Anti-TREM2 antibodies and related methods

Abstract

To provide isolated humanized anti-TREM2 antibodies and methods for preparing and using the antibodies.SOLUTION: Provided herein is an isolated humanized anti-TREM2 antibody comprising an active human IgG1 Fc region, a heavy chain including a variable heavy chain sequence having three heavy chain CDR sequences CDR-H1, CDR-H2 and CDR-H3, and a light chain including a variable light chain sequence having three light chain CDR sequences CDR-L1, CDR-L2 and CDR-L3. Also provided are methods and compositions for enhancing immune response and / or treating an immune related condition such as cancer in individuals, comprising killing, disabling or depleting non-stimulatory bone marrow cells using the antibody or antigen binding fragment thereof.SELECTED DRAWING: Figure 9A

Description

Technical Field

[0001] Cross - Reference to Related Applications This application claims the benefit of U.S. Provisional Patent Application No. 62 / 597,827, filed Dec. 12, 2017, and U.S. Provisional Patent Application No. 62 / 648,089, filed Mar. 26, 2018, each of which is hereby incorporated by reference in its entirety for all purposes.

[0002] Sequence Listing This application contains a Sequence Listing that has been submitted via EFS - Web and is hereby incorporated by reference in its entirety. The ASCII copy created on XX / XX / 20XX is named XXXXXUS_sequencelisting.txt and is X,XXX,XXX bytes in size.

Background Art

[0003] Background Immunity plays a role in preventing tumor growth. A complex microenvironment can occur within the lesion, and despite the recruitment of T cells, in many cases, there is no effective control of the developing tumor. Understanding the balance between tumor elimination and tumor escape may depend on understanding the different roles that myeloid cells play in the tumor microenvironment.

[0004] The myeloid population in the tumor microenvironment prominently includes monocytes and neutrophils (which may be broadly grouped as myeloid - derived suppressor cells), macrophages, and dendritic cells. The intratumoral myeloid population has long been considered to be non - stimulatory or suppressive as a whole, but more recently, it has been recognized that not all tumor - infiltrating myeloid cells are equivalent.

[0005] In normal tissues, many of these bone marrow cells are essential, particularly for wound repair, for the proper functioning of both innate and adaptive immunity. However, in the cancer setting, a generally described population is one of substantially excessive macrophages and dysfunctional or skewed populations of these and other cell types. “Macrophage” infiltration, when considered an aggregated population defined by a single marker such as CD68 or CD163, correlates with a worse outcome for the subject for multiple tumor types ((de Visser, Cancer Immunol Immunother, 2008;57:1531-9 (Non-Patent Document 1)); (Hanada et al., Int J Urol 2000;7:263-9 (Non-Patent Document 2)); (Yao et al., Clin Cancer Res, 520, 2001;7:4021-6 (Non-Patent Document 3)); (Ruffell et al., PNAS, 523 2012;109:2796-801 (Non-Patent Document 4))). However, the phenotypic and functional subsetting of macrophages in the tumor microenvironment is complicated by the similarity of macrophages and dendritic cells and is a problem in tumor biology. Morphological criteria are often applied to the problem, and one approach to distinguishing dendritic cells from macrophages was based, in the former case, on a more projecting or dendritic morphology and, in the latter case, on a more veiled or bulbous morphology (Bell et al., J Exp Med 555, 1999;190:1417-26 (Non-Patent Document 5)). Other groups attempt to distinguish based on genetic and cell surface markers.

[0006] The antigen-presenting compartments within tumors are diverse, and T cells can distinguish the characteristics of antigen-presenting cells (APCs). Since T cells are the major drivers of tumor immunity, it would be important to understand the precise characteristics of the APCs of the same species. Myeloid cells are prominent among the cells that can present tumor-derived antigens to T cells and thereby maintain the latter in an activated state. Antigen presentation occurs within the tumor itself and can affect the function of tumor cell cytotoxic T lymphocytes (CTLs). The activation of T cells by antigen-presenting cells (APCs) is an important component in antigen-specific immune responses and the killing of tumor cells. Since these myeloid populations represent the major T cell interaction partners and antigen-presenting cells for subsequent tumor-reactive cytotoxic T lymphocytes, understanding their identification could lead to therapeutic approaches.

[0007] Related patent applications include PCT / US2015 / 052682 (Patent Document 1) filed on September 28, 2015, and PCT / US2016 / 054104 (Patent Document 2) filed on September 28, 2016, each of which is hereby incorporated by reference in its entirety for all purposes.

[0008] All patents, patent applications, publications, documents, and papers cited herein are hereby incorporated by reference in their entirety.

Prior Art Documents

Patent Documents

[0009]

Patent Document 1

Patent Document 2

Non-Patent Documents

[0010]

Non-Patent Document 1

Non-Patent Document 2

[0011] Summary Isolated antibodies that bind to human TREM2 (SEQ ID NO: 15) and compete with the 37017 antibody (SEQ ID NO: 31 and 32) for binding to mouse TREM2 (SEQ ID NO: 17) are described herein.

[0012] In some embodiments, the antibody comprises CDR-H1 comprising the sequence set forth in SEQ ID NO: 9, CDR-H2 comprising the sequence set forth in SEQ ID NO: 10, CDR-H3 comprising the sequence set forth in SEQ ID NO: 11, CDR-L1 comprising the sequence set forth in SEQ ID NO: 12, CDR-L2 comprising the sequence set forth in SEQ ID NO: 13, and CDR-L3 comprising the sequence set forth in SEQ ID NO: 14.

[0013] In some embodiments, the antibody is afucosylated and comprises the VH sequence shown in SEQ ID NO: 1; the VL sequence shown in SEQ ID NO: 2; and an active human IgG1 Fc region.

[0014] In some embodiments, the antibody comprises all three heavy chain CDRs of the sequence shown in SEQ ID NO: 7 and all three light chain CDRs of the sequence shown in SEQ ID NO: 8.

[0015] In some embodiments, the antibody comprises a substitution of A to T at position 97 of the sequence shown in SEQ ID NO: 7; and a substitution of K to R at position 98 of the sequence shown in SEQ ID NO: 7.

[0016] In some embodiments, the antibody comprises a VH sequence as shown in SEQ ID NO:1, 3, or 5.

[0017] In some embodiments, the antibody comprises a VH sequence set forth in SEQ ID NO:1, 3, or 5 and a VL sequence set forth in SEQ ID NO:2, 4, or 6.

[0018] In some embodiments, the antibody comprises the VH sequence shown in SEQ ID NO:1.

[0019] In some embodiments, the antibody comprises the VH sequence shown in SEQ ID NO:1 and the VL sequence shown in SEQ ID NO:2.

[0020] In some embodiments, the antibody is the 37012 antibody.

[0021] In some embodiments, the antibody comprises the heavy chain sequence set forth in SEQ ID NO:25 and the light chain sequence set forth in SEQ ID NO:26.

[0022] In another aspect, described herein is an isolated antibody that binds to human TREM2 (SEQ ID NO: 15), which antibody competes with the 37017 antibody (SEQ ID NOs: 31 and 32) for binding to mouse TREM2 (SEQ ID NO: 17) and comprises an active human Fc region.

[0023] In some embodiments, the antibody is human, humanized, or chimeric.

[0024] In some embodiments, the antibody is humanized.

[0025] In some embodiments, the antibody has a cytoplasmic affinity of about 1, 2, 3, 4, or 5x10 as measured by a surface plasmon resonance (SPR) assay. -9 The following K D Binds to human TREM2 at

[0026] In some embodiments, the antibody is capable of specifically killing, depleting or neutralizing TREM2+ myeloid cells, optionally, unstimulated myeloid cells.

[0027] In some embodiments, the antibody has antibody-dependent cell-mediated cytotoxicity (ADCC) activity. In some embodiments, the antibody has antibody-mediated cell phagocytosis (ADCP) activity. In some embodiments, the antibody has complement-dependent cytotoxicity (CDC) activity.

[0028] In some embodiments, the antibody kills, inactivates, or depletes myeloid cells via antibody-dependent cell-mediated cytotoxicity (ADCC), antibody-mediated cell phagocytosis (ADCP) activity, or complement-dependent cytotoxicity (CDC).

[0029] In some embodiments, the antibody is at least one of a monoclonal antibody, a neutralizing antibody, an antagonist antibody, an agonist antibody, a polyclonal antibody, an IgG1 antibody, an IgG3 antibody, an afucosylated antibody, a bispecific antibody, a human antibody, a humanized antibody, a chimeric antibody, a full-length antibody, and antigen-binding fragments thereof.

[0030] In some embodiments, the antibody is a monoclonal antibody.

[0031] In some embodiments, the antibody is multispecific.

[0032] In some embodiments, the antibody is afucosylated.

[0033] In some embodiments, the antibody is an antigen-binding fragment thereof, Fab, Fab’, F(ab’)2, Fv, scFv, (scFv)2, a single-chain antibody molecule, a bispecific variable domain antibody, a single variable domain antibody, a linear antibody, or a V domain antibody.

[0034] In some embodiments, the antibody comprises a scaffold, and optionally, the scaffold is Fc, optionally human Fc.

[0035] In some embodiments, the antibody comprises a heavy chain constant region of a class selected from IgG, IgA, IgD, IgE, and IgM.

[0036] In some embodiments, the antibody comprises a heavy chain constant region of class IgG and a subclass selected from IgG1, IgG2, IgG3, and IgG4.

[0037] In some embodiments, the antibody comprises a heavy chain constant region of IgG1.

[0038] In some embodiments, the Fc comprises one or more modifications that result in an increase in half-life, an increase in ADCC activity, an increase in ADCP activity, or an increase in CDC activity as compared to Fc without the one or more modifications.

[0039] In some embodiments, the Fc binds to an Fcγ receptor selected from the group consisting of FcγRI, FcγRIIa, FcγRIIb, FcγRIIc, FcγRIIIa, and FcγRIIIb.

[0040] In another aspect, an isolated antibody for use in treating cancer is described herein, and the cancer is selected from solid tumors and hematological tumors.

[0041] In another aspect, an isolated antibody that competes with the antibodies described herein for binding to human TREM2 is described herein.

[0042] In another aspect, an isolated antibody that binds to the human TREM2 epitope to which the antibodies described herein bind is described herein.

[0043] In another aspect, an isolated polynucleotide or set of polynucleotides encoding the antibodies described herein, their V H their V L their light chain, their heavy chain, or their antigen-binding portion, optionally cDNA, is described herein.

[0044] In another aspect, a vector or set of vectors comprising a polynucleotide or set of polynucleotides as described herein is described herein.

[0045] In another aspect, host cells are described herein that contain a polynucleotide or set of polynucleotides as described herein, or a vector or set of vectors as described herein.

[0046] In another aspect, methods for producing an antibody are described herein that include expressing an antibody using a host cell as described herein and isolating the expressed antibody.

[0047] In another aspect, pharmaceutical compositions are described herein that contain an antibody as described herein and a pharmaceutically acceptable excipient.

[0048] In another aspect, methods for treating or preventing a disease or condition in a subject in need thereof are described herein that include administering to the subject an effective amount of an antibody or pharmaceutical composition as described herein.

[0049] In some embodiments, the disease or condition is cancer.

[0050] In some embodiments, the antibody binds to the extracellular domain of TREM2 on TREM2+ myeloid cells, and optionally, the myeloid cells are within a tumor. In one embodiment, the antibody binds to the extracellular domain of TREM2 on myeloid cells, and the myeloid cells are CD45 + , HLA-DR + , CD11c + , CD14 + , and BDCA3 - non-stimulatory myeloid cells, and the antibody kills, inactivates, or depletes the non-stimulatory myeloid cells via ADCC, CDC, and / or ADCP to a level less than the level of non-stimulatory myeloid cells present in the cancer prior to contacting the non-stimulatory myeloid cells with the antibody, and the non-stimulatory myeloid cells are CD45 + , HLA-DR + , CD14 - , CD11c + , BDCA1 - , and BDCA3 +It is present in a population of immune cells including stimulatory myeloid cells and non-stimulatory myeloid cells, and the killing, inactivation, or depletion of non-stimulatory myeloid cells treats cancer.

[0051] In some embodiments, the antibody kills, inactivates, or depletes myeloid cells via antibody-dependent cell-mediated cytotoxicity (ADCC), antibody-mediated cell phagocytosis (ADCP) activity, or complement-dependent cytotoxicity (CDC). In some embodiments, the antibody has receptor-ligand blocking, agonism, or antagonism activity.

[0052] In some embodiments, the subject is human. In some embodiments, the cancer is a solid cancer. In some embodiments, the cancer is a liquid cancer. In some embodiments, the cancer is selected from the group consisting of melanoma, kidney, hepatobiliary tract, head and neck squamous cell carcinoma (HNSC), pancreas, colon, bladder, glioblastoma, prostate, lung, breast, ovary, stomach, kidney, bladder, esophagus, kidney, melanoma, and mesothelioma. In some embodiments, the cancer is colon cancer or breast cancer.

[0053] In some embodiments, the contact enhances the subject's immune response. In some embodiments, the enhanced immune response is an adaptive immune response. In some embodiments, the enhanced immune response is a innate immune response.

[0054] In some embodiments, the subject has previously received, is receiving simultaneously, or will receive immunotherapy later. In some embodiments, the immunotherapy is a checkpoint inhibitor; a T cell checkpoint inhibitor; an anti-PD1 antibody; an anti-PDL1 antibody; an anti-CTLA4 antibody; adoptive T cell therapy; CAR-T cell therapy; dendritic cell vaccine; monocyte vaccine; an antigen-binding protein that binds to both T cells and antigen-presenting cells; a BiTE bispecific antigen-binding protein; a toll-like receptor ligand; a cytokine; a cytotoxic therapy; chemotherapy; radiotherapy; a small molecule inhibitor; a small molecule agonist; an immunomodulatory agent; and at least one of an epigenetic regulator. In some embodiments, the immunotherapy is an anti-PD1 antibody.

[0055] In another aspect, provided herein is a method of killing, inactivating, or depleting TREM2+ myeloid cells in a subject having cancer, comprising contacting the anti-TREM2 antibody described herein or the pharmaceutical composition described herein with myeloid cells, optionally, the myeloid cells are within the tumor.

[0056] In some embodiments, the antibody binds to the extracellular domain of TREM2 and the TREM2+ myeloid cells are non-stimulatory myeloid cells that are CD45 + , HLA-DR + , CD11c + , CD14 + , and BDCA3 - , and the antibody kills, inactivates, or depletes the non-stimulatory myeloid cells via ADCC, CDC, and / or ADCP to a level lower than the level of non-stimulatory myeloid cells present in the cancer prior to contacting the non-stimulatory myeloid cells with the antibody, the non-stimulatory myeloid cells are CD45 + , HLA-DR + , CD14 - , CD11c + , BDCA1 - , and BDCA3 + , and are present in a population of immune cells comprising non-stimulatory myeloid cells, and the contacting does not substantially kill, inactivate, or deplete the myeloid cells present outside the cancer and / or the stimulatory myeloid cells present in the cancer, and the killing, inactivation, or depletion of the non-stimulatory myeloid cells treats the cancer by enhancing the immune response to the cancer.

[0057] In some embodiments, the antibody kills myeloid cells by at least one of ADCC, CDC, and ADCP. In some embodiments, the antibody inactivates myeloid cells by at least one of ADCC, CDC, and ADCP. In some embodiments, the antibody depletes myeloid cells by at least one of ADCC, CDC, and ADCP. In some embodiments, the antibody has antibody-dependent cell-mediated cytotoxicity (ADCC) activity. In some embodiments, the antibody has complement-dependent cytotoxicity (CDC) activity. In some embodiments, the antibody has antibody-mediated phagocytosis (ADCP) activity. In some embodiments, the antibody has receptor-ligand blocking, agonism, or antagonism activity.

[0058] In some embodiments, the myeloid cells are stimulatory myeloid cells. In some embodiments, the myeloid cells are non-stimulatory myeloid cells. In some embodiments, the myeloid cells comprise at least one of dendritic cells, tumor-associated macrophages (TAM), neutrophils, or monocytes. In some embodiments, the myeloid cells are neutrophils. In some embodiments, the myeloid cells are tumor-associated macrophages. In some embodiments, the myeloid cells are within the tumor. In some embodiments, the myeloid cells are in a population of immune cells comprising stimulatory myeloid cells and non-stimulatory myeloid cells.

[0059] In some embodiments, the subject is human. In some embodiments, the cancer is a solid cancer. In some embodiments, the cancer is a liquid cancer. In some embodiments, the cancer is selected from the group consisting of melanoma, kidney, hepatobiliary tract, head and neck squamous cell carcinoma (HNSC), pancreas, colon, bladder, glioblastoma, prostate, lung, breast, ovary, stomach, kidney, bladder, esophagus, kidney, melanoma, and mesothelioma. In some embodiments, the cancer is colon cancer or breast cancer.

[0060] In some embodiments, the contact enhances the subject's immune response. In some embodiments, the enhanced immune response is an adaptive immune response. In some embodiments, the enhanced immune response is a innate immune response.

[0061] In some embodiments, the subject has previously received, is receiving concurrently, or will receive immunotherapy later. In some embodiments, the immunotherapy is a checkpoint inhibitor; a T cell checkpoint inhibitor; an anti-PD1 antibody; an anti-PDL1 antibody; an anti-CTLA4 antibody; adoptive T cell therapy; CAR-T cell therapy; dendritic cell vaccine; monocyte vaccine; an antigen-binding protein that binds to both T cells and antigen-presenting cells; a BiTE bispecific antigen-binding protein; a toll-like receptor ligand; a cytokine; a cytotoxic therapy; chemotherapy; radiation therapy; a small molecule inhibitor; a small molecule agonist; an immunomodulatory agent; and at least one of an epigenetic regulator. In some embodiments, the immunotherapy is an anti-PD1 antibody.

[0062] In another aspect, a method for detecting TREM2 in a subject having or suspected of having a disease or condition is described herein, the method comprising: (a) receiving a sample from the subject; and (b) detecting the presence or level of TF in the sample by contacting the sample with an antibody described herein.

[0063] In some embodiments, the disease or condition is cancer.

[0064] In some embodiments, the method described herein comprises administering a checkpoint inhibitor, optionally, the checkpoint inhibitor is an inhibitor of the PD1:PDL1 axis, optionally, the inhibitor is an antibody, and optionally, the antibody is an anti-PD1 antibody or an anti-PDL1 antibody.

