Anti-CLEC12A antibodies and uses thereof

Abstract

Described herein are antibodies, fragments thereof, and fusion proteins that specifically bind to Clec12A, and methods of making and using such antibodies. Such antibodies, fusion proteins, and fragments thereof are useful in the treatment and diagnosis of various autoimmune diseases and cancers, including leukemias, lymphomas, or myelomas, such as acute myeloid leukemia.

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This application claims priority to and the benefit of U.S. Provisional Application No. 63 / 179,755, filed April 26, 2021, the entire contents of which are incorporated herein by reference.

[0003] Sequence Listing

[0004] This application contains a Sequence Listing that has been submitted electronically in ASCII format and is hereby incorporated by reference in its entirety. The ASCII copy created on April 19, 2022 is named MIL-017WO_SL.txt and is 44,670 bytes in size. [Background technology]

[0005] Anti-Clec12A antibodies bind to the Clec12A antigen, also known as MICL or C-type lectin domain family 12 member A. The Clec12A antigen is an inhibitory C-type lectin-like receptor. Human MICL is expressed as a monomer primarily on myeloid cells such as granulocytes, monocytes, macrophages, and dendritic cells. Mouse MICL is expressed as a dimer on granulocytes, monocytes, but also on B lymphocytes and can be found on the surface of NK cells in the bone marrow.

[0006] The Clec12A gene encodes a member of the C-type lectin / C-type lectin-like domain (CTL / CTLD) superfamily, which share a common protein fold and have diverse functions, including roles in cell adhesion, cell-cell signaling, glycoprotein turnover, and inflammation and immune responses. The protein encoded by this gene inhibits the function of granulocytes and monocytes. Summary of the Invention

[0007] The present invention provides human anti-Clec12A antibodies and fragments thereof for the treatment of autoimmune diseases and cancers, such as leukemia, lymphoma, or myeloma, e.g., relapsed and refractory acute myeloid leukemia. The antibodies and fragments thereof of the present invention may be used alone, in fusion proteins, or conjugated to at least one diagnostic and / or therapeutic agent, or in combination with other therapies. Binding of human Clec12A to the anti-Clec12A antibodies or fragments thereof described herein may exhibit antibody-dependent cellular cytotoxicity (ADCC) activity, e.g., induction of apoptosis and inhibition of cancer cell proliferation.

[0008] In some embodiments, an anti-Clec12A antibody or antigen-binding fragment thereof is provided that comprises: (a) an immunoglobulin heavy chain variable (VH) region comprising an amino acid sequence at least 80% identical to QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYYWSWIRQPPGKGLEWIGYIYYSGSTKYNPSLKSRVTISVDTSKNLFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSS (SEQ ID NO: 1); and an immunoglobulin light chain variable (VL) region comprising an amino acid sequence at least about 80% identical to IQMTQSPSSLSASVGDRVTITCRASQGIRYDLGWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO:2); (b) an immunoglobulin heavy chain variable (VH) region comprising an amino acid sequence at least about 80% identical to QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYYVDSVKGRFTISRDNSKNTLYLHMNSLRAEDTAVYYCARDKGYYFDYWGQGTLVTVSS (SEQ ID NO: 3); and an immunoglobulin light chain variable (VL) region comprising an amino acid sequence at least about 80% identical to EIVMTQSPATLSLSPGERATLSCRASQSVGNRYLSWYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO:4); (c) an immunoglobulin heavy chain variable (VH) region comprising an amino acid sequence at least about 80% identical to QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYSADSVKGRFNISRDNSKNTLYLQMNSLRAEDTAVYYCARDGSRYFDYWGQGTLVTVSS (SEQ ID NO: 5); and An immunoglobulin light chain variable (VL) region comprising an amino acid sequence at least about 80% identical to EIFMTQSPATLSLSPGERATLSCRASQSVHSKYLSWYQQKPGQAPSLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPITFGQGTRLEIK (SEQ ID NO: 6); or (d) an immunoglobulin heavy chain variable (VH) region comprising an amino acid sequence at least about 80% identical to QLQLQESGPGLVKPSETLSLTCTVSGGSISSSTYYYWGWIRQPPRKGLEWIGSTHYRGSTYYNPSLKSRVTISVDTSKNQFSLKVSSVTAADTAVYYCARELTGEVFDYWGQGTLVTVSS (SEQ ID NO: 7); and an immunoglobulin light chain variable (VL) region comprising an amino acid sequence at least about 80% identical to DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSYSTPFTFGPGTKVDIK (SEQ ID NO: 8); (e) an immunoglobulin heavy chain variable (VH) region comprising an amino acid sequence at least about 80% identical to QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYYWSWIRQPPGKGLEWLGYIYFSGSTNYNPSLKSRLTISVAASKSQFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSS (SEQ ID NO: 9); and an immunoglobulin light chain variable (VL) region comprising an amino acid sequence at least about 80% identical to AIQMTQSPSSLSASVGDRVTITCRASQGIRNDLGWFQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTYFTLTISSLQPEDSATYYCLQDYNYPRTFGQGTKVEIK (SEQ ID NO: 10); (f) an immunoglobulin heavy chain variable (VH) region comprising an amino acid sequence at least about 80% identical to QVQLQESGPGLVKPSETLSLTCTVSGGSISTDYWSWIRQPPGKGLEWIGYIYFSGSTKYNPSLKSRVTISVDTSKNQFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSS (SEQ ID NO: 11); and an immunoglobulin light chain variable (VL) region comprising an amino acid sequence at least about 80% identical to AIQMTQSPSSLSASVGDRVTITCRASQDIRNDLGWFQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO: 12); (g) an immunoglobulin heavy chain variable (VH) region comprising an amino acid sequence at least about 80% identical to QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGEGLEWVTVISYDGSDKYYADSVKGRFTISRDNSKSTLFLQMNSLRAEDTAVYYCARDGQFYFDYWGQGTLVTVSS (SEQ ID NO: 13); and an immunoglobulin light chain variable (VL) region comprising an amino acid sequence at least about 80% identical to EIVMTQSPATLSLSPGESATLSCRASQSVTSRYLSWYQQKPGQAPRLLMYGASTRPTGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO: 14); (h) an immunoglobulin heavy chain variable (VH) region comprising an amino acid sequence at least about 80% identical to QVQLVESGGGVVQPGRSLRLSCAASGFTFSNYGMHWVRQAPGKGLEWVAVISYDGSDKSYKDSVKGRFTIARDNSKNTLYLQMNSLRAEDTAVYYCARDSGRYFFDYWGQGTLVTVSS (SEQ ID NO: 15); and An immunoglobulin light chain variable (VL) region comprising an amino acid sequence at least about 80% identical to EIIMTQSPATLSLSPGERATLSCRASQSVSSRSLSWYQHKPGQAPRLLIYGPSTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCHQDYNLPLTFGGGTKVEIK (SEQ ID NO: 16); or (i) an immunoglobulin heavy chain variable (VH) region comprising an amino acid sequence at least about 80% identical to QVKLVESGGGVVQPGRSLRLSCAASGFTFSKYGMHWVRQAPGKGLEWVAFIWYDGSIKNYADSVKGRFTTSRDNSKNTLYLQMNSLRAEDTAVYYCARGSLWFGEFYFDYWGQGTLVTVSS (SEQ ID NO: 17); and An immunoglobulin light chain variable (VL) region comprising an amino acid sequence that is at least about 80% identical to AIQLTQSPSSLSASVGDRVTITCRTSQGISSALAWYQQKPGKTPKLLIYDASSLESGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQFNNYPRTFGQGTKVEIK (SEQ ID NO: 18).

[0009] In some embodiments, the anti-Clec12A antibody or antibody-binding fragment thereof comprises: (a) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYYWSWIRQPPGKGLEWIGYIYYSGSTKYNPSLKSRVTISVDTSKNLFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSS (SEQ ID NO: 1); and An immunoglobulin light chain variable (VL) region comprising the amino acid sequence of IQMTQSPSSLSASVGDRVTITCRASQGIRYDLGWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO:2); (b) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYYVDSVKGRFTISRDNSKNTLYLHMNSLRAEDTAVYYCARDKGYYFDYWGQGTLVTVSS (SEQ ID NO: 3); and An immunoglobulin light chain variable (VL) region comprising the amino acid sequence of EIVMTQSPATLSLSPGERATLSCRASQSVGNRYLSWYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO: 4); (c) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYSADSVKGRFNISRDNSKNTLYLQMNSLRAEDTAVYYCARDGSRYFDYWGQGTLVTVSS (SEQ ID NO: 5); and An immunoglobulin light chain variable (VL) region comprising the amino acid sequence of: EIFMTQSPATLSLSPGERATLSCRASQSVHSKYLSWYQQKPGQAPSLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPITFGQGTRLEIK (SEQ ID NO: 6); or (d) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QLQLQESGPGLVKPSETLSLTCTVSGGSISSSTYYYWGWIRQPPRKGLEWIGSTHYRGSTYYNPSLKSRVTISVDTSKNQFSLKVSSVTAADTAVYYCARELTGEVFDYWGQGTLVTVSS (SEQ ID NO: 7); and an immunoglobulin light chain variable (VL) region comprising the amino acid sequence of DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSYSTPFTFGPGTKVDIK (SEQ ID NO: 8); (e) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYYWSWIRQPPGKGLEWLGYIYFSGSTNYNPSLKSRLTISVAASKSQFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSS (SEQ ID NO: 9); and an immunoglobulin light chain variable (VL) region comprising the amino acid sequence of AIQMTQSPSSLSASVGDRVTITCRASQGIRNDLGWFQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTYFTLTISSLQPEDSATYYCLQDYNYPRTFGQGTKVEIK (SEQ ID NO: 10); (f) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLQESGPGLVKPSETLSLTCTVSGGSISTDYWSWIRQPPGKGLEWIGYIYFSGSTKYNPSLKSRVTISVDTSKNQFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSS (SEQ ID NO: 11); and an immunoglobulin light chain variable (VL) region comprising the amino acid sequence of AIQMTQSPSSLSASVGDRVTITCRASQDIRNDLGWFQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO: 12); (g) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGEGLEWVTVISYDGSDKYYADSVKGRFTISRDNSKSTLFLQMNSLRAEDTAVYYCARDGQFYFDYWGQGTLVTVSS (SEQ ID NO: 13); and An immunoglobulin light chain variable (VL) region comprising the amino acid sequence of EIVMTQSPATLSLSPGESATLSCRASQSVTSRYLSWYQQKPGQAPRLLMYGASTRPTGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO: 14); (h) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLVESGGGVVQPGRSLRLSCAASGFTFSNYGMHWVRQAPGKGLEWVAVISYDGSDKSYKDSVKGRFTIARDNSKNTLYLQMNSLRAEDTAVYYCARDSGRYFFDYWGQGTLVTVSS (SEQ ID NO: 15); and An immunoglobulin light chain variable (VL) region comprising the amino acid sequence of EIIMTQSPATLSLSPGERATLSCRASQSVSSRSLSWYQHKPGQAPRLLIYGPSTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCHQDYNLPLTFGGGTKVEIK (SEQ ID NO: 16); or (i) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVKLVESGGGVVQPGRSLRLSCAASGFTFSKYGMHWVRQAPGKGLEWVAFIWYDGSIKNYADSVKGRFTTSRDNSKNTLYLQMNSLRAEDTAVYYCARGSLWFGEFYFDYWGQGTLVTVSS (SEQ ID NO: 17); and An immunoglobulin light chain variable (VL) region comprising the amino acid sequence of AIQLTQSPSSLSASVGDRVTITCRTSQGISSALAWYQQKPGKTPKLLIYDASSLESGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQFNNYPRTFGQGTKVEIK (SEQ ID NO: 18).

[0010] In some embodiments, the anti-Clec12A antibody or antibody-binding fragment thereof comprises: (a) a heavy chain complementarity determining region (HCDR) 1 comprising the amino acid sequence of GGSISTYY (SEQ ID NO: 19), an HCDR2 comprising the amino acid sequence of IYYSGST (SEQ ID NO: 20), and an HCDR3 comprising the amino acid sequence of AREDYYGSGSPFDY (SEQ ID NO: 21); a light chain complementarity determining region (LCDR) 1 comprising the amino acid sequence of QGIRYD (SEQ ID NO: 22), an LCDR2 comprising the amino acid sequence of AAS (SEQ ID NO: 23), and an LCDR3 comprising the amino acid sequence of LQDYNFPRT (SEQ ID NO: 24); (b) an HCDR1 comprising the amino acid sequence of GFTFSSYG (SEQ ID NO: 73), an HCDR2 comprising the amino acid sequence of ISYDGSDK (SEQ ID NO: 25), and an HCDR3 comprising the amino acid sequence of ARDKGYYFDY (SEQ ID NO: 26); and LCDR1 comprising the amino acid sequence of QSVGNRY (SEQ ID NO: 27), LCDR2 comprising the amino acid sequence of GAS (SEQ ID NO: 28), LCDR3 comprising the amino acid sequence of QQDYNLPLT (SEQ ID NO: 29); (c) HCDR1 comprising the amino acid sequence of GFTFSSYG (SEQ ID NO: 73), HCDR2 comprising the amino acid sequence of ISYDGSDK (SEQ ID NO: 25), and HCDR3 comprising the amino acid sequence of ARDGSRYFDY (SEQ ID NO: 30); and LCDR1 comprising the amino acid sequence of QSVHSKY (SEQ ID NO: 31), LCDR2 comprising the amino acid sequence of GAS (SEQ ID NO: 28), LCDR3 comprising the amino acid sequence of QQDYNLPIT (SEQ ID NO: 32); and (d) HCDR1 comprising the amino acid sequence of GGSISSSTYY (SEQ ID NO: 33), HCDR2 comprising the amino acid sequence of THYRGST (SEQ ID NO: 34), and HCDR3 comprising the amino acid sequence of ARELTGEVFDY (SEQ ID NO: 35); and LCDR1 comprising the amino acid sequence of QSISSY (SEQ ID NO: 36), LCDR2 comprising the amino acid sequence of AAS (SEQ ID NO: 23), and LCDR3 comprising the amino acid sequence of QQSYSTPFT (SEQ ID NO: 37); (e) HCDR1 comprising the amino acid sequence of GGSISTYY (SEQ ID NO: 19), HCDR2 comprising the amino acid sequence of IYFSGST (SEQ ID NO: 38), and HCDR3 comprising the amino acid sequence of AREDYYGSGSPFDY (SEQ ID NO: 21); and LCDR1 comprising the amino acid sequence of QGIRND (SEQ ID NO: 39), LCDR2 comprising the amino acid sequence of AAS (SEQ ID NO: 23), LCDR3 comprising the amino acid sequence of LQDYNYPRT (SEQ ID NO: 40); (f) HCDR1 comprising the amino acid sequence of GGSISTDY (SEQ ID NO: 41), HCDR2 comprising the amino acid sequence of IYFSGST (SEQ ID NO: 38), and HCDR3 comprising the amino acid sequence of AREDYYGSGSPFDY (SEQ ID NO: 21); and LCDR1 comprising the amino acid sequence of QDIRND (SEQ ID NO: 42), LCDR2 comprising the amino acid sequence of AAS (SEQ ID NO: 23), LCDR3 comprising the amino acid sequence of LQDYNFPRT (SEQ ID NO: 24); (g) HCDR1 comprising the amino acid sequence of GFTFSSYG (SEQ ID NO: 73), HCDR2 comprising the amino acid sequence of ISYDGSDK (SEQ ID NO: 25), and HCDR3 comprising the amino acid sequence of ARDGQFYFDY (SEQ ID NO: 43); and LCDR1 comprising the amino acid sequence of QSVTSRY (SEQ ID NO: 44), LCDR2 comprising the amino acid sequence of GAS (SEQ ID NO: 28), LCDR3 comprising the amino acid sequence of QQDYNLPLT (SEQ ID NO: 29); (h) HCDR1 comprising the amino acid sequence of GFTFSNYG (SEQ ID NO: 45), HCDR2 comprising the amino acid sequence of ISYDGSDK (SEQ ID NO: 25), and HCDR3 comprising the amino acid sequence of ARDSGRYFFDY (SEQ ID NO: 46); and LCDR1 comprising the amino acid sequence of QSVSSRS (SEQ ID NO: 47), LCDR2 comprising the amino acid sequence of GPS (SEQ ID NO: 48), LCDR3 comprising the amino acid sequence of HQDYNLPLT (SEQ ID NO: 49); or (i) an HCDR1 comprising the amino acid sequence of GFTFSKYG (SEQ ID NO: 50), an HCDR2 comprising the amino acid sequence of IWYDGSIK (SEQ ID NO: 51), and an HCDR3 comprising the amino acid sequence of ARGSLWFGEFYFDY (SEQ ID NO: 52); and LCDR1 comprising the amino acid sequence of QGISSA (sequence number 53), LCDR2 comprising the amino acid sequence of DAS (sequence number 54), and LCDR3 comprising the amino acid sequence of QQFNNYPRT (sequence number 55).

[0011] In some embodiments, the anti-Clec12A antibody or antigen-binding fragment thereof is selected from the group consisting of an IgA antibody, an IgG antibody, an IgE antibody, an IgM antibody, a bispecific or multispecific antibody, a Fab fragment, a Fab' fragment, a F(ab')2 fragment, an Fd' fragment, an Fd fragment, an isolated CDR or a set thereof; a single chain variable fragment (scFv), a polypeptide-Fc fusion, a single domain antibody, a camelid antibody, a masked antibody, a small modular immunopharmaceutical ("SMIPs™"), a single chain, a tandem diabody, a VHH, anticalin, a nanobody, a minibody, a BiTE, an ankyrin repeat protein, a DARPIN, an Avimer, a DART, a TCR-like antibody, an adnectin, an affilin, a transbody, an affibody, a TrimerX, a MicroProtein, a Fynomer, a Centyrin, and a KALBITOR.

[0012] In some embodiments, the anti-Clec12A antibody or antigen-binding fragment thereof is a monoclonal antibody or a single chain variable fragment (scFv).

[0013] In some embodiments, the anti-Clec12A antibody is an antibody that comprises an IgG constant region.

[0014] In some embodiments, the anti-Clec12A antigen-binding fragment is a single chain variable fragment (scFv).

[0015] In some embodiments, the scFv comprises a leader sequence, a heavy chain variable sequence, a GS linker, and a light chain variable sequence.

[0016] In some embodiments, the scFv comprises a sequence consisting of an amino acid sequence that is at least about 80% identical to: (a) QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYWSWIRQPPGKGLEWIGYIYYSGSTKYNPSLKSRVTISVDTSKNLFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSAIQMTQSPSSLSASVGDRVTITCRASQGIRYDLGWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO:56); (b) QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAIVISYDGSDKYYVDSVKGRFTISRDNSKNTLYLHMNSLRAEDTAVYYCARDKGYYFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSEIVMTQSPATLSLSPGERATLSCRASQSVGNRYLSWYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO:57); (c) QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYSADSVKGRFNISRDNSKNTLYLQMNSLRAEDTAVYYCARDGSRYFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSEIFMTQSPATLSLPGERATLSCRASQSVHSKYLSWYQQKPGQAPSLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPITFGQGTRLEIK (SEQ ID NO:58); or (d) QLQLQESGPGLVKPSETLSLTCTVSGGSISSSTYYWGWIRQPPRKGLEWIGSTHYRGSTYYNPSLKSRVTISVDTSKNQFSLKVSSVTAADTAVYYCARELTGEVFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSYSTPFTFGPGTKVDIK (SEQ ID NO: 59).

[0017] In some embodiments, the scFv comprises the following amino acid sequence: (a) QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYWSWIRQPPGKGLEWIGYIYYSGSTKYNPSLKSRVTISVDTSKNLFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSAIQMTQSPSSLSASVGDRVTITCRASQGIRYDLGWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO:56); (b) QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAIVISYDGSDKYYVDSVKGRFTISRDNSKNTLYLHMNSLRAEDTAVYYCARDKGYYFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSEIVMTQSPATLSLSPGERATLSCRASQSVGNRYLSWYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO:57); (c) QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYSADSVKGRFNISRDNSKNTLYLQMNSLRAEDTAVYYCARDGSRYFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSEIFMTQSPATLSLPGERATLSCRASQSVHSKYLSWYQQKPGQAPSLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPITFGQGTRLEIK (SEQ ID NO: 58); or (d) QLQLQESGPGLVKPSETLSLTCTVSGGSISSSTYYWGWIRQPPRKGLEWIGSTHYRGSTYYNPSLKSRVTISVDTSKNQFSLKVSSVTAADTAVYYCARELTGEVFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSYSTPFTFGPGTKVDIK (SEQ ID NO: 59).

[0018] In some embodiments, the anti-Clec12A antibody or antigen-binding fragment thereof is -8 Less than M, about 10 -9 Less than M, about 10 -10 Less than M, about 10 -11 Less than M, about 10 -12 Less than M or about 10 -13 Dissociation constant (K D ) and binds to Clec12A.

[0019] In some embodiments, K D is less than 0.1 pM.

[0020] In some embodiments, K D is 0.05 to 0.5 pM.

[0021] In some embodiments, K D is 0.1 nM to 5.0 nM.

[0022] In some embodiments, K D is 0.3nM to 3.5nM.

[0023] In some embodiments, the EC50 is from about 1 to about 100 nM.

[0024] In some embodiments, a method of treating cancer is provided comprising administering an anti-Clec12A antibody or antigen-binding fragment thereof described herein.

[0025] In some embodiments, the cancer is selected from leukemia, lymphoma, and myeloma.

