High affinity anti-MERTK antibodies and uses thereof

Abstract

The present disclosure provides antibodies (e.g., humanized antibodies) that specifically bind to Mer tyrosine kinase (MERTK; e.g., human MERTK), and compositions comprising such antibodies. The present disclosure also provides antibody-drug conjugates comprising (i) an anti-MERTK antibody or antigen-binding fragment thereof described herein that specifically binds to MERTK (e.g., human MERTK), and (ii) a cytotoxic agent conjugated directly to the antibody or via a linker, as well as compositions comprising such antibody-drug conjugates.

Description

[Technical Field]

[0001] 1. Field This application claims the benefit of U.S. Provisional Application No. 62 / 810,841, filed on 26 February 2019, which is incorporated herein by reference in its entirety.

[0002] This application incorporates, by reference, a sequence listing submitted with this application as an ASCII-formatted text file named "13256-008-228_SEQ_LISTING.txt", which was created on 19 February 2020 and has a size of 94,274 bytes.

[0003] This disclosure provides antibodies (e.g., humanized antibodies) that specifically bind to Mer tyrosine kinase (MERTK; e.g., human MERTK), and compositions comprising such antibodies. This disclosure also provides antibody-drug conjugates comprising (i) an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK), and (ii) a cytotoxic agent that is directly conjugated to the antibody or conjugated to the antibody via a linker, and compositions comprising such antibody-drug conjugates. This disclosure also provides a method for treating cancer, comprising the step of administering to a human subject in need of such treatment (a) an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK), or (b) an antibody-drug conjugate comprising (i) an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK), and (ii) a cytotoxic agent that is directly conjugated to the antibody or conjugated to the antibody via a linker. [Background technology]

[0004] 2.Background Mer tyrosine kinase (MERTK), also known as c-mer, MER, the proto-oncogene c-Mer, receptor tyrosine kinase MerTK, tyrosine-protein kinase Mer, STK kinase, RP38, or MGC133349, is a member of the TAM family of receptor tyrosine kinases, which also includes AXL and TYRO3 kinases. MERTK converts signals from the extracellular space through activation by ligand binding, most notably the soluble protein Gas-6. When Gas-6 binds to MERTK, autophosphorylation of MERTK is induced in its intracellular domain, leading to downstream signal activation (Cummings CT et al., (2013) Clin Cancer Res 19: 5275-5280; Verma A et al., (2011) Mol Cancer Ther 10: 1763-1773).

[0005] MERTK exists in both membrane-bound and soluble forms. It is hypothesized that the extracellular domain can be cleaved to produce a soluble extracellular domain, which acts as a decoy receptor, negatively regulating the activation of the MERTK receptor on cells by reducing the ability and / or availability of soluble Gas-6 ligands to bind to membrane-bound MERTK (Sather S et al., (2007) Blood 109: 1026-1033). As a result, MERTK has two roles in cancer progression, angiogenesis, and metastasis. On the other hand, Gas-6 activation of MERTK in endothelial cells leads to inhibition of endothelial cell recruitment by cancer cells in co-culture systems. Endothelial recruitment is a key feature of cancer cells that enables tumor angiogenesis, tumor growth, and metastasis. Downregulation of MERTK in metastatic breast cancer cells has also been shown to inhibit endothelial recruitment (Png KJ et al., (2012) Nature 481: 190-194). However, on the other hand, MERTK plays the opposite role in cancer cells, and its overexpression leads to increased metastasis, possibly by releasing cleaved MERTK to generate soluble MERTK extracellular domain protein as a decoy receptor. Furthermore, ligand-dependent MERTK activation in cancer cells stimulates oncogenic pathways, including the AKT pathway (Linger RM, Cohen RA, Cummings CT, Sather S, Migdall-Wilson J, Middleton DH, Lu X, Baron AE, Franklin WA, Merrick DT, Jedlicka P, DeRyckere D, Heasley LE, Graham DK. Mer or Axl receptor tyrosine kinase inhibition promotes apoptosis, blocks growth and enhances chemosensitivity of human non-small cell lung cancer (Oncogene. 2013 Jul 18;32(29):3420-31)). Therefore, tumor cells overexpress MERTK to promote oncogenic signaling.Furthermore, tumor cells secrete soluble extracellular MERTK receptors that act as decoy receptors, reducing the ability (and / or availability) of soluble Gas-6 ligands to activate MERTK on endothelial cells, ultimately leading to endothelial mobilization, angiogenesis, and cancer progression (Png KJ et al., (2012) Nature 481: 190-194).

[0006] MERTK is also involved in the modulation of macrophage homeostasis and the maintenance of anti-inflammatory states. In macrophages, MERTK mediates phagocytic clearance of apoptotic cells and the release of IL-10 and acute inflammatory cytokines, and its expression is stimulated by Gas-6-dependent binding to MERTK (Zizzo et al., (2012) J. Immunology Oct 1;189(7):3508-20; Cook et al., J Clin Invest. 2013;123(8):3231-3242).

[0007] Historically, MERTK was thought to function only as an oncogene, so efforts have been made to produce MERTK inhibitors for cancer treatment (e.g., compound UNC1062, a potent small molecule MERTK inhibitor developed as an anticancer compound) (Liu J et al., (2013) Eur J Med Chem 65: 83-93; Cummings CT et al., (2013) Clin Cancer Res 19: 5275-5280; Verma A et al., (2011) Mol Cancer Ther 10: 1763-1773). In recent years, antibody-drug conjugates (ADCs) have become one of the fastest-growing classes of cancer treatments (Beck A et al., (2017) Nat Rev Drug Discov 16: 315-337; Peters C and Brown S, (2015) Biosci Rep 35: art:e00225). The references made herein should not be construed as an acknowledgment that they constitute prior art to this disclosure. [Primary Technology Documents]

Non-licensed literature

[0008] [Non-licensed document 1] Cummings CT et al., (2013) Clin Cancer Res 19: 5275-5280 [Non-licensed document 2] Verma A et al., (2011) Mol Cancer Ther 10: 1763-1773 [Non-licensed document 3] Sather S et al., (2007) Blood 109: 1026-1033

Non-licensed Document 4

Non-licensed Document 5

Non-licensed Document 6

Non-licensed Document 7

[0009] 3. Overview This specification discloses antibodies that bind specifically to MERTK (e.g., human MERTK) with high affinity and antigen-binding fragments thereof. In any specific embodiment of the antibodies or antigen-binding fragments described herein, the antibody specifically recognizes the extracellular domain of MERTK (e.g., human MERTK), and the extracellular domain comprises the amino acid sequence of SEQ ID NO: 132.

[0010] In specific embodiments, the anti-MERTK antibody or its antigen-binding fragment specifically binds to MERTK (e.g., human MERTK), resulting in the internal translocation of MERTK from the cell surface. In specific embodiments, the anti-MERTK antibody or its antigen-binding fragment blocks Gas-6-induced phosphorylation of AKT. In another specific embodiment, the anti-MERTK antibody or its antigen-binding fragment prevents MERTK activation. In yet another specific embodiment, the anti-MERTK antibody or its antigen-binding fragment described herein specifically recognizes the extracellular portion of human MERTK. In yet another specific embodiment, the anti-MERTK antibody or its antigen-binding fragment reduces colonization by cancer cells. In certain embodiments, the anti-MERTK antibody or its antigen-binding fragment described herein is monoclonal. In yet another specific embodiment, the anti-MERTK antibody or its antigen-binding fragment described herein is an immunoglobulin containing two identical light chains and two identical heavy chains. In certain embodiments, the anti-MERTK antibody is a humanized antibody.

[0011] In certain embodiments, the anti-MERTK antibody or antigen-binding fragment described herein that specifically binds to MERTK (e.g., human MERTK) includes a heavy chain variable region (VH) comprising the amino acid sequence of SEQ ID NO: 105.

[0012] In another specific embodiment, the anti-MERTK antibody or antigen-binding fragment described herein that specifically binds to MERTK (e.g., human MERTK) includes a heavy chain variable region (VH) comprising the amino acid sequence of SEQ ID NO: 107.

[0013] In another specific embodiment, the anti-MERTK antibody or antigen-binding fragment described herein that specifically binds to MERTK (e.g., human MERTK) includes a heavy chain variable region (VH) comprising the amino acid sequence of SEQ ID NO: 108.

[0014] In another specific embodiment, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) includes a heavy chain variable region (VH) containing the amino acid sequence of SEQ ID NO: 109.

[0015] In another specific embodiment, the anti-MERTK antibody or antigen-binding fragment described herein that specifically binds to MERTK (e.g., human MERTK) includes a heavy chain variable region (VH) comprising the amino acid sequence of SEQ ID NO: 110.

[0016] In another embodiment, the anti-MERTK antibody or its antigen-binding fragment described herein, which specifically binds to MERTK (e.g., human MERTK), includes a light chain variable region (VL) containing the amino acid sequence of SEQ ID NO: 106. In another specific embodiment, the anti-MERTK antibody or its antigen-binding fragment described herein, which specifically binds to MERTK (e.g., human MERTK), includes a light chain variable region (VL) containing the amino acid sequence of SEQ ID NO: 111.

[0017] In another embodiment, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) comprises (a) a VH containing the amino acid sequence of SEQ ID NO: 105, and (b) a VL containing the amino acid sequence of SEQ ID NO: 106. In another embodiment, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) comprises (a) a VH containing the amino acid sequence of SEQ ID NO: 107, and (b) a VL containing the amino acid sequence of SEQ ID NO: 106. In another embodiment, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) comprises (a) a VH containing the amino acid sequence of SEQ ID NO: 108, and (b) a VL containing the amino acid sequence of SEQ ID NO: 106. In another embodiment, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) comprises (a) a VH containing the amino acid sequence of SEQ ID NO: 109, and (b) a VL containing the amino acid sequence of SEQ ID NO: 106. In another embodiment, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) comprises (a) VH containing the amino acid sequence of SEQ ID NO: 110, and (b) VL containing the amino acid sequence of SEQ ID NO: 111.

[0018] In certain embodiments, the antibodies provided herein that specifically bind to MERTK (e.g., human MERTK) include a heavy chain constant region, a light chain constant region, or both. In some embodiments, the heavy chain constant region is selected from the group of human immunoglobulins consisting of IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2. In certain embodiments, the light chain constant region is selected from the group of human immunoglobulins consisting of IgGκ and IgGλ. In some embodiments, the antibody includes a constant region in which binding affinity to one or more human Fc gamma receptors is increased.

[0019] In some embodiments, the antibody includes a constant region in which the binding affinity to one or more human Fc gamma receptors is reduced.

[0020] In a specific embodiment of the embodiment in which the antibody is an immunoglobulin comprising two identical light chains and two identical heavy chains, each heavy chain comprises a variable region (VH) comprising the amino acid sequence of SEQ ID NO: 105. In a further embodiment of such a specific embodiment, each light chain comprises a variable region (VL) comprising the amino acids of SEQ ID NO: 106.

[0021] In a specific embodiment of the embodiment in which the antibody is an immunoglobulin comprising two identical light chains and two identical heavy chains, each heavy chain comprises a variable region (VH) comprising the amino acids of SEQ ID NO: 107. In a further embodiment of such a specific embodiment, each light chain comprises a variable region (VL) comprising the amino acid sequence of SEQ ID NO: 106.

[0022] In a specific embodiment of the embodiment in which the antibody is an immunoglobulin comprising two identical light chains and two identical heavy chains, each heavy chain comprises a variable region (VH) comprising the amino acid sequence of SEQ ID NO: 108. In a further embodiment of such a specific embodiment, each light chain comprises a variable region (VL) comprising the amino acid sequence of SEQ ID NO: 106.

[0023] In a specific embodiment of the embodiment in which the antibody is an immunoglobulin comprising two identical light chains and two identical heavy chains, each heavy chain comprises a variable region (VH) comprising the amino acid sequence of SEQ ID NO: 110. In a further embodiment of such a specific embodiment, each light chain comprises a variable region (VL) comprising the amino acid sequence of SEQ ID NO: 111.

[0024] In a specific embodiment of the embodiment in which the antibody is an immunoglobulin comprising two identical light chains and two identical heavy chains, each heavy chain comprises a variable region (VH) comprising the amino acid sequence of SEQ ID NO: 109. In a further embodiment of such a specific embodiment, each light chain comprises a variable region (VL) comprising the amino acid sequence of SEQ ID NO: 106.

[0025] In a specific embodiment of the embodiment in which the antibody is an immunoglobulin comprising two identical light chains and two identical heavy chains, the light chains include a variable region (VL) comprising SEQ ID NO: 106.

[0026] In a specific embodiment of the embodiment in which the antibody is an immunoglobulin comprising two identical light chains and two identical heavy chains, the light chains include a variable region (VL) comprising SEQ ID NO: 111.

[0027] In a specific embodiment of the embodiment in which the antibody is an immunoglobulin comprising two identical light chains and two identical heavy chains, the heavy chains include a variable region (VH) comprising SEQ ID NO: 105.

[0028] In a specific embodiment of the embodiment in which the antibody is an immunoglobulin comprising two identical light chains and two identical heavy chains, the heavy chains include a variable region (VH) comprising SEQ ID NO: 107.

[0029] In a specific embodiment of the embodiment in which the antibody is an immunoglobulin comprising two identical light chains and two identical heavy chains, the heavy chains include a variable region (VH) comprising SEQ ID NO: 108.

[0030] In a specific embodiment of the embodiment in which the antibody is an immunoglobulin comprising two identical light chains and two identical heavy chains, the heavy chains include a variable region (VH) comprising SEQ ID NO: 109.

[0031] In a specific embodiment of the embodiment in which the antibody is an immunoglobulin comprising two identical light chains and two identical heavy chains, the heavy chains include a variable region (VH) comprising SEQ ID NO: 110.

[0032] In any specific embodiment of an antibody or antigen-binding fragment provided herein, where the antibody is an immunoglobulin comprising two identical light chains and two identical heavy chains, the antibody or antigen-binding fragment includes a human-derived constant region. In a further embodiment of such a specific embodiment, the heavy chain constant region has an isotype selected from the group consisting of gamma 1, gamma 2, gamma 3, and gamma 4.

[0033] In any specific embodiment of the antibody or its antigen-binding fragment provided herein, the antibody binds to MERTK on a cell (e.g., human MERTK) with an EC50 in the range of about 1 nM to 15 nM (e.g., 6.5 nM to 8 nM).

[0034] In any specific embodiment of the antibody or its antigen-binding fragment provided herein, the antibody or antigen-binding fragment reduces the expression level of MERTK in cancer cells (e.g., melanoma cell line SKMEL5). In another specific embodiment of the antibody or its antigen-binding fragment provided herein, the antibody or antigen-binding fragment reduces the expression level of MERTK in human M2 macrophages. In another specific embodiment of the antibody or its antigen-binding fragment provided herein, the antibody or antigen-binding fragment reduces the level of MERTK in human M2 macrophages differentiated in vitro.

[0035] In any specific embodiment of an antibody or its antigen-binding fragment provided herein, the antibody or its antigen-binding fragment comprises a human-derived constant region. In a further embodiment of such a specific embodiment, the antibody or its antigen-binding fragment is a humanized immunoglobulin.

[0036] In any specific embodiment of an antibody or its antigen-binding fragment provided herein, where the antibody is neither a monoclonal antibody nor an immunoglobulin containing two identical light chains and two identical heavy chains, the antibody or its antigen-binding fragment is a bispecific antibody.

[0037] In another embodiment, the Specified Information provides an immunoglobulin that specifically binds to MERTK (e.g., human MERTK), comprising (i) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 105, and (ii) a light chain variable region comprising the amino acid sequence of SEQ ID NO: 106.

[0038] In another embodiment, the Specified Information provides an immunoglobulin that specifically binds to MERTK (e.g., human MERTK), comprising (i) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 107, and (ii) a light chain variable region comprising the amino acid sequence of SEQ ID NO: 106.

[0039] In another embodiment, the Specified Information provides an immunoglobulin that specifically binds to MERTK (e.g., human MERTK), comprising (i) a heavy chain variable region containing the amino acid sequence of SEQ ID NO: 108, and (ii) a light chain variable region containing the amino acids of SEQ ID NO: 106.

[0040] In another embodiment, the Specified Information provides an immunoglobulin that specifically binds to MERTK (e.g., human MERTK), comprising (i) a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 109, and (ii) a light chain variable region comprising the amino acid sequence of SEQ ID NO: 106.

[0041] In another embodiment, a humanized immunoglobulin that specifically binds to MERTK (e.g., human MERTK) is described herein. (A)(i) a heavy chain variable region containing the amino acid sequence of SEQ ID NO: 105, and (ii) a light chain variable region containing the amino acid sequence of SEQ ID NO: 106; or (B)(i) a heavy chain variable region containing the amino acid sequence of SEQ ID NO: 107, and (ii) a light chain variable region containing the amino acid sequence of SEQ ID NO: 106; or (C)(i) a heavy chain variable region containing the amino acid sequence of SEQ ID NO: 108, and (ii) a light chain variable region containing the amino acid sequence of SEQ ID NO: 106; or (D)(i) Heavy chain variable region containing the amino acid sequence of SEQ ID NO: 109, and (ii) Light chain variable region containing the amino acid sequence of SEQ ID NO: 106 This provides humanized immunoglobulins, including those mentioned above.

[0042] In some embodiments, the antibody provided herein is IgG. In some embodiments, the antigen-binding fragment provided herein is a Fab, F(ab')2, or scFv fragment.

[0043] In another embodiment, this specification provides a chimeric antibody or an antigen-binding fragment thereof that binds to MERTK (e.g., human MERTK). In some embodiments, the chimeric antibody includes a heavy chain variable region (VH) containing the amino acids of SEQ ID NO: 110. In some embodiments, the chimeric antibody includes a variable light chain region (VL) containing the amino acids of SEQ ID NO: 111. In a specific embodiment, the chimeric antibody includes the VH of SEQ ID NO: 110 and the VL of SEQ ID NO: 111.

[0044] In another specific embodiment, the antibody comprises a heavy chain containing the amino acid sequence of SEQ ID NO: 112, 114, 115, 116, or 117. In another specific embodiment, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) comprises a light chain containing the amino acids of SEQ ID NO: 113 or 118. In yet another specific embodiment, the anti-MERTK antibody is (A) A heavy chain containing the amino acids of SEQ ID NO: 112, and a light chain containing the amino acid sequence of SEQ ID NO: 113; or (B) A heavy chain containing the amino acids of SEQ ID NO: 114, and a light chain containing the amino acid sequence of SEQ ID NO: 113; or (C) A heavy chain containing the amino acids of SEQ ID NO: 115, and a light chain containing the amino acid sequence of SEQ ID NO: 113; or (D) A heavy chain containing the amino acids of SEQ ID NO: 116, and a light chain containing the amino acid sequence of SEQ ID NO: 113; or (E) Heavy chain containing the amino acids of SEQ ID NO: 117, and light chain containing the amino acid sequence of SEQ ID NO: 113 Includes.

[0045] In a particular embodiment, the anti-MERTK antibody is a bispecific antibody, comprising two distinct antigen-binding regions, one of which specifically binds to MERTK (e.g., human MERTK) and the other binding region which binds to the antigen of interest (e.g., an immune cell receptor, a checkpoint receptor, or a tumor-associated antigen).

[0046] In specific embodiments, the binding region that binds to MERTK includes the variable region of the z10, z11, or z13 antibody described herein (for example, those shown in Tables 11, 12, and 13 below). In some specific embodiments, the other binding region of the bispecific antibody binds to CD3. In other specific embodiments, the other binding region of the bispecific antibody binds to PD-L1. In other specific embodiments, the other binding region of the bispecific antibody binds to LRP1. In other specific embodiments, the other binding region of the bispecific antibody binds to LPR8. In other specific embodiments, the other binding region of the bispecific antibody binds to TGF-β. In other specific embodiments, the other binding region of the bispecific antibody binds to ICOS. In other specific embodiments, the other binding region of the bispecific antibody binds to CD40. In other specific embodiments, the other binding region of the bispecific antibody binds to NKGD2. In other specific embodiments, the other binding region of the bispecific antibody binds to TIGIT. In certain embodiments, the binding region that binds to CD40 is an antagonist antibody. In another specific embodiment, the binding region that binds to ICOS is an antagonist antibody. In yet another specific embodiment, the binding region that binds to PD-L1 is a blocking antibody.

[0047] In another embodiment, the Specified Public Service provides, as the anti-MERTK antibody described herein, an anti-MERTK antibody or its antigen-binding fragment (e.g., a humanized or human antibody, or its antigen-binding fragment) that binds to the same epitope of MERTK (e.g., human MERTK). In another embodiment, the Specified Public Service provides an anti-MERTK antibody or its antigen-binding fragment (e.g., a humanized or human antibody, or its antigen-binding fragment) that competes with the anti-MERTK antibody or its antigen-binding fragment described herein for binding to MERTK (e.g., human MERTK). In another specific embodiment, this specification provides a first anti-MERTK antibody or antigen-binding fragment that competes with the anti-MERTK antibody or antigen-binding fragment described herein for binding to MERTK (e.g., human MERTK), wherein the competition is expressed as a reduction in the binding of the first anti-MERTK antibody or antigen-binding fragment to MERTK (e.g., human MERTK) by more than 80% (e.g., 85%, 90%, 95%, or 98%, or between 80% and 85%, 80% and 90%, 85% and 90%, or between 85% and 95%) in the presence of the anti-MERTK antibody or antigen-binding fragment described herein. In a specific embodiment, the anti-MERTK antibody or antigen-binding fragment that binds to the same epitope as MERTK (or competes with the binding of the anti-MERTK antibody or antigen-binding fragment thereof) is neither a mouse antibody nor its antigen-binding fragment.

[0048] In certain embodiments, the anti-MERTK antibody provided herein has an EC of approximately 1 nM to 10 nM in assays such as those described herein. 50The antibody specifically binds to MERTK (e.g., human MERTK). In some embodiments, the anti-MERTK antibody or its antigen-binding fragment specifically binds to human MERTK. In specific embodiments, the antibody or its antigen-binding fragment provided herein does not bind to or binds to recognizable human Axl, human Tyro3, ​​or mouse MERTK, as evaluated by assays described herein or known to those skilled in the art. In some embodiments, the anti-MERTK antibody or its antigen-binding fragment binds to human MERTK and cynomolgus monkey MERTK. In specific embodiments, the antibody provided herein is isolated. In specific embodiments, the antibody provided herein is a monoclonal antibody. In specific embodiments, the anti-MERTK antibody or its antigen-binding fragment provided herein is bivalent and can bind to two MERTK molecules (e.g., human MERTK molecules).

[0049] In another embodiment, this specification provides polynucleotide sequences encoding an antibody or an antigen-binding fragment thereof. In a specific embodiment, this specification provides a polynucleotide sequence comprising a nucleic acid molecule encoding a heavy chain variable region, wherein the nucleic acid molecule comprises the nucleic acid sequence of SEQ ID NO: 119, SEQ ID NO: 121, SEQ ID NO: 122, or SEQ ID NO: 123. In another specific embodiment, this specification provides a polynucleotide sequence comprising a nucleic acid molecule encoding a light chain variable region, wherein the nucleic acid molecule comprises the nucleic acid sequence of SEQ ID NO: 120, or SEQ ID NO: 124. In a specific embodiment, the nucleic acid molecule or polynucleotide sequence is isolated.

[0050] In another embodiment, this specification provides a vector comprising a polynucleotide sequence encoding an antibody or antigen-binding fragment described herein. In certain embodiments, the vector (e.g., an isolated vector) comprises a heavy chain variable region and / or a light chain variable region of an anti-MERTK antibody described herein, or a polynucleotide encoding a heavy chain and / or a light chain. In certain embodiments, the host cell comprises a polynucleotide or a vector. Examples of host cells include E. coli, Pseudomonas, Bacillus, Streptomyces, yeast, 293F, CHO, YB / 20, NS0, PER-C6, HEK-293, HEK-293T, NIH-3T3, HeLa, BHK, Hep G2, SP2 / 0, R1.1, BW, LM, COS1, COS7, BSC1, BSC40, BMT10 cells, plant cells, insect cells, and human cells in tissue culture. In specific embodiments, this specification provides a method for producing an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), the method comprising the step of culturing host cells such that a polynucleotide is expressed and an antibody is produced.

[0051] In another embodiment, this specification provides ex vivo cells containing one or more polynucleotides encoding any of the antibodies or antigen-binding fragments thereof described herein, or any of the immunoglobulins described herein. In a specific embodiment, the ex vivo cells are: (a) A first immunoglobulin comprising (i) a heavy chain variable region containing the amino acid sequence of SEQ ID NO: 105, and (ii) a light chain variable region containing the amino acid sequence of SEQ ID NO: 106; (b) A second immunoglobulin comprising (i) a heavy chain variable region containing the amino acid sequence of SEQ ID NO: 107, and (ii) a light chain variable region containing the amino acid sequence of SEQ ID NO: 106; and (c) A third immunoglobulin comprising (i) a heavy chain variable region containing the amino acid sequence of SEQ ID NO: 108, and (ii) a light chain variable region containing the amino acid sequence of SEQ ID NO: 106; (d) A fourth immunoglobulin comprising (i) a heavy chain region containing the amino acid sequence of SEQ ID NO: 109, and (ii) a light chain variable region containing the amino acid sequence of SEQ ID NO: 106; and (e) A fifth immunoglobulin comprising (i) a heavy chain variable region containing the amino acid sequence of SEQ ID NO: 110, and (ii) a light chain variable region containing the amino acid sequence of SEQ ID NO: 111. It contains one or more polynucleotides encoding an antibody selected from the group consisting of the following.

[0052] In another embodiment, this specification provides ex vivo cells containing a polynucleotide encoding one of the heavy chains of the antibodies described herein.

[0053] In another embodiment, this specification provides ex vivo cells containing polynucleotides encoding any of the light chains of antibodies described herein. In another embodiment, this specification provides a method for producing an antibody or an antigen-binding fragment thereof, comprising the step of culturing ex vivo cells containing one or more polynucleotides encoding any of the antibodies or antigen-binding fragments described herein under conditions such that one or more polynucleotides are expressed by the cells to produce an antibody or an antigen-binding fragment encoded by the polynucleotides. In a specific embodiment, the method for producing an antibody or an antigen-binding fragment thereof is: (a) A first immunoglobulin comprising (i) a heavy chain variable region containing the amino acid sequence of SEQ ID NO: 105, and (ii) a light chain variable region containing the amino acid sequence of SEQ ID NO: 106; (b) A second immunoglobulin comprising (i) a heavy chain variable region containing the amino acid sequence of SEQ ID NO: 107, and (ii) a light chain variable region containing the amino acid sequence of SEQ ID NO: 106; (c) A third immunoglobulin comprising (i) a heavy chain variable region containing the amino acid sequence of SEQ ID NO: 108, and (ii) a light chain variable region containing the amino acid sequence of SEQ ID NO: 106; (d) A fourth immunoglobulin comprising (i) a heavy chain region containing the amino acid sequence of SEQ ID NO: 109, and (ii) a light chain variable region containing the amino acid sequence of SEQ ID NO: 106; and (e) A fifth immunoglobulin comprising (i) a heavy chain variable region containing the amino acid sequence of SEQ ID NO: 110, and (ii) a light chain variable region containing the amino acid sequence of SEQ ID NO: 111, The method includes the step of culturing ex vivo cells containing one or more polynucleotides encoding an antibody selected from the group consisting of the following, under conditions such that one or more polynucleotides are expressed by the cells and the polynucleotides are produced.

[0054] In another embodiment, the Specified provides a method for producing an antibody heavy chain, comprising the step of culturing ex vivo cells containing a polynucleotide encoding one of the antibody heavy chains described herein, under conditions such that the polynucleotide is expressed by the cells to produce an antibody heavy chain encoded by the polynucleotide.

[0055] In another embodiment, the Specified provides a method for producing an antibody light chain, comprising the step of culturing ex vivo cells containing a polynucleotide encoding one of the antibody light chains described herein, under conditions such that the polynucleotide is expressed by the cells to produce an antibody light chain encoded by the polynucleotide.

[0056] In another embodiment, the Specified provides an antibody-drug conjugate comprising (a) an antibody moiety as described herein, which is an antibody or antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK); (b) one or more drug moieties, each of which is a cytotoxic agent; and (c) optionally a linker, wherein the cytotoxic agent is directly conjugated to the antibody moiety or conjugated to the antibody moiety via the linker.

[0057] In specific embodiments, the molar ratio of the antibody portion to the drug portion is between 1:1 and 1:12.

[0058] In specific embodiments, the molar ratio of the antibody portion to the drug portion is between 1:1 and 1:8.

[0059] In another specific embodiment, the molar ratio of the antibody portion to the drug portion is between 1:3 and 1:5.

[0060] In another specific embodiment, the molar ratio of the antibody portion to the drug portion is 1:9.

[0061] In specific embodiments, the antibody-drug conjugate includes a linker, which is a cleavage linker.

[0062] In another specific embodiment, the antibody-drug conjugate includes a linker, which is a non-cleavable linker.

[0063] In another specific embodiment, the antibody-drug conjugate includes a linker, which is maleimidocaproyl-valine-citrulline-p-aminobenzyloxycarbonyl (also known as "mc-vc-PABC") (see Figure 13A for the structure of mc-vc-PABC). In yet another specific embodiment, the antibody-drug conjugate includes a linker, which is CL2 (see Figure 13B and Cardillo et al. 2011, Clin Cancer Res; 17(10); 3157-69 for the structure of CL2). In another specific embodiment, the antibody-drug conjugate includes a linker, which is CL2A (see Figure 13B and Goldberg et al. 2018, Oncotarget; 9(48); 28989-29006, and Cardillo et al. 2011, Clin Cancer Res; 17(10); 3157-69 for the structure of CL2A).

[0064] In specific embodiments, the cytotoxic agent is a small molecule, nucleotide, peptide, or non-antibody protein. In a further embodiment of such specific embodiments, the cytotoxic agent is a small molecule.

[0065] In another specific embodiment, the cytotoxic agent is auristatin, maytansinoid, pyrrolobenzodiazepine, indolinobenzodiazepine, calicheamicin, camptothecin analog, duocalmycin, tubulin inhibitor, tubulicin or tubulicin analog, amberstatin 269, doxorubicin, antibiotic, anthracycline, microtubule inhibitor, splicostatin, or tylanstatin. In a further embodiment of such a specific embodiment, the cytotoxic agent is monomethyl auristatin E (MMAE) or monomethyl auristatin F (MMAF). In another further embodiment of such a specific embodiment, the cytotoxic agent is DM1 or DM4. In yet another further embodiment of such a specific embodiment, the cytotoxic agent is SN-38.

[0066] In another embodiment, the Specified provides a method for producing an antibody-drug conjugate as described herein, wherein a linker is absent, and the method comprises the steps of (a) directly conjugating a cytotoxic agent to the antibody moiety to produce an antibody-drug conjugate, and (b) purifying the antibody-drug conjugate.

[0067] In another embodiment, the Specified Publicly Provided Method for Producing an Antibody-Drug Conjugate as Described herein, wherein the antibody-drug conjugate comprises a linker, and the Method comprises the following steps in the order described above: (a) conjugating the linker directly to an antibody moiety to produce a linker-antibody moiety; (b) conjugating the linker of the linker-antibody moiety directly to a cytotoxic agent to produce an antibody-drug conjugate; and (c) purifying the antibody-drug conjugate.

[0068] In another embodiment, the Specified Publicly Provided Method for Producing an Antibody-Drug Conjugate as Described herein, wherein the antibody-drug conjugate comprises a linker, and the Method comprises the following steps in the order described above: (a) conjugating the linker directly to a cytotoxic agent to produce a linker-cytotoxic agent portion; (b) conjugating the linker of the linker-cytotoxic agent portion directly to an antibody portion to produce an antibody-drug conjugate; and (c) purifying the antibody-drug conjugate.

[0069] In another embodiment, this specification provides a pharmaceutical composition comprising an anti-MERTK antibody or its antigen-binding fragment, or an antibody-drug conjugate as described herein. In a specific embodiment, this specification provides a pharmaceutical composition comprising a therapeutically effective amount of either the antibody or antigen-binding fragment, or any antibody-drug conjugate as described herein. In a specific embodiment, the pharmaceutical composition is (a) A first immunoglobulin comprising (i) a heavy chain variable region containing the amino acid sequence of SEQ ID NO: 105, and (ii) a light chain variable region containing the amino acid sequence of SEQ ID NO: 106; (b) A second immunoglobulin comprising (i) a heavy chain variable region containing the amino acid sequence of SEQ ID NO: 107, and (ii) a light chain variable region containing the amino acid sequence of SEQ ID NO: 106; (c) A third immunoglobulin comprising (i) a heavy chain variable region containing the amino acid sequence of SEQ ID NO: 108, and (ii) a light chain variable region containing the amino acid sequence of SEQ ID NO: 106; (d) A fourth immunoglobulin comprising (i) a heavy chain region containing the amino acid sequence of SEQ ID NO: 109, and (ii) a light chain variable region containing the amino acid sequence of SEQ ID NO: 106; or (e) A fourth immunoglobulin comprising (i) a heavy chain variable region containing the amino acid sequence of SEQ ID NO: 110, and (ii) a light chain variable region containing the amino acid sequence of SEQ ID NO: 106, It also includes a pharmaceutically acceptable carrier.

[0070] In some embodiments, the pharmaceutical composition includes a polynucleotide sequence comprising a nucleic acid sequence encoding an antibody or an antigen-binding fragment thereof.