[0065] In another aspect, a kit comprising an antibody or pharmaceutical composition and instructions for use disclosed herein is described herein. [Invention 1001] An isolated humanized antibody that binds to human TREM2 (SEQ ID NO: 15) and competes with 37017 antibody (SEQ ID NO: 31 and 32) for binding to mouse TREM2 (SEQ ID NO: 17). [Invention 1002] a. CDR-H1 comprising the sequence set forth in SEQ ID NO: 9 b. A CDR-H2 comprising the sequence set forth in SEQ ID NO: 10, c. A CDR-H3 comprising the sequence set forth in SEQ ID NO: 11, d. A CDR-L1 comprising the sequence set forth in SEQ ID NO: 12, e. A CDR-L2 comprising the sequence set forth in SEQ ID NO: 13, and f. A CDR-L3 comprising the sequence set forth in SEQ ID NO: 14, An isolated humanized antibody of [Invention 1001] comprising [Invention 1003] Is afucosylated and Comprises a VH sequence shown in SEQ ID NO: 1; a VL sequence shown in SEQ ID NO: 2; and an active human IgG1 Fc region, An isolated antibody of [Invention 1002]. [Invention 1004] An isolated antibody of [Invention 1002] comprising a VH sequence comprising a substitution of A to T at position 97 and a substitution of K to R at position 98 of the sequence shown in SEQ ID NO: 7. [Invention 1005] An isolated antibody of [Invention 1003] comprising a VH sequence shown in SEQ ID NO: 1, 3, or 5. [Invention 1006] An isolated antibody of [Invention 1005] comprising a VH sequence shown in SEQ ID NO: 1, 3, or 5; and a VL sequence shown in SEQ ID NO: 2, 4, or 6. [Invention 1007] An isolated antibody of [Invention 1003] comprising a VH sequence shown in SEQ ID NO: 1. [Invention 1008] An isolated antibody of [Invention 1007] comprising a VH sequence shown in SEQ ID NO: 1 and a VL sequence shown in SEQ ID NO: 2. [Invention 1009] An isolated antibody of [Invention 1008] which is the 37012 antibody. [Invention 1010] An isolated antibody of [Invention 1001] comprising a heavy chain sequence shown in SEQ ID NO: 25 and a light chain sequence shown in SEQ ID NO: 26. [Invention 1011] An isolated antibody that binds to human TREM2 (SEQ ID NO: 15), i) competes with the 37017 antibody (SEQ ID NOs: 31 and 32) for binding to mouse TREM2 (SEQ ID NO: 17), ii) contains an active human Fc region, said isolated antibody. [Inventive item 1012] a. CDR-H1 containing the sequence set forth in SEQ ID NO: 9, b. CDR-H2 containing the sequence set forth in SEQ ID NO: 10, c. CDR-H3 containing the sequence set forth in SEQ ID NO: 11, d. CDR-L1 containing the sequence set forth in SEQ ID NO: 12, e. CDR-L2 containing the sequence set forth in SEQ ID NO: 13, and f. CDR-L3 containing the sequence set forth in SEQ ID NO: 14 An isolated humanized antibody comprising. [Inventive item 1013] An isolated antibody of [Inventive item 1012] comprising a VH sequence comprising a substitution of A to T at position 97 and a substitution of K to R at position 98 of the sequence shown in SEQ ID NO: 7. [Inventive item 1014] An isolated antibody of [Inventive item 1013] comprising a VH sequence shown in SEQ ID NO: 1, 3, or 5. [Inventive item 1015] An isolated antibody of [Inventive item 1014] comprising a VH sequence shown in SEQ ID NO: 1, 3, or 5; and a VL sequence shown in SEQ ID NO: 2, 4, or 6. [Inventive item 1016] An isolated antibody of [Inventive item 1015] comprising a VH sequence shown in SEQ ID NO: 1. [Inventive item 1017] An isolated antibody of [Inventive item 1016] comprising a VH sequence shown in SEQ ID NO: 1 and a VL sequence shown in SEQ ID NO: 2. [Inventive item 1018] An isolated antibody of [Inventive item 1017] which is the 37012 antibody. [Inventive item 1019] An isolated antibody of [Inventive item 1012] comprising a heavy chain sequence shown in SEQ ID NO: 25 and a light chain sequence shown in SEQ ID NO: 26. [Inventive item 1020] An isolated antibody of the invention that binds to human TREM2 with a K of about 1, 2, 3, 4, or 5 x 10 -9 M or less D as measured by surface plasmon resonance (SPR) assay. [Invention 1021] An isolated antibody of the invention that is capable of specifically killing, depleting, or inactivating TREM2+ myeloid cells, optionally non-stimulatory myeloid cells, optionally intratumoral myeloid cells. [Invention 1022] An isolated antibody of the invention having antibody-dependent cell-mediated cytotoxicity (ADCC) activity. [Invention 1023] An isolated antibody of the invention having antibody-mediated cell phagocytosis (ADCP) activity. [Invention 1024] An isolated antibody of the invention having complement-dependent cytotoxicity (CDC) activity. [Invention 1025] An isolated antibody of the invention that is at least one of a monoclonal antibody, a neutralizing antibody, an antagonist antibody, an agonist antibody, a polyclonal antibody, an IgG1 antibody, an IgG3 antibody, an afucosylated antibody, a bispecific antibody, a human antibody, a chimeric antibody, a full-length antibody, and an antigen-binding fragment thereof. [Invention 1026] An isolated antibody of the invention that is a monoclonal antibody. [Invention 1027] An isolated antibody of the invention that is multispecific. [Invention 1028] An isolated antibody of the invention that is afucosylated. [Invention 1029] An isolated antibody of the invention that is an antigen-binding fragment thereof, Fab, Fab’, F(ab’)2, Fv, scFv, (scFv)2, a single-chain antibody molecule, a bispecific variable domain antibody, a single variable domain antibody, a linear antibody, or a V domain antibody. [Invention 1030] An isolated antibody of any of the present inventions, wherein the antibody comprises a scaffold, optionally the scaffold is Fc, and optionally the Fc is human Fc. [Inventive item 1031] An isolated antibody of any of the present inventions, comprising a heavy chain constant region of a class selected from IgG, IgA, IgD, IgE, and IgM. [Inventive item 1032] An isolated antibody of any of the present inventions, comprising the IgG class and a heavy chain constant region of a subclass selected from IgG1, IgG2, IgG3, and IgG4. [Inventive item 1033] An isolated antibody of any of the present inventions, comprising the heavy chain constant region of IgG1. [Inventive item 1034] An isolated antibody of any of the present inventions, wherein the Fc comprises one or more modifications that result in an increase in half-life, an increase in ADCC activity, an increase in ADCP activity, or an increase in CDC activity as compared to the Fc without the one or more modifications. [Inventive item 1035] An isolated antibody of any of the present inventions, wherein the Fc binds to an Fcγ receptor selected from the group consisting of FcγRI, FcγRIIa, FcγRIIb, FcγRIIc, FcγRIIIa, and FcγRIIIb. [Inventive item 1036] An isolated antibody of any of the present inventions, used for the treatment of cancer, wherein the cancer is selected from solid tumors and hematological tumors. [Inventive item 1037] An isolated antibody that competes with any of the antibodies of the present inventions for binding to human TREM2. [Inventive item 1038] An isolated antibody that binds to the human TREM2 epitope to which any of the antibodies of the present inventions binds. [Inventive item 1039] An isolated polynucleotide or set of polynucleotides encoding any of the antibodies of the present inventions, optionally the polynucleotide or set of polynucleotides is cDNA, its VH, its VL, its light chain, its heavy chain, or its antigen-binding portion. [Inventive item 1040] A vector or set of vectors comprising the polynucleotide or set of polynucleotides of [the present invention 1039]. [the present invention 1041] A host cell comprising the polynucleotide or set of polynucleotides of [the present invention 1039] or the vector or set of vectors of [the present invention 1040]. [the present invention 1042] A method for producing an antibody, comprising: a step of expressing the antibody in the host cell of [the present invention 1041], and a step of isolating the expressed antibody The method comprising the above. [the present invention 1043] A pharmaceutical composition comprising any one of the antibodies of [the present invention 1001] to [the present invention 1038] and a pharmaceutically acceptable excipient. [the present invention 1044] A method for treating or preventing a disease or condition in a subject in need thereof, comprising administering to the subject an effective amount of any one of the antibodies of [the present invention 1001] to [the present invention 1038] or the pharmaceutical composition of [the present invention 1043]. [the present invention 1045] The method of [the present invention 1044], wherein the disease or condition is cancer. [the present invention 1046] The method of [the present invention 1044], wherein the antibody binds to the extracellular domain of TREM2 on TREM2+ myeloid cells, and optionally the myeloid cells are within the tumor. [the present invention 1047] The antibody binds to the extracellular domain of TREM2 on myeloid cells, the myeloid cells are non-stimulatory myeloid cells that are CD45+, HLA-DR+, CD11c+, CD14+, and BDCA3-, the antibody kills, inactivates, or depletes the non-stimulatory myeloid cells via ADCC, CDC, and / or ADCP to a level lower than the level of non-stimulatory myeloid cells present in the cancer before contacting the non-stimulatory myeloid cells with the antibody, The non-stimulatory bone marrow cells are present in a population of immune cells comprising the stimulatory bone marrow cells and the non-stimulatory bone marrow cells that are CD45+, HLA-DR+, CD14-, CD11c+, BDCA1-, and BDCA3+. Treatment of the cancer by killing, inactivating, or depleting the non-stimulatory bone marrow cells. The method of [Invention 1044].[[]] [Invention 1048] The method of [Invention 1044], wherein the antibody has antibody-dependent cell-mediated cytotoxicity (ADCC) activity. [Invention 1049] The method of [Invention 1044], wherein the antibody has complement-dependent cytotoxicity (CDC) activity. [Invention 1050] The method of [Invention 1044], wherein the antibody has antibody-mediated phagocytosis (ADCP) activity. [Invention 1051] The method of [Invention 1044], wherein the antibody has receptor-ligand blocking, agonism, or antagonism activity. [Invention 1052] The method according to any one of [Invention 1044] to [Invention 1051], wherein the subject is human. [Invention 1053] The method according to any one of [Invention 1044] to [Invention 1052], wherein the cancer is a solid cancer. [Invention 1054] The method according to any one of [Invention 1044] to [Invention 1052], wherein the cancer is a liquid cancer. [Invention 1055] The method according to any one of [Invention 1044] to [Invention 1052], wherein the cancer is selected from the group consisting of melanoma, kidney, hepatobiliary tract, head and neck squamous cell carcinoma (HNSC), pancreas, colon, bladder, glioblastoma, prostate, lung, breast, ovary, stomach, kidney, bladder, esophagus, kidney, melanoma, and mesothelioma. [Invention 1056] The method of [Invention 1055], wherein the cancer is colon cancer or breast cancer. [Invention 1057] The method according to any one of [Invention 1044] to [Invention 1056], wherein the contact enhances the immune response of the subject. [Invention 1058] The method according to [Invention 1057], wherein the enhanced immune response is an adaptive immune response. [Invention 1059] The method according to [Invention 1057], wherein the enhanced immune response is a innate immune response. [Invention 1060] The method according to any one of [Invention 1044] to [Invention 1059], wherein the subject has received, is receiving, or will receive immunotherapy. [Invention 1061] The method according to [Invention 1060], wherein the immunotherapy is at least one of a checkpoint inhibitor; a T cell checkpoint inhibitor; an anti-PD1 antibody; an anti-PDL1 antibody; an anti-CTLA4 antibody; adoptive T cell therapy; CAR-T cell therapy; dendritic cell vaccine; monocyte vaccine; an antigen-binding protein that binds to both T cells and antigen-presenting cells; a BiTE bispecific antigen-binding protein; a toll-like receptor ligand; a cytokine; a cytotoxic therapy; chemotherapy; radiotherapy; a small molecule inhibitor; a small molecule agonist; an immunomodulatory agent; and an epigenetic regulator. [Invention 1062] The method according to [Invention 1061], wherein the immunotherapy is an anti-PD1 antibody. [Invention 1063] A method of killing, inactivating, or depleting TREM2+ myeloid cells in a subject having cancer, comprising contacting the myeloid cells with an antibody according to any one of [Invention 1001] to [Invention 1038] or a pharmaceutical composition according to [Invention 1043], optionally, wherein the myeloid cells are within the tumor, the method. [Invention 1064] The antibody binds to the extracellular domain of TREM2, and the myeloid cells are non-stimulatory myeloid cells that are CD45+, HLA-DR+, CD11c+, CD14+, and BDCA3-, The antibody kills, inactivates, or depletes the non-stimulatory myeloid cells via ADCC, CDC, and / or ADCP to a level lower than the level of non-stimulatory myeloid cells present in the cancer before contacting the non-stimulatory myeloid cells with the antibody. The non-stimulatory myeloid cells are present in a population of immune cells comprising the stimulatory myeloid cells that are CD45+, HLA-DR+, CD14-, CD11c+, BDCA1-, and BDCA3+ and the non-stimulatory myeloid cells. The contacting does not substantially kill, inactivate, or deplete the myeloid cells present outside the cancer and / or the stimulatory myeloid cells present in the cancer. The killing, inactivation, or depletion of the non-stimulatory myeloid cells treats the cancer by enhancing the immune response to the cancer. The method of [Invention 1063]. [Invention 1065] The method of [Invention 1063], wherein the antibody kills the myeloid cells by at least one of ADCC, CDC, and ADCP. [Invention 1066] The method of [Invention 1063], wherein the antibody inactivates the myeloid cells by at least one of ADCC, CDC, and ADCP. [Invention 1067] The method of [Invention 1063], wherein the antibody depletes the myeloid cells by at least one of ADCC, CDC, and ADCP. [Invention 1068] The method of [Invention 1063], wherein the antibody has antibody-dependent cell-mediated cytotoxicity (ADCC) activity. [Invention 1069] The method of [Invention 1063], wherein the antibody has complement-dependent cytotoxicity (CDC) activity. [Invention 1070] The method of [Invention 1063], wherein the antibody has antibody-mediated phagocytosis (ADCP) activity. [Invention 1071] The method of [Invention 1063], wherein the antibody has receptor-ligand blocking, agonism, or antagonism activity. [The present invention 1072] Any of the methods of [the present invention 1063] to [the present invention 1071], wherein the bone marrow cells are stimulatory bone marrow cells. [The present invention 1073] Any of the methods of [the present invention 1063] to [the present invention 1071], wherein the bone marrow cells are non-stimulatory bone marrow cells. [The present invention 1074] Any of the methods of [the present invention 1063] to [the present invention 1073], wherein the bone marrow cells comprise at least one of dendritic cells, tumor-associated macrophages (TAM), neutrophils, or monocytes. [The present invention 1075] The method of [the present invention 1074], wherein the bone marrow cells are neutrophils. [The present invention 1076] The method of [the present invention 1074], wherein the bone marrow cells are tumor-associated macrophages. [The present invention 1077] Any of the methods of [the present invention 1063] to [the present invention 1076], wherein the bone marrow cells are within the tumor. [The present invention 1078] Any of the methods of [the present invention 1063] to [the present invention 1077], wherein the bone marrow cells are a population of immune cells comprising stimulatory bone marrow cells and non-stimulatory bone marrow cells. [The present invention 1079] Any of the methods of [the present invention 1063] to [the present invention 1078], wherein the contact is in vitro or in vivo. [The present invention 1080] Any of the methods of [the present invention 1063] to [the present invention 1079], wherein the contact occurs in vivo in a subject in need thereof, and optionally, the subject has cancer. [The present invention 1081] Any of the methods of [the present invention 1031] to [the present invention 1080], wherein the subject is human. [The present invention 1082] Any of the methods of [the present invention 1063] to [the present invention 1081], wherein the cancer is a solid cancer. [The present invention 1083] The method according to any one of [Invention 1063] to [Invention 1081], wherein the cancer is a liquid cancer. [Invention 1084] The method according to [Invention 1080], wherein the cancer is selected from the group consisting of melanoma, kidney, hepatobiliary tract, head and neck squamous cell carcinoma (HNSC), pancreas, colon, bladder, glioblastoma, prostate, lung, breast, ovary, stomach, kidney, bladder, esophagus, kidney, melanoma, and mesothelioma. [Invention 1085] The method according to [Invention 1084], wherein the cancer is colon cancer or breast cancer. [Invention 1086] The method according to [Invention 1080], wherein the contact enhances the immune response of the subject. [Invention 1087] The method according to [Invention 1080], wherein the enhanced immune response is an adaptive immune response. [Invention 1088] The method according to [Invention 1080], wherein the enhanced immune response is a natural immune response. [Invention 1089] The method according to any one of [Invention 1080] to [Invention 1088], wherein the subject has previously received, is currently receiving, or will receive immunotherapy thereafter. [Invention 1090] The method according to [Invention 1089], wherein the immunotherapy is at least one of a checkpoint inhibitor; a T cell checkpoint inhibitor; an anti-PD1 antibody; an anti-PDL1 antibody; an anti-CTLA4 antibody; adoptive T cell therapy; CAR-T cell therapy; dendritic cell vaccine; monocyte vaccine; an antigen-binding protein that binds to both T cells and antigen-presenting cells; a BiTE bispecific antigen-binding protein; a toll-like receptor ligand; a cytokine; a cytotoxic therapy; chemotherapy; radiation therapy; a small molecule inhibitor; a small molecule agonist; an immunomodulatory agent; and an epigenetic regulator. [Invention 1091] The method according to [Invention 1090], wherein the immunotherapy is an anti-PD1 antibody. [Invention 1092] A method for detecting TREM2 in a subject having or suspected of having a disease or condition, comprising: (a) receiving a sample from the subject; and (b) detecting the presence or level of TREM2 in the sample by contacting the sample with an antibody selected from any of [Inventive Item 1001] to [Inventive Item 1038]. The method as described above. [Inventive Item 1093] The method according to [Inventive Item 1092], wherein the disease or condition is cancer. [Inventive Item 1094] Further comprising the step of administering a checkpoint inhibitor, Optionally, the checkpoint inhibitor is an inhibitor of the PD1:PDL1 axis, optionally the inhibitor is an antibody, and optionally the antibody is an anti-PD1 antibody or an anti-PDL1 antibody. The method according to [Inventive Item 1092] or [Inventive Item 1093]. [Inventive Item 1095] A kit comprising an antibody selected from any of [Inventive Item 1001] to [Inventive Item 1038] or a pharmaceutical composition of [Inventive Item 1043] and instructions for use.

Brief Description of the Drawings

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Mode for Carrying Out the Invention

[0067] Detailed Description Definitions For the purposes of interpreting this specification, the following definitions shall apply, and whenever appropriate, terms used in the singular shall include the plural and vice versa. If any of the following definitions conflict with any document incorporated herein by reference, the defined description shall prevail.

[0068] As used herein, aspects and embodiments of the invention described herein are understood to include "comprising," "consisting of," and "consisting essentially of."

[0069] In all of the compositions described herein and all of the methods of using the compositions described herein, the composition may include any of the recited components or steps, or may "consist essentially of" the recited components or steps. When a composition is described as "consisting essentially of" a recited component, the composition contains the recited component and may contain other components that do not substantially affect the state being treated other than those explicitly recited components, but does not contain any other components that substantially affect the state being treated, or, if the composition contains additional components other than the recited components that substantially affect the state being treated, the composition does not contain the additional components in a concentration or amount sufficient to substantially affect the state being treated. When a method is described as "consisting essentially of" the recited steps, the method contains the recited steps and may contain other steps that do not substantially affect the state being treated, but does not contain any other steps that substantially affect the state being treated other than those explicitly recited steps. By way of specific non-limiting example, when a composition is described as "consisting essentially of" a component, the composition may further contain any amount of a pharmaceutically acceptable carrier, vehicle, or diluent, and such other components that do not substantially affect the state being treated.

[0070] The term "optionally", when used sequentially, means including any and all of the recited combinations, considering all sub-combinations.

[0071] As used herein, "effective amount" or "therapeutically effective amount" refers to the amount of a therapeutic compound, such as an anti-TREM2 antigen-binding agent or anti-TREM2 antibody, administered to an individual, either as a single administration or as part of a series of administrations, which is effective, either alone or in combination with another therapy, to produce the desired therapeutic effect or to cause it. Examples of desired therapeutic effects are enhancement of the immune response; deceleration or delay of tumor development; stabilization of the disease; improvement of one or more symptoms. The effective amount may be administered in one or more dosages.

[0072] As used herein, the term "treating" refers to delaying or reversing the progression of a condition such as cancer. As used herein, the term "treatment" refers to the act of treating a condition such as cancer.

[0073] As used herein, "individual" or "subject" refers to any animal classified as a mammal, including humans, domesticated animals and livestock, and pets such as animals in zoos, sports, or dogs, horses, rabbits, cows, pigs, hamsters, gerbils, mice, ferrets, rats, cats, etc. In some embodiments, the individual is a human. In some embodiments, the individual is a mouse.