[0026] In some aspects, a pharmaceutical composition is provided comprising an anti-Clec12A antibody or antigen-binding fragment thereof and a pharma- ceutically acceptable carrier, wherein the anti-Clec12A antibody or antigen-binding fragment thereof comprises: (a) an immunoglobulin heavy chain variable (VH) region comprising an amino acid sequence at least about 80% identical to QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYYWSWIRQPPGKGLEWIGYIYYSGSTKYNPSLKSRVTISVDTSKNLFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSS (SEQ ID NO: 1); and an immunoglobulin light chain variable (VL) region comprising an amino acid sequence at least about 80% identical to IQMTQSPSSLSASVGDRVTITCRASQGIRYDLGWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO:2); (b) an immunoglobulin heavy chain variable (VH) region comprising an amino acid sequence at least about 80% identical to QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYYVDSVKGRFTISRDNSKNTLYLHMNSLRAEDTAVYYCARDKGYYFDYWGQGTLVTVSS (SEQ ID NO: 3); and an immunoglobulin light chain variable (VL) region comprising an amino acid sequence at least about 80% identical to EIVMTQSPATLSLSPGERATLSCRASQSVGNRYLSWYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO:4); (c) an immunoglobulin heavy chain variable (VH) region comprising an amino acid sequence at least about 80% identical to QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYSADSVKGRFNISRDNSKNTLYLQMNSLRAEDTAVYYCARDGSRYFDYWGQGTLVTVSS (SEQ ID NO: 5); and An immunoglobulin light chain variable (VL) region comprising an amino acid sequence at least about 80% identical to EIFMTQSPATLSLSPGERATLSCRASQSVHSKYLSWYQQKPGQAPSLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPITFGQGTRLEIK (SEQ ID NO: 6); or (d) an immunoglobulin heavy chain variable (VH) region comprising an amino acid sequence at least about 80% identical to QLQLQESGPGLVKPSETLSLTCTVSGGSISSSTYYYWGWIRQPPRKGLEWIGSTHYRGSTYYNPSLKSRVTISVDTSKNQFSLKVSSVTAADTAVYYCARELTGEVFDYWGQGTLVTVSS (SEQ ID NO: 7); and an immunoglobulin light chain variable (VL) region comprising an amino acid sequence at least about 80% identical to DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSYSTPFTFGPGTKVDIK (SEQ ID NO: 8); (e) an immunoglobulin heavy chain variable (VH) region comprising an amino acid sequence at least about 80% identical to QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYYWSWIRQPPGKGLEWLGYIYFSGSTNYNPSLKSRLTISVAASKSQFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSS (SEQ ID NO: 9); and an immunoglobulin light chain variable (VL) region comprising an amino acid sequence at least about 80% identical to AIQMTQSPSSLSASVGDRVTITCRASQGIRNDLGWFQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTYFTLTISSLQPEDSATYYCLQDYNYPRTFGQGTKVEIK (SEQ ID NO: 10); (f) An immunoglobulin heavy chain variable (VH) region comprising an amino acid sequence that is at least about 80% identical to QVQLQESGPGLVKPSETLSLTCTVSGGSISTDYWSWIRQPPGKGLEWIGYIYFSGSTKYNPSLKSRVTISVDTSKNQFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSS (SEQ ID NO: 11); and an immunoglobulin light chain variable (VL) region comprising an amino acid sequence at least about 80% identical to AIQMTQSPSSLSASVGDRVTITCRASQDIRNDLGWFQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO: 12); (g) an immunoglobulin heavy chain variable (VH) region comprising an amino acid sequence at least about 80% identical to QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGEGLEWVTVISYDGSDKYYADSVKGRFTISRDNSKSTLFLQMNSLRAEDTAVYYCARDGQFYFDYWGQGTLVTVSS (SEQ ID NO: 13); and an immunoglobulin light chain variable (VL) region comprising an amino acid sequence at least about 80% identical to EIVMTQSPATLSLSPGESATLSCRASQSVTSRYLSWYQQKPGQAPRLLMYGASTRPTGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO: 14); (h) an immunoglobulin heavy chain variable (VH) region comprising an amino acid sequence at least about 80% identical to QVQLVESGGGVVQPGRSLRLSCAASGFTFSNYGMHWVRQAPGKGLEWVAVISYDGSDKSYKDSVKGRFTIARDNSKNTLYLQMNSLRAEDTAVYYCARDSGRYFFDYWGQGTLVTVSS (SEQ ID NO: 15); and An immunoglobulin light chain variable (VL) region comprising an amino acid sequence at least about 80% identical to EIIMTQSPATLSLSPGERATLSCRASQSVSSRSLSWYQHKPGQAPRLLIYGPSTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCHQDYNLPLTFGGGTKVEIK (SEQ ID NO: 16); or (i) an immunoglobulin heavy chain variable (VH) region comprising an amino acid sequence at least about 80% identical to QVKLVESGGGVVQPGRSLRLSCAASGFTFSKYGMHWVRQAPGKGLEWVAFIWYDGSIKNYADSVKGRFTTSRDNSKNTLYLQMNSLRAEDTAVYYCARGSLWFGEFYFDYWGQGTLVTVSS (SEQ ID NO: 17); and An immunoglobulin light chain variable (VL) region comprising an amino acid sequence that is at least about 80% identical to AIQLTQSPSSLSASVGDRVTITCRTSQGISSALAWYQQKPGKTPKLLIYDASSLESGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQFNNYPRTFGQGTKVEIK (SEQ ID NO: 18).

[0027] In some embodiments, the pharmaceutical composition comprises an anti-Clec12A antibody or antigen-binding fragment thereof comprising: (a) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYYWSWIRQPPGKGLEWIGYIYYSGSTKYNPSLKSRVTISVDTSKNLFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSS (SEQ ID NO: 1); and An immunoglobulin light chain variable (VL) region comprising the amino acid sequence of IQMTQSPSSLSASVGDRVTITCRASQGIRYDLGWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO:2); (b) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYYVDSVKGRFTISRDNSKNTLYLHMNSLRAEDTAVYYCARDKGYYFDYWGQGTLVTVSS (SEQ ID NO: 3); and An immunoglobulin light chain variable (VL) region comprising the amino acid sequence of EIVMTQSPATLSLSPGERATLSCRASQSVGNRYLSWYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO: 4); (c) an immunoglobulin heavy chain variable (VH) region comprising an amino acid sequence that is 80% identical to QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYSADSVKGRFNISRDNSKNTLYLQMNSLRAEDTAVYYCARDGSRYFDYWGQGTLVTVSS (SEQ ID NO: 5); and An immunoglobulin light chain variable (VL) region comprising the amino acid sequence of: EIFMTQSPATLSLSPGERATLSCRASQSVHSKYLSWYQQKPGQAPSLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPITFGQGTRLEIK (SEQ ID NO: 6); or (d) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QLQLQESGPGLVKPSETLSLTCTVSGGSISSSTYYYWGWIRQPPRKGLEWIGSTHYRGSTYYNPSLKSRVTISVDTSKNQFSLKVSSVTAADTAVYYCARELTGEVFDYWGQGTLVTVSS (SEQ ID NO: 7); and an immunoglobulin light chain variable (VL) region comprising the amino acid sequence of DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSYSTPFTFGPGTKVDIK (SEQ ID NO: 8); (e) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYYWSWIRQPPGKGLEWLGYIYFSGSTNYNPSLKSRLTISVAASKSQFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSS (SEQ ID NO: 9); and an immunoglobulin light chain variable (VL) region comprising the amino acid sequence of AIQMTQSPSSLSASVGDRVTITCRASQGIRNDLGWFQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTYFTLTISSLQPEDSATYYCLQDYNYPRTFGQGTKVEIK (SEQ ID NO: 10); (f) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLQESGPGLVKPSETLSLTCTVSGGSISTDYWSWIRQPPGKGLEWIGYIYFSGSTKYNPSLKSRVTISVDTSKNQFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSS (SEQ ID NO: 11); and an immunoglobulin light chain variable (VL) region comprising the amino acid sequence of AIQMTQSPSSLSASVGDRVTITCRASQDIRNDLGWFQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO: 12); (g) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGEGLEWVTVISYDGSDKYYADSVKGRFTISRDNSKSTLFLQMNSLRAEDTAVYYCARDGQFYFDYWGQGTLVTVSS (SEQ ID NO: 13); and An immunoglobulin light chain variable (VL) region comprising the amino acid sequence of EIVMTQSPATLSLSPGESATLSCRASQSVTSRYLSWYQQKPGQAPRLLMYGASTRPTGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO: 14); (h) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLVESGGGVVQPGRSLRLSCAASGFTFSNYGMHWVRQAPGKGLEWVAVISYDGSDKSYKDSVKGRFTIARDNSKNTLYLQMNSLRAEDTAVYYCARDSGRYFFDYWGQGTLVTVSS (SEQ ID NO: 15); and An immunoglobulin light chain variable (VL) region comprising the amino acid sequence of EIIMTQSPATLSLSPGERATLSCRASQSVSSRSLSWYQHKPGQAPRLLIYGPSTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCHQDYNLPLTFGGGTKVEIK (SEQ ID NO: 16); or (i) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVKLVESGGGVVQPGRSLRLSCAASGFTFSKYGMHWVRQAPGKGLEWVAFIWYDGSIKNYADSVKGRFTTSRDNSKNTLYLQMNSLRAEDTAVYYCARGSLWFGEFYFDYWGQGTLVTVSS (SEQ ID NO: 17); and An immunoglobulin light chain variable (VL) region comprising the amino acid sequence of AIQLTQSPSSLSASVGDRVTITCRTSQGISSALAWYQQKPGKTPKLLIYDASSLESGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQFNNYPRTFGQGTKVEIK (SEQ ID NO: 18).

[0028] In some embodiments, the pharmaceutical composition comprises an anti-Clec12A antibody or antigen-binding fragment thereof comprising: (a) a heavy chain complementarity determining region (HCDR) 1 comprising the amino acid sequence of GGSISTYY (SEQ ID NO: 19), an HCDR2 comprising the amino acid sequence of IYYSGST (SEQ ID NO: 20), and an HCDR3 comprising the amino acid sequence of AREDYYGSGSPFDY (SEQ ID NO: 21); and a light chain complementarity determining region (LCDR) 1 comprising the amino acid sequence of QGIRYD (SEQ ID NO: 22), an LCDR2 comprising the amino acid sequence of AAS (SEQ ID NO: 23), and an LCDR3 comprising the amino acid sequence of LQDYNFPRT (SEQ ID NO: 24); (b) an HCDR1 comprising the amino acid sequence of GFTFSSYG (SEQ ID NO: 73), an HCDR2 comprising the amino acid sequence of ISYDGSDK (SEQ ID NO: 25), and an HCDR3 comprising the amino acid sequence of ARDKGYYFDY (SEQ ID NO: 26); and LCDR1 comprising the amino acid sequence of QSVGNRY (SEQ ID NO: 27), LCDR2 comprising the amino acid sequence of GAS (SEQ ID NO: 28), LCDR3 comprising the amino acid sequence of QQDYNLPLT (SEQ ID NO: 29); (c) HCDR1 comprising the amino acid sequence of GFTFSSYG (SEQ ID NO: 73), HCDR2 comprising the amino acid sequence of ISYDGSDK (SEQ ID NO: 25), and HCDR3 comprising the amino acid sequence of ARDGSRYFDY (SEQ ID NO: 30); and LCDR1 comprising the amino acid sequence of QSVHSKY (SEQ ID NO: 31), LCDR2 comprising the amino acid sequence of GAS (SEQ ID NO: 28), LCDR3 comprising the amino acid sequence of QQDYNLPIT (SEQ ID NO: 32); and (d) HCDR1 comprising the amino acid sequence of GGSISSSTYY (SEQ ID NO: 33), HCDR2 comprising the amino acid sequence of THYRGST (SEQ ID NO: 34), and HCDR3 comprising the amino acid sequence of ARELTGEVFDY (SEQ ID NO: 35); and LCDR1 comprising the amino acid sequence of QSISSY (SEQ ID NO: 36), LCDR2 comprising the amino acid sequence of AAS (SEQ ID NO: 23), and LCDR3 comprising the amino acid sequence of QQSYSTPFT (SEQ ID NO: 37); (e) HCDR1 comprising the amino acid sequence of GGSISTYY (SEQ ID NO: 19), HCDR2 comprising the amino acid sequence of IYFSGST (SEQ ID NO: 38), and HCDR3 comprising the amino acid sequence of AREDYYGSGSPFDY (SEQ ID NO: 21); and LCDR1 comprising the amino acid sequence of QGIRND (SEQ ID NO: 39), LCDR2 comprising the amino acid sequence of AAS (SEQ ID NO: 23), LCDR3 comprising the amino acid sequence of LQDYNYPRT (SEQ ID NO: 40); (f) HCDR1 comprising the amino acid sequence of GGSISTDY (SEQ ID NO: 41), HCDR2 comprising the amino acid sequence of IYFSGST (SEQ ID NO: 38), and HCDR3 comprising the amino acid sequence of AREDYYGSGSPFDY (SEQ ID NO: 21); and LCDR1 comprising the amino acid sequence of QDIRND (SEQ ID NO: 42), LCDR2 comprising the amino acid sequence of AAS (SEQ ID NO: 23), LCDR3 comprising the amino acid sequence of LQDYNFPRT (SEQ ID NO: 24); (g) HCDR1 comprising the amino acid sequence of GFTFSSYG (SEQ ID NO: 73), HCDR2 comprising the amino acid sequence of ISYDGSDK (SEQ ID NO: 25), and HCDR3 comprising the amino acid sequence of ARDGQFYFDY (SEQ ID NO: 43); and LCDR1 comprising the amino acid sequence of QSVTSRY (SEQ ID NO: 44), LCDR2 comprising the amino acid sequence of GAS (SEQ ID NO: 28), LCDR3 comprising the amino acid sequence of QQDYNLPLT (SEQ ID NO: 29); (h) HCDR1 comprising the amino acid sequence of GFTFSNYG (SEQ ID NO: 45), HCDR2 comprising the amino acid sequence of ISYDGSDK (SEQ ID NO: 25), and HCDR3 comprising the amino acid sequence of ARDSGRYFFDY (SEQ ID NO: 46); and LCDR1 comprising the amino acid sequence of QSVSSRS (SEQ ID NO: 47), LCDR2 comprising the amino acid sequence of GPS (SEQ ID NO: 48), LCDR3 comprising the amino acid sequence of HQDYNLPLT (SEQ ID NO: 49); or (i) an HCDR1 comprising the amino acid sequence of GFTFSKYG (SEQ ID NO: 50), an HCDR2 comprising the amino acid sequence of IWYDGSIK (SEQ ID NO: 51), and an HCDR3 comprising the amino acid sequence of ARGSLWFGEFYFDY (SEQ ID NO: 52); and LCDR1 comprising the amino acid sequence of QGISSA (sequence number 53), LCDR2 comprising the amino acid sequence of DAS (sequence number 54), and LCDR3 comprising the amino acid sequence of QQFNNYPRT (sequence number 55).

[0029] In some embodiments, the pharmaceutical composition comprises an anti-Clec12A antibody or antigen-binding fragment thereof selected from the group consisting of an IgA antibody, an IgG antibody, an IgE antibody, an IgM antibody, a bispecific or multispecific antibody, a Fab fragment, a Fab' fragment, a F(ab')2 fragment, an Fd' fragment, an Fd fragment, an isolated CDR or set thereof; a single chain variable fragment (scFv), a polypeptide-Fc fusion, a single domain antibody, a camelid antibody, a masked antibody, a small modular immunopharmaceutical ("SMIPs™"), a single chain, a tandem diabody, a VHH, anticalin, a nanobody, a minibody, a BiTE, an ankyrin repeat protein, a DARPIN, an Avimer, a DART, a TCR-like antibody, an adnectin, an affilin, a transbody, an affibody, a TrimerX, a MicroProtein, a Fynomer, a Centyrin, and a KALBITOR.

[0030] In some embodiments, the pharmaceutical composition comprises an anti-Clec12A antibody or antigen-binding fragment as a monoclonal antibody or single-chain variable fragment (scFv).

[0031] In some embodiments, the anti-Clec12A antibody is an antibody that comprises an IgG constant region.

[0032] In some embodiments, the anti-Clec12A antigen-binding fragment is a single chain variable fragment (scFv).

[0033] In some embodiments, the scFv comprises a leader sequence, a heavy chain variable sequence, a GS linker, and a light chain variable sequence.

[0034] In some embodiments, the pharmaceutical compositions described herein comprise an scFv comprising a sequence having an amino acid sequence that is at least about 80% identical to: (a) QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYWSWIRQPPGKGLEWIGYIYYSGSTKYNPSLKSRVTISVDTSKNLFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSAIQMTQSPSSLSASVGDRVTITCRASQGIRYDLGWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO:56); (b) QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAIVISYDGSDKYYVDSVKGRFTISRDNSKNTLYLHMNSLRAEDTAVYYCARDKGYYFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSEIVMTQSPATLSLSPGERATLSCRASQSVGNRYLSWYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO:57); (c) QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYSADSVKGRFNISRDNSKNTLYLQMNSLRAEDTAVYYCARDGSRYFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSEIFMTQSPATLSLPGERATLSCRASQSVHSKYLSWYQQKPGQAPSLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPITFGQGTRLEIK (SEQ ID NO: 58); or (d) QLQLQESGPGLVKPSETLSLTCTVSGGSISSSTYYWGWIRQPPRKGLEWIGSTHYRGSTYYNPSLKSRVTISVDTSKNQFSLKVSSVTAADTAVYYCARELTGEVFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSYSTPFTFGPGTKVDIK (SEQ ID NO: 59).

[0035] In some embodiments, the pharmaceutical composition comprises an scFv comprising the following sequence: (a) QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYWSWIRQPPGKGLEWIGYIYYSGSTKYNPSLKSRVTISVDTSKNLFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSAIQMTQSPSSLSASVGDRVTITCRASQGIRYDLGWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO:56); (b) QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAIVISYDGSDKYYVDSVKGRFTISRDNSKNTLYLHMNSLRAEDTAVYYCARDKGYYFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSEIVMTQSPATLSLSPGERATLSCRASQSVGNRYLSWYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO:57); (c) QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYSADSVKGRFNISRDNSKNTLYLQMNSLRAEDTAVYYCARDGSRYFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSEIFMTQSPATLSLPGERATLSCRASQSVHSKYLSWYQQKPGQAPSLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPITFGQGTRLEIK (SEQ ID NO: 58); or (d) QLQLQESGPGLVKPSETLSLTCTVSGGSISSSTYYWGWIRQPPRKGLEWIGSTHYRGSTYYNPSLKSRVTISVDTSKNQFSLKVSSVTAADTAVYYCARELTGEVFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSYSTPFTFGPGTKVDIK (SEQ ID NO: 59).

[0036] In some embodiments, a nucleic acid sequence is provided that encodes an amino acid sequence that is at least about 90% identical to any one of SEQ ID NOs: 1-59.

[0037] In some embodiments, a vector is provided that comprises a nucleic acid sequence that encodes an amino acid sequence that is at least about 90% identical to any one of SEQ ID NOs: 1-59.

[0038] In some embodiments, a cell is provided that comprises a vector described herein.

[0039] In some embodiments, there is provided a method of treating cancer comprising administering to a subject in need thereof an anti-Clec12A antibody or antigen-binding fragment thereof, wherein the anti-Clec12A antibody or antigen-binding fragment thereof comprises: (a) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYYWSWIRQPPGKGLEWIGYIYYSGSTKYNPSLKSRVTISVDTSKNLFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSS (SEQ ID NO: 1); and An immunoglobulin light chain variable (VL) region comprising the amino acid sequence of IQMTQSPSSLSASVGDRVTITCRASQGIRYDLGWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO:2); (b) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYYVDSVKGRFTISRDNSKNTLYLHMNSLRAEDTAVYYCARDKGYYFDYWGQGTLVTVSS (SEQ ID NO: 3); and An immunoglobulin light chain variable (VL) region comprising the amino acid sequence of EIVMTQSPATLSLSPGERATLSCRASQSVGNRYLSWYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO: 4); (c) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYSADSVKGRFNISRDNSKNTLYLQMNSLRAEDTAVYYCARDGSRYFDYWGQGTLVTVSS (SEQ ID NO: 5); and An immunoglobulin light chain variable (VL) region comprising the amino acid sequence of: EIFMTQSPATLSLSPGERATLSCRASQSVHSKYLSWYQQKPGQAPSLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPITFGQGTRLEIK (SEQ ID NO: 6); or (d) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QLQLQESGPGLVKPSETLSLTCTVSGGSISSSTYYYWGWIRQPPRKGLEWIGSTHYRGSTYYNPSLKSRVTISVDTSKNQFSLKVSSVTAADTAVYYCARELTGEVFDYWGQGTLVTVSS (SEQ ID NO: 7); and an immunoglobulin light chain variable (VL) region comprising the amino acid sequence of DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSYSTPFTFGPGTKVDIK (SEQ ID NO: 8); (e) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYYWSWIRQPPGKGLEWLGYIYFSGSTNYNPSLKSRLTISVAASKSQFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSS (SEQ ID NO: 9); and an immunoglobulin light chain variable (VL) region comprising the amino acid sequence of AIQMTQSPSSLSASVGDRVTITCRASQGIRNDLGWFQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTYFTLTISSLQPEDSATYYCLQDYNYPRTFGQGTKVEIK (SEQ ID NO: 10); (f) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLQESGPGLVKPSETLSLTCTVSGGSISTDYWSWIRQPPGKGLEWIGYIYFSGSTKYNPSLKSRVTISVDTSKNQFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSS (SEQ ID NO: 11); and an immunoglobulin light chain variable (VL) region comprising the amino acid sequence of AIQMTQSPSSLSASVGDRVTITCRASQDIRNDLGWFQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO: 12); (g) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGEGLEWVTVISYDGSDKYYADSVKGRFTISRDNSKSTLFLQMNSLRAEDTAVYYCARDGQFYFDYWGQGTLVTVSS (SEQ ID NO: 13); and An immunoglobulin light chain variable (VL) region comprising the amino acid sequence of EIVMTQSPATLSLSPGESATLSCRASQSVTSRYLSWYQQKPGQAPRLLMYGASTRPTGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO: 14); (h) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLVESGGGVVQPGRSLRLSCAASGFTFSNYGMHWVRQAPGKGLEWVAVISYDGSDKSYKDSVKGRFTIARDNSKNTLYLQMNSLRAEDTAVYYCARDSGRYFFDYWGQGTLVTVSS (SEQ ID NO: 15); and An immunoglobulin light chain variable (VL) region comprising the amino acid sequence of EIIMTQSPATLSLSPGERATLSCRASQSVSSRSLSWYQHKPGQAPRLLIYGPSTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCHQDYNLPLTFGGGTKVEIK (SEQ ID NO: 16); or (i) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVKLVESGGGVVQPGRSLRLSCAASGFTFSKYGMHWVRQAPGKGLEWVAFIWYDGSIKNYADSVKGRFTTSRDNSKNTLYLQMNSLRAEDTAVYYCARGSLWFGEFYFDYWGQGTLVTVSS (SEQ ID NO: 17); and An immunoglobulin light chain variable (VL) region comprising the amino acid sequence of AIQLTQSPSSLSASVGDRVTITCRTSQGISSALAWYQQKPGKTPKLLIYDASSLESGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQFNNYPRTFGQGTKVEIK (SEQ ID NO: 18).