[0071] In another embodiment, this specification provides a method for treating cancer in a subject requiring such treatment, comprising the step of administering to the subject an anti-MERTK antibody or its antigen-binding fragment, or an antibody-drug conjugate, as described herein. In a specific embodiment, this specification provides a method for treating cancer in a subject requiring such treatment, comprising the step of administering to the subject any of the pharmaceutical compositions described herein.

[0072] In some embodiments, tumor samples derived from the subject are evaluated for overexpression of phosphorylated MERTK before treatment according to the method described herein. In certain embodiments, the subject is treated if the tumor sample overexpresses phosphorylated MERTK.

[0073] In specific embodiments of the preceding models of the method for treating cancer, the cancer is cancer of the head and neck, lung, breast, bone, ovary, stomach, pancreas, larynx, esophagus, testis, liver, parotid gland, bile duct, colon, rectum, cervix, uterus, endometrium, kidney, bladder, prostate, or thyroid. In certain embodiments of such specific models, the cancer is breast cancer. In further embodiments of such certain models, the cancer is triple-negative breast cancer.

[0074] In specific embodiments of the preceding aspects of the method for treating cancer, the cancer is sarcoma, squamous cell carcinoma, melanoma, glioma, glioblastoma, neuroblastoma, gastric cancer, colorectal cancer, non-small cell lung cancer, or Kaposi's sarcoma. In specific embodiments of the method for treating cancer, the cancer is leukemia or lymphoma. In further specific embodiments, the cancer is acute myeloid leukemia. In yet another further specific embodiment, the cancer is acute lymphoblastic leukemia. In yet another further embodiment, the cancer is multiple myeloma.

[0075] In specific embodiments, the cancer treated according to the method described herein is either overexpressing MERTK, associated with constitutively active MERTK (e.g., human MERTK), or both. In another specific embodiment, the cancerous cells of the cancer overexpress phosphorylated MERTK.

[0076] In a specific embodiment of a method for treating cancer, the method further includes the step of administering an additional therapeutic agent to the subject.

[0077] In specific embodiments of the method, which further includes the step of administering an additional therapeutic agent to a subject, the additional therapeutic agent is for treating cancer. The additional therapeutic agent may be administered in the same pharmaceutical composition, or in a different pharmaceutical composition other than an anti-MERTK antibody or its antigen-binding fragment, or an antibody-drug conjugate.

[0078] In specific embodiments of the embodiment in which the additional therapeutic agent is for treating cancer, the additional therapeutic agent is a drug used to treat breast cancer, a drug used to treat melanoma, an immunotherapy, or an angiogenesis inhibitor. In a further embodiment of such specific embodiments, the additional therapeutic agent is a drug used to treat breast cancer, selected from the group consisting of tamoxifen, raloxifene, paclitaxel, cyclophosphamide, docetaxel, vinblastine, fluorouracil, everolimus, trastuzumab, trastuzumab-emtansine, pertuzumab, and lapatinib nitosylate. In another further embodiment of such specific embodiments, the additional therapeutic agent is a drug used to treat melanoma, selected from the group consisting of BRAF inhibitors, MEK inhibitors, and dacarbazine. In yet another further embodiment of such specific embodiments, the additional therapeutic agent is a drug that blocks immune checkpoint signaling. In yet another further embodiment, the additional therapeutic agent is an anti-CTLA-4 antibody, an anti-PD-1 antibody, or an anti-PD-L1 antibody. In yet another further embodiment of such a specific embodiment, the additional therapeutic agent is an angiogenesis inhibitor selected from the group consisting of VEGF inhibitors, VEGFR2 inhibitors, sunitinib, and sorafenib.

[0079] In a specific embodiment of a method for treating cancer, the subject is human. In a specific embodiment, this specification provides an antibody-drug conjugate comprising an antibody moiety comprising a heavy chain variable region (VH) and a light chain variable region (VL), wherein VH comprises the amino acid sequence of sequence number 105, VL comprises the amino acid sequence of sequence number 106, and the antibody moiety is linked to MMAE via an mc-vc-PABC linker. In another specific embodiment, this specification provides an antibody-drug conjugate comprising an antibody moiety comprising a heavy chain variable region (VH) and a light chain variable region (VL), wherein VH comprises the amino acid sequence of sequence number 105, VL comprises the amino acid sequence of sequence number 106, and the antibody moiety is linked to SN-38 via a CL2A linker.

[0080] In certain embodiments, the antibody-drug conjugate includes an antibody moiety that is an immunoglobulin. In some embodiments, the antibody-drug conjugate includes an antibody moiety, the antibody moiety is an immunoglobulin, and the immunoglobulin includes a human constant region. 4. Brief explanation of the drawing [Brief explanation of the drawing]

[0081] [Figure 1] Determination of dissociation rates (Kd) for humanized anti-MERTK antibodies and chimeric anti-MERTK antibodies using surface plasmon resonance (SPR) measurements.

[0082] [Figure 2] Summary of protein purification for three humanized anti-MERTK antibodies (z10, z11, and z13) and a chimeric anti-MERTK antibody (xAb).

[0083] [Figure 3] Figure 3A. Flow cytometry analysis of the binding affinity of z10 antibody to human MERTK expressed by melanoma cells. Figure 3B. SPR measurement of z10 antibody binding to extracellular human MERTK.

[0084] [Figure 4] The z10 antibody specifically binds to human MERTK, as measured in affinity binding assays using ELISA (Figure 4A). However, the z10 antibody does not bind to the extracellular domains of human Axl (Figure 4B), human Tyro3 (Figure 4C), or mouse MERTK (Figure 4D), as evaluated in affinity binding assays using ELISA. The extracellular domains of human MERTK, human Axl, and mouse MERTK were conjugated with IgG Fc domains and used in ELISA.

[0085] [Figure 5A]The z10 antibody induces degradation of human MERTK in cancer cells (particularly SKMEL5 cells). SKMEL5 cells were incubated with various concentrations of z10 antibody for 24 hours (Figure 5A) or with 0.3 μg / mL of antibody for various periods (Figure 5B) before evaluating human MERTK levels by Western blot analysis. [Figure 5B] The z10 antibody induces degradation of human MERTK in cancer cells (particularly SKMEL5 cells). SKMEL5 cells were incubated with various concentrations of z10 antibody for 24 hours (Figure 5A) or with 0.3 μg / mL of antibody for various periods (Figure 5B) before evaluating human MERTK levels by Western blot analysis.

[0086] [Figure 6] The z10 antibody degrades human MERTK in vitro in differentiated human M2 macrophages. In vitro M2 macrophages were treated with 0.3 μg / mL of z10 antibody or control IgG, and the cells were incubated for varying periods before assessing human MERTK levels by Western blotting. Figure 6A shows the Western blotting results, and Figure 6B shows the relative quantification of human MERTK to tubulin.

[0087] [Figure 7] Degradation of human MERTK is mediated via internal translocation of human MERTK into SKMEL5 cells. SKMEL5 cells were incubated with control IgG or z10 antibodies, both labeled with pHrodo, which is measurable only under low pH conditions during lysosome uptake. Surface human MERTK was measured using commercially available MERTK antibodies. Figure 7A shows fluorescence over time, and Figure 7B is a representative image.

[0088] [Figure 8]The z10 antibody blocks Gas6-induced AKT phosphorylation in human cancer cells. SKMEL5 cells were incubated with 0.3 μl / mL of z10 antibody or control IgG for 2 hours, followed by incubation with 200 nM Gas6 for 10 minutes. Figure 8A shows the results of the Western blot, Figure 8B shows the amount of human MERTK compared to tubulin, and Figure 8C shows the amount of phosphorylated AKT relative to tubulin.

[0089] [Figure 9-1] The z10 antibody inhibits colony formation in human cancer cells. The figure shows the reduction in the number of colonies observed after culturing 500 SKMEL5 cells for 12 days in the presence of 0.3 μg / mL or 1 μg / mL of z10 antibody compared to the number of colonies observed when 500 SKMEL5 cells were cultured for 12 days with control IgG. Figure 9A shows the colony formation of cancer cells after 12 days of incubation with 0.3 μg / mL of z10 antibody or control IgG antibody. Figure 9B shows the colony formation of cancer cells after 12 days of incubation with 1 μg / mL of z10 antibody or control IgG antibody. Figure 9C shows the number of colonies formed after 12 days of incubation with 0.3 μg / mL of z10 antibody or control IgG antibody. Figure 9D shows the number of colonies formed after 12 days of incubation with 1 μg / mL of z10 antibody or control IgG antibody. [Figure 9-2]The z10 antibody inhibits colony formation in human cancer cells. The figure shows the reduction in the number of colonies observed after culturing 500 SKMEL5 cells for 12 days in the presence of 0.3 μg / mL or 1 μg / mL of z10 antibody compared to the number of colonies observed when 500 SKMEL5 cells were cultured for 12 days with control IgG. Figure 9A shows the colony formation of cancer cells after 12 days of incubation with 0.3 μg / mL of z10 antibody or control IgG antibody. Figure 9B shows the colony formation of cancer cells after 12 days of incubation with 1 μg / mL of z10 antibody or control IgG antibody. Figure 9C shows the number of colonies formed after 12 days of incubation with 0.3 μg / mL of z10 antibody or control IgG antibody. Figure 9D shows the number of colonies formed after 12 days of incubation with 1 μg / mL of z10 antibody or control IgG antibody.

[0090] [Figure 10] The z10 antibody induces a cytokine response in M2 macrophages and CD14+ monocytes.

[0091] [Figure 11] z10 inhibits cancer cell survival in vitro. 1,000 RPMI8226 multiple myeloma cells (Figure 11A) or 200 SKMEL5 melanoma cells (Figure 11B) were cultured in the presence of either an IgG control or z10. Survival rates were assessed on day 6 (RPMI8226) or day 4 (SKMel5) using the CellTiter-Glo 2.0 cell viability assay.

[0092] [Figure 12]Antitumor efficacy of the naked antibody against MDA-MB-231-LM2:z10. 50,000 MDA-MB-231-LM2 TNBC cells were injected into the tail vein of NSG mice. Treatment was initiated on the day of tumor cell inoculation, twice weekly via ip for the duration of the study. Figure 12A shows lung colonization of MDA-MB-231-LM2 triple-negative breast cancer cells using IVIS imaging. Figure 12B quantifies lung colonization of MDA-MB-231-LM2 triple-negative breast cancer cells. * represents a p-value less than 0.05.

[0093] [Figure 13A] Figure 13A shows the structure of maleimidocaproyl-valine-citrulline-p-aminobenzyloxycarbonyl-monomethylaulistatin E (mc-vc-PABC-MMAE) conjugated to antibody (Ab). Figure 13B shows the structure of SN-38 conjugated to antibody (Ab) via a CL2 or CL2A linker. "Ab" represents the antibody, which may be z10 or control IgG, for example, but is not limited to this. The CL2 linker has a cathepsin B cleavage site (see Figure 13B if AA is phenylalanine-lysine), but the CL2A linker does not have a cathepsin B cleavage site (see Figure 13B if AA is lysine). [Figure 13B] Figure 13A shows the structure of maleimidocaproyl-valine-citrulline-p-aminobenzyloxycarbonyl-monomethylaulistatin E (mc-vc-PABC-MMAE) conjugated to antibody (Ab). Figure 13B shows the structure of SN-38 conjugated to antibody (Ab) via a CL2 or CL2A linker. "Ab" represents the antibody, which may be z10 or control IgG, for example, but is not limited to this. The CL2 linker has a cathepsin B cleavage site (see Figure 13B if AA is phenylalanine-lysine), but the CL2A linker does not have a cathepsin B cleavage site (see Figure 13B if AA is lysine).

[0094] [Figure 14A] Figures 14A and 14B show the results of size exclusion high-pressure liquid chromatography measuring the purity of z10 conjugated to MMAE via an mc-vc-PABC linker (z10-MMAE, see Figure 13A, Ab is z10) (Figure 14A) and z10 conjugated to SN-38 via a CL2A linker (z10-SN-38, see Figure 13B, Ab is z10, AA is lysine) (Figure 14B). [Figure 14B] Figures 14A and 14B show the results of size exclusion high-pressure liquid chromatography measuring the purity of z10 conjugated to MMAE via an mc-vc-PABC linker (z10-MMAE, see Figure 13A, Ab is z10) (Figure 14A) and z10 conjugated to SN-38 via a CL2A linker (z10-SN-38, see Figure 13B, Ab is z10, AA is lysine) (Figure 14B).

[0095] [Figure 15] z10-ADC binds to MERTK with a Kd similar to that of z10. Figure 15 provides binding affinity data for z10-ADC. Recombinant hMER proteins (Mer-Fc conjugates) (50 nM to 0.05 nM) were equilibrated with 0.3 nM z10, z10-MMAE, or z10-SN-38, respectively, and then added to ELISA plates pre-coated with the hMER recombinant protein. Antibody binding was detected using an AP-conjugated anti-human IgG secondary antibody and colorimetrically developed using a GloMax microplate reader.

[0096] [Figure 16]Figures 16A and 16B show that MERTK is internally translocated by SKMel5 cells (Figure 16A) and degraded (Figure 16B) after incubation with z10, z10-MMAE, or z10-SN-38. z10, z10-MMAE, and z10-SN-38 degrade MERTK to a similar degree. SKMel5 cells were incubated with 6.7 nM pHrodo-labeled z10, z10-MMAE, or z10-SN-38. The pHrodo signal is measurable only under low pH conditions, for example, during lysosome uptake. Surface MERTK was stained with BV421 conjugated MERTK antibody. Binding was measured by flow cytometry.

[0097] [Figure 17-1] z10-MMAE and z10-SN-38 kill MERTK-expressing cancer cells. SKMel5 (Figure 17A) or RPMI8226 (Figure 17B) cells were incubated for 7 days with an IgG control, an IgG control conjugated to MMAE via an mc-mv-PABC linker (IgG-MMAE, see Figure 13A, Ab is IgG), or z10-MMAE. Cell viability was measured using CellTiterGlo. SKMel5 (Figure 17C) or RPMI8226 (Figure 17D) cells were incubated for 7 days with the indicated concentrations of an IgG control, an IgG control conjugated to SN-38 via a CL2A linker (IgG-SN-38, see Figure 13B, Ab is IgG, AA is lysine), or z10-SN-38. Cell viability was measured using CellTiterGlo. Figure 17E shows the Western blot analysis of whole cell lysates prepared from SKMel5 or RPMI8226 cells. [Figure 17-2]z10-MMAE and z10-SN-38 kill MERTK-expressing cancer cells. SKMel5 (Figure 17A) or RPMI8226 (Figure 17B) cells were incubated for 7 days with an IgG control, an IgG control conjugated to MMAE via an mc-mv-PABC linker (IgG-MMAE, see Figure 13A, Ab is IgG), or z10-MMAE. Cell viability was measured using CellTiterGlo. SKMel5 (Figure 17C) or RPMI8226 (Figure 17D) cells were incubated for 7 days with the indicated concentrations of an IgG control, an IgG control conjugated to SN-38 via a CL2A linker (IgG-SN-38, see Figure 13B, Ab is IgG, AA is lysine), or z10-SN-38. Cell viability was measured using CellTiterGlo. Figure 17E shows the Western blot analysis of whole cell lysates prepared from SKMel5 or RPMI8226 cells. [Figure 17-3] z10-MMAE and z10-SN-38 kill MERTK-expressing cancer cells. SKMel5 (Figure 17A) or RPMI8226 (Figure 17B) cells were incubated for 7 days with an IgG control, an IgG control conjugated to MMAE via an mc-mv-PABC linker (IgG-MMAE, see Figure 13A, Ab is IgG), or z10-MMAE. Cell viability was measured using CellTiterGlo. SKMel5 (Figure 17C) or RPMI8226 (Figure 17D) cells were incubated for 7 days with the indicated concentrations of an IgG control, an IgG control conjugated to SN-38 via a CL2A linker (IgG-SN-38, see Figure 13B, Ab is IgG, AA is lysine), or z10-SN-38. Cell viability was measured using CellTiterGlo. Figure 17E shows the Western blot analysis of whole cell lysates prepared from SKMel5 or RPMI8226 cells.

[0098] [Figure 18]z10-MMAE inhibits lung colonization of MDA-MB-231-LM2 TNBC cells. Figure 18A shows lung colonization of MDA-MB-231-LM2 triple-negative breast cancer cells using IVIS imaging. Figure 18B quantifies lung colonization of MDA-MB-231-LM2 triple-negative breast cancer cells. 50,000 MDA-MB-231-LM2 TNBC cells were injected into the tail vein of NSG mice. Treatment was initiated on the day of tumor cell inoculation, and lung metastatic colonization was monitored by bioluminescence imaging. Arrows in Figure 18B indicate the day of administration. ** indicates a p-value less than 0.01.

[0099] [Figure 19] z10 / z10-ADC did not affect the viability of MERTK-expressing M1 macrophages (Figure 19A) or M2 macrophages (Figure 19B). M2 macrophages were differentiated by culturing 8 × 10⁴ monocytes in PBMC medium with M-CSF (50 ng / mL) for 8 days. M1 macrophages were differentiated by culturing 8 × 10⁴ monocytes in PBMC medium with GM-CSF (20 ng / mL) for 5 days, and then cultured for a further 3 days with GM-CSF (20 ng / mL), IFNg (20 ng / mL), IL-6 (20 ng / mL), and LPS (10 pg / mL). M2 and M1 macrophages were treated with the indicated antibody or ADC in differentiation medium for 4 days, and viability was evaluated using the CellTiter-Glo 2.0 cell viability assay. [Modes for carrying out the invention]

[0100] 5. Detailed explanation This specification provides an anti-MERTK antibody (e.g., a monoclonal antibody) that specifically binds to MERTK (e.g., human MERTK), and an antigen-binding fragment thereof.

[0101] In certain embodiments, the anti-MERTK antibody (e.g., humanized antibody) or its antigen-binding fragment described herein specifically recognizes the extracellular domain of MERTK (e.g., human MERTK). In certain embodiments, the anti-MERTK antibody (e.g., humanized antibody) or its antigen-binding fragment described herein does not bind to human Axl, human Tyro3, ​​or mouse MERTK, as can be detected by techniques known to those skilled in the art or described herein. In specific embodiments, the anti-MERTK antibody (e.g., humanized antibody) or its antigen-binding fragment described herein does not bind to human Axl, human Tyro3, ​​or mouse MERTK, as can be evaluated by affinity binding assays using ELISA as described in Section 6.2 below.

[0102] In specific embodiments, the anti-MERTK antibody (e.g., a humanized antibody) or its antigen-binding fragment described herein specifically binds to the human MERTK protein containing the amino acid sequence of SEQ ID NO: 131. In another specific embodiment, the anti-MERTK antibody (e.g., a humanized antibody) or its antigen-binding fragment described herein specifically binds to the extracellular region of human MERTK containing the amino acid sequence of SEQ ID NO: 132. In yet another specific embodiment, the anti-MERTK antibody (e.g., a humanized antibody) or its antigen-binding fragment described herein specifically binds to SEQ ID NO: 132. In specific embodiments, the anti-MERTK antibody (e.g., a humanized antibody) or its antigen-binding fragment is bivalent. In specific embodiments, the anti-MERTK antibody (e.g., a humanized antibody) or its antigen-binding fragment described herein contains two antigen-binding sites that bind to MERTK (e.g., human MERTK). In a particular embodiment, the two antigen-binding sites bind to the same epitope on MERTK (e.g., human MERTK). In a particular embodiment, the two antigen-binding sites contain the same CDR.

[0103] In certain embodiments, an anti-MERTK antibody (e.g., a humanized antibody) or its antigen-binding fragment is determined to have a K content of about 1 pM to 10 nM, which is known to those skilled in the art or determined using techniques described herein. DThen, it binds to a recombinant MERTK-Fc chimera (e.g., a recombinant human MERTK-Fc chimera). In specific embodiments, the anti-MERTK antibody (e.g., a humanized antibody) or its antigen-binding fragment described herein binds to a K2 antibody of about 1 pM to about 6 pM, determined using surface plasmon resonance (SPR) as described in Section 6 below. D Then, it binds to a recombinant MERTK-Fc chimera (e.g., a recombinant human MERTK-Fc chimera). In certain embodiments, the anti-MERTK antibody (e.g., a humanized antibody) or its antigen-binding fragment is subjected to about 1 nM to 20 nM EC using an assay described herein or known to those skilled in the art. 50 Then, it binds to MERTK (e.g., human MERTK). In a specific embodiment, the anti-MERTK antibody (e.g., humanized antibody) or its antigen-binding fragment described herein binds to approximately 1 nM to 10 nM of K in the assay described in Section 6 below. D Then, it binds to MERTK (for example, human MERTK).

[0104] In certain embodiments, the anti-MERTK antibody (e.g., humanized antibody) or antibody fragment described herein has an EC of about 1 nM to 20 nM, as evaluated by flow cytometry, for example, as described herein or known to those skilled in the art. 50 The antibody then binds to human MERTK-expressing cells (e.g., human MERTK cancer cells or M2 macrophages). In specific embodiments, the anti-MERTK antibody (e.g., a humanized antibody) or its antigen-binding fragment described herein has an EC of 1 nm to 10 nM, as evaluated by flow cytometry as described in Section 6.2 below, for example. 50 It then binds to human MERTK-expressing cells (e.g., human MERTK cancer cells or M2 macrophages).

[0105] In certain embodiments, the anti-MERTK antibody (e.g., a humanized antibody) or its antigen-binding fragment described herein reduces the expression level of human MERTK in cancer cells, as evaluated by assays described herein (e.g., Section 6.2 below) or assays known to those skilled in the art. In certain embodiments, the anti-MERTK antibody (e.g., a humanized antibody) or its antigen-binding fragment describes herein reduces the expression level of human MERTK in SKMEL5 of melanoma cell lines, as evaluated by assays described herein (e.g., Section 6.2 below) or assays known to those skilled in the art. In certain embodiments, the anti-MERTK antibody (e.g., a humanized antibody) or its antigen-binding fragment reduces the expression level of human MERTK in human M2 macrophages, as evaluated by assays described herein (e.g., Section 6.2 below) or assays known to those skilled in the art. In certain embodiments, the anti-MERTK antibody (e.g., a humanized antibody) or its antigen-binding fragment reduces the expression level of human MERTK by internal migration of MERTK, as evaluated by assays described herein (e.g., Section 6.2 below) or assays known to those skilled in the art. In certain embodiments, an anti-MERTK antibody (e.g., a humanized antibody) or its antigen-binding fragment reduces the expression level of human MERTK by internal translocation to lysosomes, as evaluated by assays described herein (e.g., Section 6.2 below) or assays known to those skilled in the art.

[0106] In certain embodiments, the anti-MERTK antibody (e.g., a humanized antibody) or its antigen-binding fragment described herein reduces Gas-6-induced phosphorylation of AKT in human MERTK-expressing cells (e.g., cancer cells), as evaluated by assays described herein (e.g., Section 6.2 below) or assays known to those skilled in the art. In specific embodiments, the anti-MERTK antibody (e.g., a humanized antibody) or its antigen-binding fragment described herein prevents Gas-6-induced phosphorylation of AKT in human MERTK-expressing cancer cells, such as melanoma SKMEL5 cells, as evaluated by assays described herein (e.g., Section 6.2 below) or assays known to those skilled in the art. In certain embodiments, the anti-MERTK antibody (e.g., a humanized antibody) or its antigen-binding fragment described herein reduces Gas-6-induced phosphorylation of human MERTK in human MERTK-expressing cells, as evaluated by assays known to those skilled in the art. In specific embodiments, the anti-MERTK antibody (e.g., a humanized antibody) or its antigen-binding fragment described herein blocks Gas6-induced MERTK activation in human MERTK-expressing cancer cells (e.g., melanoma cells, e.g., SKMEL5, etc.) as evaluated by assays known to those skilled in the art.

[0107] In certain embodiments, the anti-MERTK antibody (e.g., a humanized antibody) or antigen-binding fragment described herein reduces the colony-forming ability of cancer cells, as evaluated by assays described herein (e.g., Section 6.2 below) or assays known to those skilled in the art. In certain embodiments, the anti-MERTK antibody (e.g., a humanized antibody) or antigen-binding fragment described herein inhibits colony formation of cancer cells (e.g., melanoma cells such as SKMEL5).

[0108] In certain embodiments, the anti-MERTK antibody (e.g., a humanized antibody) or its antigen-binding fragment described herein induces a cytokine response in M2 macrophages, CD14+ monocytes, or both, as evaluated using assays described herein or known to those skilled in the art, as provided in Figure 10.

[0109] In specific embodiments, the anti-MERTK antibody (e.g., a humanized antibody) or its antigen-binding fragment described herein is isolated.

[0110] The antibodies described herein may include monoclonal or polyclonal antibodies. In specific embodiments, antibodies may be immunoglobulins, tetrameric antibodies containing two heavy chain molecules and two light chain molecules, antibody light chain monomers, antibody heavy chain monomers, antibody light chain dimers, antibody heavy chain dimers, antibody light chain-antibody heavy chain pairs, single-domain antibodies, monovalent antibodies, single-chain antibodies, single-chain Fv(scFv), disulfide-linked Fv(scFv), and anti-idiotype (anti-Id) antibodies (e.g., anti-Id antibodies).

[0111] In certain embodiments, the antibodies described herein may be multispecific or bispecific. In certain embodiments, the antibodies described herein are bispecific monoclonal antibodies. In certain embodiments, the antibodies described herein are monovalent. In specific embodiments, the antibodies described herein are bivalent. In specific embodiments, the antibodies described herein bind to human MERTK on cells. In some embodiments, the antibodies described herein are monospecific. In certain embodiments, the antibodies described herein are recombinantly produced. In some embodiments, the antibodies described herein are synthetic antibodies. In specific embodiments, the antibodies described herein are humanized antibodies. In other embodiments, the antibodies described herein may be human antibodies.

[0112] The antibodies described herein can be of any type of immunoglobulin molecule (e.g., IgG, IgE, IgM, IgD, IgA or IgY), any class (e.g., IgG1, IgG2, IgG3, IgG4, IgA1 or IgA2), or any subclass (e.g., IgG 2a or IgG 2b ). In certain embodiments, the antibodies described herein are IgG antibodies, or a class or subclass thereof. In specific embodiments, the antibodies described herein (e.g., humanized antibodies) are monoclonal antibodies. In a specific embodiment, the anti-MERTK antibody described herein is an IgG antibody.

[0113] As used herein, the terms "antigen-binding fragment," "antigen-binding region" and similar terms refer to a part of an antibody molecule that contains the amino acid residues that confer its specificity for an antigen (e.g., complementarity determining regions (CDRs)). The antigen-binding region can be derived from any animal species such as rodents (e.g., mouse, rat or hamster) and humans. By way of example, antigen-binding fragments include Fab fragments, F(ab’)2 fragments, and antigen-binding fragments of any of the antibodies described above. In a specific embodiment, the antigen-binding fragment of a humanized antibody comprises the variable heavy chain region, variable light chain region or both of the z10, z11 or z13 antibodies described herein.

[0114] As used herein, the terms "variable region" or "variable domain" are used interchangeably and are common in the art. The variable region typically refers to a part of an antibody, generally part of a light or heavy chain, which generally varies significantly in sequence between antibodies and is used in the binding and specificity of a particular antibody for a particular antigen. The variability in the sequence is concentrated in regions called complementarity determining regions (CDRs), while the more highly conserved regions in the variable domain are called framework regions (FRs). Without wishing to be bound by any particular mechanism or theory, the CDRs of the light and heavy chains are thought to be the main cause of antibody-antigen interaction and specificity.

[0115] CDR is defined in various ways within the art, including Kabat, Chothia, AbM, contact, IMGT, and Exemplary definitions. Kabat's definition is based on sequence variability and is the most commonly used definition for predicting CDR regions (Kabat, Elvin A. et al., Sequences of Proteins of Immunological Interest. Bethesda: National Institutes of Health, 1983). Chothia's definition is based on the location of structural loop regions (Chothia et al., (1987) J Mol Biol 196: 901-917). AbM's definition, a compromise between Kabat and Chothia's definitions, is an integrated software package for antibody structure modeling created by Oxford Molecular Group (bioinf.org.uk / abs) (Martin ACR et al., (1989) PNAS 86: 9268-9272). The definition of contact is based on the analysis of available complex crystal structures (bioinf.org.uk / abs) (see MacCallum RM et al., (1996) J Mol Biol 5: 732-745). The definition of IMGT is from IMGT ("IMGT®, the international ImMunoGeneTics information system® website imgt.org, founder and director: Marie-Paule Lefranc, Montpellier, France"). The definition of example is a combination of AbM and Kabat (Presta et al., (1997) Cancer Res 57: 4593-4599).

[0116] In specific embodiments, the humanized antibody or antigen-binding fragment described herein comprises a variable heavy chain region, a variable light chain region, or both, wherein the variable heavy chain region, the variable light chain region, or both are the variable heavy chain region, the variable heavy chain region, or both of the z10, z11, or z12 antibody, which are provided in Tables 11, 12, and 13, respectively. In another specific embodiment, the humanized antibody or antigen-binding fragment described herein comprises a variable heavy chain region, a variable light chain region, or both, wherein the variable heavy chain comprises the amino acid sequence of the variable heavy chain region in Table 14, and the variable light chain region comprises the amino acid sequence of the variable light chain region in Tables 11, 12, or 13 below. In specific embodiments, the humanized antibody is monoclonal. In some embodiments, the humanized antibody is a tetrameric antibody containing immunoglobulin, two heavy chain molecules, and two light chain molecules, an antibody light chain monomer, an antibody heavy chain monomer, an antibody light chain dimer, and an antibody heavy chain dimer, an antibody light chain-antibody heavy chain pair, a single-domain antibody, a single-chain antibody, a single-chain Fv, a disulfide-linked Fv, or an anti-idiotype antibody. The humanized antibody is any type (e.g., IgG, IgE, IgM, IgD, IgA, or IgY), any class (e.g., IgG1, IgG2, IgG3, IgG4, IgA, or IgA2), or any subclass (e.g., IgG) of the immunoglobulin molecule. 2a or IgG 2b ) may be of the same nature. In specific embodiments, the humanized antibody described herein is IgG, or a class or subclass thereof.

[0117] This application envisions not only antibodies and their antigen-binding fragments comprising sequences disclosed herein (e.g., CDRs, framework regions, variable regions), but also antibodies and their antigen-binding fragments consisting of or essentially derived from sequences disclosed herein. For example, the antibody may consist of the light and heavy chain sequences of the z10, z11, or z13 antibodies described herein.

[0118] In specific embodiments, this specification provides a chimeric anti-MERTK antibody or its antigen-binding fragment comprising a variable heavy chain region, a variable light chain region, or both, wherein the variable light chain comprises the amino acid sequence of the variable light chain region shown in Table 15 below, and the variable heavy chain comprises the amino acid sequence of the variable heavy chain region shown in Table 15 below. In specific embodiments, the chimeric antibody is monoclonal. In some embodiments, the chimeric antibody is a tetrameric antibody comprising immunoglobulin, two heavy chain molecules, and two light chain molecules, an antibody light chain monomer, an antibody heavy chain monomer, an antibody light chain dimer, an antibody heavy chain dimer, an antibody light chain-antibody heavy chain pair, a single-domain antibody, a single-chain antibody, a single-chain Fv, a disulfide-linked Fv, or an anti-idiotype antibody. Chimeric antibodies are any type (e.g., IgG, IgE, IgM, IgD, IgA, or IgY), any class (e.g., IgG1, IgG2, IgG3, IgG4, IgA, or IgA2), or any subclass (e.g., IgG) of an immunoglobulin molecule. 2a or IgG 2b ) may be of the same nature. In certain embodiments, the chimeric antibody described herein is an IgG antibody, or a class or subclass thereof.

[0119] In other embodiments, this specification provides multispecific antibodies and heteroconjugate antibodies. In specific embodiments, this specification provides a bispecific antibody comprising two different antigen-binding regions, wherein one binding region binds to MERTK (e.g., human MERTK) and comprises a variable heavy chain region, a variable light chain region, or both of the antibodies described herein (e.g., z10, z11, z13, or xAb antibodies), and the other binding region binds to the antigen of interest. In some embodiments, the bispecific antibody comprises two different antigen-binding regions, where one binding region specifically binds to MERTK (e.g., human MERTK), and the other binding region binds to the antigen of interest, wherein the binding region that specifically binds to MERTK (e.g., human MERTK) comprises a variable heavy chain region and a variable light chain region of the z10, z12, or z13 antibodies described herein. In specific embodiments, the antigen of interest is an immune cell receptor or a tumor-associated antigen. For bispecific antibodies, see Section 5.1.3 below.

[0120] Heteroconjugate antibodies are also provided herein. In specific embodiments, the heteroconjugate antibody comprises two monoclonal antibodies having specificity for two different antigens, one of which comprises a variable heavy chain region and a variable light chain region as described herein (e.g., z10, z11, z13, or xAb antibody).

[0121] We also provide isolated nucleic acids (polynucleotides), such as the anti-MERTK antibodies described herein and their complementary DNA (cDNA) encoding the antigen-binding fragments. In certain embodiments, we provide polynucleotides comprising the VH, VL, or both nucleic acid sequences of the z10, z11, z13, or xAb antibodies, as shown in Tables 21, 22, 23, or 24. In some embodiments, we provide polynucleotides comprising the heavy chain, light chain, or both, of the z10, z11, z13, or xAb antibodies, as shown in Tables 25, 26, 27, or 28.