[0074] The terms "modulate" and "modulation" refer to reducing or inhibiting, or alternatively activating or increasing, the recited variable element.

[0075] The terms "increase" and "activate" refer to an increase of 10%, 20%, 30%, 40%, 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 2-fold, 3-fold, 4-fold, 5-fold, 10-fold, 20-fold, 50-fold, 100-fold, or more of the recited variable element.

[0076] The terms "reduce" and "inhibit" refer to a decrease of 10%, 20%, 30%, 40%, 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 2-fold, 3-fold, 4-fold, 5-fold, 10-fold, 20-fold, 50-fold, 100-fold, or more of the recited variable element.

[0077] The term "stimulate" refers to the activation of receptor signaling to induce a biological response associated with the activation of a receptor. An "agonist" is an entity that binds to a receptor and stimulates it.

[0078] The term "antagonize" refers to the inhibition of receptor signaling to inhibit a biological response associated with receptor activation. An "antagonist" is an entity that binds to a receptor and antagonizes it.

[0079] As used herein, the term "about" refers to the normal error range of each value that would be immediately apparent to those skilled in the art. An exemplary error range is plus or minus 5%. References to "about" values or parameters herein include (and describe) embodiments directed to the value or parameter itself.

[0080] It should be noted that, as used in this specification and the appended claims, the singular forms "a", "an", and "the" include plural referents unless the context clearly dictates otherwise.

[0081] For any of the structural and functional characteristics described herein, methods for determining these characteristics are known in the art.

[0082] antibody structure This application provides antibodies and compositions comprising antibodies that bind to the TREM2 protein, including antibodies that inactivate non-stimulatory bone marrow cells.

[0083] The term "antibody" is used herein in the broadest sense and includes certain types of immunoglobulin molecules that contain one or more antigen-binding domains that specifically bind to an antigen or epitope. Antibodies specifically include intact antibodies (e.g., intact immunoglobulins), antibody fragments, and multispecific antibodies.

[0084] The recognized immunoglobulin genes include kappa, lambda, alpha, gamma, delta, epsilon, and mu constant region genes, as well as numerous immunoglobulin variable region genes. The light chains are classified into either kappa or lambda. The "class" of an antibody or immunoglobulin refers to the type of constant domain or constant region that the heavy chain has. The five main classes of antibodies: IgA, IgD, IgE, IgG, and IgM, and some of these can be further divided into subclasses (isotypes), for example, IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2. The heavy chain constant domains corresponding to the different classes of immunoglobulins are called alpha, delta, epsilon, gamma, and mu, respectively.

[0085] An exemplary immunoglobulin (antibody) structural unit consists of two pairs of polypeptide chains, each pair having one "light" chain (about 25 kD) and one "heavy" chain (about 50 - 70 kD). The N-terminal domain of each chain defines a variable region of about 100 - 110 or more amino acids that is mainly involved in antigen recognition. The terms variable light chain (VL) and variable heavy chain (VH) refer to these light and heavy chain domains, respectively. The IgG1 heavy chain consists of VH, CH1, CH2, and CH3 domains from the N-terminus to the C-terminus. The light chain consists of VL and CL domains from the N-terminus to the C-terminus. The IgG1 heavy chain contains a hinge between the CH1 domain and the CH2 domain. In certain embodiments, the immunoglobulin construct includes at least one immunoglobulin domain derived from IgG, IgM, IgA, IgD, or IgE that is linked to a therapeutic polypeptide. In some embodiments, the immunoglobulin domains found in the antibodies provided herein are derived from or induced in immunoglobulin-based constructs such as diabodies or nanobodies. In certain embodiments, the immunoglobulin constructs described herein include at least one immunoglobulin domain derived from heavy chain antibodies such as camelid antibodies. In certain embodiments, the immunoglobulin constructs provided herein include at least one immunoglobulin domain derived from mammalian antibodies such as bovine antibodies, human antibodies, camel antibodies, mouse antibodies, or any chimeric antibody.

[0086] In some embodiments, the antibodies provided herein include heavy chains. In one embodiment, the heavy chain is IgA. In one embodiment, the heavy chain is IgD. In one embodiment, the heavy chain is IgE. In one embodiment, the heavy chain is IgG. In one embodiment, the heavy chain is IgM. In one embodiment, the heavy chain is IgG1. In one embodiment, the heavy chain is IgG2. In one embodiment, the heavy chain is IgG3. In one embodiment, the heavy chain is IgG4. In one embodiment, the heavy chain is IgA1. In one embodiment, the heavy chain is IgA2.

[0087] As used herein, the terms "hypervariable region" or "HVR" refer to each of the regions of an antibody variable domain where the sequences are hypervariable and / or form structurally defined loops ("hypervariable loops"). Generally, native four-chain antibodies contain six HVRs: three of VH (H1, H2, H3) and three of VL (L1, L2, L3). HVRs generally contain amino acid residues from hypervariable loops and / or complementarity determining regions (CDRs), the latter being those with the highest sequence variability and / or involved in antigen recognition. Except for CDR1 of VH, CDRs generally contain amino acid residues that form hypervariable loops. The hypervariable regions (HVRs) are also referred to as "complementary determining regions" (CDRs), and these terms are used interchangeably herein with respect to the portions of the variable region that form the antigen-binding region. This particular region has been described by Kabat et al., U.S. Dept. of Health and Human Services, Sequences of Proteins of Immunological Interest (1983) and Chothia et al., J Mol Biol 196:901-917 (1987), and the definitions include amino acid residue overlaps or subsets when compared to each other. Nevertheless, the application of any definition referring to CDRs of an antibody or its variants is intended to be within the scope of the terms defined and used herein. The exact residue numbers encompassing a particular CDR will vary depending on the sequence and size of the CDR. Those skilled in the art can determine, as per the specification, which residues are included in a particular CDR given the amino acid sequence of the variable region of the antibody.

[0088] The boundaries of the amino acid sequences of the CDRs can be determined by one of ordinary skill in the art using any of the known numbering schemes, including those described in Kabat et al., supra (the "Kabat" numbering scheme); Al-Lazikani et al., 1997, J. Mol. Biol., 273:927-948 (the "Chothia" numbering scheme); MacCallum et al., 1996, J. Mol. Biol. 262:732-745 (the "Contact" numbering scheme); Lefranc et al., Dev. Comp. Immunol., 2003, 27:55-77 (the "IMGT" numbering scheme); and Honegge and Pluckthun, J. Mol. Biol., 2001, 309:657-70 (the "AHo" numbering scheme), each of which is incorporated by reference in its entirety.

[0089] Table A provides the positions of CDR-L1, CDR-L2, CDR-L3, CDR-H1, CDR-H2, and CDR-H3 identified by the Kabat and Chothia schemes. For CDR-H1, residue numbering is provided using both the Kabat and Chothia numbering schemes.

[0090] The CDRs may be assigned, for example, using antibody numbering software such as Abnum, described in Abhinandan and Martin, Immunology, 2008, 45:3832-3839, which is available at www.bioinf.org.uk / abs / abnum / and is incorporated by reference in its entirety.

[0091] (Table A) Residues of CDRs according to the Kabat and Chothia numbering schemes TIFF2025109867000002.tif42150 * The C-terminus of CDR-H1 varies between H32 and H34 depending on the length of the CDR when numbered using the Kabat numbering rules.

[0092] The "EU numbering scheme" is generally used when referring to residues within the constant region of an antibody heavy chain (e.g., as reported in Kabat et al. supra). Unless otherwise specified, the EU numbering scheme is used to refer to the residues of the antibody heavy chain constant region described herein.

[0093] As used herein, the term "single-chain" refers to a molecule comprising amino acid monomers linearly linked by peptide bonds. In such a particular embodiment, the C-terminus of the Fab light chain is linked to the N-terminus of the Fab heavy chain of the single-chain Fab molecule. As described in more detail herein, an scFv has a variable domain of the light chain (VL) linked by a polypeptide chain from the C-terminus to the N-terminus of the variable domain of the heavy chain (VH). Alternatively, an scFv is composed of a polypeptide chain in which the C-terminus of VH is linked to the N-terminus of VL by a polypeptide chain.

[0094] A "Fab fragment" (also called fragment antigen-binding) contains the constant domain of the light chain (CL) and the first constant domain of the heavy chain (CH1) along with the variable domains VL and VH on the light and heavy chains, respectively. The variable domains contain complementarity-determining loops (CDRs, also called hypervariable regions) involved in antigen binding. A Fab' fragment differs from a Fab fragment by the addition of several residues to the carboxy terminus of the heavy chain CH1 domain that includes one or more cysteines from the antibody hinge region.

[0095] An "F(ab')2" fragment contains two Fab' fragments linked near the hinge region by disulfide bonds. An F(ab')2 fragment may be produced, for example, by recombinant methods or by pepsin digestion of an intact antibody. An F(ab') fragment can be dissociated, for example, by treatment with β-mercaptoethanol.

[0096] An "Fv" fragment contains a dimer of one heavy chain variable domain and one light chain variable domain non-covalently linked.

[0097] "Single-chain Fv" or "scFv" contains the VH and VL domains of an antibody, and these domains are present in a single polypeptide chain. In one embodiment, the Fv polypeptide further contains a polypeptide linker between the VH and VL domains that enables the scFv to form the desired structure for antigen binding. For a review of scFv, see Pluckthun in The Pharmacology of Monoclonal Antibodies, vol. 113, Rosenberg and Moore eds., Springer-Verlag, New York, pp. 269-315 (1994). HER2 antibody scFv fragments are described in WO93 / 16185; U.S. Patent No. 5,571,894; and U.S. Patent No. 5,587,458.

[0098] "scFv-Fc" fragment contains an scFv bound to an Fc domain. For example, the Fc domain may be bound to the C-terminus of the scFv. The Fc domain may follow V H -V L or V L -V H depending on the orientation of the variable domains in (i.e., V H or V L ). Any suitable Fc domain known in the art or described herein may be used. In some cases, the Fc domain contains an IgG4 Fc domain.

[0099] The term "single-domain antibody" or "sdAb" refers to a molecule in which one variable domain of an antibody binds specifically to an antigen in the absence of other variable domains. Single-domain antibodies and fragments thereof are described in Arabi Ghahroudi et al., FEBS Letters, 1998, 414:521-526 and Muyldermans et al., Trends in Biochem. Sci., 2001, 26:230-245, each of which is incorporated herein by reference in its entirety. Single-domain antibodies are also known as sdAbs or nanobodies. SdAbs are quite stable and are easy to express as fusion partners with the Fc chain of an antibody (Harmsen MM, De Haard HJ (2007). “Properties, production, and applications of camelid single-domain antibody fragments”. Appl. Microbiol Biotechnol. 77(1):13-22).

[0100] The terms "full-length antibody", "intact antibody", and "whole antibody" are used interchangeably herein to refer to an antibody having a structure substantially similar to a naturally occurring antibody structure and having a heavy chain that includes an Fc region. For example, when used to refer to an IgG molecule, a "full-length antibody" is an antibody that includes two heavy chains and two light chains.

[0101] The term "epitope" means a part of an antigen that binds specifically to an antibody. Epitopes often consist of surface-accessible amino acid residues and / or sugar side chains and may have specific three-dimensional structural characteristics and specific charge characteristics. Conformational and non-conformational epitopes are distinguished in that binding to the former, but not the latter, can be lost in the presence of a denaturing solvent. An epitope may include amino acid residues that are directly involved in binding and other amino acid residues that are not directly involved in binding. The epitope to which an antibody binds can be determined using known techniques for epitope determination, such as testing antibody binding to TREM2 variants or chimeric TREM2 variants having different point mutations.

[0102] A "multispecific antibody" is an antibody that includes two or more different antigen-binding domains that specifically bind collectively to two or more different epitopes. The two or more different epitopes can be epitopes on the same antigen (e.g., a single TREM2 molecule expressed by a cell) or different antigens (e.g., different TREM2 molecules expressed by the same cell, or a TREM2 molecule and a non-TREM2 molecule). In some embodiments, the multispecific antibody binds to two different epitopes (i.e., a "bispecific antibody"). In some embodiments, the multispecific antibody binds to three different epitopes (i.e., a "trispecific antibody").

[0103] A "monospecific antibody" is an antibody that includes one or more binding sites that specifically bind to a single epitope. Examples of monospecific antibodies are naturally occurring IgG molecules that are bivalent (i.e., have two antigen-binding domains) but recognize the same epitope with each of the two antigen-binding domains. The binding specificity can be present at any suitable valence.

[0104] The term "monoclonal antibody" refers to an antibody derived from a substantially homogeneous population of antibodies. A substantially homogeneous population of antibodies includes antibodies that are substantially similar and bind to the same epitope(s), excluding variants that can typically occur during the production of monoclonal antibodies. Such variants generally exist only in small amounts. Monoclonal antibodies are usually obtained by a process that includes selecting a single antibody from a plurality of antibodies. For example, the selection process can be the selection of a unique clone from a pool of multiple clones, such as hybridoma clones, phage clones, yeast clones, bacterial clones, or other recombinant DNA clones. The selected antibody can further be modified, for example, to improve the affinity for the target ("affinity maturation"), to humanize the antibody, to improve production in cell culture, and / or to reduce immunogenicity in a subject.

[0105] "Effector function" refers to the biological activity mediated by the Fc region of an antibody, which can vary depending on the antibody isotype. Examples of antibody effector functions include C1q binding that activates complement-dependent cytotoxicity (CDC), Fc receptor binding that activates antibody-dependent cell-mediated cytotoxicity (ADCC), and antibody-dependent cell phagocytosis (ADCP), receptor ligand blockade, agonism, or antagonism.

[0106] An anti-TREM2 antibody can include those described herein, such as the clones listed in the table. In some embodiments, the antibody includes an alternative scaffold. In some embodiments, the antibody consists of an alternative scaffold. In some embodiments, the antibody consists essentially of an alternative scaffold. In some embodiments, the antibody includes an antibody fragment. In some embodiments, the antibody consists of an antibody fragment. In some embodiments, the antibody consists essentially of an antibody fragment. A "TREM2 antibody", "anti-TREM2 antibody", or "TREM2-specific antibody" is an antibody as provided herein that specifically binds to the antigen TREM2. In some embodiments, the antibody binds to the extracellular domain of TREM2. In certain embodiments, the TREM2 antibodies provided herein bind to an epitope of TREM2 that is conserved between or among TREM2 proteins from different species.

[0107] The term "chimeric antibody" refers to an antibody in which a portion of the heavy and / or light chain is derived from a particular source or species, while the remainder of the heavy and / or light chain is derived from a different source or species.

[0108] The "humanized" form of a non-human antibody is a chimeric antibody containing a minimal sequence derived from a non-human antibody. A humanized antibody is generally a human antibody (recipient antibody) in which one or more residues derived from one or more CDRs are replaced by residues derived from one or more CDRs of a non-human antibody (donor antibody). The donor antibody can be any suitable non-human antibody, e.g., a mouse, rat, rabbit, chicken, or non-human primate antibody having the desired specificity, affinity, or biological effect. A humanized antibody, when administered to a human subject, is less likely to induce an immune response and / or induces a less severe immune response as compared to a non-human species antibody. In some cases, selected framework region residues of the recipient antibody are replaced by the corresponding framework region residues from the donor antibody. A humanized antibody may also contain residues not found in either the recipient antibody or the donor antibody. Such modifications may be made to further purify antibody function. Examples of methods for making humanized antibodies can be found in U.S. Patent Nos. 6,054,297, 5,886,152, and 5,877,293, each of which is incorporated herein by reference in its entirety. For further details, see Jones et al., Nature, 1986, 321:522-525; Riechmann et al., Nature, 1988, 332:323-329; and Presta, Curr. Op. Struct. Biol., 1992, 2:593-596 (each of which is incorporated herein by reference in its entirety).

[0109] In one embodiment, a constant domain(s) derived from a human antibody is fused to a variable domain(s) of a non-human species. In another embodiment, one or more amino acid residues in one or more CDR sequences of a non-human antibody are changed such that when it is administered to a human subject, it reduces the likelihood of immunogenicity of the non-human antibody, and none of the changed amino acid residues are critical for the immunospecific binding of the antibody to the antigen, or the changes made to the amino acid sequence are conservative changes, such that the binding of the humanized antibody to the antigen is significantly lower than the binding of the non-human antibody to the antigen.

[0110] A "human antibody" is one produced by a human or human cell, or having an amino acid sequence corresponding to the amino acid sequence of an antibody derived from a non-human source that utilizes a human antibody repertoire or human antibody-encoding sequence (e.g., obtained from a human source or designed de novo). Human antibodies specifically exclude humanized antibodies. In one embodiment, all of the variable and constant domains are derived from human immunoglobulin sequences (fully human antibodies). These antibodies may be prepared by various methods including immunization of a mouse genetically modified to express antibodies derived from genes encoding human heavy and / or light chains with the antigen of interest.

[0111] In some embodiments, the antibodies provided herein include antibody fragments. In some embodiments, the antibodies provided herein consist of antibody fragments. In some embodiments, the antibodies provided herein consist essentially of antibody fragments. In some embodiments, the antibody fragment is an Fv fragment. In some embodiments, the antibody fragment is a Fab fragment. In some embodiments, the antibody fragment is an F(ab’)2 fragment. In some embodiments, the antibody fragment is a Fab’ fragment. In some embodiments, the antibody fragment is a scFv (sFv) fragment. In some embodiments, the antibody fragment is a scFv-Fc fragment. In some embodiments, the antibody fragment is a fragment of a single domain antibody.

[0112] Sequence of the TREM2 antibody V H domain In some embodiments, the antibodies provided herein include a V H sequence selected from SEQ ID NOs: 1, 3, 5, and 7. In some embodiments, the antibodies provided herein include the V H sequence of SEQ ID NO: 1. In some embodiments, the antibodies provided herein include the V H sequence of SEQ ID NO: 3. In some embodiments, the antibodies provided herein include the VH It includes a sequence. In some embodiments, the antibodies provided herein have a V of SEQ ID NO: 7 H sequence.

[0113] In some embodiments, the antibodies provided herein have a V exemplified by SEQ ID NOs: 1, 3, 5, and 7 H sequence with at least about 50%, 60%, 70%, 80%, 90%, 95%, or 99% identity to the V H sequence. In some embodiments, the antibodies provided herein have a V provided by SEQ ID NOs: 1, 3, 5, and 7 with up to 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, or 25 amino acid substitutions H sequence. In some aspects, the amino acid substitutions are conservative amino acid substitutions. In some embodiments, the antibodies described in this paragraph are referred to herein as "variants". In some embodiments, such variants are derived from the sequences provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis, or any other method known in the art or described herein. In some embodiments, such variants are not derived from the sequences provided herein and may be isolated de novo, for example, according to the methods provided herein for obtaining antibodies.

[0114] V L domain In some embodiments, the antibodies provided herein have a V selected from SEQ ID NOs: 2, 4, 6, and 8 L sequence. In some embodiments, the antibodies provided herein have a V of SEQ ID NO: 2 L sequence. In some embodiments, the antibodies provided herein have a V of SEQ ID NO: 4 L sequence. In some embodiments, the antibodies provided herein have a V of SEQ ID NO: 6 L sequence. In some embodiments, the antibodies provided herein have a V of SEQ ID NO: 8 L sequence.

[0115] In some embodiments, the antibodies provided herein have a V sequence provided in SEQ ID NOs: 2, 4, 6, and 8 with at least about 50%, 60%, 70%, 80%, 90%, 95%, or 99% identity. L In some embodiments, the antibodies provided herein include a V sequence having up to 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, or 25 amino acid substitutions in the V sequences provided in SEQ ID NOs: 2, 4, 6, and 8. L In some aspects, the amino acid substitutions are conservative amino acid substitutions. In some embodiments, the antibodies described in this paragraph are referred to herein as "variants." In some embodiments, such variants are derived from the sequences provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis, or any other method known in the art or described herein. In some embodiments, such variants are not derived from the sequences provided herein and may be isolated de novo, for example, according to the methods provided herein for obtaining antibodies. L In some embodiments, the antibodies provided herein include a combination of V-V.

