cells (TM4 cells as described, e.g., in Mather, IP., Biol. Reprod. 23 (1980) 243-252); monkey kidney cells (C VI); African green monkey kidney cells (VERO-76); human cervical carcinoma cells (HELA); canine kidney cells (MDCK; buffalo rat liver cells (BRL 3A); human lung ceils (W138); human liver cells (Hep G2); mouse mammary tumor (MMT 060562); TRI cells (as described, e.g., in Mather, J.P. et al.. Annals N.Y. Acad. Sci. 383 (1982) 44-68); MRC 5 cells; and FS4 cells. Other useful mammalian host cell lines include Chinese hamster ovary (CHO) cells, including DHFR- CHO cells (Urlaub, G. et al., Proc. Natl. Acad. Sci. US A 77 (1980) 4216- 422.0); and myeloma cell lines such as Y0, NS0 and Sp2/0. For a review of certain mammalian host cell lines suitable for antibody production, see, e.g., Yazaki, P. and Wu, A.M., Methods in Molecular Biology, Vol. 2.48, Lo, B.K.C. (ed.), Humana Press, Totowa, NJ (2004), pp. 255-268.

[0218] In one aspect, the host cell is eukaryotic, e.g., a Chinese Hamster Ovary (CHO) cell or lymphoid cell (e.g., Y0, NS0, Sp20 cell).

I). Assays

[0211] Anti-MerTK antibodies provided herein may be identified, screened for, or characterized for their phy sical/chemicai properties and/or biological activities by various assays known in the art.

1. Binding assays and other assays

[0212] In one aspect, an antibody of the invention is tested for its antigen binding activity, e.g., by known methods such as ELISA, Western blot, etc.

[0213] In another aspect, competition assays may be used to identify an antibody that competes with any of the anti-MerTK antibodies described supra for binding to MerTK. In certain aspects, such a competing antibody binds to the same epitope (e.g., a linear or a conformational epitope) that is bound by any of the anti-MerTK antibodies described supra. Detailed exemplary methods for mapping an epitope to which an antibody binds are provided in Morris (1996) “Epitope Mapping Protocols”, in Methods in Molecular Biology vol. 66 (Humana Press, Totowa, N.I). [0214] In an exemplary competition assay, immobilized MerTK is incubated in a solution comprising a first labeled antibody’ that binds to MerTK (e.g., any’ of the anti-MerTK antibodies described supra) and a second unlabeled antibody that is being tested for its ability' to compete with the first antibody for binding to MerTK, The second antibody may be present in a hybridoma supernatant. As a control, immobilized MerTK is incubated in a solution comprising the first labeled antibody but not the second unlabeled antibody. After incubation under conditions permissive for binding of the first antibody to MerTK, excess unbound antibody is removed, and tire amount of label associated with immobilized MerTK is measured. If the amount of label associated with immobilized MerTK is substantially reduced in the test sample relative to the control sample, then that indicates that the second antibody is competing with the first antibody for binding to MerTK. See Harlow and Lane (1988) Antibodies: A Laboratory Manual ch.14 (Cold Spring Harbor Laboratory', Cold Spring Harbor, NY).

2. Activity assays

[0215] In one aspect, assays are provided for identifying anti-MerTK antibodies thereof having biological activity. Biological activity' may include, e.g., reducing MeiTK-mediated phagocytic activity, reducing MerTK-mediated clearance of apoptotic cells, and/or enhancing tumor immunogenicity’ of a checkpoint inhibitor. Antibodies having such biological activity' in vivo and/or in vitro are also provided.

[0216] In certain aspects, an antibody of the invention is tested for such biological activity. Examples of assays suitable for measuring such biological activity are described further herein, including the Exemplification section below.

E. Pharmaceutical compositions

[0217] In a further aspect, provided are pharmaceutical compositions compri sing any of the antibodies provided herein, e.g., for use in any of the below therapeutic methods. In one aspect, a pharmaceutical composition comprises any of the antibodies provided herein and a pharmaceutically acceptable carrier. In another aspect, a pharmaceutical composition comprises any of the antibodies provided herein and at least one additional therapeutic agent, e.g., as described below.

[0218] Pharmaceutical compositions of an anti-MerTK antibody as described herein are prepared by mixing such antibody having the desired degree of purity witli one or more optional pharmaceutically acceptable earners (Remington's Pharmaceutical Sciences 16th edition, Osol, A. Ed. (1980)), in the form of lyophilized compositions or aqueous solutions. Pharmaceutically acceptable earners are generally nontoxic to recipients at the dosages and concentrations employed, and include, but are not limited to: buffers such as histidine, phosphate, citrate, acetate, and other organic acids; antioxidants including ascorbic acid and methionine; preservatives (such as octadecyldimethylbenzyl ammonium chloride; hexamethonium chloride; benzalkonium chloride; benzethonium chloride; phenol, butyl or benzyl alcohol; alkyl parabens such as methyl or propyl paraben; catechol; resorcinol; cyclohexanol; 3-pentanol; and m-cresol); low molecular weight (less than about 10 residues) polypeptides; proteins, such as serum albumin, gelatin, or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, histidine, arginine, or lysine; monosaccharides, di saccharides, and other caibohy dates including glucose, mannose, or dextrins; chelating agents such as EDTA; sugars such as sucrose, mannitol, trehalose or sorbitol; saltforming counter-ions such as sodium; metal complexes (e.g., Zn-protein complexes); and/or non-ionic surfactants such as polyethylene glycol (PEG). Exemplary pharmaceutically acceptable carriers herein further include insterstitial drug dispersion agents such as soluble neutral-active hyaluronidase glycoproteins (sHASEGP), for example, human soluble PH-20 hyaluronidase glycoproteins, such as rHuPH20 (HYLENEX®', Halozyme, Inc.). Certain exemplary sHASEGPs and methods of use, including rHuPH20, are described in US Patent Publication Nos. 2005/0260186 and 2006/0104968. In one aspect, a sHASEGP is combined with one or more additional glycosaminoglv canases such as chondroitinases.

[0219] Exemplary lyophilized antibody compositions are described in US Patent No. 6,267,958. Aqueous antibody compositions include those described in US Patent No, 6,171,586 and WO 2006/044908, the latter compositions including a histidine-acetate buffer,

[0220] The plrarmaceutical composition herein may also contain more than one active ingredients as necessary for the particular indication being treated, preferably those with complementary activities that do not adversely affect each other. For example, it may be desirable to further provide immune checkpoint inhibitor(s), e.g. , an anti-PDL 1 antibody. Such active ingredients are suitably present in combination in amounts that are effective for the purpose intended.

[0221] Active ingredients may be entrapped in microcapsules prepared, for example, by coacervation techniques or by interfacial polymerization, for example, hydroxymethylcellulose or gelatin-microcapsules and poly-(methylmethacylate) microcapsules, respectively, in colloidal ding delivery systems (for example, liposomes, albumin microspheres, microemulsions, nano-particles and nanocapsules) or in macroemulsions. Such techniques are disclosed in Remington’s Pharmaceutical Sciences 16th edition, Osol, A. Ed, (1980).

[0222] Pharmaceutical compositions for sustained-release may be prepared. Suitable examples of sustained-release preparations include semipermeable matrices of solid hydrophobic polymers containing the antibody, which matrices are in the form of shaped articles, e.g., films, or microcapsules.

[0223] The pharmaceutical compositions to be used for in vivo administration are generally sterile. Sterility may be readily accomplished, e.g., by filtration through sterile filtration membranes.

F. Therapeutic Methods and Routes of Administration [0224] Any of the anti-MeiTK antibodies provided herein may be used in therapeutic methods. In any of the methods disclosed herein, in some embodiments, the methods (e. g. , using a PEGylated anti-MerTK antibody of the present disclosure) result in decreased retinal toxicity, as compared to simitar methods using a non-PEGylated anti-MerTK antibody.

[0225] In one aspect, an anti-MerTK antibody for use as a medicament is provided. In further aspects, an anti-MerTK antibody for use in treating cancer is provided. In certain aspects, an anti- MerTK antibody for use in a method of treatment is provided. In certain aspects, the invention provides an anti-MerTK antibody for use in a method of treating an individual having cancer comprising administering to the individual an effective amount of the anti-MerTK antibody . In one such aspect, the method further comprises administering to the individual an effective amount of at least one additional therapeutic agent (e.g., one, two, three, four, five, or six additional therapeutic agents), e.g., as described below. In further aspects, the invention provides an anti-MerTK antibody for use in enhancing immune function and/or reducing Me rTK -mediated clearance of apoptotic cells. In certain aspects, the invention provides an anti-MerTK antibody for use in a method of enhancing immune function and/or reducing MerTK-mediated clearance of apoptotic cells in an individual comprising administering to the individual an effective amount of the anti-MerTK antibody to enhance immune function and/or reduce MerTK-mediated clearance of apoptotic cells. An “individual” according to any of the above aspects is preferably a human.

[0226] In a further aspect, the invention provides for the use of an anti-MerTK antibody in the manufacture or preparation of a medicament. In one aspect, the medicament is for treatment of cancer. In a further aspect, the medicament is for use in a method of treating cancer comprising administering to an individual having cancer an effective amount of the medicament. In one such aspect, the method further comprises administering to the individual an effective amount of at least one additional therapeutic agent, e.g., as described below. In a further aspect, the medicament is for enhancing immune function and/or reducing MerTK-mediated clearance of apoptotic cells. In a further aspect, the medicament is for use in a method of enhancing immune function and/or reducing MerTK-mediated clearance of apoptotic cells in an individual comprising administering to the individual an effective amount of the medicament to entrance immune function and/or reduce MerTK- mediated clearance of apoptotic cells. An “individual” according to any of the above aspects may be a human,

[0227] In a further aspect, the invention provides a method for treating a cancer. In one aspect, the method comprises administering to an individual having such cancer an effective amount of an anti-MerTK antibody . In one such aspect, the method further comprises administering to the individual an effective amount of at least one additional therapeutic agent, as described below.

[0228] An “individual” according to any of the above aspects may be a human.

[0229] In a further aspect, the invention provides a method for enhancing immune function and/or reducing MerTK-mediated clearance of apoptotic cells in an individual, e.g. , an individual having cancer. In one aspect, the method comprises administering to the individual an effective amount of an anti-MerTK antibody to enhance immune function and/or reduce MerTK-mediated clearance of apoptotic cells. In one aspect, an “individual” is a human.

[0230] In a further aspect, the invention provides pharmaceutical compositions comprising any of the anti-MerTK antibodies provided herein, e.g., for use in any of the above therapeutic methods. In one aspect, a pharmaceutical composition comprises any of the anti-MerTK antibodies provided herein and a pharmaceutically acceptable carrier. In another aspect, a pharmaceutical composition comprises any of the anti-MerTK antibodies provided herein and at least one additional therapeutic agent, e.g., as described below.

1. Monotherapy

[0231] In some embodiments, an anti-MerTK antibody of the present disclosure (e.g., a PEGylated anti-MerTK antibody) is administered as a monotherapy to treat an individual having cancer. As used herein, “cancer” refers to or describes the physiological condition in mammals that is typical ly characterized by unregulated cell growth. In certain embodiments, the cancer may be a solid cancer or a hematologic cancer. Solid cancers are generally characterized by tumor mass formation in specific tissues. “Tumor,” as used herein, refers to all neoplastic cell growth and proliferation, whether malignant or benign, and all pre-cancerous and cancerous cells and tissues. Non-limiting examples of solid cancers to be treated with an anti-MerTK antibody of the present disclosure include carcinoma, lymphoma, blastoma, and sarcoma. More particular examples of such cancers include, but not limited to, squamous cell cancer (e.g, epithelial squamous cell cancer), lung cancer including small-cell lung cancer, non-small cell lung cancer, adenocarcinoma of the lung and squamous carcinoma of the lung, cancer of the peritoneum, hepatocellular cancer, gastric or stomach cancer including gastrointestinal cancer and gastrointestinal stromal cancer, pancreatic cancer, glioblastoma, cervical cancer, ovarian cancer, liver cancer, bladder cancer, cancer of the urinary tract, hepatoma, breast cancer, colon cancer, rectal cancer, colorectal cancer, endometrial or uterine carcinoma, salivary gland carcinoma, kidney or renal cancer, prostate cancer, vulval cancer, thyroid cancer, hepatic carcinoma, anal carcinoma, penile carcinoma, melanoma, superficial spreading melanoma, lentigo maligna melanoma, acral lentiginous melanomas, nodular melanomas, as well as abnormal vascular proliferation associated with phakomatoses, edema (such as that associated with brain tumors), Meigs’ syndrome, brain, head and neck cancer, and associated metastases. In certain embodiments, cancers that are amenable to treatment by anti-MerTK antibodies of the present disclosure include breast cancer, colorectal cancer, rectal cancer, non-small cell lung cancer, glioblastoma, renal cell cancer, prostate cancer, liver cancer, pancreatic cancer, soft-tissue sarcoma, kaposi’s sarcoma, carcinoid carcinoma, head and neck cancer, ovarian cancer, and mesothelioma. In some embodiments, the cancer is selected from: small cell lung cancer, glioblastoma, neuroblastomas, melanoma, breast carcinoma, gastric cancer, colorectal cancer (CRC), and hepatocellular carcinoma. Yet, in some embodiments, the cancer is selected from: non-small cell lung cancer, colorectal cancer, glioblastoma and breast carcinoma, including metastatic forms of those cancers. In some embodiments, the cancer is colorectal cancer, including coion cancer and rectal cancer. In some embodiments, the cancer is urothelial carcinoma, non-small cell lung cancer, triple negative breast cancer, small cell lung cancer, hepatocellular carcinoma, or melanoma.

[0232] In contrast, hematologic cancers originate in the blood or bone marrow. In some embodiments, the hematologic cancer to be treated with an anti-MerTK antibody of the present disclosure is leukemia. Examples of leukemias include, without limitation, chronic lymphocytic leukemia (CLL); acute lymphoblastic leukemia (ALL); hairy cell leukemia; chronic myeloblastic leukemia; and acute myeloblastic leukemia. In some embodiments, the hematologic cancer to be treated with an anti-MerTK antibody of the present disclosure is lymphoma. N on-limiting examples of lymphoma include T-cell lymphoma (such as adult T-cell leukemia/lymphoma; hepatosplenic T- cell lymphoma; peripheral T-cell lymphoma, anaplastic large ceil lymphoma; and angioimmunoblastic T ceil lymphoma), B-ceil lymphoma (including low' grade/follicular nonHodgkin’s lymphoma (NHL); small lymphocytic (SL) NHL; intermediate grade/follicular NHL; intermediate grade diffuse NHL; high grade immunoblastic NHL; high grade lymphoblastic NHL; high grade small non-cleaved cell NHL; bulky disease NHL; diffuse large B-cell lymphoma; mantle cell lymphoma; Burkitt lymphoma; AIDS-related lymphoma; and Waldenstrom’s Macroglobulinemia), Hodgkin’s lymphoma, and post-transplant lymphoproliferative disorder (PTLD). In some embodiments, the hematologic cancer to be treated with an anti-MerTK antibody of die present disclosure is myeloma. In a specific embodiment, the myeloma is plasmacytoma or multiple myeloma. In certain embodiments, cancers that are amenable to treatment by anti-MerTK antibodies of the present disclosure include non-Hodgkin’s lymphoma and multiple myeloma.

[0233] In another aspect, provided herein are methods for treating or delaying progression of cancer in an individual comprising administering to the individual an effective amount of an anti- MerTK antibody as described in the present disclosure. In some embodiments, the treatment results in a sustained response in the individual after cessation of the treatment. The methods described herein may find use in treating conditions where enhanced immunogenicity' is desired such as increasing tumor immunogenicity for the treatment of cancer. Also provided herein are methods of enhancing immune function in an individual having cancer comprising administering to the individual an effective amount of an anti-MerTK antibody as described in the present disclosure. In some embodiments, the cancer expresses functional STING, functional Cx43, and functional cGAS polypeptides. Functional proteins are proteins that are able to carry out their regular functions in a cell. Examples of functional proteins may include wild-type proteins, tagged proteins, and mutated proteins that retain or improve protein function as compared to a wild-type protein. Protein function can be measured by any method known to those of skill in the art, including assaying for protein or mRNA expression and sequencing genomic DNA or rnRN A. In some embodiments, the cancer comprises tumor-associated macrophages that express functional STING polypeptides. In some embodiments, the cancer comprises tumor cells that express functional cGAS polypeptides. In some embodiments, the cancer comprises tumor cells that express functional Cx43 polypeptides. In some embodiments, the cancer is colorectal cancer, including colon cancer and rectal cancer. In some embodiments, the cancer is urothelial carcinoma, non-small cell lung cancer, triple negative breast cancer, small cell lung cancer, hepatocellular carcinoma, or melanoma.