[0122] Furthermore, this specification provides vectors (e.g., expression vectors) and cells (e.g., host cells) containing nucleic acids (polynucleotides) encoding the anti-MERTK antibody or its antigen-binding fragment described herein. Methods for producing such antibodies are also provided.

[0123] In another aspect, the Specified provides an antibody-drug conjugate comprising (a) an antibody moiety which is an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to a MERTK as Specified herein (e.g., human MERTK), (b) one or more drug moieties, each drug moiety which is a cytotoxic agent, and (c) optionally a linker, wherein the cytotoxic agent is directly conjugated to the antibody moiety or conjugated to the antibody moiety via the linker. As used in this disclosure, the term “conjugated” means a covalent bond which may be direct or via an intervening covalent structure.

[0124] Where the cytotoxic agent is a peptide or protein, or where the cytotoxic agent and linker (if present) are peptides or proteins, nucleic acids encoding an antibody-drug conjugate are also provided. In other embodiments, methods for producing an antibody-drug conjugate comprising (i) an anti-MERTK antibody or its antigen-binding fragment, and (ii) a cytotoxic agent directly conjugated to the anti-MERTK antibody or antigen-binding fragment, or conjugated to the anti-MERTK antibody or antigen-binding fragment via a linker are provided herein.

[0125] In another embodiment, this specification provides compositions (e.g., pharmaceutical compositions) comprising the anti-MERTK antibody or its antigen-binding fragment as described herein. In specific embodiments, the pharmaceutical composition comprises the anti-MERTK antibody or its antigen-binding fragment as described herein, one or more pharmaceutically acceptable carriers, excipients or both, and optionally one or more other therapeutic agents.

[0126] In another embodiment, this specification provides compositions (e.g., pharmaceutical compositions) comprising the antibody-drug conjugate described herein. In specific embodiments, the pharmaceutical composition comprises the antibody-drug conjugate described herein, one or more pharmaceutically acceptable carriers, excipients, or both, and optionally one or more other therapeutic agents.

[0127] In another embodiment, this specification provides a method for treating cancer in a subject, comprising the step of administering to the subject an effective amount of an anti-MERTK antibody or its antigen-binding fragment, or a composition thereof, as described herein. In specific embodiments, the anti-MERTK antibody or its antigen-binding fragment is a humanized antibody or antigen-binding fragment as described herein. In other embodiments, the anti-MERTK antibody or its antigen-binding fragment is a bispecific antibody or heteroconjugate antibody as described herein. In some embodiments, the anti-MERTK antibody or its antigen-binding fragment is a chimeric antibody or its antigen-binding fragment.

[0128] In another embodiment, the Specified provides a method for treating cancer in a subject, comprising the step of administering to the subject an effective amount of the antibody-drug conjugate described herein. In a specific embodiment, the Specified provides a method for treating cancer in a subject, comprising the step of administering to the subject an effective amount of the antibody-drug conjugate comprising (i) an anti-MERTK antibody or its antigen-binding fragment, and (ii) a cytotoxic agent that is directly conjugated to the anti-MERTK antibody or its antigen-binding fragment, or conjugated to the anti-MERTK antibody or its antigen-binding fragment via a linker. 5.1. Antibodies 5.1.1. Sequences and Variants

[0129] Tables 1 and 7 or 8 below provide VH CDRs and VL CDRs, respectively, of anti-MERTK antibodies or their antigen-binding fragments, defined using different systems. Tables 2, 3, 4, 5, and 6 below provide VH framework regions, defined using different systems, which may be combined with the CDRs in Table 1. In specific embodiments, the CDRs in Table 1 are combined with VH framework regions 1, 2, and 4 using the same numbering system, and with VH framework region 3 of the consensus sequence using one numbering system, which are provided in Tables 2, 3, 4, or 6. Tables 9 and 10 below provide VL framework regions, defined using different systems, which may be combined with the CDRs in Table 7 or 8. Tables 11, 12, 13, and 15 provide VH and VL amino acid sequences for specific antibodies. Table 14 provides VH amino acid sequences, which may be combined with the VL provided in Tables 11, 12, 13, or 15. Tables 16, 17, 18, and 20 provide amino acid sequences for the heavy and light chains of specific antibodies. Table 19 provides heavy chain amino acid sequences that may be combined with the light chain amino acid sequences of Tables 16, 17, or 18. Tables 21, 22, 23, and 24 provide nucleic acid sequences encoding the variable region of specific antibodies. Tables 25, 26, 27, and 28 provide nucleic acid sequences encoding the heavy and light chains of specific antibodies. Tables 29 and 30 provide exemplary human MERTK amino acid sequences to which the anti-MERTK antibodies or their antigen-binding fragments described herein may bind.

[0130] In specific embodiments, the anti-MERTK antibody or its antigen-binding fragment provided herein includes VH CDR1 of the antibody in Table 1, as defined by Kabat, Chothia, AbM, Contact, IMGT, or Exemplary. In some embodiments, the anti-MERTK antibody or its antigen-binding fragment provided herein includes VH CDR2 of the antibody in Table 1, as defined by Kabat, Chothia, AbM, Contact, IMGT, or Exemplary. In some embodiments, the anti-MERTK antibody or its antigen-binding fragment provided herein includes VH CDR3 of the antibody in Table 1, as defined by Kabat, Chothia, AbM, Contact, IMGT, or Exemplary. In some embodiments, the anti-MERTK antibody or its antigen-binding fragment provided herein includes one, two, or all three of the VH CDRs of the antibodies in Table 1, as determined by a numbering system.

[0131] In specific embodiments, the anti-MERTK antibody or its antigen-binding fragment provided herein includes VL CDR1 of the antibody in Table 7, as defined by Kabat, Chothia, AbM, Contact, IMGT, or Exemplary. In some embodiments, the anti-MERTK antibody or its antigen-binding fragment provided herein includes VL CDR2 of the antibody in Table 7, as defined by Kabat, Chothia, AbM, Contact, IMGT, or Exemplary. In some embodiments, the anti-MERTK antibody or its antigen-binding fragment provided herein includes VL CDR3 of the antibody in Table 7, as defined by Kabat, Chothia, AbM, Contact, IMGT, or Exemplary. In some embodiments, the anti-MERTK antibody or its antigen-binding fragment provided herein includes one, two, or all three of the VL CDRs of the antibodies in Table 7, as determined by a numbering system.

[0132] [Table 1]

[0133] Table 2-1 Table 2-2

[0134] Table 3

[0135] Table 4

[0136] Table 5-1 Table 5-2

[0137] Table 6

[0138] Table 7-1 Table 7-2

[0139] Table 8

[0140] Table 9-1 Table 9-2

[0141] Table 10

[0142] Table 11-1 Table 11-2

[0143] Table 12

[0144] Table 13

[0145] Table 14

[0146] Table 15

[0147] Table 16

[0148] Table 17

[0149] Table 18-1 Table 18-2

[0150] Table 19

[0151] Table 20

[0152] Table 21-1 Table 21-2

[0153] Table 22

[0154] Table 23

[0155] Table 24-1 Table 24-2

[0156] Table 25

[0157] Table 26-1 Table 26-2

[0158] Table 27

[0159] Table 28

[0160] Table 29-1 Table 29-2

[0161] In specific embodiments, this specification provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or antigen-binding fragment comprises a framework region (FR) 1, variable heavy chain regions (VH) CDR1, FR2, VH CDR2, FR3, VH CDR3 and FR4, wherein VH CDR1, VH CDR2, and VH CDR3 comprise the amino acid sequences of VH CDR1, VH CDR2 and VH CDR3 shown in Table 1 using Exemplary's numbering system, and FR1, FR2, FR3 and FR4 comprise the amino acid sequences of FR1, FR2, FR3 and FR4 shown in Tables 2, 3, 4, or 5 using Exemplary's numbering system. In another specific embodiment, the Specified herein provides an anti-MERTK or antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or antigen-binding fragment comprises a framework region (FR)1, variable heavy chain regions (VH)CDR1, FR2, VH CDR2, FR3, VH CDR3 and FR4, wherein VH CDR1, VH CDR2, and VH CDR3 comprise the amino acid sequences of VH CDR1, VH CDR2 and VH CDR3 shown in Table 1 using the Kabat numbering system, and FR1, FR2, FR3 and FR4 comprise the amino acid sequences of FR1, FR2, FR3 and FR4 shown in Tables 2, 3, 4, or 5 using the Kabat numbering system.In another specific embodiment, the Specified herein provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or the antigen-binding fragment comprises a framework region (FR) 1, variable heavy chain regions (VH) CDR1, FR2, VH CDR2, FR3, VH CDR3 and FR4, wherein VH CDR1, VH CDR2, and VH CDR3 comprise the amino acid sequences of VH CDR1, VH CDR2 and VH CDR3 shown in Table 1 using Chothia's numbering system, and FR1, FR2, FR3 and FR4 comprise the amino acid sequences of FR1, FR2, FR3 and FR4 shown in Tables 2, 3, 4, or 5 using Chothia's numbering system. In another specific embodiment, the Specified herein provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or the antigen-binding fragment comprises a framework region (FR) 1, variable heavy chain regions (VH) CDR1, FR2, VH CDR2, FR3, VH CDR3 and FR4, wherein VH CDR1, VH CDR2, and VH CDR3 comprise the amino acid sequences of VH CDR1, VH CDR2 and VH CDR3 shown in Table 1 using the AbM numbering system, and FR1, FR2, FR3 and FR4 comprise the amino acid sequences of FR1, FR2, FR3 and FR4 shown in Tables 2, 3, 4, or 5 using the AbM numbering system.In another specific embodiment, the Specified Information provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or the antigen-binding fragment comprises a framework region (FR) 1, variable heavy chain regions (VH) CDR1, FR2, VH CDR2, FR3, VH CDR3 and FR4, wherein VH CDR1, VH CDR2, and VH CDR3 comprise the amino acid sequences of VH CDR1, VH CDR2 and VH CDR3 shown in Table 1 using Contact's numbering system, and FR1, FR2, FR3 and FR4 comprise the amino acid sequences of FR1, FR2, FR3 and FR4 shown in Tables 2, 3, 4, or 5 using Contact's numbering system. In another specific embodiment, the Specified herein provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or the antigen-binding fragment comprises a framework region (FR) 1, variable heavy chain regions (VH) CDR1, FR2, VH CDR2, FR3, VH CDR3 and FR4, wherein VH CDR1, VH CDR2, and VH CDR3 comprise the amino acid sequences of VH CDR1, VH CDR2 and VH CDR3 shown in Table 1 using the IMGT numbering system, and FR1, FR2, FR3 and FR4 comprise the amino acid sequences of FR1, FR2, FR3 and FR4 shown in Tables 2, 3, 4, or 5 using the IMGT numbering system.

[0162] In another specific embodiment, the Specified Information provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or the antigen-binding fragment comprises a framework region (FR) 1, variable heavy chain regions (VH) CDR1, FR2, VH CDR2, FR3, VH CDR3, and FR4, wherein VH CDR1, VH CDR2, and VH CDR3 comprise the amino acid sequences of VH CDR1, VH CDR2, and VH CDR3 shown in Table 1 using a single numbering system, FR1, FR2, and FR4 comprise the amino acid sequences of FR1, FR2, and FR4 shown in Tables 2, 3, or 4 using the same numbering system, and FR3 comprises the amino acid sequence of FR3 shown in Table 5 using the same numbering system.

[0163] In specific embodiments, this specification provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or antigen-binding fragment comprises a variable heavy chain region (VH), the VH comprising a framework region (FR) 1, VH CDR1, FR2, VH CDR2, FR3, VH CDR3, and FR4, the VH CDR1, VH CDR2, and VH CDR3 comprising the amino acid sequences of VH CDR1, VH CDR2, and VH CDR3 shown in Table 1 using Exemplary's numbering system, and the FR1, FR2, FR3, and FR4 comprising the amino acid sequences of FR1, FR2, FR3, and FR4 shown in Tables 2, 3, 4, or 5 using Exemplary's numbering system. In another specific embodiment, the Specified Information provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or antigen-binding fragment comprises a variable heavy chain region (VH), the VH comprising a framework region (FR) 1, VH CDR1, FR2, VH CDR2, FR3, VH CDR3, and FR4, the VH CDR1, VH CDR2, and VH CDR3 comprising the amino acid sequences of VH CDR1, VH CDR2, and VH CDR3 shown in Table 1 using the Kabat numbering system, and the FR1, FR2, FR3, and FR4 comprising the amino acid sequences of FR1, FR2, FR3, and FR4 shown in Tables 2, 3, 4, or 5 using the Kabat numbering system.In another specific embodiment, the Specified Public Service provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or the antigen-binding fragment comprises a variable heavy chain region (VH), the VH comprising a framework region (FR) 1, VH CDR1, FR2, VH CDR2, FR3, VH CDR3, and FR4, the VH CDR1, VH CDR2, and VH CDR3 comprising the amino acid sequences of VH CDR1, VH CDR2, and VH CDR3 shown in Table 1 using Chothia's numbering system, and the FR1, FR2, FR3, and FR4 comprising the amino acid sequences of FR1, FR2, FR3, and FR4 shown in Tables 2, 3, 4, or 5 using Chothia's numbering system. In another specific embodiment, the Specified Public Service provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or the antigen-binding fragment comprises a variable heavy chain region (VH), the VH comprising a framework region (FR) 1, VH CDR1, FR2, VH CDR2, FR3, VH CDR3, and FR4, the VH CDR1, VH CDR2, and VH CDR3 comprising the amino acid sequences of VH CDR1, VH CDR2, and VH CDR3 shown in Table 1 using the AbM numbering system, and the FR1, FR2, FR3, and FR4 comprising the amino acid sequences of FR1, FR2, FR3, and FR4 shown in Tables 2, 3, 4, or 5 using the AbM numbering system.In another specific embodiment, the Specified Public Service provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or the antigen-binding fragment comprises a variable heavy chain region (VH), the VH comprising a framework region (FR) 1, VH CDR1, FR2, VH CDR2, FR3, VH CDR3, and FR4, the VH CDR1, VH CDR2, and VH CDR3 comprising the amino acid sequences of VH CDR1, VH CDR2, and VH CDR3 shown in Table 1 using Contact's numbering system, and the FR1, FR2, FR3, and FR4 comprising the amino acid sequences of FR1, FR2, FR3, and FR4 shown in Tables 2, 3, 4, or 5 using Contact's numbering system. In another specific embodiment, the Specified herein provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or the antigen-binding fragment comprises a variable heavy chain region (VH), the VH comprising a framework region (FR) 1, VH CDR1, FR2, VH CDR2, FR3, VH CDR3, and FR4, the VH CDR1, VH CDR2, and VH CDR3 comprising the amino acid sequences of VH CDR1, VH CDR2, and VH CDR3 shown in Table 1 using the IMGT numbering system, and the FR1, FR2, FR3, and FR4 comprising the amino acid sequences of FR1, FR2, FR3, and FR4 shown in Tables 2, 3, 4, or 5 using the IMGT numbering system.

[0164] In another specific embodiment, the Specified Information provides an anti-MERTK antibody or antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or antigen-binding fragment comprises a variable heavy chain region (VH), the VH comprising framework region (FR) 1, VH CDR1, FR2, VH CDR2, FR3, VH CDR3, and FR4, the VH CDR1, VH CDR2, and VH CDR3 comprising the amino acid sequences of VH CDR1, VH CDR2, and VH CDR3 shown in Table 1 using a single numbering system, the FR1, FR2, and FR4 comprising the amino acid sequences of FR1, FR2, and FR4 shown in Tables 2, 3, or 4 using the same numbering system, and the FR3 comprising the amino acid sequence of FR3 shown in Table 5 using the same numbering system.

[0165] In specific embodiments, this specification provides an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or its antigen-binding fragment comprises a framework region (FR) 1, variable light chain regions (VL) CDR1, FR2, VL CDR2, FR3, VL CDR3 and FR4, wherein VL CDR1, VL CDR2, and VL CDR3 comprise the amino acid sequences of VL CDR1, VL CDR2 and VL CDR3 shown in Table 7 using Exemplary's numbering system, and FR1, FR2, FR3 and FR4 comprise the amino acid sequences of FR1, FR2, FR3 and FR4 shown in Table 9 using Exemplary's numbering system. In another specific embodiment, the Specified herein provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or the antigen-binding fragment comprises a framework region (FR) 1, variable light chain regions (VL) CDR1, FR2, VL CDR2, FR3, VL CDR3 and FR4, wherein VL CDR1, VL CDR2, and VL CDR3 comprise the amino acid sequences of VL CDR1, VL CDR2 and VL CDR3 shown in Table 7 using the Kabat numbering system, and FR1, FR2, FR3 and FR4 comprise the amino acid sequences of FR1, FR2, FR3 and FR4 shown in Table 9 using the Kabat numbering system.In another specific embodiment, the Specified herein provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or the antigen-binding fragment comprises a framework region (FR) 1, variable light chain regions (VL) CDR1, FR2, VL CDR2, FR3, VL CDR3 and FR4, wherein VL CDR1, VL CDR2, and VL CDR3 comprise the amino acid sequences of VL CDR1, VL CDR2 and VL CDR3 shown in Table 7 using Chothia's numbering system, and FR1, FR2, FR3 and FR4 comprise the amino acid sequences of FR1, FR2, FR3 and FR4 shown in Table 9 using Chothia's numbering system. In another specific embodiment, the Specified herein provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or the antigen-binding fragment comprises a framework region (FR) 1, variable light chain regions (VL) CDR1, FR2, VL CDR2, FR3, VL CDR3 and FR4, wherein VL CDR1, VL CDR2, and VL CDR3 comprise the amino acid sequences of VL CDR1, VL CDR2 and VL CDR3 shown in Table 7 using the AbM numbering system, and FR1, FR2, FR3 and FR4 comprise the amino acid sequences of FR1, FR2, FR3 and FR4 shown in Table 9 using the AbM numbering system.In another specific embodiment, the Specified Information provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or the antigen-binding fragment comprises a framework region (FR) 1, variable light chain regions (VL) CDR1, FR2, VL CDR2, FR3, VL CDR3 and FR4, wherein VL CDR1, VL CDR2, and VL CDR3 comprise the amino acid sequences of VL CDR1, VL CDR2 and VL CDR3 shown in Table 7 using Contact's numbering system, and FR1, FR2, FR3 and FR4 comprise the amino acid sequences of FR1, FR2, FR3 and FR4 shown in Table 9 using Contact's numbering system. In another specific embodiment, the Specified herein provides an anti-MERTK antibody or antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or antigen-binding fragment comprises a framework region (FR) 1, variable light chain regions (VL) CDR1, FR2, VL CDR2, FR3, VL CDR3 and FR4, wherein VL CDR1, VL CDR2, and VL CDR3 comprise the amino acid sequences of VL CDR1, VL CDR2 and VL CDR3 shown in Table 7 using the IMGT numbering system, and FR1, FR2, FR3 and FR4 comprise the amino acid sequences of FR1, FR2, FR3 and FR4 shown in Table 9 using the IMGT numbering system.

[0166] In specific embodiments, this specification provides an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or its antigen-binding fragment comprises a variable light chain region (VL), the VL comprising a framework region (FR) 1, variable light chain region (VL) CDR1, FR2, VL CDR2, FR3, VL CDR3, and FR4, the VL CDR1, VL CDR2, and VL CDR3 comprising the amino acid sequences of VL CDR1, VL CDR2, and VL CDR3 shown in Table 7 using Exemplary's numbering system, and the FR1, FR2, FR3, and FR4 comprising the amino acid sequences of FR1, FR2, FR3, and FR4 shown in Table 9 using Exemplary's numbering system. In another specific embodiment, the Specified Information provides an anti-MERTK antibody or antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or antigen-binding fragment comprises a variable light chain region (VL), the VL comprising a framework region (FR) 1, variable light chain region (VL) CDR1, FR2, VL CDR2, FR3, VL CDR3, and FR4, the VL CDR1, VL CDR2, and VL CDR3 comprising the amino acid sequences of VL CDR1, VL CDR2, and VL CDR3 shown in Table 7 using the Kabat numbering system, and the FR1, FR2, FR3, and FR4 comprising the amino acid sequences of FR1, FR2, FR3, and FR4 shown in Table 9 using the Kabat numbering system.In another specific embodiment, the Specified herein provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or the antigen-binding fragment comprises a variable light chain region (VL), the VL comprising a framework region (FR) 1, variable light chain region (VL) CDR1, FR2, VL CDR2, FR3, VL CDR3, and FR4, the VL CDR1, VL CDR2, and VL CDR3 comprising the amino acid sequences of VL CDR1, VL CDR2, and VL CDR3 shown in Table 7 using Chothia's numbering system, and the FR1, FR2, FR3, and FR4 comprising the amino acid sequences of FR1, FR2, FR3, and FR4 shown in Table 9 using Chothia's numbering system. In another specific embodiment, the Specified Public Service provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or the antigen-binding fragment comprises a variable light chain region (VL), the VL comprising a framework region (FR) 1, variable light chain region (VL) CDR1, FR2, VL CDR2, FR3, VL CDR3, and FR4, the VL CDR1, VL CDR2, and VL CDR3 comprising the amino acid sequences of VL CDR1, VL CDR2, and VL CDR3 shown in Table 7 using the AbM numbering system, and the FR1, FR2, FR3, and FR4 comprising the amino acid sequences of FR1, FR2, FR3, and FR4 shown in Table 9 using the AbM numbering system.In another specific embodiment, the Specified herein provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or the antigen-binding fragment comprises a variable light chain region (VL), the VL comprising a framework region (FR) 1, variable light chain region (VL) CDR1, FR2, VL CDR2, FR3, VL CDR3 and FR4, the VL CDR1, VL CDR2, VL CDR3 comprising the amino acid sequences of VL CDR1, VL CDR2 and VL CDR3 shown in Table 7 using Contact's numbering system, and the FR1, FR2, FR3 and FR4 comprising the amino acid sequences of FR1, FR2, FR3 and FR4 shown in Table 9 using Contact's numbering system. In another specific embodiment, the Specified Information provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or antigen-binding fragment comprises a variable light chain region (VL), the VL comprising a framework region (FR) 1, variable light chain region (VL) CDR1, FR2, VL CDR2, FR3, VL CDR3, and FR4, the VL CDR1, VL CDR2, and VL CDR3 comprising the amino acid sequences of VL CDR1, VL CDR2, and VL CDR3 shown in Table 7 using the IGMT numbering system, and the FR1, FR2, FR3, and FR4 comprising the amino acid sequences of FR1, FR2, FR3, and FR4 shown in Table 9 using the IMGT numbering system.

[0167] In specific embodiments, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or its antigen-binding fragment comprises a variable heavy chain region (VH) and a variable light chain region (VL), where VH comprises VH framework region (FR) 1, VH CDR1, VH FR2, VH CDR2, VH FR3, VH CDR3 and VH FR4, and VL comprises VL FR1, VL CDR1, VL FR2, VL CDR2, VL FR3, VL CDR3 and VL FR4, where VH CDR1, VH CDR2, VH CDR3 comprises the amino acid sequences of VH CDR1, VH CDR2, and VH CDR3 shown in Table 1 using the Exemplary numbering system, and VH FR1, VH FR2, VH FR3 and VH The present invention provides an anti-MERTK antibody or its antigen-binding fragment, wherein FR4 comprises the amino acid sequences of VH FR1, VH FR2, VH FR3, and VH FR4 shown in Tables 2, 3, 4, or 5 using the Exemplary numbering system; VL CDR1, VL CDR2, and VL CDR3 comprises the amino acid sequences of VL CDR1, VL CDR2, and VL CDR3 shown in Table 7 using the Exemplary numbering system; and VL FR1, VL FR2, VL FR3, and VL FR4 comprises the amino acid sequences of VL FR1, VL FR2, VL FR3, and VL FR4 shown in Table 9 using the Exemplary numbering system.In another specific embodiment, as described herein, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or its antigen-binding fragment comprises a variable heavy chain region (VH) and a variable light chain region (VL), where VH comprises VH framework region (FR) 1, VH CDR1, VH FR2, VH CDR2, VH FR3, VH CDR3 and VH FR4, and VL comprises VL FR1, VL CDR1, VL FR2, VL CDR2, VL FR3, VL CDR3 and VL FR4, where VH CDR1, VH CDR2, VH CDR3 comprises the amino acid sequences of VH CDR1, VH CDR2, and VH CDR3 shown in Table 1 using the Kabat numbering system, and VH FR1, VH FR2, VH FR3 and VH The present invention provides an anti-MERTK antibody or its antigen-binding fragment, wherein FR4 comprises the amino acid sequences of VH FR1, VH FR2, VH FR3, and VH FR4 as shown in Tables 2, 3, 4, or 5 using the Kabat numbering system; VL CDR1, VL CDR2, and VL CDR3 comprises the amino acid sequences of VL CDR1, VL CDR2, and VL CDR3 as shown in Table 7 using the Kabat numbering system; and VL FR1, VL FR2, VL FR3, and VL FR4 comprises the amino acid sequences of VL FR1, VL FR2, VL FR3, and VL FR4 as shown in Table 9 using the Kabat numbering system.In another specific embodiment, as described herein, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or its antigen-binding fragment comprises a variable heavy chain region (VH) and a variable light chain region (VL), where VH comprises VH framework region (FR) 1, VH CDR1, VH FR2, VH CDR2, VH FR3, VH CDR3 and VH FR4, and VL comprises VL FR1, VL CDR1, VL FR2, VL CDR2, VL FR3, VL CDR3 and VL FR4, where VH CDR1, VH CDR2, VH CDR3 comprises the amino acid sequences of VH CDR1, VH CDR2, and VH CDR3 shown in Table 1 using Chothia's numbering system, and VH FR1, VH FR2, VH FR3 and VH The present invention provides an anti-MERTK antibody or its antigen-binding fragment, wherein FR4 comprises the amino acid sequences of VH FR1, VH FR2, VH FR3, and VH FR4 shown in Tables 2, 3, 4, or 5 using the Chothia numbering system; VL CDR1, VL CDR2, and VL CDR3 comprises the amino acid sequences of VL CDR1, VL CDR2, and VL CDR3 shown in Table 7 using the Chothia numbering system; and VL FR1, VL FR2, VL FR3, and VL FR4 comprises the amino acid sequences of VL FR1, VL FR2, VL FR3, and VL FR4 shown in Table 9 using the Chothia numbering system.In another specific embodiment, as described herein, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or its antigen-binding fragment comprises a variable heavy chain region (VH) and a variable light chain region (VL), where VH comprises VH framework region (FR)1, VH CDR1, VH FR2, VH CDR2, VH FR3, VH CDR3 and VH FR4, and VL comprises VL FR1, VL CDR1, VL FR2, VL CDR2, VL FR3, VL CDR3 and VL FR4, where VH CDR1, VH CDR2, VH CDR3 comprises the amino acid sequences of VH CDR1, VH CDR2, and VH CDR3 shown in Table 1 using the AbM numbering system, and VH FR1, VH FR2, VH FR3 and VH The present invention provides an anti-MERTK antibody or its antigen-binding fragment, wherein FR4 comprises the amino acid sequences of VH FR1, VH FR2, VH FR3, and VH FR4 shown in Tables 2, 3, 4, or 5 using the AbM numbering system; VL CDR1, VL CDR2, and VL CDR3 comprises the amino acid sequences of VL CDR1, VL CDR2, and VL CDR3 shown in Table 7 using the AbM numbering system; and VL FR1, VL FR2, VL FR3, and VL FR4 comprises the amino acid sequences of VL FR1, VL FR2, VL FR3, and VL FR4 shown in Table 9 using the AbM numbering system.In another specific embodiment, as described herein, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or its antigen-binding fragment comprises a variable heavy chain region (VH) and a variable light chain region (VL), where VH comprises VH framework region (FR) 1, VH CDR1, VH FR2, VH CDR2, VH FR3, VH CDR3 and VH FR4, and VL comprises VL FR1, VL CDR1, VL FR2, VL CDR2, VL FR3, VL CDR3 and VL FR4, where VH CDR1, VH CDR2, VH CDR3 comprises the amino acid sequences of VH CDR1, VH CDR2, and VH CDR3 shown in Table 1 using Contact's numbering system, and VH FR1, VH FR2, VH FR3 and VH The present invention provides an anti-MERTK antibody or its antigen-binding fragment, wherein FR4 comprises the amino acid sequences of VH FR1, VH FR2, VH FR3, and VH FR4 shown in Tables 2, 3, 4, or 5 using Contact's numbering system; VL CDR1, VL CDR2, and VL CDR3 comprises the amino acid sequences of VL CDR1, VL CDR2, and VL CDR3 shown in Table 7 using Contact's numbering system; and VL FR1, VL FR2, VL FR3, and VL FR4 comprises the amino acid sequences of VL FR1, VL FR2, VL FR3, and VL FR4 shown in Table 9 using Contact's numbering system.In another specific embodiment, as described herein, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or its antigen-binding fragment comprises a variable heavy chain region (VH) and a variable light chain region (VL), where VH comprises VH framework region (FR) 1, VH CDR1, VH FR2, VH CDR2, VH FR3, VH CDR3 and VH FR4, and VL comprises VL FR1, VL CDR1, VL FR2, VL CDR2, VL FR3, VL CDR3 and VL FR4, where VH CDR1, VH CDR2, VH CDR3 comprises the amino acid sequences of VH CDR1, VH CDR2, and VH CDR3 shown in Table 1 using the IMGT numbering system, and VH FR1, VH FR2, VH FR3 and VH The present invention provides an anti-MERTK antibody or its antigen-binding fragment, wherein FR4 comprises the amino acid sequences of VH FR1, VH FR2, VH FR3, and VH FR4 shown in Tables 2, 3, 4, or 5 using the IMGT numbering system; VL CDR1, VL CDR2, and VL CDR3 comprises the amino acid sequences of VL CDR1, VL CDR2, and VL CDR3 shown in Table 7 using the IMGT numbering system; and VL FR1, VL FR2, VL FR3, and VL FR4 comprises the amino acid sequences of VL FR1, VL FR2, VL FR3, and VL FR4 shown in Table 9 using the IMGT numbering system.

[0168] In another specific embodiment, as described herein, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or its antigen-binding fragment comprises a variable heavy chain region (VH) and a variable light chain region (VL), where VH comprises VH framework region (FR) 1, VH CDR1, VH FR2, VH CDR2, VH FR3, VH CDR3 and VH FR4, and VL comprises VL FR1, VL CDR1, VL FR2, VL CDR2, VL FR3, VL CDR3 and VL FR4, where VH CDR1, VH CDR2, VH CDR3 comprises the amino acid sequences of VH CDR1, VH CDR2, and VH CDR3 shown in Table 1 using a single numbering system, and VH FR1, VH FR2, VH FR3 and VH The present invention provides an anti-MERTK antibody or its antigen-binding fragment, wherein FR4 comprises the amino acid sequences of VH FR1, VH FR2, and VH FR4 shown in Tables 2, 3, or 4 using the same numbering system; VH FR3 comprises the amino acid sequence of VH FR3 shown in Table 5; VL CDR1, VL CDR2, and VL CDR3 comprises the amino acid sequences of VL CDR1, VL CDR2, and VL CDR3 shown in Table 7 using the same numbering system; and VL FR1, VL FR2, VL FR3, and VL FR4 comprises the amino acid sequences of VL FR1, VL FR2, VL FR3, and VL FR4 shown in Table 9 using the same numbering system.

[0169] In another specific embodiment, the Specified Information provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or the antigen-binding fragment comprises a variable heavy chain region (VH) and a variable light chain region (VL), where VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1, VH CDR2 comprises the amino acid sequence of SEQ ID NO: 6, VH CDR3 comprises the amino acid sequence of SEQ ID NO: 11, VL CDR1 comprises the amino acid sequence of SEQ ID NO: 68, VL CDR2 comprises the amino acid sequence of SEQ ID NO: 69, and VL CDR3 comprises the amino acid sequence of SEQ ID NO: 70.

[0170] In another specific embodiment, the Specified Information provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or the antigen-binding fragment comprises a variable heavy chain region (VH) and a variable light chain region (VL), where VH CDR1 comprises the amino acid sequence of SEQ ID NO: 2, VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7, VH CDR3 comprises the amino acid sequence of SEQ ID NO: 12, VL CDR1 comprises the amino acid sequence of SEQ ID NO: 71, VL CDR2 comprises the amino acid sequence of SEQ ID NO: 72, and VL CDR3 comprises the amino acid sequence of SEQ ID NO: 73.

[0171] In another specific embodiment, the Specified Information provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or the antigen-binding fragment comprises a variable heavy chain region (VH) and a variable light chain region (VL), where VH CDR1 comprises the amino acid sequence of SEQ ID NO: 3, VH CDR2 comprises the amino acid sequence of SEQ ID NO: 8, VH CDR3 comprises the amino acid sequence of SEQ ID NO: 11, VL CDR1 comprises the amino acid sequence of SEQ ID NO: 68, VL CDR2 comprises the amino acid sequence of SEQ ID NO: 69, and VL CDR3 comprises the amino acid sequence of SEQ ID NO: 70.

[0172] In another specific embodiment, the Specified Information provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or the antigen-binding fragment comprises a variable heavy chain region (VH) and a variable light chain region (VL), where VH CDR1 comprises the amino acid sequence of SEQ ID NO: 4, VH CDR2 comprises the amino acid sequence of SEQ ID NO: 9, VH CDR3 comprises the amino acid sequence of SEQ ID NO: 13, VL CDR1 comprises the amino acid sequence of SEQ ID NO: 74, VL CDR2 comprises the amino acid sequence of SEQ ID NO: 75, and VL CDR3 comprises the amino acid sequence of SEQ ID NO: 76.