[0116] V H -V L combination In some embodiments, the antibodies provided herein include a V sequence selected from SEQ ID NOs: 1, 3, 5, and 7, and a V sequence selected from SEQ ID NOs: 2, 4, 6, and 8. H In some embodiments, the antibodies provided herein include the V sequence of SEQ ID NO: 1 and the V sequence of SEQ ID NO: 2. In some embodiments, the antibodies provided herein include the V sequence of SEQ ID NO: 3 and the V sequence of SEQ ID NO: 4. In some embodiments, the antibodies provided herein include the V sequence of SEQ ID NO: 5 and the V sequence of SEQ ID NO: 6. L In some embodiments, the antibodies provided herein include the V sequence of SEQ ID NO: 5 and the V sequence of SEQ ID NO: 6.

[0117] In some embodiments, the antibodies provided herein include the V sequence of SEQ ID NO: 1 and the V sequence of SEQ ID NO: 2. H In some embodiments, the antibodies provided herein include the V sequence of SEQ ID NO: 3 and the V sequence of SEQ ID NO: 4. L In some embodiments, the antibodies provided herein include the V sequence of SEQ ID NO: 5 and the V sequence of SEQ ID NO: 6. H In some embodiments, the antibodies provided herein include the V sequence of SEQ ID NO: 3 and the V sequence of SEQ ID NO: 4. L In some embodiments, the antibodies provided herein include the V sequence of SEQ ID NO: 5 and the V sequence of SEQ ID NO: 6. H In some embodiments, the antibodies provided herein include the V sequence of SEQ ID NO: 5 and the V sequence of SEQ ID NO: 6. LIt includes sequences. In some embodiments, the antibodies provided herein have a V of SEQ ID NO: 7 H sequence and a V of SEQ ID NO: 8 L sequence. In certain aspects, any one of SEQ ID NOs: 1, 3, 5, and 7 can be combined with any one of SEQ ID NOs: 2, 4, 6, and 8. For example, SEQ ID NO: 1 can be combined with any one of SEQ ID NOs: 2, 4, 6, or 8. As another example, SEQ ID NO: 2 can be combined with any one of SEQ ID NOs: 1, 3, 5, or 7.

[0118] In some embodiments, the antibodies provided herein have a V of SEQ ID NOs: 1, 3, 5, and 7 H sequences having at least about 50%, 60%, 70%, 80%, 90%, 95%, or 99% identity to the exemplified V H sequences, and a V of SEQ ID NOs: 2, 4, 6, and 8 having at least about 50%, 60%, 70%, 80%, 90%, 95%, or 99% identity to the exemplified VL sequences L sequences. In some embodiments, the antibodies provided herein include a VH sequence provided by SEQ ID NOs: 1, 3, 5, and 7 having a maximum of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, or 25 amino acid substitutions, and a VL sequence provided by SEQ ID NOs: 2, 4, 6, and 8 having a maximum of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, or 25 amino acid substitutions. In some aspects, the amino acid substitutions are conservative amino acid substitutions. In some embodiments, the antibodies described in this paragraph are referred to herein as "variants". In some embodiments, such variants are derived from the sequences provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis, or any other method known in the art or described herein. In some embodiments, such variants are not derived from the sequences provided herein and may be isolated de novo, for example, according to the methods provided herein for obtaining antibodies.

[0119] CDR In some embodiments, the antibodies provided herein comprise 1 to 3 CDRs of the V H domains selected from SEQ ID NOs: 1, 3, 5, and 7. In some embodiments, the antibodies provided herein comprise 2 to 3 CDRs of the V H domains selected from SEQ ID NOs: 1, 3, 5, and 7. In some embodiments, the antibodies provided herein comprise 3 CDRs of the VH domain selected from SEQ ID NOs: 1, 3, 5, and 7. In some aspects, the CDRs are exemplary CDRs. In some aspects, the CDRs are Kabat CDRs. In some aspects, the CDRs are Chothia CDRs. In some aspects, the CDRs are AbM CDRs. In some aspects, the CDRs are Contact CDRs. In some aspects, the CDRs are IMGT CDRs.

[0120] In some embodiments, the CDR is a CDR having at least about 50%, 75%, 80%, 85%, 90%, or 95% identity with the CDR-H1, CDR-H2, or CDR-H3 of SEQ ID NOs: 1, 3, 5, and 7. In some embodiments, CDR-H1 is the CDR-H1 of the VH domain selected from SEQ ID NOs: 1, 3, 5, and 7 having a maximum of 1, 2, 3, 4, or 5 amino acid substitutions. In some embodiments, CDR-H2 is the CDR-H2 of the VH domain selected from SEQ ID NOs: 1, 3, 5, and 7 having a maximum of 1, 2, 3, 4, 5, 6, 7, or 8 amino acid substitutions. In some embodiments, CDR-H3 is the CDR-H3 of the VH domain selected from SEQ ID NOs: 1, 3, 5, and 7 having a maximum of 1, 2, 3, 4, 5, 6, 7, or 8 amino acid substitutions. In some aspects, the amino acid substitutions are conservative amino acid substitutions. In some embodiments, the antibodies described in this paragraph are referred to herein as "variants". In some embodiments, such variants are derived from the sequences provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis, or any other method known in the art or described herein. In some embodiments, such variants are not derived from the sequences provided herein and may be isolated de novo, for example, according to the methods provided herein for obtaining antibodies.

[0121] In some embodiments, the antibodies provided herein comprise 1 to 3 CDRs of the VL domain selected from SEQ ID NOs: 2, 4, 6, and 8. In some embodiments, the antibodies provided herein comprise 2 to 3 CDRs of the VL domain selected from SEQ ID NOs: 2, 4, 6, and 8. In some embodiments, the antibodies provided herein comprise 3 CDRs of the VL domain selected from SEQ ID NOs: 2, 4, 6, and 8. In some aspects, the CDR is an exemplary CDR. In some aspects, the CDR is a Kabat CDR. In some aspects, the CDR is a Chothia CDR. In some aspects, the CDR is an AbM CDR. In some aspects, the CDR is a Contact CDR. In some aspects, the CDR is an IMGT CDR.

[0122] In some embodiments, the CDR is a CDR having at least about 50%, 75%, 80%, 85%, 90%, or 95% identity with the CDR-L1, CDR-L2, or CDR-L3 of SEQ ID NOs: 2, 4, 6, and 8. In some embodiments, CDR-L1 is the CDR-L1 of the VL domain selected from SEQ ID NOs: 2, 4, 6, and 8 having a maximum of 1, 2, 3, 4, or 5 amino acid substitutions. In some embodiments, CDR-L2 is the CDR-L2 of the VL domain selected from SEQ ID NOs: 2, 4, 6, and 8 having a maximum of 1, 2, 3, 4, 5, 6, 7, or 8 amino acid substitutions. In some embodiments, CDR-L3 is the CDR-L3 of the VL domain selected from SEQ ID NOs: 2, 4, 6, and 8 having a maximum of 1, 2, 3, 4, 5, 6, 7, or 8 amino acid substitutions. In some aspects, the amino acid substitutions are conservative amino acid substitutions. In some embodiments, the antibodies described in this paragraph are referred to herein as "variants". In some embodiments, such variants are derived from the sequences provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis, or any other method known in the art or described herein. In some embodiments, such variants are not derived from the sequences provided herein and may be isolated de novo, for example, according to the methods provided herein for obtaining antibodies.

[0123] In some embodiments, the antibodies provided herein comprise one to three CDRs of the VH domain selected from SEQ ID NOs: 1, 3, 5, and 7, and one to three CDRs of the VL domain selected from SEQ ID NOs: 2, 4, 6, and 8. In some embodiments, the antibodies provided herein comprise two to three CDRs of the VH domain selected from SEQ ID NOs: 1, 3, 5, and 7, and two to three CDRs of the VL domain selected from SEQ ID NOs: 2, 4, 6, and 8. In some embodiments, the antibodies provided herein comprise three CDRs of the VH domain selected from SEQ ID NOs: 1, 3, 5, and 7, and three CDRs of the VL domain selected from SEQ ID NOs: 2, 4, 6, and 8. In some aspects, the CDRs are exemplary CDRs. In some aspects, the CDRs are Kabat CDRs. In some aspects, the CDRs are Chothia CDRs. In some aspects, the CDRs are AbM CDRs. In some aspects, the CDRs are Contact CDRs. In some aspects, the CDRs are IMGT CDRs.

[0124] In some embodiments, the antibodies provided herein comprise a CDR-H3 selected from SEQ ID NO: 11. In some aspects, the CDR-H3 has at least about 50%, 75%, 80%, 85%, 90%, or 95% identity to the CDR-H3 of SEQ ID NO: 11. In some embodiments, the CDR-H3 is a CDR-H3 selected from SEQ ID NO: 11 having up to 1, 2, 3, 4, 5, 6, 7, or 8 amino acid substitutions. In some aspects, the amino acid substitutions are conservative amino acid substitutions. In some embodiments, the antibodies described in this paragraph are referred to herein as "variants". In some embodiments, such variants are derived from the sequences provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis, or any other method known in the art or described herein. In some embodiments, such variants are not derived from the sequences provided herein and may be isolated de novo, for example, according to the methods provided herein for obtaining antibodies.

[0125] In some embodiments, the antibodies provided herein include the CDR-H2 of SEQ ID NO: 10. In some aspects, the CDR-H2 has at least about 50%, 75%, 80%, 85%, 90%, or 95% identity to the CDR-H2 of SEQ ID NO: 10. In some embodiments, the CDR-H2 is the CDR-H2 of SEQ ID NO: 10 having up to 1, 2, 3, 4, 5, 6, 7, or 8 amino acid substitutions. In some aspects, the amino acid substitutions are conservative amino acid substitutions. In some embodiments, the antibodies described in this paragraph are referred to herein as "variants". In some embodiments, such variants are derived from the sequences provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis, or any other method known in the art or described herein. In some embodiments, such variants are not derived from the sequences provided herein and may be isolated de novo, for example, according to the methods provided herein for obtaining antibodies.

[0126] In some embodiments, the antibodies provided herein include the CDR-H1 of SEQ ID NO: 9. In some aspects, the CDR-H1 has at least about 50%, 75%, 80%, 85%, 90%, or 95% identity to the CDR-H1 of SEQ ID NO: 9. In some embodiments, the CDR-H1 is the CDR-H1 of SEQ ID NO: 9 having up to 1, 2, 3, 4, 5, 6, 7, or 8 amino acid substitutions. In some aspects, the amino acid substitutions are conservative amino acid substitutions. In some embodiments, the antibodies described in this paragraph are referred to herein as "variants". In some embodiments, such variants are derived from the sequences provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis, or any other method known in the art or described herein. In some embodiments, such variants are not derived from the sequences provided herein and may be isolated de novo, for example, according to the methods provided herein for obtaining antibodies.

[0127] In some embodiments, the antibodies provided herein comprise CDR-H3 of SEQ ID NO: 11 and CDR-H2 of SEQ ID NO: 10. In some embodiments, the antibodies provided herein comprise CDR-H3 of SEQ ID NO: 11, CDR-H2 of SEQ ID NO: 10, and CDR-H1 of SEQ ID NO: 9. In some embodiments, CDR-H3 has at least about 50%, 75%, 80%, 85%, 90%, or 95% identity with CDR-H3 of SEQ ID NO: 11, CDR-H2 has at least about 50%, 75%, 80%, 85%, 90%, or 95% identity with CDR-H2 of SEQ ID NO: 10, and CDR-H1 has at least about 50%, 75%, 80%, 85%, 90%, or 95% identity with CDR-H1 of SEQ ID NO: 9. In some embodiments, CDR-H3 is CDR-H3 of SEQ ID NO: 11 having up to 1, 2, 3, 4, 5, 6, 7, or 8 amino acid substitutions; CDR-H2 is CDR-H2 of SEQ ID NO: 10 having up to 1, 2, 3, 4, 5, 6, 7, or 8 amino acid substitutions; and CDR-H1 is CDR-H1 of SEQ ID NO: 9 having up to 1, 2, 3, 4, or 5 amino acid substitutions. In some aspects, the amino acid substitutions are conservative amino acid substitutions. In some embodiments, the antibodies described in this paragraph are referred to herein as "variants". In some embodiments, such variants are derived from the sequences provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis, or any other method known in the art or described herein. In some embodiments, such variants are not derived from the sequences provided herein and may be isolated de novo, for example, according to the methods provided herein for obtaining antibodies.

[0128] In some embodiments, the antibodies provided herein include the CDR-L3 of SEQ ID NO: 14. In some aspects, the CDR-L3 has at least about 50%, 75%, 80%, 85%, 90%, or 95% identity to the CDR-L3 of SEQ ID NO: 14. In some embodiments, the CDR-L3 is the CDR-L3 of SEQ ID NO: 14 having up to 1, 2, 3, 4, 5, 6, 7, or 8 amino acid substitutions. In some aspects, the amino acid substitutions are conservative amino acid substitutions. In some embodiments, the antibodies described in this paragraph are referred to herein as "variants". In some embodiments, such variants are derived from the sequences provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis, or any other method known in the art or described herein. In some embodiments, such variants are not derived from the sequences provided herein and may be isolated de novo, for example, according to the methods provided herein for obtaining antibodies.

[0129] In some embodiments, the antibodies provided herein include the CDR-L2 of SEQ ID NO: 13. In some aspects, the CDR-L2 has at least about 50%, 75%, 80%, 85%, 90%, or 95% identity to the CDR-L2 of SEQ ID NO: 13. In some embodiments, the CDR-L2 is the CDR-L2 of SEQ ID NO: 13 having up to 1, 2, 3, 4, 5, 6, 7, or 8 amino acid substitutions. In some aspects, the amino acid substitutions are conservative amino acid substitutions. In some embodiments, the antibodies described in this paragraph are referred to herein as "variants". In some embodiments, such variants are derived from the sequences provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis, or any other method known in the art or described herein. In some embodiments, such variants are not derived from the sequences provided herein and may be isolated de novo, for example, according to the methods provided herein for obtaining antibodies.

[0130] In some embodiments, the antibodies provided herein include the CDR-L1 of SEQ ID NO: 12. In some aspects, the CDR-L1 has at least about 50%, 75%, 80%, 85%, 90%, or 95% identity to the CDR-L1 of SEQ ID NO: 12. In some embodiments, the CDR-L1 is the CDR-L1 of SEQ ID NO: 12 having up to 1, 2, 3, 4, 5, 6, 7, or 8 amino acid substitutions. In some aspects, the amino acid substitutions are conservative amino acid substitutions. In some embodiments, the antibodies described in this paragraph are referred to herein as "variants". In some embodiments, such variants are derived from the sequences provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis, or any other method known in the art or described herein. In some embodiments, such variants are not derived from the sequences provided herein and may be isolated de novo, for example, according to the methods provided herein for obtaining antibodies.

[0131] In some embodiments, the antibodies provided herein comprise CDR-L3 of SEQ ID NO: 14 and CDR-L2 of SEQ ID NO: 13. In some embodiments, the antibodies provided herein comprise CDR-L3 of SEQ ID NO: 14, CDR-L2 of SEQ ID NO: 13, and CDR-L1 of SEQ ID NO: 12. In some embodiments, CDR-L3 has at least about 50%, 75%, 80%, 85%, 90%, or 95% identity to CDR-L3 of SEQ ID NO: 14, CDR-L2 has at least about 50%, 75%, 80%, 85%, 90%, or 95% identity to CDR-L2 of SEQ ID NO: 13, and CDR-L1 has at least about 50%, 75%, 80%, 85%, 90%, or 95% identity to CDR-L1 of SEQ ID NO: 12. In some embodiments, CDR-L3 is CDR-L3 of SEQ ID NO: 14 having up to 1, 2, 3, 4, or 5 amino acid substitutions; CDR-L2 is CDR-L2 of SEQ ID NO: 13 having up to 1, 2, 3, or 4 amino acid substitutions; CDR-L1 is CDR-L1 of SEQ ID NO: 12 having up to 1, 2, 3, 4, 5, or 6 amino acid substitutions. In some aspects, the amino acid substitutions are conservative amino acid substitutions. In some embodiments, the antibodies described in this paragraph are referred to herein as "variants". In some embodiments, such variants are derived from the sequences provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis, or any other method known in the art or described herein. In some embodiments, such variants are not derived from the sequences provided herein and may be isolated de novo, for example, according to the methods provided herein for obtaining antibodies.

[0132] In some embodiments, the antibodies provided herein comprise CDR-H3 of SEQ ID NO: 11, CDR-H2 of SEQ ID NO: 10, CDR-H1 of SEQ ID NO: 9, CDR-L3 of SEQ ID NO: 14, CDR-L2 of SEQ ID NO: 13, and CDR-L1 of SEQ ID NO: 12. In some embodiments, CDR-H3 has at least about 50%, 75%, 80%, 85%, 90%, or 95% identity with CDR-H3 of SEQ ID NO: 11, CDR-H2 has at least about 50%, 75%, 80%, 85%, 90%, or 95% identity with CDR-H2 of SEQ ID NO: 10, CDR-H1 has at least about 50%, 75%, 80%, 85%, 90%, or 95% identity with CDR-H1 of SEQ ID NO: 9, CDR-L3 has at least about 50%, 75%, 80%, 85%, 90%, or 95% identity with CDR-L3 of SEQ ID NO: 14, CDR-L2 has at least about 50%, 75%, 80%, 85%, 90%, or 95% identity with CDR-L2 of SEQ ID NO: 13, and CDR-L1 has at least about 50%, 75%, 80%, 85%, 90%, or 95% identity with CDR-L1 of SEQ ID NO: 12. In some embodiments, CDR-H3 is CDR-H3 of SEQ ID NO: 11 having a maximum of 1, 2, 3, 4, 5, 6, 7, or 8 amino acid substitutions; CDR-H2 is CDR-H2 of SEQ ID NO: 10 having a maximum of 1, 2, 3, 4, 5, 6, 7, or 8 amino acid substitutions; CDR-H1 is CDR-H1 of SEQ ID NO: 9 having a maximum of 1, 2, 3, 4, or 5 amino acid substitutions; CDR-L3 is CDR-L3 of SEQ ID NO: 14 having a maximum of 1, 2, 3, 4, or 5 amino acid substitutions; CDR-L2 is CDR-L2 of SEQ ID NO: 13 having a maximum of 1, 2, 3, or 4 amino acid substitutions; CDR-L1 is CDR-L1 of SEQ ID NO: 12 having a maximum of 1, 2, 3, 4, 5, or 6 amino acid substitutions. In some aspects, the amino acid substitutions are conservative amino acid substitutions. In some embodiments, the antibodies described in this paragraph are referred to herein as "variants." In some embodiments, such variants are derived from the sequences provided herein, for example, by affinity maturation, site-directed mutagenesis, random mutagenesis, or any other method known in the art or described herein.In some embodiments, such a manifold is not derived from the sequences provided herein and may be isolated de novo, for example, according to the methods provided herein for obtaining antibodies.

[0133] In some embodiments, the antibodies provided herein comprise CDR-H1 of SEQ ID NO: 9, CDR-H2 of SEQ ID NO: 10, CDR-H3 of SEQ ID NO: 11, CDR-L1 of SEQ ID NO: 12, CDR-L2 of SEQ ID NO: 13, and CDR-L1 of SEQ ID NO: 14.