[0040] In some embodiments, the method of treating cancer comprises administering to a subject in need thereof an anti-Clec12A antibody or antigen-binding fragment thereof, comprising: (a) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYYWSWIRQPPGKGLEWIGYIYYSGSTKYNPSLKSRVTISVDTSKNLFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSS (SEQ ID NO: 1); and An immunoglobulin light chain variable (VL) region comprising the amino acid sequence of IQMTQSPSSLSASVGDRVTITCRASQGIRYDLGWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO:2); (b) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYYVDSVKGRFTISRDNSKNTLYLHMNSLRAEDTAVYYCARDKGYYFDYWGQGTLVTVSS (SEQ ID NO: 3); and An immunoglobulin light chain variable (VL) region comprising the amino acid sequence of EIVMTQSPATLSLSPGERATLSCRASQSVGNRYLSWYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO: 4); (c) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYSADSVKGRFNISRDNSKNTLYLQMNSLRAEDTAVYYCARDGSRYFDYWGQGTLVTVSS (SEQ ID NO: 5); and An immunoglobulin light chain variable (VL) region comprising the amino acid sequence of: EIFMTQSPATLSLSPGERATLSCRASQSVHSKYLSWYQQKPGQAPSLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPITFGQGTRLEIK (SEQ ID NO: 6); or (d) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QLQLQESGPGLVKPSETLSLTCTVSGGSISSSTYYYWGWIRQPPRKGLEWIGSTHYRGSTYYNPSLKSRVTISVDTSKNQFSLKVSSVTAADTAVYYCARELTGEVFDYWGQGTLVTVSS (SEQ ID NO: 7); and an immunoglobulin light chain variable (VL) region comprising the amino acid sequence of DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSYSTPFTFGPGTKVDIK (SEQ ID NO: 8); (e) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYYWSWIRQPPGKGLEWLGYIYFSGSTNYNPSLKSRLTISVAASKSQFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSS (SEQ ID NO: 9); and an immunoglobulin light chain variable (VL) region comprising the amino acid sequence of AIQMTQSPSSLSASVGDRVTITCRASQGIRNDLGWFQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTYFTLTISSLQPEDSATYYCLQDYNYPRTFGQGTKVEIK (SEQ ID NO: 10); (f) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLQESGPGLVKPSETLSLTCTVSGGSISTDYWSWIRQPPGKGLEWIGYIYFSGSTKYNPSLKSRVTISVDTSKNQFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSS (SEQ ID NO: 11); and an immunoglobulin light chain variable (VL) region comprising the amino acid sequence of AIQMTQSPSSLSASVGDRVTITCRASQDIRNDLGWFQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO: 12); (g) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGEGLEWVTVISYDGSDKYYADSVKGRFTISRDNSKSTLFLQMNSLRAEDTAVYYCARDGQFYFDYWGQGTLVTVSS (SEQ ID NO: 13); and An immunoglobulin light chain variable (VL) region comprising the amino acid sequence of EIVMTQSPATLSLSPGESATLSCRASQSVTSRYLSWYQQKPGQAPRLLMYGASTRPTGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO: 14); (h) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVQLVESGGGVVQPGRSLRLSCAASGFTFSNYGMHWVRQAPGKGLEWVAVISYDGSDKSYKDSVKGRFTIARDNSKNTLYLQMNSLRAEDTAVYYCARDSGRYFFDYWGQGTLVTVSS (SEQ ID NO: 15); and An immunoglobulin light chain variable (VL) region comprising the amino acid sequence of EIIMTQSPATLSLSPGERATLSCRASQSVSSRSLSWYQHKPGQAPRLLIYGPSTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCHQDYNLPLTFGGGTKVEIK (SEQ ID NO: 16); or (i) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QVKLVESGGGVVQPGRSLRLSCAASGFTFSKYGMHWVRQAPGKGLEWVAFIWYDGSIKNYADSVKGRFTTSRDNSKNTLYLQMNSLRAEDTAVYYCARGSLWFGEFYFDYWGQGTLVTVSS (SEQ ID NO: 17); and An immunoglobulin light chain variable (VL) region comprising the amino acid sequence of AIQLTQSPSSLSASVGDRVTITCRTSQGISSALAWYQQKPGKTPKLLIYDASSLESGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQFNNYPRTFGQGTKVEIK (SEQ ID NO: 18).

[0041] In some embodiments, the method of treating cancer comprises administering to a subject in need thereof an anti-Clec12A antibody or fragment thereof, comprising: (a) a heavy chain complementarity determining region (HCDR) 1 comprising the amino acid sequence of GGSISTYY (SEQ ID NO: 19), an HCDR2 comprising the amino acid sequence of IYYSGST (SEQ ID NO: 20), and an HCDR3 comprising the amino acid sequence of AREDYYGSGSPFDY (SEQ ID NO: 21); and a light chain complementarity determining region (LCDR) 1 comprising the amino acid sequence of QGIRYD (SEQ ID NO: 22), an LCDR2 comprising the amino acid sequence of AAS (SEQ ID NO: 23), and an LCDR3 comprising the amino acid sequence of LQDYNFPRT (SEQ ID NO: 24); (b) an HCDR1 comprising the amino acid sequence of GFTFSSYG (SEQ ID NO: 73), an HCDR2 comprising the amino acid sequence of ISYDGSDK (SEQ ID NO: 25), and an HCDR3 comprising the amino acid sequence of ARDKGYYFDY (SEQ ID NO: 26); and LCDR1 comprising the amino acid sequence of QSVGNRY (SEQ ID NO: 27), LCDR2 comprising the amino acid sequence of GAS (SEQ ID NO: 28), LCDR3 comprising the amino acid sequence of QQDYNLPLT (SEQ ID NO: 29); (c) HCDR1 comprising the amino acid sequence of GFTFSSYG (SEQ ID NO: 73), HCDR2 comprising the amino acid sequence of ISYDGSDK (SEQ ID NO: 25), and HCDR3 comprising the amino acid sequence of ARDGSRYFDY (SEQ ID NO: 30); and LCDR1 comprising the amino acid sequence of QSVHSKY (SEQ ID NO: 31), LCDR2 comprising the amino acid sequence of GAS (SEQ ID NO: 28), LCDR3 comprising the amino acid sequence of QQDYNLPIT (SEQ ID NO: 32); and (d) HCDR1 comprising the amino acid sequence of GGSISSSTYY (SEQ ID NO: 33), HCDR2 comprising the amino acid sequence of THYRGST (SEQ ID NO: 34), and HCDR3 comprising the amino acid sequence of ARELTGEVFDY (SEQ ID NO: 35); and LCDR1 comprising the amino acid sequence of QSISSY (SEQ ID NO: 36), LCDR2 comprising the amino acid sequence of AAS (SEQ ID NO: 23), and LCDR3 comprising the amino acid sequence of QQSYSTPFT (SEQ ID NO: 37); (e) HCDR1 comprising the amino acid sequence of GGSISTYY (SEQ ID NO: 19), HCDR2 comprising the amino acid sequence of IYFSGST (SEQ ID NO: 38), and HCDR3 comprising the amino acid sequence of AREDYYGSGSPFDY (SEQ ID NO: 21); and LCDR1 comprising the amino acid sequence of QGIRND (SEQ ID NO: 39), LCDR2 comprising the amino acid sequence of AAS (SEQ ID NO: 23), LCDR3 comprising the amino acid sequence of LQDYNYPRT (SEQ ID NO: 40); (f) HCDR1 comprising the amino acid sequence of GGSISTDY (SEQ ID NO: 41), HCDR2 comprising the amino acid sequence of IYFSGST (SEQ ID NO: 38), and HCDR3 comprising the amino acid sequence of AREDYYGSGSPFDY (SEQ ID NO: 21); and LCDR1 comprising the amino acid sequence of QDIRND (SEQ ID NO: 42), LCDR2 comprising the amino acid sequence of AAS (SEQ ID NO: 23), LCDR3 comprising the amino acid sequence of LQDYNFPRT (SEQ ID NO: 24); (g) HCDR1 comprising the amino acid sequence of GFTFSSYG (SEQ ID NO: 73), HCDR2 comprising the amino acid sequence of ISYDGSDK (SEQ ID NO: 25), and HCDR3 comprising the amino acid sequence of ARDGQFYFDY (SEQ ID NO: 43); and LCDR1 comprising the amino acid sequence of QSVTSRY (SEQ ID NO: 44), LCDR2 comprising the amino acid sequence of GAS (SEQ ID NO: 28), LCDR3 comprising the amino acid sequence of QQDYNLPLT (SEQ ID NO: 29); (h) HCDR1 comprising the amino acid sequence of GFTFSNYG (SEQ ID NO: 45), HCDR2 comprising the amino acid sequence of ISYDGSDK (SEQ ID NO: 25), and HCDR3 comprising the amino acid sequence of ARDSGRYFFDY (SEQ ID NO: 46); and LCDR1 comprising the amino acid sequence of QSVSSRS (SEQ ID NO: 47), LCDR2 comprising the amino acid sequence of GPS (SEQ ID NO: 48), LCDR3 comprising the amino acid sequence of HQDYNLPLT (SEQ ID NO: 49); or (i) an HCDR1 comprising the amino acid sequence of GFTFSKYG (SEQ ID NO: 50), an HCDR2 comprising the amino acid sequence of IWYDGSIK (SEQ ID NO: 51), and an HCDR3 comprising the amino acid sequence of ARGSLWFGEFYFDY (SEQ ID NO: 52); and LCDR1 comprising the amino acid sequence of QGISSA (sequence number 53), LCDR2 comprising the amino acid sequence of DAS (sequence number 54), and LCDR3 comprising the amino acid sequence of QQFNNYPRT (sequence number 55).

[0042] In some embodiments, the anti-Clec12A antibody or antigen-binding fragment thereof is a monoclonal antibody or a single chain variable fragment (scFv).

[0043] In some embodiments, the anti-Clec12A antibody is an antibody that comprises an IgG constant region.

[0044] In some embodiments, the antigen-binding fragment is a single chain variable fragment (scFv).

[0045] In some embodiments, the scFv comprises a leader sequence, a heavy chain variable sequence, a GS linker, and a light chain variable sequence.

[0046] In some embodiments, the method of treating cancer comprises administering to a subject in need thereof an scFv comprising a sequence having at least about 80% identity to: (a) QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYWSWIRQPPGKGLEWIGYIYYSGSTKYNPSLKSRVTISVDTSKNLFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSAIQMTQSPSSLSASVGDRVTITCRASQGIRYDLGWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO:56); (b) QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAIVISYDGSDKYYVDSVKGRFTISRDNSKNTLYLHMNSLRAEDTAVYYCARDKGYYFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSEIVMTQSPATLSLSPGERATLSCRASQSVGNRYLSWYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO:57); (c) QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYSADSVKGRFNISRDNSKNTLYLQMNSLRAEDTAVYYCARDGSRYFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSEIFMTQSPATLSLPGERATLSCRASQSVHSKYLSWYQQKPGQAPSLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPITFGQGTRLEIK (SEQ ID NO: 58); or (d) QLQLQESGPGLVKPSETLSLTCTVSGGSISSSTYYWGWIRQPPRKGLEWIGSTHYRGSTYYNPSLKSRVTISVDTSKNQFSLKVSSVTAADTAVYYCARELTGEVFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSYSTPFTFGPGTKVDIK (SEQ ID NO: 59).

[0047] In some embodiments, the method of treating cancer comprises administering to a subject in need thereof an scFv comprising the following sequence: (a) QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYWSWIRQPPGKGLEWIGYIYYSGSTKYNPSLKSRVTISVDTSKNLFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSAIQMTQSPSSLSASVGDRVTITCRASQGIRYDLGWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO:56); (b) QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAIVISYDGSDKYYVDSVKGRFTISRDNSKNTLYLHMNSLRAEDTAVYYCARDKGYYFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSEIVMTQSPATLSLSPGERATLSCRASQSVGNRYLSWYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO:57); (c) QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYSADSVKGRFNISRDNSKNTLYLQMNSLRAEDTAVYYCARDGSRYFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSEIFMTQSPATLSLPGERATLSCRASQSVHSKYLSWYQQKPGQAPSLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPITFGQGTRLEIK (SEQ ID NO: 58); or (d) QLQLQESGPGLVKPSETLSLTCTVSGGSISSSTYYWGWIRQPPRKGLEWIGSTHYRGSTYYNPSLKSRVTISVDTSKNQFSLKVSSVTAADTAVYYCARELTGEVFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSYSTPFTFGPGTKVDIK (SEQ ID NO: 59).

[0048] In some embodiments, the cancer is selected from leukemia, lymphoma, and myeloma.

[0049] In some embodiments, the treatment comprises administration of one or more additional agents.

[0050] In some embodiments, the one or more additional agents are selected from an antibody, a chemotherapeutic agent, and radiation therapy.

[0051] definition "A" or "An": The articles "a" and "an" are used herein to refer to one or to more than one (i.e., to at least one) of the grammatical object of the article. By way of example, "an element" means one element or more than two elements.

[0052] Affinity: As used herein, the term "affinity" refers to a characteristic of the binding interaction between a binding moiety (e.g., an antigen binding moiety (e.g., a variable domain described herein) and / or an Fc receptor binding moiety (e.g., an FcRn binding moiety described herein)) and a target (e.g., an antigen (e.g., Clec12A) and / or an FcR (e.g., FcRn)), which indicates the strength of the binding interaction. In some embodiments, a measure of affinity is the dissociation constant (K D In some embodiments, the binding moiety has a high affinity for the target (e.g., about 10 -7 Less than M, about 10 -8 Less than M or about 10 -9 K under M D In some embodiments, the binding moiety has a low affinity for the target (e.g., about 10 -7 Higher than M, about 10 -6 Higher than M, about 10 -5 Higher than M or about 10 -4 K is higher than M D In some embodiments, the binding moiety has a high affinity for the target at a first pH, a low affinity for the target at a second pH, and an intermediate affinity for the target at pH levels between the first and second pH.

[0053] Approximately or about: As used herein, when applied to one or more values ​​of interest, the term "approximately" or "about" refers to a value similar to the reference value mentioned. In certain embodiments, the term "approximately" or "about" refers to a range of values ​​within a range of 25%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1% or less in either direction (greater or smaller) of the reference value mentioned, unless otherwise specified or clear from the context (except when such number exceeds 100% of possible values).

[0054] Antibody: As used herein, the term "antibody" refers to a polypeptide that includes at least one immunoglobulin variable region, e.g., an amino acid sequence that provides an immunoglobulin variable domain or an immunoglobulin variable domain sequence. For example, an antibody can include a heavy (H) chain variable region (abbreviated herein as VH) and a light (L) chain variable region (abbreviated herein as VL). In another example, an antibody includes two heavy (H) chain variable regions and two light (L) chain variable regions. The term "antibody" encompasses antigen-binding fragments of antibodies (e.g., single chain antibodies, Fab, F(ab')2, Fd, Fv, and dAb fragments), as well as complete antibodies, e.g., intact immunoglobulins of the types IgA, IgG, IgE, IgD, IgM (and subtypes thereof). The light chain of the immunoglobulin may be of type kappa or lambda.

[0055] Binding moiety: As used herein, a "binding moiety" is any molecule or portion of a molecule that can specifically bind to a target, e.g., a target of interest (e.g., an antigen (e.g., Clec12A) and / or an FcR (e.g., FcRn)). Binding moieties include, for example, antibodies, antigen-binding fragments thereof, Fc regions or Fc fragments thereof, antibody mimetics, peptides, and aptamers.

[0056] An antibody-binding fragment or antibody fragment thereof refers to a portion of an intact antibody. An antibody-binding fragment or antibody fragment thereof refers to a portion of an intact antibody that binds to an antigen (e.g., Clec12A). An antigen-binding fragment may include the antigen-determining variable region of an intact antibody. Examples of antibody fragments include, but are not limited to, Fab, Fab', F(ab')2, and Fv fragments, linear antibodies, antibody mimetics, scFv, and single chain antibodies.

[0057] Complementarity Determining Region (CDR): The "CDRs" of a variable domain are those amino acid residues within the variable region identified according to Kabat, Chothia, the combined Kabat and Chothia, i.e. AbM, contact and / or conformation definitions, or any method of CDR determination known in the art. The CDRs of an antibody may be identified as hypervariable regions as originally defined by Kabat et al. See, e.g., Kabat et al., 1992, Sequences of Proteins of Immunological Interest, 5th ed., Public Health Service, NIH, Washington DC. The locations of CDRs may also be identified as structural loop structures originally described by Chothia and others. See, e.g., Chothia et al., Nature 342:877-883, 1989. Other approaches to CDR identification include the "AbM definition," a compromise between Kabat and Chothia, derived using Oxford Molecular's AbM antibody modeling software (now Accelrys®), or the "contact definition" of CDRs based on observed antigen contact sites, as described in MacCallum et al., J.Mol.Biol.,262:732-745,1996. In another approach, referred to herein as the "conformational definition" of CDRs, the positions of CDRs may be identified as residues that contribute enthalpic to antigen binding. See, for example, Makabe et al., Journal of Biological Chemistry,283:1 156-1166,2008. Still other CDR boundary definitions may not strictly follow one of the above approaches, but may be shortened or extended in light of predictions or experimental findings that certain residues or groups of residues or even the entire CDR do not significantly affect antigen binding, but still overlap with at least a portion of Kabat's CDR. As used herein, CDR may refer to a CDR defined by any approach known in the art, including a combination of approaches.The methods used herein may utilize CDRs defined according to any of these approaches. For any given embodiment that includes multiple CDRs, the CDRs may be defined according to the IMGT, Kabat, Chothia, extended, AbM, contact, and / or conformational definitions. In certain embodiments, the CDRs are identified or numbered according to the IMGT numbering system.

[0058] Constant region: As used herein, the term "constant region" refers to a polypeptide corresponding to or derived from one or more constant region immunoglobulin domains of an antibody. The constant region can include any or all of the following immunoglobulin domains: CH1 domain, hinge region, CH2 domain, CH3 domain (derived from IgA, IgD, IgG, IgE, or IgM), and CH4 domain (derived from IgE or IgM).

[0059] Epitope: As used herein, "epitope" is a term of the art and refers to a localized region of an antigen to which an antibody can specifically bind. An epitope can be, for example, consecutive amino acids of a polypeptide (linear or consecutive epitope), or an epitope can be, for example, a polypeptide or two or more non-contiguous regions of a polypeptide together (conformational, non-linear, discontinuous, or discontinuous epitope). In certain embodiments, the epitope to which an antibody binds can be determined, for example, by NMR spectroscopy, X-ray diffraction crystallography studies, ELISA assays, hydrogen / deuterium exchange combined with mass spectrometry (e.g., liquid chromatography electrospray mass spectrometry), array-based oligopeptide scanning assays, and / or mutagenesis mapping (e.g., site-directed mutagenesis mapping). In the case of X-ray crystallography, crystallization can be achieved using any method known 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 can be studied using well-known X-ray diffraction techniques and refined using computer software well-known in the art, such as Refmac and Phenix. Mutagenesis mapping studies can be achieved using any method known to those skilled in the art. See, e.g., Champe M et al, (1995) J Biol Chem 270:1388-1394 and Cunningham BC & Wells JA (1989) Science 244:1081-1085 for a description of mutagenesis techniques, including alanine scanning mutagenesis techniques.

[0060] Fc region: As used herein, the term "Fc region" refers to a dimer of two "Fc polypeptides", each of which comprises the constant region of an antibody, excluding the first constant region immunoglobulin domain. In some embodiments, an "Fc region" comprises two Fc polypeptides linked by one or more disulfide bonds, chemical linkers, or peptide linkers. "Fc polypeptide" refers to the last two constant region immunoglobulin domains of IgA, IgD, and IgG, and the last three constant region immunoglobulin domains of IgE and IgM, and may include some or all of the flexible hinge N-terminal to these domains. In the case of IgG, the "Fc polypeptide" includes immunoglobulin domains C gamma 2 (Cγ2) and C gamma 3 (Cγ3), as well as the lower part of the hinge between C gamma 1 (Cγ1) and Cγ2. Although the boundaries of the Fc polypeptide may vary, the human IgG heavy chain Fc polypeptide is usually defined to include the residues from T223 or C226 or P230 to its carboxyl terminus, numbering according to the EU index of Kabat et al. (1991, NIH Publication 91-3242, National Technical Information Services, Springfield, VA). For IgA, the Fc polypeptide includes the immunoglobulin domains C alpha 2 (Cα2) and C alpha 3 (Cα3), as well as the lower part of the hinge between C alpha 1 (Cα1) and Cα2. The Fc region may be synthetic, recombinant, or derived from a natural source such as IVIG.

[0061] K a As used herein, "K a " refers to the rate of association of a particular binding moiety with a target to form a binding moiety / target complex.

[0062] K d As used herein, "K d " refers to the dissociation rate of a particular binding moiety / target complex.

[0063] K DAs used herein, "K D " refers to the dissociation constant, which is K d K a (i.e., K d / K a ) and expressed as molar concentration (M). D The value can be determined using methods well established in the art, for example using surface plasmon resonance or using a biosensor system such as a Biacore® system.

[0064] Reference: A "reference" entity, system, amount, set of conditions, etc., to which a test entity, system, amount, set of conditions, etc., as described herein, is compared. For example, in some embodiments, a "reference" antibody is a control antibody that has not been engineered as described herein.

[0065] Selective binding: As used herein, "selective binding," "selectively binds," "specific binding," or "specifically binds," in reference to a binding moiety and a target, refers to the preferential association of the binding moiety with the target and not with non-target entities. Some degree of non-specific binding may occur between the binding moiety and the non-target. In some embodiments, a binding moiety selectively binds to a target if the binding between the binding moiety and the target is more than 2-fold, more than 5-fold, more than 10-fold, or more than 100-fold greater than the binding between the binding moiety and the non-target. In some embodiments, a binding affinity is greater than about 10 -5 Less than M, about 10 -6 Less than M, about 10 -7 Less than M, about 10 -8 Less than M or about 10 -9A binding moiety selectively binds to a target when M is less than M. In some embodiments, a molecule that specifically binds to an antigen may generally bind to other peptides or polypeptides with lower affinity, as determined, for example, by immunoassays, a BIACORE®, KinExA 3000 instrument (Sapidyne Instruments, Boise, Id.), or other assays known in the art.

[0066] Single-chain variable fragment (scFv): As used herein, the term "single-chain variable fragment" or "scFv" refers to a V H Heavy chains (VL) of immunoglobulins (e.g., mouse or human) covalently linked to form a VL heterodimer. H ) and light chain (V L It refers to a fusion protein of the variable region of the heavy chain (V H ) and light chain (V L ) is directly bonded or V H N-terminus of V L or at the C-terminus of V H The C-terminus of V L The linker is typically glycine-rich for flexibility and serine or threonine-rich for solubility. The linker can link the heavy and light chain variable regions of the extracellular antigen-binding domain. Non-limiting examples of linkers are disclosed in Shen et al., Anal. Chem. 80(6):1910-1917(2008) and WO2014 / 087010, the contents of which are incorporated herein by reference in their entirety.