[0234] Also provided herein are methods of reducing MerTK-mediated clearance of apoptotic cells in an individual comprising administering to the individual an effective amount of an anti- MerTK antibody as described in the present disclosure to reduce MerTK-mediated clearance of apoptotic cells. In some embodiments, the clearance of apoptotic cells is reduced by 1-10 fold, 1-8 fold, 1-5 fold, 1-4 fold, 1-3 fold, 1-2 fold, 2-10 fold, 2-8 fold, 2-5 fold, 2-4 fold, 2-3 fold, 3-10 fold, 3- 8 fold, 3-5 fold, 3-4 fold, or by at least about 1.1 fold, 1.2 fold, 1.3 fold, 1.4 fold, 1.5 fold, 1.6 fold, 1.7 fold, 1.8 fold, 1.9 fold, 2.0 fold, 2.1 fold, 2.2 fold, 2.3 fold, 2.4 fold, 2.5 fold, 2.6 fold, 2.7 fold, 2.8 fold, 2.9 fold, 3.0 fold, 3.1 fold, 3.2 fold, 3.3 fold, 3.4 fold, 3.5 fold, 3.6 fold, 3.7 fold, 3.8 fold, 3.9 fold, 4.0 fold, 4.1 fold, 4.2 fold, 4.3 fold, 4.4 fold, 4.5 fold, 4.6 fold, 4.7 fold, 4.8 fold, 4.9 fold, 5.0 fold, 5. 1 fold, 5.2 fold, 5.3 fold, 5.4 fold, 5.5 fold, 5.6 fold, 5.7 fold, 5.8 fold, 5.9 fold, 6.0 fold, 6.1 fold, 6.2 fold, 6.3 fold, 6,4 fold, 6,5 fold, 6.6 fold, 6.7 fold, 6.8 fold, 6.9 fold, 7.0 fold, 7.1 fold, 7.2. fold, 7.3 fold, 7.4 fold, 7,5 fold, 7,6 fold, 7.7 fold, 7.8 fold, 7.9 fold, or 8.0 fold. Reduction of MerTK-mediated clearance of apoptotic cells may be determined by comparing the level of MerTK- mediated clearance of apoptotic cells in a sample from an individual after administration of an effective amount of an anti-MerTK antibody or an immunoconjugate t hereof to a reference level of MerTK-mediated clearance of apoptotic cells. In some embodiments, the reference level is the level of MerTK-mediated clearance of apoptotic cells a reference sample. In some embodiments, the reference sample is taken from the subject taken prior to administration of an effective amount of an anti-MerTK antibody or an immunoconjugate thereof. In some embodiments, the sample comprises tumor tissue or tumor cells.

[0235] In some embodiments, an anti-MerTK antibody of the present disclosure reduces phagocytic activity of apoptotic cells by about 10-100%, 20-100%, 30-100%, 40-100%, 50-100%, 60- 100%, 70-100%, 75-100%, 80- 100%, 85-100%, 90-100%, 95-100%, 10-95%, 20-95%, 30-95%, 40- 95%, 50-95%, 60-95%, 70-95%, 75-95%, 80-95%, 85-95%, 90-95%, 10-90%, 20-90%, 30-90%, 40-

90%, 50-90%, 60-90%, 70-90%, 75-90%, 80-90%, 85-90%, 10-85%, 2.0-85%, 30-85%, 40-85%, 50-

85%, 60-85%, 70-85%, 75-85%, 80-85%, 10-80%, 20-80%, 30-80%, 40-80%, 50-80%, 60-80%, 70-

80%, 75-80%, 10-75%, 20-75%, 30-75%, 40-75%, 50-75%, 60-75%, 70-75%, 10-70%, 20-70%, 30-

70%, 40-70%, 50-70%, 60-70%, 10-65%, 20-65%, 30-65%, 40-65%, 50-65%, 60-65%, 10-60%, 20-

60%, 30-60%, 40-60%, 50-60%, 10-55%, 20-55%. 30-55%, 40-55%, 50-55%, 10-40%, 20-40%, or

30-40%, or by at least about 10%, 20%, 30%, 40%, 50%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98% or 99%. In some embodiments, the anti-MerTK antibody has a half maximal inhibitory’ concentration (IC50) for reducing phagocytic activity of apoptotic ceils of about 1 pM - 50 pM, 1 pM - 100 pM, 1 pM - 500 pM, 1 pM - 1 nM, 1 pM - 1.5 nM, 5 pM - 50 pM, 5 pM - 100 pM, 5 pM - 500 pM, 5 pM - 1 nM, 5 pM - 1 .5 nM, 10 pM - 50 pM, 10 pM - 100 pM, 10 pM - 500 pM, 10 pM - 1 nM, 10 pM - 1.5 nM, 50 pM - 100 pM, 50 pM - 500 pM, 50 pM - 1 nM, 50 pM - 1.5 nM, 100 pM - 500 pM, 100 pM - I nM, or 100 pM - 1.5 nM.

[0236] The anti-MerTK antibody may be administered intravenously , intramuscularly, subcutaneously, topically, orally, transdermally, intraperitoneally, intraorbitaily, by implantation, by inhalation, intrathecaily, intraventncularly, or intranasally. The appropriate dosage of the anti-MerTK antibody may be determined based on the type of disease to be treated, the severity and course of the disease, the clinical condition of the individual, the individual’s clinical history' and response to the treatment, and the discretion of the attending physician.

2. Combinations with an additional therapy

[0237] Antibodies of the invention can be administered alone or used in a combination therapy. For instance, the combination therapy includes administering an antibody of the invention and administering at least one additional therapeutic agent (e.g. one, two, three, four, five, or six additional therapeutic agents). In certain aspects, the combination therapy comprises administering an antibody of the invention and administering at least one additional therapeutic agent, such as an immune checkpoint inhibitor.

[0238] In some embodiments, the uses and methods may further comprise an additional therapy or administration of an effective amount of an additional therapeutic agent. The additional therapy may be radiation therapy, surgery (e.g., lumpectomy and a mastectomy), chemotherapy, gene therapy, DNA therapy, viral therapy, RNA therapy, immunotherapy, bone marrow transplantation, nanotherapy, monoclonal antibody therapy, or a combination of the foregoing. The additional therapy may be in the form of adjuvant or neoadjuvant therapy. In some embodiments, the additional therapy is the administration of small molecule enzymatic inhibitor or anti -meta static agent. In some embodiments, the additional therapy is the administration of side-effect limiting agents (e.g., agents intended to lessen the occurrence and/or severity of side effects of treatment, such as anti -nausea agents, etc.). In some embodiments, the additional therapy is radiation therapy. In some embodiments, the additional therapy is surgery. In some embodiments, the additional therapy is a combination of radiation therapy and surgery. In some embodiments, the additional therapy is gamma irradiation. In some embodiments, the additional therapy is therapy targeting PI3K/AKT/mTOR pathway, I-ISP90 inhibitor, tubulin inhibitor, apoptosis inhibitor, and/or chemopreventative agent. [0239] In some embodiments, the additional therapy is an antagonist directed against B7-H3 (also known as CD276), e.g., a blocking antibody, MGA271, an antagonist directed against a TGF beta, e.g., metelimumab (also known as CAT-192), fresolimumab (also known as GCI008), or LY2157299, a treatment comprising adoptive transfer of a T cell (e.g., a cytotoxic T cell or CTL) expressing a chimeric antigen receptor (CAR), a treatment comprising adoptive transfer of a T cell comprising a dominant-negative TGF beta receptor, e.g, a dominant-negative TGF beta type II receptor, a treatment comprising a HERCREEM protocol (see, e.g., ClinicalTrials.gov Identifier NCT00889954), an agonist directed against CD137 (also known as TNFRSF9, 4- IBB, or ILA), e.g., an activating antibody, urelumab (also known as BMS-663513), an agonist directed against CD40, e.g., an activating antibody, CP-870893, an agonist directed against 0X40 (also known as CD 134), e.g., an activating antibody, administered in conjunction with a different anti-OX40 antibody (e.g., AgonOX)., an agonist directed against CD27, e.g., an activating antibody, CDX-1127, indoleamine- 2,3 -dioxygenase (IDO), 1-melhy i-D-tiyptophan (also known as 1-D-MT), an antibody-drug conjugate (in some embodiments, comprising mertansine or monomethy 1 auristatin E (MMAE)), an anti-NaPi2b antibody -MMAE conjugate (also known as DNIB0600A orRG7599), trastuzumab emtansine (also known as T-DM1, ado-trastuzumab emtansine, orKADCYLA®, Genentech), DMUC5754 A, an antibody -drug conjugate targeting the endothelin B receptor (EDNBR), e.g., an antibody directed against EDNBR conjugated with MMAE, an angiogenesis inhibitor , an antibody directed against a VEGF, e.g., VEGF-A, bevacizumab (also known as AVASTIN®, Genentech), an antibody directed against angiopoietin 2 (also known as Ang2), MEDI3617, an anti neo plastic agent, an agent targeting CSF-1R (also known as M-CSFR or CD115), anti-CSF-lR (also known as IMC- CS4), an interferon, for example interferon alplra or interferon gamma, Roferon-A, GM-CSF (also known as recombinant human granulocyte macrophage colony stimulating factor, rhu GM-CSF, sargramostim, or Leukine®), IL-2 (also known as aldesleukin or Proleukin®), IL-12, an antibody targeting CD20 (in some embodiments, the antibody targeting CD20 is obinntuzumab (also known as GA 101 or Gazyva®) or rituximab), an antibody targeting G1TR (in some embodiments, the antibody targeting GITR is TRX518), in conjunction with a cancer vaccine (in some embodiments, the cancer vaccine is a peptide cancer vaccine, which in some embodiments is a personalized peptide vaccine; in some embodiments the peptide cancer vaccine is a multivalent long peptide, a multi -peptide, a peptide cocktail, a hybrid peptide, or a peptide-pulsed dendritic cell vaccine (see, e.g., Yamada et al., Cancer Sci, 104: 14-21, 2013)), in conjunction with an adjuvant, a TLR agonist, e.g., Poly-ICLC (also known as Hiltonol®), LPS, MPL, or CpG ODN, tumor necrosis factor (TNF) alpha, IL-1, HMGB1, an IL-10 antagonist, an 11,-4 antagonist, an 11,-13 antagonist, an HVEM antagonist, an ICOS agonist, e.g., by administration of ICOS-L, or an agonistic antibody directed against ICOS, a treatment targeting CX3CL1, a treatment targeting CXCL10, a treatment targeting CCL5, anLFA-1 or ICAM1 agonist, a Selectin agonist, a targeted therapy, an inhibitor of B-Raf, vemurafenib (also known as Zelboraf®, dabrafenib (also known as Tafinlar®), erlotinib (also known as Tarceva®), an inhibitor of a MEK, such as MEKI (also known as MAP2K1) or MEK2 (also known as MAP2K2). cobimetinib (also known as GDC-0973 or XL-518), trametinib (also known as Mekinist®), an inhibitor of K-Ras, an inhibitor of c-Met, onartuzumab (also known as MetMAb), an inhibitor of Aik, AF802 (also known as CH5424802 or alectinib), an inhibitor of a phosphatidylinositol 3-kinase (PI3K), BKM120, idelalisib (also known as GS-1101 or CAL-iOl), perifosine (also known as KRX-0401), an Akt, MK2206, GSK690693, GDC-0941, an inhibitor of mTOR, sirolimus (also known as rapamycin), temsirolimus (also known as CCI-779 or Torisel®), everolimus (also known as RAD001), ridaforolimus (also known as AP-23573, MK-8669, or deforolimus), OSI-027, AZD8055, INK 128, a dual PI3K/mTOR inhibitor, XI., 765, GDC-0980, BEZ235 (also known as NVP-BEZ235), BGT226, GSK2126458, PF-04691502, or PF-05212384 (also known as PKI-587). In some embodiments, the additional therapeutic agent is CT- Oil (also known as Pidilizumab orMDV9300; CAS Registry' No. 1036730-42-3; CureTeclv'Medivation). CT-011, also known as IiBAT or IiBAT-1, is an antibody described in W02009/101611.

3. Combinations with checkpoint, inhibitors

[0240] In some embodiments, the additional therapeutic agent is an immune checkpoint inhibitor. In certain aspects, the application provides methods for enhancing immune function in an individual having cancer comprising administering an effective amount of a combination of an anti- MerTK antibody and an immune checkpoint inhibitor. In certain embodiments, the anti-AffiRTK antibody increases die immune effect of an immune checkpoint inhibitor by about 2 fold, 3 fold, 5 fold, 8 fold, 10 fold, 15 fold or 20 fold. In certain embodiments, the anti-AffiRTK antibody increases the immune effect of an immune checkpoint inhibitor by about 1-2 fold, 1-5 fold, 1-10 fold, 1-15 fold, 1-20 fold, 1-25 fold, 1-30 fold, 1-50 Ibid, 1-75 Ibid, 1-100 fold, 1-150 fold, 1-200 fold, 1-250 fold, 1.5-2 fold, 1.5-5 fold, 1.5-10 fold, 1.5-15 fold, 1.5-20 fold, 1.5-25 fold, 1.5-30 fold, 1.5-50 fold, 1.5- 75 fold, 1.5-100 fold, 1.5-150 fold, 1.5-200 fold, 1.5-250 fold, 2-5 fold, 2-10 fold, 2-15 fold, 2-20 fold, 2-25 fold, 2-30 fold, 2-50 fold, 2-75 fold, 2-100 fold, 2-150 fold, 2-200 fold, 2-250 fold, 2.5-5 fold, 2.5-10 fold, 2.5-15 fold, 2.5-20 fold, 2.5-25 fold, 2.5-30 fold, 2.5-50 fold, 2.5-75 fold, 2.5-100 fold, 2.5-150 fold, 2.5-200 fold, 2.5-250 fold, 5-10 fold, 5-15 fold, 5-20 fold, 5-25 fold, 5-30 fold, 5- 50 fold, 5-75 fold, 5-100 fold, 5-150 fold, 5-2.00 fold, 5-250 fold, 10-15 fold, 10-20 fold, 10-25 fold, 10-30 fold, 10-50 fold, 10-75 fold, 10-100 fold, 10-150 fold, 10-2.00 fold, 10-250 fold, 20-25 fold, 2.0- 30 fold, 20-50 foid, 20-75 fold, 20-100 fold, 20-150 foid, 20-200 fold, 20-250 fold, 25-30 fold, 25-50 fold, 25-75 fold, 25-100 fold, 25-150 fold, 25-200 foid, or 25-250 fold or by at least about 1 fold, 2 fold, 5 fold, 10 fold, 15 foid 20 fold 25 fold, 30 fold, 40 fold, 50 fold 60 fold, 70 fold, 75 foid, 80 fold, 90 fold, 100 fold, 125 fold, 150 foid, 200 fold, 225 fold or 250 foid.

[0241] In some embodiments, the individual has cancer that is resistant (has been demonstrated to be resistant) to one or more immune checkpoint inhibitors. In some embodiments, resistance to immune checkpoint inhibitors includes recurrence of cancer or refractory' cancer. Recurrence may refer to the reappearance of cancer, in the original site or a new site, after treatment. In some embodiments, resistance to immune checkpoint inhibitors includes progression of the cancer during treatment with the immune checkpoint inhibitors. In some embodiments, resistance to immune checkpoint inhibitors includes cancer that does not respond to treatment. The cancer may be resistant at the beginning of treatment or it may become resistant during treatment. In some embodiments, the cancer is at early stage or at late stage.