[0134] Fc region The terms “Fc domain” or “Fc region” herein are used to define the C-terminal region of an immunoglobulin heavy chain that contains at least a portion of the constant region. The terms include native sequence Fc regions and variant Fc regions. Unless otherwise specified herein, the numbering of amino acid residues in the Fc region or constant region is by the EU numbering system, also called the EU index, as described in Kabat et al, Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD, 1991. The “Fc polypeptide” of a dimeric Fc herein refers to one of the two polypeptides that form the dimeric Fc domain, i.e., a polypeptide that includes the C-terminal constant region of an immunoglobulin heavy chain capable of stable self-association. For example, the Fc polypeptide of a dimeric IgG Fc includes the IgG CH2 and IgG CH3 constant domain sequences. Fc can be of the classes IgA, IgD, IgE, IgG, and IgM, some of which may be further divided into subclasses (isotypes), e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2.

[0135] The terms "Fc receptor" and "FcR" are used to describe receptors that bind to the Fc region of an antibody. For example, the FcR can be a native sequence human FcR. In general, FcRs bind to IgG antibodies (gamma receptors) and include receptors of the FcγRI, FcγRII, and FcγRIII subclasses, including allelic variants and alternatively spliced forms of these receptors. The FcγRII receptor includes FcγRIIA ("activating receptor") and FcγRIIB ("inhibitory receptor"), which mainly have similar amino acid sequences with different cytoplasmic domains. Other isotypes of immunoglobulins can also bind to specific FcRs (see, e.g., Janeway et al., Immuno Biology: the immune system in health and disease, (Elsevier Science Ltd., NY) (4th ed., 1999)). The activating receptor FcγRIIA contains an immunoreceptor tyrosine-based activation motif (ITAM) in its cytoplasmic domain. The inhibitory receptor FcγRIIB contains an immunoreceptor tyrosine-based inhibitory motif (ITIM) in its cytoplasmic domain (reviewed in Daeron, Annu. Rev. Immunol. 15:203 - 234 (1997)). FcRs are reviewed in Ravetch and Kinet, Annu. Rev. Immunol 9:457 - 92 (1991); Capel et al., Immunomethods 4:25 - 34 (1994), and de Haas et al., J. Lab. Clin. Med. 126:330 - 41 (1995). Other FcRs, including those to be identified in the future, are included within the term "FcR" herein. The term also includes the neonatal receptor FcRn, which is involved in the transfer of maternal IgG to the fetus (Guyer et al., J. Immunol. 117:587 (1976); and Kim et al., J. Immunol. 24:249 (1994)).

[0136] In some embodiments, the antibody is an IgG1 antibody.

[0137] In some embodiments, the antibody is an IgG3 antibody.

[0138] In some embodiments, the antibody is an IgG2 antibody.

[0139] In some embodiments, the antibody is an IgG4 antibody.

[0140] Modifications of the CH2 domain can affect the binding of FcRs to Fc. A number of amino acid modifications of the Fc region for selectively modifying the affinity of Fc for different Fc-gamma (Fcγ) receptors are known in the art. In one embodiment, the Fc comprises one or more modifications to facilitate selective binding of Fc-gamma receptors.

[0141] Exemplary mutations that alter the binding of FcRs to Fc are listed below: S298A / E333A / K334A, S298A / E333A / K334A / K326A (Lu Y, Vernes JM, Chiang N, et al. J Immunol Methods. 2011 Feb 28;365(1-2):132-41); F243L / R292P / Y300L / V305I / P396L, F243L / R292P / Y300L / L235V / P396L (Stavenhagen JB, Gorlatov S, Tuaillon N, et al. Cancer Res. 2007 Sep 15;67(18):8882-90; Nordstrom JL, Gorlatov S, Zhang W, et al. Breast Cancer Res. 2011 Nov 30;13(6):R123); F243L (Stewart R, Thom G, Levens M, et al. Protein Eng Des Sel. 2011 Sep;24(9):671-8.), S298A / E333A / K334A (Shields RL, Namenuk AK, Hong K, et al. J Biol Chem. 2001 Mar 2;276(9):6591-604); S239D / I332E / A330L, S239D / I332E (Lazar GA, Dang W, Karki S, et al. Proc Natl Acad Sci U S A. 2006 Mar 14;103(11):4005-10); S239D / S267E, S267E / L328F (Chu SY, Vostiar I, Karki S, et al. Mol Immunol. 2008 Sep;45(15):3926-33); S239D / D265S / S298A / I332E, S239E / S298A / K326A / A327H, G237F / S298A / A330L / I332E, S239D / I332E / S298A, S239D / K326E / A330L / I332E / S298A, G236A / S239D / D270L / I332E, S239E / S267E / H268D, L234F / S267E / N325L, G237F / V266L / S267D and other mutations listed in WO2011 / 120134 and WO2011 / 120135 (incorporated herein by reference). Therapeutic Antibody Engineering (William R. Strohl and Lila M. Strohl, Woodhead Publishing series in Biomedicine No 11, ISBN 1 907568 37 9, Oct 2012) lists mutations on page 283.

[0142] In some embodiments, the antibodies described herein include modifications that improve the ability to mediate effector functions. Such modifications are known in the art and include afucosylation, or manipulation of the affinity of Fc for activating receptors, mainly FCGR3a in the case of ADCC and C1q in the case of CDC. Table B below summarizes various designs reported in the literature regarding effector function manipulation.

[0143] In certain embodiments, the antibodies provided herein include an Fc region having one or more amino acid substitutions that improve ADCC, such as substitutions at one or more of positions 298, 333, and 334 of the Fc region. In some embodiments, the antibodies provided herein include an Fc region having one or more amino acid substitutions at positions 239, 332, and 330, as described in Lazar et al., Proc. Natl. Acad. Sci. USA, 2006, 103:4005 - 4010, which is incorporated herein by reference in its entirety.

[0144] In some embodiments, the antibodies provided herein include one or more modifications that improve or decrease C1q binding and / or CDC. See U.S. Patent No. 6,194,551, WO99 / 51642, and Idusogie et al., J. Immunol., 2000, 164:4178 - 4184, each of which is incorporated herein by reference in its entirety.

[0145] Accordingly, in one embodiment, the antibodies described herein may include a dimeric Fc comprising one or more amino acid modifications as described in Table B that confer improved effector function. In another embodiment, the antibody can be afucosylated to improve effector function.

[0146] (Table B) CH2 Domain and Effector Function Manipulation TIFF2025109867000003.tif127148

[0147] Fc modifications that reduce FcγR and / or complement binding and / or effector functions are known in the art. Recent publications describe strategies that have been used to engineer antibodies with reduced or silenced effector activity (see Strohl, WR (2009), Curr Opin Biotech 20:685-691, and Strohl, WR and Strohl LM, “Antibody Fc engineering for optimal antibody performance” In Therapeutic Antibody Engineering, Cambridge: Woodhead Publishing (2012), pp 225-249). These strategies include reducing effector function by modification of glycosylation, use of IgG2 / IgG4 scaffolds, or introduction of mutations into the hinge or CH2 region of the Fc. For example, US Patent Publication No. 2011 / 0212087 (Strohl), International Patent Publication No. WO2006 / 105338 (Xencor), US Patent Publication No. 2012 / 0225058 (Xencor), US Patent Publication No. 2012 / 0251531 (Genentech), and Strop et al ((2012) J. Mol. Biol. 420:204-219) describe specific modifications that reduce FcγR or complement binding to the Fc.

[0148] Specific non-limiting examples of known amino acid modifications that reduce FcγR or complement binding to the Fc include those identified in Table C below.

[0149] (Table C) Modifications that reduce FcγR or complement binding to the Fc TIFF2025109867000004.tif109128

[0150] Methods for producing antibodies that have little or no fucose at the Fc glycosylation site (Asn297 EU numbering) without modifying the amino acid sequence are well known in the art. The GlymaxX® technology (ProBioGen AG) is based on the introduction of the gene of an enzyme that biases the cellular pathway of fucose biosynthesis into the cells used for antibody production. This prevents the addition of the sugar "fucose" to the N-linked antibody carbohydrate moiety by the antibody-producing cells (von Horsten et al. (2010) Glycobiology. 2010 Dec;20(12):1607-18). Examples of cell lines capable of producing defucosylated antibodies include CHO-DG44 with stable overexpression of the bacterial redox enzyme GDP-6-deoxy-D-lyxo-4-hexulose reductase (RMD) that is deficient in protein fucosylation (see Henning von Horsten et al., Glycobiol 2010, 20:1607-1618) or Lec13 CHO cells (Ripka et al., Arch. Biochem. Biophys., 1986, 249:533-545; US Patent Publication No. 2003 / 0157108; WO2004 / 056312 (each of which is incorporated herein by reference in its entirety)), as well as knockout cell lines such as the alpha-1,6-fucosyltransferase gene or FUT8 knockout CHO cells (Yamane-Ohnuki et al., Biotech. Bioeng., 2004, 87:614-622; Kanda et al., Biotechnol. Bioeng., 2006, 94:680-688; and WO 2003 / 085107 (each of which is incorporated herein by reference in its entirety)). Another approach for obtaining antibodies with reduced levels of fucosylation can be found in US Patent No. 8,409,572, which teaches selecting a cell line for antibody production based on its ability to cause a decrease in the level of fucosylation of the antibody.

[0151] The antibodies can be fully afucosylated (meaning that they do not contain detectable fucose), or they can be partially afucosylated, which means that the isolated antibody contains less than 95%, less than 85%, less than 75%, less than 65%, less than 55%, less than 45%, less than 35%, less than 25%, less than 15%, or less than 5% of the amount of fucose that is typically detected for similar antibodies produced by mammalian expression systems.

[0152] In some embodiments, the antibodies provided herein comprise an IgG1 domain with reduced fucose content at position Asn297 as compared to a naturally occurring IgG1 domain. Such Fc domains are known to have improved ADCC. See Shields et al., J. Biol. Chem., 2002, 277:26733 - 26740, which is incorporated herein by reference in its entirety. In some embodiments, such antibodies do not contain any fucose at position Asn297. The amount of fucose can be determined using any suitable method, such as that described in WO2008 / 077546, which is incorporated herein by reference in its entirety.

[0153] In some embodiments, the antibodies provided herein comprise a bisecting - type oligosaccharide, such as a bisecting - type oligosaccharide that binds to the Fc region of the antibody and is bisected with GlcNAc. Such antibody variants can have reduced fucosylation and / or improved ADCC function. Examples of such antibody variants are described, for example, in WO2003 / 011878; U.S. Patent No. 6,602,684, and U.S. Patent Publication No. 2005 / 0123546, each of which is incorporated herein by reference in its entirety.

[0154] Other exemplary glycosylation variants that can be incorporated into the antibodies provided herein are described, for example, in U.S. Patent Publication Nos. 2003 / 0157108, 2004 / 0093621, 2003 / 0157108, 2003 / 0115614, 2002 / 0164328, 2004 / 0093621, 2004 / 0132140, 2004 / 0110704, 2004 / 0110282, 2004 / 0109865; International Patent Publication Nos. 2000 / 61739, 2001 / 29246, 2003 / 085119, 2003 / 084570, 2005 / 035586, 2005 / 035778, 2005 / 053742, 2002 / 031140, Okazaki et al., J. Mol. Biol., 2004, 336:1239-1249, and Yamane-Ohnuki et al., Biotech. Bioeng., 2004, 87:614-622, each of which is incorporated herein by reference in its entirety.

[0155] In some embodiments, the antibodies provided herein comprise an Fc region having at least one galactose residue in the oligosaccharide attached to the Fc region. Such antibody variants can have improved CDC function. Examples of such antibody variants are described, for example, in WO1997 / 30087; WO1998 / 58964; and WO1999 / 22764, each of which is incorporated herein by reference in its entirety.

[0156] Examples of cell lines capable of producing defucosylated antibodies include CHO-DG44 with stable overexpression of the bacterial redox enzyme GDP-6-deoxy-D-lyxo-4-hexulose reductase (RMD) lacking protein fucosylation (see Henning von Horsten et al., Glycobiol 2010, 20:1607-1618) or Lec13 CHO cells (Ripka et al., Arch. Biochem. Biophys., 1986, 249:533-545; US Patent Publication No. 2003 / 0157108; WO2004 / 056312, each of which is incorporated by reference in its entirety), as well as knockout cell lines such as the alpha-1,6-fucosyltransferase gene or FUT8 knockout CHO cells (Yamane-Ohnuki et al., Biotech. Bioeng., 2004, 87:614-622; Kanda et al., Biotechnol. Bioeng., 2006, 94:680-688; and WO2003 / 085107, each of which is incorporated by reference in its entirety).

[0157] In some embodiments, the antibody has antibody-dependent cell phagocytosis (ADCP) activity. ADCP can occur when an antibody binds to an antigen on the surface of a pathogenic or tumorigenic target cell. Phagocytic cells that possess Fc receptors on their cell surface, including monocytes and macrophages, recognize and bind to the Fc region of the antibody bound to the target cell. When the Fc receptor binds to the antibody-bound target cell, phagocytosis of the target cell can be initiated. ADCP can be considered a form of ADCC.

[0158] In some embodiments, the antibody is capable of forming immune complexes. For example, the immune complex can be a tumor cell coated with the antibody.

[0159] In some embodiments, the anti-TREM2 antibody does not substantially bind to bone marrow cells present outside of the cancer tissue. In some embodiments, the anti-TREM2 antibody does not substantially bind to stimulatory bone marrow cells present in the cancer tissue.

[0160] In some embodiments, the antibody is a monoclonal antibody.

[0161] In some embodiments, the antibody is a polyclonal antibody.

[0162] In some embodiments, the antibody is produced by a hybridoma. In other embodiments, the antibody is produced by recombinant cells engineered to express the desired variable and constant domains.

[0163] In some embodiments, the antibody can be a single-chain antibody or other antibody derivative that retains antigen specificity and a lower hinge region or a variant thereof.

[0164] In some embodiments, the antibody can be a multifunctional antibody, recombinant antibody, human antibody, humanized antibody, fragment, or a variant thereof. In certain embodiments, the antibody fragment or a derivative thereof is selected from Fab fragment, Fab’2 fragment, CDR, and ScFv.

[0165] In some embodiments, the antibody is specific for a surface antigen such as the TREM2 protein. In some embodiments, the therapeutic antibody is specific for a tumor antigen (e.g., a molecule specifically expressed by tumor cells). In certain embodiments, the therapeutic antibody may have a human or non-human primate IgG1 or IgG3 Fc portion.

[0166] Binding Regarding the binding of an antibody to a target molecule, the terms "binds to", "specifically binds to", "specifically binds specifically to", "is specific for", "selectively binds to", and "is selective for" with respect to a particular antigen (e.g., a polypeptide target) or an epitope on a particular antigen mean binding that is somewhat different from non-specific or non-selective interactions (e.g., with non-target molecules). Specific binding can be measured, for example, by measuring the binding to the target molecule and comparing it to the binding to non-target molecules. Specific binding can also be determined by competition with a competitor molecule that mimics the epitope recognized on the target molecule. In that case, specific binding is indicated if the binding of the antibody to the target molecule is competitively inhibited by the competitor molecule.

[0167] "Affinity" refers to the total strength of non-covalent interactions between a single binding site of a molecule (e.g., an antibody) and its binding partner (e.g., an antigen or an epitope). Unless otherwise specified, as used herein, "affinity" refers to the intrinsic binding affinity that reflects a 1:1 interaction between members of a binding pair (e.g., an antibody and an antigen or an epitope). The affinity of molecule X for partner Y can be expressed by the dissociation equilibrium constant (K D ). The kinetic components contributing to the dissociation equilibrium constant are described in more detail below. Affinity can be measured by common methods known in the art, including those described herein, such as surface plasmon resonance (SPR) technology (e.g., BIACORE®) or biolayer interferometry (e.g., FORTEBIO®).

[0168] As used herein, the term "k d " (sec -1 ) refers to the dissociation rate constant of a particular antibody-antigen interaction. This value is also called the k off value.

[0169] As used herein, the term "k a " (M -1 ×sec -1 ) refers to the association rate constant of a particular antibody-antigen interaction. This value is the kon is also referred to as a value.

[0170] As used herein, the term "K D "(M) refers to the dissociation equilibrium constant of a particular antibody-antigen interaction. K D = k d / k a . In some embodiments, the affinity of an antibody is described with respect to the K D for the interaction between such an antibody and its antigen. For clarity, as is known in the art, a smaller K D value indicates a high affinity interaction, while a larger K D value indicates a low affinity interaction.

[0171] As used herein, the term "K A "(M -1 ) refers to the association equilibrium constant of a particular antibody-antigen interaction. K A = k a / k d .

[0172] When used in the context of two or more antibodies in this specification, the terms "competes with" or "cross-competes with" indicate that two or more antibodies compete for binding to an antigen (e.g., TREM2). In one exemplary assay, TREM2 is coated on a surface, contacted with a first TREM2 antibody, and then a second TREM2 antibody is added. In another exemplary assay, a first TREM2 antibody is coated on a surface, contacted with TREM2, and then a second TREM2 antibody is added. In either assay, if the presence of the first TREM2 antibody reduces the binding of the second TREM2 antibody, the antibodies compete with each other. The term "competes with" also includes combinations of antibodies where one antibody reduces the binding of another antibody, but no competition is observed when the antibodies are added in the reverse order. However, in some embodiments, the first and second antibodies inhibit each other's binding regardless of the order in which they are added. In some embodiments, one antibody reduces the binding of another antibody to its antigen by at least 25%, at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, or at least 95%. One of ordinary skill in the art can select the concentrations of the antibodies used in a competition assay based on the affinity of the antibodies for TREM2 and the valency of the antibodies. The assays described in this definition are exemplary, and one of ordinary skill in the art can utilize any suitable assay to determine whether antibodies compete with each other. Suitable assays are described, for example, in Cox et al., "Immunoassay Methods," in Assay Guidance Manual [Internet], Updated December 24, 2014 (www.ncbi.nlm.nih.gov / books / NBK92434 / ; accessed September 29, 2015); Silman et al., Cytometry, 2001, 44:30-37; and Finco et al., J. Pharm. Biomed. Anal., 2011, 54:351-358, each of which is incorporated by reference in its entirety.

[0173] In some embodiments, the antibodies provided herein have a mAb content of about 0.001, 0.01, 0.02, 0.03, 0.04, 0.05, 0.06, 0.07, 0.08, 0.09, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 1.95, 2, 3, 4, 5, 6, 7, 8, 9, or 10x10 as measured by a Biacore assay. -9 K below M D In some embodiments, the K of the antibodies provided herein binds to human TREM2. D is approximately 0.001-0.01, 0.01-0.1, 0.01-0.05, 0.05-0.1, 0.1-0.5, 0.5-1, 0.25-0.75, 0.25-0.5, 0.5-0.75, 0.75-1, 0.75-2, 1.1-1.2, 1.2-1.3, 1.3-1.4, 1.4-1.5, 1.5-1.6, 1.6-1.7, 1.7-1.8, 1.8-1.9, 1.9-2, 1-2, 1-5, 2-7, 3-8, 3-5, 4-6, 5-7, 6-8, 7-9, 7-10, or 5-10x10 as measured by Biacore assay. -9 It's M.