[0067] Subject: As used herein, the term "subject" refers to any subject for which diagnosis, prognosis, or treatment is desired. For example, the subject can be a mammal, such as a human or non-human primate (such as an ape, monkey, orangutan, or chimpanzee), dog, cat, guinea pig, rabbit, rat, mouse, horse, cow, or bovine.

[0068] Target: As used herein, a "target" is any molecule that is specifically bound by a binding portion of an antibody or an antigen-binding fragment thereof. In some embodiments, the target is an antigen described herein (e.g., Clec12A). In some embodiments, the target is an FcR (e.g., FcRn). The terms "primary target" and "secondary target" are used herein to refer to molecules of two different molecular species, rather than two molecules of the same molecular species. For example, in some embodiments, the primary target is a serum protein and the secondary target is FcRn.

[0069] Therapeutically effective amount: As used herein, the term "therapeutically effective amount" refers to an amount of a therapeutic molecule (e.g., an anti-Clec12A antibody described herein) that confers a therapeutic effect on the treated subject at a reasonable benefit / risk ratio applicable to any medical treatment. The therapeutic effect can be objective (i.e., measurable by some test or marker) or subjective (i.e., the subject shows signs of or feels an effect). In particular, a "therapeutically effective amount" refers to an amount of a therapeutic molecule or composition effective to treat, ameliorate, or prevent a particular disease or condition, or to exhibit a therapeutic or prophylactic effect detectable, for example, by improving symptoms associated with the disease, preventing or delaying the onset of the disease, and / or reducing the severity or frequency of symptoms of the disease. The therapeutically effective amount may be administered in a dosing regimen that may be composed of multiple unit doses. For any particular therapeutic molecule, the therapeutically effective amount (and / or the appropriate unit dose within an effective dosing regimen) may vary depending, for example, on the route of administration, combination with other pharmaceutical agents. In addition, the specific therapeutically effective amount (and / or unit dose) for any particular subject will depend on a variety of factors, including the disorder being treated and the severity of the disorder; the activity of the particular pharmaceutical agent used; the particular composition used; the age, weight, general health, sex, and diet of the subject; the time of administration, route of administration, and / or rate of excretion or metabolism of the particular therapeutic molecule used; the duration of treatment; and similar factors well known in the medical arts.

[0070] Treatment: As used herein, the term "treatment" (also "treat" or "treating") refers to any administration of a therapeutic molecule (e.g., an anti-Clec12A antibody described herein) that partially or completely relieves, ameliorates, alleviates, inhibits, delays the onset of, reduces the severity of, and / or reduces the incidence of, one or more symptoms, characteristics, and / or features of a particular disease, disorder, and / or condition. Such treatment may be of subjects who do not exhibit symptoms of the relevant disease, disorder, and / or condition, and / or of subjects who exhibit only early signs of the disease, disorder, and / or condition. Alternatively or additionally, such treatment may be of subjects who exhibit one or more established signs of the relevant disease, disorder, and / or condition.

[0071] The drawings are for illustrative purposes only and not for limiting purposes. [Brief description of the drawings]

[0072] [Figure 1-1] 1 shows Biacore Clec12A-His binding traces from a surface plasmon resonance assay. [Figure 1-2] 1 shows Biacore Clec12A-His binding traces from a surface plasmon resonance assay. [Diagram 2] Exemplary flow cytometry histograms for wild-type and Clec12A-overexpressing cell lines are shown. [Diagram 3] 1 shows an exemplary on-cell binding affinity concentration response curve in U937 cells. [Figure 4] 1 shows an exemplary ELISA epitope binning assay format. [Figure 5A] 1 shows a soluble membrane protein (SMP) assay. [Figure 5B] 1 shows a soluble membrane protein (SMP) assay. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0073] The present disclosure is based, in part, on the discovery of engineered antibodies and antigen-binding fragments thereof that bind to Clec12A (e.g., human Clec12A), member A of the C-type lectin domain family 12, a cell surface receptor that contains ITIM motifs, regulates signaling cascades, and mediates tyrosine phosphorylation of target MAP kinases.

[0074] Clec12A is highly expressed in cancerous cells such as acute myeloid leukemia cells, but not in normal hematopoietic stem cells. In addition, CLL-1 is also expressed on the surface of leukemia stem cells (LSCs), which can self-generate and generate more leukemia cells, which is associated with relapse.

[0075] In some embodiments, Clec12A is used to treat cancer: anti-Clec12A antibodies are used to recruit a patient's unstimulated primary T cells to cancer cells that have Clec12A on their surface using bispecific antibodies such as CD3 / Clec12A, or to generate CART cells specific for the Clec12A antigen to target AML.

[0076] antibody The antibodies described herein are designed to bind to Clec12A. In certain embodiments, the anti-Clec12A antibodies and antigen-binding fragments thereof bind to human Clec12A. Human Clec12A has been described in the art, for example in NCBI Reference No. NP_612210.4, NCBI Reference No. NP_963917.2, NCBI Reference No. NP_001193939.1, and NCBI Reference No. NP_001287659.1.

[0077] The anti-Clec12A antibodies described herein may be immunoglobulins, heavy chain antibodies, light chain antibodies, LRR-based antibodies, or other protein scaffolds with antibody-like properties, as well as other immunological binding moieties known in the art, including, for example, Fab, Fab', Fab'2, Fab2, Fab3, F(ab')2, Fd, Fv, Feb, scFv, SMIPs, antibodies, diabodies, triabodies, tetrabodies, minibodies, maxibodies, tandabs, DVDs, BiTe, TandAbs, etc., or any combination thereof. The subunit structures and three-dimensional configurations of different classes of antibodies are known in the art.

[0078] An antibody can be an immunoglobulin molecule of four polypeptide chains, for example, two heavy (H) chains and two light (L) chains. The heavy chain can include a heavy chain variable domain and a heavy chain constant domain. The heavy chain constant domain can include CH1, hinge, CH2, CH3, and optionally CH4 regions. A suitable heavy chain constant region can be derived from any immunoglobulin (e.g., IgA, IgG, or IgE). In some embodiments, a suitable heavy chain constant region can be derived from IgG1, IgG2, or IgG4. In certain embodiments, a suitable heavy chain constant region is derived from IgG1. The light chain can include a light chain variable domain and a light chain constant domain. The light chain constant domain can include either a kappa light chain or a lambda light chain. The heavy chain variable domain of the heavy chain and the light chain variable domain of the light chain can be further divided into regions of variability, called complementarity determining regions (CDRs), typically interspersed with more conserved regions, called framework regions (FRs). Each of such heavy and light chain variable domains may comprise three CDRs and four framework regions, arranged from amino terminus to carboxyl terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. One or more of these may be engineered as described herein. The assignment of amino acids to each domain follows the definitions in Kabat Sequences of Proteins of Immunological Interest (National Institutes of Health, Bethesda, Md. (1987 and 1991)), or Chothia & Lesk J. Mol. Biol. 196:901-917 (1987); Chothia et al. Nature 342:878-883 (1989). As used herein, CDRs refer to each of the heavy chain (HCDR1, HCDR2, HCDR3) and light chain (LCDR1, LCDR2, LCDR3).

[0079] Embodiments of the present invention include antibodies comprising the CDRs found in the VH and VL domains described herein, identified using conventional numbering systems such as the IMGT, Kabat, and Clothia numbering systems. Such numbering systems are well known in the art. In certain embodiments, the CDRs are identified or numbered according to the IMGT numbering system.

[0080] Exemplary anti-Clec12A antibody Exemplary anti-Clec12A antibodies are shown in the table below. The variable heavy (VH) and variable light (VL) sequences of anti-Clec12A antibodies AI are shown below.

[0081] JPEG2024516645000001.jpg196158

[0082] JPEG2024516645000002.jpg97157

[0083] In some embodiments, the anti-Clec12A antibody comprises a heavy and / or light chain amino acid sequence having at least about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, or about 99% sequence identity to any one of SEQ ID NOs:1-18.

[0084] In some embodiments, the anti-Clec12A antibody contains no more than 30, 29, 28, 27, 26, 25, 24, 23, 22, 21, 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, or 2 amino acid substitutions relative to SEQ ID NOs: 1-18.

[0085] As will be appreciated by one of ordinary skill in the art, any such heavy chain CDR sequence can be readily combined, for example by molecular biology techniques, with any other antibody sequence or domain provided herein or otherwise known in the art, including any framework region, CDR, or constant domain, or portion thereof, disclosed herein or otherwise known in the art, as may be present in any format of antibody or antigen-binding fragment thereof disclosed herein or otherwise known in the art.

[0086] In the various engineered antibodies described herein, the heavy chain constant domain can be of any class (or subclass). In the various engineered antibodies described herein, the heavy chain constant domain can comprise the amino acid sequence of any one or more of IgG, IgM, IgA, IgD, or IgE, including subclasses such as IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2. In various embodiments, the constant domain of the engineered antibodies described herein can comprise a mixture of two or more classes (or subclasses) of immunoglobulin heavy chain constant domains. For example, an anti-Clec12A antibody can comprise a first portion of a constant domain having a sequence of an immunoglobulin constant domain selected from the constant domains of the IgG, IgM, IgA, IgD, or IgE classes, and a second portion of a constant domain having a sequence of an immunoglobulin constant domain selected from the constant domains of the IgG, IgM, IgA, IgD, or IgE classes that is different from the first portion. In some cases, the constant domain of an anti-Clec12A antibody described herein may comprise a mixture of two or more subclasses of a particular class of constant domain, for example a first portion of a constant domain having a sequence of an immunoglobulin constant domain selected from an IgG1, IgG2, IgG3, or IgG4 subclass constant domain, and a second portion of a constant domain having a sequence of an immunoglobulin constant domain selected from an IgG1, IgG2, IgG3, or IgG4 subclass constant domain that is different from the first. In some particular embodiments, the constant domain comprises all or a portion of an IgG2 constant domain and all or a portion of an IgG4 constant domain.

[0087] In some cases, the anti-Clec12A antibody comprises an antibody constant region, Fc region, or Fc fragment that exhibits altered binding (compared to a reference constant region) to one or more Fc receptors (e.g., an FcγRI, FcγRIIA, FcγRIIB, FcγRIIIA, FcγRIIIB, FcγRIV, or FcRn receptor). In some embodiments, the constant region, Fc region, or Fc fragment is engineered to bind to a target (e.g., an FcRn receptor) in an altered manner (e.g., in a pH-sensitive manner (e.g., in a more or less pH-sensitive manner) and / or with decreased or increased binding compared to a reference constant region, Fc region, or Fc fragment. In some embodiments, the anti-Clec12A antibody comprises an antibody constant region, Fc region, or Fc fragment that exhibits decreased binding (compared to a reference constant region) to one or more Fcγ receptors (e.g., FcγRI, FcγRIIA, FcγRIIB, FcγRIIIA, FcγRIIIB, or FcγRIV). In some embodiments, the anti-Clec12A antibody comprises an antibody constant region, Fc region, or Fc fragment that exhibits increased binding to an FcRn receptor at serum pH and / or at intracellular pH (compared to a reference constant region).

[0088] For example, an anti-Clec12A antibody may comprise a constant region, Fc region, or Fc fragment of an IgG antibody engineered to include an amino acid addition, deletion, or substitution of one or more of amino acid residues 251-256, 285-290, 308-314, 385-389, and 428-436 (Kabat numbering (Kabat et al., (1991) Sequences of Proteins of Immunological Interest, NIH)). Without wishing to be bound by theory, it is believed that one or more of these constant region, Fc region, or Fc fragment amino acids mediate interaction with an Fc receptor, e.g., FcRn. In some embodiments, one or more of these disclosed amino acids are substituted with histidine, arginine, lysine, aspartic acid, glutamic acid, serine, threonine, asparagine, or glutamine. In some embodiments, a non-histidine residue is substituted with a histidine residue. In some embodiments, the histidine residue is replaced with a non-histidine residue.

[0089] In some embodiments, the anti-Clec12A antibody comprises an IgG antibody constant region, Fc region, or Fc fragment having an amino acid modification at one or more of positions 308, 309, 311, 312, and 314, more specifically, a substitution of one or more of positions 308, 309, 311, 312, and 314 with threonine, proline, serine, aspartic acid, and leucine, respectively. In some embodiments, one or more of the residues at positions 308, 309, and 311 are substituted with isoleucine, proline, and glutamic acid, respectively. In still other embodiments, one or more of the residues at positions 308, 309, 311, 312, and 314 are substituted with threonine, proline, serine, aspartic acid, and leucine, respectively.

[0090] In some embodiments, the anti-Clec12A antibody comprises an IgG antibody constant region, Fc region, or Fc fragment having an amino acid modification, more particularly a substitution, at one or more of positions 251, 252, 254, 255, and 256. In some embodiments, residue 251 is substituted with leucine or arginine, residue 252 is substituted with leucine, tyrosine, phenylalanine, serine, tryptophan, or threonine, residue 254 is substituted with threonine or serine, residue 255 is substituted with leucine, glycine, isoleucine, or arginine, and / or residue 256 is substituted with serine, phenylalanine, arginine, glutamine, glutamic acid, aspartic acid, alanine, asparagine, or threonine. In some embodiments, residue 251 is substituted with leucine, residue 252 is substituted with tyrosine or leucine, residue 254 is substituted with threonine or serine, and / or residue 255 is substituted with arginine. In still other embodiments, residue 252 is substituted with phenylalanine, and / or residue 256 is substituted with aspartic acid. In some embodiments, residue 251 is substituted with leucine, residue 252 is substituted with tyrosine, residue 254 is substituted with threonine or serine, and / or residue 255 is substituted with arginine.

[0091] In some embodiments, the anti-Clec12A antibody comprises an IgG antibody constant region, Fc region, or Fc fragment having an amino acid modification, more particularly a substitution, at one or more of positions 428, 433, 434, 435, and 436. In some embodiments, residue 428 is substituted with methionine, threonine, leucine, phenylalanine, or serine, residue 433 is substituted with lysine, arginine, serine, isoleucine, proline, glutamine, or histidine, residue 434 is substituted with phenylalanine, tyrosine, or histidine, residue 435 is substituted with tyrosine, and / or residue 436 is substituted with histidine, asparagine, arginine, threonine, lysine, methionine, or threonine. In some embodiments, one or more of residues 433, 434, 435, and 436 are substituted with lysine, phenylalanine, tyrosine, and histidine, respectively. In some embodiments, residue 428 is substituted with methionine and / or residue 434 is substituted with tyrosine.

[0092] In some embodiments, the anti-Clec12A antibody comprises an IgG antibody constant region, Fc region, or Fc fragment having an amino acid modification at one or more of positions 385, 386, 387, and 389, more particularly a substitution at one or more of these positions. In some embodiments, residue 385 is substituted with arginine, aspartic acid, serine, threonine, histidine, lysine, or alanine, residue 386 is substituted with threonine, proline, aspartic acid, serine, lysine, arginine, isoleucine, or methionine, residue 387 is substituted with arginine, histidine, serine, threonine, alanine, or proline, and / or residue 389 is substituted with proline or serine. In some embodiments, one or more of residues 385, 386, 387, and 389 are substituted with arginine, threonine, arginine, and proline, respectively. In some embodiments, one or more of residues 385, 386, and 389 are substituted with aspartic acid, proline, and serine, respectively.

[0093] In some embodiments, the anti-Clec12A antibody comprises an IgG antibody constant region, Fc region, or Fc fragment having one or more of the following substitutions: a leucine at residue 251, a tyrosine or leucine at residue 252, a threonine or serine at residue 254, an arginine at residue 255, a threonine at residue 308, a proline at residue 309, a serine at residue 311, an aspartic acid at residue 312, a leucine at residue 314, an arginine at residue 385, a threonine at residue 386, an arginine at residue 387, a proline at residue 389, a methionine at residue 428, a lysine at residue 433, a phenylalanine or tyrosine at residue 434, a tyrosine at position 435, and / or a tyrosine at position 436. Additional amino acid substitutions that can be included in the constant region, Fc region, or Fc fragment include those described in, for example, US Pat. Nos. 6,277,375; 8,012,476; and 8,163,881.

[0094] In some embodiments, the anti-Clec12A antibodies described herein comprise a heavy chain constant domain comprising Ala-Ala mutations, e.g., as described in PCT Publication Nos. WO94 / 28027 and WO98 / 47531; and Xu et al. (2000) Cell Immunol 200:16-26. Thus, in some embodiments, anti-Clec12A antibodies having one or more mutations in the heavy chain constant region, including Ala-Ala mutations, have reduced or no effector function. In accordance with these embodiments, the constant region of the anti-Clec12A antibodies described herein may comprise a substitution at position 234 to alanine and / or a mutation at position 235 (EU numbering) to alanine.

[0095] As will be appreciated by one of ordinary skill in the art, any such heavy chain constant domain sequence can be readily combined, for example by molecular biology techniques, with any other antibody sequence or domain provided herein or otherwise known in the art, including any framework region, CDR, or constant domain, or portion thereof, disclosed herein or otherwise known in the art, as may be present in any format of antibody or antigen-binding fragment thereof disclosed herein or otherwise known in the art.

[0096] Exemplary Antibodies The engineered antibodies can comprise various heavy and light chains as described herein. In some embodiments, an anti-Clec12A antibody can comprise two heavy and light chains. In various embodiments, the disclosure encompasses antibodies comprising at least one heavy and / or light chain disclosed herein, at least one heavy and / or light chain framework domain disclosed herein, at least one heavy and / or light chain CDR domain disclosed herein, and / or any heavy and / or light chain constant domain disclosed herein.

[0097] In various embodiments, the anti-Clec12A antibodies disclosed herein are homodimeric monoclonal antibodies. In various embodiments, the anti-Clec12A antibodies disclosed herein are heterodimeric antibodies. In various embodiments, the anti-Clec12A antibodies are, for example, typical antibodies, or diabodies, triabodies, tetrabodies, minibodies, maxibodies, tandibodies, DVDs, BiTe, scFvs, TandAbs, scFvs, Fabs, Fab2s, Fab3s, F(ab')2s, etc., or any combination thereof.

[0098] In some embodiments, the disclosure provides a fusion protein comprising one or more variable domains or engineered antibodies described herein, or portions thereof, and one or more additional polypeptides.

[0099] Exemplary Single Chain Variable Fragments In some embodiments, the disclosure provides a single chain variable fragment. In some embodiments, the scFv is a human scFv. The term "single chain variable fragment" or "scFv" refers to a V H Heavy chains (VL) of immunoglobulins (e.g., mouse or human) covalently linked to form a VL heterodimer. H ) and light chain (V L It refers to a fusion protein of the variable region of the heavy chain (V H ) and light chain (V L ) is directly bonded or V H N-terminus of V L or at the C-terminus of V H The C-terminus of V LThe linker is linked by a peptide-encoded linker (e.g., 10, 15, 20, 25 amino acids) that connects the heavy chain variable region and the light chain variable region of the extracellular antigen-binding domain to the N-terminus of the G4S linker (SEQ ID NO: 78). The linker is usually rich in glycine for flexibility and rich in serine or threonine for solubility. The linker can link the heavy chain variable region and the light chain variable region of the extracellular antigen-binding domain. Non-limiting examples of linkers are disclosed in Shen et al., Anal. Chem. 80(6):1910-1917 (2008) and WO2014 / 087010, the contents of which are incorporated herein by reference in their entirety. In a particular embodiment, the linker is a G4S linker (SEQ ID NO: 78).

[0100] Alternatively, or additionally, the scFv may be derived from a Fab (e.g., not from an antibody obtained from a Fab library). In certain embodiments, the anti-Clec12A antibody or antigen-binding fragment thereof is a Fab. In certain embodiments, the Fab is cross-linked. In certain embodiments, the anti-Clec12A antibody or antigen-binding fragment thereof is a F(ab)2. Any of the foregoing molecules can be included in a fusion protein with a heterologous sequence to form an anti-Clec12A antigen antibody or antigen-binding fragment thereof.

[0101] In certain embodiments, the anti-Clec12A antibody or antigen-binding fragment thereof is at least about 1×10 -6 M, at least about 1x10 -7 M, at least about 1x10 -8 M, at least about 1x10 -9 M, or at least about 1x10 -10 Dissociation constant of M (K D In certain embodiments, the anti-Clec12A antibody or antigen-binding fragment thereof binds to Clec12A (e.g., human Clec12A) at a binding affinity of at least about 2×10 -8 Dissociation constant of M (K D In certain embodiments, the anti-Clec12A antibody or antigen-binding fragment thereof binds to Clec12A (e.g., human Clec12A) at a binding affinity of at least about 2×10 -8 M ~ approx. 8x10 -9 Dissociation constant of M (KD ) binds to Clec12A (e.g., human Clec12A).

[0102] In some embodiments, the anti-Clec12A antibody or antigen-binding fragment thereof has a dissociation constant (K D In some embodiments, the anti-Clec12A antibody or antigen-binding fragment thereof binds to Clec12A (e.g., human Clec12A) with a dissociation constant (K) of at least about 50 nM, at least about 40 nM, at least about 35 nM, at least about 30 nM, at least about 25 nM, at least about 20 nM, at least about 19 nM, at least about 18 nM, at least about 17 nM, at least about 16 nM, at least about 15 nM, at least about 14 nM, at least about 13 nM, at least about 12 nM, at least about 11 nM, at least about 10 nM, at least about 9 nM, at least about 8 nM, at least about 7 nM, at least about 6 nM, at least about 5 nM. D ) binds to Clec12A (e.g., human Clec12A).

[0103] In some embodiments, the anti-Clec12A scFv comprises a sequence consisting of an amino acid sequence having about 80% or greater identity to SEQ ID NO:56, SEQ ID NO:57, SEQ ID NO:58, or SEQ ID NO:59. In some embodiments, the anti-Clec12A scFv comprises a sequence consisting of SEQ ID NO:56, SEQ ID NO:57, SEQ ID NO:58, or SEQ ID NO:59.

[0104] In some embodiments, the anti-Clec12A scFv comprises a linker comprising or consisting of the amino acid sequence represented by SEQ ID NO:60, as provided below: GGGGSGGGGSGGGGS (SEQ ID NO:60)

[0105] In some embodiments, the linker comprises or consists of the amino acid sequence set forth in SEQ ID NO:61, provided below: GGGGSGGGGSGGGSGGGGS (sequence number 61).