[0242] Further details regarding therapeutic immune checkpoint inhibitors are provided below and in, e.g., Byun et al. (2017) Nat Rev Endocrinol. 13: 195-207; La-Beck et al. (2015) Pharmacotherapy. 35(10): 963-976; Buchbinder et al. (2016) Am J Clin Oncol. 39(1): 98-106; Michot et al. (2016) Eur J Cancer. 54: 139-148, and Topalian el al. (2.016) Nat Rev Cancer. 16: 275-287. [0243] In some embodiments, the immune checkpoint inhibitor is a cytotoxic T-lymphocyte- associated protein 4 (CTLA4) (also known as CD 152) inhibitor. In some embodiments, the C'TLA-4 inhibitor is a blocking antibody, ipilimumab (also known as MDX-010, MDX-101, or Yen'oy®), tremelimumab (also known as ticilimumab or CP-675, 206).

[0244] In some embodiments, the immune checkpoint inhibitor is a PD-1 axis binding antagonist.

[0245] Provided herein are methods for treating cancer in an individual comprising administering to the individual an effective amount of a PD-1 axis binding antagonist and an anti-MerTK antibody of the present disclosure (e.g., a PEGylated anti-MerTK antibody). Also provided herein are methods of enhancing immune function or response in an individual (e.g., an individual having cancer) comprising administering to the individual an effective amount of a PD-1 axis binding antagonist and an anti-MerTK antibody of the present disclosure (e.g., a PEGylated anti-MerTK antibody).

[0246] In such methods, the PD-1 axis binding antagonist includes a PD-1 binding antagonist, a PDL1 binding antagonist, and/or a PDL2 binding antagonist. Alternative names for “PD-1” include CD279 and SLEB2. Alternative names for “PDL1” include B7-H1, B7-4, CD274, and B7-H. Alternative names for “PDL2” include B7-DC, Btdc, and CD273. In some embodiments, PD-1, PDL1, and PDL2 are human PD-1, PDL1 and PDL2.

[0247] In some embodiments, the PD-1 binding antagonist is a molecule tliat inhibits the binding of PD-1 to its ligand binding partner(s). In a specific aspect the PD-1 ligand binding partners are PDL1 and/or PDL2. In another embodiment, a PDL1 binding antagonist is a molecule that inhibits the binding of PDL1 to its binding partner(s). In a specific aspect, PDL1 binding partner(s) are PD-1 and/or B7- 1 . In another embodiment, the PDL2 binding antagonist is a molecule that inhibits the binding of PDL2 to its binding partner(s). In a specific aspect, a PDL2 binding partner is PD-1. The antagonist may be an antibody, an antigen binding fragment thereof, an immunoadhesin, a fusion protein, an oligopeptide or a small molecule. If the antagonist is an antibody, in some embodiments the antibody comprises a human constant region selected from die group consisting of IgGl, IgG2, IgG3 and IgG4.

[0248] In some embodiments, the PD-1 binding antagonist is an anti-PD-1 antibody (e.g., a human antibody, a humanized antibody, or a chimeric antibody). A variety of anti-PD-1 antibodies can be utilized in the methods disclosed herein. In any of the embodiments herein, the PD-I antibody can bind to a human PD-1 or a variant thereof. In some embodiments the anti-PD- i antibody is a monoclonal antibody. In some embodiments, the anti-PD-1 antibody is an antibody fragment selected from the group consisting of Fab, Fab’, Fab’-SH, Fv, scFv, and (Fab’)i fragments. In some embodiments, the anti-PD-1 antibody is a chimeric or humanized antibody. In other embodiments, the anti-PD- 1 antibody is a human antibody.

[0249] In some embodiments, the anti-PD-1 antibody is nivolumab (CAS Regisby Number: 946414-94-4). Nivolumab (Bristol-Myers Squibb/Ono), also known as MDX-1106-04, MDX-1106, ONO-4538, BMS-936558, and OPDIVO®, is an anti-PD-1 antibody described in W02006/121168.

In some embodiments, the anti-PD-1 antibody comprises a heavy chain and a light chain sequence, wherein:

(a) the heavy chain comprises the amino acid sequence:

QVQLVESGGGVVQPGRSLRLDCKASGITFSNSGMI-IWVRQAPGKGLEWVAVIWY DGSKRYYADSVKGRFTISRDNSKNTLFLQMNSLRAEDTAVYYCATNDDYWGQGTLVTVSS ASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLY SLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPPCPPCPAPEFLGGPSVFLFPPKP KDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTV LHQDWLNGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVK GFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEAL HNHYTQKSLSLSLGK (SEQ ID NO: 24), and

(b) ilie light chain comprises the amino acid sequence:

EIVLTQSPATLSLSPGERATLSCRASQSVSSYLAWYQQKPGQAPRLLIYDASNRAT GIPARFSGSGSGTOFTLTISSLEPEDFAVYYCQQSSNWPRTFGQGTKVEIKRTVAAPSVFIFPPS DEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSK ADVT.KHKVY ACEV THQGI -SSPVT 'KSFNRGEC (SEQ ID NO: 25).

[0250] In some embodiments, the anti-PD-1 antibody comprises the six HVR sequences from SEQ ID NO: 24 and SEQ ID NO: 25 (e.g., the three heavy chain HVRs from SEQ ID NO:24 and the three light chain HVRs from SEQ ID NO: 25). In some embodiments, the anti-PD-1 antibody comprises the heavy chain variable domain from SEQ ID NO: 24 and the light chain variable domain from SEQ ID NO: 25.

[0251] In some embodiments, the anti-PD-1 antibody is pembrolizumab (CAS Registry Number: 1374853-91-4). Pembrolizumab (Merck), also known as MK-3475, Merck 3475, lambrolizumab, SCH-900475, and KEYTRUDA®, is an anti-PD-1 antibody described in W02009/114335. In some embodiments, die anti-PD-1 antibody comprises a heavy chain and a light chain sequence, wherein:

(a) the heavy chain comprises the amino acid sequence:

QVQLVQSGVEVKKPGASVKVSCKASGYTFTNYYMYWVRQAPGQGLEWMGG INPSNGGTNFNEKFKNRVTLTTDSSTTTAYMELKSLQFDDTAVYYCARRDYRFDMGFDYW GQGTTVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGV HTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPPCPPCP APEFLGGPSWLFPPKPKDTL.MISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTK PREEQFNSTYRWSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISKAK GQPREPQ\^ZYTI.PPSQEEMTKNQVSLTCLVKGFYPSDIA\⁷EWESNGQPENN

WTTPPVI,DSDGSFFLYSRI..TWKSRWQEGNVTSCSVMHEAL.HNHYTQKSLSLSI..GK (SEQ ID NO: 26), and

(b) the light chain comprises the amino acid sequence: EIVLTQSPAT LSLSPGERATLSCRASKGVSTSGYSYLHWYQQKPGQAPRLLrYLASYLES GVPARFSGSGSGTDFTLTISSLEPEDFAVYYCQHSRDLPLTFGGGTKVEIKRTVAAPSVF IFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVl’EQ DSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 27). [0252] In some embodiments, the anti-PD-1 antibody comprises the six HVR sequences from SEQ ID NO: 26 and SEQ ID NO: 27 (e.g., the three heavy chain HVRs from SEQ ID NO: 26 and the three light chain HVRs from SEQ ID NO:27). In some embodiments, the anti-PD-1 antibody comprises the heavy chain variable domain from SEQ ID NO: 26 and the light chain variable domain from SEQ ID NO: 27.

[0253] In some embodiments, the anti-PD-1 antibody is MEDI-0680 (AMP-514: AstraZeneca). MEDI-0680 is a humanized IgG4 anti-PD-1 antibody.

[0254] In some embodiments, the anti-PD-1 antibody is PDR001 (CAS Registry No. 1859072- 53-9; Novartis). PDR001 is a humanized IgG4 anti-PD-1 antibody that blocks the binding of PDL1 and PDL2 to PD-1.

[0255] In some embodiments, the anti-PD-1 antibody is REGN2810 (Regeneron). REGN28I0 is a human anti-PD-1 antibody.

[0256] In some embodiments, the anti-PD-1 antibody is BGB-108 (BeiGene). In some embodiments, the anti-PD-1 antibody is BGB-A317 (BeiGene).

[0257] In some embodiments, the anti-PD-1 antibody is JS-001 (Shanghai Junshi). JS-001 is a humanized anti-PD-1 antibody.

[0258] In some embodiments, the anti-PD-1 antibody is STI-A1110 (Sorrento). STI-Ali lO is a human anti-PD- 1 antibody ,

[0259] In some embodiments, the anti-PD-1 antibody is INCSHR-1210 (Incyte). INCSHR-1210 is a human lgG4 anti-PD-1 antibody.

[0260] In some embodiments, the anti-PD-1 antibody is PF-06801591 (Pfizer).

[0261] In some embodiments, the anti-PD- 1 antibody is TSR-042 (also known as ANB011;

Tesaro/AnaptysBio).

[0262] In some embodiments, the anti-PD-1 antibody is AM0001 (ARMO Biosciences). [0263] In some embodiments, the anti-PD-1 antibody is ENUM 244C8 (Enumeral Biomedical Holdings). ENUM 244C8 is an anti-PD-1 antibody that inhibits PD-1 function without blocking binding of PDL1 to PD-1.

[0264] In some embodiments, the anti-PD-1 antibody is ENUM 388D4 (Enumeral Biomedical Holdings), ENUM 388D4 is an anti-PD-1 antibody that competitively inhibits binding of PDL1 to PD-1.

[0265] In some embodiments, the PD-1 antibody comprises the six H VR sequences (e.g. , the three heavy chain H VRs and the three light chain HVRs) and/or the heavy chain variable domain and light chain variable domain from a PD-1 antibody described in W02015/112800 (Applicant: Regeneron), W02015/112805 (Applicant: Regeneron), WO2015/112900 (Applicant: Novartis), US20150210769 (Assigned to Novartis), WO2016/089873 (Applicant: Celgene), W02015/035606 (Applicant: Beigene), WO2015/085847 (Applicants: Shanghai Hengrui Pharmaceutical/Jiangsu Hengrui Medicine), W02014/206107 (Applicants: Shanghai Junshi Biosciences/Junmeng Biosciences), WO2012/145493 (Applicant: Amplimmune), US9205148 (Assigned to Medlmmune), W02015/119930 (Applicants: Pfizer, 'Merck), WO2015/119923 (Applicants: Pfizer/Merck), WO2016/032927 (Applicants: Pfizer/Merck), WO2014/179664 (Applicant: AnaptysBio), W02016/106160 (Applicant: Enumeral), and WO2014/194302 (Applicant: Sorrento).

[0266] In some embodiments, the PD-1 axis binding antagonist is an anti-PDLl antibody. A variety of anti-PDLl antibodies are contemplated and described herein. In any of the embodiments herein, the isolated anti-PDLl antibody can bind to a human PDL1, for example a human PDL1 as shown in UniProtKB/Swiss-Prol Accession No.Q9NZQ7.1, or a variant thereof. In some embodiments, the anti-PDLl antibody is capable of inhibiting binding between PDL1 and PD-1 and/or between PDL1 and B7-1. In some embodiments, the anti-PDLl antibody is a monoclonal antibody. In some embodiments, the anti-PDLl antibody is an antibody fragment selected from the group consisting of Fab, Fab’-SH, Fv, scFv, and (Fab’)j fragments. In some embodiments, the anti- PDLl antibody is a humanized antibody. In some embodiments, the anti-PDLl antibody is a human antibody. Examples of anti-PDLl antibodies useful for the methods of the present disclosure, and methods for making thereof are described in PCT patent application WO 2010/077634 A 1 and US Patent No. 8,217, 149, which are incorporated herein by reference.

[0267] In some embodiments, the anti-PDLl antibody is atezolizumab (CAS Registry Number: 1422185-06-5). Atezolizumab (Genentech), also known as MPDL3280A, is an anti-PDLl antibody, [0268] In some embodiments, the anti-PDLl antibody comprises a heavy chain variable region and a light chain variable region, wherein:

(a) the heavy chain variable region comprises an l-IVR-Hl, HVR-H2, and HVR-H3 sequence of GFTFSDSW1H (SEQ ID NO: 28), AWISPYGGSTYYADSVKG (SEQ ID NO: 29) and RHWPGGFDY (SEQ ID NO: 30), respectively, and (b) the light chain variable region comprises an HVR-L 1 , HVR-L2, and HVR-L3 sequence of RASQDVSTA VA (SEQ ID NO: 31), SASFLYS (SEQ TD NO: 32) and QQYLYHPAT (SEQ ID NO: 33), respective!}'.

[0269] In some embodiments, the anti-PDLl antibody is MPDL3280A, also known as atezolizumab and TECENTRIQ® (CAS Regisriy Number: 1422185-06-5). In some embodiments, the anti-PDLl antibody comprises a heavy chain and a light drain sequence, wherein:

(a) the heavy drain variable region sequence comprises the amino acid sequence:

EVQLVESGGGLVQPGGSLRLSCAASGFTFSDSWIHWVRQAPGKGLEWVAWISPYGGSTYYA DSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCrXRRHWPGGFDYWGQGTLVrVSS (SEQ ID NO: 34), and

(b) the light chain variable region sequence comprises the amino acid sequence: DIQMTQSPSSLSASVGDRVTITCRASQDVSTAVAWYQQKPGKAPKLL1Y SASF LYSGVPSRFSGSGSGTOFTLTISSLQPEDFATYYCQQYLYHPATFGQGTKVEIKR (SEQ ID NO: 35).

[0270] In some embodiments, the anti-PDLl antibody comprises a heavy drain and a light drain sequence, wherein:

(a) the heavy chain comprises the amino acid sequence:

EVQLVESGGGLVQPGGSLRLSCAASGFTFSDSWIHWVRQAPGKGLEWVAWISPYGGSTYYA DSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCARRHWPGGFDYWGQGTLVTVSSAST KGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVTITFPAVLQSSGLYSLS SVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKP

KDTLMISRTPEVTCVWDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYASTYRVVSVLTV LHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVK GFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEAL HNHYTQKSLSLSPG (SEQ ID NO: 36), and

(b) the light chain comprises the amino acid sequence:

DIQMTQSPSSLSASVGDRVTITCRASQDVSTAVAWYQQKPGKAPKLLIYSASFLYSGVPSRFS GSGSGTOFTLTISSLQPEDFATYYCQQYLYHPATFGQGTKVEIKRTVAAPSVFIFPPSDEQLKS GTASWCLLNNFYPREAKVQWVDNAI.QSGNSQESVTEQDSKDSTYSLSSTI^'n.SKADYEK HKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 37)

[0271] In some embodiments, the anti-PDLl antibody is avelumab (CAS Registry Number:

1537032-82-8). Avelumab, also known as MSB0010718C, is a human monoclonal IgGl anti-PDLl antibody (Merck KGaA, Pfizer). In some embodiments, the anti-PDLl antibody comprises a heavy drain and a light chain sequence, wherein:

(a) the heavy chain comprises the amino acid sequence:

EVQLLESGGGLVQPGGSLRLSCAASGFTFSSY1MMWVRQAPGKGLEWVSS1YPSGG1TFYAD TVKGRFT1SRDNSKNTLYLQMNSLRAEDTAVYYCARIKLGTVTTVDYWGQGTLVTVSSAST KGPSVFPLAPSSKSTSGGTAALGCLV'KDYFPEPVTVSWNSGALTSGV'HTFPAVLQSSGLYSLS SVVTVPSSSLGTQTYTCNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKP KDTLMISRTPEVTCVWDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWSVLTV LHQDWI.NGKEYKCKVSNKALPAPIEK’nSKAKGQPREPQVYTIRPSRDELTKNQVSLTCLVK GFYPSDIAVEWESNGQPENNYKTTPPVT.,DSDGSFFI>YSKLTVDKSRWQQGNVFSCSVMHEAL HNHYTQKSLSLSPG (SEQ ID NO: 38), and

(b) Hie light chain comprises the amino acid sequence:

QSALTQPASVSGSPGQSmSCTGTSSDVGGYNYVSWYQQHPGKAPKLMIYDVSNRPSGVSN RFSGSKSGNTASLTISGLQAEDEADYYCSSYTSSSTRVFGTGTKVTVLGQPKANPTVTLFPPSS EELQANKATLVCLISDFYPGAVTVAWKADGSPVKAGVETTKPSKQSNNKYAASSYLSLTPEQ WKSHRSYSCQVTHEGSTVEKTVAPTECS (SEQ ID NO: 39).