[0174] In some embodiments, the antibodies provided herein have a cytoplasmic affinity of about 2, 1.98, 1.95, 1.9, 1.85, 1.8, 1.75, 1.7, 1.65, 1.6, 1.55, 1.50, 1.45, or 1.4x10 as measured by a Biacore assay. -9 K below M D In some embodiments, the antibody binds to human TREM2 at 1.9-1.8, 1.8-1.7, 1.7-1.6, 1.6-1.5, or 1.9-1.5x10 as measured by a Biacore assay. -9 K of M D Provided herein are antibodies that bind to human TREM2 at about 10, 9.56, 9.5, 9.0, 8.88, 8.84, 8.5, 8, 7.5, 7.32, 7, 6.5, 6, 5.5, 5, 4.5, 4, 3.5, 3, 2.5, 2, 1.5, or 1x10 as measured by a Biacore assay. -4K at (1 / s) below d binds to human TREM2. In some embodiments, the antibodies provided herein have a K of 7-10, 7-8, 8-9, 9-10, 7-7.5, 7.5-8, 8-8.5, 8.5-9, 9-9.5, or 9.5-10 x 10 -4 (1 / s) for binding to human TREM2. In some embodiments, the antibodies provided herein have a K of about 4, 4.1, 4.2, 4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, 45, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6, 7, 8, 9, or 10 x 10 d (1 / Ms) or greater for binding to human TREM2. In some embodiments, the antibodies provided herein have a K of 4-7, 4-4.5, 4.5-5, 5-5.5, 5.5-6, 6-6.5, or 6.5-7, 7-8, 8-9, or 9-10 x 10 5 (1 / Ms) for binding to human TREM2. a In some embodiments, the antibodies provided herein bind to human TREM2 with an EC50 of 2, 1.9, 1.8, 1.7, 1.6, 1.5, 1.4, 1.3, 1.2, 1.1, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, or 0.1 nM or less as measured by flow cytometry. In some embodiments, the antibody binds to human TREM2 with an EC50 of 0.6-1.4 nM as measured by flow cytometry. In some embodiments, the antibody binds to human TREM2 with an EC50 of about 0.5, 0.6, 0.9, 1.1, 1.2, 1.3, 1.4, or 1.5 nM as measured by flow cytometry. 5 (1 / Ms) for binding to human TREM2. a for binding to human TREM2.

[0175] In some embodiments, the antibodies provided herein bind to human TREM2 with an EC50 of 2, 1.9, 1.8, 1.7, 1.6, 1.5, 1.4, 1.3, 1.2, 1.1, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, or 0.1 nM or less as measured by flow cytometry. In some embodiments, the antibody binds to human TREM2 with an EC50 of 0.6-1.4 nM as measured by flow cytometry. In some embodiments, the antibody binds to human TREM2 with an EC50 of about 0.5, 0.6, 0.9, 1.1, 1.2, 1.3, 1.4, or 1.5 nM as measured by flow cytometry.

[0176] In some embodiments, the antibodies provided herein bind to mouse TREM2 with an EC50 of 2, 1.9, 1.8, 1.7, 1.6, 1.5, 1.4, 1.3, 1.2, 1.1, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, or 0.1 nM or less as measured by flow cytometry. In some embodiments, the antibody binds to mouse TREM2 with an EC50 of 0.6 - 1.4 nM as measured by flow cytometry. In some embodiments, the antibody binds to mouse TREM2 with an EC50 of approximately 0.5, 0.6, 0.9, 1.1, 1.2, 1.3, 1.4, or 1.5 nM as measured by flow cytometry.

[0177] In some embodiments, the antibodies provided herein do not bind to human TREM2 with an EC50 of 20 nM or greater as measured by flow cytometry. In some embodiments, the antibodies provided herein do not bind to mouse TREM2 with an EC50 of 3 nM or greater as measured by flow cytometry.

[0178] To screen for antibodies that bind to an epitope on a target antigen to which the antibody of interest (e.g., TREM2) binds, a standard cross - blocking assay as described in Antibodies, A Laboratory Manual, Cold Spring Harbor Laboratory, Ed Harlow and David Lane (1988) can be performed. Alternatively or additionally, epitope mapping can be performed by methods known in the art.

[0179] Competition between antibodies can be determined in an assay where the test antibody inhibits or blocks the specific binding of a reference antibody to a common antigen (see, e.g., Junghans et al., Cancer Res. 50:1495, 1990; Fendly et al., Cancer Research 50:1550 - 1558; US 6,949,245). A test antibody competes with the reference antibody if excess test antibody (e.g., at least 2x, 5x, 10x, 20x, or 100x) inhibits or blocks the binding of the reference antibody, as measured in a competitive binding assay, by, for example, at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, or 99%. Antibodies identified by a competition assay (competing antibodies) include antibodies that bind to the same epitope as the reference antibody and antibodies that bind to an adjacent epitope that is close enough to the epitope to which the reference antibody binds such that steric hindrance occurs. For example, a second competing antibody that competes with the first antibody described herein for binding to TREM2 can be identified. In certain cases, the second antibody can block or inhibit the binding of the first antibody by, for example, at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, or 99%, as measured in a competitive binding assay. In certain cases, the second antibody can displace the first antibody by more than 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, or 99%.

[0180] Function In some embodiments, the antibody has antibody-dependent cell-mediated cytotoxicity (ADCC) activity. ADCC can occur when an antibody binds to an antigen on the surface of a pathogenic or tumor-forming target cell. Effector cells having an Fc gamma receptor (FcγR or FCGR) on their cell surface, including cytotoxic T cells, natural killer (NK) cells, macrophages, neutrophils, eosinophils, dendritic cells, or monocytes, recognize and bind to the Fc region of the antibody bound to the target cell. Such binding can trigger activation of intracellular signaling pathways leading to cell death. In certain embodiments, the immunoglobulin Fc region subtype (isotype) of the antibody includes human IgG1 and IgG3. As used herein, ADCC refers to a cell-mediated reaction in which non-specific cytotoxic cells (e.g., natural killer (NK) cells, neutrophils, and macrophages) that express an Fc receptor (FcR) recognize an antibody bound to a target cell and subsequently cause lysis of the target cell. NK cells, which are primary cells that mediate ADCC, express only FcγRIII, while monocytes express FcγRI, FcγRII, and FcγRIII. FcR expression on hematopoietic cells is summarized in Table 3 on page 464 of Ravetch and Kinet, Annu. Rev. Immunol 9:457-92 (1991). To assess the ADCC activity of a molecule of interest, an in vitro ADCC assay such as that described in U.S. Patent No. 5,500,362 or 5,821,337 may be performed. Effector cells useful in such assays include peripheral blood mononuclear cells (PBMC) and natural killer (NK) cells. Alternatively or additionally, the ADCC activity of a molecule of interest may be evaluated in vivo in an animal model such as that disclosed in Clynes et al., Proc. Natl. Acad. Sci. (USA) 95:652-656 (1998).

[0181] In some embodiments, the antibody has complement-dependent cytotoxicity (CDC) activity. Antibody-mediated CDC is mediated by proteins of the classical complement cascade and is triggered by the binding of the complement protein C1q to the antibody. The antibody Fc region that binds Clq can induce activation of the complement cascade. In certain embodiments, the immunoglobulin Fc region subtypes (isotypes) of the antibody include human IgG1 and IgG3. As used herein, CDC refers to the ability of a molecule to lyse a target in the presence of complement. The complement activation pathway is initiated by the binding of the first component of the complement system (C1q) to a molecule (e.g., a polypeptide such as an antibody) complexed with a homologous antigen. To assess complement activation, a CDC assay may be performed, such as that described in Gazzano-Santoro et al., J. Immunol. Methods 202:163 (1996).

[0182] In some embodiments, the antibody is an agonist antibody. An agonist antibody can induce (e.g., increase) one or more activities or functions of NSM after the antibody binds to the TREM2 protein expressed on the cell. An agonist antibody can bind and activate NSM, causing a change in cell proliferation or modifying antigen presentation ability. An agonist antibody can bind and activate NSM, inducing an intracellular signaling pathway that leads to modification of cell growth or apoptosis.

[0183] In some embodiments, the antibody is an antagonist antibody. An antagonist antibody can block (e.g., decrease) one or more activities or functions of NSM after the antibody binds to the TREM2 protein expressed on the cell. For example, an antagonist antibody can bind and block a ligand that binds to one or more NSM proteins, preventing cell differentiation and proliferation or modifying antigen presentation ability. An antagonist antibody can bind to the TREM2 protein by a ligand to prevent activation, prevent binding and activation, and modify an intracellular signaling pathway that contributes to cell growth and survival.

[0184] In some embodiments, the antibody is a depletion antibody. The depletion antibody kills non-stimulatory myeloid cells upon contact via the interaction of the antibody with other immune cells of the molecule. For example, when the antibody binds to cells carrying the TREM2 protein, it can capture complement proteins and induce complement-dependent cell lysis. When the antibody binds to cells with the TREM2 protein, it can also induce neighboring cells with Fc receptors to kill them by antibody-dependent cell-mediated cytotoxicity (ADCC).

[0185] In some embodiments, the antibody is a neutralizing antibody, and the antibody neutralizes one or more biological activities of NSM. In some embodiments, the TREM2 protein is expressed on the surface of non-stimulatory myeloid cells, and the antibody recognizes the extracellular domain of the TREM2 protein.

[0186] In some embodiments, the antibody is selective for NSM (preferentially binds to TREM2). In certain embodiments, the antibody that selectively binds to NSM has a dissociation constant (Kd) in the range of 0.0001 nM to 1 μM. In certain embodiments, the antibody specifically binds to an epitope on the TREM2 protein that is conserved among proteins from different species. In another embodiment, the selective binding includes, but does not require, exclusive binding.

[0187] In one embodiment, the anti-TREM2 antibody that is bound to the target causes the depletion of non-stimulatory myeloid cells in vivo. In some embodiments, the effector proteins induced by the clustered antibody can induce various responses, including the release of inflammatory cytokines, the control of antigen production, endocytosis, or cell killing. In one embodiment, the antibody can mobilize and activate complement, or mediate antibody-dependent cell-mediated cytotoxicity (ADCC) in vivo, or mediate phagocytosis by binding to Fc receptors in vivo. The antibody can also deplete non-stimulatory myeloid cells by inducing apoptosis or necrosis of non-stimulatory myeloid cells upon binding.

[0188] In some embodiments, inactivation of non-stimulatory myeloid cells is in vitro and is achieved by a) killing the non-stimulatory myeloid cells, b) magnetic bead depletion of the non-stimulatory myeloid cells; or c) fluorescence-activated cell sorting (FACS) sorting of the non-stimulatory myeloid cells.

[0189] In some embodiments, the antibody is bound to or conjugated to an effector molecule. In certain embodiments, the antibody is conjugated to at least one therapeutic agent selected from the group consisting of a radionuclide, a cytotoxin, a chemotherapeutic agent, a drug, a prodrug, a toxin, an enzyme, an immunomodulatory agent, an anti-angiogenic agent, a pro-apoptotic agent, a cytokine, a hormone, an oligonucleotide, an antisense molecule, siRNA, a second antibody, and a second antibody fragment.

[0190] In certain embodiments, the antibody is conjugated to a drug, such as a toxin, a chemotherapeutic agent, an immunomodulator, or a radioisotope. Some methods of preparing ADCs (antibody-drug conjugates) are known in the art and are described, for example, in U.S. Patent Nos. 8,624,003 (pot method), 8,163,888 (one-step), and 5,208,020 (two-step method). The antibody or an antigen-binding fragment thereof can be conjugated to at least one agent including a radionuclide, a cytotoxin, a chemotherapeutic agent, a drug, a prodrug, a toxin, an enzyme, an immunomodulatory agent, an anti-angiogenic agent, a pro-apoptotic agent, a cytokine, a hormone, an oligonucleotide, an antisense molecule, siRNA, a second antibody, and a second antibody fragment that is antigen-binding.

[0191] Non-stimulatory myeloid cells (NSM) Methods and compositions for inactivating and / or detecting non-stimulatory myeloid cells (NSM) including the use of an anti-TREM2 antibody are described herein. Methods and compositions for targeting and / or detecting non-stimulatory myeloid cells that express the NSM protein are also provided herein.

[0192] Also provided herein are methods and compositions for ablating and / or detecting non-stimulatory myeloid cells, including the use of antibodies directed to non-human homologs of human NSM protein in said non-human individual.

[0193] As used herein, non-stimulatory myeloid cells are myeloid cells that are not sufficiently effective at stimulating an immune response (e.g., not effective at stimulating an anti-tumor response in the tumor microenvironment as compared to stimulatory myeloid cells). In some embodiments, non-stimulatory myeloid cells are not effective at presenting antigen (e.g., tumor antigen) to T cells or stimulating a tumor-specific T cell response as compared to stimulatory myeloid cells. In some embodiments, non-stimulatory myeloid cells may exhibit a decrease in the ability to uptake, process, and / or present tumor-associated antigen to T cells as compared to stimulatory myeloid cells. Non-stimulatory myeloid cells may have a reduced ability or no ability to re-prime cytotoxic T lymphocytes, or in some cases, may not be capable of stimulating effective killing of tumor cells. Non-stimulatory myeloid cells may exhibit a decrease in the expression of genes and cell surface markers involved in antigen processing, antigen presentation, and / or antigen co-stimulation, including but not limited to CD80, CD86, MHC I, and MHC II, as compared to stimulatory myeloid cells.

[0194] Non-stimulatory myeloid cells, when compared to stimulatory myeloid cells, although not limited, may show a decrease in the expression of genes related to cross-presentation, co-stimulation, and / or stimulatory cytokines, including any one or more of TAP1, TAP2, PSMB8, PSMB9, TAPBP, PSME2, CD24a, CD274, BTLA, CD40, CD244, ICOS-L, ICAM-1, TIM-3, PDL-2, RANK, FLT3, CSF2RB, CSF2RB2, CSF2RA, IL12b, XCR1, CCR7, CCR2, CCL22, CXCL9, and CCL5, as well as an increase in the expression of the anti-inflammatory cytokine IL-10. In some embodiments, non-stimulatory myeloid cells are determined by the transcription factor IRF4 and the cytokines GM-CSF or CSF-1 for differentiation and survival. In some embodiments, non-stimulatory myeloid cells may contribute to tumor angiogenesis by secreting vascular endothelial growth factor (VEGF) and nitric oxide synthase (NOS), and support tumor growth by secreting epidermal growth factor (EGF).

[0195] In some embodiments, non-stimulatory myeloid cells are tumor-associated macrophages (TAMs), neutrophils, monocytes, or dendritic cells (DCs). In some embodiments, non-stimulatory myeloid cells are not dendritic cells (DCs). In some embodiments, non-stimulatory myeloid cells are neutrophils.

[0196] In some embodiments, non-stimulatory myeloid cells are tumor-associated macrophages (TAMs). TAMs are macrophages that are present near or within a cancerous tumor and are derived from circulating monocytes or resident tissue macrophages.

[0197] In some embodiments, non-stimulatory bone marrow cells and stimulatory bone marrow cells are distinguished based on markers they express or markers they selectively express. The expression of a cell surface marker can be described as '+', or 'positive'. The absence of a cell surface marker can be described as '-', or 'negative'. The expression of a cell surface marker can further be described as 'high' (cells that express a high level of the marker) or 'low' (cells that express a low level of the marker), which indicates the relative expression of each marker on the cell surface. The level of a marker may be determined by various methods known in the art, such as immunostaining and FACS analysis, or gel electrophoresis and Western blotting.

[0198] In some embodiments, non-stimulatory bone marrow cells are dendritic cells (DCs). In some embodiments, dendritic cells can be distinguished by the morphology of their projections or dendrites. In one embodiment, non-stimulatory dendritic cells are at least CD45+, HLA-DR+, CD14-, CD11c+, and BDCA1+ (also referred to as DC1 cells). In one embodiment, non-stimulatory dendritic cells are not CD45+, HLA-DR+, CD14-, CD11c+, and BDCA3+ (also referred to as DC2 cells). In one embodiment, dendritic cells that are CD45+, HLA-DR+, CD14-, CD11c+, and BDCA3+ are stimulatory bone marrow cells.

[0199] In some embodiments, non-stimulatory bone marrow cells are tumor-associated macrophages. In some embodiments, for example, in humans, non-stimulatory tumor-associated macrophages are at least CD45+, HLA-DR+, CD14+. In some embodiments, non-stimulatory tumor-associated macrophages are at least CD45 + , HLA-DR + , CD14 + , CD11b + . In some embodiments, non-stimulatory tumor-associated macrophages are at least CD45 + , HLA-DR + , CD14 + , CD11c+ is. In some embodiments, the non-stimulatory tumor-associated macrophages are at least CD45 + , HLA-DR + , CD14 + , BDCA3 - is. In some embodiments, the non-stimulatory tumor-associated macrophages are at least CD45 + , HLA-DR + , CD14 + , BDCA3 - , CD11b + is. In some embodiments, the non-stimulatory tumor-associated macrophages are at least CD45 + , HLA-DR + , CD14 + , BDCA3 - , CD11c + is. In some embodiments, the non-stimulatory tumor-associated macrophages are at least CD45 + , HLA-DR + , CD14 + , CD11b + , and CD11c + is. In some embodiments, the non-stimulatory tumor-associated macrophages are at least CD45 + , HLA-DR + , CD14 + , BDCA3 - , CD11b + , and CD11c + is.

[0200] In some embodiments, the methods and compositions of the invention are useful for targeting TAM and DC in other mammals, such as mice. In such embodiments, mouse TAM and DC are contacted with a TREM2 antibody. In one embodiment, for example, in mice, tumor-associated macrophages are at least CD45+, HLA-DR+, CD14+, CD11b high , and CD11c low (also referred to as TAM1). In one embodiment, for example, in mice, tumor-associated macrophages are at least CD45+, HLA-DR+, CD14+, CD11b low , and CD11chigh (also referred to as TAM2). As used herein, the term "CD11b high macrophage" refers to macrophages that express high levels of CD11b. As used herein, the term "CD11b low macrophage" refers to CD11b high macrophages that express substantially lower levels of CD11b on their surface than CD11b high macrophages. As used herein, the term "CD11c low " refers to macrophages that express high levels of CD11c. As used herein, the term "CD11c low macrophage" refers to macrophages that express substantially lower levels of CD11c on their surface than Cd11c high macrophages.

[0201] In some embodiments, the non-stimulatory bone marrow cells of the present invention comprise one or more of TAM and DC1 cells.

[0202] In some embodiments, for example, in mice, the non-stimulatory bone marrow cells of the present invention comprise one or more of TAM1, TAM2, and DC1 cells. In such embodiments, the non-stimulatory bone marrow cells of the present invention are contacted with a TREM2 antibody.

[0203] In some embodiments, the non-stimulatory bone marrow cells are bone marrow cells that are tumor cells.

[0204] In some embodiments, the non-stimulatory bone marrow cells are localized within the periphery of the tumor lesion or within the transformed tumor vessels, and they contact autologous T cells. In one embodiment, the localization of the non-stimulatory bone marrow cells is modified such that the cells are no longer localized at the edge of the tumor or no longer in contact with T cells.

[0205] In some embodiments, the non-stimulatory bone marrow cells are in a population of immune cells that includes stimulatory and non-stimulatory bone marrow cells. In some embodiments, the non-stimulatory bone marrow cells are in a population of immune cells that includes only non-stimulatory bone marrow cells. The populations of immune cells of the invention are pure, homogeneous, non-homogeneous, derived from various sources (e.g., diseased tissue, tumor tissue, healthy tissue, cell banks), maintained in primary cell culture, and / or maintained in ex vivo culture.

[0206] In some embodiments, the non-stimulatory bone marrow cells are tumor-associated macrophages.

[0207] In some embodiments, the non-stimulatory bone marrow cells are dendritic cells.