[0173] In another specific embodiment, the Specified Information provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or the antigen-binding fragment comprises a variable heavy chain region (VH) and a variable light chain region (VL), where VH CDR1 comprises the amino acid sequence of SEQ ID NO: 5, VH CDR2 comprises the amino acid sequence of SEQ ID NO: 10, VH CDR3 comprises the amino acid sequence of SEQ ID NO: 14, VL CDR1 comprises the amino acid sequence of SEQ ID NO: 77, VL CDR2 comprises the amino acid sequence of SEQ ID NO: 72, and VL CDR3 comprises the amino acid sequence of SEQ ID NO: 70.

[0174] In another specific embodiment, the Specified Information provides an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or the antigen-binding fragment comprises a variable heavy chain region (VH) and a variable light chain region (VL), where VH CDR1 comprises the amino acid sequence of SEQ ID NO: 3, VH CDR2 comprises the amino acid sequence of SEQ ID NO: 6, VH CDR3 comprises the amino acid sequence of SEQ ID NO: 11, VL CDR1 comprises the amino acid sequence of SEQ ID NO: 68, VL CDR2 comprises the amino acid sequence of SEQ ID NO: 69, and VL CDR3 comprises the amino acid sequence of SEQ ID NO: 70.

[0175] In certain embodiments, the anti-MERTK antibody or its antigen-binding fragment described herein, which specifically binds to MERTK (e.g., human MERTK), includes a heavy chain variable region (VH) containing the amino acid sequence of SEQ ID NO: 105. In another specific embodiment, the anti-MERTK antibody or its antigen-binding fragment described herein, which specifically binds to MERTK (e.g., human MERTK), includes a heavy chain variable region (VH) containing the amino acid sequence of SEQ ID NO: 107. In yet another specific embodiment, the anti-MERTK antibody or its antigen-binding fragment described herein, which specifically binds to MERTK (e.g., human MERTK), includes a heavy chain variable region (VH) containing the amino acid sequence of SEQ ID NO: 108. In yet another specific embodiment, the humanized anti-MERTK or its antigen-binding fragment described herein, which specifically binds to MERTK (e.g., human MERTK), includes a heavy chain variable region (VH) containing the amino acid sequence of SEQ ID NO: 109.

[0176] In certain embodiments, the anti-MERTK antibody or antigen-binding fragment described herein that specifically binds to MERTK (e.g., human MERTK) includes a heavy chain variable region (VH) comprising the amino acid sequence of SEQ ID NO: 110.

[0177] In certain embodiments, the anti-MERTK antibody or its antigen-binding fragment described herein that specifically binds to MERTK (e.g., human MERTK) includes a variable light chain region (VL) containing the amino acid sequence of SEQ ID NO: 106. In other embodiments, the anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) includes a variable light chain region (VL) containing the amino acid sequence of SEQ ID NO: 111.

[0178] In a specific embodiment, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) comprises a heavy chain variable region (VH) and a light chain variable region (VL), where VH comprises the amino acid sequence of SEQ ID NO: 105 and VL comprises the amino acid sequence of SEQ ID NO: 106. In another specific embodiment, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) comprises a heavy chain variable region (VH) and a light chain variable region (VL), where VH comprises the amino acid sequence of SEQ ID NO: 107 and VL comprises the amino acid sequence of SEQ ID NO: 106. In yet another embodiment, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) comprises a heavy chain variable region (VH) and a light chain variable region (VL), where VH comprises the amino acid sequence of SEQ ID NO: 108 and VL comprises the amino acid sequence of SEQ ID NO: 106. In another embodiment, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) comprises a heavy chain variable region (VH) and a light chain variable region (VL), where VH comprises the amino acid sequence of SEQ ID NO: 110 and VL comprises the amino acid sequence of SEQ ID NO: 111.

[0179] In another specific embodiment, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) comprises a heavy chain variable region (VH) and a light chain variable region (VL), where VH comprises the amino acid sequence of SEQ ID NO: 109 and VL comprises the amino acid sequence of SEQ ID NO: 106 or 111.

[0180] In another specific embodiment, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) comprises a heavy chain variable region (VH) and a light chain variable region (VL), where VH comprises the amino acid sequence of VH shown in Table 11, and VL comprises the amino acid sequence of VL shown in Table 11. In another specific embodiment, a humanized anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) comprises a heavy chain variable region (VH) and a light chain variable region (VL), where VH comprises the amino acid sequence of VH shown in Table 12, and VL comprises the amino acid sequence of VL shown in Table 12. In yet another specific embodiment, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) comprises a heavy chain variable region (VH) and a light chain variable region (VL), where VH comprises the amino acid sequence of VH shown in Table 13, and VL comprises the amino acid sequence of VL shown in Table 13. In another specific embodiment, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) comprises a heavy chain variable region (VH) and a light chain variable region (VL), wherein VH comprises the amino acid sequence of VH shown in Table 14, and VL comprises the amino acid sequence of VL shown in Tables 11, 12, or 13.

[0181] In certain embodiments, the antibodies described herein may be described by their VH domain alone or their VL domain alone. For example, see Rader C et al., (1998) PNAS 95: 8910-8915, incorporated herein by reference in its entirety, which describes how humanization of a mouse anti-αvβ3 antibody by identifying complementary light or heavy chains from a library of human light or heavy chains results in a humanized antibody variant with high affinity or higher affinity than the original antibody. See also Clackson T et al., (1991) Nature 352: 624-628, incorporated herein by reference in its entirety, which describes a method for producing an antibody that binds to a specific antigen by using a specific VH domain (or VL domain) and screening a library for complementary variable domains. See also Kim SJ & Hong HJ, (2007) J Microbiol 45: 572-577, which describes a method for producing antibodies that bind to a specific antigen by using a specific VH domain to screen a library (e.g., a human VL library) for complementary VL domains, and then using the selected VL domains to lead to the selection of additional complementary (e.g., human) VH domains. This entire work is incorporated herein by reference.

[0182] As used herein, the terms “immunely binding,” “immunely recognizing,” “specifically binding,” and “specifically recognizing” are analogous terms in the context of antibodies, as understood by those skilled in the art, and refer to antibodies and their antigen-binding fragments that bind to an antigen (e.g., an epitope or immune complex) via an antigen-binding site, and do not exclude cross-reactivity between the antibody or antigen-binding fragment and other antigens. Whether or not immunospecific binding to MERTK (e.g., human MERTK) is maintained can be determined using any method known in the art.

[0183] In specific embodiments, this specification provides anti-MERTK antibodies or antigen-binding fragments thereof, comprising the heavy chain and / or light chain of an antibody, for example, the heavy chain alone, the light chain alone, or both the heavy chain and the light chain. With respect to the light chain, in specific embodiments, the light chain of the anti-MERTK antibody or antigen-binding fragment described herein is a kappa light chain. In another specific embodiment, the light chain of the anti-MERTK antibody or antigen-binding fragment described herein is a lambda light chain. In yet another specific embodiment, the light chain of the anti-MERTK antibody or antigen-binding fragment described herein is a human kappa light chain or a human lambda light chain.

[0184] In another specific embodiment, the anti-MERTK antibody or antigen-binding fragment described herein that specifically binds to MERTK (e.g., human MERTK) comprises a light chain, the light chain comprising the amino acid sequences of a variable light chain region (VL) and a kappa light chain constant region, the VL comprising the amino acid sequence of SEQ ID NO: 106 or SEQ ID NO: 111. Where used herein, the terms “constant region” or “constant domain” are interchangeable and have their common meanings in the art. The constant region is a portion of the antibody, for example, the carboxyl-terminal portion of the light and / or heavy chain, which does not directly participate in the binding of the antibody to the antigen but may exhibit various effector functions, such as interaction with the Fc receptor. The constant region of an immunoglobulin molecule generally has a more conserved amino acid sequence compared to the immunoglobulin variable domain.

[0185] In specific embodiments, the anti-MERTK antibody or antigen-binding fragment described herein that specifically binds to MERTK (e.g., human MERTK) comprises a light chain comprising a variable light chain region (VL) and a human kappa or lambda light chain constant region, wherein the VL comprises the amino acid sequence of SEQ ID NO: 106 or SEQ ID NO: 111. Non-limiting examples of human constant region sequences have been described in the Art; see, for example, Kabat EA et al., (1991).

[0186] In another specific embodiment, the anti-MERTK antibody or antigen-binding fragment described herein that specifically binds to MERTK (e.g., human MERTK) comprises a light chain, the light chain comprising the amino acid sequence of SEQ ID NO: 113 or SEQ ID NO: 118.

[0187] With respect to the heavy chain, in specific embodiments, the heavy chain of the anti-MERTK antibody or its antigen-binding fragment described herein may be a human alpha (α), delta (δ), epsilon (ε), gamma (γ), or mu (μ) heavy chain. In specific embodiments, the anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) includes a heavy chain, the heavy chain being a constant region or a part thereof as described herein or known in the art (e.g., C H 1, C H 2 or C H 3) or a combination thereof), and the variable heavy chain region (VH) of the anti-MERTK antibody described herein (e.g., SEQ ID NO: 105, SEQ ID NO: 107, SEQ ID NO: 108, 109 or SEQ ID NO: 110).

[0188] In certain embodiments, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) comprises a heavy chain, the heavy chain comprising the variable heavy chain region (VH) of the anti-MERTK antibody described herein (e.g., SEQ ID NOs: 105, 107, 108, 109, or 110), and the constant region or portion of the variable region of the human gamma heavy chain constant region (e.g., C H 1, C H 2 or C H 3) or a combination thereof. In specific embodiments, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) comprises the amino acid sequence of SEQ ID NO: 112, SEQ ID NO: 114, SEQ ID NO: 115, SEQ ID NO: 116, or SEQ ID NO: 117. Non-limiting examples of human constant region sequences have been described in the Art; see, for example, Kabat EA et al., (1991) above.

[0189] In specific embodiments, the anti-MERTK antibody described herein that specifically binds to MERTK (e.g., human MERTK) comprises (i) a heavy chain containing the amino acid sequence of SEQ ID NO: 112, and (ii) a light chain containing the amino acids of SEQ ID NO: 113. In specific embodiments, the anti-MERTK antibody described herein that specifically binds to MERTK (e.g., human MERTK) comprises (i) a heavy chain containing the amino acid sequence of SEQ ID NO: 114, and (ii) a light chain containing the amino acids of SEQ ID NO: 113. In specific embodiments, the anti-MERTK antibody described herein that specifically binds to MERTK (e.g., human MERTK) comprises (i) a heavy chain containing the amino acid sequence of SEQ ID NO: 115, and (ii) a light chain containing the amino acid sequence of SEQ ID NO: 113. In specific embodiments, the anti-MERTK antibody described herein that specifically binds to MERTK (e.g., human MERTK) comprises (i) a heavy chain containing the amino acid sequence of SEQ ID NO: 116, and (ii) a light chain containing the amino acids of SEQ ID NO: 113. In a specific embodiment, the anti-MERTK antibody described herein that specifically binds to MERTK (e.g., human MERTK) comprises (i) a heavy chain containing the amino acid sequence of SEQ ID NO: 117, and (ii) a light chain containing the amino acids of SEQ ID NO: 118.

[0190] In a specific embodiment, the anti-MERTK antibody or antigen-binding fragment described herein that immunospecifically binds to MERTK (e.g., human MERTK) comprises a heavy chain variable region (VH) and a light chain variable region (VL) containing any amino acid sequence described herein, and the constant region comprises the amino acid sequence of the constant region of a human IgG, IgE, IgM, IgD, IgA, or IgY immunoglobulin molecule. In another specific embodiment, the anti-MERTK antibody or antigen-binding fragment described herein that immunospecifically binds to MERTK (e.g., human MERTK) comprises VH and VL containing any amino acid sequence described herein, and the constant region comprises a human IgG, IgE, IgM, IgD, IgA, or IgY immunoglobulin molecule, any class of immunoglobulin molecule (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2), or any subclass (e.g., IgG 2a and IgG2b It includes the amino acid sequence of the steady region of (for example, those shown in Tables 11, 12, 13 or 15).

[0191] In certain embodiments, one, two or more mutations (for example, amino acid substitutions) are introduced into the Fc region of the anti-MERTK antibody or its antigen-binding fragment described herein to alter one or more functional properties of the antibody.

[0192] In some embodiments, one, two or more mutations (for example, amino acid substitutions) are introduced into the Fc region (for example, C numbered according to the Kabat numbering system (for example, Kabat's EU index)) of the anti-MERTK antibody or its antigen-binding fragment described herein. H 2 domain (residues 231-340 of human IgG1) and / or C H 3 domain (residues 341-447 of human IgG1) and / or hinge region) to increase or decrease the affinity of the antibody or its antigen-binding fragment for Fc receptors (for example, activating Fc receptors) on the surface of effector cells. Mutations in the Fc region of the anti-MERTK antibody or its antigen-binding fragment that decrease or increase the affinity of the antibody for Fc receptors, and techniques for introducing such mutations into Fc receptors or their fragments, are known to those of skill in the art. Examples of mutations in the Fc region of the anti-MERTK antibody or its antigen-binding fragment that can alter the affinity of the antibody or its antigen-binding fragment for Fc receptors are described, for example, in Smith P et al., (2012) PNAS 109: 6181-6186, US Patent No. 6,737,056, and International Publications WO02 / 060919; WO98 / 23289; and WO97 / 34631, which are incorporated herein by reference.

[0193] In some embodiments, the antibodies or antigen-binding fragments described herein include a glycosylation constant region. In some embodiments, the antibodies or antigen-binding fragments described herein include a non-glycosylation constant region. Antibodies having a reduced fucose content have been reported to have increased affinity for Fc receptors, such as FcγRIIIa. Therefore, in certain embodiments, the antibodies or antigen-binding fragments described herein may have a reduced fucose content or no fucose content at all. Such antibodies or antigen-binding fragments can be produced using techniques known to those skilled in the art. For example, antibodies or antigen-binding fragments can be expressed in cells that have insufficient or deficient fucosylation ability. In a specific example, antibodies having a reduced fucose content can be produced using a cell system having knockouts of both alleles of α1,6-fucosyltransferase. The Potelligent® system (Lonza) is an example of such a system that can be used to produce antibodies or antigen-binding fragments having a reduced fucose content. Alternatively, antibodies or antigen-binding fragments having reduced or no fucose content can be produced, for example, by (i) culturing cells under conditions that prevent or reduce fucosylation; (ii) post-translational removal of fucose (e.g., using a fucosidase enzyme); (iii) post-translational addition of a desired carbohydrate after recombinant expression of a non-glycosylated glycoprotein (e.g.); or (iv) purification of glycoproteins for selection of unfucosylated antibodies or their antigen-binding fragments. For methods for producing antibodies or their antigen-binding fragments having no or reduced fucose content, see, for example, Longmore GD & Schachter H (1982) Carbohydr Res 100: 365-92 and Imai-Nishiya H et al., (2007) BMC Biotechnol. 7: 84.

[0194] In another specific embodiment, the anti-MERTK antibody or antigen-binding fragment described herein that specifically binds to MERTK (e.g., human MERTK) comprises a heavy chain or a light chain, where (i) the heavy chain comprises a variable region comprising (a) the amino acid sequence of SEQ ID NO: 105, SEQ ID NO: 107, SEQ ID NO: 108, or SEQ ID NO: 109, and (b) a constant heavy chain domain comprising the amino acid sequence of the constant domain of human IgG; or (ii) the light chain comprises a variable region comprising (a) the amino acid sequence of SEQ ID NO: 106, and (b) a constant light chain domain comprising the amino acid sequence of the constant domain of a human light chain.

[0195] In another specific embodiment, the anti-MERTK antibody or antigen-binding fragment described herein that specifically binds to MERTK (e.g., human MERTK) comprises a heavy chain and a light chain, (i) the heavy chain comprising a variable region comprising (a) the amino acid sequence of SEQ ID NO: 105, SEQ ID NO: 107, SEQ ID NO: 108 or SEQ ID NO: 109, and (b) a constant heavy chain domain comprising the amino acid sequence of the constant domain of human IgG; and (ii) the light chain comprising a variable region comprising the amino acid sequence of SEQ ID NO: 106, and (b) a constant light chain domain comprising the amino acid sequence of the constant domain of human kappa light chain.

[0196] In another specific embodiment, the anti-MERTK antibody or antigen-binding fragment described herein that specifically binds to MERTK (e.g., human MERTK) comprises a heavy chain or a light chain, where (i) the heavy chain comprises a variable region comprising (a) the amino acid sequence of SEQ ID NO: 110 and (b) a constant heavy chain domain comprising the amino acid sequence of the constant domain of human IgG; or (ii) the heavy chain comprises a variable region comprising (a) the amino acid sequence of SEQ ID NO: 111 and (b) a constant light chain domain comprising the amino acid sequence of the constant domain of human kappa light chain. In another specific embodiment, the anti-MERTK antibody or antigen-binding fragment described herein that specifically binds to MERTK (e.g., human MERTK) comprises a heavy chain and a light chain, (i) the heavy chain comprising a variable region comprising (a) the amino acid sequence of SEQ ID NO: 110 and (b) a constant heavy chain domain comprising the amino acid sequence of the constant domain of human IgG; and (ii) the light chain comprising a variable region comprising (a) the amino acid sequence of SEQ ID NO: 111 and (b) a constant light chain domain comprising the amino acid sequence of the constant domain of human kappa light chain.

[0197] The determination of the percentage of identity between two sequences (e.g., amino acid sequences or nucleic acid sequences) can also be achieved using mathematical algorithms. A specific non-restrictive example of a mathematical algorithm used for comparing two sequences is the algorithm in Karlin S & Altschul SF (1990) PNAS 87: 2264-2268, modified in Karlin S & Altschul SF (1993) PNAS 90: 5873-5877. Such algorithms are incorporated into the NBLAST and XBLAST programs in Altschul SF et al., (1990) J Mol Biol 215: 403. A BLAST nucleotide search can be performed using the parameters of the NBLAST nucleotide program, for example, set to score=100 and word length=12, to obtain nucleotide sequences homologous to the nucleic acid molecules described herein. A BLAST protein search can be performed using, for example, the parameters of the XBLAST program set to score 50 and word length = 3 to obtain amino acid sequences homologous to the protein molecules described herein. To obtain gapped alignments for comparison purposes, gapped BLAST can be used as described in Altschul SF et al., (1997) Nuc Acids Res 25: 3389 3402. Alternatively, PSI BLAST can be used to perform iterative searches to detect distant relationships between molecules (as described above). When using the BLAST, gapped BLAST, and PSI Blast programs, the default parameters of each program (e.g., XBLAST and NBLAST) can be used (see, for example, the National Center for Biotechnology Information (NCBI) on the World Wide Web, ncbi.nlm.nih.gov). Another specific non-restrictive example of a mathematical algorithm used for sequence comparison is the algorithm in Myers and Miller, 1988, CABIOS 4:11 17. Such algorithms are incorporated into the ALIGN program (version 2.0), which is part of the GCG sequence alignment software package.When using the ALIGN program to compare amino acid sequences, the PAM120 weighted residual table, a gap length penalty of 12, and a gap penalty of 4 can be used.

[0198] The percentage of identity between two sequences can be determined using techniques similar to those described above, with or without gaps. Typically, only exact matches are counted in the calculation of the percentage of identity.

[0199] In certain embodiments, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) includes a VH having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of VH in SEQ ID NO: 105, SEQ ID NO: 107, SEQ ID NO: 108, or SEQ ID NO: 109. In some embodiments, an anti-MERTK antibody or its antigen-binding fragment includes a VH having at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of VH in SEQ ID NO: 110.

[0200] In certain embodiments, an anti-MERTK antibody or its antigen-binding fragment that immunospecifically binds to MERTK (e.g., human MERTK) includes a VL having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the VL of SEQ ID NO: 106. In certain embodiments, an anti-MERTK antibody or its antigen-binding fragment includes a VL having at least 99% sequence identity with the VH of SEQ ID NO: 111.

[0201] In a particular embodiment, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) comprises (i) a VH domain having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the amino acid sequence of the VH domain of SEQ ID NO: 105, SEQ ID NO: 107, SEQ ID NO: 108, or SEQ ID NO: 109; and (ii) a VL domain having at least 80%, at least 85%, at least 90%, at least 95%, or at least 98% sequence identity with the amino acid sequence of the VL domain of SEQ ID NO: 106.

[0202] In certain embodiments, the anti-MERTK antibody or antigen-binding fragment described herein that specifically binds to MERTK (e.g., human MERTK) comprises (i) a VH domain having at least 96%, at least 97%, at least 98%, or at least 99% sequence identity with the VH domain of SEQ ID NO: 110, and (ii) a VL domain having at least 99% sequence identity with the VL domain of SEQ ID NO: 111.

[0203] In another embodiment, the Antibodies herein provide antibodies that bind to the same or overlapping epitopes of MERTK (e.g., human MERTK). As used herein, “epitope” is a term used in the art and refers to a localized region of an antigen to which an antibody can specifically bind. An epitope may be, for example, an adjacent amino acid of a polypeptide (linear or adjacent epitope), or an epitope may be coupled with, for example, two or more non-adjacent regions of a polypeptide (conformal, nonlinear, discontinuous, or non-adjacent epitope). In certain embodiments, the epitopes of an antibody may be determined, for example, by NMR spectroscopy, X-ray diffraction crystallography studies, ELISA assays, hydrogen / deuterium exchange combined with mass spectrometry (e.g., MALDI mass spectrometry), array-based oligopeptide scanning assays, and / or mutagenesis mapping (e.g., site-directed mutagenesis mapping). Regarding X-ray crystallography, crystallization can be achieved using any known method in the art (e.g., Giege R et al., (1994) Acta Crystallogr D Biol Crystallogr 50(Pt 4): 339-350; McPherson A (1990) Eur J Biochem 189: 1-23; Chayen NE (1997) Structure 5: 1269-1274; McPherson A (1976) J Biol Chem 251: 6300-6303).Antibody: Antigen crystals may be studied using well-known X-ray diffraction techniques and refined using computer software such as X-PLOR (Yale University, 1992, distributed by Molecular Simulations, Inc.; see, for example, Meth Enzymol (1985) volumes 114 & 115, eds Wyckoff HW et al.; see U.S. Patent Application No. 2004 / 0014194), and BUSTER (Bricogne G (1993) Acta Crystallogr D Biol Crystallogr 49(Pt 1): 37-60; Bricogne G (1997) Meth Enzymol 276A: 361-423, eds Carter CW; Roversi P et al., (2000) Acta Crystallogr D Biol Crystallogr 56(Pt 10): 1316-1323). Mutagenesis mapping studies may be achieved using any method known to those skilled in the art. For example, see Champe M et al., (1995) and Cunningham BC & Wells JA (1989) for a description of mutagenesis techniques, including the alanine scanning mutagenesis technique. In addition, antibodies that recognize and bind to the same or overlapping epitopes of MERTK (e.g., human MERTK) can be identified using routine techniques such as immunoassays, for example, by demonstrating the ability of one antibody to block the binding of another antibody to its target antigen, i.e., by competitive binding assays. Competitive binding assays can also be used to determine whether two antibodies have similar binding specificity to an epitope. Competitive binding can be determined in assays in which the immunoglobulin under test inhibits the specific binding of a reference antibody to a common antigen such as MERTK.Many types of competitive binding assays exist, such as solid-phase direct or indirect radioimmunoassays (RIAs), solid-phase direct or indirect enzyme immunoassays (EIAs), sandwich competitive assays (see Stahli C et al., (1983) Methods Enzymol 9: 242-253); solid-phase direct biotin-avidin EIA (see Kirkland TN et al., (1986) J Immunol 137: 3614-9); solid-phase direct labeling assays, solid-phase direct labeling sandwich assays (see Harlow E & Lane D, (1988) Antibodies: A Laboratory Manual, Cold Spring Harbor Press); solid-phase direct labeling RIA using I-125 labeling (see Morel GA et al., (1988) Mol Immunol 25(1): 7-15); and solid-phase direct biotin-avidin EIA (see Cheung RC (1990) Virology 176: 546-52); and directly labeled RIA (Moldenhauer G et al., (1990) Scand J Immunol 32: 77-82) are known. Typically, such assays involve the use of purified antigen (e.g., MERTK (e.g., human MERTK)) or cells having either of these bound to a solid surface, unlabeled test immunoglobulin, and labeled reference immunoglobulin. Competitive inhibition can be measured by determining the amount of label bound to the solid surface or cells in the presence of test immunoglobulin. Typically, test immunoglobulin is present in excess. Typically, when competing antibodies are present in excess, they inhibit the specific binding of the reference antibody to the common antigen by at least 50-55%, 55-60%, 60-65%, 65-70%, 70-75%, or more. Competitive binding assays can consist of a number of different forms using either labeled antigen or labeled antibody. In a typical version of this assay, the antigen is immobilized in a 96-well plate. The ability of an unlabeled antibody to block the binding of a labeled antibody to the antigen is then measured, using radiolabeling or enzymatic labeling.For further details, see, for example, Wagener C et al., (1983) J Immunol 130: 2308-2315; Wagener C et al., (1984) J Immunol Methods 68: 269-274; Kuroki M et al., (1990) Cancer Res 50: 4872-4879; Kuroki M et al., (1992) Immunol Invest 21: 523-538; Kuroki M et al., (1992) Hybridoma 11: 391-407 and Antibodies: A Laboratory Manual, Ed Harlow E & Lane D editors, pp. 386-389.

[0204] In certain embodiments, when two antibodies recognize the same or sterically overlapping epitopes in a competitive binding assay, such as a competitive ELISA assay, which may consist of several different forms using either a labeled antigen or a labeled antibody, the competitive binding assay can be used to determine whether the antibody is competitively blocked, for example, in a dose-dependent manner, by another antibody, such as an antibody that binds to essentially the same or overlapping epitope as the reference antibody. In certain embodiments, the antibody can be tested in a competitive binding assay using the antibodies described herein (e.g., antibodies z10, z11, or z13).

[0205] In another embodiment, this specification provides antibodies that compete (e.g., in a dose-dependent manner) for the binding of MERTK (e.g., human MERTK) to an anti-MERTK antibody (e.g., z10, z11, z13, or xAb) as described herein, as determined using an assay known to those skilled in the art or described herein (e.g., an ELISA competitive assay or a surface plasmon resonance assay). In another embodiment, this specification provides antibodies that competitively inhibit (e.g., in a dose-dependent manner) the binding of an anti-MERTK antibody (e.g., z10, z11, z13, or xAb) as described herein to MERTK (e.g., human MERTK), as determined using an assay known to those skilled in the art or described herein (e.g., an ELISA competitive assay, or a suspension array, or a surface plasmon resonance assay).

[0206] In certain embodiments, this specification provides an anti-MERTK antibody that competes with the antibody described herein for binding to MERTK (e.g., human MERTK) to the same extent that the antibody described herein self-competes with MERTK (e.g., human MERTK) for binding to MERTK (e.g., human MERTK). In certain embodiments, this specification provides a first antibody that competes with the anti-MERTK antibody described herein for binding to MERTK (e.g., human MERTK), wherein the competition is greater than 80% (e.g., 85%, 90%, 95%, or 98%, or between 80% and 85%, 80% and 90%, 85% and 90%, or between 85% and 95%), and is indicated as a reduction in the binding of the first antibody to MERTK (e.g., human MERTK).

[0207] In a specific embodiment, this specification provides an anti-MERTK antibody that competes (for example, in a dose-dependent manner) with an antibody comprising a VH domain containing the amino acid sequence of SEQ ID NO: 105, SEQ ID NO: 107, SEQ ID NO: 108, SEQ ID NO: 109, or SEQ ID NO: 110, and a VL domain containing the amino acid sequence of SEQ ID NO: 106, or SEQ ID NO: 111, for specific binding to MERTK (e.g., human MERTK).

[0208] In a specific embodiment, an anti-MERTK antibody (e.g., a humanized antibody or a human antibody) or its antigen-binding fragment specifically binds to the same or overlapping epitopes of an antibody comprising a VH domain having the amino acid sequence of SEQ ID NO: 105 and a VL domain having the amino acid sequence of SEQ ID NO: 106. In another specific embodiment, the anti-MERTK antibody or its antigen-binding fragment described herein immunospecifically binds to the same or overlapping epitopes of an antibody comprising a VH domain having the amino acid sequence of SEQ ID NO: 107 and a VL domain having the amino acid sequence of SEQ ID NO: 106. In yet another specific embodiment, the anti-MERTK antibody or its antigen-binding fragment described herein immunospecifically binds to the same or overlapping epitopes of an antibody comprising a VH domain having the amino acid sequence of SEQ ID NO: 108 and a VL domain having the amino acid sequence of SEQ ID NO: 106. In yet another specific embodiment, the anti-MERTK antibody or its antigen-binding fragment described herein immunospecifically binds to the same or overlapping epitopes of an antibody comprising a VH domain having the amino acid sequence of SEQ ID NO: 110 and a VL domain having the amino acid sequence of SEQ ID NO: 111. It is possible to determine whether two antibodies bind to the same epitope using assays known to those skilled in the art or described herein (e.g., X-ray crystallography, ELISA assay, etc.). In specific embodiments, an antibody or its antigen-binding fragment that competes with the antibody described herein for binding to MERTK (e.g., human MERTK) is not a mouse antibody. In specific embodiments, an antibody or its antigen-binding fragment that binds to the same or overlapping epitopes of the antibody described herein (e.g., human MERTK) is not a mouse antibody.

[0209] Affinity is not limited, but the equilibrium dissociation constant (K D ) and equilibrium association constant (K A It can be measured and / or represented by several methods known in the art, including ). D This can be determined by techniques known to those skilled in the art, such as biolayer interference spectroscopy or surface plasmon resonance.

[0210] In certain embodiments, an anti-MERTK antibody (e.g., a humanized antibody) or its antigen-binding fragment that competes with the antibody described herein for binding to MERTK (e.g., human MERTK), or an anti-MERTK antibody or its antigen-binding fragment that binds to the same or overlapping epitopes of the anti-MERTK antibody described herein, has a K content of approximately 8 nM, 7 nM, 6 nM, 5 nM, 4.5 nM, 4 nM, 3.5 nM, 3 nM, 2.5 nM, 2 nM, 1.5 nM, 1 nM, 0.75 nM, 0.5 nM, 0.25 nM, or 0.1 nM. D Then, it binds to MERTK (e.g., human MERTK). In some embodiments, the anti-MERTK antibody or its antigen-binding fragment described herein, which competes with the antibody described herein for binding to MERTK (e.g., human MERTK), or the anti-MERTK antibody or its antigen-binding fragment which binds to the same or overlapping epitopes of the antibody described herein, is used in assays such as those described herein or known to those skilled in the art, with K concentrations of 0.1nM to 10nM, 0.5nM to 10nM, 1nM to 10nM, 2nM to 10nM, 2nM to 5nM, and 4nM to 10nM. D Then, it binds to MERTK (e.g., human MERTK). In some embodiments, an anti-MERTK antibody or its antigen-binding fragment described herein that competes with the antibody described herein for binding to MERTK (e.g., human MERTK), or an anti-MERTK antibody or its antigen-binding fragment that binds to the same or overlapping epitopes of the antibody described herein, is subjected to approximately 2 pM to 8 pM K using an assay described herein or known to those skilled in the art. D It binds to MERTK (e.g., human MERTK) via avidity.

[0211] In specific embodiments, the anti-MERTK antibody or its antigen-binding fragment described herein contains about 3 nM of K in assays described herein or known to those skilled in the art. DThen, it binds to MERTK (e.g., human MERTK). In some embodiments, the anti-MERTK antibody or its antigen-binding fragment described herein binds to about 1 pM to 15 pM K in assays described herein or known in the art. D Avidity binds to MERTK (e.g., human MERTK). In another specific embodiment, the anti-MERTK antibody or its antigen-binding fragment described herein binds to about 1.46 nM K in assays described herein or known to those skilled in the art. D Avidity binds to MERTK (e.g., human MERTK). In another specific embodiment, the anti-MERTK antibody or its antigen-binding fragment described herein binds to about 5.74 pM K in assays described herein or known to those skilled in the art. D Avidity binds to MERTK (e.g., human MERTK). In another specific embodiment, the anti-MERTK antibody or its antigen-binding fragment described herein binds to about 3.68 pM K in assays described herein or known to those skilled in the art. D Avidity binds to MERTK (e.g., human MERTK). In another specific embodiment, the anti-MERTK antibody or its antigen-binding fragment described herein binds to about 2.99 pM K in assays described herein or known to those skilled in the art. D Avidity is bound to MERTK (e.g., human MERTK). Where used herein, the term “approximately” indicates that deviations of 5% to 10% above and 5% to 10% below a value or range, when used to modify a number or range, remain within the intended meaning of the enumerated value or range.

[0212] In a specific embodiment, the anti-MERTK antibody or an antigen-binding fragment thereof binds to MERTK (e.g., human MERTK) with an EC50 of 1 nM to 20 nM, as evaluated by an assay described herein or known in the art. In another specific embodiment, the anti-MERTK antibody or an antigen-binding fragment thereof binds to MERTK (e.g., human MERTK) with an EC50 of 1 nM to 10 nM, as evaluated by an assay described herein or known in the art. In another specific embodiment, the anti-MERTK antibody or an antigen-binding fragment thereof binds to MERTK (e.g., human MERTK) with an EC50 of about 1 nM, about 2 nM, about 3 nM, about 4 nM, about 5 nM, about 6 nM, about 7 nM, about 8 nM, about 9 nM or about 10 nM, as evaluated by an assay described herein or known in the art.