[0272] In some embodiments, the anti-PDLl antibody comprises the six HVR sequences from SEQ ID NO: 38 and SEQ ID NO: 39 (e.g., the three heavy chain HVRs from SEQ ID NO:38 and the three light chain HVRs from SEQ ID NO: 39). In some embodiments, the anti-PDLl antibody comprises the heavy chain variable domain from SEQ ID NO: 38 and the light chain variable domain from SEQ ID NO: 39.

[0273] In some embodiments, the anti-PDLl antibody is durvalumab (CAS Registry Number: 1428935-60-7), Durvalumab, also known as MEDI4736, is an Fc optimized human monoclonal IgGl kappa anti-PDLl antibody (Medlmmune, AstraZeneca) described in WO2011/066389 and US2013/034559. In some embodiments, the anti-PDLl antibody comprises a heavy chain and a light chain sequence, wherein:

(a) the heavy chain comprises the amino acid sequence:

EVQLVESGGGLVQPGGSLRLSCAASGFTFSRYWMSWVRQAPGKGLEWVAN1KQDGSEKYY VDSVKGRFT1SRDNAKNSLYLQMNSLRAEDTAVYYCAREGGWFGELAFDYWGQGTLVTVS SASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGL YSLSSVVTVPSSSLGTQ’nqCNVNHKPSNTKV'DKRVEPKSCDKTHTCPPCPAPEFEGGPSVFLF PPKPKDTLMISRTPEVTCVVVT)VSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVA⁷S VLTVLHQDWLNGKEYKCKVSNKALPASTEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLT CLVKGFYPSDIA\RWESNGQPENNYKTTPPVTDSDGSFFI,YSKLTVDKSRWQQGNVFSCSV MHEALHNHYTQKSLSLSPG (SEQ ID NO: 40), and

(b) the light chain comprises the amino acid sequence: EIVLTQSPGTLSLSPGERATLSCRASQRVSSSYLAWYQQKPGQAPRLLIYDASSRATGIPDRFS GSGSGTOFTLTISRLEPEDFAVYYCQQYGSLPWTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKS GTASWCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEK HKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 41).

[0274] In some embodiments, the anti-PDLl antibody comprises the six HVR sequences from SEQ ID NO:40 and SEQ ID NO:41 (e.g., the three heavy chain HVRs from SEQ ID NO:40 and the three light chain HVRs from SEQ ID N0:41). In some embodiments, the anti-PDLl antibody comprises the heavy chain variable domain from SEQ ID NO: 40 and the light chain variable domain from SEQ ID NO: 41.

[0275] In some embodiments, the anti-PDLl antibody is MDX-1105 (Bristol Myers Squibb). MDX-1105, also known as BMS-936559, is an anti-PDLl antibody described in W02007/005874. [0276] In some embodiments, the anti-PDLl antibody is LY3300054 (Eli Lilly).

[0277] In some embodiments, the anti-PDLl antibody is STI-A1014 (Sorrento). STI-A1014 is a human anti-PDLl antibody.

[0278] In some embodiments, the anti-PDLl antibody is KN035 (Suzhou Alphamab). KN035 is single-domain antibody (dAB) generated from a camel phage display libraiy.

[0279] In some embodiments, the anti-PDLl antibody comprises a cleavable moiety or linker that, when cleaved (e.g. , by a protease in the tumor microenvironment), activates an antibody antigen binding domain to allow it to bind its antigen, e.g., by removing a non-binding steric moiety. In some embodiments, the anti-PDLl antibody is CX-072 (CytomX Therapeutics).

[0280] In some embodiments, the PDL1 antibody comprises the six HVR sequences (e.g., the three heavy drain H VRs and the three light chain HVRs) and/or the heavy chain variable domain and light chain variable domain from a PDL 1 antibody described in US20160108123 (Assigned to Novartis), W02016/000619 (Applicant: Beigene), WO2012/145493 (Applicant: Amplimmune), US9205148 (Assigned to Medlmmune), WO2013/181634 (Applicant: Sorrento), and W02016/061142 (Applicant: Novartis).

[0281] In a still further specific aspect, the PD-1 or PDL 1 antibody lias reduced or minimal effector function. In a still further specific aspect the minimal effector function results from an “effector-less Fc mutation” or a glycosylation mutation. In still a further embodiment, the effectorless Fc mutation is an N297A or D265A/N297A substitution in the constant region. In some embodiments, the isolated anti-PDLl antibody is aglycosvlated. Glycosylation of antibodies is typically either N-linked or O-linked. N-linked refers to the attachment of the carbohydrate moiety to the side chain of an asparagine residue. The tripeptide sequences asparagine-X-serine and asparagine- X-threonine, where X is any amino acid except proline, are the recognition sequences for enzymatic attachment of the carbohydrate moiety to the asparagine side drain. Thus, the presence of either of these tripeptide sequences in a polypeptide creates a potential glycosylation site. O-linked glycosylation refers to the attachment of one of the sugars N-aceylgalactosamine, galactose, or xylose to a hydroxyamino acid, most commonly serine or threonine, although 5 -hydroxy proline or 5- hydroxylysine may also be used. Removal of glycosylation sites form an antibody is conveniently accomplished by altering the amino acid sequence such that one of the above-described tripeptide sequences (for N-linked glycosylation sites) is removed. The alteration may be made by substitution of an asparagine, serine or threonine residue within the glycosylation site another amino acid residue (e.g., glycine, alanine or a conservative substitution). [0282] In some embodiments, the anti-MERTK antibody increases the immune effect of the anti- PDL1 antibody about 3 fold after 20 days of combination treatment. In some embodiments, the anti- MERTK antibody increases the immune effect of the anti-PDLl antibody about 10 fold after 30 days of treatment.

[0283] In some embodiments, the PD-1 binding antagonist is an immunoadhesin (e.g., an immunoadhesin comprising an extracellular or PD- 1 binding portion of PDL 1 or PDL2 fused to a constant region (e.g., an Fc region of an immunoglobulin sequence). In some embodiments, the PD-1 binding antagonist is AMP-224. AMP-224 (CAS Registry No. 1422184-00-6;

GlaxoSnrithKline/Medlimnune), also known as B7-DCIg, is a PDL2-Fc fusion soluble receptor described in W02010/027827 and WO2011/066342.

[0284] In some embodiments, the PD-1 binding antagonist is a peptide or small molecule compound. In some embodiments, the PD-1 binding antagonist is AUNP-I2 (PierreFabre/Aurigene). See, e.g., WO2012/ 168944, WO2015/036927, WO2015/044900, W02015/033303, WO2013/144704, WO2013/132317, and WO2011/161699.

[0285] In some embodiments, the PDL1 binding antagonist is a small molecule that inhibits PD- 1 . In some embodiments, the PDL I binding antagonist is a small molecule that inhibits PDL1 . In some embodiments, the PDL1 binding antagonist is a small molecule that inhibits PDL1 and VISTA. In some embodiments, the PDL1 binding antagonist is CA-170 (also known as AUPM-170). In some embodiments, the PDL1 binding antagonist is a small molecule tliat inhibits PDL1 and TIM3, In some embodiments, the small molecule is a compound described in W02015/033301 and WO2015/033299.

[0286] In another aspect, provided herein are methods for enhancing immune function in an individual having cancer comprising administering an effective amount of a combination of an anti- MerTK antibody (e.g., a PEGylated anti-MerTK antibody) and an immune checkpoint inhibitor. Various aspects of immune function that may be enhanced by the anti-MerTK antibodies described herein and methods for measuring such enhancement are described below.

[0287] In some embodiments of the methods of the present disclosure, the cancer (in some embodiments, a sample of the patient’s cancer as examined using a diagnostic test) lias elevated levels of T cell infiltration. As used herein, T cell infiltration of a cancer may refer to the presence of T cells, such as tumor-infiltrating lymphocytes (TILs), within or otherwise associated with the cancer tissue. It is known in the art that T cell infiltration may be associated with improved clinical outcome in certain cancers (see, e.g., Zhang etal., N. Engl. J. Med. 348(3):203-213 (2003)).

[0288] However, T cell exhaustion is aiso a major immunological feature of cancer, with many tumor-infiltrating lymphocytes (TILs) expressing high levels of inhibitory co-receptors and lacking the capacity to produce effector cytokines (Wherry', E.J. Nature immunology 12: 492-499 (2011); Rabinovich, G.A., et al., Annual review of immunology 25:267-296 (2007)). In some embodiments of the methods of the present disclosure, the individual has a T cell dysfunctional disorder. In some embodiments of the methods of the present disclosure, the T ceil dysfunctional disorder is characterized by T cell anergy or decreased ability’ to secrete cytokines, proliferate or execute cytolytic activity’. In some embodiments of the methods of the present disclosure, the T cell dysfunctional disorder is characterized by T ceil exhaustion. In some embodiments of the methods of the present disclosure, the T cells are CD4+ and CD8+ T cells. In some embodiments, the T cells are CD4-1- and/or CD8+ T cells.

[0289] In some embodiments, CD8+ T cells are characterized, e.g., by presence of CD8b expression (e.g., by rtPCR using e.g., Fluidigm) (Cd8b is also known as T-celi surface glycoprotein CD8 beta chain; CD8 antigen, alpha polypeptide p37; Accession No. is NM_172213). In some embodiments, CD8+ T ceils aie from peripheral blood. In some embodiments, CD8+ T cells are from tumor.

[0290] In some embodiments, Treg cells are characterized, e.g., by presence of Fox3p expression (e.g., by rtPCR e.g., using Fluidigm) (Foxp3 is also known as forkhead box protein P3; scurfin;

FOXP3delta7; immunodeficiency, polyendocrinopathy, enteropathy, X-linked; the accession no. is NM 014009). In some embodiments, Treg are from peripheral blood. In some embodiments, Treg cells are from tumor.

[0291] In some embodiments, inflammatory T cells are characterized, e.g,, by presence of Tbet and/or CXCR3 expression (e.g., by rtPCR using, e.g., Fluidigm). In some embodiments, inflammatory' T cells are from peripheral blood. In some embodiments, inflammatojy T ceils are from tumor.

[0292] In some embodiments of the methods of the present disclosure, CD4 and/or CD8 T cells exhibit increased release of cy tokines selected from the group consisting of IFN- y, TNF-a and interleukins. Cytokine release may be measured by any means known in the art, e.g., using Western blot, ELISA, or immunohistochemical assays to detect the presence of released cytokines in a sample containing CD4 and/or CD8 T cells.

[0293] In some embodiments of the methods of the present disclosure, the CD4 and/or CD8 T ceils are effector memoiy T cells. In some embodiments of the methods of the present disclosure, the CD4 and/or CD8 effector memoiy T ceils are characterized by having the expression of CD44^hlgh CD62L^|OW. Expression of CD44^mgh CD62L^!OB may be detected by any means known in the art, e.g., by preparing single cell suspensions of tissue (e.g., a cancer tissue) and performing surface staining and flow cytometry using commercial antibodies against CD44 and CD62L, In some embodiments of the methods of the present disclosure, the CD4 and/or CD8 effector memory T cells are characterized by having expression of CXCR3 (also known as C-X-C chemokine receptor type 3; Mig receptor; IP10 receptor; G protein-coupled receptor 9; interferon-inducible protein 10 receptor; Accession No.

NM 001504). In some embodiments, the CD4 and/or CD8 effector memory’ T cells are from peripheral blood. In some embodiments, the CD4 and/or CD8 effector memoiy T cells are from tumor. [0294] In some embodiments of the methods of the present disclosure, Treg function is suppressed relative to prior to the administration of the combination. In some embodiments, T cell exhaustion is decreased relative to prior to the administration of the combination.

[0295] In some embodiments, number of Treg is decreased relative to prior to the administration of the combination. In some embodiments, plasma interferon gamma is increased relative to prior to the administration of the combination, Treg number may be assessed, e.g., by determining percentage of CD4+Fox3p-t- CD454- cells (e.g., by FACS analysis). In some embodiments, absolute number of Treg, e.g., in a sample, is determined. In some embodiments, Treg are from peripheral blood. In some embodiments, Treg are from tumor.

[0296] In some embodiments, T cell priming, activation and/or proliferation is increased relative to prior to the administration of the combination. In some embodiments, the T cells are CD4+ and/or CD8+ T cells. In some embodiments, T cell proliferation is detected by determining percentage of K167+ CD8+ T cells (e.g., by FACS analysis). In some embodiments, T cell proliferation is detected by determining percentage of Ki67+ CD4+ T cells (e.g., by FACS analysis). In some embodiments, the T cells are from peripheral blood. In some embodiments, the T cells are from tumor.

[0297] Such combination therapies noted above encompass combined administration (where two or more therapeutic agents are included in the same or separate pharmaceutical compositions), and separate administration, in which case, administration of the antibody of the invention can occur prior to, simultaneously, and/or following, administration of the additional therapeutic agent or agents. In one aspect, administration of the anti-MerTK antibody and administration of an additional therapeutic agent occur within about one month, or within about one, two or three weeks, or within about one, two, three, four, five, or six days, of each other. In one aspect, the antibody and additional therapeutic agent are administered to the patient on Day 1 of the treatment. Antibodies of the invention can also be used in combination with radiation therapy.

[0298] An antibody of the invention (and any additional therapeutic agent) can be administered by any suitable means, including parenteral, intrapulmonary, and intranasal, and, if desired for local treatment, intralesional administration. Parenteral infusions include intramuscular, intravenous, intraarterial, intraperitoneal, or subcutaneous administration. Dosing can be by any suitable route, e.g., by injections, such as intravenous or subcutaneous injections, depending in part on whether the administration is brief or chronic. Various dosing schedules including but not limited to single or multiple administrations over various time-points, bolus administration, and pulse infusion are contemplated herein.

[0299] Armbodies of the invention would be formulated, dosed, and administered in a fashion consistent with good medical practice. Factors for consideration in this context include the particular disorder being treated, the particular mammal being treated, the clinical condition of the individual patient, the cause of the disorder, the site of delivery of the agent, die method of administration, the scheduling of administration, and other factors known to medical practitioners. The antibody need not be, but is optionally formulated with one or more agents currently used to prevent or treat the disorder in question. The effective amount of such other agents depends on the amount of antibody present in the pharmaceutical composition, the type of disorder or treatment, and other factors discussed above. These are generally used in the same dosages and with administration routes as described herein, or about from 1 io 99% of the dosages described herein, or in any dosage and by any route that is empirically /clinically determined to be appropriate.

[0300] For the prevention or treatment of disease, the appropriate dosage of an antibody of the invention (when used alone or in combination with one or more other additional therapeutic agents) will depend on the type of disease to be treated, the type of antibody', the seventy and course of the disease, whether the antibody is administered for preventive or therapeutic purposes, previous therapy, the patient’s clinical history' and response to the antibody, and the discretion of the attending physician. The antibody is suitably administered to the patient at one time or over a series of treatments. Depending on the type and severity of the disease, about 1 p.g/kg to 15 mg/kg (e.g., 0. i mg/kg- lOmg/kg) of antibody can be an initial candidate dosage for administration to the patient, whether, for example, by one or more separate administrations, or by continuous infusion. One typical daily dosage might range from about 1 pg/kg to 100 mg/kg or more, depending on the factors mentioned above. For repeated administrations over several days or longer, depending on the condition, the treatment would generally be sustained until a desired suppression of disease sy mptoms occurs. One exemplary' dosage of the antibody would be in the range from about 0.05 mg/kg to about 10 mg/kg. Thus, one or more doses of about 0.5 mg/kg, 2.0 mg/kg, 4.0 mg/kg or 10 mg/kg (or any combination thereof) may be administered to the patient. Such doses may be administered intermittently, e.g., every' week or every three weeks (e.g., such that the patient receives from about tw'O to about twenty, or, e.g., about six doses of the antibody). An initial higher loading dose, followed by one or more lower doses may be administered.