[0106] In some embodiments, the linker comprises or consists of the amino acid sequence set forth in SEQ ID NO:62, provided below: GGGGSGGGGSGGGGSGGGSGGGGS (sequence number 63).

[0107] In some embodiments, the linker comprises or consists of the amino acid sequence set forth in SEQ ID NO:63, provided below: GGGGSGGGGSGGGGSGGGGSGGGSGGGGS (sequence number 64).

[0108] In some embodiments, the anti-Clec12A antibody or antigen-binding fragment thereof comprises a conservative sequence modification (e.g., an anti-Clec12A antibody or fragment thereof described herein). In some embodiments, a conservative sequence modification is an amino acid modification that does not significantly affect or change the binding properties of the anti-Clec12A antibody or antigen-binding fragment thereof disclosed herein (e.g., an antibody or antigen-binding fragment thereof) comprising the amino acid sequence. Conservative modifications may include amino acid substitutions, additions, and deletions. Modifications can be introduced into the anti-Clec12A antibody or antigen-binding fragment thereof by standard techniques known in the art, such as site-directed mutagenesis and PCR-mediated mutagenesis. Amino acids can be classified into groups according to their physicochemical properties, such as their charge and polarity. Conservative amino acid substitutions are those in which an amino acid residue is replaced with an amino acid from the same group. For example, amino acids can be classified by charge: positively charged amino acids include lysine, arginine, and histidine, negatively charged amino acids include aspartic acid and glutamic acid, and medium charged amino acids include alanine, asparagine, cysteine, glutamine, glycine, isoleucine, leucine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, and valine. Additionally, amino acids can be classified by polarity: polar amino acids include arginine (basic polar), asparagine, aspartic acid (acidic polar), glutamic acid (acidic polar), glutamine, histidine (basic polar), lysine (basic polar), serine, threonine, and tyrosine; nonpolar amino acids include alanine, cysteine, glycine, isoleucine, leucine, methionine, phenylalanine, proline, tryptophan, and valine. Thus, one or more amino acid residues in a CDR region can be replaced with other amino acid residues from the same group, and the altered antibody can be tested for retained function. In certain embodiments, no more than one, no more than two, no more than three, no more than four, no more than five residues in a particular sequence or CDR region are altered.

[0109] In some embodiments, the light and / or heavy chain of the anti-Clec12A scFv comprises a signal peptide. In some embodiments, the signal peptide comprises an amino acid sequence having at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 91%, at least about 91%, at least about 92%, at least about 93%, at least about 94%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% homology or identity to one amino acid sequence selected from METDTLLLWVLLLWVPGSTG (SEQ ID NO:75), MYRMQLLSCIALSLALVTNS (SEQ ID NO:62), MYSMQLASCVTLTLVLLVNS (SEQ ID NO:76), METPAQLLFLLLLWLPDTTG (SEQ ID NO:77), MALPVTALLLPLALLLHAARP (SEQ ID NO:65), MKWVTFISLLFSSAYS (SEQ ID NO:66), MDSKGSSQKGSRLLLLLVVSNLLLCQGVVS (SEQ ID NO:67). In some embodiments, the signal sequence is METDTLLLWVLLLWVPGSTG (SEQ ID NO: 75). In some embodiments, the signal sequence is MYRMQLLSCIALSLALVTNS (SEQ ID NO: 62). In some embodiments, the signal sequence is MYSMQLASCVTLTLVLLVNS (SEQ ID NO: 76). In some embodiments, the signal sequence is METPAQLLFLLLLWLPDTTG (SEQ ID NO: 77). In some embodiments, the signal sequence is MALPVTALLLPLALLLHAARP (SEQ ID NO: 65). In some embodiments, the signal sequence is MKWVTFISLLFSSAYS (SEQ ID NO: 66).

[0110] In some embodiments, the anti-Clec12A scFv comprises the following amino acid sequence: QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYWSWIRQPPGKGLEWIGYIYYSGSTKYNPSLKSRVTISVDTSKNLFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSAIQMTQSPSSLSASVGDRVTITCRASQGIRYDLGWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO:56);

[0111] QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAIVISYDGSDKYYVDSVKGRFTISRDNSKNTLYLHMNSLRAEDTAVYYCARDKGYYFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSEIVMTQSPATLSLSPGERATLSCRASQSVGNRYLSWYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO:57);

[0112] QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYSADSVKGRFNISRDNSKNTLYLQMNSLRAEDTAVYYCARDGSRYFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSEIFMTQSPATLSLPGERATLSCRASQSVHSKYLSWYQQKPGQAPSLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPITFGQGTRLEIK (SEQ ID NO:58); or

[0113] QLQLQESGPGLVKPSETLSLTCTVSGGSISSSTYYWGWIRQPPRKGLEWIGSTHYRGSTYYNPSLKSRVTISVDTSKNQFSLKVSSVTAADTAVYYCARELTGEVFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSYSTPFTFGPGTKVDIK (sequence number 59).

[0114] In some embodiments, the anti-Clec12A scFv comprises an amino acid sequence having at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 91%, at least about 92%, at least about 93%, at least about 94%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% homology or identity to one or more of SEQ ID NOs:56-59.

[0115] In some embodiments the anti-Clec12A scFv comprises the amino acid sequence set forth in SEQ ID NO: 56. In some embodiments the anti-Clec12A scFv comprises the amino acid sequence set forth in SEQ ID NO: 57. In some embodiments the anti-Clec12A scFv comprises the amino acid sequence set forth in SEQ ID NO: 58. In some embodiments the anti-Clec12A scFv comprises the amino acid sequence set forth in SEQ ID NO: 59.

[0116] Nucleotide sequence The present disclosure includes nucleotide sequences encoding one or more heavy chains, heavy chain variable domains, heavy chain framework regions, heavy chain CDRs, heavy chain constant domains, light chains, light chain variable domains, light chain framework regions, light chain CDRs, light chain constant domains, or other immunoglobulin-like sequences or antibodies disclosed herein. In various embodiments, such nucleotide sequences may be present in a vector. In various embodiments, such nucleotides may be present in the genome of a cell, for example, a subject's cell in need of treatment, or a cell for antibody production, for example, a mammalian cell for antibody production.

[0117] Engineered antibodies and fusion proteins In some embodiments, the disclosure provides fusion proteins comprising (i) one or more antigen-binding regions as described herein (e.g., antigen-binding regions of immunoglobulins, heavy chain antibodies, light chain antibodies, LRR-based antibodies, or other protein scaffolds with antibody-like properties, as well as other antigen-binding moieties known in the art, including, e.g., Fab, Fab', Fab'2, Fab2, Fab3, F(ab')2, Fd, Fv, Feb, scFv, SMIP, antibodies, diabodies, triabodies, tetrabodies, minibodies, maxibodies, tandb, DVD, BiTe, TandAb, etc.), e.g., one or more variable domains as described herein, or portions thereof (e.g., one or more CDRs as described herein), and (ii) one or more additional polypeptides. For example, albumin is an abundant serum protein and is protected from degradation by pH-dependent recycling mediated by its interaction with FcRn. In some embodiments, one or more variable domains or engineered antibodies described herein, or portions thereof (e.g., one or more CDRs described herein) are fused to albumin, a portion thereof (such as a portion of albumin that binds to FcRn), and / or an engineered variant of albumin that binds to FcRn with improved affinity. In other examples, one or more variable domains or engineered antibodies described herein, or portions thereof (e.g., one or more CDRs described herein) are fused to a polypeptide that binds to albumin to form a fusion protein-albumin complex, which can then bind to FcRn. In some embodiments, the polypeptide that binds to albumin is a single chain variable fragment (scFv). Albumin or a portion thereof can include one or more amino acid mutations that can alter binding to FcRn. Such mutations are known in the art (see, e.g., Andersen et al., Nature Communications 3:610 doi:10.1038 / nocmms1607(2012)). In other examples, one or more variable domains or engineered antibodies described herein, or portions thereof (eg, one or more CDRs described herein) are fused to transferrin.Transferrin is recycled by binding to the transferrin receptor (see, e.g., Widera et al., Adv. Drug Deliv. Rev. 55:1439-66 (2003)).

[0118] Engineered antibodies and fragments thereof Anti-Clec12A antibodies and antigen-binding fragments thereof according to the present disclosure are engineered to contain one or more binding moieties that specifically bind to one or more targets of interest. Clec12A antibodies and antigen-binding fragments thereof include nucleic acids (e.g., RNA and DNA), proteins (e.g., antibodies), and combinations thereof. In some embodiments, the pH-dependent binding moiety may be or may include, for example, nucleic acids (e.g., RNA and DNA), as well as aptamers, polypeptides (e.g., antibodies or fragments thereof, albumin, receptors, ligands, signal peptides, avidin, and protein A), polysaccharides, biotin, hydrophobic groups, hydrophilic groups, drugs, and any organic molecule that binds to a receptor.

[0119] Antibodies or fragments thereof as binding moieties In some embodiments, the antibody or fragment thereof described herein is an anti-Clec12A antibody. In some cases, one or more of the binding moieties described herein is or includes an antibody, an antigen-binding fragment thereof, and / or an Fc region (or Fc fragment) thereof. The basic structure of an IgG antibody consists of two identical light polypeptide chains and two identical heavy polypeptide chains linked together by disulfide bonds. The first domains located at the amino terminus of each chain are variable in amino acid sequence and provide the antibody binding specificity found in an individual antibody. These are known as the heavy chain variable (VH) region and the light chain variable (VL) region. The other domains of each chain are relatively invariant in amino acid sequence and are known as the constant heavy (CH) region and the constant light (CL) region. In the case of an IgG antibody, the light chain includes one variable region (VL) and one constant region (CL). The IgG heavy chain includes a variable region (VH), a first constant region (CH1), a hinge region, a second constant region (CH2), and a third constant region (CH3). In IgE and IgM antibodies, the heavy chain contains an additional constant region (CH4).

[0120] Antibodies may include, for example, monoclonal antibodies, recombinantly produced antibodies, monospecific antibodies, multispecific antibodies (including bispecific antibodies), human antibodies, engineered antibodies, humanized antibodies, chimeric antibodies, immunoglobulins, synthetic antibodies, tetrameric antibodies comprising two heavy chain 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, intrabodies, antibody fusions (sometimes referred to herein as "antibody conjugates"), heteroconjugate antibodies, antibody-drug conjugates, single domain antibodies, monovalent antibodies, single chain antibodies or single chain Fvs (scFvs), camelized antibodies, affibodies, Fab fragments, F(ab')2 fragments, disulfide-linked Fvs (sdFvs), anti-idiotypic (anti-Id) antibodies (including, for example, anti-anti-Id antibodies), minibodies, domain antibodies, synthetic antibodies (sometimes referred to as "antibody mimetics"), and antigen-binding fragments of any of the above. In certain embodiments, the antibodies described herein refer to polyclonal antibody populations.

[0121] As used herein, the term "Fc fragment" refers to one or more fragments of the Fc region that retain Fc function and / or activity as described herein, such as binding to an Fc receptor. As used herein, the term "antigen-binding portion" of an antibody refers to one or more fragments of an antibody that retain the ability to specifically bind to an antigen. Examples of binding fragments encompassed within the term "antigen-binding fragment" of an antibody include Fab fragments, F(ab')2 fragments, Fd fragments, Fv fragments, scFv fragments, dAb fragments (Ward et al., (1989) Nature 341:544-546), and isolated complementarity determining regions (CDRs). These antibody fragments can be obtained using conventional techniques known to those of skill in the art, and the fragments can be screened for utility in the same manner as intact antibodies.

[0122] In some aspects, the invention provides an antibody or antigen-binding fragment thereof that binds to human Clec12A, comprising a human heavy chain and / or light constant region, wherein the human heavy constant region comprises an isotype variant comprising a human IgG1, human IgG2, human IgG3, or human IgG4 Fc region.

[0123] In a further aspect, the invention provides a humanized antibody or fragment thereof that binds to human Clec12A, wherein the antibody comprises a variant human IgG Fc region comprising the amino acid substitution S324N, which replaces the serine at amino acid position 324 of the parent antibody with an asparagine, whereas the antibody comprising the variant human IgG Fc region exhibits improved complement dependent cytotoxicity (CDC) compared to the parent antibody.

[0124] The antibody or fragment can be produced by any method known in the art for synthesizing antibodies (see, e.g., Harlow et al., Antibodies: A Laboratory Manual, (Cold Spring Harbor Laboratory Press, 2nd ed. 1988); Brinkman et al., 1995, J. Immunol. Methods 182:41-50; WO92 / 22324; WO98 / 46645). Chimeric antibodies can be produced using the methods described, for example, in Morrison, 1985, Science 229:1202, and humanized antibodies by the methods described, for example, in U.S. Patent No. 6,180,370.

[0125] Further compositions and methods described herein are bispecific and multivalent antibodies, e.g., as described in Segal et al., J. Immunol. Methods 248:1-6 (2001); and Tutt et al., J. Immunol. 147:60 (1991).

[0126] Engineered antigen-binding regions In some embodiments, the binding moiety is or comprises an antibody (e.g., an IgG antibody, e.g., an IgG1, IgG2, or IgG3 antibody), or an antigen-binding fragment, engineered to bind a target (i.e., antigen) in an altered manner (e.g., in a pH-sensitive manner, e.g., in a more or less pH-sensitive manner) compared to a reference antibody or antigen-binding fragment. For example, an antibody can be engineered by modifying (e.g., by adding, deleting, or substituting) amino acids within one or more antibody CDRs and / or at positions involved in the antibody's CDR structure. Exemplary non-limiting sites of an antibody that can be modified include (amino acid positions are indicated according to Kabat numbering (Kabat et al., (1991) Sequences of Proteins of Immunological Interest, NIH)):

[0127] Heavy chain: H27, H31, H32, H33, H35, H50, H58, H59, H61, H62, H63, H64, H65, H99, H100b, and H102

[0128] Light chain: L24, L27, L28, L32, L53, L54, L56, L90, L92, and L94.

[0129] In some embodiments, one or more of these disclosed amino acids may be replaced with histidine, arginine, lysine, aspartic acid, glutamic acid, serine, threonine, asparagine, or glutamine. Without wishing to be bound by theory, it is believed that replacing one or more amino acids at these positions with histidine results in an antibody with pH-dependent antigen-binding properties. In some embodiments, a non-histidine residue is replaced with a histidine residue. In some embodiments, a histidine residue is replaced with a non-histidine residue. Additional engineered antigen-binding regions include, for example, those described in US Patent Publication No. 20110229489.

[0130] Engineered constant regions In some cases, the binding moiety is or comprises an antibody constant region, Fc region, or Fc fragment that binds to one or more Fc receptors (e.g., FcγRI, FcγRIIA, FcγRIIB, FcγRIIIA, FcγRIIIB, FcγRIV, or FcRn receptors). In some embodiments, the constant region, Fc region, or Fc fragment is engineered to bind to a target (e.g., an Fc receptor) in an altered manner (e.g., in a pH-sensitive manner (e.g., in a more or less pH-sensitive manner) compared to a reference constant region, Fc region, or Fc fragment.

[0131] In some examples, the binding moiety may be or include a constant region, Fc region, or Fc fragment of an IgG antibody that has been engineered to include amino acid additions, deletions, or substitutions of one or more of the amino acid residues described herein (e.g., 251-256, 285-290, 308-314, 385-389, and 428-436 (Kabat numbering (Kabat et al., (1991) Sequences of Proteins of Immunological Interest, NIH))).

[0132] Production of Clec12A antibodies and their fragments In some embodiments, the antibody or fragment thereof described herein is engineered by mutagenesis using known techniques to include one or more binding moieties that exhibit binding to one or more targets. For example, the sequence of a reference polypeptide (e.g., a therapeutic antibody or a therapeutic fusion protein) can be obtained, and one or more amino acid residues can be added, deleted, or substituted. In some embodiments, one or more of the amino acid residues are substituted with histidine, arginine, lysine, aspartic acid, glutamic acid, serine, threonine, asparagine, or glutamine. In some embodiments, one or more amino acids are substituted with histidine.

[0133] In some embodiments, the substitution of an amino acid residue with histidine results in the insertion of a protonation site, which may increase the pH sensitivity of the binding moiety. Polypeptides can be produced using standard methods and assayed for binding to a target of interest as described herein. Additional methods for increasing the pH sensitivity of binding moieties are described, for example, in Sarkar et al., Nature Biotechnology 20:908-913 (2002); Murtaugh et al., Protein Science 20:1619-1631 (2011); and U.S. Patent Publication No. 20110229489.

[0134] In some embodiments, a first target of interest is selected, an antibody that selectively binds to the target is provided, obtained, and / or generated (e.g., using known methods described herein), one or more amino acids in the antigen binding region and / or Fc region are substituted (e.g., with histidine, arginine, lysine, aspartic acid, glutamic acid, serine, threonine, asparagine, or glutamine), and the pH sensitivity of binding to the target (and, additionally or alternatively, to FcRn) is determined.

[0135] In some embodiments, a polypeptide that naturally binds to a target of interest is provided, obtained, and / or produced. The polypeptide is conjugated to an Fc region or Fc fragment (e.g., binds to FcRn with a desired binding affinity) described herein using known methods. For example, the polypeptide and the Fc region or Fc fragment can be conjugated by chemical means or by recombinant expression as a fusion protein. Additionally or alternatively, one or more amino acids of the polypeptide can be substituted (e.g., with histidine, arginine, lysine, aspartic acid, glutamic acid, serine, threonine, asparagine, or glutamine) to determine the pH sensitivity and target of the binding of the polypeptide.

[0136] In some embodiments, the antibodies or fragments thereof described herein are engineered to contain one or more binding moieties identified and / or selected by screening. For example, antigen-binding moieties that bind to an antigen can be identified using libraries expressing antigen-binding moieties, e.g., phage libraries. Methods for screening recombinant antibody libraries are known (see, e.g., Hoogenboom, Nature Biotech. 23:1105-1116 (2005); U.S. Patent No. 5,837,500; U.S. Patent No. 5,571,698; WO2012 / 044831).

[0137] PEGylation In certain embodiments, the anti-Clec12A antibodies described herein can be PEGylated to contain mono- or poly (e.g., 2-4) PEG moieties. Such PEGylated antibodies can exhibit increased half-life compared to a non-PEGylated reference antibody, e.g., an antibody having the same amino acid sequence but different, a different amount of PEGylation, or no PEGylation.

[0138] PEGylation can be carried out by any suitable reaction known in the art.The method for preparing PEGylated protein can generally include: (a) reacting a polypeptide with polyethylene glycol (such as a reactive ester or aldehyde derivative of PEG) under conditions that allow the polypeptide to be bound to one or more PEG groups; (b) obtaining a reaction product(s).In general, the reaction conditions can be determined on a case-by-case basis based on known parameters and desired results.

[0139] There are numerous methods of PEG attachment available to one of skill in the art. For example, PEGylating an antibody or fragment thereof described herein can be carried out via an acylation reaction or an alkylation reaction with a reactive polyethylene glycol molecule.

[0140] Measuring binding moiety-target interactions The binding properties of the antibodies or fragments thereof described herein (e.g., anti-Clec12A antibodies described herein) to a target (e.g., Clec12A) can be measured by methods known in the art, such as one of the following methods: BIACORE analysis, enzyme-linked immunosorbent assay (ELISA), X-ray crystallography, sequence analysis, and scanning mutagenesis. The binding interaction of the antibody with Clec12A and / or FcRn can be analyzed using surface plasmon resonance (SPR). SPR, or biomolecular interaction analysis (BIA), detects biospecific interactions in real time, without labeling any of the interactants. A change in mass at the binding surface of the BIA chip (indicative of a binding event) results in a change in the refractive index of light near the surface. The change in refractive index generates a detectable signal, which is measured as an indication of a real-time reaction between biomolecules. Methods for using SPR are described, for example, in U.S. Patent No. 5,641,640; Raether (1988) Surface Plasmons Springer Verlag; Sjolander and Urbaniczky (1991) Anal. Chem. 63:2338-2345; Szabo et al. (1995) Curr. Opin. Struct. Biol. 5:699-705, and in online materials provided by BIAcore International AB (Uppsala, Sweden). In addition, KinExA® (Kinexclusion Assay) assays available from Sapidyne Instruments (Boise, Id.) can also be used.

[0141] Information from SPR can be used to determine the equilibrium dissociation constant (K D ), and K on and K. offSuch data can be used to compare different molecules. Information from SPR can also be used to develop structure-activity relationships (SAR). For example, the kinetic and equilibrium binding parameters of a particular binding moiety to a target at various pH levels can be evaluated. At a particular pH level, certain binding parameters, e.g., high affinity, low affinity, and slow K off It is possible to identify variant amino acids at a given position that correlate with

[0142] Treatment method In some embodiments, the anti-Clec12A antibodies or antigen-binding fragments thereof described herein are used in a method for treating one or more Clec12A-associated conditions. In some embodiments, the anti-Clec12A antibodies or antigen-binding fragments thereof described herein are for use as a medicament. Clec12A-associated conditions may include, but are not limited to, conditions caused by Clec12A expression, conditions involving symptoms that are due in whole or in part to Clec12A expression, or conditions known to occur in association with Clec12A expression.

[0143] In some aspects, the invention provides methods for treating cancer comprising administering an anti-Clec12A antibody or antigen-binding fragment thereof described herein. Cancer is a broad group of diverse diseases characterized by the uncontrolled proliferation of abnormal cells in the body. Uncontrolled cell division and proliferation can result in the formation of malignant tumors that can invade adjacent tissues and metastasize to distant parts of the body via the lymphatic system and bloodstream. In some embodiments, "cancer" or "cancer tissue" includes solid tumors. Examples of cancers that can be treated by the methods of the invention include, but are not limited to, cancers of the immune system, including lymphomas, leukemias, myelomas, and other white blood cell malignancies. In some embodiments, the cancer is acute myeloid leukemia. In some embodiments, the cancer is relapsed and refractory acute myeloid leukemia.