[0213] In a specific embodiment, the anti-MERTK antibody or an antigen-binding fragment thereof is as described in Section 6 below. 5.1.2. Functional characteristics

[0214] In some embodiments, anti-MERTK antibodies or antigen-binding fragments described herein that specifically bind to MERTK (e.g., human MERTK) reduce the expression level of human MERTK in cancer cells (e.g., SKMEL5 melanoma cells) by more than 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or 85%, as known to those skilled in the art or as assessed by assays described herein. In some embodiments, the anti-MERTK antibody or its antigen-binding fragment described herein, which specifically binds to human MERTK, reduces the expression level of human MERTK in human M2 macrophages by more than 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or 85%, as known to those skilled in the art or as evaluated by assays described herein. In specific embodiments, the anti-MERTK antibody or its antigen-binding fragment binds to MERTK (e.g., human MERTK) but not to human Axl, human Tyro3, ​​or mouse MERTK, as evaluated by methods described herein or known in the art.

[0215] In some embodiments, an anti-MERTK antibody or its antigen-binding fragment binds to human MERTK and cynomolgus monkey MERTK.

[0216] In some embodiments, anti-MERTK antibodies or antigen-binding fragments described herein that specifically bind to MERTK (e.g., human MERTK) induce a cytokine secretion response by M2 macrophages, CD14+ monocytes, or both, as shown in Figure 10. In certain embodiments, anti-MERTK antibodies or antigen-binding fragments alter the expression of certain cytokines by M2 macrophages, CD14+ monocytes, or both, by more than 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or 85%, as known to those skilled in the art or as assessed by assays described herein (e.g., the antibody or antigen-binding fragment increases the expression of certain cytokines).

[0217] In some embodiments, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) blocks (e.g., partially or completely) the level of Gas-6-induced MERTK phosphorylation in human MERTK-expressing cells, such as cancer cells (e.g., melanoma cells), M2 macrophages, or both, in assays known to those skilled in the art. In specific embodiments, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) reduces the level of Gas-6-induced human MERTK phosphorylation in human MERTK-expressing cells, such as cancer cells (e.g., melanoma cells), M2 macrophages, or both, by at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or 85% in assays known to those skilled in the art.

[0218] In certain embodiments, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) induces degradation of human MERTK in human MERTK-expressing cells, such as cancer cells (e.g., melanoma cells), M2 macrophages, or both. In specific embodiments, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) reduces the level of human MERTK on the surface of human MERTK-expressing cells, such as cancer cells (e.g., melanoma cells), M2 macrophages, or both, by at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or 85% in assays described herein or known to those skilled in the art.

[0219] In certain embodiments, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) induces the internal translocation of human MERTK in human MERTK-expressing cells, such as cancer cells (e.g., melanoma cells), M2 macrophages, or both. In specific embodiments, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) increases the internal translocation of human MERTK by human MERTK-expressing cells, such as cancer cells (e.g., melanoma cells), M2 macrophages, or both, by at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or 85% in assays described herein or known to those skilled in the art.

[0220] In some embodiments, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) inhibits (e.g., partially or completely) colonization by cancer cells (e.g., melanoma cells) in assays described herein or known to those skilled in the art. In specific embodiments, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) reduces the level of colonization by cancer cells (e.g., melanoma cells) by at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or 85% in assays described herein or known to those skilled in the art. In some embodiments, the anti-MERTK antibody or antigen-binding fragment described herein that immunospecifically binds to MERTK (e.g., human MERTK) reduces the colony-forming ability of cancer cells (e.g., SKMEL5 melanoma cells) by more than 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or 85%, as evaluated by assays known to those skilled in the art or described herein.

[0221] In some embodiments, the anti-MERTK antibody or its antigen-binding fragment described herein, which specifically binds to MERTK (e.g., human MERTK), prevents Gas6-induced phosphorylation of AKT in assays described herein or known in the art. In certain embodiments, the anti-MERTK antibody or its antigen-binding fragment described herein, which specifically binds to MERTK (e.g., human MERTK), competes with Gas-6 (e.g., human Gas-6) for binding to human MERTK. In some embodiments, the anti-MERTK antibody or its antigen-binding fragment described herein also inhibits Gas-6 from binding to human MERTK (e.g., completely inhibiting or partially inhibiting). In some embodiments, the anti-MERTK antibody or its antigen-binding fragment described herein, which specifically binds to MERTK (e.g., human MERTK), inhibits the binding of Gas-6 (e.g., human or mouse Gas-6) to human MERTK by more than 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or 85%, as evaluated by assays known to those skilled in the art or described herein. In specific embodiments, the assay used to evaluate the inhibition of the binding of Gas-6 (e.g., human Gas-6) to human MERTK in the presence of the anti-MERTK antibody or its antigen-binding fragment described herein is an antibody capture ELISA known to those skilled in the art, such as that described in International Publication WO2016 / 106221 (e.g., Example 1 of International Publication WO2016 / 106221).

[0222] In some embodiments, an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) reduces the level of Gas-6-induced AKT phosphorylation in human MERTK-expressing cancer cells (e.g., melanoma cells) by at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or 85% in assays described herein or known to those skilled in the art.

[0223] In specific embodiments, anti-MERTK antibodies or antigen-binding fragments described herein that specifically bind to MERTK (e.g., human MERTK) inhibit angiogenesis within tumors. In some embodiments, the inhibition of angiogenesis is at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or 85%. In specific embodiments, the inhibition of angiogenesis is at least 50%, 55%, 60%, 65%, or 70%. The inhibition of angiogenesis can be evaluated by methods described herein and / or known to those skilled in the art. The inhibition can be compared to the level of angiogenesis without any antibody or with an unrelated antibody (e.g., an antibody that does not immune-specifically bind to MERTK).

[0224] In certain embodiments, anti-MERTK antibodies or antigen-binding fragments described herein that specifically bind to MERTK (e.g., human MERTK) inhibit tumor progression. The inhibition of tumor progression is at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or 85%. Tumor progression can be evaluated by methods known to those skilled in the art. Tumor progression can be compared to the cancer state with or without any antibody or with an unrelated antibody (e.g., an antibody that does not specifically bind to human MERTK). In specific embodiments, anti-MERTK antibodies or antigen-binding fragments have one, two, three, or more features of the antibodies described in Section 6 below. 5.1.3. Bispecific antibodies

[0225] In a specific embodiment, this specification provides a bispecific antibody comprising two different antigen-binding regions, wherein one binding region specifically binds to human MERTK, the other binding region binds to another target antigen, and the binding region that specifically binds to MERTK (e.g., human MERTK) is the anti-MERTK antibody or its antigen-binding fragment described herein. In a specific embodiment, this specification provides a bispecific antibody comprising two different antigen-binding regions, wherein one binding region specifically binds to MERTK (e.g., human MERTK), the other binding region binds to another target antigen, and the binding region that specifically binds to MERTK (e.g., human MERTK) comprises the variable heavy chain region of the anti-MERTK antibody or its antigen-binding fragment described herein. In another specific embodiment, this specification provides a bispecific antibody comprising two different antigen-binding regions, wherein one binding region specifically binds to MERTK (e.g., human MERTK), and the other binding region binds to another target antigen, and the binding region that specifically binds to MERTK (e.g., human MERTK) comprises the amino acid sequence of the variable heavy chain region of antibody z10 shown in Table 11. In yet another specific embodiment, this specification provides a bispecific antibody comprising two different binding regions, wherein one binding region specifically binds to MERTK (e.g., human MERTK), and the other binding region binds to another target antigen, and the binding region that specifically binds to MERTK (e.g., human MERTK) comprises the amino acid sequence of the variable heavy chain region of antibody z11 shown in Table 12. In another specific embodiment, this specification provides a bispecific antibody comprising two different antigen-binding regions, wherein one binding region specifically binds to MERTK (e.g., human MERTK), and the other binding region binds to another target antigen, and the binding region that specifically binds to MERTK (e.g., human MERTK) comprises the amino acid sequence of the variable heavy chain region of antibody z13 shown in Table 13.In another specific embodiment, this specification provides a bispecific antibody comprising two different binding regions, wherein one binding region specifically binds to MERTK (e.g., human MERTK), and the other binding region binds to another target antigen, and the binding region that specifically binds to MERTK (e.g., human MERTK) comprises a variable heavy chain region having the amino acid sequence shown in Table 14.

[0226] In another specific embodiment, this specification provides a bispecific antibody comprising two different antigen-binding regions, wherein one binding region specifically binds to MERTK (e.g., human MERTK), the other binding region binds to another target antigen, and the binding region that specifically binds to MERTK (e.g., human MERTK) comprises the variable light chain region of the anti-MERTK antibody or its antigen-binding fragment described herein. In another specific embodiment, this specification provides a bispecific antibody comprising two different antigen-binding regions, wherein one binding region specifically binds to MERTK (e.g., human MERTK), the other binding region binds to another target antigen, and the binding region that specifically binds to MERTK (e.g., human MERTK) comprises the amino acid sequence of the variable light chain region of antibody z10 shown in Table 11. In another specific embodiment, this specification provides a bispecific antibody comprising two different antigen-binding regions, wherein one binding region specifically binds to MERTK (e.g., human MERTK), and the other binding region binds to another target antigen, and the binding region that specifically binds to MERTK (e.g., human MERTK) comprises the amino acid sequence of the variable light chain region of antibody z11 shown in Table 12. In yet another specific embodiment, this specification provides a bispecific antibody comprising two different antigen-binding regions, wherein one binding region specifically binds to MERTK (e.g., human MERTK), and the other binding region binds to another target antigen, and the binding region that specifically binds to MERTK (e.g., human MERTK) comprises the amino acid sequence of the variable light chain region of antibody z13 shown in Table 13. In another specific embodiment, the Specified provides a bispecific antibody comprising two different binding regions, wherein one binding region specifically binds to MERTK (e.g., human MERTK), and the other binding region binds to another antigen of interest, and the binding region that specifically binds to MERTK (e.g., human MERTK) comprises a variable light chain region, the variable light chain region comprising the VL amino acid sequence shown in Tables 11, 12, or 13.

[0227] In another specific embodiment, this specification provides a bispecific antibody comprising two different antigen-binding regions, wherein one binding region specifically binds to MERTK (e.g., human MERTK), and the other binding region binds to another target antigen, and the binding region that specifically binds to MERTK (e.g., human MERTK) comprises the variable light chain region and variable heavy chain region of the anti-MERTK antibody or its antigen-binding fragment described herein. In another specific embodiment, this specification provides a bispecific antibody comprising two different antigen-binding regions, wherein one binding region specifically binds to MERTK (e.g., human MERTK), and the other binding region binds to another target antigen, and the binding region that specifically binds to MERTK (e.g., human MERTK) comprises the amino acid sequences of the variable light chain region and variable heavy chain region of antibody z10 shown in Table 11. In another specific embodiment, this specification provides a bispecific antibody comprising two different antigen-binding regions, wherein one binding region specifically binds to MERTK (e.g., human MERTK), and the other binding region binds to another target antigen, and the binding region that specifically binds to MERTK (e.g., human MERTK) comprises the amino acid sequences of the variable light chain region and variable heavy chain region of antibody z11 shown in Table 12. In another specific embodiment, this specification provides a bispecific antibody comprising two different antigen-binding regions, wherein one binding region specifically binds to MERTK (e.g., human MERTK), and the other binding region binds to another target antigen, and the binding region that specifically binds to MERTK (e.g., human MERTK) comprises the amino acid sequences of the variable light chain region and variable heavy chain region of antibody z13 shown in Table 13.In another specific embodiment, the Specified provides a bispecific antibody comprising two different antigen-binding regions, wherein one binding region specifically binds to MERTK (e.g., human MERTK), and the other binding region binds to another antigen of interest, and the binding region that specifically binds to MERTK (e.g., human MERTK) comprises a variable light chain region and a variable heavy chain region, the variable heavy chain region comprising the amino acid sequence shown in Table 14, and the variable light chain region comprising the VL amino acid sequence shown in Tables 11, 12, or 13.

[0228] In certain embodiments, the target antigen to which the other binding region of the bispecific antibody described herein binds is an antigen present on immune cells (e.g., T cells, NK cells, or dendritic cells). In some embodiments, the target antigen to which the other binding region of the bispecific antibody described herein binds is an immune checkpoint receptor (e.g., PD-1, PD-L1, CTLA-4, LAG3, Tim3, ICOS, CD40, GITR, or OX40). In specific embodiments, the other binding region of the bispecific antibody described herein includes an agonist anti-GITR antibody, an agonist anti-OX40 antibody, or an agonist anti-CD40 antibody. In specific embodiments, the other binding region of the bispecific antibody includes a blocking antibody or its antigen-binding fragment that binds to PD-1, PD-L1, CTLA-4, or LAG3. In certain embodiments, the target antigen to which the other binding region of the bispecific antibody described herein binds is a T cell receptor. In specific embodiments, the target antigen to which the other binding domain of the bispecific antibody described herein binds is CD3, PD-L1, LRP1, LRP8, TGF-β, NKGD2, or TIGIT. In some embodiments, the target antigen is a tumor-associated antigen.

[0229] In a specific embodiment, the other binding region of the bispecific antibody described herein is nivolumab or its antigen-binding fragment. In another specific embodiment, the other binding region of the bispecific antibody described herein is lambrolizumab or its antigen-binding fragment. In another specific embodiment, the other binding region of the bispecific antibody described herein is MEDI-4736 or its antigen-binding fragment. In another specific embodiment, the other binding region of the bispecific antibody described herein is ipilimumab or its antigen-binding fragment. In another specific embodiment, the other binding region of the bispecific antibody described herein is MPDL-3280A or its antigen-binding fragment. In another specific embodiment, the other binding region of the bispecific antibody described herein is pidilizumab or its antigen-binding fragment. In another specific embodiment, the other binding region of the bispecific antibody described herein is avelumab or its antigen-binding fragment. In another specific embodiment, the other binding region of the bispecific antibody described herein is pembrolizumab or its antigen-binding fragment. 5.2. Antibody-drug conjugates

[0230] This specification provides an antibody-drug conjugate comprising (a) an antibody moiety which is an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), (b) one or more drug moieties, each drug moiety which is a cytotoxic agent, and (c) optionally a linker, wherein the cytotoxic agent is directly conjugated to the antibody moiety or conjugated to the antibody moiety via the linker. As used in this disclosure, the term “conjugated” means covalent, which may be directly or via an intervening covalent structure. The antibody moiety of the antibody-drug conjugate provided herein may be an antibody as described in Section 5.1 or Section 6. The antibody-drug conjugate may be an antibody-drug conjugate as described in Section 6.

[0231] In certain embodiments, the antibody-drug conjugate provided herein has a molar ratio of antibody portion to drug portion between 1:1 and 1:20. In specific embodiments, the antibody-drug conjugate provided herein has a molar ratio of antibody portion to drug portion between 1:1 and 1:15. In specific embodiments, the antibody-drug conjugate provided herein has a molar ratio of antibody portion to drug portion between 1:1 and 1:12. In specific embodiments, the antibody-drug conjugate provided herein has a molar ratio of antibody portion to drug portion between 1:1 and 1:8. In preferred embodiments, the antibody-drug conjugate provided herein has a molar ratio of antibody portion to drug portion between 1:3 and 1:5. In specific embodiments, the antibody-drug conjugate provided herein has a molar ratio of antibody portion to drug portion of 1:3. In another specific embodiment, the antibody-drug conjugate provided herein has a molar ratio of antibody portion to drug portion of 1:4. In another specific embodiment, the antibody-drug conjugate provided herein has a molar ratio of antibody portion to drug portion of 1:5. In yet another specific embodiment, the antibody-drug conjugate provided herein has a molar ratio of antibody portion to drug portion of 1:9.

[0232] The drug portion is conjugated to one or more chains of the antibody portion. In some embodiments, the drug portion is conjugated to one chain of the antibody portion (for example, when the antibody portion is an scFv, or when the antibody portion is a multichain antibody such as an immunoglobulin (a tetramer) or its antigen-binding fragment). In other embodiments, the drug portion is conjugated to two or more chains of the antibody portion (when the antibody portion is a multichain antibody such as an immunoglobulin or its antigen-binding fragment). In specific embodiments, the drug portion is conjugated to two identical chains of the immunoglobulin, for example, the heavy chain or the light chain. In other embodiments, the drug portion is conjugated to all chains of the antibody portion (when the antibody portion is a multichain antibody such as an immunoglobulin or its antigen-binding fragment). In some embodiments, the drug portion is conjugated to the antibody by the method described in Section 6.4 below.

[0233] In specific embodiments, the drug moiety is conjugated to one or more sites in the constant region of the antibody. In specific embodiments, the drug moiety is conjugated to one or more sites in the variable region of the antibody. In specific embodiments, the drug moiety is conjugated to one or more lysine residues in the antibody. In specific embodiments, the drug moiety is conjugated to one or more cysteine ​​residues in the antibody, for example, if MMAE or SN-38 is the drug moiety. In specific embodiments, the drug moiety is conjugated to the Fc region of the antibody, which is an immunoglobulin. In specific embodiments, the drug moiety is a peptide or protein fused (directly or via a linker, which is a peptide or protein) to the N-terminus or C-terminus of one or more chains of the antibody moiety.

[0234] For example, if the antibody portion is an scFv, the drug portion may be fused to the N or C terminus of the scFv (directly or via a linker that is a peptide or protein). In a specific embodiment, if the antibody portion is a multichain antibody or its antigen-binding fragment, the drug portion may be fused to one of the chains of the antibody portion (directly or via a linker that is a peptide or protein). In the case of an antibody that is an immunoglobulin, in a specific embodiment, both heavy chains may be fused to the drug portion (directly or via a linker that is a peptide or protein), and / or both light chains may be fused to the drug portion (directly or via a linker that is a peptide or protein). In such specific embodiments, this specification provides a vector (e.g., an expression vector) comprising a polynucleotide sequence encoding a fusion protein consisting of a drug portion and an antibody portion or their chains, and a linker (if present) between the drug portion and the antibody portion. This specification also provides ex vivo host cells comprising such a vector for recombinant expression of a fusion protein consisting of a drug portion and an antibody portion or their chains, and a linker (if present) between the drug portion and the antibody portion. The characteristics of such vectors and ex vivo host cells, and the methods for producing them, may be the same as those described below for anti-MERTK antibodies. Furthermore, this specification provides a method for producing a fusion protein comprising a drug portion and an antibody portion or their chains, and a linker (if present) between the drug portion and the antibody portion, comprising the step of culturing such ex vivo host cells under conditions such that a polynucleotide containing a nucleotide sequence encoding the fusion protein is expressed by the ex vivo host cells to produce the fusion protein. The antibody-drug conjugates described herein may be produced by methods known in the art or by methods described in Section 6 below.

[0235] In some embodiments, the antibody-drug conjugate has the structure shown in Figure 13A or Figure 13B, particularly in Figure 13A. Figure 13A shows the structure of MMAE linked to the antibody (Ab) via an mc-vc-PABC linker. Figure 13B shows the structure of SN-38 linked to the antibody (Ab) via a CL2 or CL2A linker. When "AA" in Figure 13B is phenylalanine-lysine, Figure 13B shows the antibody linked to SN-38 via a CL2 linker (containing a cathepsin B cleavage site). When "AA" in Figure 13B is lysine, Figure 13B shows the antibody linked to SN-38 via a CL2A linker (not containing a cathepsin B cleavage site).

[0236] In a specific embodiment, the antibody portion of the antibody-drug conjugate includes a variable region of antibody z10. In a specific embodiment, the antibody-drug conjugate provided herein includes an antibody portion comprising a heavy chain variable region (VH) and a light chain variable region (VL), wherein VH comprises the amino acid sequence of sequence number 105, and VL comprises the amino acid sequence of sequence number 106, and the antibody portion is conjugated to MMAE via an mc-vc-PABC linker. In another specific embodiment, the antibody-drug conjugate provided herein includes an antibody portion comprising a heavy chain variable region (VH) and a light chain variable region (VL), wherein VH comprises the amino acid sequence of sequence number 105, and VL comprises the amino acid sequence of sequence number 106, and the antibody portion is conjugated to SN-38 via a CL2A linker.

[0237] In alternative embodiments of the antibody-drug conjugates described throughout this disclosure, the drug portion is not necessarily a cytotoxic agent. For example, the drug portion may be any drug known in the art that is suitable for use, but may include, but is not limited to, those described in Section 5.2.3 below, or may be an imaging agent, for example, for the purpose of diagnosing or monitoring a disease.

[0238] In preferred embodiments of the antibody-drug conjugates described throughout this disclosure, the drug portion is conjugated to an antibody (also referred to herein as the “antibody portion”) via a linker. 5.2.1. Functional characteristics of antibody-drug conjugates

[0239] In certain embodiments, the antibody-drug conjugate described herein is approximately 8 nM, 7 nM, 6 nM, 5 nM, 4.5 nM, 4 nM, 3.5 nM, 3 nM, 2.5 nM, 2 nM, 1.5 nM, 1 nM, 0.75 nM, 0.5 nM, 0.25 nM, 0.17 nM, 0.1 nM, 0.081 nM, or 0.021 nM of K D The antibody-drug conjugate then binds to MERTK (e.g., human MERTK). In some embodiments, the antibody-drug conjugate provided herein reduces the expression level of human MERTK in cancer cells (e.g., SKMEL5 melanoma cells) by more than 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or 85%, as described in Section 6 below, as evaluated by assays known to those skilled in the art or described herein. In specific embodiments, the antibody-drug conjugate provided herein binds to MERTK (e.g., binds to human MERTK but not to human Axl, human Tyro3, ​​or mouse MERTK), as evaluated by methods described herein or known in the art. In certain embodiments, the antibody-drug conjugate provided herein includes an antibody moiety, and the antibody-drug conjugate binds to MERTK with higher affinity than the unconjugated antibody moiety.

[0240] In certain embodiments, the antibody-drug conjugate provided herein induces cell death in MERTK (e.g., human MERTK) expressing cells (e.g., M2 macrophages, cancer cells, or both) in vitro. In some specific embodiments, at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or 85% of MERTK expressing cells cultured for a certain period in the presence of the antibody-drug conjugate described herein die at the end of the period, as evaluated by methods known to those skilled in the art for measuring cell death in vitro. In other specific embodiments, the percentage of MERTK-expressing cells that have undergone cell death (e.g., M2 macrophages, cancer cells, or both) is at least 50%, 100%, 2x, 3x, 5x, 10x, 20x, 50x, 100x, 200x, 500x, or 1000x higher when cultured for a certain period in the presence of the antibody-drug conjugate described herein compared to when cultured without the antibody-drug conjugate (e.g., cultured without any antibody-drug conjugate, or with an antibody-drug conjugate containing an unrelated antibody (e.g., an antibody that does not immune-specifically bind to MERTK (e.g., human MERTK)), or with an unconjugated anti-MERTK antibody or unconjugated anti-MERTK antigen-binding fragment described herein). The period may be, for example, about 1 hour, 2 hours, 4 hours, 8 hours, 12 hours, 16 hours, 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, or 1 week.

[0241] In some embodiments, the antibody-drug conjugates provided herein do not affect the viability of MERTK-expressing macrophages, as can be determined by methods known to those skilled in the art for measuring cell death in vitro, or by the methods described herein, for example, in Section 6 below. In some embodiments, the antibody-drug conjugates provided herein do not affect the viability of M1 macrophages, as can be determined by methods known to those skilled in the art for measuring cell death in vitro, or by the methods described herein, for example, in Section 6 below. In some embodiments, the antibody-drug conjugates provided herein do not affect the viability of M2 macrophages, as can be determined by methods known to those skilled in the art for measuring cell death in vitro, or by the methods described herein, for example, in Section 6 below.

[0242] In certain embodiments, the antibody-drug conjugate provided herein induces the degradation of MERTK in MERTK-expressing cells, such as cancer cells (e.g., melanoma cells). In specific embodiments, the antibody-drug conjugate provided herein reduces the level of human MERTK on the surface of human MERTK-expressing cells, such as cancer cells (e.g., melanoma cells), by at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or 85% in assays described herein or known to those skilled in the art. In certain embodiments, the antibody-drug conjugate provided herein induces a higher degradation of MERTK in MERTK-expressing cells than the unconjugated antibody portion, as evaluated in assays described herein or known in the art.

[0243] In certain embodiments, the antibody-drug conjugate provided herein induces internal translocation of MERTK in MERTK-expressing cells, such as cancer cells (e.g., melanoma cells). In specific embodiments, the antibody-drug conjugate provided herein increases the internal translocation of human MERTK by human MERTK-expressing cells, such as cancer cells (e.g., melanoma cells), by at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or 85% in assays described herein or known to those skilled in the art. In certain embodiments, the antibody-drug conjugate provided herein comprises an antibody portion, and the antibody-drug induces higher internal translocation of MERTK-expressing cells than the unconjugated antibody portion, as evaluated by assays described herein or known to those skilled in the art.

[0244] In certain embodiments, the antibody-drug conjugates provided herein inhibit tumor progression. The inhibition of tumor progression is at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or 85%. Tumor progression can be evaluated by methods known to those skilled in the art. Tumor progression can be compared to the state of cancer without treatment with any antibody, treated with an unrelated antibody (e.g., an antibody that does not specifically bind to human MERTK), treated with SN-38, treated with MMAE, or treated with the unconjugated antibody portion of the antibody-drug conjugate. In specific embodiments, the anti-MERTK antibody or its antigen-binding fragment has one, two, three or more features of the antibodies described in Section 6 below.

[0245] In specific embodiments, the antibody-drug conjugates described herein inhibit angiogenesis within tumors. In some embodiments, the inhibition of angiogenesis is at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or 85%. In specific embodiments, the inhibition of angiogenesis is at least 50%, 55%, 60%, 65%, or 70%. The inhibition of angiogenesis can be evaluated by methods described herein and / or known to those skilled in the art. The inhibition can be compared to the level of angiogenesis without an antibody-drug conjugate (e.g., cultured without any antibody-drug conjugate, or with an antibody-drug conjugate containing an unrelated antibody (e.g., an antibody that does not specifically bind to MERTK (e.g., human MERTK)), or with an unconjugated anti-MERTK antibody or unconjugated anti-MERTK antigen-binding fragment as described herein).

[0246] In certain embodiments, the antibody-drug conjugates described herein inhibit the progression of a tumor (e.g., a tumor of human breast cancer cells). The inhibition of tumor progression is at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or 85%. Tumor progression can be evaluated by methods known to those skilled in the art. Tumor progression can be compared to the state of cancer without an antibody-drug conjugate (e.g., cultured without any antibody-drug conjugate, or cultured with an antibody-drug conjugate containing an unrelated antibody (e.g., an antibody that does not specifically bind to MERTK (e.g., human MERTK)), or cultured with an unconjugated anti-MERTK antibody or unconjugated anti-MERTK antigen-binding fragment described herein). In specific embodiments, the assay used to evaluate tumor progression is a mouse tumor transplantation model. 5.2.2. Antibody part

[0247] The antibody-drug conjugates provided herein may include any anti-MERTK antibody provided herein. See, for example, Section 5.1.1 for an example of the antibody portion of an antibody-drug conjugate provided herein. In a specific embodiment, the antibody portion is an immunoglobulin, for example, an immunoglobulin or humanized immunoglobulin containing a human constant domain. In another specific embodiment, the antibody portion is, for example, an scFv containing the VH and VL domains of the antibody described in Section 5.1.1 of this specification.

[0248] In some embodiments, the antibody-drug conjugates provided herein include a drug portion conjugated via a linker to an antibody portion containing VH, for example, the VH described in Section 5.1.1. In some embodiments, the antibody-drug conjugates provided herein include a drug portion conjugated via a linker to an antibody portion containing VL, for example, the VL described in Section 5.1.1. In some embodiments, the antibody-drug conjugates provided herein include a drug portion conjugated via a linker to an antibody portion containing VH and VL of an antibody disclosed herein, for example, as described in Section 5.1.1.

[0249] In some embodiments, the antibody-drug conjugate provided herein comprises a drug moiety optionally conjugated via a linker to an antibody moiety comprising a VH containing the amino acid sequence of SEQ ID NO: 105. In some embodiments, the antibody-drug conjugate provided herein comprises a drug moiety optionally conjugated via a linker to an antibody moiety comprising a VL containing the amino acid sequence of SEQ ID NO: 106. In some embodiments, the antibody-drug conjugate provided herein comprises (i) a VH containing the amino acid sequence of SEQ ID NO: 105, and (ii) a drug moiety optionally conjugated via a linker to an antibody moiety comprising a VL containing the amino acid sequence of SEQ ID NO: 106.

[0250] In some embodiments, the antibody-drug conjugate provided herein comprises an MMAE conjugated via a linker to an antibody portion containing VH, wherein VH comprises the amino acid sequence of SEQ ID NO: 105. In some embodiments, the antibody-drug conjugate provided herein comprises an MMAE conjugated via a linker to an antibody portion containing VL, wherein VL comprises the amino acid sequence of SEQ ID NO: 106. In some embodiments, the antibody-drug conjugate provided herein comprises an SN-38 conjugated via a linker to an antibody portion containing VH, wherein VH comprises the amino acid sequence of SEQ ID NO: 105. In some embodiments, the antibody-drug conjugate provided herein comprises an SN-38 conjugated via a linker to an antibody portion containing VL, wherein VL comprises the amino acid sequence of SEQ ID NO: 106.

[0251] In a specific embodiment, the antibody-drug conjugate provided herein comprises MMAE conjugated via a linker to an antibody portion comprising VH and VL, wherein VH comprises the amino acid sequence of SEQ ID NO: 105 and VL comprises the amino acid sequence of SEQ ID NO: 106. In another specific embodiment, the antibody-drug conjugate provided herein comprises SN-38 conjugated via a linker to an antibody portion comprising VH and VL, wherein VH comprises the amino acid sequence of SEQ ID NO: 105 and VL comprises the amino acid sequence of SEQ ID NO: 106.

[0252] In some embodiments, the antibody-drug conjugate provided herein includes a drug portion conjugated via a linker to an antibody portion containing a heavy chain, for example, the heavy chain described in Section 5.1.1. In some embodiments, the antibody-drug conjugate provided herein includes MMAE conjugated via a linker to an antibody portion containing a heavy chain, for example, the heavy chain described in Section 5.1.1. In some embodiments, the antibody-drug conjugate provided herein includes SN-38 conjugated via a linker to an antibody portion containing a heavy chain, for example, the heavy chain described in Section 5.1.1. In some embodiments, the antibody-drug conjugate provided herein includes a drug portion conjugated via a linker to an antibody portion containing a light chain, for example, the light chain described in Section 5.1.1. In some embodiments, the antibody-drug conjugate provided herein includes MMAE conjugated via a linker to an antibody portion containing a light chain, for example, the light chain described in Section 5.1.1. In some embodiments, the antibody-drug conjugate provided herein includes an SN-38 conjugated via a linker to an antibody portion containing a light chain, for example, the light chain described in Section 5.1.1. In some embodiments, the antibody-drug conjugate provided herein includes (i) a heavy chain, and (ii) a drug portion conjugated via a linker to an antibody portion containing a light chain, for example, the heavy chain and light chain described in Section 5.1.1. In some embodiments, the antibody-drug conjugate provided herein includes (i) a heavy chain, and (ii) an MMAE conjugated via a linker to an antibody portion containing a light chain, for example, the heavy chain and light chain described in Section 5.1.1. In some embodiments, the antibody-drug conjugate provided herein includes (i) a heavy chain, and (ii) a light chain, for example, an SN-38 conjugated via a linker to an antibody portion containing a heavy chain and light chain described in Section 5.1.1.

[0253] In some embodiments, the antibody-drug conjugate provided herein includes a drug moiety conjugated via a linker to an antibody moiety comprising a heavy chain containing the amino acid sequence of SEQ ID NO: 112. In some specific embodiments, the antibody-drug conjugate provided herein includes MMAE conjugated via a linker to an antibody moiety comprising a heavy chain containing the amino acid sequence of SEQ ID NO: 112. In some specific embodiments, the antibody-drug conjugate provided herein includes SN-38 conjugated via a linker to an antibody moiety comprising a heavy chain containing the amino acid sequence of SEQ ID NO: 112. In some embodiments, the antibody-drug conjugate provided herein includes a drug moiety conjugated via a linker to an antibody moiety comprising a light chain containing the amino acids of SEQ ID NO: 113. In some specific embodiments, the antibody-drug conjugate provided herein includes MMAE conjugated via a linker to a light chain containing the amino acids of SEQ ID NO: 113. In some specific embodiments, the antibody-drug conjugate provided herein includes SN-38 conjugated via a linker to a light chain containing the amino acids of SEQ ID NO: 113. In some embodiments, the antibody-drug conjugate provided herein comprises a drug moiety conjugated via a linker to an antibody moiety comprising (i) a heavy chain comprising the amino acid sequence of SEQ ID NO: 112 and (ii) a light chain comprising the amino acids of SEQ ID NO: 113. In a specific embodiment, the antibody-drug conjugate provided herein comprises MMAE conjugated via a linker to an antibody moiety comprising (i) a heavy chain comprising the amino acid sequence of SEQ ID NO: 112 and (ii) a light chain comprising the amino acids of SEQ ID NO: 113. In another specific embodiment, the antibody-drug conjugate provided herein comprises SN-38 conjugated via a linker to an antibody moiety comprising (i) a heavy chain comprising the amino acid sequence of SEQ ID NO: 112 and (ii) a light chain comprising the amino acids of SEQ ID NO: 113. 5.2.3. Drug portion and linker

[0254] The cytotoxic agent used in antibody-drug conjugates is often also called the payload or warhead. Most cytotoxic agents used in antibody-drug conjugates target DNA or microtubules and have high cytotoxic efficacy (approximately 10%). -10 ~10 -12 M IC 50 (See Beck A et al., (2017) Nat Rev Drug Discov 16: 315-337). However, less potent cytotoxic agents may be used. Cytotoxic agents may be small molecules, nucleotides, peptides, or non-antibody proteins. In a specific embodiment, the cytotoxic agent is a small molecule. In another specific embodiment, the cytotoxic agent is a non-antibody protein. Non-limiting exemplary cytotoxic agents that can be used in antibody-drug conjugates according to the present invention are described in Beck A et al., (2017) Nat Rev Drug Discov 16: 315-337; Peters C and Brown S, (2015) Biosci Rep 35: art:e00225; McCombs JR and Owen SC (2015) The AAPS Journal 17: 339-351; Jackson DY (2016) Org Process Res Dev 20: 852-866; and Olivier KJ and Hurvitz SA ed., (2016) Antibody-Drug Conjugates: Fundamentals, Drug Development, and Clinical, Wiley.