[0301] Any of the anti-MerTK antibodies described herein and any immune checkpoint inhibitors known in the art or described herein may be used in the methods of the present disclosure. [0302] In some embodiments, the combination therapy of the present disclosure comprises administration of an anti-MerTK antibody and an immune checkpoint inhibitor. The anti-MerTK antibody and the immune checkpoint inhibitor may be administered in any suitable manner known in the art. For example, the anti-MerTK antibody and the immune checkpoint inhibitor may be administered sequentially (at different times) or concurrently (at the same time). In some embodiments, the immune checkpoint inhibitor is in a separate composition as the anti-MerTK antibody. In some embodiments, the immune checkpoint inhibitor is in the same composition as the anti-MerTK antibody .

[0303] The anti-MerTK antibody and the immune checkpoint inhibitor may be administered by the same route of administration or by different routes of administration. In some embodiments, the immune checkpoint inhibitor is administered intravenously , intramuscularly, subcutaneously, topically, orally, transdermally, intraperitoneally, intraorb itallv, by implantation, by inhalation, intrathecally, intraventricuiarly, or intranasally. In some embodiments, the anti-MerTK antibody is administered intravenously, intramuscularly, subcutaneously, topically, orally, transdermally, intraperitoneally, intraorbitally, by implantation, by inhalation, intrathecally, intraventricnlarly, or intranasally. An effective amount of the immune checkpoint inhibitor and the anti-MerTK antibody may be administered for prevention or treatment of disease. The appropriate dosage of the anti- MerTK antibody and/or the immune checkpoint inhibitor may be determined based on the type of disease to be treated, the type of the immune checkpoint inhibitor and the anti-MerTK antibody, the severity and course of the disease, the clinical condition of the individual, the individual’s clinical histoiy and response to the treatment, and the discretion of the attending physician. In some embodiments, combination treatment with anti-MerTK antibody and an immune checkpoint inhibitor (e.g., anti- PD-1 or anti-PDLl antibody) are synergistic, whereby an efficacious dose of an anti- MerTK antibody in the combination is reduced relative to efficacious dose of the anti-MerTK antibody as a single agent.

[0304] As a general proposition, the therapeutically effective amount of the antibody administered to human will be in the range of about 0.01 to about 50 mg/kg of patient body weight whether by one or more administrations. In some embodiments, the antibody used is about 0.01 to about 45 mg/kg, about 0.01 to about 40 mg/kg, about 0.01 to about 35 mg/kg, about 0.01 to about 30 mg/kg, about 0.01 to about 25 mg/kg, about 0.01 to about 20 mg/kg, about 0.01 to about 15 mg/kg, about 0.01 to about 10 mg/kg, about 0.01 to about 5 mg/kg, or about 0.01 to about 1 mg/kg administered daily, for example. In some embodiments, the antibody is administered at 15 mg/kg. However, other dosage regimens may be useful. In one embodiment, an anti-MerTK antibody described herein or an anti-PDLl antibody described herein is administered to a human at a dose of about 100 mg, about 200 mg, about 300 mg, about 400 mg, about 500 mg, about 600 mg, about 700 mg, about 800 mg, about 900 mg, about 1000 mg, about 1100 mg, about 1200 mg, about 1300 mg or about 1400 mg on day 1 of 21 -day cycles. The dose may be administered as a single dose or as multiple doses (e.g., 2 or 3 doses), such as infusions. The dose of the antibody administered in a combination treatment may be reduced as compared to a single treatment. The progress of this therapy is easily monitored by conventional techniques.

[0305] The progress of this therapy is easily monitored by conventional techniques and assays, [0306] In one aspect, the present disclosure provides the anti-MerTK antibodies as described above (e.g. , PEGylated anti-MerTK antibodies) for use as a medicament. In some embodiments, the use is in treating cancer. In some embodiments, the use is in reducing MerTK-mediated clearance of apoptolic cells. Further provided herein are uses of the anti-MerTK antibodies as described above in the manufacture of a medicament. In some embodiments, the medicament is for treatment of cancer. In some embodiments, the cancer expresses functional cGAS-STING cytosolic DNA sensing pathway proteins. These proteins are part of the cGAS-STING signaling pathway and function in innate immunity to detect the presence of cytosolic DNA in order to trigger the expression of inflammatory genes. Examples of cGAS-STTNG cytosolic DNA sensing pathway proteins include but are not limited to cGAS, STING, TBK-1, IRF3, p50, p60, p65, NF-KB, ISRE, IKK, and STAT6. In some embodiments, the cancer expresses functional STING, functional Cx43, and functional cGAS polypeptides. Functional proteins are proteins that are able to carry out their regular functions in a cell. Examples of functional proteins may include wild-type proteins, tagged proteins, and mutated proteins that retain or improve protein function as compared to a wild-type protein. Protein function can be measured by any methods known to those of skill in the art, including assaying for protein or mRNA expression and sequencing genomic DNA or mRNA. In some embodiments, the cancer comprises tumor-associated macrophages that express functional STING polypeptides. In some embodiments, the cancer comprises tumor cells that express functional cGAS polypeptides. In some embodiments, the cancer comprises tumor cells that express functional Cx43 polypeptides. In certain embodiments, the cancer is colon cancer. In some embodiments, the cancer is urothelial carcinoma, non-small cell lung cancer, triple negative breast cancer, small cell lung cancer, hepatocellular carcinoma, or melanoma. In some embodiments, the medicament is for reducing MerTK-mediated clearance of apoptotic ceils.

[0307] In another aspect, the individual has cancer that expresses (has been shown to express e.g., in a diagnostic test) PDL1 biomarker. In some embodiments, the patient’s cancer expresses low PDLI biomarker. In some embodiments, the patient’s cancer expresses high PDLI biomarker. In some embodiments of any of the methods, assays and/or kits, the PDL1 biomarker is absent from the sample when it comprises 0% of the sample.

[0308] In some embodiments of any of the methods, assays and'br kits, the PDL1 bio marker is present in the sample when it comprises more than 0% of the sample. In some embodiments, the PDLI biomarker is present in at least 1% of the sample. In some embodiments, the PDLI biomarker is present in at least 5% of the sample. In some embodiments, the PDLI biomarker is present in at least 10% oftbe sample.

[0309] In some embodiments of any of the methods, assays and/or kits, the PDLI biomarker is detected in the sample using a method selected from the group consisting of FACS, Western biot, El, ISA, immunoprecipitation, immunohistochemistty, immunofluorescence, radioimmunoassay, dot blotting, immunodetection methods, HPLC, surface plasmon resonance, optical spectroscopy, mass spectre metery, HPLC, qPCR, RT-qPCR, multiplex qPCR or RT-qPCR, RNA-seq, microarray analysis, SAGE, MassARRAY technique, and FISH, and combinations thereof.

[0310] In some embodiments of any of the methods, assays and/or kits, the PDLI bio marker is detected in the sample by protein expression. In some embodiments, protein expression is determined by immunohistochemistty (IHC). In some embodiments, the PDLI biomarker is detected using an anti-PDLl antibody. In some embodiments, the PDLI biomarker is detected as a weak staining intensity by IHC, In some embodiments, the PDL 1 biomarker is detected as a moderate staining intensity by IHC. In some embodiments, the PDL1 biomarker is detected as a strong staining intensity by IHC. In some embodiments, the PDL1 biomarker is detected on tumor cells, tumor infiltrating immune cells, stromal cells and any combinations thereof. In some embodiments, the staining is membrane staining, cytoplasmic staining or combinations thereof.

[0311] In some embodiments of any of the methods, assays and/or kits, the absence of the PDL1 biomarker is detected as absent or no staining in the sample. In some embodiments of any of the methods, assays and/or kits, the presence of the PDL1 biomarker is detected as any staining in the sample.

G. Articles of Manufacture

[0312] In another aspect of the invention, an article of manufacture containing materials useful for the treatment and/or prevention of the disorders described above is provided. The article of manufacture comprises a container and a label or package insert on or associated with the container. Suitable containers include, for example, bottles, vials, sy ringes, IV solution bags, etc. The containers may be formed from a variety of materials such as glass or plastic. The container holds a composition which is by itself or combined with another composition effective for treating, preventing and/or diagnosing the condition and may have a sterile access port (for example the container may be an intravenous solution bag or a vial having a stopper pierceable by a hypodermic injection needle). At least one active agent in the composition is an antibody of the invention. The label or package insert indicates that the composition is used for treating the condition of choice. Moreover, the article of manufacture may comprise (a) a first container with a composition contained therein, wherein the composition comprises an antibody of the invention; and (b) a second container with a composition contained therein, wherein the composition comprises a further cytotoxic or otherwise therapeutic agent. The article of manufacture in this aspect of the invention may further comprise a package insert indicating that the compositions can be used to treat a particular condition. Alternatively, or additionally, the article of manufacture may further comprise a second (or third) container comprising a pharmaceutically -acceptable buffer, such as bacteriostatic water for injection (B WFI), phosphate- buffered saline, Ringer's solution and dextrose solution. It may further include other materials desirable from a commercial and user standpoint, including other buffers, diluents, filters, needles, and syringes.

[0313] The specification is considered to be sufficient to enable one skilled in the art to practice the compositions and methods of the present disclosure. Various modifications in addition to those shown and described herein will become apparent to those skilled in the art from the foregoing description and fall within the scope of the appended claims. All publications, patents, and patent applications cited herein are hereby incorporated by reference in their entirety⁷ for all purposes. EXAMPLES

[0314] The present disclosure will be more fully understood by reference to the following examples. They should not, however, be construed as limiting the scope of the disclosure. It is understood that the examples and embodiments described herein are for illustrative purposes only and tliat various modifica tions or changes in light thereof will be suggested to persons skilled in the art and are to be included within the spirit and purview of this application and scope of the appended claims.

Example 1: On-target ocular toxicity with anti-MerTK antibody treatment

[0315] MerTK is expressed on the apical membrane of RPE cells and plays a critical role in the phagocytosis of shedding photoreceptor cells. The RPE of MerTK loss-of-function and constitutive knockout rodents is unable to remove shedding photoreceptor outer segments by phagocytosis resulting in retinal degeneration. Humans with defective MerTK function develop night blindness in early childhood followed by a rapid decrease in visual acuity into early adulthood. Retinal toxicities have been observed with MerTK blockade with a small molecule MerTK inhibitor and an anti- MERTK monoclonal antibody (mAb).

[0316] This example describes the assessment of potential retinal toxicities associated with administration of an anti-MerTK monoclonal antibody.

Materials & Methods

First pilot toxicology study

[0317] In a first pilot toxicology study (Toxicology Study 1) with C57BL/6N mice, 14C9 anti- MerTK mAb was intravenously (TV) administered twice weekly (BIW) for 4 weeks (9 total doses) at doses of 0 (control), 10 and 30 mg/kg, Male and female mice (n=4/sex/group) w ere assigned to the toxicity groups and we re scheduled fo r necropsy at the end of the dosing period (Study Day 32). Blood for from additional mice in the 14C9 mAb treated groups (m=6/sex/group) was collected for toxicokmetic evaluation. The evaluation criteria included die following parameters: clinical observations, body weight, clinical pathology (hematology and clinical chemistry ), anatomic pathology of major organs of interest, and toxicokinetics.

Second pilot toxicology study

[0318] To determine whether the retinal lesions observed in Toxicology Study 1 were associated with functional impairment of the retina, a subsequent pilot toxicology study (Toxicology study 2) with BALB/c mice incorporating full-field electroretinogram (ffERG) was conducted. In this study, 14C9 mAh was administered IV at 0, 5 or 30 mg/kg twice weekly (BIW; total 8 doses) or 45 mg/kg thrice weekly (TIW; 12 total doses) for 4 weeks, with a 6-week recovery' period. Male and female mice (n=4/sex/group) were assigned to the toxicity groups at all dose levels, and terminal and recovery' necropsies were on Study Days 30 and 72, respectively. Blood for toxicokinetic evaluation was collected from additional mice in the 14C9 mAb-treated groups (n=6/sex/group). Criteria for evaluation included the following parameters: clinical observations, body weight, clinical pathology (hematology and clinical chemistry'), anatomic pathology of the eye and testis, ffERG, and toxicokinetics.

Third pilot toxicology study

[0319] A third pilot toxicology study was conducted in cynomolgus monkeys. 13B4 ami-MerTK mAb was intravenously (IV) administered as a single dose or repeat dose every 3 weeks (Q3 W) for 6 weeks. Female monkeys (n=3/group) were assigned to each dose group, and terminal and recovery necropsies were on Study Day 45. Animals receiving a single dose were administered 10 mg/kg, and animals dosed Q3W were administered 10 or 30 mg/kg (total of 3 administrations). The evaluation ccriteria included the following parameters: clinical observations, food consumption, body weight, physical examinations, ophthalmologic examinations, fundus ocular photography, intraocular pressure, ERG (full-field [ffERG] and mult-focal [mfERG]), optical coherence tomography (OCT), clinical pathology (e.g. hematology, clinical chemistry and urinalysis), anatomic pathology', toxicokinetics, and anti-drug antibody measurements.

Results

[0320] MerTK is required for turnover of photoreceptor outer segment (POS) cells, and continuous blockade of MerTK leads to retinal toxicity (FIG. 1A). To assess the potential retinal toxicities associated with administration of an anti-MerTK mAb, three toxicology studies were conducted.

[0321] The first of these toxicology studies. Toxicology study 1, was conducted in C57BL/6N mice with the mouse surrogate anti-MerTK mAb, 14C9. In this first study, mice were administered 14C9 mAb was twice weekly for 4 weeks at doses of 0, 10 and 30 mg/kg. An increase in the area under the curve (AUG) was observed in the toxocokinetic curve for 14C9 exposure. This increase was dose-proportional, as it was observed with an increase in dose from 10 to 30 mg/kg.

One male mouse in the 30 mg/kg dose group was found dead on Study Day 5, with the cause of death considered unrelated to 14C9 treatment. Two male mice in the 10 mg/kg dose group and one male mouse in the 30 mg/kg dose group were moribund and were euthanized early on Study Day s 22 and 18, respectively. The cause of death in these animals was considered to be procedure-related and not related to 14C9 administration. For the remaining animals which survived until terminal necropsy, analysis of the necropsy sampels revealed 14C9-related toxicity in the eye and testes. 14C9~related outer retinal degeneration, consisting of minimal -to -mild photoreceptor (PR) vacuolation, increased cellularity in the PR layer, and drop down of the outer nuclear layer was observed in these mice, with a dose -dependent increase in incidence and severity. Additionally, in the testes, moderate-to-marked seminiferous tubule atrophy and degeneration (correlating with decreased testis weight) was observed in all 14C9-treated male mice without any dose-dependency. Based on these findings, it was concluded tlrat administration of 14C9 mAb to C57BL/6N mice at 10 and 30 mg/kg BIW for 4-weeks resulted in retinal and testicular toxicity.

[0322] To determine whether the retinal lesions observed in Toxicology Study 1 were associated with functional impairment of the retina, a subsequent pilot toxicology study (Toxicology study 2) was conducted. This second toxicology study was conducted with BALB/c mice and incorporated full-field electroretinogram (ffERG). In tills study, 14C9 mAb was administered at 0, 5 or 30 mg/kg twice weekly or 45 mg/kg thrice weekly for 4 weeks, with a 6-week recovery’ period. Toxicokinetic analysis revealed a dose-proportional increase in AUC for 14C9 exposure in this study, witli an increase in dose from 10 to 30 mg/kg BIW, and 45 mg/kg TIW.