[0144] In some embodiments, the lymphoma is selected from the group consisting of acute lymphoblastic leukemia (ALL), AIDS-related lymphoma, ALK-positive large B-cell lymphoma, Burkitt's lymphoma, chronic lymphocytic leukemia (CLL), classical Hodgkin lymphoma, diffuse large B-cell lymphoma (DLBCL), follicular lymphoma, intravascular large B-cell lymphoma, large B-cell lymphoma arising in HHV8-associated multicentric Castleman disease, lymphomatoid granulomatosis, and lymphomas with leukemia, bronchitis ... The cancer is selected from the group consisting of lymphoplasmacytic lymphoma, mantle cell lymphoma (MCL), marginal zone B-cell lymphoma (MZL), mucosa-associated lymphoid tissue lymphoma (MALT), nodal marginal zone B-cell lymphoma (NMZL), nodal lymphocyte-predominant Hodgkin lymphoma, non-Hodgkin lymphoma, plasmablastic lymphoma, primary central nervous system lymphoma, primary effusion lymphoma, splenic marginal zone lymphoma (SMZL), and Waldenstrom's macroglobulinemia. In some embodiments, the lymphoma is selected from the group consisting of acute lymphoblastic leukemia (ALL), chronic lymphocytic leukemia (CLL), diffuse large B-cell lymphoma (DLBCL), follicular lymphoma, mantle cell lymphoma (MCL), marginal zone B-cell lymphoma (MZL), mucosa-associated lymphoid tissue lymphoma (MALT), and non-Hodgkin's lymphoma. In some embodiments, the lymphoma is non-Hodgkin's lymphoma. In some embodiments, the cancer is relapsed and refractory acute myeloid leukemia.

[0145] In certain embodiments, the tumor is cancer. In certain embodiments, the tumor is a blood cancer. In certain embodiments, the tumor is selected from the group consisting of multiple myeloma, leukemia, lymphoma, and myeloid malignancies. Non-limiting examples of blood cancers include multiple myeloma, leukemia, and lymphoma. Non-limiting examples of leukemias include acute myeloid leukemia (AML), chronic myeloid leukemia (CML), acute lymphocytic leukemia (ALL), chronic lymphocytic leukemia (CLL), acute promyelocytic leukemia (APL), mixed phenotype acute leukemia (MLL), hairy cell leukemia, and B-cell prolymphocytic leukemia. The lymphoma can be Hodgkin's lymphoma or non-Hodgkin's lymphoma. Non-limiting examples of myeloid malignancies include myelodysplastic syndromes (MDS), myeloproliferative neoplasms (MPN), myeloid / lymphoid neoplasms (e.g., myeloid / lymphoid neoplasms with eosinophilia and rearrangements of platelet-derived growth factor receptor alpha (PDGFRA), platelet-derived growth factor receptor beta (PDGFRB), or fibroblast growth receptor 1 (FGFR1), or myeloid / lymphoid neoplasms with PCM1-JAK2), acute myeloid leukemia (AML), blastic plasmacytoid dendritic cell neoplasm, B-lymphoblastic leukemia / lymphoma, and T-lymphoblastic leukemia / lymphoma. In certain embodiments, the myeloid malignancies include myelodysplastic syndromes.

[0146] In certain embodiments, the tumor is a B-cell malignancy. Non-limiting examples of B-cell malignancies include B-cell lymphoma (BCL), B-cell acute lymphocytic leukemia (ALL), B-cell chronic lymphocytic leukemia (CLL), multiple myeloma (MM), CLL with Richter's transformation, and CNS lymphoma. B-cell lymphomas include B-cell non-Hodgkin's lymphoma (NHL) and B-cell Hodgkin's lymphoma.

[0147] In various embodiments, administration of an anti-Clec12A antibody or antigen-binding fragment thereof described herein, or a fragment thereof, results in a reduction in the prevalence, frequency, level and / or amount of one or more symptoms or biomarkers of a Clec12A associated condition as described herein or known in the art, for example at least a 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 60%, 70%, 80%, 90%, 95%, 99% or 100% reduction in the one or more symptoms or biomarkers compared to a previous measurement or baseline in the subject.

[0148] In some embodiments, administration of an anti-Clec12A antibody or antigen-binding fragment thereof described herein to a subject having cancer results in a greater reduction or amelioration of one or more symptoms or biomarkers of cancer than a reference antibody (e.g., an antibody that cross-competes for Clec12A binding) under comparable conditions.

[0149] In some embodiments, the anti-Clec12A antibodies or antigen-binding fragments thereof described herein exhibit a reduced effective dose compared to a reference protein (e.g., an antibody that cross-competes for Clec12A binding). For example, the effective dose of the anti-Clec12A antibodies described herein may be, for example, less than 1,000 mg / dose, for example, less than 900 mg / dose, 800 mg / dose, 700 mg / dose, 600 mg / dose, 500 mg / dose, 550 mg / dose, 400 mg / dose, 350 mg / dose, 300 mg / dose, 200 mg / dose, 100 mg / dose, 50 mg / dose, 25 mg / dose, or less. In certain examples, the effective dose of the anti-Clec12A antibodies disclosed herein is lower than the effective or recommended or approved dose of the reference antibody, and the dose of the reference antibody may be, for example, 900 mg / dose, or 600 mg / dose. Alternatively, or in combination with the dosages disclosed herein, the anti-Clec12A antibodies described herein can be effectively or usefully administered less than once per week, e.g., less than once per week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks, 11 weeks, 12 weeks, 3 months, 4 months, 5 months, 6 months, 7 months, 8 months, 9 months, 10 months, 11 months, or year. In certain instances, the effective or useful administration frequency of the anti-Clec12A antibodies disclosed herein is less than the effective or recommended or approved administration frequency of the reference antibody, which can be administered weekly (e.g., at a dosage of 300-600 mg depending on the subject's weight) or every two weeks (e.g., at a dosage of 300-1200 mg depending on the subject's weight).

[0150] In some embodiments, the anti-Clec12A antibodies or antigen-binding fragments thereof described herein can be administered at lower doses compared to a reference protein, e.g., an antibody that cross-competes for Clec12A binding, to achieve a similar, equally effective, equally effective, or substantially effective outcome, when the anti-Clec12A antibody is administered in the same, equivalent, or substantially equivalent formulation and / or by the same, equivalent, or substantially equivalent route of administration as the reference (e.g., an antibody that cross-competes for Clec12A binding). In some embodiments, the anti-Clec12A antibodies described herein can be administered at extended intervals compared to a reference antibody (e.g., an antibody that cross-competes for Clec12A binding) to achieve a similar, equally effective, equally effective, or substantially effective outcome, when the anti-Clec12A antibody is administered in the same, equivalent, or substantially equivalent formulation and / or by the same, equivalent, or substantially equivalent route of administration as the reference. In some embodiments, the anti-Clec12A antibodies described herein may be administered in a reduced number of unit doses and / or for a shortened treatment period to achieve a similar, equally effective, equally effective or substantially effective outcome compared to a reference antibody, when the anti-Clec12A antibody is administered in the same, equivalent or substantially equivalent formulation and / or by the same, equivalent or substantially equivalent route of administration as the reference (e.g. an antibody that cross-competes for Clec12A binding).

[0151] According to some such embodiments, an administered dose of an anti-Clec12A antibody described herein may be less likely to induce an adverse reaction, e.g., an adverse immune reaction, when administered to a subject than an effective dose of a reference antibody, e.g., an antibody that cross-competes for Clec12A binding. Thus, in various embodiments, an anti-Clec12A antibody disclosed herein may be less likely to induce an adverse reaction or side effect, per unit of activity administered, than a reference antibody. In various embodiments, an anti-Clec12A antibody disclosed herein may be less likely to induce an adverse reaction or side effect of a particular severity, per unit of activity administered, than a reference antibody. In various embodiments, an anti-Clec12A antibody disclosed herein may induce one or more adverse reactions or side effects to a lesser extent, or in fewer patients, than a reference antibody, per unit of activity administered. Examples of possible adverse reactions or side effects associated with administration of an antibody capable of binding to Clec12A may include headache, nasopharyngitis, back pain, nausea, diarrhea, hypertension, upper respiratory tract infection, abdominal pain, vomiting, anemia, cough, peripheral edema, and / or urinary tract infection.

[0152] In some embodiments, upon administration to a subject (e.g., in a single dose), an anti-Clec12A antibody or antigen-binding fragment thereof described herein is measured at an increased level in plasma at a defined time (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14 days or more) after administration compared to the level of a control (e.g., an antibody that cross-competes for Clec12A binding) at the same defined time. For example, at a given time point after administration of a single dose, the level of an anti-Clec12A antibody described herein is at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, 200%, 300%, 400%, or 500% higher than the corresponding level of the reference antibody.

[0153] In some embodiments, the anti-Clec12A antibodies or antigen-binding fragments thereof described herein are measured at increased levels in plasma at a defined time (e.g. 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14 days or more) after administration (e.g. of a single dose) compared to the level of a control at the same defined time. For example, at a given time point after administration of a single dose, the level of an anti-Clec12A antibody described herein is at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, 200%, 300%, 400%, or 500% higher than the corresponding level of a reference antibody.

[0154] In some embodiments, the anti-Clec12A antibodies described herein have an increased half-life (e.g. compared to a control, e.g. a reference antibody, e.g. an antibody that cross-competes for Clec12A binding) and therefore the anti-Clec12A antibody can be administered to a subject over extended intervals, for example the anti-Clec12A antibody can be administered once every week, every 2 weeks, every 3 weeks, every 4 weeks, every 6 weeks, every 8 weeks, or more.

[0155] In some embodiments, a therapeutically effective amount of an anti-Clec12A antibody or antigen-binding fragment thereof described herein is about 90%, 80%, 70%, 60%, 50%, 40%, 30%, 20%, 10%, or 5% of the effective amount of a reference therapeutic protein, e.g., an antibody that cross-competes for Clec12A binding. In some embodiments, a single dose of an anti-Clec12A antibody or antigen-binding fragment described herein achieves the same therapeutic effect as two or more doses of the reference antibody.

[0156] In some embodiments, the anti-Clec12A antibodies or antigen-binding fragments thereof described herein are administered at a dose that is about 90%, 80%, 70%, 60%, 50%, 40%, 30%, 20%, 10%, or 5% of the concentration of the target antigen (e.g., Clec12A) in the subject.

[0157] In some embodiments, the anti-Clec12A antibodies or antigen-binding fragments thereof described herein can be physically introduced into a subject using any of a variety of methods and delivery systems known to those skilled in the art. Exemplary routes of administration of the formulations disclosed herein include intravenous, intramuscular, subcutaneous, intraperitoneal, spinal or other parenteral routes of administration, such as by injection or infusion. As used herein, the phrase "parenteral administration" refers to a method of administration other than enteral and topical administration, usually by injection, and includes, but is not limited to, intravenous, intramuscular, intraarterial, intrathecal, intralymphatic, intralesional, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intraarticular, subcapsular, subarachnoid, intraspinal, epidural, and intrasternal injection and infusion, and in vivo electroporation. In some embodiments, the formulations are administered by a parenteral route, such as a topical, epidermal, or mucosal route of administration, for example, intranasally, orally, intravaginally, rectally, sublingually, or topically. Also, administration can be, for example, once, multiple times, and / or over one or more extended periods of time.

[0158] In some embodiments, the anti-Clec12A antibodies or antigen-binding fragments thereof described herein can be used in a number of diagnostic and therapeutic applications. For example, detectably labeled versions of the engineered antibodies described herein can be used in assays to detect the presence or amount of Clec12A in a sample (e.g., a biological sample). The engineered antibodies described herein can be used in in vitro assays to study binding to Clec12A. In some embodiments, the anti-Clec12A antibodies described herein can be used as positive controls in assays designed to identify additional novel compounds that would otherwise be useful in treating Clec12A-associated disorders. For example, the anti-Clec12A antibodies described herein can be used as positive controls in assays to identify additional compounds (e.g., small molecules, aptamers, or antibodies) that bind to Clec12A.

[0159] The anti-Clec12A antibodies or antigen-binding fragments thereof described herein can be used to monitor a subject, e.g., a subject having, suspected of having, at risk of developing, or undergoing treatment for one or more Clec12A-associated conditions. Monitoring can include determining the amount or activity of Clec12A in the subject's, e.g., serum, of the subject. In some embodiments, the evaluation is performed at least 1 hour, e.g., at least 2, 4, 6, 8, 12, 24, or 48 hours, or at least 1 day, 2 days, 4 days, 10 days, 13 days, 20 days, or more, or at least 1 week, 2 weeks, 4 weeks, 10 weeks, 13 weeks, 20 weeks, or more, after administration of an anti-Clec12A antibody described herein. The subject can be evaluated at one or more of the following time periods: before the start of treatment; during treatment; or after one or more elements of the treatment have been administered. The evaluation can include evaluating the need for further treatment, e.g., evaluating whether the dosage, frequency of administration, or duration of treatment should be changed. It may also include evaluating the need to add or remove selected therapies, for example, adding or removing any treatments for a Clec12A-associated disorder described herein.

[0160] Formulation and Administration In various embodiments, the antibodies or antigen-binding fragments thereof described herein (e.g., anti-Clec12A antibodies described herein) can be incorporated into pharmaceutical compositions. Such pharmaceutical compositions can be useful, for example, for the prevention and / or treatment of diseases, such as Clec12A-associated disorders. Pharmaceutical compositions can be formulated by methods known to those of skill in the art (such as those described in Remington's Pharmaceutical Sciences, 17th edition, ed. Alfonso R. Gennaro, Mack Publishing Company, Easton, Pa. (1985)).

[0161] An appropriate means of administration can be selected based on the age and condition of the subject. A single dose of a pharmaceutical composition comprising an antibody or fragment thereof described herein (e.g., an anti-Clec12A antibody described herein) can be selected from the range of 0.001 to 1000 mg per kg of body weight. Meanwhile, the dose can be selected from the range of 0.001 to 100,000 mg per kg of body weight, but the present disclosure is not limited to such a range. The dose and administration method vary depending on the patient's body weight, age, condition, etc., and can be appropriately selected by a person skilled in the art as needed.

[0162] In various embodiments, the pharmaceutical composition can be formulated to include a pharma- ceutically acceptable carrier or excipient. Examples of pharma-ceutically acceptable carriers include, but are not limited to, any and all physiologically compatible solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, etc. The compositions of the present invention can include pharma-ceutically acceptable salts, such as acid addition salts or base addition salts.

[0163] In various embodiments, compositions comprising the antibodies described herein, e.g., sterile preparations for injection, can be formulated according to conventional pharmaceutical practice using distilled water for injection as a vehicle. For example, isotonic solutions containing saline or glucose and other supplements such as D-sorbitol, D-mannose, D-mannitol, sodium chloride, and the like, can be used as aqueous solutions for injection, optionally in combination with suitable solubilizers, e.g., alcohols such as ethanol and polyhydric alcohols such as propylene glycol or polyethylene glycol, non-ionic surfactants such as polysorbate 80™, HCO-50, and the like.

[0164] As disclosed herein, the pharmaceutical compositions may be in any form known in the art, including, for example, liquid, semi-solid, and solid dosage forms, such as liquid solutions (e.g., injectable and infusion solutions), dispersions or suspensions, tablets, pills, powders, liposomes, and suppositories.

[0165] The selection or use of any particular form may depend, in part, on the intended mode of administration and therapeutic application. For example, compositions, including compositions intended for systemic or local delivery, may be in the form of injections or drops. For example, compositions may be formulated for administration by parenteral modes (e.g., intravenous, subcutaneous, intraperitoneal, or intramuscular injection). As used herein, parenteral administration refers to modes of administration other than enteral and topical administration, usually by injection, and includes, but is not limited to, intravenous, intranasal, intraocular, intrapulmonary, intramuscular, intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intrapulmonary, intraperitoneal, transtracheal, subcutaneous, subcuticular, intraarticular, subcapsular, subarachnoid, intraspinal, epidural, intracerebral, intracranial, intracarotid, and intrasternal injection and infusion.

[0166] The route of administration may be parenteral, e.g., by injection, intranasal, pulmonary, or transdermal. Administration may be systemic or local, by intravenous, intramuscular, intraperitoneal, or subcutaneous injection.

[0167] In various embodiments, the pharmaceutical composition of the present invention can be formulated as a solution, microemulsion, dispersion, liposome, or other ordered structure suitable for stable storage at high concentration. Sterile injectables can be prepared by incorporating the required amount of the composition described herein with one or a combination of the ingredients listed above in an appropriate solvent, followed by filtration sterilization as necessary. In general, dispersions are prepared by mixing the composition described herein with a basic dispersion medium and other required ingredients from those listed above into a sterile vehicle. In the case of sterile powders for preparing sterile injectables, the preparation method includes vacuum drying and freeze-drying, which produces a powder of the composition described herein and any desired additional ingredients (see below) from a previously sterile-filtered solution of the composition described herein and any desired additional ingredients. The proper fluidity of the solution can be maintained, for example, by the use of a coating such as lecithin, by maintaining the required particle size in the case of dispersions, and by the use of surfactants. Prolonged absorption of the injectable composition can be achieved by including an agent that delays absorption, for example, monostearate salts and gelatin, in the composition.

[0168] Pharmaceutical compositions can be administered parenterally in the form of injectable formulations, including sterile solutions or suspensions in water or another pharma- ceutically acceptable liquid. For example, pharmaceutical compositions can be formulated by suitably combining therapeutic molecules with pharma- ceutically acceptable vehicles or media, such as sterile water and saline, vegetable oils, emulsifiers, suspending agents, surfactants, stabilizers, flavoring excipients, diluents, vehicles, preservatives, binders, and then mixing in unit dose forms required by generally accepted pharmaceutical practice. The amount of active ingredient contained in the pharmaceutical formulation is such that a suitable dose within the specified range is provided. Non-limiting examples of oily liquids include sesame oil and soybean oil, which may be combined with benzyl benzoate or benzyl alcohol as a solubilizer. Other items that may be included are buffers, such as phosphate buffer or sodium acetate buffer, soothing agents, such as procaine hydrochloride, stabilizers, such as benzyl alcohol or phenol, and antioxidants. Formulated injections can be packaged in suitable ampoules.

[0169] In some embodiments, the compositions can be formulated for storage at temperatures below 0° C. (e.g., −20° C. or −80° C.). In some embodiments, the compositions can be formulated for storage at 2-8° C. (e.g., 4° C.) for up to 2 years (e.g., 1 month, 2 months, 3 months, 4 months, 5 months, 6 months, 7 months, 8 months, 9 months, 10 months, 11 months, 1 year, 11 / 2 years, or 2 years). Thus, in some embodiments, the compositions described herein are storage stable at 2-8° C. (e.g., 4° C.) for at least 1 year.

[0170] In certain instances, the pharmaceutical composition can be formulated as a solution. In some embodiments, the composition can be formulated as a buffered solution suitable for storage at, for example, 2-8° C. (e.g., 4° C.) at an appropriate concentration.

[0171] The composition comprising one or more engineered antibodies described herein can be formulated into an immunoliposome composition. Such formulations can be prepared by methods known in the art. Liposomes with enhanced circulation time are disclosed, for example, in U.S. Patent No. 5,013,556.

[0172] In certain embodiments, the composition can be formulated with a carrier that protects the compound from rapid release, such as a controlled release formulation, including implants and microencapsulated delivery systems.Biodegradable biocompatible polymers can be used, such as ethylene vinyl acetate, polyanhydrides, polyglycolic acid, collagen, polyorthoesters, and polylactic acid.Many methods for preparing such formulations are known in the art.See, for example, JR Robinson (1978) "Sustained and Controlled Release Drug Delivery Systems", Marcel Dekker, Inc., New York.

[0173] In some embodiments, the composition can be formulated into a composition suitable for pulmonary administration (e.g., administration by inhaler or nebulizer) to a mammal, such as a human. Methods for formulating such compositions are well known in the art. Dry powder inhalation formulations and systems suitable for administering the formulations are also known in the art. Pulmonary administration can be oral and / or nasal administration. Examples of pharmaceutical devices for pulmonary delivery include metered dose inhalers, dry powder inhalers (DPIs), and nebulizers. For example, the compositions described herein can be administered to the lungs of a subject by dry powder inhalers. These inhalers are propellant-free devices that deliver dispersible and stable dry powder formulations to the lungs. Dry powder inhalers are well known in the medical arts and include, but are not limited to, TURBOHALER® (AstraZeneca; London, England), AIR® inhaler (ALKERMES®; Cambridge, Mass.); ROTAHALER® (GlaxoSmithKline; London, England); and ECLIPSE™ (Sanofi-Aventis; Paris, France). See, e.g., PCT Publication Nos. WO04 / 026380, WO04 / 024156, and WO01 / 78693. DPI devices have been used for pulmonary administration of polypeptides such as insulin and growth hormone. In some embodiments, the compositions described herein can be administered intrapulmonary by a metered dose inhaler. These inhalers rely on propellants to deliver discrete doses of the compound to the lungs. Additional devices and methods of pulmonary administration are described, for example, in US Patent Application Publication Nos. 20050271660 and 20090110679, the disclosures of each of which are incorporated by reference herein in their entireties.

[0174] In some embodiments, the composition can be formulated for delivery to the eye, for example, in the form of a pharma- ceutically acceptable solution, suspension, or ointment.The formulation used for eye treatment can be in the form of a sterile aqueous solution, with additional components, such as, but not limited to, preservatives, buffers, tonicity agents, antioxidants and stabilizers, non-ionic wetting or clarifying agents, and thickeners.The preparations described herein can be administered topically to the eye of a subject in need of treatment (e.g., a subject suffering from AMD) in a conventional manner, for example, in the form of drops, or by immersing the eye in a treatment solution containing one or more compositions.

[0175] In certain embodiments, various devices for introducing drugs into the vitreous cavity of eye can be suitable for administering the compositions described herein.For example, US Patent Application Publication No. 2002 / 0026176 describes a drug-containing plug that can be inserted through the sclera to protrude into the vitreous cavity and deliver pharmaceuticals into the vitreous cavity.In another example, US Patent No. 5,443,505 describes an implantable device for introduction into the suprachoroidal space or avascular area for sustained release of drugs into the interior of the eye.US Patent Nos. 5,773,019 and 6,001,386 each disclose an implantable drug delivery device that can be attached to the scleral surface of eye. Additional methods and devices for delivering therapeutic agents to the eye (e.g., delivery via transcleral patches and contact lenses) are described, for example, in Ambati and Adamis (2002) Prog Retin Eye Res 21(2):145-151; Ranta and Urtti (2006) Adv Drug Delivery Rev 58(11):1164-1181; Barocas and Balachandran (2008) Expert Opin Drug Delivery 5(1):1-10(10); Gulsen and Chauhan (2004) Invest Opthalmol Vis Sci 45:2342-2347; Kim et al. (2007) Ophthalmic Res 39:244-254; and PCT Publication No. WO04 / 073551, the disclosures of which are incorporated herein by reference in their entireties.