[0255] In certain embodiments, the cytotoxic agent is one of the drugs listed in Table 30 or 31. In specific embodiments, the cytotoxic agents used in the antibody-drug conjugate are auristatin (such as monomethyl auristatin E (MMAE), monomethyl auristatin F (MMAF), Aur0101, PF06380101, auristatin W, or auristatin F) or its derivatives, maytansinoids (such as DM1 or DM4), pyrrolobenzodiazepines (PBD) (such as SGD1882 or SG3199), indolinobenzodiazepines (such as DGN462 or DGN549), calicheamicin (ozogamicin) (such as CM1), and camptothecin analogs (SN-38, DX These include -8951f or DX-8951f derivatives, duocalmycin (seco-duocalmycin-hydroxy-benzamide-azaindole (seco-DUBA), minor groove-binding alkylating agent (MGBA), or MED-2460, etc.), tubulin inhibitors (such as cryptophycin), tubulicin or tubulicin analogs (such as AZ13599185), amberstatin 269, doxorubicin, antibiotics (such as Refalog), anthracyclines (such as PNU-159682), microtubule inhibitors (such as rhizoxin), splicesostatin, or tylanstatin. In a specific embodiment, the cytotoxic agent used in the antibody-drug conjugate is MMAE or MMAF. In a specific embodiment, the cytotoxic agent used in the antibody-drug conjugate is MMAE (see, for example, Figure 13A). In another specific embodiment, the cytotoxic agent used in the antibody-drug conjugate is DM1 or DM4. In another specific embodiment, the cytotoxic agent used in the antibody-drug conjugate is SN-38. In another specific embodiment, the cytotoxic agent used in the antibody-drug conjugate is SN-38, as shown in Figure 13B. [Table 30-1] [Table 30-2] [Table 30-3] [Table 30-4] [Table 31-1] [Table 31-2]

[0256] As used herein, the term “small molecule” refers to an organic or inorganic compound (which may be a heteroorganic compound or an organometallic compound) having a molecular weight of less than about 10,000 grams per mole. In specific embodiments, the small molecule has a molecular weight of less than about 5,000 grams per mole, or less than about 2,000 grams per mole, or less than about 1,000 grams per mole, or less than about 500 grams per mole, or less than about 100 grams per mole.

[0257] In some embodiments, the cytotoxic agent is directly conjugated to the antibody moiety. In other embodiments, the cytotoxic agent is conjugated to the antibody moiety via a linker. A suitable linker used in the present invention is preferably stable in the bloodstream to limit off-target toxicity and unstable at the cancer site to allow for the release of the cytotoxic agent (see Peters C and Brown S, (2015) Biosci Rep 35: art:e00225). Non-limiting exemplary linkers that can be used in accordance with the present invention are described in Beck A et al., (2017) Nat Rev Drug Discov 16: 315-337; Peters C and Brown S, (2015) Biosci Rep 35: art:e00225; McCombs JR and Owen SC (2015) The AAPS Journal 17: 339-351; Jackson DY (2016) Org Process Res Dev 20: 852-866; and Olivier KJ and Hurvitz SA ed., (2016) Antibody-Drug Conjugates: Fundamentals, Drug Development, and Clinical, Wiley. In some embodiments, the linker is a cleavable linker having a motif that is sensitive to lysosomal proteases (e.g., cathepsin B), a motif that is sensitive to acidic pH (e.g., hydrazone), or a motif that contains a disulfide crosslink that can be reduced by glutathione. In other embodiments, the linker is a non-cleavable linker. Examples, but not limited to, include small molecules, nucleotides, peptides, or non-antibody proteins. In specific embodiments, the linker is a small molecule.In specific embodiments, the linker used in the antibody-drug conjugate is cathepsin B, hydrazone, succinimidyl-4-(N-maleimidomethyl)cyclohexane-1-carboxylate (SMCC), maleimidocaproic acid (mc), valine-citrulline (vc), N-hydroxysuccinimidyl-4-(2-pyridyldithio)-2-sulfobutanoate (sulfoSPDB), N-hydroxysuccinimidyl-4-(2-pyridydithio)butanoate (SPDB), N-succinimidyl-4-(2-pyridyldithio)pentanoate (SPP), valine-alanine (va), polyethylene glycol 8-valine-citrulline (PEG8-va), mb-vc, CL2A, cleavable vc-based linker, or fleximer polymer linker. In specific embodiments, the linker selected for use in the antibody-drug conjugate described herein does not interfere with the binding of the antibody moiety to the antigen, cytotoxic agent, or both. In specific embodiments, the linker selected for use in the antibody-drug conjugate described herein is maleimidocaproyl-valine-citrulline-p-aminobenzyloxycarbonyl (mc-vc-PABC). In other specific embodiments, the linker selected for use in the antibody-drug conjugate described herein is CL2. In other specific embodiments, the linker selected for use in the antibody-drug conjugate described herein is CL2A. In some specific embodiments, the linker selected for use in the antibody-drug conjugate described herein is the linker shown in Figure 13A. In other specific embodiments, the linker selected for use in the antibody-drug conjugate described herein is the linker shown in Figure 13B, where AA is lysine. In other specific embodiments, the linker selected for use in the antibody-drug conjugate described herein is the linker shown in Figure 13B, where AA is phenylalanine-lysine.In a specific embodiment, the antibody-drug conjugate includes a linker, which is maleimidocaproyl-valine-citrulline-p-aminobenzyloxycarbonyl (also known as "mc-vc-PABC") (see Figure 13A for the structure of mc-vc-PABC). In another specific embodiment, the antibody-drug conjugate includes a linker, which is CL2 (see Figure 13B and Cardillo et al. 2011, Clin Cancer Res; 17(10); 3157-69 for the structure of CL2). In another specific embodiment, the antibody-drug conjugate includes a linker, which is CL2A (see Figure 13B and Goldberg et al. 2018, Oncotarget; 9(48); 28989-29006, and Cardillo et al. 2011, Clin Cancer Res; 17(10); 3157-69 for the structure of CL2A).

[0258] In some embodiments, the linker selected for use in the antibody-drug conjugate described herein comprises one or more cleavage sites. In specific embodiments, the linker selected for use in the antibody-drug conjugate described herein contains a cathepsin B protease site. In other specific embodiments, the linker selected for use in the antibody-drug conjugate described herein contains a cathepsin B protease site and a pH-dependent cleavage site.

[0259] Non-limiting exemplary linker-drug moiety pairs that can be used in the antibody-drug conjugate described in the present invention are described in Beck A et al., (2017) Nat Rev Drug Discov 16: 315-337; Peters C and Brown S, (2015) Biosci Rep 35: art:e00225; McCombs JR and Owen SC (2015) The AAPS Journal 17: 339-351; Jackson DY (2016) Org Process Res Dev 20: 852-866; and Olivier KJ and Hurvitz SA ed., (2016) Antibody-Drug Conjugates: Fundamentals, Drug Development, and Clinical, Wiley. In specific embodiments, the linker-drug moiety pair in the antibody-drug conjugate is a peptide linker having vc-MMAE, mc-MMAF, SMCC-DM1, sulfoSPDB-DM4, SPDB-DM4, SPP-DM1, va-SGD1882, polyethylene glycol 8(PEG8)-va-SG3199, sulfoSPDB-DGN462, hydrazone-CM1, vc-seco-DUBA, mb-vc-MGBA, CL2A-SN38, DX-8951 derivative, cleavable vc-based linker having hydrazone-doxorubicin, Aur0101, vc-PF06380101, a fleximer polymer linker having auristatin F, cleavable linker-tubulin inhibitor, or vc-reflag. In specific embodiments, the linker-drug moiety pair in the antibody-drug conjugate is mc-vc-PABC-MMAE (as shown in Figure 13A). In other specific embodiments, the linker-drug moiety pair in the antibody-drug conjugate is CL2-SN-38 (as shown in Figure 13B, where AA is phenylalanine-lysine). In other specific embodiments, the linker-drug moiety pair in the antibody-drug conjugate is CL2A-SN-38 (as shown in Figure 13B, where AA is lysine).

[0260] In specific embodiments, the drug portion of the antibody-drug conjugate described herein does not interfere with the binding of the antibody portion to its antigen (e.g., MERTK (e.g., human MERTK)). 5.3. Antibody Production 5.3.1. Antibody Production and Screening

[0261] In another embodiment, this specification provides a method for producing an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK).

[0262] The antibodies or antigen-binding fragments described herein can be produced by any method known in the art for antibody synthesis, for example, by chemical synthesis or by recombinant expression techniques. Unless otherwise indicated, the methods described herein utilize conventional techniques in the relevant fields within the scope of the art, including molecular biology, microbiology, genetic analysis, recombinant DNA, organic chemistry, biochemistry, PCR, oligonucleotide synthesis and modification, nucleic acid hybridization, and other fields within the scope of the art. These techniques are described and fully explained in the references cited herein, for example.For example, see Maniatis T et al., (1982) Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press; Sambrook J et al., (1989), Molecular Cloning: A Laboratory Manual, Second Edition, Cold Spring Harbor Laboratory Press; Sambrook J et al., (2001) Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY; Ausubel FM et al., Current Protocols in Molecular Biology, John Wiley & Sons (1987 and annual updates); Current Protocols in Immunology, John Wiley & Sons (1987 and annual updates) Gait (ed.) (1984) Oligonucleotide Synthesis: A Practical Approach, IRL Press; Eckstein (ed.) (1991) Oligonucleotides and Analogues: A Practical Approach, IRL Press; Birren B et al., (eds.) (1999) Genome Analysis: A Laboratory Manual, Cold Spring Harbor Laboratory Press.

[0263] In specific embodiments, the antibodies described herein are antibodies (e.g., recombinant antibodies) prepared, expressed, produced, or isolated by any means including, for example, DNA sequence synthesis, gene manipulation, or production. In certain embodiments, such antibodies include sequences encoded by DNA sequences that are not naturally present in the in vivo antibody germline repertoire of animals or mammals (e.g., humans). In specific embodiments, the antibodies described herein are produced by a method comprising the step of using human MERTK (SEQ ID NO: 131) or its extracellular domain (SEQ ID NO: 132) as an immunogen.

[0264] In certain embodiments, this specification provides a method for producing an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK), comprising the step of culturing cells or host cells as described herein. In certain embodiments, this specification provides a method for producing an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK), comprising the step of expressing the antibody or its antigen-binding fragment using cells or host cells as described herein (e.g., cells or host cells containing a polynucleotide encoding the antibody as described herein). In certain embodiments, the cells are isolated cells. In certain embodiments, an exogenous polynucleotide is introduced into the cells. In certain embodiments, the method further comprises the step of purifying the antibody or its antigen-binding fragment obtained from the cells or host cells.

[0265] Methods for producing polyclonal antibodies are known in the art (see, for example, Chapter 11 in: Short Protocols in Molecular Biology, (2002) 5th Ed., Ausubel FM et al., eds., John Wiley and Sons, New York).

[0266] The term "monoclonal antibody," as used herein, is not limited to antibodies produced by hybridoma technology. Monoclonal antibodies can be prepared using a wide variety of techniques known in the art, including the use of hybridoma, recombinant and phage display technologies, or combinations thereof. For example, monoclonal antibodies can be produced recombinantly from host cells exogenously expressing the antibodies or fragments thereof described herein, e.g., the light and / or heavy chains of such antibodies. Methods for preparing clonal cell lines and the monoclonal antibodies expressed thereby are well known in the art (see, for example, Chapter 11 in Short Protocols in Molecular Biology, (2002) 5th Ed., Ausubel FM et al. above). For example, monoclonal antibodies can be produced using techniques known in the art, and hybridoma techniques, including, for example, Harlow E & Lane D, Antibodies: A Laboratory Manual, (Cold Spring Harbor Laboratory Press, 2nd ed. 1988); Hammerling GJ et al., in: Monoclonal Antibodies and T-Cell Hybridomas 563 681 (Elsevier, NY, 1981); and Kohler G & Milstein C (1975) Nature 256: 495. Methods for producing and screening specific antibodies using hybridoma techniques are commonplace and well-known in the art. In some embodiments, mice (or other animals such as rats, monkeys, donkeys, pigs, sheep, hamsters, or dogs) can be immunized with an antigen (e.g., human), and once an immune response is detected, for example, once antigen-specific antibodies are detected in mouse serum, the mouse spleen is harvested and spleen cells are isolated.Next, spleen cells are fused with any suitable myeloma cells, for example, cells from cell line SP2 / 0 available from the American Cell Culture and Preservation Center (ATCC®) (Manassas, VA), using known techniques to form hybridomas. The hybridomas are selected and cloned by limiting dilution.

[0267] The hybridoma cells thus prepared are preferably seeded and grown in a suitable culture medium containing one or more substances that inhibit the growth or survival of unfused parent myeloma cells. The culture medium in which the hybridoma cells are growing is assayed for the production of monoclonal antibodies against MERTK (e.g., human MERTK). After identifying hybridoma cells that produce antibodies with desired specificity, affinity, and / or activity, clones can be subcloned, grown, and isolated from the culture medium by standard methods (Goding JW (Ed), Monoclonal Antibodies: Principles and Practice, above). The binding specificity of the monoclonal antibodies produced by the hybridoma cells is determined by methods known in the art, such as immunoprecipitation, or by in vitro binding assays such as radioimmunoassay (RIA) or enzyme-linked immunosorbent assay (ELISA).

[0268] In specific embodiments, this specification discloses monoclonal antibodies produced by a single cell (e.g., a hybridoma or host cell producing recombinant antibodies) which bind immunospecifically to MERTK (e.g., human MERTK) as determined, for example, by an ELISA known in the art or by an ELISA or other antigen-binding or competitive binding assay described in Section 6 of this Spec. In certain embodiments, the monoclonal antibody is a monovalent or polyvalent (e.g., bivalent) antibody. In certain embodiments, the monoclonal antibody is a monospecific antibody or a multispecific antibody (e.g., bispecific antibody).

[0269] Antibody fragments that recognize MERTK (e.g., human MERTK) can be produced by any technique known to those skilled in the art. For example, the Fab and F(ab')2 fragments described herein can be produced by protein cleavage of an immunoglobulin molecule using an enzyme such as papain (which produces the Fab fragment) or pepsin (which produces the F(ab')2 fragment). The Fab fragment corresponds to one of the two identical arms of the antibody molecule and contains a complete light chain paired with the VH and CH1 domains of the heavy chain. The F(ab')2 fragment contains two antigen-binding arms of the antibody molecule linked by a disulfide bond in the hinge region.

[0270] Furthermore, the anti-MERTK antibodies or their antigen-binding fragments described herein can also be generated using various phage display methods known in the art. In phage display methods, a functional antibody domain is displayed on the surface of a phage particle carrying the encoding polynucleotide sequence. In particular, the DNA sequences encoding the VH and VL domains are amplified from an animal cDNA library (e.g., a human or mouse cDNA library of affected tissue). The DNA encoding the VH and VL domains is recombined with an scFv linker by PCR and cloned into a phagemide vector. The vector is electroporated into E. coli cells, and E. coli infects with helper phages. The phages used in these methods are typically filamentous phages containing fd and M13, and the VH and VL domains are usually recombinantly fused to either gene III or gene VIII of the phage. Phages expressing antigen-binding domains that bind to a specific antigen can be selected or identified by antigen, for example, using a labeled antigen or an antigen bound to or captured on a solid surface or beads.Examples of phage display methods that can be used to produce the antibodies described herein include: Brinkman U et al., (1995) J Immunol Methods 182: 41-50; Ames RS et al., (1995) J Immunol Methods 184: 177-186; Kettleborough CA et al., (1994) Eur J Immunol 24: 952-958; Persic L et al., (1997) Gene 187: 9-18; Burton DR & Barbas CF (1994) Advan Immunol 57: 191-280; PCT application No. PCT / GB91 / 001134; International publications No. WO90 / 02809, WO91 / 10737, WO92 / 01047, 92 / 18619, WO93 / 11236, WO95 / 15982, WO95 / 20401 and WO97 / 13844; and U.S. Patents No. 5,698,426, 5,223, Examples include those disclosed in Nos. 409, 5,403,484, 5,580,717, 5,427,908, 5,750,753, 5,821,047, 5,571,698, 5,427,908, 5,516,637, 5,780,225, 5,658,727, 5,733,743 and 5,969,108.

[0271] As described in the above references, after phage selection, the antibody-coding region from the phage is isolated and used to generate a whole antibody containing a humanized antibody, a chimeric antibody, or any other desired antigen-binding fragment, which can then be expressed in any desired host, including mammalian cells, insect cells, plant cells, yeast, and bacteria, as described below. Techniques for recombinantly producing antibody fragments such as Fab, Fab', and F(ab')2 fragments can also be used, employing methods known in the art, such as those disclosed in PCT Publication WO92 / 22324; Mullinax RL et al., (1992) BioTechniques 12(6): 864-9; Sawai H et al., (1995) Am J Reprod Immunol 34: 26-34; and Better M et al., (1988) Science 240: 1041-1043.

[0272] In one embodiment, the VH or VL sequence can be amplified from a template, e.g., an scFv clone, using PCR primers containing a VH or VL nucleotide sequence, a restriction site, and a flanking sequence protecting the restriction site, in order to generate the entire antibody. Using cloning techniques known to those skilled in the art, the PCR-amplified VH domain can be cloned into a vector expressing the VH constant region, and the PCR-amplified VL domain can be cloned into a vector expressing the VL constant region, e.g., a human kappa or lambda constant region. The VH and VL domains can also be cloned into a single vector expressing the required constant regions. The heavy-chain and light-chain conversion vectors are then co-transfected into cell lines using techniques known to those skilled in the art to generate stable or transient cell lines expressing the full-length antibody, e.g., IgG.

[0273] Chimeric antibodies are molecules in which different parts of the antibody are derived from different immunoglobulin molecules. For example, a chimeric antibody may contain the variable region of a mouse or rat monoclonal antibody fused to the constant region of a human antibody. Methods for producing chimeric antibodies are known in the art. See, for example, Morrison SL (1985) Science 229: 1202-7; Oi VT & Morrison SL (1986) BioTechniques 4: 214-221; Gillies SD et al., (1989) J Immunol Methods 125: 191-202; and U.S. Patents 5,807,715, 4,816,567, 4,816,397 and 6,331,415. In a specific embodiment, the chimeric antibody comprises a variable heavy chain region (VH) and a variable light chain region (VL), where VH and VL each contain the amino acid sequences shown in Table 13.

[0274] A humanized antibody comprises a framework region capable of binding to a given antigen and substantially having the amino acid sequence of a human immunoglobulin, and a CDR substantially having the amino acid sequence of a non-human immunoglobulin (e.g., mouse immunoglobulin). In certain embodiments, the humanized antibody also comprises at least a portion of the immunoglobulin constant region (Fc), typically that of a human immunoglobulin. The antibody may also comprise the CH1, hinge, CH2, CH3, and CH4 regions of the heavy chain. The humanized antibody can be selected from any class of immunoglobulin, including IgM, IgG, IgD, IgA, and IgE, as well as any isotype, including IgG1, IgG2, IgG3, and IgG4.Humanized antibodies are used in, but are not limited to, CDR grafting (European Patent No. EP239400, International Publication No. WO91 / 09967; and U.S. Patents Nos. 5,225,539, 5,530,101 and 5,585,089), veneering, or resurfacing (European Patent Nos. EP592106 and EP519596; Padlan EA (1991) Mol Immunol 28(4 / 5): 489-498; Studnicka GM et al., (1994) Prot Engineering 7(6): 805-814; and Roguska MA et al., (1994) PNAS 91: 969-973), chain shuffling (U.S. Pat. No. 5,565,332), as well as e.g. U.S. Pat. No. 6,407,213, U.S. Pat. (2000) Protein Eng. 13(5): 353-60;Morea V et al., (2000) Methods 20(3): 267-79;Baca M et al., (1997) J Biol Chem 272(16): 10678-84;Roguska MA et al., (1996) Protein Eng 9(10): 895 904;Couto JR et al., (1995) It can be produced using a variety of techniques known in the art, including the techniques disclosed in Cancer Res. 55 (23 Supp): 5973s-5977s; Couto JR et al., (1995) Cancer Res 55(8): 1717-22; Sandhu JS (1994) Gene 150(2): 409-10 and Pedersen JT et al., (1994) J Mol Biol 235(3): 959-73. See also U.S. Patent Application Publication No. US2005 / 0042664(A1) (February 24, 2005), which is incorporated herein by reference in its entirety.

[0275] In specific embodiments, a humanized antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK) includes VH and VL of antibody z10 (see Table 11), VH and VL of antibody z11 (see Table 12), or VH and VL of antibody z13 (see Table 13). Methods for producing multispecific antibodies (e.g., bispecific antibodies) are described, see, for example, U.S. Patents 7,951,917; 7,183,076; 8,227,577; 5,837,242; 5,989,830; 5,869,620; 6,132,992 and 8,586,713.

[0276] Single-domain antibodies, such as antibodies lacking a light chain, can be produced by methods well known in the art. See Riechmann L & Muyldermans S (1999) J Immunol 231: 25-38; Nuttall SD et al., (2000) Curr Pharm Biotechnol 1(3): 253-263; Muyldermans S, (2001) J Biotechnol 74(4): 277-302; U.S. Patent No. 6,005,079; and International Publications WO94 / 04678, WO94 / 25591, and WO01 / 44301.

[0277] Furthermore, using antibodies that immunospecifically bind to the MERTK antigen (e.g., human MERTK), anti-idiotype antibodies that "mimic" the antigen can be generated using techniques well known to those skilled in the art. (See, for example, Greenspan NS & Bona CA (1989) FASEB J 7(5): 437-444; and Nissinoff A (1991) J Immunol 147(8): 2429-2438).

[0278] In certain embodiments, the anti-MERTK antibody or its antigen-binding fragment described herein binds to the same or overlapping epitopes of MERTK (e.g., human MERTK) as the anti-MERTK antibody or its antigen-binding fragment described herein. In certain embodiments, the antibody described herein competitively blocks any one of the antibodies described herein (e.g., z10, z11, or z13, or xAb) from binding to MERTK (e.g., human MERTK) (e.g., in a dose-dependent manner) is a human anti-MERTK antibody or its antigen-binding fragment. Human antibodies can be produced using any method known in the art. For example, transgenic mice that cannot express functional endogenous immunoglobulins but can express human immunoglobulin genes can be used. In particular, complexes of human heavy-chain and light-chain immunoglobulin genes can be introduced into mouse embryonic stem cells by random or homologous recombination. Alternatively, human variable regions, constant regions, and diversity regions can be introduced into mouse embryonic stem cells in addition to human heavy-chain and light-chain genes. Mouse heavy and light chain immunoglobulin genes can be rendered nonfunctional, either separately from or simultaneously with the introduction of human immunoglobulin loci by homologous recombination. In particular, J HHomozygous deletion of the region prevents endogenous antibody production. Modified embryonic stem cells are enlarged and microinjected into blastocysts to produce chimeric mice. The chimeric mice are then bred to produce homozygous offspring expressing human antibodies. Transgenic mice are immunized in the usual manner with all or part of a selected antigen, e.g., the antigen (e.g., MERTK). Monoclonal antibodies against the antigen can be obtained from the immunized transgenic mice using conventional hybridoma techniques. The human immunoglobulin transgenes carried by the transgenic mice are rearranged during B cell differentiation and subsequently undergo class switching and somatic mutation. Therefore, it is possible to produce therapeutically useful IgG, IgA, IgM, and IgE antibodies using such techniques. For an overview of this technique for producing human antibodies, see, for example, Lonberg N & Huszar D (1995) Int Rev Immunol 13:65-93. For a detailed discussion of this technique for producing human antibodies and human monoclonal antibodies, and of protocols for producing such antibodies, see, for example, International Publications WO98 / 24893, WO96 / 34096 and WO96 / 33735; and U.S. Patents 5,413,923, 5,625,126, 5,633,425, 5,569,825, 5,661,016, 5,545,806, 5,814,318 and 5,939,598. Examples of mice capable of producing human antibodies include Xenomouse® (Abgenix, Inc.; U.S. Patent Nos. 6,075,181 and 6,150,184), HuAb-Mouse® (Medarex, Inc. / Gen Pharm; U.S. Patent Nos. 5,545,806 and 5,569,825), Trans Chromo Mouse® (Kirin), and KM Mouse® (Medarex / Kirin).

[0279] Human antibodies that specifically bind to MERTK (e.g., human MERTK) can be prepared using antibody libraries derived from human immunoglobulin sequences by various methods known in the art, including the phage display method described above. See also U.S. Patents 4,444,887, 4,716,111 and 5,885,793; and International Publications WO98 / 46645, WO98 / 50433, WO98 / 24893, WO98 / 16654, WO96 / 34096, WO96 / 33735 and WO91 / 10741.

[0280] In some embodiments, human antibodies can be produced using mouse-human hybridomas. For example, human peripheral blood lymphocytes transformed with Epstein-Barr virus (EBV) can be fused with mouse myeloma cells to produce mouse-human hybridomas that secrete human monoclonal antibodies. These mouse-human hybridomas can then be screened to determine which secrete human monoclonal antibodies that immunospecifically bind to a target antigen (e.g., human MERTK). Such methods are known and described in the art; see, for example, Shinmoto H et al., (2004) Cytotechnology 46: 19-23; Naganawa Y et al., (2005) Human Antibodies 14: 27-31.

[0281] In a specific embodiment, a method for screening and selecting an antibody or antigen-binding fragment described herein that specifically binds to MERTK (e.g., human MERTK) is as described in Example 1 below.

[0282] Once the anti-MERTK antibody or its antigen-binding fragment described herein is produced, it can be purified by any method known in the art for the purification of immunoglobulin molecules, for example, by chromatography (e.g., by ion exchange, affinity, particularly affinity to specific antigens after protein A, and size column chromatography), centrifugation, differential solubility, or any other standard technique for protein purification. Furthermore, the antibodies described herein can be fused to heterologous polypeptide sequences described herein or otherwise known in the art to facilitate purification.

[0283] In specific embodiments, the anti-MERTK antibody or its antigen-binding fragment described herein is isolated or purified. Generally, an isolated antibody is an antibody that substantially does not contain other antibodies having different antigen specificity from the isolated antibody. For example, in certain embodiments, the antibody preparation described herein is substantially free of cellular material and / or chemical precursors. The phrase “substantially free of cellular material” includes antibody preparations isolated from cellular components of cells produced by isolation or recombination. Thus, an antibody that is substantially free of cellular material includes antibody preparations and / or antibody variants having less than about 30%, 20%, 10%, 5%, 2%, 1%, 0.5%, or 0.1% (dry weight) of heterogeneous proteins (also referred to herein as “contamination proteins”), such as different post-translational modified forms of the antibody or other different versions of the antibody (e.g., antibody fragments). When antibodies are produced recombinantly, they generally contain substantially no culture medium; i.e., the culture medium accounts for less than about 20%, 10%, 2%, 1%, 0.5%, or 0.1% of the volume of the protein preparation. When antibodies are produced by chemosynthesis, they generally contain substantially no chemical precursors or other chemicals; i.e., they are separated from chemical precursors or other chemicals involved in protein synthesis. Thus, such preparations of antibodies contain less than about 30%, 20%, 10%, or 5% (by dry weight) of chemical precursors or compounds other than the antibody of interest. In specific embodiments, the antibodies described herein are isolated or purified. 5.3.2. Polynucleotides

[0284] In certain embodiments, this specification provides polynucleotides comprising nucleotide sequences encoding the anti-MERTK antibody described herein or its antigen-binding fragments that specifically bind to the MERTK (e.g., human MERTK) antigen, and vectors, for example, vectors containing such polynucleotides for their highly efficient expression in host cells (e.g., E. coli and mammalian cells). In some embodiments, the polynucleotides are isolated or purified.

[0285] As used herein, “isolated” polynucleotide or nucleic acid molecule is one isolated from other nucleic acid molecules present in the natural origin of the nucleic acid molecule (e.g., mouse or human). “Isolated” nucleic acid molecules, such as cDNA molecules, may not substantially contain other cellular material or culture medium if produced by recombinant techniques, and may not substantially contain chemical precursors or other chemicals if chemically synthesized. For example, the phrase “substantially contained” includes preparations of polynucleotide or nucleic acid molecules containing less than about 15%, 10%, 5%, 2%, 1%, 0.5%, or 0.1% of other material, such as cellular material, culture medium, other nucleic acid molecules, chemical precursors, and / or other chemicals.

[0286] In certain embodiments, the Specified provides a polynucleotide comprising a nucleotide sequence encoding an anti-MERTK antibody or its antigen-binding fragment, which specifically binds to a MERTK (e.g., human MERTK) polynucleotide and comprises an amino acid sequence described herein, and an antibody that competes with such antibody for binding (e.g., in a dose-dependent manner) to an epitope that is the same as or overlaps with that of such antibody.

[0287] In certain embodiments, this specification provides polynucleotides comprising nucleotide sequences encoding the light or heavy chain of an antibody described herein.

[0288] In certain embodiments, the polynucleotide described herein comprises a nucleotide sequence encoding an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK), and the antibody or its antigen-binding fragment comprises a heavy chain variable region containing the amino acid sequence described herein (SEQ ID NO: 105, SEQ ID NO: 107, SEQ ID NO: 108, or SEQ ID NO: 109). In certain embodiments, the polynucleotide described herein comprises a nucleotide sequence encoding an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK), and the anti-MERTK antibody or its antigen-binding fragment comprises a light chain variable region containing the amino acid sequence described herein (SEQ ID NO: 106, or SEQ ID NO: 111). In certain embodiments, the polynucleotide described herein comprises a nucleotide sequence encoding an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK), and the antibody or its antigen-binding fragment comprises a heavy chain variable region containing the amino acid sequence of SEQ ID NO: 105 and a light chain variable region containing the amino acid sequence of SEQ ID NO: 106. In certain embodiments, the polynucleotide described herein comprises a nucleotide sequence encoding the anti-MERTK antibody described herein or its antigen-binding fragment, which specifically binds to MERTK (e.g., human MERTK), and the antibody or its antigen-binding fragment comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 107 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 106. In certain embodiments, the polynucleotide described herein comprises a nucleotide sequence encoding the anti-MERTK antibody described herein or its antigen-binding fragment, which specifically binds to MERTK (e.g., human MERTK), and the antibody or its antigen-binding fragment comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 108 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 106.In certain embodiments, the polynucleotide described herein comprises a nucleotide sequence encoding an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or its antigen-binding fragment comprises a heavy chain variable region containing the amino acid sequence of SEQ ID NO: 110 and a light chain variable region containing the amino acid sequence of SEQ ID NO: 111. In certain embodiments, the polynucleotide described herein comprises a nucleotide sequence encoding an anti-MERTK antibody or its antigen-binding fragment that specifically binds to MERTK (e.g., human MERTK), wherein the antibody or its antigen-binding fragment comprises a heavy chain variable region containing the amino acid sequence of SEQ ID NO: 109 and a light chain variable region containing the amino acid sequence of SEQ ID NO: 106.

[0289] In certain embodiments, the polynucleotide described herein codes for VH, and the polynucleotide includes the nucleic acid sequence of SEQ ID NO: 119, SEQ ID NO: 121, SEQ ID NO: 122, or SEQ ID NO: 123. In certain embodiments, the polynucleotide described herein codes for VL, and the polynucleotide includes the nucleic acid sequence of SEQ ID NO: 120, or SEQ ID NO: 124. In specific embodiments, the polynucleotide described herein codes for VH and VL, and the polynucleotide includes the nucleic acid sequence of SEQ ID NO: 119, and the polynucleotide includes the nucleic acid sequence of SEQ ID NO: 120. In another specific embodiment, the polynucleotide described herein codes for VH and VL, and the polynucleotide includes the nucleic acid sequence of SEQ ID NO: 121, and the polynucleotide includes the nucleic acid sequence of SEQ ID NO: 120. In yet another specific embodiment, the polynucleotide described herein codes for VH and VL, and the polynucleotide includes the nucleic acid sequence of SEQ ID NO: 122, and the polynucleotide includes the nucleic acid sequence of SEQ ID NO: 120. In another embodiment, the polynucleotide described herein encodes VH and VL, the polynucleotide comprises the nucleic acid of SEQ ID NO: 123, and the polynucleotide comprises the nucleic acid sequence of SEQ ID NO: 124.

[0290] In specific embodiments, this specification provides polynucleotides comprising nucleotide sequences encoding antibodies, for example, separate light and heavy chains. With respect to the light chain, in specific embodiments, the polynucleotides provided herein comprise nucleotide sequences encoding antibodies described herein, comprising human kappa light chains or human lambda light chains.