[0323] One female mouse in the 30 mg/kg dose group was found dead on Study Day 15, with the cause of death not considered as related to 14C9 treatment. One male mouse in the 30 mg/kg dose group was moribund and euthanized early on Study Day 21 , with the cause of death considered to be anesthesia-related. For the remaining animals in Toxicology Study 2. that survived until terminal necropsy, the clinical pathology findings were limited to a reversible, minimally -to-mildly increase in alanine transaminase (ALT) and aspartate transaminase (AST) levels in individual animals, as well as minimally elevated globulins and a decrease in tire albumin/globulin ratio at a 45 mg/kg dose. As shown in FIG. 1C, 14C9-reIated outer retinal degeneration, consisting of minimal-to-marked PR vacuolization, increased cellularity in the PR layer, drop down of the outer nuclear layer, and decreased cellularity with degeneration or necrosis of the outer nuclear layer, was also observed, with a dose-dependent increase in severity. Similar findings were observed in one control male. The retinal lesions correlated with decreases in PR func tion as assessed by ffERG, neither of which resolved during the recovery- period. Marked seminiferous tubule atrophy and degeneration without any dose- dependency was observed in the testes of all 14C9 administered male mice.

[0324] Based on the results of Toxicology Study 2, it was concluded that administration of 14C9 mAb to BALB/c mice at 10 or 30 mg/kg BIW, or 45 mg/kg TIW for 4-weeks resulted in irreversible retinal and testicular lesions, and functional impairment of the PRs. The retinal lesions observed in BALB/c mice (albino strain) were significantly more extensive than those observed in C57BL6/N mice (pigmented strain) during Toxicology Study 1, suggesting that there may be strain-related differences in susceptibility to anti-MerTK mAb retinal effects.

[0325] Finally, a third toxicology study (Toxicology Study 3) was conducted in cymologus monkeys witli unconjugated I3B4 anti-MerTK mAb having a hlgGl isotype with LALAPG mutation. Anti-MerTK mAb was intravenously (IV) administered as a single dose of 10 mg/kg or a repeat dose of 10 or 30 mg/kg every 3 weeks (Q3 W) for 6 weeks. Female monkeys (n=3/group) were assigned to each dose group, and terminal and recovery necropsies were on Study Day 45.

[0326] All animals survived to their scheduled necropsies on Study Days 45. Changes in clinical pathology parameters were limited to minimally increased AST and mildly increased ALT levels (without histologic correlates) in all dose groups, which returned to baseline in animals receiving a single administration of 10 mg/kg of 13B4. 13B4-related microscopic findings in lymphoid tissues were observed in animals administered 13B4 > 10 mg/kg Q3W and included minimal or slight decreased lymphocytes in the thymus (correlating with decreased thymus weights in animals administered 30 mg/kg), and minimally increased kaiyorrhectic debris in follicular germinal centers in the spleen, mesenteric and mandibular lymph nodes, and GALT/Peyer’s patch.

[0327] As shown in FIG. IB, minimal macrophage infiltration was observed in the photoreceptor layer in one animal at 30 mg/kg Q3W. No evidence of photoreceptor damage and no changes in ffERG, mfERG and OCT were observed for this group. In addition, no changes in ERG were observed for the 10 mg/kg Q3W group. Evidence of pharmacodynamic (PD) effect was observed at both dose levels, presented as increased cell-free DN and Increased apoptotic debris in follicular germinal center, presumably due to impairment of clearance by resident macrophages. From the results of Toxicology Study 3, it can be concluded that, administration of 13B4 mAb to cynomolgus monkeys as either a single 10 mg/kg dose or repeat dose at 10 or 30 mg/kg Q3W (3 total administrations) was well-tolerated, with 13B4-related microscopic findings in ly mphoid tissues and tiie eye of animals receiving multiple doses of 30 mg/kg 13B4.

[0328] Based on the results of these toxicology studies, the key safety risk of retinal toxicity is thought to be due to on-target inhibition of MerTK -mediated RPE engulfment of shedding photoreceptor outer segments. The retinal toxicity may also be driven by time over threshold, and based on the available data, there is a lack of monitorability in cynomolgus and mouse by either ERG or OCT analysis.

Example 2: Conjugating large PEG polymers to anti-MerTK antibodies via engineered cysteines

[0329] This example described the production of polymer-conjugated anti-MerTK antibodies.

Materials and Methods

Preparation ofTHIOMAB antibody for conjugation

[0330] Antibodies were produced with engineered cysteine at various positions, expressed in

CHO cells and purified via standard methods, including Protein A affinity chromatography followed by size exclusion chromatography. Conjugatable PEG polymers with a reactive maieimide moiety and nominal molecular weight of 40 kDa were purchased from NOF America, catalog numbers GL2- 400MA (2 -arm branched) and ME-400MA (linear). [0331] The engineered cysteine in the THIOMAB antibody format is commonly blocked with cysteine or glutathione that occurs during the expression in mammalian cells. A standard process for deblocking was earned out to remove the cysteine or glutathione from the engineered cysteine for conjugation of the desired moiety. Briefly, 50- 100 molar excess of reducing agent, DTT, was added to a 10 mg/mL antibody at alkaline pH of 7.5-8,5 in 100 mM Tris pH 8.5, 150 mM NaCl, 2 mM EDTA, and the mix was incubated at room temperature for approximately 18 hours. The antibody was then purified using cationic exchange to remove the DTT and the reduced cysteine & glutathione. The partially reduced antibody was re-oxidized using 15 molar excess of DHAA for 2-3 hours at room temperature and in 20 mM Tris pH 7. The oxidation stale was assessed using LC-MS to check for the mass of intact oxidized antibody and monitor the presence of free light chain. The re-oxdized antibody was then purified by cationic exchange chromatography and formulated with 10 mM succinate pH 5, 150 mM NaCl, 2 mM EDTA.

Preparation of PEG conjugates

[0332] Conjugation of PEG polymers to the THIOMAB antibody were produced as follows. A 4-

6 molar equivalents of PEG dissolved in 10 mM succinate pH 5 was added to a 10 mg/mL antibody solution, which had the pH was adjusted with IM Hepes pH 7.2 to a final concentration of 100 mM Hepes pH 7.2. The conjugation reaction proceeded at room temperature for approximately 3 to 18 hours until a maximal antibody -to-poly mer ratio was achieved. The progress of the conjugation reaction was monitored using an HPLC equipped with a YARRA SEC-4000 column that can resolve antibody to polymer ratios of 0, 1 and 2. The conjugation reaction was then purified using hydrophobic interaction chromatography. The pH of the conjugation reaction was adjusted to pH 6.5 with IM sodium acetate pH 5.0 and a final concentration of 800 mM ammonium sulfate was added. The conjugate was then purified using a ProPAC HIC-10 bonded silica 10x150 mm column and formulated into 20 rnM histidine acetate pH 5.5, 240 mM sucrose, 0.02% Tween-20 using dialysis. UV absorbance at 280 nm was used to determine the antibody -based concentration of the formulated conjugates, given that PEG does not absorb at 280 nm, in combination with extinction coefficient of the antibody^ component,

[0333] The polymer to antibody ratio and percent aggregation were characterized using HPLC equipped with YARRA SEC-4000 with 0.2. M Potassium phosphate, 0.25M Potassium chloride, pH 6,2, 15% ispropanol. The endotoxin levels were determined using Charles River EndoSafe cartridges. The hydrody namic radius of the conjugates were determined using an UPLC SEC-MALS/QELS Wyatt system equipped with a Acclaim-1000 column, with the resulting data being processed and analyzed using the Astra 7.1.2 software package.

Binding affinity determination

[0334] For binding affinity determinations of 14C9.C90S THIOMAB conjugates in IgGs, Surface Plasmon Resonance (SRP) measurements with a Biacore'™-8K instrument (GE Healthcare) were used. Briefly, each IgG variant was captured by Series S sensor chip Protein A (29127555, GE Healthcare) to achieve approximately 100RU, and then 3-fold serial dilutions of mouse MerTK (0.6 nM to 50 nM) were injected in HBS-EP buffer (100 mM 4-(2-hydroxyethyl)-l- piperazineethanesulfonic acid (HEPES) pH 7.4, 150 mM NaCl, 3 mM EDTA, 0.05% (v/v) Surfactant P20) at 25°C with a flow rate of 50ul/min. Association rates (k_on) and dissociation rates (k₀£f) were calculated using a simple one-to-one Langmuir binding model (Biacore Insight software version 2.0). The equilibrium dissociation constant (K_D) was calculated as the ratio k_Off/k_c,_n.

[0335] For binding affinity determinations of 14C9.C90S_LC_K149C series of THIOMAB conjugates in Fabs, Surface Plasmon Resonance (SRP) measurement with a Biacore™-T200 instrument (GE Healthcare) was used. Briefly, human Fc -tagged mouse MerTK (R&D 591 -MR) was captured by protein A sensor chip to achieve approximately 50RIJ, and then 3-fold serial dilutions of each Fab variant (0.6 nM to 50 nM) were injected in HBS-EP buffer (100 mM 4-(2-hydroxyethyl)-l- piperazineethanesulfonic acid (HEPES) pH 7.4, 150 mM NaCl, 3 mM EDTA, 0.05% (v/v) Surfactant P20) at 25°C with a flow rate of 50j.il/min. Association rates (k_on) and dissociation rates (k_Off) were calculated using a simple one-to-one Langmuir binding model (BIAcore T2.00 evaluation software version 2.0). The equilibrium dissociation constant (K_D) was calculated as the ratio koa/kon.

Results

[0336] It was hypothesized that increasing the size (e.g., hydrodynamic radius) of the antibody via conjugation of polymers may reduce the ocular uptake of the anti-MerTK antibody, and reduce the on-target ocular toxicity observed with the parental antibody. Hydrophilic polymers such as PEG have been used traditionally to increase the systemic half-life of small molecules and antibody fragments, and more recently to increase the vitreous half-life of intravitreal administered biologies. The rationale is that by increasing the size of the drug through conjugation of PEG, clearance is significantly reduced due to bypassing glomerular filtration cutoff for systemic administration and bypassing retinal clearance mechanisms for IVT. Increasing the size of the anti-MerTK antibody may decrease the distribution to the eye by preventing the antibody from crossing the blood-retinal barrier, while maintaining the ability of the antibody to bind MerTK on tumor associated macrophages in the tumor micro-environment, though there has not been a systematic study addressing the effect of size on penetration of the blood-retinal barrier (del Amo el at. (2018) Progress in Retinal and Eye Research 57:134-185).

[0337] Hydrophilic polymers, such as PEG, are available in a wide variety of formats with different sizes available, formats (linear, branched) and reactive moieties for conjugation to the antibody. Compared to other hydrophilic polymers, PEG has been widely used in the clinic with several approvals forPEGylated proteins and antibody fragments. Two formats of PEG, linear and 2- arm branched format, at a nominal molecular weight of 40 kDa were chosen for assessment, as previous data has suggested that the format of the PEG can impact the pharmacokinetics and biodistribution (Leong, S.R. et al. (2001) Cytokine 16: 106- 119; Vugmeyster, Y. et al. (2012) Bioconjugate chemistry 23: 1452-1462).

[0338] Site-specific cysteines were engineered into the antibody framework for conjugation for both the murine and human anti-MerTK antibodies using the cysteine-engineered (THIOMAB) antibody technology. The cysteine-maleimide conjugation chemistry in combination with the THIOMAB antibody allows for site-specific conjugation that enables production of homogenous products regarding antibody -to-polymer ratio and enables control of the stability of the connection between the antibody and the polymer. A set of predicted stable sites for the anti-murine MerTK m!gG2a format that would likely translate to the known stable sites on the anti-human MerTK huIgGl format were screened (Ohri, R. et al. (2018) Bioconjugate Chemistry 29:473-485), with selection of these sites based on sequence and structure alignment of the m!gG2a framework and human IgGl framework. Three sites on the constant domain of the Fab region of the antibody were chosen for engineering a cysteine, LC K183, LC K149C and HC T182C.

[0339] Both the linear and 2 -arm branched, 40 kDa PEG polymers with maleimide moiety reacted readily with the deblocked, reduced cysteines of THIOMAB antibodies to produce polymer- antibody conjugates with a polymer to antibody ratio of at least 1 .8 after 3-18 hour incubation at ambient temperature with less than 10% aggregation observed during the course of the reaction. The maximum expected polymer to antibody ratio is 2, since each single engineered cysteine appears twice in the heterodimeric antibody. Conjugation of the 40 kDa PEG was monitored using SEC- HPLC with a YARRA SEC-4000 column that was able to separate antibody -polymer ratios of 2 and 1 from unconjugated antibody. After purification and formulation, the antibody -to-polymer ratio was enriched to an average of 2.0 with 0% aggregation, routinely observed across the different conjugation sites, in both tire anti -murine 14C9 m!gG2a and the anti-human 13134 huIgGl antibodies. A schematic of the PEG-conjugates formed following this approach is shown in FIG. 2.

[0340] After conjugation, the hydrodynamic ratio of the anti-MerTK PEG conjugates was characterized. The hydrodynamic radius and molecular weight of the anti-MerTK PEG conjugates was determined using UPLC equipped with MATS and QELS detector, and samples were runned over an ACCLAIM-1000 SEC column with phosphate buffered saline. The molecular weight of the anti-MerTK 14C9 antibody and the PEG conjugates were observed to agree with the theoretical molecular weight values (Table 2). The hydrodynamic radius of the unconjugated antibody was determined to 5 nm, in good agreement with reported values. The hydrodynamic radii of the PEGylated conjugates was determined to be twice tliat observed for the unconjugated antibody, with the linear PEG-40K slightly larger at 10.8 nm compared to the 2-ann branched PEG-40K at 10.3 nm (Table 2).

Table 2. Hydrodynamic radius of the parental anti-murine MerTK antibody and THIOMAB antibody- PEG conjugates as measured by SEC-MALS/QELS.

[0341] The binding affinitities of the anti-MerTK PEG conjugates was assessed by Biacore analysis. As shown in FIGs. 3A-3B, conjugation of PEG to 14C9 appeared to have moderate improvement (3-5 fold) in binding affinity to mouse MerTK extracellular domain (ECD). No difference was observed between different sites of engineered cysteines on the Fab domain.

Furthermore, as shown in FIG. 4, a moderate increase (3-fold) in binding affinity’ to MerTK ECD was observed when binding was analyzed with surface plasmon resonance (SPR).

Conclusions

[0342] Both the murine anti-MerTK and human anti-MerTK THIOMAB antibodies readily conjugated with maleimide linked linear and 2-arm branched 40 kDa PEG polymers, producing conjugates with an antibody to polymer ratio of 2.0 and < 5% aggregation in tire purified, formulated conjugates. The hydrodynamic radius of the conjugates was approximately two-times larger than the unconjugated antibody with the linear PEGvlated antibody with a marginally larger radius than the branched version.

Example 3: In vitro efferocytosis and macrophage binding analyses ofPEGylated anti-MerTK antibodies

[0343] This example describes the inhibition of macrophage-mediated efferocytosis by anti- MerTK antibody conjugates.

Materials and Methods

Reagents

[0344] Cytokine mouse M-CSF was obtained from PeproTech (Rocky Hill, NJ). Staurosporine from Streptomyces sp. was obtained from Sigma-Aldrich (Saint Louis, MI). pHrodo Red, succinimidyl ester (pHrodo Red SE) was obtained from Thermo Scientific, (Whaltham, MA).

Mouse Peritoneal Macrophages

[0345] Mouse peritoneal macrophages (MPMs) were either obtained from Cell Biologies (Cat" C57-6032TF, Chicago, IL) or isoiated freshly in-house. For isolation of MPMs, 8 weeks old C57-BL6 mice were used. The mice were first euthanized by COr and 10 mL of chilled PBS (with 3% FBS) was injected into the peritoneal cavity of each mouse. After drawing the fluid back to the same syringe, the collected peritoneal cell suspensions were centrifuged at 1,400 rpm for 5 min and the pelleted cells were resuspended inRPMI medium, supplemented with 10% FBS. Commercial MPMs (6 million cells) were cultured in RPMI medium supplemented with 10% FBS and recombinant mouse M-CSF (30 ng/mL) in a temperature-sensitive Nunc UpCell 10-cm dish (Thermo Scientific, Wbaltham, MA) for 3 days to promote recovery. After recovery, MPMs were harvested by washing with chilled-PBS without the use of dissociation enzymes.