[0176] In certain embodiments, administration of the anti-Clec12A antibodies described herein is accomplished by administering to a subject a nucleic acid encoding the antibody. The nucleic acid encoding the therapeutic antibodies described herein can be incorporated into a genetic construct used as part of a gene therapy protocol to deliver a nucleic acid that can be used to express and produce the antibody in a cell. Such component expression constructs can be administered in a therapeutically effective carrier, e.g., any formulation or composition that can effectively deliver the component gene to a cell in vivo. Approaches include viral vectors, including recombinant retroviruses, adenoviruses, adeno-associated viruses, lentiviruses, and herpes simplex virus-1 (HSV-1), or insertion of the gene of interest into a recombinant bacterial or eukaryotic plasmid. Viral vectors can be directly transfected into cells. Plasmid DNA can be delivered, for example, with the aid of cationic liposomes (lipofectin) or derivatized polylysine conjugates, gramicidin S, artificial viral envelopes, or other such intracellular carriers, and can also be delivered by direct injection or CaPO4 precipitation of the gene construct (see, for example, WO04 / 060407).Examples of suitable retroviruses include pLJ, pZIP, pWE, and pEM, which are known to those of skill in the art (see, e.g., Eglitis et al. (1985) Science 230:1395-1398; Danos and Mulligan (1988) Proc Natl Acad Sci USA 85:6460-6464; Wilson et al. (1988) Proc Natl Acad Sci USA 85:3014-3018; Armentano et al. (1990) Proc Natl Acad Sci USA 87:6141-6145; Huber et al. (1991) Proc Natl Acad Sci USA 88:8039-8043; Ferry et al. (1991) Proc Natl Acad Sci USA 88:8377-8381; Chowdhury et al. (1999) Proc Natl Acad Sci USA 88:8377-8381; Chowdhury et al. (1999) Proc Natl Acad Sci USA 88:8377-8381). (1991) Science 254:1802-1805; van Beusechem et al. (1992) Proc Natl Acad Sci USA 89:7640-7644; Kay et al. (1992) Human Gene Therapy 3:641-647; Dai et al. (1992) Proc Natl Acad Sci USA 89:10892-10895; Hwu et al. (1993) J Immunol 150:4104-4115; U.S. Patent Nos. 4,868,116, and 4,980,286; and PCT Publication Nos. WO89 / 07136, WO89 / 02468, WO89 / 05345, and WO92 / 07573). Another viral gene delivery system utilizes vectors derived from adenovirus (see, e.g., Berkner et al. (1988) BioTechniques 6:616; Rosenfeld et al. (1991) Science 252:431-434; and Rosenfeld et al. (1992) Cell 68:143-155). Suitable adenoviral vectors derived from the adenovirus strain Ad type 5 dl324 or other strains of adenovirus (e.g., Ad2, Ad3, Ad7, etc.) are known to those of skill in the art.Yet another viral vector system useful for delivery of a gene of interest is the adeno-associated virus (AAV). See, e.g., Flotte et al. (1992) Am J Respir Cell Mol Biol 7:349-356; Samulski et al. (1989) J Virol 63:3822-3828; and McLaughlin et al. (1989) J Virol 62:1963-1973.

[0177] In various embodiments, subcutaneous administration may be accomplished by a device such as a syringe, a prefilled syringe, an auto-injector (e.g., disposable or reusable), a pen injector, a patch injector, a wearable injector, a portable syringe infusion pump with a subcutaneous infusion set, or other device for combining with an antibody drug for subcutaneous injection.

[0178] The injection system of the present disclosure may use a delivery pen as described in U.S. Pat. No. 5,308,341. Pen devices most commonly used for self-delivery of insulin to diabetic patients are well known in the art. Such devices may include at least one injection needle (e.g., a 31-gauge needle about 5-8 mm in length) and are typically pre-filled with one or more therapeutic unit doses of a therapeutic solution, useful for rapid delivery of the solution to a subject with as little pain as possible. One drug delivery pen includes a vial holder that can receive a vial of a therapeutic drug or other drug. The pen may be a completely mechanical device or may be combined with electronic circuitry to precisely set and / or indicate to the user the dosage of the medication to be injected. See, e.g., U.S. Pat. No. 6,192,891. In some embodiments, the needle of the pen device is disposable and the kit includes one or more disposable replacement needles. Pen devices suitable for delivery of any one of the presently characterized compositions are also described, e.g., in U.S. Pat. Nos. 6,277,099; 6,200,296; and 6,146,361. The disclosures of each of these are incorporated herein by reference in their entirety. Microneedle-based pen devices are described, for example, in U.S. Patent No. 7,556,615, the disclosure of which is incorporated herein by reference in its entirety. See also Precision Pen Injector (PPI) device, MOLLY™, manufactured by Scandinavian Health Ltd.

[0179] In some embodiments, the compositions described herein can be therapeutically delivered to a subject by local administration. As used herein, "local administration" or "local delivery" can refer to delivery of a composition or agent that does not depend on transport to its intended target tissue or site via the vascular system. For example, the composition can be delivered by injection or implantation of the composition or agent, or by injection or implantation of a device containing the composition or agent. In certain embodiments, after local administration near the target tissue or site, the composition or agent, or one or more components thereof, can diffuse to the intended target tissue or site that is not the site of administration.

[0180] In some embodiments, the compositions provided herein can be in unit dosage form suitable for self-administration.Such unit dosage form can typically be provided in a container such as vial, cartridge, pre-filled syringe or disposable pen.Administration device such as the administration device described in U.S. Patent No. 6,302,855 can also be used with the injection system described herein, for example.

[0181] The appropriate dose of the compositions described herein that can treat or prevent a disorder in a subject may depend on a variety of factors, including, for example, the age, sex, and weight of the subject to be treated, as well as the particular inhibitor compound used. For example, a different dose of a composition comprising an antibody described herein may be required to treat a subject with a Clec12A-associated disorder, compared to a dose of a different formulation of the antibody. Other factors that affect the dose administered to a subject include, for example, the type or severity of the disorder. For example, a subject with a Clec12A-associated disorder may require administration of a different dosage than a subject with another Clec12A-associated disorder. Other factors may include, for example, other medical disorders that may be concurrently or previously affecting the subject, the overall health of the subject, the genetic predisposition of the subject, diet, duration of administration, excretion rate, drug combination, and any other additional therapies administered to the subject. It should also be understood that the specific dosage and treatment regimen for any particular subject may also be adjusted based on the judgment of the treating physician.

[0182] The compositions described herein can be administered as a fixed dose or in milligrams per kilogram (mg / kg) doses. In some embodiments, the dose can be selected to reduce or avoid antibody production or other host immune responses to one or more of the antibodies or antigen-binding fragments thereof in the composition. While not intended to be limiting in any way, exemplary dosages of antibodies, such as the compositions described herein, include, for example, 1-1000 mg / kg, 1-100 mg / kg, 0.5-50 mg / kg, 0.1-100 mg / kg, 0.5-25 mg / kg, 1-20 mg / kg, and 1-10 mg / kg. Exemplary dosages of the compositions described herein include, but are not limited to, 0.1 mg / kg, 0.5 mg / kg, 1.0 mg / kg, 2.0 mg / kg, 4 mg / kg, 8 mg / kg, or 20 mg / kg.

[0183] The pharmaceutical solution may include a therapeutically effective amount of the compositions described herein. Such effective amounts can be readily determined by one of skill in the art based in part on the effect of the administered composition or the combined effect of the composition and one or more additional active agents when multiple agents are used. The therapeutically effective amount of the compositions described herein may also vary depending on factors such as the individual's medical condition, age, sex, and weight, as well as the ability of the composition (and one or more additional active agents) to induce a desired response in the individual, e.g., improvement of at least one condition parameter, e.g., improvement of at least one symptom of a Clec12A-associated disorder. For example, a therapeutically effective amount of the compositions described herein may inhibit (reduce the severity or eliminate the occurrence) and / or prevent a particular disorder and / or any one of the symptoms of a particular disorder known in the art or described herein. A therapeutically effective amount is also an amount in which any toxic or deleterious effects of the composition are outweighed by the therapeutically beneficial effects.

[0184] Suitable human doses of any of the compositions described herein can be further evaluated, for example, in a Phase I dose escalation study. See, e.g., van Gurp et al. (2008) Am J Transplantation 8(8):1711-1718; Hanouska et al. (2007) Clin Cancer Res 13(2,part 1):523-531; and Hetherington et al. (2006) Antimicrobial Agents and Chemotherapy 50(10):3499-3500.

[0185] Toxicity and therapeutic efficacy of the compositions can be determined by known pharmaceutical procedures in cell cultures or experimental animals (e.g., animal models of any Clec12A-associated disorder). These procedures can be used, for example, in the LD 50 (lethal dose for 50% of the population) and ED 50 It can be used to determine the therapeutically effective dose in 50% of the population. The dose ratio between toxic and therapeutic effects is the therapeutic index, which is the LD50 / ED 50 The therapeutic index can be expressed as a ratio of 0.01 to 0.01. Compositions described herein that exhibit high therapeutic indices are preferred. Compositions that exhibit toxic side effects may be used, but care must be taken to design a delivery system that targets such compounds to the site of affected tissue, minimizing potential damage to normal cells, thereby reducing side effects.

[0186] Those skilled in the art will appreciate that the data obtained from cell culture assays and animal studies can be used in formulating a range of dosages for use in humans. Appropriate dosages of the compositions described herein will generally be administered at ED 100 mg with little or no toxicity. 50 The circulating concentration of the composition is within a range including 100 mg / kg / day. The dosage may vary within this range depending on the dosage form employed and / or the route of administration used. For the compositions described herein, a therapeutically effective amount can be initially estimated from cell culture analysis. A dose can be formulated in an animal model to achieve a circulating plasma concentration range that includes I0 (i.e., the concentration of antibody that achieves half-maximal inhibition of symptoms) as determined in cell culture. Such information can be used to more accurately determine useful doses in humans. Levels in plasma can be measured, for example, by high performance liquid chromatography. In some embodiments, for example, when local administration (e.g., to the eye or joint) is desired, cell culture or animal models can be used to determine the dosage required to achieve a therapeutically effective concentration in the local site.

[0187] Combination therapy In various embodiments, the anti-Clec12A antibody described herein may be included in a course of treatment that further comprises administering to the subject at least one additional agent.In various embodiments, the additional agent administered in combination with the anti-Clec12A antibody described herein may be a drug chemotherapeutic agent.In various embodiments, the additional agent administered in combination with the antibody described herein may be an agent that inhibits inflammation.

[0188] In some embodiments, the anti-Clec12A antigen-binding fragment is a single chain variable fragment (scFv) with specificity for human Clec12A. In some embodiments, the anti-Clec12A scFv can be conjugated (e.g. linked) to a therapeutic agent (e.g. chemotherapeutic agents and radioactive atoms) to bind to, deliver the therapeutic agent to, and kill cancer cells expressing human Clec12A. In some embodiments, the anti-Clec12A antibody is linked to a therapeutic agent. In some embodiments, the therapeutic agent is a chemotherapeutic agent, a cytokine, a radioactive atom, an siRNA, or a toxin. In some embodiments, the therapeutic agent is a chemotherapeutic agent. In some embodiments, the therapeutic agent is a radioactive atom.

[0189] In some embodiments, the method can be performed in combination with other therapies for Clec12A-associated disorders.For example, the composition can be administered to the subject simultaneously with, before, or after chemotherapy.In some embodiments, the composition can be administered to the subject simultaneously with, before, or after adoptive therapy.

[0190] In various embodiments, the additional agent administered in combination with the anti-Clec12A antibody described herein may be administered simultaneously with the anti-Clec12A antibody, on the same day as the anti-Clec12A antibody, or in the same week as the anti-Clec12A antibody. In various embodiments, the additional agent administered in combination with the anti-Clec12A antibody described herein may be administered in a single formulation with the anti-Clec12A antibody. In certain embodiments, the additional agent is administered in a manner that is temporally separated from the administration of the anti-Clec12A antibody described herein, for example, one or more hours before or after, one or more days before or after, one or more weeks before or after, or one or more months before or after the administration of the anti-Clec12A antibody. In various embodiments, the frequency of administration of one or more additional agents may be the same as, similar to, or different from the frequency of administration of the anti-Clec12A antibody described herein.

[0191] Included in combination therapy are therapeutic regimens that include the administration of two different antibodies as described herein and / or therapeutic regimens that include the administration of an antibody as described herein via multiple formulations and / or routes of administration.

[0192] In some embodiments, the composition may be formulated with one or more additional therapeutic agents, such as additional therapies for treating or preventing Clec12A-associated disorders (e.g., cancer or autoimmune disorders) in a subject. Additional agents for treating Clec12A-associated disorders in a subject may vary depending on the specific disorder to be treated, but include, but are not limited to, rituximab, cyclophosphamide, doxorubicin, vincristine, prednisone, osfamide, carboplatin, etoposide, dexamethasone, cytarabine, cisplatin, cyclophosphamide, or fludarabine.

[0193] The compositions described herein may replace or enhance previous or currently administered therapy.For example, when treated with the compositions described herein, administration of one or more additional active agents may be stopped or reduced following administration of the anti-Clec12A antibody described herein, for example, administered at a lower level, for example, at a lower level of a reference antibody that cross-competes for Clec12A binding.In some embodiments, administration of the previous therapy may be maintained.In some embodiments, the previous therapy is maintained until the level of the composition reaches a sufficient level to provide a therapeutic effect.The two therapies may be administered in combination.

[0194] Recombinant Gene Technology In accordance with the present disclosure there may be employed conventional molecular biology, microbiology, and recombinant DNA techniques within the skill of the art. Such techniques are described in the literature (e.g., Sambrook, Fritsch & Maniatis, Molecular Cloning: A Laboratory Manual, Second Edition (1989); Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY; DNA Cloning: A Practical Approach, Volumes I and II (DNGlover ed.1985); Oligonucleotide Synthesis (MJGait ed.1984); Nucleic Acid Hybridization(BDHames&S.J.Higgins eds.(1985));Transcription And Translation(BDHames&S.J.Higgins, eds.(1984));Animal Cell Culture(RIFreshney, ed.(1986));Immobilized Cells and Enzymes(IRL Press,(1986));B.Perbal,A Practical Guide To Molecular Cloning(1984);FMAusubel et al.(eds.),Current Protocols in Molecular Biology,John Wiley & Sons, Inc. (1994).

[0195] Recombinant expression of a gene, such as a nucleic acid encoding a polypeptide, such as an anti-Clec12A antibody described herein, may include the construction of an expression vector comprising a nucleic acid encoding the polypeptide. Once a polynucleotide is obtained, a vector for the production of the polypeptide can be produced by recombinant DNA technology using techniques known in the art. Known methods can be used to construct an expression vector comprising a polypeptide coding sequence and appropriate transcriptional and translational control signals. These methods include, for example, in vitro recombinant DNA techniques, synthetic techniques, and in vivo genetic recombination.

[0196] The expression vector can be transferred to a host cell by conventional techniques and the transfected cells can then be cultured by conventional techniques to produce the polypeptide.

[0197] All publications, patent applications, patents, and other references mentioned herein are incorporated by reference in their entirety.Furthermore, the materials, methods, and examples are illustrative only and are not intended to be limiting.Unless otherwise defined, technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which this invention belongs.Although methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, suitable methods and materials are described herein. EXAMPLES

[0198] The following examples illustrate some of the preferred modes of making and practicing the present invention, however, it should be understood that these examples are for illustrative purposes only and are not intended to limit the scope of the invention.

[0199] Example 1 Identification and characterization of anti-Clec12A antibodies This example demonstrates the derivation and characterization of the binding affinity of anti-Clec12A antibodies.

[0200] Hybridoma technology was used to select and screen Clec12A antibodies. Selections were performed on CHO-S cells overexpressing Clec12A. Antibodies were selected against a variety of sequences. From a range of 74 antibodies, 16 antibodies were selected for soluble and cellular Clec12A binding affinity.

[0201] The binding affinity of Clec12A antibodies was determined by Biacore analysis.

[0202] Briefly, human anti-Clec12A antibodies derived from and produced in hybridomas were tested for binding affinity to soluble Clec12A in a Biacore SPR system. Briefly, antibodies were immobilized on an anti-mouse IgG sensor chip and bound to soluble Clec12A-His protein (Clec12A extracellular domain fused to a 6xHis tag (SEQ ID NO: 79)) at 100 nM in solution phase. The resulting Biacore trace is shown in Figure 1 and a summary of the binding kinetics is given in Table 1.

[0203] [Table 1]

[0204] Overall, the affinities of the anti-Clec12A antibodies ranged from a high of 0.1 pM (clone C) to a low of 3.8 nM (clone B).

[0205] Example 2. On-cell binding of anti-Clec12A antibodies This example shows on-cell binding of anti-Clec12A antibodies measured by flow cytometry.

[0206] On-cell binding of anti-Clec12A antibodies was assessed by flow cytometry on Clec12A-overexpressing and parental CHO-S cell lines labeled with 100 nM Calcein-AM. Each antibody was tested on both cell lines to confirm target-specific binding on cells.

[0207] Flow cytometry chromatograms are shown in Figure 2. Results showed a 2-3 log shift of Clec12A-positive strains relative to background binding on the corresponding parental cell lines.

[0208] Overall, serial dilution flow cytometry results demonstrated saturable binding to U937 cells that endogenously express Clec12A.

[0209] [Table 2]

[0210] As shown in Table 2, the EC50 values ​​calculated from the curves indicate that each of the four antibodies binds to U937 cells with high affinity, ranging from 0.2 nM (Antibody D) to 3.3 nM (Antibody B). The higher affinity observed in Clec12A-positive cells versus soluble protein suggests that these antibodies A and D bind to epitopes that may be more naturally presented on cells than in Clec12A-His fusions.

[0211] Example 3. Epitope binning by cross-competition A cross-competition epitope binning assay was performed to determine whether any of the anti-Clec12A antibodies bound to unique, non-competing epitopes derived from the two reference anti-Clec12A antibodies (Reference 1 and Reference 2).

[0212] Epitope binning / ligand blocking was performed using a sandwich-format cross-blocking assay. Control anti-target IgG was loaded onto the AHQ sensor, and unoccupied Fc binding sites on the sensor were blocked with an irrelevant human IgG1 antibody. The sensor was then exposed to 100 nM of target antigen, followed by exposure to a secondary anti-target antibody or ligand.

[0213] Further binding by a second antibody or ligand following antigen binding indicates an unoccupied epitope (non-competition), whereas lack of binding indicates epitope blocking (competition or ligand blocking).

[0214] [Table 3]

[0215] The ELISA assay method used for testing included that shown in the schematic diagram in Figure 4. Briefly, an anti-Clec12A reference control antibody was immobilized on an ELISA plate and the plate was blocked with BSA. Each test antibody was pre-incubated with soluble biotinylated Clec12A-His protein, and then the mixture was added to the plate. Bound soluble Clec12A was detected using HRP-conjugated streptavidin. Antibodies that bind to a non-competing epitope from the reference antibody will reflect the binding signal of the soluble protein to the reference antibody, while antibodies with the same epitope are blocked by the reference antibody reflected by no or low binding signal.

[0216] The results demonstrated that anti-Clec12A antibodies B, C and D bind to epitopes competing with Reference Antibody 1 and Reference Antibody 2, respectively, while antibody A binds to a unique epitope.

[0217] Example 4. Characterization of Clec12A binding of anti-Clec12A scFv In this example, we evaluate the non-specific binding of anti-Clec12A antibodies. To further evaluate the potential for anti-Clec12A antibodies to bind to non-specific membrane proteins, scFvs derived from the four lead antibodies were evaluated in a surface membrane protein (SMP) assay (Figure 5A).

[0218] The SMP assay used was an ELISA-based assay with human HEK-293 or insect SF9 cell membranes coated onto plates to test for non-specific binding to these membranes by the test antibodies, and included internal control high and low non-specific binding antibodies.

[0219] The high non-specific binding control used, sc209, was an antibody with off-target toxins in the clinic, whereas the low non-specific binding control, 5f9, showed no off-target toxins in the clinic (Figure 5B).

[0220] Overall, the results showed low non-specific binding with anti-Clec12A antibodies B, C and D (Figure 5B).

[0221] Example 5. Off-target screening panel assay This example demonstrates the specificity of anti-Clec12A scFv in an off-target binding assay.

[0222] Briefly, off-target binding assays were performed to test the specificity of anti-Clec12A scFv for Clec12A and other membrane proteins. Two selected Clec12A scFv clones were used in a "cut-down assay" to screen for binding to over 3000 human receptors.

[0223] In cut-down assays, the higher the binding, the more likely that an interaction is real. In general, hits labeled "V.weak" are unlikely to be real interactions. Anti-Clec12A antibody D did not show any non-specific interactions with any receptors other than Clec12A or receptors that would appear as artifacts in this assay.

[0224] This cut-down assay was used to screen clones for off-target activity, and selected scFv-Fc clones were tested more comprehensively in additional screening and confirmation assays.

[0225] Example 6. Sequencing of anti-Clec12A antibody clones The anti-Clec12A antibodies were sequenced and the variable region amino acid sequences are shown in Table 5 (heavy chain CDR sequences), Table 6 (heavy chain variable region sequences), Table 7 (light chain CDR sequences), and Table 8 (light chain variable region sequences).

[0226] All four heavy chain sequences have a unique CDR3, anti-Clec12A antibodies B and C are all derived from the same germline framework, and anti-Clec12A antibodies A and D are unique frameworks. All four light chain sequences have a unique CDR3, anti-Clec12A antibodies B and C are all derived from the same germline framework, and anti-Clec12A antibodies A and D are unique frameworks.

[0227] [Table 5]

[0228] [Table 6-1]

[0229] [Table 6-2]

[0230] [Table 7]

[0231] [Table 8]

[0232] Example 7. Expression of Clec12A protein In this example, expression of Clec12A protein is demonstrated.

[0233] A DNA fragment encoding human Clec12A ECD (H65-A265) was fused with an N-terminal 6x histidine tag (SEQ ID NO: 79) and then cloned into the pcDNA3.4 vector. The expression plasmid was transiently transfected into Expi293F cells using 293fection, and the expressed protein was purified from the culture using a Ni column followed by SEC.

[0234] [Table 9]

[0235] The results of the SMP assay and off-target binding assessment showed that the anti-Clec12A Clone D antibody is a high affinity and specific antibody. The antibody shows high affinity for Clec12A positive cells, but the affinity for soluble Clec12A is slightly lower, possibly due to the non-native Clec12A conformation in the soluble fusion protein. Furthermore, epitope binning data showed that the Clone D antibody competes with the epitopes targeted by the Ref. 1 and Ref. 2 antibodies.

[0236] Other embodiments Although many embodiments of the present invention are described herein, the disclosure and examples may be modified to provide other methods and compositions of the present invention. It will therefore be understood that the scope of the present invention should be defined by the appended claims in addition to the specific embodiments presented by way of example. All references cited herein are incorporated by reference in their entirety.