[0291] In a specific embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding the light chain of an antibody, wherein the nucleotide sequence comprises the variable light chain region of SEQ ID NO: 106 and the nucleotide sequence of the human light chain constant region of the human kappa or lambda light chain. In another embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding the light chain of an antibody, wherein the light chain comprises the variable light chain region of SEQ ID NO: 111 and the human light chain constant region of the human kappa or lambda light chain.

[0292] In certain embodiments, the polynucleotide provided herein comprises a nucleotide sequence encoding an antibody described herein that specifically binds to MERTK (e.g., human MERTK), the antibody comprising a light chain, the amino acid sequence of the variable region of the light chain may comprise any amino acid sequence of SEQ ID NO: 106 or SEQ ID NO: 111, and the constant region of the light chain comprising the amino acid sequence of the human kappa light chain constant region. In another particular embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding an antibody described herein that specifically binds to MERTK (e.g., human MERTK) and comprises a light chain, the amino acid sequence of the variable region of the light chain may comprise any amino acid sequence of SEQ ID NO: 106 or SEQ ID NO: 111, and the constant region of the light chain comprising the amino acid sequence of the human lambda light chain constant region. In yet another embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding the light chain of an antibody, the light chain comprising the amino acid sequence of SEQ ID NO: 113 or 118. In another embodiment, the polynucleotides provided herein include a nucleotide sequence encoding the light chain of an antibody that specifically binds to MERTK (e.g., human MERTK), the nucleotide sequence including SEQ ID NO: 126 or 130.

[0293] In certain embodiments, the polynucleotide provided herein comprises a nucleotide sequence encoding an antibody described herein that specifically binds to MERTK (e.g., human MERTK), the antibody comprising a heavy chain, the heavy chain comprising the amino acid sequence of SEQ ID NO: 105 and a heavy chain constant region. In another particular embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding an antibody described herein that specifically binds to MERTK (e.g., human MERTK) and includes a heavy chain, the heavy chain comprising the amino acid sequence of SEQ ID NO: 107 and a heavy chain constant region. In yet another particular embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding an antibody described herein that specifically binds to MERTK (e.g., human MERTK) and includes a heavy chain, the heavy chain comprising the amino acid sequence of SEQ ID NO: 108 and a heavy chain constant region. In yet another particular embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding an antibody described herein that specifically binds to MERTK (e.g., human MERTK) and includes a heavy chain, the heavy chain comprising the amino acid sequence of SEQ ID NO: 109 and a heavy chain constant region. In another specific embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding an antibody described herein that specifically binds to MERTK (e.g., human MERTK) and includes a heavy chain, the heavy chain comprising the amino acid sequence of SEQ ID NO: 110 and a heavy chain constant region.

[0294] In certain embodiments, the polynucleotide provided herein comprises a nucleotide sequence encoding an antibody described herein that specifically binds to MERTK (e.g., human MERTK), the antibody comprising a heavy chain, the heavy chain comprising the amino acid sequence of SEQ ID NO: 112. In certain embodiments, the polynucleotide provided herein comprises a nucleotide sequence encoding an antibody described herein that specifically binds to MERTK (e.g., human MERTK), the antibody comprising a heavy chain, the heavy chain comprising the amino acid sequence of SEQ ID NO: 114. In certain embodiments, the polynucleotide provided herein comprises a nucleotide sequence encoding an antibody described herein that specifically binds to MERTK (e.g., human MERTK), the antibody comprising a heavy chain, the heavy chain comprising the amino acid sequence of SEQ ID NO: 115. In certain embodiments, the polynucleotide provided herein comprises a nucleotide sequence encoding an antibody described herein that specifically binds to MERTK (e.g., human MERTK), the antibody comprising a heavy chain, the heavy chain comprising the amino acid sequence of SEQ ID NO: 116. In certain embodiments, the polynucleotide provided herein comprises a nucleotide sequence encoding an antibody described herein that specifically binds to MERTK (e.g., human MERTK), the antibody comprising a heavy chain, the heavy chain comprising the amino acid sequence of SEQ ID NO: 117.

[0295] In a specific embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding the heavy chain of an antibody that specifically binds to MERTK (e.g., human MERTK), wherein the nucleotide sequence comprises SEQ ID NO: 119 and the nucleotide sequence of the human heavy chain constant region. In another embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding the heavy chain of an antibody that specifically binds to MERTK, wherein the nucleotide sequence comprises SEQ ID NO: 125.

[0296] In a specific embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding the heavy chain of an antibody that specifically binds to MERTK (e.g., human MERTK), wherein the nucleotide sequence comprises SEQ ID NO: 121 and the nucleotide sequence of the human heavy chain constant region. In another embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding the heavy chain of an antibody that specifically binds to MERTK, wherein the nucleotide comprises SEQ ID NO: 127.

[0297] In a specific embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding the heavy chain of an antibody that specifically binds to MERTK (e.g., human MERTK), wherein the nucleotide sequence comprises SEQ ID NO: 122 and the nucleotide sequence of the human heavy chain constant region. In another embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding the heavy chain of an antibody that specifically binds to MERTK (e.g., human MERTK), wherein the nucleotide sequence comprises SEQ ID NO: 128.

[0298] In a specific embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding the heavy chain of an antibody that specifically binds to MERTK (e.g., human MERTK), wherein the nucleotide sequence comprises SEQ ID NO: 123 and the nucleotide sequence of the human heavy chain constant region. In another embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding the heavy chain of an antibody that specifically binds to MERTK (e.g., human MERTK), wherein the nucleotide comprises SEQ ID NO: 129.

[0299] In a specific embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding the light chain of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), and a nucleotide sequence encoding the heavy chain of the antibody, wherein the nucleotide sequence encoding the heavy chain comprises SEQ ID NO: 119 and the nucleotide sequence of the human heavy chain constant region, and the nucleotide sequence encoding the light chain comprises SEQ ID NO: 120 and the nucleotide sequence of the human heavy chain constant region. In another specific embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding the light chain of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), and a nucleotide sequence encoding the heavy chain of the antibody, wherein the nucleotide sequence encoding the light chain comprises SEQ ID NO: 120 and the nucleotide sequence of the human light chain constant region, and the nucleotide sequence encoding the heavy chain comprises SEQ ID NO: 121 and the nucleotide sequence of the human heavy chain constant region. In another specific embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding the light chain of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), and a nucleotide sequence encoding the heavy chain of the antibody, wherein the nucleotide sequence encoding the light chain comprises SEQ ID NO: 120 and the nucleotide sequence of the human light chain constant region, and the nucleotide sequence encoding the heavy chain comprises SEQ ID NO: 122 and the nucleotide sequence of the human heavy chain constant region. In another specific embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding the light chain of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), and a nucleotide sequence encoding the heavy chain of the antibody, wherein the nucleotide sequence encoding the light chain comprises SEQ ID NO: 124 and the nucleotide sequence of the human light chain constant region, and the nucleotide sequence encoding the heavy chain comprises SEQ ID NO: 123 and the nucleotide sequence of the human heavy chain constant region.

[0300] In a specific embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding an antibody that specifically binds to MERTK (e.g., human MERTK), wherein the antibody comprises (1) a variable heavy chain region and a human heavy chain constant region comprising the amino acid sequence of SEQ ID NO: 105, and (2) a variable light chain region and a human light chain constant region comprising the amino acid sequence of SEQ ID NO: 106. In another specific embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding an antibody that specifically binds to MERTK (e.g., human MERTK), wherein the antibody comprises (1) a variable heavy chain region and a human heavy chain constant region comprising the amino acid sequence of SEQ ID NO: 107, and (2) a variable light chain region and a human light chain constant region comprising the amino acid sequence of SEQ ID NO: 106. In yet another specific embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding an antibody that specifically binds to MERTK (e.g., human MERTK), wherein the antibody comprises (1) a variable heavy chain region and a human heavy chain constant region comprising the amino acid sequence of SEQ ID NO: 108, and (2) a variable light chain region and a human light chain constant region comprising the amino acid sequence of SEQ ID NO: 106. In another specific embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding an antibody that specifically binds to MERTK (e.g., human MERTK), wherein the antibody comprises (1) a variable heavy chain region and a human heavy chain constant region comprising the amino acid sequence of SEQ ID NO: 110, and (2) a variable light chain region and a human light chain constant region comprising the amino acid sequence of SEQ ID NO: 111.

[0301] In another specific embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding an antibody that specifically binds to MERTK (e.g., human MERTK), wherein the antibody comprises (1) a variable heavy chain region and a human heavy chain constant region comprising the amino acid sequence of SEQ ID NO: 109, and (2) a variable light chain region and a human light chain constant region comprising the amino acid sequence of SEQ ID NO: 106.

[0302] In another specific embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), where one nucleotide sequence encodes the heavy chain of the antibody and the other nucleotide sequence encodes the light chain of the antibody, the nucleotide sequence encoding the heavy chain comprising SEQ ID NO: 125 and the nucleotide sequence encoding the light chain comprising SEQ ID NO: 126. In another specific embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), where one nucleotide sequence encodes the heavy chain of the antibody and the other nucleotide sequence encodes the light chain of the antibody, the nucleotide sequence encoding the heavy chain comprising SEQ ID NO: 127 and the nucleotide sequence encoding the light chain comprising SEQ ID NO: 126. In another specific embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), where one nucleotide sequence encodes the heavy chain of the antibody and the other nucleotide sequence encodes the light chain of the antibody, the nucleotide sequence encoding the heavy chain includes SEQ ID NO: 128 and the nucleotide sequence encoding the light chain includes SEQ ID NO: 126. In another specific embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), where one nucleotide sequence encodes the heavy chain of the antibody and the other nucleotide sequence encodes the light chain of the antibody, the nucleotide sequence encoding the heavy chain includes SEQ ID NO: 129 and the nucleotide sequence encoding the light chain includes SEQ ID NO: 130.

[0303] In another specific embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), where one nucleotide sequence encodes the heavy chain of the antibody and the other encodes the light chain of the antibody, the heavy chain comprising the amino acid sequence of SEQ ID NO: 112 and the light chain comprising the amino acid sequence of SEQ ID NO: 113. In another specific embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), where one nucleotide sequence encodes the heavy chain of the antibody and the other encodes the light chain of the antibody, the heavy chain comprising the amino acid sequence of SEQ ID NO: 114 and the light chain comprising the amino acid sequence of SEQ ID NO: 113. In another specific embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein one nucleotide sequence encodes the heavy chain of the antibody and the other encodes the light chain of the antibody, the heavy chain comprising the amino acid sequence of SEQ ID NO: 115 and the light chain comprising the amino acid sequence of SEQ ID NO: 113. In another specific embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), wherein one nucleotide sequence encodes the heavy chain of the antibody and the other encodes the light chain of the antibody, the heavy chain comprising the amino acid sequence of SEQ ID NO: 116 and the light chain comprising the amino acid sequence of SEQ ID NO: 113. In another specific embodiment, the polynucleotide provided herein comprises a nucleotide sequence encoding an anti-MERTK antibody or an antigen-binding fragment thereof that specifically binds to MERTK (e.g., human MERTK), where one nucleotide sequence encodes the heavy chain of the antibody and the other encodes the light chain of the antibody, the heavy chain comprising the amino acid sequence of SEQ ID NO: 117 and the light chain comprising the amino acid sequence of SEQ ID NO: 118.

[0304] In certain embodiments, polynucleotides, nucleic acids, or nucleotides include deoxyribonucleic acid, ribonucleic acid, ribonucleotides, and polymeric forms thereof. In some embodiments, polynucleotides, nucleic acids, or nucleotides are single-stranded or double-stranded. In specific embodiments, the sequence of the polynucleotide, nucleic acid, or nucleotide is a cDNA sequence.

[0305] In some embodiments, the polynucleotide sequence (e.g., nucleic acid sequence) described herein that encodes the anti-MERTK antibody or its antigen-binding fragment is codon-optimized using methodologies known to those skilled in the art. In certain embodiments, the optimized polynucleotide sequence encoding the anti-MERTK antibody or its antigen-binding fragment, or its antigen-binding fragment (e.g., the VH domain and / or VL domain), can be hybridized to an antisense (e.g., complementary) polynucleotide of the unoptimized polynucleotide sequence encoding the anti-MERTK antibody or its antigen-binding fragment (e.g., the VH domain and / or VL domain). In specific embodiments, the optimized nucleotide sequence encoding the anti-MERTK antibody or its antigen-binding fragment is hybridized to an antisense polynucleotide of the unoptimized polynucleotide sequence encoding the anti-MERTK antibody or its antigen-binding fragment under high stringency conditions. In specific embodiments, an optimized nucleotide sequence encoding an anti-MERTK antibody or its antigen-binding fragment is hybridized to an antisense polynucleotide of a non-optimized nucleotide sequence encoding the anti-MERTK antibody or its antigen-binding fragment described herein, under high-stringency, medium-stringency, or low-stringency hybridization conditions. Information regarding hybridization conditions is described herein; see, for example, U.S. Patent Application Publication US2005 / 0048549 (e.g., paragraphs 72-73), incorporated herein by reference.

[0306] Polynucleotides can be obtained and their nucleotide sequences determined by any method known in the art. The nucleotide sequences encoding the antibodies described herein and modified versions thereof can be determined using methods known in the art, i.e., nucleotide codons known to encode specific amino acids are assembled in such a manner to generate nucleic acid sequences encoding antibodies. Such polynucleotides encoding antibodies can be assembled from chemically synthesized oligonucleotides (e.g., as described in Kutmeier G et al., (1994), BioTechniques 17: 242-6), and in short, this involves a synthesis in which oligonucleotides containing portions of the antibody-encoding sequence are duplicated, these oligonucleotides are annealed and ligated, and the ligated oligonucleotides are then amplified by PCR.

[0307] Alternatively, the polynucleotides encoding the antibodies described herein can be generated from nucleic acids from a suitable source (e.g., hybridomas) using methods well known in the art (e.g., PCR and other molecular cloning methods). For example, PCR amplification using synthetic primers hybridizable to the 3' and 5' ends of known sequences can be performed using genomic DNA obtained from hybridoma cells producing the antibody of interest. Using such a PCR amplification method, nucleic acids containing sequences encoding the light and / or heavy chains of the antibody can be obtained. Using such a PCR amplification method, nucleic acids containing sequences encoding the variable light and / or variable heavy chain regions of the antibody can be obtained. The amplified nucleic acids can be cloned into vectors for expression in host cells and further cloning, for example, to generate humanized antibodies.

[0308] If clones containing nucleic acid sequences encoding a specific antibody are not available, but the sequence of the antibody molecule is known, nucleic acids encoding immunoglobulins can be obtained by chemical synthesis or by PCR amplification using synthetic primers that can hybridize to the 3' and 5' ends of the sequence, preferably poly(A+RNA), from a suitable source (e.g., an antibody cDNA library, or a cDNA library generated from any tissue or cell expressing the antibody, such as hybridoma cells selected to express the antibody described herein, or nucleic acids isolated therefrom, preferably poly(A+RNA)), or by cloning using oligonucleotide probes specific to a particular gene sequence to identify cDNA clones from an antibody-encoding cDNA library. The amplified nucleic acids produced by PCR can then be cloned into a replicable cloning vector using any method known in the art.

[0309] The DNA encoding the anti-MERTK antibody described herein can be readily isolated and sequenced using conventional procedures (e.g., by using oligonucleotide probes that can specifically bind to the genes encoding the heavy and light chains of the anti-MERTK antibody). Hybridoma cells can serve as the source of such DNA. Once isolated, the DNA can be placed in an expression vector and then transfected into host cells that do not otherwise produce immunoglobulin proteins, such as E. coli cells, monkey COS cells, Chinese hamster ovary (CHO) cells (e.g., CHO cells from CHO GS System® (Lonza)), 293F cells, HEK293 cells, or myeloma cells, to obtain a synthesis of the MERTK agonist antibody in recombinant host cells.

[0310] To generate the entire antibody, the VH or VL sequence can be amplified in the scFv clone using PCR primers containing the VH or VL nucleotide sequence, a restriction site, and a flanking sequence protecting the restriction site. Using cloning techniques known to those skilled in the art, the PCR-amplified VH domain can be cloned into a vector expressing a heavy chain constant region, e.g., the human gamma-4 constant region, and the PCR-amplified VL domain can be cloned into a vector expressing a light chain constant region, e.g., the human kappa or lambda constant region. In certain embodiments, the vector for expressing the VH or VL domain includes a promoter, a secretion signal, a cloning site for a variable domain, a constant domain, and a selection marker. The VH and VL domains can also be cloned into a single vector expressing the required constant region. The heavy chain and light chain conversion vectors are then co-transfected into cell lines using techniques known to those skilled in the art to generate stable or transient cell lines expressing a full-length antibody, e.g., IgG.

[0311] DNA can also be modified, for example, by substituting the mouse sequence with the coding sequence for the human heavy and light chain constant domains, or by covalently joining all or part of the coding sequence for a non-immunoglobulin polypeptide to the immunoglobulin coding sequence.

[0312] Furthermore, this specification provides polynucleotides that hybridize to the antibodies or antigen-binding fragments thereof described herein under high-stringency, medium-stringency, or low-stringency hybridization conditions. In specific embodiments, the polynucleotides described herein hybridize to the polynucleotides encoding VH (SEQ ID NOs. 119, 121, 122, or 123) and / or VL (SEQ ID NOs. 120 or 124) provided herein under high-stringency, medium-stringency, or low-stringency hybridization conditions.

[0313] Hybridization conditions have been described in the art and are known to those skilled in the art. For example, hybridization under stringent conditions may include hybridization to filter-bound DNA in 6× sodium chloride / sodium citrate (SSC) at about 45°C, followed by one or more washes in 0.2× SSC / 0.1% SDS at about 50–65°C; hybridization under highly stringent conditions may include hybridization to filter-bound nucleic acids in 6× SSC at about 45°C, followed by one or more washes in 0.1× SSC / 0.2% SDS at about 68°C. Hybridization under other stringent hybridization conditions is known and described to those skilled in the art; see, for example, Ausubel FM et al., eds., (1989) Current Protocols in Molecular Biology, Vol. I, Green Publishing Associates, Inc. and John Wiley & Sons, Inc., New York at pages 6.3.1–6.3.6 and 2.10.3. 5.3.3. Cells and Vectors

[0314] In certain embodiments, the Specified herein provides a vector (e.g., an expression vector) comprising a polynucleotide containing a nucleotide sequence encoding the anti-MERTK antibody or its antigen-binding fragment as described herein for recombinant expression in host cells, preferably mammalian cells. The expression vector may be, for example, a plasmid or a viral vector (e.g., Newcastle disease virus, adenovirus, adeno-associated virus, vaccinia, etc.). The Specified herein also provides host cells comprising such a vector for recombinant expression of the anti-MERTK antibody or its antigen-binding fragment as described herein.

[0315] Recombinant expression of antibodies described herein (e.g., full-length antibodies, heavy and / or light chains of antibodies, or single-chain antibodies) that specifically bind to MERTK (e.g., human MERTK) involves the construction of an expression vector containing a polynucleotide encoding the antibody. Once polynucleotides encoding the antibody molecule, the heavy and / or light chain of an antibody, or their antigen-binding fragments (e.g., heavy and / or light chain variable regions) are obtained, a vector for the production of the antibody molecule can be produced by recombinant DNA techniques using techniques well known in the art. Therefore, a method for preparing a protein by expressing a polynucleotide containing an antibody or antibody fragment (e.g., a light chain or heavy chain, or both) encoding a nucleotide sequence is described herein. Using methods well known to those skilled in the art, an expression vector containing the coding sequence of the antibody or antibody fragment (e.g., a light chain or heavy chain, or both), as well as appropriate transcription and translational regulatory signals, can be constructed. These methods include, for example, in vitro recombinant DNA techniques, synthetic techniques, and in vivo genetic recombination. We also provide a replicable vector comprising a nucleotide sequence operably linked to a promoter, the antibody molecule described herein, the heavy or light chain of the antibody, or the heavy or light chain variable domain of an anti-MERTK antibody or its antigen-binding fragment. Such a vector may, for example, contain a nucleotide sequence encoding the constant region of the antibody molecule (see, for example, International Publication Nos. WO86 / 05807 and WO89 / 01036; and U.S. Patent No. 5,122,464), and the variable domain of the antibody can be cloned into such a vector for the expression of the entire heavy chain, the entire light chain, or both the entire heavy chain and the entire light chain.

[0316] Expression vectors can be transferred into cells (e.g., host cells) by conventional techniques, and the resulting cells can then be cultured by conventional techniques to produce the anti-MERTK antibody or its antigen-binding fragment described herein. Therefore, this specification provides host cells containing the antibody or its antigen-binding fragment, its heavy or light chain, or fragment thereof, or a polynucleotide encoding the single-chain antibody described herein, operably linked to a promoter for the expression of such a sequence in the host cell. Where used herein, the term “host cell” can be any type of cell, e.g., primary cells, cells in culture, or cells derived from a cell line. In specific embodiments, the term “host cell” refers to cells transfected with nucleic acid molecules and their offspring or potential offspring. Such offspring may not be identical to the parent cells transfected with nucleic acid molecules due to mutations or environmental influences that may occur in later generations or in the integration of nucleic acid molecules into the host cell genome.

[0317] The antibody molecules described herein can be expressed using a variety of host-expression vector systems. Such host-expression systems represent a medium by which the desired coding sequence can be produced and subsequently purified, but also represent cells that can express the antibody molecules described herein in situ when transformed or transfected with the appropriate nucleotide coding sequence. These include, but are not limited to, bacteria (e.g., E. coli and B. subtilis) transformed with recombinant bacteriophage DNA, plasmid DNA, or cosmid DNA expression vectors containing the antibody coding sequence; yeast (e.g., Saccharomyces Pichia) transformed with recombinant yeast expression vectors containing the antibody coding sequence; insect cell lines infected with recombinant virus expression vectors (e.g., baculovirus) containing the antibody coding sequence; and plant cell lines (e.g., Chlamydomonas) infected with recombinant virus expression vectors (e.g., cauliflower mosaic virus, CaMV; tobacco mosaic virus, TMV) or transformed with recombinant plasmid expression vectors (e.g., Ti plasmid) containing the antibody coding sequence. Examples include green algae such as reinhardtii; or mammalian cell lines possessing recombinant expression constructs containing promoters derived from the genome of mammalian cells (e.g., metallothionein promoter) or promoters derived from mammalian viruses (e.g., late adenovirus promoter; vaccinia virus 7.5K promoter) (e.g., COS (e.g., COS1 or COS), CHO, BHK, MDCK, HEK293, NS0, PER.C6, VERO, CRL7O3O, HsS78Bst, HeLa, and NIH3T3, HEK-293T, 293F, HepG2, SP210, R1.1, BW, LM, BSC1, BSC40, YB / 20, and BMT10 cells). In specific embodiments, the cells for expressing the antibodies or antigen-binding fragments described herein are CHO cells, for example, CHO cells from the CHO GS System (Lonza). In certain embodiments, the cells for expressing the antibodies described herein are human cells, for example, human cell lines.In specific embodiments, the mammalian expression vector is pOptiVEC® or pcDNA3.3. In certain embodiments, bacterial cells such as Escherichia coli or eukaryotic cells (e.g., mammalian cells) are used for the expression of recombinant antibody molecules, particularly for the expression of the entire recombinant antibody molecule. Mammalian cells such as Chinese hamster ovary (CHO) cells, combined with a vector such as a major intermediate early gene promoter element derived from human cytomegalovirus, are an effective expression system for antibodies (Foecking MK & Hofstetter H (1986) Gene 45: 101-5; and Cockett MI et al., (1990) Biotechnology 8(7): 662-7). In certain embodiments, the antibodies described herein are produced by CHO cells or NS0 cells. In specific embodiments, the expression of the nucleotide sequence encoding the antibodies described herein that immunospecifically bind MERTK (e.g., human MERTK) is regulated by a constitutive promoter, an inducible promoter, or a tissue-specific promoter.

[0318] In bacterial systems, several expression vectors can be advantageously selected depending on the intended use of the antibody molecule being expressed. For example, when producing large quantities of such antibodies for the production of pharmaceutical compositions of antibody molecules, a vector that directs the expression of a high level of fusion protein product that can be easily purified may be desirable. Such vectors include, but are not limited to, the E. coli expression vector pUR278 (Ruether U & Mueller-Hill B (1983) EMBO J 2: 1791-1794), where the antibody coding sequence can be individually ligated into the vector in frame with the lac Z coding region, thereby producing the fusion protein; and the pIN vector (Inouye S & Inouye M (1985) Nuc Acids Res 13: 3101-3109; Van Heeke G & Schuster SM (1989) J Biol Chem 24: 5503-5509). For example, a pGEX vector can be used to express an exogenous polypeptide as a fusion protein with glutathione 5-transferase (GST). Generally, such fusion proteins are soluble and can be easily purified from lysed cells by adsorption and binding to matrix glutathione agarose beads, followed by elution in the presence of free glutathione. The pGEX vector is designed to include a thrombin or factor Xa protease cleavage site so that the cloned target gene product can be released from the GST portion.

[0319] In insects, for example, the nuclear polyhedron virus (AcNPV) of Autographa californica can be used as a vector to express foreign genes. The virus grows in Spodoptera frugiperda cells. The coding sequence of the antibody can be individually cloned into a non-essential region of the virus (e.g., the polyhedrin gene) and placed under the control of the AcNPV promoter (e.g., the polyhedrin promoter).

[0320] Several virus-based expression systems can be used in mammalian host cells. When using adenovirus as an expression vector, the coding sequence of the antibody of interest can be ligated to the adenovirus's transcription / translation regulatory complex, such as the late promoter and trielemental reader sequence. This chimeric gene can then be inserted into the adenovirus genome by recombination in vitro or in vivo. Insertion into a non-essential region of the viral genome (e.g., region E1 or E3) produces a recombinant virus that is viable and can express the antibody molecule in an infected host (see, e.g., Logan J & Shenk T (1984) PNAS 81(12): 3655-9). Specific start signals may also be required for efficient translation of the inserted antibody coding sequence. These signals include the ATG start codon and adjacent sequences. Furthermore, the start codon must coincide with the reading frame of the desired coding sequence to ensure translation of the entire insert. These exogenous translational regulatory signals and start codons can be of various origins, both natural and synthetic. Expression efficiency can be enhanced by the inclusion of appropriate transcriptional enhancer elements, transcriptional terminators, etc. (see, for example, Bitter G et al., (1987) Methods Enzymol. 153: 516-544).

[0321] In addition, host cell lines can be selected that modulate the expression of the inserted sequence or modify and process the gene product in a desired specific manner. Such modification (e.g., glycosylation) and processing (e.g., cleavage) of protein products may be important for protein function. Different host cells have characteristic and specific mechanisms for post-translational processing and modification of proteins and gene products. By selecting an appropriate cell line or host line, the correct modification and processing of expressed foreign proteins can be ensured. For this purpose, eukaryotic host cells with cellular mechanisms for appropriate processing, glycosylation, and phosphorylation of the primary transcript of the gene product can be used. Such mammalian host cells include, but are not limited to, CHO, VERO, BHK, Hela, MDCK, HEK293, NIH3T3, W138, 293F, BT483, Hs578T, HTB2, BT2O and T47D, NS0 (mouse myeloma cell line that does not endogenously produce any immunoglobulin chains), CRL7O3O, COS (e.g., COS1 or COS), PER.C6, VERO, HsS78Bst, HEK-293T, HEK293, HepG2, SP210, R1.1, BW, LM, BSC1, BSC40, YB / 20, BMT10 and HsS78Bst cells. In certain embodiments, the anti-MERTK antibody described herein is produced in mammalian cells such as CHO cells.

[0322] In specific embodiments, the antibodies or antigen-binding fragments described herein have a reduced fucose content or no fucose content at all. Such antibodies can be produced using techniques known to those skilled in the art. For example, the antibodies can be expressed in cells that have insufficient or deficient ability to fucosylate. In a specific example, a cell system having knockouts of both alleles of α1,6-fucosyltransferase can be used to produce a reduced antibody or antigen-binding fragment having a reduced fucose content. The Potelligent® system (Lonza) is an example of such a system that can be used to produce an antibody or antigen-binding fragment having a reduced fucose content.

[0323] For long-term, high-yield production of recombinant proteins, stable expression cells can be generated. For example, a cell line that stably expresses the anti-MERTK antibody or its antigen-binding fragment described herein, or the chimeric anti-MERTK antibody or its antigen-binding fragment described herein, can be manipulated. In specific embodiments, the cells provided herein stably express a light chain / light chain variable domain and a heavy chain / heavy chain variable domain that associate to form the antibody or its antigen-binding fragment described herein.

[0324] In certain embodiments, instead of using an expression vector containing a viral replication origin, host cells can be transformed with DNA controlled by appropriate expression regulatory elements (e.g., promoters, enhancers, sequences, transcription terminators, polyadenylation sites, etc.) and selectable markers. After introduction of exogenous DNA / polynucleotides, the engineered cells can be grown in enriched medium for 1-2 days and then switched to selective medium. Selectable markers in recombinant plasmids confer resistance to selection, allowing cells to stably incorporate the plasmid into their chromosomes, grow, and form proliferation foci that can then be cloned and expanded into cell lines. This method can be advantageously used to engineer cell lines expressing the humanized anti-MERTK antibody or its antigen-binding fragment described herein, or the chimeric anti-MERTK antibody or its antigen-binding fragment described herein. Such engineered cell lines may be particularly useful in screening and evaluating compositions that interact directly or indirectly with antibody molecules.

[0325] Several select systems can be used, but are not limited to, the genes for herpes simplex virus thymidine kinase (Wigler M et al., (1977) Cell 11(1): 223-32), hypoxanthine guanine phosphoribosyltransferase (Szybalska EH & Szybalski W (1962) PNAS 48(12): 2026-2034), and adenine phosphoribosyltransferase (Lowy I et al., (1980) Cell 22(3): 817-23), which can be used in TK cells, HGPRT cells, or APRT cells, respectively. Furthermore, antimetabolite resistance can be used as a basis for selection for the following genes: dhfr (Wigler M et al., (1980) PNAS 77(6): 3567-70; O'Hare K et al., (1981) PNAS 78: 1527-31) which confers resistance to methotrexate; gpt (Mulligan RC & Berg P (1981) PNAS 78(4): 2072-6) which confers resistance to mycophenolate; neo (Wu GY & Wu CH (1991) Biotherapy 3: 87-95; Tolstoshev P (1993) Ann Rev Pharmacol Toxicol 32: 573-596; Mulligan RC (1993) Science 260: 926-932; and Morgan RA & Anderson WF (1993) Ann Rev Biochem 62: 191-217; Nabel GJ & Felgner PL (1993) Trends Biotechnol 11(5): 211-5); and hygro (Santerre RF et al., (1984) Gene 30(1-3): 147-56) which confers resistance to hygromycin.Methods generally known in the field of recombinant DNA technology can be routinely applied to select desired recombinant clones, and such methods are described, for example, in Ausubel FM et al., (eds.), Current Protocols in Molecular Biology, John Wiley & Sons, NY (1993); Kriegler M, Gene Transfer and Expression, A Laboratory Manual, Stockton Press, NY (1990); and Chapters 12 and 13, Dracopoli NC et al., (eds.), Current Protocols in Human Genetics, John Wiley & Sons, NY (1994); and Colbere-Garapin F et al., (1981) J Mol Biol 150: 1-14, which are incorporated herein by reference in their entirety.

[0326] The expression level of antibody molecules can be increased by vector amplification (see Bebbington CR & Hentschel CCG, The use of vectors based on gene amplification for the expression of cloned genes in mammalian cells in DNA cloning, Vol. 3 (Academic Press, New York, 1987) for a review). If the marker in an antibody-expressing vector system is amplified, an increase in the level of the inhibitor present in the host cell culture increases the number of copies of the marker gene. Since the amplified region is related to the antibody gene, antibody production also increases (Crouse GF et al., (1983) Mol Cell Biol 3: 257-66).

[0327] Host cells can be co-transfected with two or more expression vectors described herein, namely a first vector encoding a heavy-chain polypeptide and a second vector encoding a light-chain polypeptide. The two vectors may contain identical selectable markers that enable equivalent expression of the heavy-chain and light-chain polypeptides. Host cells can be co-transfected with two or more expression vectors in different amounts. For example, host cells can be transfected with any one of the following ratios of the first and second expression vectors: 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:9, 1:10, 1:12, 1:15, 1:20, 1:25, 1:30, 1:35, 1:40, 1:45, or 1:50.

[0328] In a specific embodiment, the host cell provided herein comprises a vector comprising a nucleotide sequence encoding a variable light chain region (VL) of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), and a nucleotide sequence encoding a variable heavy chain region (VH) of the antibody, wherein VL comprises the amino acid sequence of SEQ ID NO: 106, and VH comprises the amino acid sequence of SEQ ID NO: 105. In another specific embodiment, the host cell provided herein comprises a vector comprising a nucleotide sequence encoding a light chain of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), and a nucleotide sequence encoding a heavy chain of the antibody, wherein the light chain comprises a variable light chain region comprising the amino acids of SEQ ID NO: 106, and a human light chain constant region, and the heavy chain comprises a variable heavy chain region comprising the amino acid sequence of SEQ ID NO: 105, and a human heavy chain constant region.