Preparation of pHrodo-Labeled Apoptotic Jurkat Cells

[0346] Jurkat cells (ATCC #T1B-I5zs were cultured in RPMI medium supplemented with 10% FBS. Cells from an exponentially growing culture were harvested and induced to undergo apoptosis by treatment with 1.0 uM Staurosporine for a period of 4 hours at room temperature. The cells were then washed twice and re-suspended in PBS to a density of 1.0 x 10⁶ cells/ml. The apoptotic cells were then labeled by incubating in 1.0 pM of pHrodo Red, succinimidyl ester in die dark, at room temperature for 1 hour. After labeling, the apoptotic cells were washed with PBS and a 10 min slow- spin centrifugation (750xg) 3 times. The cells were then re-suspended in RPMI medium + 10% FBS and then used in the efferocytosis assays.

Real-Time Imaging Efferocytosis Assay via IncuCyte Zoom

[0347] Macrophages were seeded overnight at a density of 4.0 x 10⁴ cells/well on a 96-well, low- evaporation Nunclon Delta Surface plate (Thermo Scientific) in RPMI medium supplemented with 10% FBS. Serial dilutions of antibodies were prepared in RPMI media supplemented with 10% FBS and then added to the 96-well plate containing the differentiated macrophages for 1 hr. After incubation with blocking antibodies, freshly prepared pHrodo-labeled apoptotic cells were added to the macrophages at a density of 8.0 x 10^s cells/well. Upon addition of apoptotic cells, the 96-well plate was placed in the IncuCyte Zoom instrument (Essen Biosciences; Ann Harbor, MI) and images were obtained every 15 minutes for a period of 24-48 hours using the lOx objective lens and the red channel. Total red fluorescence intensity was quantified using the IncuCyte Basic Software (2016B), with the background noise subtracted using the Top-Hat method. Cell confluency in each image was also quantified and was used to normalize the total red fluorescence intensity from different wells.

The efferocytosis activity was quantified as (total red fluorescence intensity’ - background fluorescence intensity’ of apoptotic cells)/(macrophage ceil confluence), and the maximum activity’ (100%) was defined as the value acquired in wells with untreated macrophages + apoptotic cells. Inhibition ciuves were generated using efferocytosis activities recorded 4-8 hours after addition of apoptotic cells. The efferocytosis activity from duplicate or triplicate wells were plotted as a function of antibody concentration and the data were fitted to a sigmoidal model with Prism (Graphpad Software; La Jolla, CA). The ICw value was calculated as die concentration of test material required to reduce the efferocytosis activity of macrophages by 50%,

Results

[0348] THIOMAB antibody -PEG conjugates (FIG. 2) were prepared as described in Example 2.

The ability of die PEG-conjugated anti-MerTK antibodies to inhibit mouse peritoneal macrophage- mediated efferocytosis was then evaluated. All conjugated antibodies inhibited mouse peritoneal macrophage-mediated efferocytosis (FIG. 5A). In particular, the PEG40K-B ranched conjugated 14C9 antibodies showed comparable potency as the parental mAb in inhibiting efferocytosis. The potency’ and efficacy of the 14C9 parental and PEG-conjugated antibodies are summarized in FIG. SB.

[0349] The 14C9 LC K 149C conjugates were chosen for further evaluation. These 14C9 PEG- antibody conjugates showed comparable inhibitory activities as the parent antibody in the mouse efferocytosis assay (FIGs. 6A-6B). Both branched and linear PEG conjugated antibodies inhibited mouse pentoneal macrophage-mediated efferocytosis. PEG40K-Branched conjugated 14C9 antibody showed a potency comparable to that of the parental mAb and was more potent than the PEG40K- Linear conjugated 14C9 antibody. The 14C9 LC K149C conjugates also exhibited around a 10-fold increase in EC50 cell binding as determined by flow cytometry (FIG. 7). The EC50s determined for binding to murine macrophages are summarized in Table 3.

Table 3. Cell-based binding of the parental anti-murine MerTK antibody and the TH1OMAB antibody -PEG conjugates to murine MerTK on primary peritoneal murine macrophages.

[0350] The efferocytosis assay was also performed to evaluate 14C9 Fab conjugates. As shown in FIGs. 8A-8B, the 14C9 Fab conjugates inhibited efferocytosis with similar potency as the unconjugated 14C9 Fab. A decrease in potency was observed for all Fabs as compared to the parental 14C9 antibody.

Example 4: In vivo characterization of'PEGylated anti-MerTK antibodies

[0351] This example describes the in vivo evaluation of PEG-conjugated anti-MerTK antibodies.

Materials and Methods

Single high dose study

[0352] For this study, female C57BL/6J mice were subcutaneously inoculated with 0. 1 million MC38 cells on the lower right flank. Animals were grouped based on weight and tumor volume to ensure similar weight and starting tumor volume distribution before treatment. Anti-gpl20 (control antibody), anti-MerTK or antibody conjugates were administered via intravenous (i.v) injection at a dose of 30 mg/kg of the antibody part. Two day later, tumors and eyes were collect.

[0353] For receptor occupancy assay, half tumors and eyes were dissociated to obtain single cell suspension. The cells were stained with live/dead dye (L 10119) and specific markers, followed by wash and fixation steps. Cells were then stained with anti-MerTK(14C9)-AF647 to assess MerTK receptor occupancy on tumor-associated macrophages (TAMs) and RPEs. For tumor pharmacodynamic assays, RNA was isolated from half tumors and qPCR assays were used to examine gene expression. Antibodies used: anti-CD45(clone 30-F1 1), anti-CDl Ibtclone MI/70), anti-CDl lc(clone HL3), anti-Ly6G(clone 1A8), anti-L.y6C(clone FIK1.4), anti-CD90.2 (clone 30- H12), anti-MHC Class II (M5/114.15.1), anti-F4/80 (clone BM8), anti-CD24 (clone GK1.5), anti- CD31(clone 390), and anti-RPE65 (401.8B11.3D9). Taqman probes used: IFNb (Mm00439546_sI), IFIT1 (Mm00515153_ml), USP18 (Mm01188805_ ml) and IRF7 (Mm00516793_gl).

Repeated high dose study

[0354] In this study, anti-gp!20 (control antibody), anti-MerTK or antibody conjugates were administered via intravenous (IV) injection at a dose of 45 rng/kg of the antibody part to naive (non- tumor-bearing) female C57BL/6J mice on both day 1 and day 5. On day 7, eyes were collected, and MerTK occupancy on RPEs was assessed as described in the single high dose study (16-3045 Y).

Repeated low dose study

[0355] A repeated low' dose study was earned out as described for the repeated high dose study except that the antibody or antibody’ conjugates were administered at a dose of 2.5 mg/'kg of the antibody part on both day 1 and day’ 5. On day 7, receptor occupancy on tumor associated macrophages (TAMs) and tumor pliarmacodynamic response were examined.

Results

[0356] During an initial study, the PEG-conjugated 14C9 anti-MerTK antibodies were administered as a single 30 rng/kg dose intravenously to MC38-turnor bearing mice. Eye and tumor samples were obtained to evaluate MerTK receptor occupancy, as well as for evaluation of induction of an interferon response in the tumors. As shown in FIG. 9, the parental antibody and the PEG- conjugated antibodies exhibited a comparable induction of interferon beta (IFNb) and interferon- stimulated genes (ISGs) in tumors. The PEG-conjugated antibodies marginally maintained occupancy of MerTK on retinal pigment epithelium cells (RPEs), but maintained occupancy on tumor-associated macrophages (TAMs) (FIGs. 10A-10C). Moreover, the mean fluorescence intensity (MFI) suggested partial occupancy, with around 70% or higher unoccupied MerTK remaining with the antibody conjugates as compared to around 27% with the parental antibody. With the parental antibody, around 20% of RPEs maintained occupancy of around 30% MerTK receptors, while w ith the antibody conjugates, at least 90% RPEs maintained occupancy of at least 70% MerTK receptors.

[0357] Following the initial single dose study, a repeated high dose study was conducted, with the purpose of assessing MerTK occupancy on RPEs after repeated treatment with high dose of anti- MerTK mAb (14C9) or Ab conjugates in non-tumor-bearing mice. In this study, anti-gpl20 (control antibody), anti-MerTK or antibody conjugates were administered by IV injection at a dose of 45 mg/'kg non-tumor-bearing mice on both day 1 and day 5 of the study. On day 7, eye samples were collected to evaluate MerTK occupancy on RPEs. [0358] The PEG-conjugated antibodies reduced MeiTK occupancy on RPEs following a repeated high dose of antibody (FIGs. 11A-11B). Following a repeated dose of 45mg/kg, the parental antibody nearly completely occupied MerTK on RPEs. in contrast, the antibody conjugates maintained at least 60% free-MerTK+ RPEs, with PEG-40K Branched (PEG-40K-B) maintaining around 90% free-MerTK+ RPEs. On RPEs bearing functional or unoccupied MerTK (free-MerTK+ cells), the levels of unoccupied MerTK were improved with the antibody conjugates as compared to the parental antibody (FIG. 11C). PEG-40K-Branched conjugates were relatively better, with around 90% of RPEs maintaining occupancy of around 57% MerTK receptors. As described above, following single high dose (30 mg/kg) administration, all of the antibody conjugates resulted in at least 90% RPEs maintaining occupancy of at least 70% MerTK receptors.

[0359] Finally, the PEG-conjugated antibodies were evaluated in a repeated low dose study, with administration of 2.5 mg/kg to tumor-bearing mice on Day 1 and Day 5. Both the PEG40K-Linear and PEG40K-Branched-conjugated antibodies demonstrated a lower MerTK occupancy on TAMs as compared to the parental Ab following repeated low⁷ dose administration (FIGs. 12A-12C). Around 31% of TAMs remained with around 35.8% of free MerTK receptors, and around 69% TAMs were completely occupied w⁷ith low dose (2.5 mg/kg x2) of the parental antibody. In contrast, PEG40K- Linear- and PEG-40K-Branched-conjugated antibodies resulted in around 73% of TAMs remaining with around 59% of free MerTK receptors, and in around 27% TAMs with complete occupancy. With the low⁷ repeated dose, the parental and PEG-conjugated antibodies exhibited comparable induction of ISGs in tumors (FIG. 13).

[0368] /Although the present disclosure lias been described in some detail by w'ay of illustration and example for purposes of clarity of understanding, the descriptions and examples should not be construed as limiting the scope of the present disclosure. The disclosures of all patent and scientific literature cited herein are expressly incorporated in their entirety by reference.

Claims

CLAIMS What is claimed is:

1. A PEGylated antibody that binds to MerTK, wherein the antibody is conjugated to one or more polyethylene glycol (PEG) polymer(s); wherein each of the PEG polymer(s) is conjugated to a heavy chain or light chain of the antibody at an engineered cysteine residue; and wherein the antibody comprises:

(1) a heavy chain variable domain (VH) comprising (a) a CDR-H1 comprising the amino acid sequence of SYAMG (SEQ ID NO: 1), (b) a CDR-H2 comprising the amino acid sequence of IINSYGNTYYANWAKG (SEQ ID NO: 2), and (c) a CDR-H3 comprising the amino acid sequence of DPGVSSNL (SEQ ID NO: 3), and a light chain variable domain (VL) comprising (d) a CDR-L1 comprising the amino acid sequence of QASQNIYSGLA (SEQ ID NO: 4), (e) a CDR-L2 comprising the amino acid sequence of GASKLAS (SEQ ID NO: 5), and (f) a CDR-L3 comprising the amino acid sequence of QATYYSSNSVA (SEQ ID NO: 6); or

(2) a heavy chain variable domain (VH) comprising (a) a CDR-H1 comprising the amino acid sequence of ANTMN (SEQ ID NO: 7), (b) a CDR-H2 comprising the amino acid sequence of IFTATGSTYYATWVNG (SEQ ID NO: 8), and (c) a CDR-H3 comprising the amino acid sequence of SGSGSSSGAFNI (SEQ ID NO: 9), and a light chain variable domain (VL) comprising (d) a CDR-Ll comprising the amino acid sequence of QASQSISSSLA (SEQ ID NO: 10), (e) a CDR-L2 comprising the amino acid sequence of AASILAS (SEQ ID NO: i 1), and (I) a CDR-L3 comprising the amino acid sequence of QCTSYGSLFLGP (SEQ ID NO: 12).

2. The antibody of claim I, wherein tlie VH domain comprises a sequence having at least 95% sequence identity to the amino acid sequence of EVQLVESGEGLVQPGGSLRLSCAASGFSLSSYAMGWVRQAPGKGLEY VGI1NSYGNTYYAN WAKGRFTISRDNSKNTVYLQMGSLRAEDN'LAVYYCARDPGVSSNLWGRGTLVTVSS (SEQ ID NO: 13); and/or the VL domain comprises a sequence having at least 95% sequence identity to the amino acid sequence of

DIQMTQSPSTLSASVGDRVTITCQASQNTYSGLAWYQQKPGKAPKLLIYGASKLASGVPSRFS GSGSGTEFTLTISSLQPDDFATYYCQATYYSSNSVAFGGGTKVEIK (SEQ ID NO: 14).

3. The antibody of claim 2, wherein tlie VH domain comprises the amino acid sequence of EVQLVESGEGLVQPGGSLRLSCAASGFSLSSYAMGWVRQAPGKGLEY VGIINSYGNTYYAN WAKGRFTISRDNSKNTVYLQMGSLRAEDMAVYYCARDPGVSSNLWGRGTLVTVSS (SEQ ID NO: 13); and the VL domain comprises the amino acid sequence of

DIQMTQSPSTLSASVGDRVTITCQASQNIYSGLAWYQQKPGKAPKLLIYGASKLASGVPSRFS GSGSGTEFTI.TISSLQPDDFATYYCQATYYSSNSVAFGGG’rKVEIK (SEQ ID NO: 14).

4. The antibody of claim 2 or claim 3, wherein the heavy chain comprises the amino acid sequence of

EVQLVESGEGLVQPGGSLRLSCAASGFSLSSYAMGWVRQAPGKGLEYVGIINSYGNTYYAN WAKGRFTISRDNSKNTVYLQMGSLRAEDMAVYYCARDPGVSSNLWGRGTLVTVSSASTKG PSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSV VTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEAAGGPSVFLFPPKPK DTI,MISRTPEVTCWWVSHEDPEVKFNWYVDGVEVHNAKTKPRF.EQYNSTYRWSVI>T\a, HQDWT.,NGKEYKCKVSNKALGAPIEKTISKAKGQPREPQVYTT.,PPSREEMTKNQVSLTCL\^ZK GFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEAL HNHYTQKSLSLSPG (SEQ ID NO: 21).

5. The antibody of claim 2 or claim 3, wherein the heavy chain comprises the amino acid sequence of

EVQLVESGEGLVQPGGSLRLSCAASGFSLSSYAMGWVRQAPGKGLEYVGTINSYGNTYYAN WAKGRFTISRDNSKNTVYLQMGSLRAEDMAVYYCARDPGVSSNLWGRGTLVTVSSASTKG PSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSV VT\^ZPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFI..FPPKPK DTI,MISRTPEVTCWWVSHEDPEVKFNWYVDGVEVEINAKTKPRF.EQYGSTYRVVSVI>T\a, HQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQWTLPPSREEMTKNQVSLTCLVKG FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALH NHYTQKSLSLSPG (SEQ ID NO:22).

6. The antibody of any one of claims 2-5, wherein the light chain comprises the amino acid sequence of

DIQMTQSPS’n.SASVGDRVTITCQASQNIYSGLAWYQQKPGKAPKLLIYGASKLASGVPSRFS GSGSGTEFTI,TISSLQPDDFATYYCQATYYSSNSVAFGGGrKVEIKRTVAAPSVFIFPPSDEQL KSGTASWCLLNNFYPREzXKVQWKWNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADY EKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 23).

7. The antibody of any one of claims i -6, wherein the antibody is conjugated to one or two PEG polymers.

8. The antibody of any one of claims 1 -7, wherein the PEG poly mer(s) are linear PEG polymers.

9. The antibody of any one of claims 1-7, wherein the PEG polymer(s) are branched PEG polymers.

10. The antibody of claim 9, wherein the branched PEG polymer(s) comprise 2 branches.

11. The antibody of any one of claims 1-10, wherein the PEGylated antibody has a hydrodynamic radius of greater than about 6 nm.