Claims

1. An anti-Clec12A antibody or antigen-binding fragment thereof, comprising: (a) an HCDR1 comprising the amino acid sequence of GGSISSSTYY (SEQ ID NO: 33), an HCDR2 comprising the amino acid sequence of THYRGST (SEQ ID NO: 34), and an HCDR3 comprising the amino acid sequence of ARELTGEVFDY (SEQ ID NO: 35); and an LCDR1 comprising the amino acid sequence of QSISSY (SEQ ID NO: 36), an LCDR2 comprising the amino acid sequence of AAS (SEQ ID NO: 23), and an LCDR3 comprising the amino acid sequence of QQSYSTPFT (SEQ ID NO: 37); (b) an HCDR1 comprising the amino acid sequence of GFTFSSYG (SEQ ID NO: 73), an HCDR2 comprising the amino acid sequence of ISYDGSDK (SEQ ID NO: 25), and an HCDR3 comprising the amino acid sequence of ARDKGYYFDY (SEQ ID NO: 26); and an LCDR1 comprising the amino acid sequence of QSVGNRY (SEQ ID NO: 27), an LCDR2 comprising the amino acid sequence of GAS (SEQ ID NO: 28), and an LCDR3 comprising the amino acid sequence of QQDYNLPLT (SEQ ID NO: 29); (c) an HCDR1 comprising the amino acid sequence of GFTFSSYG (SEQ ID NO: 73), an HCDR2 comprising the amino acid sequence of ISYDGSDK (SEQ ID NO: 25), and an HCDR3 comprising the amino acid sequence of ARDGSRYFDY (SEQ ID NO: 30); and an LCDR1 comprising the amino acid sequence of QSVHSKY (SEQ ID NO: 31), an LCDR2 comprising the amino acid sequence of GAS (SEQ ID NO: 28), and an LCDR3 comprising the amino acid sequence of QQDYNLPIT (SEQ ID NO: 32); (d) a heavy chain complementarity determining region (HCDR) 1 comprising the amino acid sequence of GGSISTYY (SEQ ID NO: 19), a HCDR2 comprising the amino acid sequence of IYYSGST (SEQ ID NO: 20), and a HCDR3 comprising the amino acid sequence of AREDYYGSGSPFDY (SEQ ID NO: 21); and a light chain complementarity determining region (LCDR) 1 comprising the amino acid sequence of QGIRYD (SEQ ID NO: 22), a LCDR2 comprising the amino acid sequence of AAS (SEQ ID NO: 23), and a LCDR3 comprising the amino acid sequence of LQDYNFPRT (SEQ ID NO: 24); (e) an HCDR1 comprising the amino acid sequence of GGSISTYY (SEQ ID NO: 19), an HCDR2 comprising the amino acid sequence of IYFSGST (SEQ ID NO: 38), and an HCDR3 comprising the amino acid sequence of AREDYYGSGSPFDY (SEQ ID NO: 21); and an LCDR1 comprising the amino acid sequence of QGIRND (SEQ ID NO: 39), an LCDR2 comprising the amino acid sequence of AAS (SEQ ID NO: 23), and an LCDR3 comprising the amino acid sequence of LQDYNYPRT (SEQ ID NO: 40); (f) an HCDR1 comprising the amino acid sequence of GGSISTDY (SEQ ID NO: 41), an HCDR2 comprising the amino acid sequence of IYFSGST (SEQ ID NO: 38), and an HCDR3 comprising the amino acid sequence of AREDYYGSGSPFDY (SEQ ID NO: 21); and an LCDR1 comprising the amino acid sequence of QDIRND (SEQ ID NO: 42), an LCDR2 comprising the amino acid sequence of AAS (SEQ ID NO: 23), and an LCDR3 comprising the amino acid sequence of LQDYNFPRT (SEQ ID NO: 24); (g) an HCDR1 comprising the amino acid sequence of GFTFSSYG (SEQ ID NO: 73), an HCDR2 comprising the amino acid sequence of ISYDGSDK (SEQ ID NO: 25), and an HCDR3 comprising the amino acid sequence of ARDGQFYFDY (SEQ ID NO: 43); and an LCDR1 comprising the amino acid sequence of QSVTSRY (SEQ ID NO: 44), an LCDR2 comprising the amino acid sequence of GAS (SEQ ID NO: 28), and an LCDR3 comprising the amino acid sequence of QQDYNLPLT (SEQ ID NO: 29); (h) an HCDR1 comprising the amino acid sequence of GFTFSNYG (SEQ ID NO: 45), an HCDR2 comprising the amino acid sequence of ISYDGSDK (SEQ ID NO: 25), and an HCDR3 comprising the amino acid sequence of ARDSGRYFFDY (SEQ ID NO: 46); and an LCDR1 comprising the amino acid sequence of QSVSSRS (SEQ ID NO: 47), an LCDR2 comprising the amino acid sequence of GPS (SEQ ID NO: 48), and an LCDR3 comprising the amino acid sequence of HQDYNLPLT (SEQ ID NO: 49); or (i) an HCDR1 comprising the amino acid sequence of GFTFSKYG (SEQ ID NO:50), an HCDR2 comprising the amino acid sequence of IWYDGSIK (SEQ ID NO:51), and an HCDR3 comprising the amino acid sequence of ARGSLWFGEFYFDY (SEQ ID NO:52); and an LCDR1 comprising the amino acid sequence of QGISSA (SEQ ID NO:53), an LCDR2 comprising the amino acid sequence of DAS (SEQ ID NO:54), and an LCDR3 comprising the amino acid sequence of QQFNNYPRT (SEQ ID NO:55).

2. 2. The anti-Clec12A antibody or antigen-binding fragment thereof of claim 1, comprising: (a) an immunoglobulin heavy chain variable (VH) region comprising an amino acid sequence at least about 80% identical to QLQLQESGPGLVKPSETLSLTCTVSGGSISSSTYYWGWIRQPPRKGLEWIGSTHYRGSTYYNPSLKSRVTISVDTSKNQFSLKVSSVTAADTAVYYCARELTGEVFDYWGQGTLVTVSS (SEQ ID NO: 7); and an immunoglobulin light chain variable (V L ) region comprising an amino acid sequence at least about 80% identical to DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSYSTPFTFGPGTKVDIK (SEQ ID NO:8); (b) an immunoglobulin heavy chain variable (V) comprising an amino acid sequence that is at least about 80% identical to QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYYVDSVKGRFTISRDNSKNTLYLHMNSLRAEDTAVYYCARDKGYYFDYWGQGTLVTVSS (SEQ ID NO: 3). Ｈ ) area, and An immunoglobulin light chain variable (V) comprising an amino acid sequence that is at least about 80% identical to EIVMTQSPATHLSPGERATLSCRASQSVGNRYLSWYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO: 4). Ｌ )region; (c) an immunoglobulin heavy chain variable (V) comprising an amino acid sequence that is at least about 80% identical to QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYSADSVKGRFNISRDNSKNTLYLQMNSLRAEDTAVYYCARDGSRYFDYWGQGTLVTVSS (SEQ ID NO: 5); Ｈ ) area, and An immunoglobulin light chain variable (V) comprising an amino acid sequence that is at least about 80% identical to EIFMTQSPATHLSPGERATLSCRASQSVHSKYLSWYQQKPGQAPSLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPITFGQGTRLEIK (SEQ ID NO: 6). Ｌ )region; (d) an immunoglobulin heavy chain variable (VH) region comprising an amino acid sequence at least about 80% identical to QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYWSWIRQPPGKGLEWIGYIYYSGSTKYNPSLKSRVTISVDTSKNLFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSS (SEQ ID NO: 1); and an immunoglobulin light chain variable (V L ) region comprising an amino acid sequence at least about 80% identical to IQMTQSPSSLSASVGDRVTITCRASQGIRYDLGWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO:2); (e) an immunoglobulin heavy chain variable (V) comprising an amino acid sequence that is at least about 80% identical to QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYWSWIRQPPGKGLEWLGYIYFSGSTNYNPSLKSRLTISVAASKSQFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSS (SEQ ID NO: 9); Ｈ ) area, and An immunoglobulin light chain variable (V) comprising an amino acid sequence that is at least about 80% identical to AIQMTQSPSSLSASVGDRVTITCRASQGIRNDLGWFQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTYFTLTISSLQPEDSATYYCLQDYNYPRTFGQGTKVEIK (SEQ ID NO: 10). Ｌ )region; (f) an immunoglobulin heavy chain variable (V) comprising an amino acid sequence that is at least about 80% identical to QVQLQESGPGLVKPSETLSLTCTVSGGGSISTDYWSWIRQPPGKGLEWIGYIYFSGSTKYNPSLKSRVTISVDTSKNQFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSS (SEQ ID NO: 11); Ｈ ) area, and An immunoglobulin light chain variable (V) comprising an amino acid sequence that is at least about 80% identical to AIQMTQSPSSLSASVGDRVTITCRASQDIRNDLGWFQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO: 12). Ｌ )region; (g) an immunoglobulin heavy chain variable (V) comprising an amino acid sequence that is at least about 80% identical to QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGEGLEWVTVISYDGSDKYYADSVKGRFTISRDNSKSTLFLQMNSLRAEDTAVYYCARDGQFYFDYWGQGTLVTVSS (SEQ ID NO: 13). Ｈ ) area, and An immunoglobulin light chain variable (V) comprising an amino acid sequence that is at least about 80% identical to EIVMTQSPATHLSPGESATLSCRASQSVTSRYLSWYQQKPGQAPRLLMYGASTRPTGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO: 14). Ｌ )region; (h) an immunoglobulin heavy chain variable (V) comprising an amino acid sequence that is at least about 80% identical to QVQLVESGGGVVQPGRSLRLSCAASGFTFSNYGMHWVRQAPGKGLEWVAVISYDGSDKSYKDSVKGRFTIARDDNSKNTLYLQMNSLRAEDTAVYYCARDSGRYFFDYWGQGTLVTVSS (SEQ ID NO: 15). Ｈ ) area, and An immunoglobulin light chain variable (V) comprising an amino acid sequence that is at least about 80% identical to Ｌ ) area; or (i) an immunoglobulin heavy chain variable (V) fragment comprising an amino acid sequence that is at least about 80% identical to QVKLVESGGGVVQPGRSLRLSCAASGFTFSKYGMHWVRQAPGKGLEWVAFIWYDGSIKNYADSVKGRFTTSRDNSKNTLYLQMNSLRAEDTAVYYCARGSLWFGEFYFDYWGQGTLVTVSS (SEQ ID NO: 17); Ｈ ) area, and An immunoglobulin light chain variable (V) comprising an amino acid sequence that is at least about 80% identical to AIQLTQSPSSLSASVGDRVTITCRTSQGISSALAWYQQKPGKTPKLLIYDASSLESGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQFNNYPRTFGQGTKVEIK (SEQ ID NO: 18). Ｌ )region.

3. 2. The anti-Clec12A antibody or antigen-binding fragment thereof of claim 1, comprising: (a) an immunoglobulin heavy chain variable (VH) region comprising the amino acid sequence of QLQLQESGPGLVKPSETLSLTCTVSGGSISSSTYYWGWIRQPPRKGLEWIGSTHYRGSTYYNPSLKSRVTISVDTSKNQFSLKVSSVTAADTAVYYCARELTGEVFDYWGQGTLVTVSS (SEQ ID NO: 7); and an immunoglobulin light chain variable (V L ) region comprising the amino acid sequence of: DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSYSTPFTFGPGTKVDIK (SEQ ID NO:8); (b) an immunoglobulin heavy chain variable (V) comprising the amino acid sequence of QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYYVDSVKGRFTISRDNSKNTLYLHMNSLRAEDTAVYYCARDKGYYFDYWGQGTLVTVSS (SEQ ID NO: 3); Ｈ ) area, and Immunoglobulin light chain variable (V) comprising the amino acid sequence of EIVMTQSPATHLSPGERATLSCRASQSVGNRYLSWYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO: 4). Ｌ )region; (c) an immunoglobulin heavy chain variable (V) comprising the amino acid sequence of QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYSADSVKGRFNISRDNSKNTLYLQMNSLRAEDTAVYYCARDGSRYFDYWGQGTLVTVSS (SEQ ID NO: 5); Ｈ ) area, and Immunoglobulin light chain variable (V) comprising the amino acid sequence of EIFMTQSPATHLSPGERATLSCRASQSVHSKYLSWYQQKPGQAPSLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPITFGQGTRLEIK (SEQ ID NO: 6). Ｌ ) area; or (d) an immunoglobulin heavy chain variable (V H ) comprising the amino acid sequence of QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYWSWIRQPPGKGLEWIGYIYYSGSTKYNPSLKSRVTISVDTSKNLFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSS (SEQ ID NO: 1); an immunoglobulin light chain variable (V L ) region comprising the amino acid sequence of: IQMTQSPSSLSASVGDRVTITCRASQGIRYDLGWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO:2); (e) an immunoglobulin heavy chain variable (V) comprising the amino acid sequence of QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYWSWIRQPPGKGLEWLGYIYFSGSTNYNPSLKSRLTISVAASKSQFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSS (SEQ ID NO: 9); Ｈ ) region, and an immunoglobulin light chain variable (V) region comprising the amino acid sequence of AIQMTQSPSSLSASVGDRVTITCRASQGIRNDLGWFQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTYFTLTISSLQPEDSATYYCLQDYNYPRTFGQGTKVEIK (SEQ ID NO: 10). Ｌ )region; (f) an immunoglobulin heavy chain variable (V) comprising the amino acid sequence of QVQLQESGPGLVKPSETLSLTCTVSGGGSISTDYWSWIRQPPGKGLEWIGYIYFSGSTKYNPSLKSRVTISVDTSKNQFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSS (SEQ ID NO: 11); Ｈ ) area, and Immunoglobulin light chain variable (V) comprising the amino acid sequence of AIQMTQSPSSLSASVGDRVTITCRASQDIRNDLGWFQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO: 12). Ｌ )region; (g) an immunoglobulin heavy chain variable (V) comprising the amino acid sequence of QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGEGLEWVTVISYDGSDKYYADSVKGRFTISRDNSKSTLFLQMNSLRAEDTAVYYCARDGQFYFDYWGQGTLVTVSS (SEQ ID NO: 13); Ｈ ) area, and An immunoglobulin light chain variable (V) comprising the amino acid sequence of EIVMTQSPATHLSPGESATLSCRASQSVTSRYLSWYQQKPGQAPRLLMYGASTRPTGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO: 14). Ｌ )region; (h) an immunoglobulin heavy chain variable (V) comprising the amino acid sequence of QVQLVESGGGVVQPGRSLRLSCAASGFTFSNYGMHWVRQAPGKGLEWVAVISYDGSDKSYKDSVKGRFTIARDDNSKNTLYLQMNSLRAEDTAVYYCARDSGRYFFDYWGQGTLVTVSS (SEQ ID NO: 15); Ｈ ) area, and An immunoglobulin light chain variable (V) comprising the amino acid sequence of: Ｌ ) area; or (i) an immunoglobulin heavy chain variable (V) comprising the amino acid sequence of QVKLVESGGGVVQPGRSLRLSCAASGFTFSKYGMHWVRQAPGKGLEWVAFIWYDGSIKNYADSVKGRFTTSRDNSKNTLYLQMNSLRAEDTAVYYCARGSLWFGEFYFDYWGQGTLVTVSS (SEQ ID NO: 17); Ｈ ) area, and Immunoglobulin light chain variable (V) comprising the amino acid sequence of AIQLTQSPSSLSASVGDRVTITCRTSQGISSALAWYQQKPGKTPKLLIYDASSLESGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQFNNYPRTFGQGTKVEIK (SEQ ID NO: 18). Ｌ )region.

4. The anti-Clec12A antibody or antigen-binding fragment thereof is an IgA antibody, an IgG antibody, an IgE antibody, an IgM antibody, a bispecific or multispecific antibody, a Fab fragment, a Fab' fragment, an F(ab')2 fragment, an Fd' fragment, an Fd fragment, an isolated CDR or set thereof; a single chain variable fragment (scFv), a polypeptide-Fc fusion, a single domain antibody, a camelid antibody, a masked antibody, a small modular immunopharmaceutical ("SMIPs™"), a single chain, a tandem 2. The anti-Clec12A antibody or antigen-binding fragment thereof of claim 1, wherein the antibody or antigen-binding fragment is selected from the group consisting of: Mudiabody, VHH, Anticalin, Nanobody, Minibody, BiTE, Ankyrin repeat protein, DARPIN, Avimer, DART, TCR-like antibody, Adnectin, Affilin, Transbody, Affibody, TrimerX, MicroProtein, Fynomer, Centyrin, and KALBITOR.

5. The anti-Clec12A antibody or antigen-binding fragment thereof of claim 1 , wherein the anti-Clec12A antibody or antigen-binding fragment thereof is a monoclonal antibody or a single-chain variable fragment (scFv).

6. The anti-Clec12A antibody or antigen-binding fragment thereof according to claim 1 , wherein the anti-Clec12A antibody is an antibody comprising an IgG constant region.

7. The anti-Clec12A antibody or antigen-binding fragment thereof of claim 1 , wherein the anti-Clec12A antigen-binding fragment is a single-chain variable fragment (scFv).

8. The anti-Clec12A antibody or antigen-binding fragment thereof of claim 7, wherein the scFv comprises a leader sequence, a heavy chain variable sequence, a GS linker, and a light chain variable sequence.

9. 9. The anti-Clec12A antibody or antigen-binding fragment of claim 8, wherein the scFv comprises a sequence having at least about 80% identity to: (a) QLQLQESGPGLVKPSETLSLTCTVSGGSISSSTYYWGWIRQPPRKGLEWIGSTHYRGSTYYNPSLKSRVTISVDTSKNQFSLKVSSVTAADTAVYYCARELTGEVFDYWGQGTLVTVSSASTGGGGSGGGGGSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSYSSTPFTFGPGTKVDIK (SEQ ID NO:59); (b) QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYWSWIRQPPGKGLEWIGYIYYSGSTKYNPSLKSRVTISVDTSKNLFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSSASTGGGGSGGGGGSGGGGSAIQMTQSPSSLSASVGDRVTITCRASQGIRYDLGWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO:56); (c) QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAIVISYDGSDKYYVDSVKGRFTISRDNSKNTLYLHMNSLRAEDTAVYYCARDKGYYFDYWGQGTLVTVSSASTGGGGSGGGGGSGGGGGSEIVMTQSPATLSLSPGERATLSCRASQSVGNRYLSWYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO:57); or (d) QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYSADSVKGRFNISRDNSKNTLYLQMNSLRAEDTAVYYCARDGSRYFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSEIFMTQSPATHSLPGERATLSCRASQSVHSKYLSWYQQKPGQAPSLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPITFGQGTRLEIK (SEQ ID NO:58).

10. 10. The anti-Clec12A antibody or antigen-binding fragment thereof of claim 9, wherein the scFv comprises the following sequence: (a) QLQLQESGPGLVKPSETLSLTCTVSGGSISSSTYYWGWIRQPPRKGLEWIGSTHYRGSTYYNPSLKSRVTISVDTSKNQFSLKVSSVTAADTAVYYCARELTGEVFDYWGQGTLVTVSSASTGGGGSGGGGGSGGGGSDIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSYSSTPFTFGPGTKVDIK (SEQ ID NO:59); (b) QVQLQESGPGLVKPSETLSLTCTVSGGSISTYYWSWIRQPPGKGLEWIGYIYYSGSTKYNPSLKSRVTISVDTSKNLFSLKLSSVTAADTAVYYCAREDYYGSGSPFDYWGQGTLVTVSSASTGGGGSGGGGGSGGGGSAIQMTQSPSSLSASVGDRVTITCRASQGIRYDLGWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQDYNFPRTFGQGTKVEIK (SEQ ID NO:56); (c) QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAIVISYDGSDKYYVDSVKGRFTISRDNSKNTLYLHMNSLRAEDTAVYYCARDKGYYFDYWGQGTLVTVSSASTGGGGSGGGGGSGGGGGSEIVMTQSPATLSLSPGERATLSCRASQSVGNRYLSWYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPLTFGGGTKVEIK (SEQ ID NO:57); or (d) QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAPGKGLEWVAVISYDGSDKYSADSVKGRFNISRDNSKNTLYLQMNSLRAEDTAVYYCARDGSRYFDYWGQGTLVTVSSASTGGGGSGGGGSGGGGSEIFMTQSPATHSLPGERATLSCRASQSVHSKYLSWYQQKPGQAPSLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQPEDFAVYYCQQDYNLPITFGQGTRLEIK (SEQ ID NO:58).

11. The anti-Clec12A or fragment thereof is －８ Less than M, about 10 －９ Less than M, about 10 －１０ Less than M, about 10 －１１ Less than M, about 10 －１２ Less than M or about 10 －１３ Dissociation constant (K Ｄ 2. The anti-Clec12A antibody of claim 1 , which binds to Clec12A at the C1A1 domain.

12. The above K Ｄ The anti-Clec12A antibody of claim 11, wherein the Clec12A antibody has a specific binding affinity of about 0.1 pM or less.

13. The above K Ｄ The anti-Clec12A antibody of claim 11, wherein the antibody has a concentration of about 0.05 to about 0.5 pM.

14. The above K Ｄ The anti-Clec12A antibody of claim 11, wherein the antibody has a concentration of about 0.1 nM to about 5.0 nM.

15. The above K Ｄ The anti-Clec12A antibody of claim 14, wherein the antibody has a binding affinity of about 0.3 nM to about 3.5 nM.

16. The anti-Clec12A antibody of claim 1, wherein the EC50 is from about 1 to about 100 nM.

17. A composition for treating cancer in a subject, comprising an anti-Clec12A antibody or antigen-binding fragment thereof according to any one of claims 1 to 16.

18. 18. The composition of claim 17, wherein the cancer is selected from leukemia, lymphoma, and myeloma.

19. The composition described in claim 17, wherein the composition is administered together with one or more additional agents.

20. The composition of claim 19, wherein the one or more additional agents are selected from an antibody, a chemotherapeutic agent, and radiation therapy.

21. A pharmaceutical composition comprising the anti-Clec12A antibody or antigen-binding fragment thereof according to any one of claims 1 to 16, and a pharma- ceutically acceptable carrier.

22. A nucleic acid sequence encoding an amino acid sequence that is at least about 90% identical to any one of the amino acid sequences according to any one of claims 1 to 16.

23. A vector comprising the nucleic acid sequence of claim 22.

24. A cell comprising the vector of claim 23.

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