[0208] In some embodiments, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 - , CD11c + , and BDCA1 + . In some embodiments, the non-stimulatory bone marrow cells include cells that are CD45 + , HLA-DR + , CD14 - , CD11c + , and BDCA1 + . In some embodiments, the non-stimulatory bone marrow cells consist of cells that are CD45 + , HLA-DR + , CD14 - , CD11c + , and BDCA1 + . In some embodiments, the non-stimulatory bone marrow cells consist essentially of cells that are CD45 + , HLA-DR + , CD14 - , CD11c + , and BDCA1 + .

[0209] In some embodiments, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 + , BDCA3- It is so. In some embodiments, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 + , BDCA3 - and include cells that are so. In some embodiments, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 + , BDCA3 - and consist of cells that are so. In some embodiments, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 + , BDCA3 - and are essentially composed of cells that are so.

[0210] In some embodiments, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 + , CD11b + and are so. In some embodiments, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 + , CD11b + and include cells that are so. In some embodiments, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 + , CD11b + and consist of cells that are so. In some embodiments, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 + , CD11b + and are essentially composed of cells that are so.

[0211] In some embodiments, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 + , CD11c + and are so. In some embodiments, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 + , CD11c+ comprises cells that are... In some embodiments, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 + , CD11c + cells. In some embodiments, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 + , CD11c + and consist essentially of cells.

[0212] In some embodiments, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 + , BDCA3 - , and CD11c + cells. In some embodiments, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 + , BDCA3 - , and CD11c + cells. In some embodiments, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 + , BDCA3 - , and CD11c + cells. In some embodiments, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 + , BDCA3 - , and CD11c + and consist essentially of cells.

[0213] In some embodiments, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 + , BDCA3 - , CD11b + cells. In some embodiments, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 +, BDCA3 - , CD11b + comprises cells that are... In some embodiments, the non-stimulatory myeloid cells are CD45 + , HLA-DR + , CD14 + , BDCA3 - , CD11b + consist of cells that are... In some embodiments, the non-stimulatory myeloid cells are CD45 + , HLA-DR + , CD14 + , BDCA3 - , CD11b + consist essentially of cells that are...

[0214] In some embodiments, the non-stimulatory myeloid cells are CD45 + , HLA-DR + , CD14 + , CD11b + , and CD11c + In some embodiments, the non-stimulatory myeloid cells are CD45 + , HLA-DR + , CD14 + , CD11b + , and CD11c + comprise cells that are... In some embodiments, the non-stimulatory myeloid cells are CD45 + , HLA-DR + , CD14 + , CD11b + , and CD11c + consist of cells that are... In some embodiments, the non-stimulatory myeloid cells are CD45 + , HLA-DR + , CD14 + , CD11b + , and CD11c + consist essentially of cells that are...

[0215] In some embodiments, the non-stimulatory myeloid cells are CD45 + , HLA-DR + , CD14 + , BDCA3 - , CD11b + , and CD11c+ It is. In some embodiments, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 + , BDCA3 - , CD11b + , and CD11c + cells. In some embodiments, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 + , BDCA3 - , CD11b + , and CD11c + cells. In some embodiments, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 + , BDCA3 - , CD11b + , and CD11c + consisting essentially of cells.

[0216] In some embodiments, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 - , CD11c + , and BDCA3 + are not. In some embodiments, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 - , CD11c + , and BDCA3 + cells not including.

[0217] In some embodiments, for example, in mice, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 + , CD11b high , and CD11c low are. In some embodiments, for example, in mice, the non-stimulatory bone marrow cells are CD45 + , HLA-DR + , CD14 + , CD11bhigh and CD11c low comprises cells that are. In some embodiments, for example, in mice, non-stimulatory bone marrow cells are CD45 + HLA-DR + CD14 + CD11b high and CD11c low consist of cells that are. In some embodiments, for example, in mice, non-stimulatory bone marrow cells are CD45 + HLA-DR + CD14 + CD11b high and CD11c low consist essentially of cells that are. In such embodiments, non-stimulatory mouse bone marrow cells are contacted with a TREM2 antibody.

[0218] In some embodiments, for example, in mice, non-stimulatory bone marrow cells are CD45 + HLA-DR + CD14 + CD11b low and CD11c high are. In some embodiments, for example, in mice, non-stimulatory bone marrow cells are CD45 + HLA-DR + CD14 + CD11b low and CD11c high comprise cells that are. In some embodiments, for example, in mice, non-stimulatory bone marrow cells are CD45 + HLA-DR + CD14 + CD11b low and CD11c high consist of cells that are. In some embodiments, for example, in mice, non-stimulatory bone marrow cells are CD45 + HLA-DR + CD14 + CD11b low and CD11c high consist essentially of cells that are. In such embodiments, non-stimulatory mouse bone marrow cells are contacted with a TREM2 antibody.

[0219] In some embodiments, the non-stimulatory bone marrow cells are in the cancer tissue.

[0220] In some embodiments, the population of immune cells is in the cancer tissue.

[0221] In some embodiments, the non-stimulatory cells and the stimulatory bone marrow cells are in the cancer tissue.

[0222] In some embodiments, the biological sample comprises a population of immune cells comprising non-stimulatory bone marrow cells and stimulatory bone marrow cells.

[0223] NSM cells can generically refer to DC1, TAM1, and TAM2 cells that are present in tumor tissue and can be distinguished from other cell types by the expression of NSM cell markers. For example, genes and related proteins that are more abundantly expressed or translated in NSM cells than in SDC can act as NSM markers. An exemplary NSM marker is CD11b. Additional exemplary NSM markers are shown in Table A. NSM cells can express TREM2, MS4A7, C5AR1, LYVE1, ABCC3, LILRB4, MRC1 / CD206, SIGLEC1, STAB1, TMEM37, MERTK, and TMEM119 on the cell surface. In some aspects, NSM cells do not express at least one of KIT, CCR7, BATF3, FLT3, ZBTB46, IRF8, BTLA, MYCL1, CLEC9A, BDCA3, and XCR1.

[0224] In one embodiment, NSM cells express one or more of the NSM marker genes listed in Table A. In another embodiment, NSM cells express 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or more of the NSM markers listed in Table D. In another embodiment, NSM cells express most or all of the NSM markers listed in Table D. In another embodiment, NSM cells are identified as expressing MRC1, MS4A7, C1QC, APOE, C1QB, C1QA, and C5AR1.

[0225] (Table D) TIFF2025109867000005.tif78128

[0226] Stimulatory bone marrow cells As used herein, stimulatory bone marrow cells (also referred to as SDC in certain embodiments) are bone marrow cells effective to stimulate an immune response (e.g., more effective to stimulate an anti-tumor response in the tumor microenvironment as compared to non-stimulatory bone marrow cells). In some embodiments, stimulatory bone marrow cells are effective to present an antigen (e.g., a tumor antigen) to T cells or to stimulate a tumor-specific T cell response as compared to non-stimulatory bone marrow cells. In some embodiments, stimulatory bone marrow cells may exhibit an increase in the ability to uptake, process, and / or present tumor-associated antigens to T cells as compared to non-stimulatory bone marrow cells. Stimulatory bone marrow cells may re-prime cytotoxic T lymphocytes or, in some cases, have an increased ability to stimulate effective tumor cell killing as compared to non-stimulatory bone marrow cells. Stimulatory bone marrow cells may exhibit higher expression of genes and cell surface markers involved in antigen processing, antigen presentation, and / or antigen co-stimulation, including but not limited to CD80, CD86, MHC I, and MHC II, as compared to non-stimulatory bone marrow cells.

[0227] Exemplary stimulatory bone marrow cell markers are listed in Table A. For example, in human SDC, the expression of Xcr1, Clec9a, and BDCA3 (CD141) are markers for SDC identification. In mice, CD103 can also be used as a strong marker of SDC identity, although it will be noted that it is not expressed in human SDC.

[0228] In one embodiment, the SDC is a tumor-infiltrating myeloid cell that has dendritic cell identity and expresses one or more of the SDC markers listed in Table A. In another embodiment, the SDC is a tumor-infiltrating myeloid cell that has dendritic cell identity and expresses 2, 3, 4, 5, 6, 7, 8, 9, or all of the SDC markers listed in Table A. In another embodiment, the SDC is identified as a tumor-infiltrating myeloid dendritic cell that expresses BDCA3, KIT, CCR7, BATF3, FLT3, ZBTB46, IRF8, BTLA, MYCL1, XCR1, and CLEC9A. Cells of the SDC may express at least one of KIT, CCR7, BATF3, FLT3, ZBTB46, IRF8, BTLA, MYCL1, CLEC9A, BDCA3, and XCR1. In some embodiments, the SDC does not substantially express TREM2, MS4A7, C5AR1, LYVE1, ABCC3, LILRB4, MRC1 / CD206, SIGLEC1, STAB1, TMEM37, MERTK, and / or TMEM119 on the cell surface. In some embodiments, the SDC does not substantially express C5AR1, LYVE1, ABCC3, MRC1, SIGLEC1, STAB1, C1QB, C1QA, TMEM37, MERTK, C1QC, TMEM119, MS4A7, APOE, CYP4F18, TREM2, TLR7, and / or LILRB4. Flow cytometry and PCR can be used to evaluate the expression of the markers disclosed herein among other technically acceptable assays.

[0229] The stimulated myeloid cells are CD45 + , HLA-DR + , CD14 - , CD11c + , and BDCA3 + and may be. The stimulated myeloid cells are CD45 + , HLA-DR + , and BDCA3 + and may be. The stimulated myeloid cells are CD45 + , HLA-DR + , CD14 - , and BDCA3 + and may be. The stimulated myeloid cells are CD45+ , HLA-DR + , CD11c + , and BDCA3 + may be.

[0230] Proteins, nucleotides, and homologs Methods and compositions are provided herein for inactivating and / or detecting non-stimulatory human bone marrow cells that express NSM protein. In some embodiments, the invention relates to inactivating and / or detecting non-stimulatory bone marrow cells from non-human mammalian cells that express an NSM protein homolog. For example, the mouse NSM protein may express a restricted expression pattern equivalent to that of the human homolog. Accordingly, in one embodiment, methods and compositions are provided herein for inactivating and / or detecting non-stimulatory mouse bone marrow cells that express NSM protein. Similar methods and compositions are also provided herein for inactivating and / or detecting non-stimulatory cells from any individual that express an NSM protein homolog having a similar expression pattern, the cells of which exhibit an expression pattern equivalent to that of the NSM protein.

[0231] The NSM protein or nucleotide may comprise at least one or more of C5AR1, LYVE1, ABCC3, MRC1, SIGLEC1, STAB1, C1QB, C1QA, TMEM37, MERTK, C1QC, TMEM119, MS4A7, APOE, CYP4F18, TREM2, TLR7, and LILRB4, and homologs thereof. The SDC protein or nucleotide may comprise at least one or more of KIT, CCR7, BATF3, FLT3, ZBTB46, IRF8, BTLA, MYCL1, CLEC9A, BDCA3, and XCR1, and homologs thereof. The cell surface NSM protein may comprise at least one or more of TREM2, MS4A7, C5AR1, LYVE1, ABCC3, LILRB4, MRC1 / CD206, SIGLEC1, STAB1, TMEM37, MERTK, and TMEM119. The cell surface NSM protein can be targeted with one or more anti-TREM2 antibodies alone or in combination. Generally, NSM is positive for the NSM protein or nucleotide and negative for the SDC protein or nucleotide, and conversely, SDC is generally positive for the SDC protein or nucleotide and negative for the NSM protein or nucleotide.

[0232] The antibodies described herein comprise at least one polypeptide, but they typically comprise an HC / LC dimer, i.e., four polypeptides. Polynucleotides encoding the polypeptides described herein are also described. Antibodies are typically isolated.

[0233] As used herein, "isolated" means a drug (e.g., a polypeptide or polynucleotide) that has been identified and separated and / or recovered from the components of its natural cell culture environment. Contaminating components of the natural environment are substances that interfere with the diagnostic or therapeutic use of the antibody and may include enzymes, hormones, and other proteinaceous or non-proteinaceous solutes. Isolated also refers to a drug that has been synthetically produced, for example, by human intervention.

[0234] The terms "polypeptide", "peptide", and "protein" are used interchangeably herein to refer to polymers of amino acid residues. That is, descriptions of polypeptides apply equally to descriptions of peptides and descriptions of proteins, and vice versa. The terms apply to both naturally occurring amino acid polymers and amino acid polymers in which one or more amino acid residues are amino acids encoded non-naturally. As used herein, the terms encompass amino acid chains of any length, including full-length proteins, in which the amino acid residues are linked by covalent peptide bonds.

[0235] The term "amino acid" refers to naturally occurring amino acids and non-naturally occurring amino acids, as well as amino acid analogs and mimetics that function in a manner similar to naturally occurring amino acids. Naturally encoded amino acids are the 20 common amino acids (alanine, arginine, asparagine, aspartic acid, cysteine, glutamine, glutamic acid, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, and valine) as well as pyrrolysine and selenocysteine. Amino acid analogs are compounds that have the same basic chemical structure as naturally occurring amino acids, i.e., hydrogen, a carboxyl group, an amino group, and an R group, e.g., carbon bonded to homoserine, norleucine, methionine sulfoxide, methionine methyl sulfonium. Such analogs have a modified R group (e.g., norleucine) or a modified peptide backbone, but retain the same basic chemical structure as naturally occurring amino acids. References to amino acids include, for example, naturally occurring proteinogenic L-amino acids; chemically modified amino acids such as D-amino acids, amino acid variants and derivatives; naturally occurring non-proteinogenic amino acids such as β-alanine, ornithine; and chemically synthesized compounds having properties known in the art that are characteristic of amino acids. Examples of non-naturally occurring amino acids include, but are not limited to, α-methyl amino acids (e.g., α-methyl alanine), D-amino acids, histidine-like amino acids (e.g., 2-amino-histidine, β-hydroxy-histidine, homohistidine), amino acids having additional methylene in the side chain ("homo" amino acids), and amino acids in which the carboxylic acid functional group of the side chain is replaced by a sulfonic acid group (e.g., cysteic acid). The incorporation of non-naturally occurring amino acids, including synthetic non-natural amino acids, substituted amino acids, or one or more D-amino acids, into the proteins of the present invention can be advantageous in a number of different ways. D-amino acid-containing peptides, for example, exhibit increased stability in vitro or in vivo compared to their L-amino acid-containing counterparts. Thus, constructs of peptides incorporating D-amino acids can be particularly useful when higher intracellular stability is required or desired.More specifically, D-peptides and the like exhibit resistance to endogenous peptidases and proteases, thereby providing improved biological availability of the molecule and extended in vivo lifetimes when such properties are required. Furthermore, D-peptides and the like cannot be efficiently processed for major histocompatibility complex class II-restricted presentation to helper T cells and, therefore, are less likely to induce a humoral immune response in the whole organism.

[0236] Amino acids may be referred to herein by either the commonly known three-letter symbols or the one-letter symbols recommended by the IUPAC-IUB Biochemical Nomenclature Commission. Similarly, nucleotides may be referred to by the generally recognized one-letter codes.

[0237] The present invention also encompasses polynucleotides encoding the polypeptides of the antibodies. The term "polynucleotide" or "nucleotide sequence" is intended to denote a continuous stretch of two or more nucleotide molecules. The nucleotide sequence can be of genomic, cDNA, RNA, semi-synthetic or synthetic origin, or any combination thereof.

[0238] The term "nucleic acid" refers to deoxyribonucleotides, deoxyribonucleosides, ribonucleosides, or ribonucleotides, and polymers thereof in either single-stranded or double-stranded form. Unless otherwise specifically limited, the term includes nucleic acids containing known analogs of natural nucleotides that have binding properties similar to the reference nucleic acid and are metabolized in a manner similar to naturally occurring nucleotides. Unless otherwise specifically limited, the term also refers to oligonucleotide analogs including PNA (peptide nucleic acid), and analogs of DNA used in antisense technology (such as phosphorothioates, phosphoramidates, etc.). Unless otherwise specified, a particular nucleic acid sequence implicitly encompasses conservatively modified variants thereof (including, but not limited to, degenerate codon substitutions), and complementary sequences, as well as the explicitly shown sequences. Specifically, degenerate codon substitutions may be achieved by generating sequences in which the third position of one or more selected (or all) codons is substituted with a mixture of bases and / or deoxyinosine residues (Batzer et al., Nucleic Acid Res. 19:5081 (1991); Ohtsuka et al., J. Biol. Chem. 260:2605-2608 (1985); Rossolini et al., Mol. Cell. Probes 8:91-98 (1994)).

[0239] "Conservatively modified variants" applies to both amino acid and nucleic acid sequences. With respect to a particular nucleic acid sequence, "conservatively modified variants" refers to nucleic acids that encode the same or essentially the same amino acid sequence or, when the nucleic acid does not encode an amino acid sequence, to essentially the same sequence. Because of the degeneracy of the genetic code, a large number of functionally identical nucleic acids encode any given protein. For example, the codons GCA, GCC, GCG, and GCU all code for the amino acid alanine. Thus, at every position where an alanine is specified by a codon, the codon can be modified to any of the corresponding codons described without changing the encoded polypeptide. Such nucleic acid variations are "silent mutations" and represent one species of conservatively modified change. Every nucleic acid sequence herein that encodes a polypeptide also describes every possible silent mutation of the nucleic acid. One of ordinary skill in the art will recognize that each codon in a nucleic acid (except for AUG, which is ordinarily the only codon for methionine, and TGG, which is ordinarily the only codon for tryptophan) can be modified to yield a functionally identical molecule. Thus, each silent mutation of a nucleic acid that encodes a polypeptide is implicit in each described sequence.

[0240] With respect to amino acid sequences, one of ordinary skill in the art will recognize that individual substitutions, deletions, or additions to a nucleic acid, peptide, polypeptide, or protein sequence which alter, add, or delete a single amino acid or a small percentage of amino acids in the encoded sequence are "conservatively modified variants" where the alteration results in the deletion of an amino acid, the addition of an amino acid, or the substitution of an amino acid with a chemically similar amino acid. Tables of conserved substitutions providing functionally similar amino acids are known to those of ordinary skill in the art. Such conservatively modified variants are in addition to and do not exclude polymorphic variants, interspecies homologs, and alleles herein described.

[0241] Tables of conservative substitutions providing functionally similar amino acids are known to those of ordinary skill in the art. The following eight groups contain amino acids that are conservative substitutions for one another: 1) alanine (A), glycine (G); 2) aspartic acid (D), glutamic acid (E); 3) asparagine (N), glutamine (Q); 4) arginine (R), lysine (K); 5) isoleucine (I), leucine (L), methionine (M), valine (V); 6) phenylalanine (F), tyrosine (Y), tryptophan (W); 7) serine (S), threonine (T); and 8) cysteine (C), methionine (M) (see, e.g., Creighton, Proteins: Structures and Molecular Properties (W H Freeman & Co.; 2nd edition (December 1993))).

[0242] The terms "identical" or percent "identity" in the context of two or more nucleic acid or polypeptide sequences refer to two or more sequences or subsequences that are the same. When aligned to obtain maximal correspondence over a comparison window, or designated region as measured using the following sequence comparison algorithms (or other algorithms available to those of skill in the art), or by manual alignment and visual inspection, the sequences are "substantially identical" if they have an amino acid residue or nucleotide percentage that is the same (i.e., about 60% identity, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, or about 95% identity over the designated region). Alignment for purposes of determining percent amino acid sequence identity can be achieved in various ways within the skill in the art, e.g., using publicly available computer software such as BLAST, BLAST-2, ALIGN, MEGALIGN (DNASTAR), CLUSTALW, CLUSTAL OMEGA, or MUSCLE software. This definition also refers to the complement of a test sequence. Identity can exist over a region of at least about 50 amino acids or nucleotides in length, or over a region of 75 - 100 amino acids or nucleotides in length, or over the full length of a polynucleotide or polypeptide if so specified. Polynucleotides encoding the polypeptides of the invention, including homologs from species other than human, may be obtained by a process that includes screening a library using a labeled probe having the polynucleotide sequence described herein or a fragment thereof under stringent hybridization conditions, and isolating full-length cDNA and genomic clones containing the polynucleotide sequence. Such hybridization techniques are well known to those of skill in the art.