[0329] In a specific embodiment, the host cell provided herein comprises a vector comprising a nucleotide sequence encoding a variable light chain region (VL) of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), and a nucleotide sequence encoding a variable heavy chain region (VH) of the antibody, wherein VL comprises the amino acid sequence of SEQ ID NO: 106, and VH comprises the amino acid sequence of SEQ ID NO: 109. In another specific embodiment, the host cell provided herein comprises a vector comprising a nucleotide sequence encoding a light chain of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), and a nucleotide sequence encoding a heavy chain of the antibody, wherein the light chain comprises a variable light chain region comprising the amino acid sequence of SEQ ID NO: 106, and a human light chain constant region, and the heavy chain comprises a variable heavy chain region comprising the amino acid sequence of SEQ ID NO: 109, and a human heavy chain constant region.

[0330] In a specific embodiment, the host cell provided herein comprises a vector comprising a nucleotide sequence encoding the VL of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), and a nucleotide sequence encoding the VH of the antibody, wherein the VL comprises the amino acid sequence of SEQ ID NO: 106, and the VH comprises the amino acid sequence of SEQ ID NO: 107. In another specific embodiment, the host cell provided herein comprises a vector comprising a nucleotide sequence encoding the light chain of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), and a nucleotide sequence encoding the heavy chain of the antibody, wherein the light chain comprises the VL comprising the amino acid sequence of SEQ ID NO: 106, and a human light chain constant region, and the heavy chain comprises the VH comprising the amino acid sequence of SEQ ID NO: 107, and a human heavy chain constant region.

[0331] In a specific embodiment, the host cell provided herein comprises a vector comprising a nucleotide sequence encoding the VL of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), and a nucleotide sequence encoding the VH of the antibody, wherein the VL comprises the amino acid sequence of SEQ ID NO: 106, and the VH comprises the amino acid sequence of SEQ ID NO: 108. In another specific embodiment, the host cell provided herein comprises a vector comprising a nucleotide sequence encoding the light chain of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), and a nucleotide sequence encoding the heavy chain of the antibody, wherein the light chain comprises the variable light chain region of SEQ ID NO: 108 and the human light chain constant region, and the heavy chain comprises the nucleotide sequence of the variable heavy chain region of SEQ ID NO: 105 and the human heavy chain constant region.

[0332] In a specific embodiment, the host cell provided herein comprises a vector comprising a nucleotide sequence encoding VL of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), and a nucleotide sequence encoding VH of the antibody, wherein VL comprises the amino acid sequence of SEQ ID NO: 111, and VH comprises the amino acid sequence of SEQ ID NO: 110.

[0333] In another specific embodiment, a host cell provided herein comprises a vector comprising a nucleotide sequence encoding the light chain of an antibody described herein that specifically binds to MERTK (e.g., human MERTK), and a nucleotide sequence encoding the heavy chain of the antibody, wherein the nucleotide sequence encoding the light chain comprises SEQ ID NO: 120 and the nucleotide sequence of the human light chain constant region, and the nucleotide sequence encoding the heavy chain comprises SEQ ID NO: 119 and the nucleotide sequence of the human heavy chain constant region.

[0334] In another specific embodiment, the host cell provided herein comprises a vector comprising a nucleotide sequence encoding the light chain of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), and a nucleotide sequence encoding the heavy chain of the antibody, wherein the nucleotide sequence encoding the light chain comprises SEQ ID NO: 120 and the nucleotide sequence of the human light chain constant region, and the nucleotide sequence encoding the heavy chain comprises SEQ ID NO: 121 and the nucleotide sequence of the human heavy chain constant region. In another specific embodiment, the host cell provided herein comprises a vector comprising a nucleotide sequence encoding the light chain of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), and a nucleotide sequence encoding the heavy chain of the antibody, wherein the nucleotide sequence encoding the light chain comprises SEQ ID NO: 120 and the nucleotide sequence of the human light chain constant region, and the nucleotide sequence encoding the heavy chain comprises SEQ ID NO: 122 and the nucleotide sequence of the human heavy chain constant region. In another specific embodiment, a host cell provided herein comprises a vector comprising a nucleotide sequence encoding the light chain of an antibody described herein that specifically binds to MERTK (e.g., human MERTK), and a nucleotide sequence encoding the heavy chain of the antibody, wherein the nucleotide sequence encoding the light chain comprises SEQ ID NO: 124 and the nucleotide sequence of the human light chain constant region, and the nucleotide sequence encoding the heavy chain comprises SEQ ID NO: 123 and the nucleotide sequence of the human heavy chain constant region.

[0335] In a specific embodiment, the host cell provided herein comprises a first vector and a second vector, the first vector comprising a nucleotide sequence encoding the variable heavy chain region of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), and the second vector comprising the variable light chain region of the antibody, the variable heavy chain region comprising the amino acid sequence of SEQ ID NO: 105, and the variable light chain region comprising the amino acid sequence of SEQ ID NO: 106. In another specific embodiment, the host cell provided herein comprises a first vector and a second vector, the first vector comprising a nucleotide sequence encoding the variable heavy chain region of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), the second vector comprising the variable light chain region of the antibody, the variable heavy chain region comprising the amino acid sequence of SEQ ID NO: 107, and the variable light chain region comprising the amino acid sequence of SEQ ID NO: 106. In another specific embodiment, the host cell provided herein comprises a first vector and a second vector, the first vector comprising a nucleotide sequence encoding the variable heavy chain region of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), and the second vector comprising the variable light chain region of the antibody, the variable heavy chain region comprising the amino acid sequence of SEQ ID NO: 108, and the variable light chain region comprising the amino acid sequence of SEQ ID NO: 106. In another specific embodiment, the host cell provided herein comprises a first vector and a second vector, the first vector comprising a nucleotide sequence encoding the variable heavy chain region of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), the second vector comprising the variable light chain region of the antibody, the variable heavy chain region comprising the amino acid sequence of SEQ ID NO: 109, and the variable light chain region comprising the amino acid sequence of SEQ ID NO: 106. In another specific embodiment, the host cell provided herein comprises a first vector and a second vector, the first vector comprising a nucleotide sequence encoding a variable heavy chain region of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), and the second vector comprising a variable light chain region of the antibody, the variable heavy chain region comprising the amino acid sequence of SEQ ID NO: 110, and the variable light chain region comprising the amino acid sequence of SEQ ID NO: 111.In some embodiments, the first vector includes a heavy chain steady region, and / or the second vector includes a light chain steady region.

[0336] Alternatively, a single vector can be used that encodes and expresses both heavy and light chain polypeptides. In such an expression vector, the transcription of both genes may be driven by a common promoter, but the translation of mRNA from the first gene may be by a cap-dependent scanning mechanism, while the translation of mRNA from the second gene may be by a cap-independent mechanism, such as IRES.

[0337] In a specific embodiment, the host cell provided herein comprises a vector comprising a nucleotide sequence encoding the variable heavy chain region of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), and a nucleotide encoding the variable light chain region of the antibody, wherein the variable heavy chain region comprises the amino acid sequence of SEQ ID NO: 105, and the variable light chain region comprises the amino acid sequence of SEQ ID NO: 106. In another specific embodiment, the host cell provided herein comprises a vector comprising a nucleotide sequence encoding the variable heavy chain region of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), and a nucleotide encoding the variable light chain region of the antibody, wherein the variable heavy chain region comprises the amino acid sequence of SEQ ID NO: 107, and the variable light chain region comprises the amino acid sequence of SEQ ID NO: 106. In another specific embodiment, the host cell provided herein comprises a vector comprising a nucleotide sequence encoding a variable heavy chain region of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), and a nucleotide encoding a variable light chain region of the antibody, wherein the variable heavy chain region comprises the amino acid sequence of SEQ ID NO: 108, and the variable light chain region comprises the amino acid sequence of SEQ ID NO: 106. In another specific embodiment, the host cell provided herein comprises a vector comprising a nucleotide sequence encoding a variable heavy chain region of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), and a nucleotide encoding a variable light chain region of the antibody, wherein the variable heavy chain region comprises the amino acid sequence of SEQ ID NO: 109, and the variable light chain region comprises the amino acid sequence of SEQ ID NO: 106. In another specific embodiment, the host cell provided herein comprises a vector comprising a nucleotide sequence encoding a variable heavy chain region of the antibody described herein that specifically binds to MERTK (e.g., human MERTK), and a nucleotide encoding a variable light chain region of the antibody, wherein the variable heavy chain region comprises the amino acid sequence of SEQ ID NO: 110, and the variable light chain region comprises the amino acid sequence of SEQ ID NO: 111. In some embodiments, the vector comprises a heavy chain constant region and / or a light chain constant region.

[0338] In specific embodiments, the host cells described herein (e.g., ex vivo host cells) are cultured under conditions that produce an antibody or its antigen-binding fragment encoded by a polynucleotide sequence contained in the host cells, using techniques known in the art. In certain embodiments, the antibody or its antigen-binding fragment is isolated or purified from the host cells using techniques known in the art. 5.4 Production of Antibody-Drug Conjugates

[0339] In another embodiment, this specification provides a method for producing an antibody-drug conjugate as described herein, wherein a linker is absent, and the method comprises the steps of (a) directly conjugating a cytotoxic agent to the antibody moiety to produce an antibody-drug conjugate, and (b) purifying the antibody-drug conjugate. An example of a method for producing an antibody-drug conjugate is described in Section 6 (e.g., Example 4).

[0340] In another embodiment, the Specified Publicly Provided Method for Producing an Antibody-Drug Conjugate as Described herein, wherein the antibody-drug conjugate comprises a linker, and the Method comprises the following steps in the order described above: (a) conjugating the linker directly to an antibody moiety to produce a linker-antibody moiety; (b) conjugating the linker of the linker-antibody moiety directly to a cytotoxic agent to produce an antibody-drug conjugate; and (c) purifying the antibody-drug conjugate.

[0341] In another embodiment, the Specified Publicly Provided Method for Producing an Antibody-Drug Conjugate as Described herein, wherein the antibody-drug conjugate comprises a linker, and the Method comprises the following steps in the order described above: (a) conjugating the linker directly to a cytotoxic agent to produce a linker-cytotoxic agent portion; (b) conjugating the linker of the linker-cytotoxic agent portion directly to an antibody portion to produce an antibody-drug conjugate; and (c) purifying the antibody-drug conjugate.

[0342] The steps of conjugating and purifying can be carried out by methods known in the art used to produce antibody-drug conjugates, for example, Beck A et al., (2017) Nat Rev Drug Discov 16: 315-337; Peters C and Brown S, (2015) Biosci Rep 35: art:e00225; McCombs JR and Owen SC (2015) The AAPS Journal 17: 339-351; Jackson DY (2016) Org Process Res Dev 20: 852-866; or Olivier KJ and Hurvitz SA ed., (2016) Antibody-Drug Conjugates: Fundamentals, Drug Development, and Clinical, Wiley. In some embodiments, the drug portion is conjugated to one chain of the antibody portion (for example, when the antibody portion is an scFv, or when the antibody portion is a multichain antibody such as an immunoglobulin (which is a tetramer) or its antigen-binding fragment). In other embodiments, the drug portion is conjugated to two or more chains of the antibody portion (when the antibody portion is a multichain antibody such as an immunoglobulin or its antigen-binding fragment). In specific embodiments, the drug portion is conjugated to two identical chains of the immunoglobulin, for example, the heavy chain or the light chain. In other embodiments, the drug portion is conjugated to all chains of the antibody portion (when the antibody portion is a multichain antibody such as an immunoglobulin or its antigen-binding fragment). In specific embodiments, the antibody-drug conjugate described herein is purified by chromatography. In another specific embodiment, the antibody-drug conjugate described herein is purified by centrifugation.Conjugation chemistry, which can be used to generate antibody-drug conjugates, but is not limited to, includes: thiols and maleimides, thiols and self-hydrolyzed maleimides, thiols and phenyloxadiazole sulfone, oxime ligation, reactions of alkoxyamines and keto groups, strain-accelerated azide-alkyne cycloaddition (SPAAC), copper-free click chemistry, cysteine ​​oxidized to formylglycine, hydrazino-iso-picte-spengler (HIPS) ligation, ligation of γ-carboxyamide groups from glutamine residues and primary amines, ligation of LPETG and polyg This could involve lysine-motif primary amines, maleimide and 6-thiofucose, fucose-specific conjugation of hydrazide, periodate oxidation (aldehyde) and aminooxy payload, strain-promoting alkyne-azide cycloaddition, C2-keto-gal oxime formation, site-selective aldehyde oxidation and oxime ligation, oxime ligation NH2 and indole-based 5-difluoro-2,4-dinitrobenzene derivatives, thiol and bissulfone, thiol and dibromomaleimide, thiol and maleimide followed by pH 9.2 treatment (45°C, 48 hours), or thiol and arylpropionitrile (see Beck A et al., (2017) Nat Rev Drug Discov 16: 315-337).

[0343] The anti-MERTK antibody or its antigen-binding fragment described herein can be produced as described in 5.3 below. In specific embodiments, the anti-MERTK antibody or its antigen-binding fragment described herein is manipulated or modified by methods known in the art, for example, by the method described in Beck A et al., (2017) Nat Rev Drug Discov 16: 315-337; Peters C and Brown S, (2015) Biosci Rep 35: art:e00225; McCombs JR and Owen SC (2015) The AAPS Journal 17: 339-351; Jackson DY (2016) Org Process Res Dev 20: 852-866; or Olivier KJ and Hurvitz SA ed., (2016) Antibody-Drug Conjugates: Fundamentals, Drug Development, and Clinical, Wiley, in order to facilitate conjugation with a drug moiety, and in particular to promote site-specific conjugation with a drug moiety. Non-limiting exemplary methods that can be used to manipulate or modify an anti-MERTK antibody or its antigen-binding fragment to facilitate conjugation with a drug moiety (see Beck A et al., (2017) Nat Rev Drug Discov 16: 315-337) include the addition of one or more additional cysteine ​​or selenocysteine, non-natural amino acid manipulation, the addition of one or more amino acid tags recognizable by certain enzymes that can assist in conjugation, glycan remodeling, the addition of amino-terminal serines, and native cysteine ​​crosslinking.

[0344] The linker and the cytotoxic agent can be produced, respectively, by any method known in the art for producing the linker and the cytotoxic agent. 5.5. Pharmaceutical Compositions

[0345] This specification provides for a pharmaceutical composition comprising (a) an anti-MERTK antibody (e.g., a humanized antibody) or its antigen-binding fragment as described herein, and (b) a pharmaceutically acceptable carrier. In specific embodiments, the antibody or its antigen-binding fragment is purified. In specific embodiments, the antibody or its antigen-binding fragment is present in the pharmaceutical composition in a therapeutically effective amount. In specific embodiments, the pharmaceutical composition comprises an anti-MERTK antibody or its antigen-binding fragment (e.g., antibody z10) comprising VH containing the amino acid sequence of SEQ ID NO: 105 and VL containing the amino acid sequence of SEQ ID NO: 106. In another specific embodiment, the pharmaceutical composition comprises an anti-MERTK antibody or its antigen-binding fragment (e.g., antibody z11) comprising VH containing the amino acid sequence of SEQ ID NO: 107 and VL containing the amino acid sequence of SEQ ID NO: 106. In yet another specific embodiment, the pharmaceutical composition comprises an anti-MERTK antibody or its antigen-binding fragment (e.g., antibody z13) comprising VH containing the amino acid sequence of SEQ ID NO: 108 and VL containing the amino acid sequence of SEQ ID NO: 106. In another specific embodiment, the pharmaceutical composition comprises an anti-MERTK antibody or its antigen-binding fragment, comprising a VL containing the amino acid sequence of SEQ ID NO: 109 and the amino acid sequence of SEQ ID NO: 106.

[0346] This specification also provides pharmaceutical compositions comprising (a) a chimeric anti-MERTK antibody or its antigen-binding fragment as described herein, and (b) a pharmaceutically acceptable carrier. In specific embodiments, the antibody or its antigen-binding fragment is purified. In specific embodiments, the antibody or its antigen-binding fragment is present in the pharmaceutical composition in a therapeutically effective amount. In specific embodiments, the pharmaceutical composition comprises a chimeric antibody or its antigen-binding fragment (e.g., antibody xAb) comprising VH, which comprises the amino acid sequence of SEQ ID NO: 110, and VL, which comprises the amino acid sequence of SEQ ID NO: 111. This specification also provides pharmaceutical compositions comprising a polynucleotide or vector as described herein, and a pharmaceutically acceptable carrier.

[0347] This specification also provides a pharmaceutical composition comprising (a) an antibody-drug conjugate described herein, and (b) a pharmaceutically acceptable carrier. In specific embodiments, the antibody-drug conjugate is present in the pharmaceutical composition in a therapeutically effective amount.

[0348] This specification also provides a pharmaceutical composition comprising (a) a bispecific antibody (e.g., as described herein) that binds to MERTK (e.g., human MERTK) and another antigen of interest, and (b) a pharmaceutically acceptable carrier. In specific embodiments, the bispecific antibody is purified. In specific embodiments, the bispecific antibody is present in the pharmaceutical composition in a therapeutically effective amount.

[0349] In a specific embodiment, the group of antibody-drug conjugates contained in the pharmaceutical composition has a drug-to-antibody ratio (DAR) between 1 and 20. The DAR is the average number of cytotoxic agents conjugated to the antibody. In another specific embodiment, the group of antibody-drug conjugates contained in the pharmaceutical composition has a DAR between 1 and 15. In yet another specific embodiment, the group of antibody-drug conjugates contained in the pharmaceutical composition has a DAR between 1 and 12. In yet another specific embodiment, the group of antibody-drug conjugates contained in the pharmaceutical composition has a DAR between 1 and 8. In yet another specific embodiment, the group of antibody-drug conjugates contained in the pharmaceutical composition has a DAR between 3 and 5. In yet another specific embodiment, the group of antibody-drug conjugates contained in the pharmaceutical composition has a DAR between 3.5 and 4.

[0350] Acceptable carriers, which may be excipients or stabilizers, are non-toxic to the recipient at the dosage and concentration used and are not limited to, but include: buffers such as phosphates, citrates and other organic acids; antioxidants including ascorbic acid and methionine; preservatives (octadecyldimethylbenzylammonium chloride; hexamethonium chloride; benzalkonium chloride, benzethonium chloride; phenol, butyl alcohol or benzyl alcohol; alkylparabens such as methylparaben or propylparaben; catechol; resorcinol; cyclohexanol; 3-pentanol; and m-cresol); and low molecular weight (less than approximately 10 residues) polypeptides. Examples include: 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, disaccharides, and other carbohydrates including glucose, mannose, or dextrin; chelating agents such as EDTA; sugars such as sucrose, mannitol, trehalose, or sorbitol; salt-forming counterions such as sodium; metal complexes (e.g., Zn-protein complexes); and / or nonionic surfactants such as TWEEN®, PLURONICS®, or polyethylene glycol (PEG).

[0351] In specific embodiments, the pharmaceutical composition comprises, in a pharmaceutically acceptable carrier, an anti-MERTK antibody or its antigen-binding fragment as described herein, and optionally one or more additional prophylactic or therapeutic agents. In specific embodiments, the pharmaceutical composition comprises, in a pharmaceutically acceptable carrier, an effective amount of the anti-MERTK antibody or its antigen-binding fragment as described herein, and optionally one or more additional prophylactic or therapeutic agents. In specific embodiments, the pharmaceutical composition and / or antibody or its antigen-binding fragment as described herein may be combined with any of the additional therapeutic agents as described herein in a therapeutically effective amount (see Section 5.4.2 below).

[0352] In specific embodiments, the pharmaceutical composition comprises an antibody-drug conjugate described herein and, optionally, one or more additional prophylactic or therapeutic agents in a pharmaceutically acceptable carrier. In specific embodiments, the pharmaceutical composition comprises an effective amount of an antibody-drug conjugate described herein and, optionally, one or more additional prophylactic or therapeutic agents in a pharmaceutically acceptable carrier. In specific embodiments, the pharmaceutical composition and / or antibody-drug conjugate described herein may be combined with any of the additional therapeutic agents described herein in a therapeutically effective amount (see Section 5.4.2 below).

[0353] In some embodiments, the antibody-drug conjugate is the sole active ingredient in the pharmaceutical composition.

[0354] In some embodiments, the anti-MERTK antibody is the sole active ingredient in the pharmaceutical composition. In specific embodiments, the humanized antibody or its antigen-binding fragment described herein is the sole active ingredient in the pharmaceutical composition. In other embodiments, the polynucleotide or vector encoding the antibody or its antigen-binding fragment is the sole active ingredient in the pharmaceutical composition.

[0355] The pharmaceutical compositions described herein can be used to treat cancer. The pharmaceutical compositions may be formulated for any route of administration (e.g., parenteral, topical, intratumor, etc.).

[0356] Pharmaceutically acceptable carriers used in parenteral preparations include aqueous and non-aqueous media, antimicrobial agents, isotonic agents, buffers, antioxidants, local anesthetics, suspending and dispersing agents, emulsifiers, metal ion sequestering or chelating agents, and other pharmaceutically acceptable substances. Examples of aqueous media include sodium chloride injection, Ringer's injection, isotonic dextrose injection, sterile water injection, dextrose, and Ringer's lactate injection. Non-aqueous parenteral media include plant-derived fixative oils, cottonseed oil, corn oil, sesame oil, and peanut oil. Antimicrobial agents in bacteriostatic or fungiostatic concentrations may be added to parenteral preparations packaged in multi-dose containers, containing phenol or cresol, mercury preparations, benzyl alcohol, chlorobutanol, methyl and propyl p-hydroxybenzoates, thimerosal, benzalkonium chloride, and benzethonium chloride. Examples of isotonic agents include sodium ch...

Claims

1. An antibody or antigen-binding fragment thereof that specifically binds to human Mer tyrosine kinase (MERTK), wherein the antibody or antigen-binding fragment comprises a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VH comprises the amino acid sequence of SEQ ID NO: 105 and the VL comprises the amino acid sequence of SEQ ID NO:

106.

2. An antibody or antigen-binding fragment thereof that specifically binds to human MERTK, wherein the antibody or antigen-binding fragment comprises a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VH comprises the amino acid sequence of SEQ ID NO: 107 or 108, and the VL comprises the amino acid sequence of SEQ ID NO:

106.

3. An antibody or antigen-binding fragment thereof that specifically binds to human MERTK, wherein the antibody or antigen-binding fragment comprises a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VH comprises the amino acid sequence of SEQ ID NO: 108 or 109, and the VL comprises the amino acid sequence of SEQ ID NO:

106.

4. An antibody or antigen-binding fragment thereof that specifically binds to human MERTK, wherein the antibody or antigen-binding fragment comprises a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VH comprises the amino acid sequence of SEQ ID NO: 110, and the VL comprises the amino acid sequence of SEQ ID NO:

120.

5. The antibody or antigen-binding fragment thereof according to any one of claims 1 to 4, wherein the antibody is a monoclonal antibody.

6. 6. The antibody or antigen-binding fragment thereof according to claim 1, wherein the antibody is an immunoglobulin comprising two identical heavy chains and two identical light chains.

7. The antibody or antigen-binding fragment thereof according to any one of claims 1 to 6, wherein the antibody comprises a heavy chain constant region and a light chain constant region of human origin.

8. The antibody or antigen-binding fragment thereof according to any one of claims 1 to 7, wherein the antibody is an IgG.

9. The antibody or antigen-binding fragment thereof according to any one of claims 1 to 8, wherein the antibody comprises a portion of a heavy chain constant region and a portion of a light chain constant region of human origin.

10. The antibody or antigen-binding fragment thereof according to any one of claims 1 to 9, wherein the antigen-binding fragment is a Fab or F(ab')2 fragment.

11. A bispecific antibody comprising two different antigen-binding regions, one of which comprises the antibody or antigen-binding fragment of any one of claims 1 to 4, and the other of which binds to an antigen of interest.

12. 12. The bispecific antibody of claim 11, wherein the antigen of interest is an immune cell receptor or a tumor-associated antigen.

13. 12. The bispecific antibody of claim 11, wherein the antigen of interest is CD3, PD-L1, LRP1, LPR8, TGF-β, ICOS, CD40, NKGD2, or TIGIT.

14. 11. An antibody-drug conjugate comprising: (a) an antibody moiety which is the antibody or antigen-binding fragment thereof of any one of claims 1 to 10; (b) one or more drug moieties, each drug moiety being a cytotoxic agent; and (c) optionally a linker, wherein the cytotoxic agent is conjugated to the antibody moiety directly or via the linker.

15. 15. The antibody-drug conjugate of claim 14, having a molar ratio of said antibody moiety to said drug moiety between 1:3 and 1:

12.

16. 15. The antibody-drug conjugate of claim 14, having a molar ratio of said antibody moiety to said drug moiety of 1:

9.

17. The antibody-drug conjugate of any one of claims 14 to 16, wherein the cytotoxic agent is a small molecule, a nucleotide, a peptide, or a non-antibody protein.

18. The antibody-drug conjugate of any one of claims 14 to 16, wherein the cytotoxic agent is a small molecule.

19. 17. The antibody-drug conjugate of any one of claims 14 to 16, wherein the cytotoxic agent is an auristatin, a maytansinoid, a pyrrolobenzodiazepine, an indolinobenzodiazepine, a calicheamicin, a camptothecin analog, a duocarmycin, a tubulin inhibitor, a tublysin or a tublysin analog, amberstatin 269, doxorubicin, SN-38, an antibiotic, an anthracycline, a microtubule inhibitor, a splicesostatin, or a tylanstatin.

20. The antibody-drug conjugate of any one of claims 14 to 16, wherein the cytotoxic agent is monomethyl auristatin E (MMAE) or monomethyl auristatin F (MMAF).

21. The antibody-drug conjugate of any one of claims 14 to 16, wherein the cytotoxic agent is DM1 or DM4.

22. The antibody-drug conjugate of any one of claims 14 to 16, wherein the cytotoxic agent is monomethyl auristatin E.

23. The antibody-drug conjugate of any one of claims 14 to 16, wherein the cytotoxic agent is SN-38.

24. The antibody-drug conjugate of any one of claims 14 to 23, wherein the antibody-drug conjugate comprises the linker, and the linker is a cleavable linker.

25. The antibody-drug conjugate of any one of claims 14 to 23, wherein the antibody-drug conjugate comprises the linker, and the linker is a non-cleavable linker.

26. The antibody-drug conjugate of any one of claims 14 to 23, wherein the antibody-drug conjugate comprises the linker, and the linker is maleimidocaproyl-valine-citrulline-p-aminobenzyloxycarbonyl.

27. The antibody-drug conjugate of any one of claims 14 to 23, wherein the antibody-drug conjugate comprises the linker, and the linker is CL2.

28. The antibody-drug conjugate of any one of claims 14 to 23, wherein the antibody-drug conjugate comprises the linker, and the linker is CL2A.

29. 11. An isolated nucleic acid sequence comprising a polynucleotide encoding the antibody or antigen-binding fragment thereof of any one of claims 1 to 10.

30. 11. An isolated nucleic acid sequence comprising a polynucleotide encoding a VH according to any one of claims 1 to 10.

31. 11. An isolated nucleic acid sequence comprising a polynucleotide encoding a VL according to any one of claims 1 to 10.

32. An ex vivo cell containing one or more polynucleotides encoding the antibody or antigen-binding fragment of any one of SEQ ID NOs: 1 to 10.

33. 27. A method for producing an antibody or antigen-binding fragment, comprising culturing the cell of claim 26 under conditions such that the one or more polynucleotides are expressed by the cell to produce the antibody or antigen-binding fragment encoded by the polynucleotide.

34. 29. A method of producing the antibody-drug conjugate of any one of claims 14 to 28, wherein the linker is absent, the method comprising the steps of: (a) conjugating the cytotoxic agent directly to the antibody moiety to produce the antibody-drug conjugate; and (b) purifying the antibody-drug conjugate.

35. 29. A method of producing the antibody-drug conjugate of any one of claims 14 to 28, wherein the antibody-drug conjugate comprises the linker, the method comprising the following steps, in the order stated: (a) conjugating the linker directly to the antibody moiety to produce a linker-antibody moiety, (b) conjugating the linker of the linker-antibody moiety directly to the cytotoxic agent to produce the antibody-drug conjugate, and (c) purifying the antibody-drug conjugate.

36. 29. A method of producing the antibody-drug conjugate of any one of claims 14 to 28, wherein the antibody-drug conjugate comprises the linker, the method comprising the following steps in the order stated: (a) conjugating the linker directly to the cytotoxic agent to produce a linker-cytotoxic agent moiety, (b) conjugating the linker of the linker-cytotoxic agent moiety directly to the antibody moiety to produce the antibody-drug conjugate, and (c) purifying the antibody-drug conjugate.

37. A pharmaceutical composition comprising a therapeutically effective amount of the antibody or antigen-binding fragment of any one of claims 1 to 10 or the antibody-drug conjugate of any one of claims 14 to 28, and a pharmaceutically acceptable carrier.

38. 40. A method of treating cancer in a subject in need thereof, comprising administering to the subject the pharmaceutical composition of claim 37.

39. 39. The method of claim 38, wherein the cancer is cancer of the head and neck, lung, breast, bone, ovary, stomach, pancreas, larynx, esophagus, testis, liver, parotid gland, bile duct, colon, rectum, cervix, uterus, endometrium, kidney, bladder, prostate, or thyroid, or the cancer is melanoma, sarcoma, or leukemia.

40. 39. The method of claim 38, wherein the cancer is sarcoma, squamous cell carcinoma, melanoma, glioma, glioblastoma, neuroblastoma, Kaposi's sarcoma, gastric cancer, colorectal cancer, non-small cell lung cancer, head and neck cancer, or multiple myeloma.

41. 39. The method of claim 38, wherein the cancer is leukemia or lymphoma.

42. 42. The method of claim 41, wherein the leukemia is acute myeloid leukemia or acute lymphocytic leukemia.

43. 39. The method of claim 38, wherein the cancer is breast cancer.

44. 39. The method of claim 38, wherein the cancer is triple-negative breast cancer.

45. 45. The method of any one of claims 38 to 44, wherein cancerous cells of the cancer overexpress MERTK.

46. 45. The method of any one of claims 38 to 44, wherein cancerous cells of the cancer overexpress phosphorylated MERTK.

47. 47. The method of any one of claims 38 to 46, wherein MERTK is constitutively active on cancerous cells of the cancer.

48. 45. The method of any one of claims 38 to 44, wherein the cancer is associated with overexpression of MERTK.

49. 49. The method of any one of claims 38-44 or 48, wherein the cancer is associated with constitutively active MERTK.

50. 45. The method of any one of claims 38 to 44, further comprising the steps of assessing whether phosphorylated MERTK is overexpressed in a tumor sample from the subject before treating the subject, and treating the subject if phosphorylated MERTK is overexpressed in the tumor sample.

51. 51. The method of any one of claims 38 to 50, further comprising administering to the subject an additional therapeutic agent.

52. 52. The method of claim 51, wherein the additional therapeutic agent is for treating the cancer.

53. 53. The method of claim 52, wherein the additional therapeutic agent is an agent used to treat breast cancer, an agent used to treat melanoma, an immunotherapy, or an angiogenesis inhibitor.

54. 53. The method of claim 52, wherein the additional therapeutic agent is an angiogenesis inhibitor selected from the group consisting of a VEGF inhibitor, a VEGFR2 inhibitor, sunitinib, and sorafenib.

55. 53. The method of claim 52, wherein the additional therapeutic agent is an agent used to treat breast cancer selected from the group consisting of tamoxifen, raloxifene, paclitaxel, cyclophosphamide, docetaxel, vinblastine, fluorouracil, everolimus, trastuzumab, trastuzumab-emtansine, pertuzumab, and lapatinib ditosylate.

56. 53. The method of claim 52, wherein the additional therapeutic agent is an agent used to treat melanoma selected from the group consisting of a BRAF inhibitor, a MEK inhibitor, and dacarbazine.

57. 53. The method of claim 52, wherein the additional therapeutic agent is an agent that blocks immune checkpoint signaling.

58. 58. The method of claim 57, wherein the additional therapeutic agent is an anti-CTLA-4 antibody, an anti-PD1 antibody, or an anti-PDL1 antibody.

59. 59. The method of any one of claims 38 to 58, wherein the additional therapeutic agent is an anti-Lag3 antibody, an agonistic GITR antibody, an agonistic OX40 antibody, an anti-ICOS antibody, an anti-NKGD2 antibody, an anti-TIGIT antibody, an anti-TGF-β antibody, CAR-T therapy, an ApoE mimetic, RGX-104, or RGX-202.

60. 60. The method of any one of claims 38 to 59, wherein the subject is a human.

61. 1. An antibody-drug conjugate comprising an antibody portion comprising a heavy chain variable region (VH) and a light chain variable region (VL), wherein said VH comprises the amino acid sequence of SEQ ID NO: 105 and said VL comprises the amino acid sequence of SEQ ID NO: 106, wherein said antibody portion is conjugated to MMAE via an mc-vc-PABC linker.

62. 1. An antibody-drug conjugate comprising an antibody portion comprising a heavy chain variable region (VH) and a light chain variable region (VL), wherein said VH comprises the amino acid sequence of SEQ ID NO: 105 and said VL comprises the amino acid sequence of SEQ ID NO: 106, wherein said antibody portion is conjugated to SN-38 via a CL2A linker.

63. The antibody-drug conjugate of claim 61 or 62, wherein the antibody moiety is an immunoglobulin.

64. 64. The antibody-drug conjugate of claim 63, wherein the immunoglobulin comprises a human constant region.

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