12. The antibody of any one of claims 1-11, wherein the PEGylated antibody has a hydrodynamic radius of greater than or equal to about 10 nm.

13. The antibody of any one of claims 1-12, wherein the PEGylated antibody has a hydrodynamic radius of between about 9 nm and about 11 nm.

14. The antibody of any one of claims 1-13, wherein the PEG polymer(s) each have a molecular weight of between about 10 kDa and about 40 kDa.

15. The antibody of any one of claims 1-14, wherein each of the PEG polymer(s) is conjugated to a heavy chain or light chain of the antibody al an engineered cysteine residue via maleimide-cysteine conjugation.

16. The antibody of any one of claims 1-14, wherein each of the PEG polymer(s) is conjugated to a heavy drain or light chain of the antibody at an engineered cysteine residue via iodoacetamidecysteine conjugation

17. The antibody of any one of claims 1-16, wherein the engineered cysteine residue is selected from the group consisting of K149C of the light chain, K183C of the light drain, T186C of the heavy chain, and Y373C of the heavy drain, numbering of the light chain according to Kabat and numbering of the heavy chain according to EU index.

18. The antibody of claim 17, wherein the antibody comprises two heavy chains, two light chains, and two PEG polymers; and wherein both light chains of tire antibody comprise a K149C engineered cysteine conjugated to one of the two PEG polymers.

19. The antibody of claim 17, wherein the antibody comprises two heavy drains, two tight drains, and two PEG polymers; and wherein both light chains of the antibody comprise a K183C engineered cysteine conjugated to one of the two PEG polymers.

20. The antibody of claim 17, wherein the antibody comprises two heavy drains, two light drains, and two PEG polymers; and wherein both heavy drains of the antibody comprise a T186C engineered cysteine conjugated to one of the two PEG polymers.

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21. The antibody of claim 17, wherein the antibody comprises two heavy drains, two light drains, and two PEG polymers; and wherein both heavy drains of the antibody comprise a Y373C engineered cysteine conjugated to one of the two PEG polymers.

22. A method of producing a PEGylated antibody that binds to MerTK, comprising contacting an antibody comprising one or more free engineered cysteine residues with one or more polyethylene glycol (PEG) polymer(s) comprising a maleimide moiety under conditions suitable for each of the PEG polymer(s) to be conjugated to an engineered cysteine residue of tlie antibody via thioether linkage; wherein the antibody comprises:

(1) a heavy chain variable domain (VH) comprising (a) a CDR-H1 comprising the amino acid sequence of SYAMG (SEQ ID NO: 1), (b) a CDR-H2 comprising the amino acid sequence of IINS YGNTYYANWAKG (SEQ ID NO: 2), and (c) a CDR-H3 comprising the amino acid sequence of DPGVSSNL (SEQ ID NO: 3), and a light chain vanable domain (VL) comprising (d) a CDR-L1 comprising the amino acid sequence of QASQNIYSGLA (SEQ ID NO: 4), (e) a CDR-L2 comprising the amino acid sequence of GASKLAS (SEQ ID NO: 5), and (f) a CDR-L3 comprising the amino acid sequence of QATYYSSNSVA (SEQ ID NO: 6); or

(2) a heavy chain variable domain (VH) comprising (a) a CDR-H1 comprising the amino acid sequence of ANTMN (SEQ ID NO: 7), (b) a CDR-H2 comprising the amino acid sequence of IFTATGSTYYATWVNG (SEQ ID NO: 8), and (c) a CDR-H3 comprising the amino acid sequence of SGSGSSSGAFNI (SEQ ID NO: 9), and a light chain variable domain (VL) comprising (d) a CDR-L1 comprising the amino acid sequence of QASQSISSSLA (SEQ ID NO: 10), (e) a CDR-L2 comprising the amino acid sequence of AASILAS (SEQ ID NO: 1 1), and (I) a CDR-L3 comprising the amino acid sequence of QCTSYGSLFLGP (SEQ ID NO: 12).

23. The method of claim 22, wherein the one or more free engineered cysteine residues of the antibody are contacted with the maleimide moiety of the one or more PEG polymer(s) at a pH between 7.0 and 7.5.

24. A method of producing a PEGylated antibody that binds to MerTK, comprising contacting an antibody comprising one or more free engineered cysteine residues with one or more polyethylene glycol (PEG) poly mens) comprising an iodoacetamide moiety under conditions suitable for each of tiie PEG poly mer(s) to be conjugated to an engineered cysteine residue of the antibody via thioether linkage; wherein the antibody comprises:

-92- (1) a heavy chain variable domain (VH) comprising (a) a CDR-H1 comprising the amino acid sequence of SYAMG (SEQ ID NO: 1), (b) a CDR-H2 comprising the amino acid sequence of IINSYGNTY^YANWAKG (SEQ ID NO: 2), and (c) a CDR-H3 comprising the amino acid sequence of DPGVSSNL (SEQ ID NO: 3 ), and a light chain variable domain (VL) comprising (d) a CDR-LI comprising the amino acid sequence of QASQNIYSGLA (SEQ ID NO: 4), (e) a CDR-L2 comprising the amino acid sequence of GASKLAS (SEQ ID NO: 5), and (f) a CDR-L3 comprising the amino acid sequence of QATYYSSNSVA (SEQ ID NO: 6); or

(2) a heavy chain variable domain (VH) comprising (a) a CDR-H1 comprising the amino acid sequence of ANTMN (SEQ ID NO: 7), (b) a CDR-H2 comprising the amino acid sequence of IFTATGSTYYATWVNG (SEQ ID NO: 8), and (c) a CDR-H3 comprising the amino acid sequence of SGSGSSSGAFNI (SEQ ID NO: 9), and a light chain variable domain (VL) comprising (d) a CDR- L1 comprising the amino acid sequence of QASQSISSSLA (SEQ ID NO: 10), (e) a CDR-L2 comprising the amino acid sequence of AASILAS (SEQ ID NO: 11), and (I) a CDR-L3 comprising the amino acid sequence of QCTS YGSLFLGP (SEQ ID NO: 12).

25. The method of any one of claims 22-24, wherein the PEG polymer(s) are conjugated to the antibody at a ratio of 2.0 polymers per antibody.

26. The method of any one of claims 22-25, wherein, prior to contacting the antibody with the PEG polymer(s), each of the free engineered cysteine residues is blocked with a cysteine or glutathione moiety; and the method further comprises deblocking the engineered cysteine residue.

2.7. The method of any one of clams 22-26, further comprising, after contacting the antibody with die PEG polymer(s), purifying the PEGy lated antibody from unconjugated antibody and PEG polymer.

28. The method of claim 27, wherein the PEGylated antibody is purified by hydrophobic interaction chromatography .

29. The method of any one of clams 22-28, wherein the VH domain comprises a sequence having at least 95% sequence identity to the amino acid sequence of

EVQLVESGEGLVQPGGSLRLSCAASGFSLSSYAMGWVRQAPGKGLEYVG1INSYGNTYYAN

WAKGRFTISRDNSKNTVYLQMGSLRAEDMAVYYCARDPGVSSNLWGRGTLVTVSS (SEQ ID NO: 13); and/or the VL domain comprises a sequence having at least 95% sequence identity to the amino acid sequence of

DIQMTQSPSTLSASVGDRVTITCQASQNIYSGLAWYQQKPGKAPKLLIYGASKLASGVTSRFS GSGSGTEFTLTISSLQPDDFATYYCQATYYSSNSVAFGGGTKVEIK (SEQ ID NO: 14).

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30. The method of claim 29, wherein the VH domain comprises the amino acid sequence of

EVQLVESGEGLVQPGGSLRLSCAASGFSLSSYAMGWVRQAPGK.GLEYVGIINSYGNTYYAN WAKGRFTTSRDNSKNTVYLQMGSLRAEDMAVYYCARDPGVSSNLWGRGTLVTVSS (SEQ ID NO: 13); and the VL domain comprises the amino acid sequence of

DIQMTQSPS’n.SASVGDRVTITCQASQNIYSGLAWYQQKPGKAPKLLIYGASKLASGVPSRFS GSGSGTEFTI.TISSLQPDDFATYYCQATYYSSNSVAFGGG’rKVEIK (SEQ ID NO: 14).

31. The method of claim 29 or claim 30, wherein the heavy chain comprises the amino acid sequence of

EVQLVESGEGLVQPGGSLRLSCAASGFSLSSYAMGWVRQAPGKGLEYVGIINSYGNTYYAN WAKGRFTISRDNSKNTVYLQMGSLRAEDMAVYYCARDPGVSSNLWGRGTLVTVSSASTKG PSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSV VTXTSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPEAAGGPSVFLFPPKPK

DIEMISRTPEVTCVVVDVSHEDPEVIH'NWYVDGVEVI-INAKTKPREEQYNSTYRVVSVLTVL HQDWLNGKEYKCKVSNKALGAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVK GFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEAL HNHYTQKSLSLSPG (SEQ ID NO: 21).

32. The method of claim 29 or claim 30, wherein the heavy drain comprises the amino acid sequence of

EVQIATiSGEGLVQPGGSLRLSCAASGFSLSSYAMGWVRQAPGKGLEYVGIINSYGNTYYAN WAKGRimSRDNSKNTVYLQMGSLRAEDMAVYYCARDPGVSSNLWGRGTLVTVSSASTKG PSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSV VTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPK

DTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYGSTYRWSVLTVL HQDWLNGKEYKCKVSNKALPAPffiKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKG FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALH NHYTQKSLSLSPG (SEQ ID NO:22).

33. The method of any one of claims 29-32, wherein the light chain comprises the amino acid sequence of

DIQMTQSPSTLSASVGDRVTITCQASQNIYSGLAWYQQKPGKAPKLLIYGASKLASGVPSRFS GSGSGTEFTLTISSLQPDDFATYYCQATYYSSNSVAFGGGTKVEIKRTVAAPSVFIFPPSDEQL KSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADY EKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 23).

34. The method of any one of clams 22-33, wherein the PEG polymer(s) are linear PEG polymers.

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35. The method of any one of clams 22-33, wherein the PEG polymer(s) are branched PEG polymers.

36. The method of claim 35, wherein the branched PEG polymer(s) comprise 2 branches.

37. The method of any one of dams 22-36, wherein the PEGylated antibody has a hydrodynamic radius of greater than about 6 mu.

38. The method of any one of dams 22-37, wherein the PEGylated antibody has a hydrodynamic radius of greater than or equal to about 10 mn.

39. The method of any one of dams 22-38, wherein the PEGylated antibody has a hydrodynamic radius of between about 9 mu and about 11 nm.

40. The method of any one of clams 22-39, wherein the PEG polymer(s) each have a molecular weight of between about 10 kDa and about 40 kDa.

41. The method of any one of clams 22-40, wherein the one or more free engineered cysteine residues are selected from the group consisting of K149C of the light chain, K183C of the light chain, T186C of the heavy chain, and Y373C of the heavy chain, numbering of the light chain according to

Rabat and numbering of the heavy chain according to EU index.

42. The method of claim 41 , wherein the antibody comprises two heavy chains, two light chains, and two PEG polymers; and wherein both light chains of the antibody comprise a K149C engineered cysteine conjugated to one of the two PEG polymers.

43. The method of claim 41, wherein the antibody comprises two heavy chains, two light chains, and two PEG polymers; and wherein both light chains of the antibody comprise a K183C engineered cysteine conjugated to one of the two PEG polymers.

44. The method of claim 41, wherein the antibody comprises two heavy chains, two light chains, and two PEG polymers; and wherein both heavy chains of the antibody comprise a T186C engineered cysteine conjugated to one of the two PEG polymers.

45. The method of claim 41, wherein the antibody comprises two heavy chains, two light chains, and two PEG polymers; and wherein both heavy chains of the antibody comprise a Y373C engineered cysteine conjugated to one of the two PEG polymers.

46. A PEGylated antibody that binds to MerTK produced by the method of any one of claims 22-

45.

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47. A pharmaceutical composition comprising the PEGylated antibody of any one of claims 1-21 and 46 and a pharmaceutically acceptable carrier.

48. The pharmaceutical composition of claim 47, further comprising an additional therapeutic agent.

49. The pharmaceutical composition of claim 48, wherein the additional therapeutic agent is selected from one or more of tamoxifen, letrozole, exemestane, anastrozole, irinotecan, cetuximab, fulvestrant, vinorelbine, erlotinib, bevacizumab, vincristine, imatinib mesylate, sorafenib, lapatinib, trastuzumab, cisplatin, gemcitabine, methotrexate, vinblastine, carboplatin, paclitaxel, 5-fluorouracil, doxorubicin, bortezomib, melphalan, prednisone, and docetaxel.

50. The pharmaceutical composition of claim 48, wherein the additional therapeutic agent is an immune checkpoint inhibitor.

51. The PEGylated antibody of any one of claims 1-21 and 46 or the pharmaceutical composition of any of claims 47-50 for use as a medicament.

52. The PEGy lated antibody of any one of claims 1-21 and 46 or the pharmaceutical composition of any of claims 47-50 for use i n treating cancer.

53. Use of the PEGylated antibody of any one of claims 1-21 and 46 or the pharmaceutical composition of any of claims 47-50 in the manufacture of a medicament for treating cancer.

54. Use of the PEGylated antibody of any one of claims 1-21 and 46 or the pharmaceutical composition of any of claims 47-50 in the manufacture of a medicament for reducing MerTK- mediated clearance of apoptotic cells in an individual.

55. A method of treating an individual having cancer comprising administering to the individual an effective amount of the PEGylated antibody of any one of claims 1-21 and 46 or the pharmaceutical composition of any of claims 47-50.

56. The method of claim 55, wherein administration of the PEGylated antibody causes reduced retinal toxicity' in the individual, as compared to administration of an antibody that binds to MerTK that is not PEGylated.

57. The method of claim 55 or claim 56, further comprising administering an additional therapeutic agent to the individual.

58. The method of claim 57, wherein the additional therapeutic agent is selected from one or more of tamoxifen, letrozole, exemestane, anastrozole, irinotecan, cetuximab, fulvestrant, vinorelbine,

-96- erlotinib, bevacizumab, vincristine, imatinib mesylate, sorafenib, lapatinib, trastuzumab, cisplatin, gemcitabine, methotrexate, vinblastine, carboplatin, paclitaxel, 5-fluorouracil, doxorubicin, bortezomib, melphalan, prednisone, and docetaxel.

59. The method of claim 57, wherein the additional therapeutic agent is an immune checkpoint inhibitor.

60. The method of claim 59, wherein the immune checkpoint inhibitor is selected from the group consisting of a cytotoxic T-lymphocyte-associated protein 4 (CTLA4) inhibitor, a programmed cell death protein 1 (PD-1) binding antagonist, and a programmed death-ligand 1 (PDL1) binding antagonist.

61. The method of claim 59, wherein the immune checkpoint inhibitor is a PDL1 binding antagonist.

62. The method of claim 61, wherein the PDL1 binding antagonist is an anti-PDLl antibody.

63. The method of claim 62, wherein the anti-PDL 1 antibody is atezolizumab.

64. The method of any one of claims 59-63, further comprising administering an effective amount of an additional chemotherapeutic agent to the individual.

65. The method of any one of claims 55-64, wherein the cancer is colon cancer.

66. The method of any one of claims 55-64, wherein the cancer is selected from the group consisting of coion cancer, urothelial carcinoma, non-small cell lung cancer, triple negative breast cancer, small cell lung cancer, hepatocellular carcinoma, and melanoma.

67. A method of reducing MerTK-mediated clearance of apoptotic cells in an individual comprising administering to the individual an effective amount of the PEGylated antibody of any one of claims 1-21 and 46 or the pharmaceutical composition of any of claims 47-50 to reduce MerTK- mediated clearance of apoptotic cells.

68. The method of claim 67, wherein administration of the PEGylated antibody causes reduced retinal toxicity in the individual, as compared to administration of an antibody that binds to MerTK that is not PEGylated.

69. The method of claim 67 or claim 68, wherein the clearance of apoptotic cells is reduced by about 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, 4.0, 4.1, 4.2, 4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, or 8,0 fold.