[0243] For array comparison, usually one array against which a test array is compared functions as a reference array. When using an array comparison algorithm, the test array and the reference array are input into a computer, and optionally, subsequence coordinates are specified and array algorithm program parameters are specified. Default program parameters can be used or alternative parameters can be specified. Next, the array comparison algorithm calculates the percent sequence identity for the test array compared to the reference array based on the program parameters.

[0244] As used herein, a "comparison window" includes reference to any one segment of a number of contiguous positions consisting of from 20 to 600, usually about 50 to about 200, more usually about 100 to about 150, which can be compared to a reference sequence of the same number of contiguous positions after two sequences are optimally aligned. Methods of aligning sequences for comparison are known to those of skill in the art. Without limitation, those by the local homology algorithm of Smith and Waterman (1970) Adv. Appl. Math. 2:482c, by the homology alignment algorithm of Needleman and Wunsch (1970) J. Mol. Biol. 48:443, by the method of searching for similarity of Pearson and Lipman (1988) Proc. Nat'l. Acad. Sci. USA 85:2444, by computerized implementations of these algorithms (GAP, BESTFIT, FASTA, and TFASTA in the Wisconsin Genetics Software Package, Genetics Computer Group (575 Science Drive, Madison, Wisconsin)), or by manual alignment and visual inspection (see, e.g., Ausubel et al., Current Protocols in Molecular Biology (1995 supplement)) can perform an optimal alignment of sequences for comparison.

[0245] An example of an algorithm suitable for determining percent sequence identity and sequence similarity is the BLAST and BLAST 2.0 algorithms, which are described in Altschul et al. (1997) Nuc. Acids Res. 25:3389-3402 and Altschul et al. (1990) J. Mol. Biol. 215:403-410, respectively. Software for performing BLAST analyses is publicly available through the National Center for Biotechnology Information, which can be found on the World Wide Web at ncbi.nlm.nih.gov. The BLAST algorithm parameters W, T, and X determine the sensitivity and speed of the alignment. The BLASTN program (for nucleotide sequences) uses, by default, a word length (W) of 11, an expectation (E) of 10, or M = 5, N = -4, and a comparison of both strands. For amino acid sequences, the BLASTP program uses, by default, a word length of 3, and an expectation (E) of 10, and the BLOSUM62 scoring matrix uses an alignment (B) of 50 (see Henikoff and Henikoff (1992) Proc. Natl. Acad. Sci. USA 89:10915), an expectation (E) of 10, M = 5, N = -4, and a comparison of both strands. The BLAST algorithm is typically performed with the "low complexity" filter turned off.

[0246] The BLAST algorithm also performs a statistical analysis of the similarity between two sequences (see, e.g., Karlin and Altschul (1993) Proc. Natl. Acad. Sci. USA 90:5873-5787). One measure of similarity provided by the BLAST algorithm is the minimum total probability (P(N)), which provides an indication of the probability that a match between two nucleotide or amino acid sequences occurred by chance. For example, a nucleic acid is considered similar to a reference sequence if the minimum total probability in a comparison of the test nucleic acid to the reference nucleic acid is less than about 0.2, or less than about 0.01, or less than about 0.001.

[0247] The phrase "selectively (or specifically) hybridizes" refers to the binding, duplexing, or hybridization of a molecule to only a particular nucleotide sequence under stringent hybridization conditions when the sequence is present in a complex mixture (including, but not limited to, cells or library DNA or RNA).

[0248] The phrase "stringent hybridization conditions" refers to the hybridization of DNA, RNA, or other nucleic acids, or combinations thereof, under conditions of low ionic strength and high temperature, as known in the art. Generally, under stringent conditions, a probe will hybridize to the target subsequence of a complex mixture of nucleic acids (including, but not limited to, total cellular or library DNA or RNA), but not to other sequences in the complex mixture. Stringent conditions are sequence-dependent and will be different in different circumstances. Longer sequences will hybridize particularly at high temperatures. A general guide to nucleic acid hybridization is found in Tijssen, Laboratory Techniques in Biochemistry and Molecular Biology--Hybridization with Nucleic Probes, “Overview of principles of hybridization and the strategy of nucleic acid assays” (1993).

[0249] As used herein, the terms "engineer, engineered, engineering" are considered to include any manipulation of the peptide backbone, or post-translational modification of a naturally occurring or recombinant polypeptide or fragment thereof. Engineering includes changes in the amino acid sequence, changes in the glycosylation pattern, or changes in the side chain groups of individual amino acids, and combinations of these approaches. Engineered proteins are expressed and produced by standard molecular biology techniques.

[0250] "Isolated nucleic acid molecule or polynucleotide" means a nucleic acid molecule, DNA, or RNA that has been removed from its natural environment. For example, a recombinant polynucleotide encoding a polypeptide contained in a vector is considered to be isolated. Further examples of isolated polynucleotides include recombinant polynucleotides maintained within a polynucleotide that has been (partially or substantially) purified in a heterologous host cell or in solution. Isolated polynucleotides include polynucleotide molecules contained in a cell that normally contains the polynucleotide molecule, but the polynucleotide molecule is present extrachromosomally or at a chromosomal location different from its natural chromosomal location. Isolated RNA molecules include in vivo or in vitro RNA transcripts, as well as positive and negative strand forms, and double-stranded forms. The isolated polynucleotides or nucleic acids described herein further include molecules that are synthetically produced, for example, via PCR or chemical synthesis. In addition, the polynucleotide or nucleic acid, in certain embodiments, includes control elements such as a promoter, ribosome binding site, or transcription terminator.

[0251] The term "polymerase chain reaction" or "PCR" generally refers to a method for amplifying a desired nucleotide sequence in vitro, for example, as described in U.S. Patent No. 4,683,195. Generally, the PCR method involves repeated cycles of primer extension synthesis using oligonucleotide primers capable of preferentially hybridizing to a template nucleic acid.

[0252] A polynucleotide nucleotide sequence is intended to be identical to a reference nucleotide sequence, except that the polynucleotide nucleotide sequence may contain up to 5 point mutations per 100 nucleotides of the reference nucleotide sequence, with a nucleic acid or polynucleotide having a nucleotide sequence that is at least, for example, 95% "identical" to the reference nucleotide sequence of the present invention. In other words, to obtain a polynucleotide having a nucleotide sequence that is at least 95% identical to the reference nucleotide sequence, up to 5% of the nucleotides in the reference sequence may be deleted or replaced with another nucleotide, or up to 5% of the total number of nucleotides in the reference sequence may be inserted into the reference sequence. These changes to the reference sequence can occur between the residues of the reference sequence, at the 5' or 3' ends of the reference nucleotide sequence that are interspersed among one or more adjacent groups within the reference sequence, or anywhere between those terminal positions. As a practical matter, whether a particular polynucleotide sequence is at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to the nucleotide sequence of the present invention can usually be determined using known computer programs such as those discussed above for polypeptides (e.g., ALIGN-2).

[0253] A derivative or variant of a polypeptide is said to "share homology" with, or be "homologous to", the peptide if the amino acid sequence of the derivative or variant has at least 50% identity with a 100 - amino acid sequence derived from the original peptide. In certain embodiments, the derivative or variant is at least 75% identical to either a peptide or a fragment of a peptide having the same number of amino acid residues as the derivative. In certain embodiments, the derivative or variant is at least 85% identical to either a peptide or a fragment of a peptide having the same number of amino acid residues as the derivative. In certain embodiments, the amino acid sequence of the derivative is at least 90% identical to a peptide or a fragment of a peptide having the same number of amino acid residues as the derivative. In some embodiments, the amino acid sequence of the derivative is at least 95% identical to a peptide or a fragment of a peptide having the same number of amino acid residues as the derivative. In certain embodiments, the derivative or variant is at least 99% identical to either a peptide or a fragment of a peptide having the same number of amino acid residues as the derivative.

[0254] As used herein, the term "modified" refers to any change made to a given polypeptide, such as a change in the length, amino acid sequence, chemical structure, co - translational modification, or post - translational modification of the polypeptide. The form "(modified)" means that the polypeptide under consideration may be optionally modified, i.e., the polypeptide under consideration may be either modified or unmodified.

[0255] In some embodiments, a polypeptide comprises an amino acid sequence that is at least 80, 85, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to a relevant (e.g., polypeptide and / or antibody) amino acid sequence or a fragment thereof described in a table(s) or accession number(s) disclosed herein. In some embodiments, an isolated antibody or protein disclosed herein comprises an amino acid sequence encoded by a polynucleotide that is at least 80, 85, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to a relevant nucleotide sequence or a fragment thereof described in a table(s) or accession number(s) disclosed herein. In some embodiments, a nucleotide sequence comprises a nucleotide sequence that is at least 80, 85, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% identical to a nucleotide sequence disclosed herein as described in a table(s) or accession number(s) disclosed herein.

[0256] Pharmaceutical composition The present application provides a composition comprising an antibody, which comprises one or more of the antibodies described herein, together with one or more pharmaceutically acceptable excipients. In some embodiments, the composition is sterile. A pharmaceutical composition generally comprises an effective amount of an antibody.

[0257] These compositions may include, in addition to one or more of the antibodies disclosed herein, pharmaceutically acceptable excipients, carriers, buffers, stabilizers, or other substances well known to those of skill in the art. Such substances should be non-toxic and should not interfere with the effectiveness of the active ingredient. The exact nature of the carrier or other substance may depend on the route of administration, e.g., oral, intravenous, cutaneous or subcutaneous, nasal, intramuscular, intraperitoneal routes.

[0258] Pharmaceutical compositions for oral administration can be in the form of tablets, capsules, powders, or liquids. Tablets can contain solid carriers such as gelatin or adjuvants. Liquid pharmaceutical compositions generally contain liquid carriers such as water, petroleum, animal or vegetable oils, mineral oils, or synthetic oils. They can include physiological saline, dextrose or other sugar solutions, or glycols such as ethylene glycol, propylene glycol, or polyethylene glycol.

[0259] For intravenous, cutaneous or subcutaneous injection, or injection at the site of affliction, the active ingredient will be in the form of a parenterally acceptable aqueous solution that is free of pyrogenic substances and has suitable pH, isotonicity, and stability. Those skilled in the art can adequately prepare suitable solutions using isotonic vehicles such as, for example, sodium chloride injection solution, Ringer's injection solution, lactated Ringer's injection solution, etc. Optionally, preservatives, stabilizers, buffers, antioxidants, and / or other additives can be included.

[0260] Regardless of whether it is a polypeptide, antibody (e.g., anti-TREM2 antibody), nucleic acid, small molecule, or other pharmaceutically useful compound to be administered to an individual, the administration is preferably at a "therapeutically effective amount" or "prophylactically effective amount" (in some cases, it may be therapeutic, but prophylaxis can be considered as treatment), which is sufficient to show benefit to the individual. The actual amount administered, as well as the rate and course of administration, will depend on the nature and severity of the protein aggregation disease being treated. The formulation of the treatment, such as determination of the dosage, etc., is within the scope of the responsibility of general practitioners and other physicians and usually takes into account the disorder to be treated, the condition of the individual subject, the site of delivery, the method of administration, and other factors known to the practitioner. Examples of the above techniques and protocols can be found in Remington’s Pharmaceutical Sciences, 16th edition, Osol, A. (ed), 1980.

[0261] The composition can be administered alone or in combination with other treatments, simultaneously or sequentially, depending on the condition to be treated.

[0262] Method Preparation method The antibodies described herein can be produced using recombinant methods and compositions, such as those described in U.S. Patent No. 4,816,567, for example.

[0263] In one embodiment, an isolated nucleic acid encoding an antibody described herein is provided. Such a nucleic acid may encode an amino acid sequence comprising a VL of the antibody and / or an amino acid sequence comprising a VH (e.g., a light chain and / or a heavy chain of the antibody) or an amino acid sequence comprising a VHH of a single domain antibody. In further embodiments, one or more vectors (e.g., expression vectors) comprising such a nucleic acid are provided. In one embodiment, the nucleic acid is provided in a multicistronic vector. In further embodiments, a host cell comprising such a nucleic acid is provided. In such an embodiment, the host cell comprises (1) a vector comprising a nucleic acid encoding an amino acid sequence comprising a VL of the antibody and an amino acid sequence comprising a VH of an antigen-binding polypeptide construct, or (2) a first vector comprising a nucleic acid encoding an amino acid sequence comprising a VL of an antigen-binding polypeptide construct and a second vector comprising a nucleic acid encoding an amino acid sequence comprising a VH of an antigen-binding polypeptide construct (e.g., is transformed). In one embodiment, the host cell is a eukaryotic cell, such as a Chinese hamster ovary (CHO) cell, or a human embryonic kidney (HEK) cell, or a lymphoid cell (e.g., Y0, NS0, Sp20 cells). In one embodiment, a method of making an antibody is provided, the method comprising culturing a host cell comprising a nucleic acid encoding the antibody, as provided above, under conditions suitable for expression of the antibody, and optionally recovering the antibody from the host cell (or host cell culture medium).

[0264] In the case of recombinant production of antibodies, for example, the nucleic acid encoding the antibody as described above is isolated and inserted into one or more vectors for further cloning and / or expression in a host cell. Such nucleic acids may be readily isolated and sequenced using conventional procedures (e.g., by using oligonucleotide probes capable of specifically binding to the genes encoding the heavy and light chains of the antibody).

[0265] The term "substantially purified" refers to constructs or variants thereof described herein that may be substantially or essentially free of components normally associated with or interacting with the protein as found in its natural environment, i.e., in the case of a recombinantly produced heteromultimer that is substantially free of extracellular substances in a natural cell or in a particular embodiment, the host cell has less than about 30%, less than about 25%, less than about 20%, less than about 15%, less than about 10%, less than about 5%, less than about 4%, less than about 3%, less than about 2%, or less than about 1% (by dry weight) of contaminating protein, including preparations of protein. When a heteromultimer or a variant thereof is recombinantly produced by a host cell, the protein in a particular embodiment is present at about 30%, about 25%, about 20%, about 15%, about 10%, about 5%, about 4%, about 3%, about 2%, or about 1% or less of the dry weight of the cell. When a heteromultimer or a variant thereof is recombinantly produced by a host cell, the protein in a particular embodiment is present in the medium at about 5 g / L, about 4 g / L, about 3 g / L, about 2 g / L, about 1 g / L, about 750 mg / L, about 500 mg / L, about 250 mg / L, about 100 mg / L, about 50 mg / L, about 10 mg / L, or about 1 mg / L or less of the dry weight of the cell. In a particular embodiment, a "substantially purified" heteromultimer produced by the methods described herein has a purification level of at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, specifically at least about 75%, 80%, 85%, more specifically at least about 90%, at least about 95%, at least about 99%, or more, as determined by suitable methods such as SDS / PAGE analysis, RP-HPLC, SEC, and capillary electrophoresis.

[0266] Host cells suitable for cloning or expressing vectors encoding antibodies include prokaryotic or eukaryotic cells as described herein.

[0267] "Recombinant host cell" or "host cell" refers to a cell containing an exogenous polynucleotide, regardless of the method used for insertion, e.g., direct uptake, transduction, f-mating, or other methods known in the art for creating recombinant host cells. The exogenous polynucleotide may be maintained as a non-integrated vector, e.g., a plasmid, or integrated into the host genome. Host cells may include CHO, derivatives of CHO, NS0, Sp2O, CV-1, VERO-76, HeLa, HepG2, Per.C6, or BHK.

[0268] As used herein, the term "eukaryote" refers to organisms belonging to the phylogenetic domain Eucarya, e.g., animals (including, but not limited to, mammals, insects, reptiles, birds, etc.), ciliates, plants (including, but not limited to, monocots, dicots, algae, etc.), fungi, yeast, flagellates, microsporidia, protists, etc.

[0269] As used herein, the term "prokaryote" refers to prokaryotic organisms. For example, non-eukaryotes may belong to the Eubacteria (including, but not limited to, Escherichia coli, Thermus thermophilus, Bacillus stearothermophilus, Pseudomonas fluorescens, Pseudomonas aeruginosa, Pseudomonas putida, etc.) phylogenetic domain, or the Archaea (including, but not limited to, Methanococcus jannaschii, Methanobacterium thermoautotrophicum, Halobacterium, e.g., Haloferax volcanii and Halobacterium species NRC-1, Archaeoglobus fulgidus, Pyrococcus furiosus, Pyrococcus horikoshii, Aeuropyrum pernix, etc.) phylogenetic domain.

[0270] For example, antibodies may be produced in bacteria, particularly when glycosylation and Fc effector functions are not required. For the expression of antibody fragments and polypeptides in bacteria, see, for example, U.S. Patent Nos. 5,648,237, 5,789,199, and 5,840,523, and also Charlton, Methods in Molecular Biology, Vol. 248 (B.K.C. Lo, ed., Humana Press, Totowa, N.J., 2003), pp. 245-254, which describes the expression of antibody fragments in E. coli. After expression, the antibody may be isolated from the bacterial cell paste in the soluble fraction and further purified.

[0271] In addition to prokaryotes, eukaryotic microorganisms such as filamentous fungi or yeast are suitable cloning or expression hosts for vectors encoding antibodies, including fungal and yeast lineages, and this glycosylation pathway is "humanized" and results in the production of antibodies that partially or fully contain the human glycosylation pattern. See Gerngross, Nat. Biotech. 22:1409-1414 (2004), and Li et al., Nat. Biotech. 24:210-215 (2006).

[0272] Host cells suitable for the expression of glycosylated antibodies are also derived from multicellular organisms (invertebrates and vertebrates). Examples of invertebrate cells include plant and insect cells. A number of baculovirus strains have been identified that can be used with insect cells, particularly Spodoptera frugiperda cells in the case of transfection.

[0273] Plant cell cultures can also be utilized as hosts. See, for example, U.S. Patent Nos. 5,959,177, 6,040,498, 6,420,548, 7,125,978, and 6,417,429, which describe the PLANTIBODIES™ technology for producing antibodies in transgenic plants.

[0274] Vertebrate cells may also be used as hosts. For example, mammalian cell lines adapted to grow in suspension may be useful. Other examples of useful mammalian host cell lines are monkey kidney CV1 line transformed by SV40 (COS-7); human embryonic kidney lines (e.g., 293 or 293 cells as described in Graham et al., J. Gen Virol. 36:59 (1977)); baby hamster kidney cells (BHK); mouse Sertoli cells (e.g., TM4 cells as described in Mather, Biol. Reprod. 23:243-251 (1980)); monkey kidney cells (CV1); African green monkey kidney cells (VERO-76); human cervical cancer cells (HELA); dog kidney cells (MDCK); buffalo rat liver cells (BRL 3A); human lung cells (W138); human liver cells (Hep G2); mouse mammary tumor (MMT 060562); TRI cells as described in, for example, Mather et al., Annals N.Y. Acad. Sci. 383:44-68 (1982); MRC5 cells; and FS4 cells. Other useful mammalian host cell lines include DHFR - Chinese hamster ovary (CHO) cells including DHFR CHO cells (Urlaub et al., Proc. Natl. Acad. Sci. USA 77:4216 (1980)); and myeloma cell lines such as Y0, NS0, and SP2 / 0. For a review of specific mammalian host cell lines suitable for antibody production, see, for example, Yazaki and Wu, Methods in Molecular Biology, Vol. 248 (B.K.C. Lo, ed., Humana Press, Totowa, N.J.), pp. 255-268 (2003).

[0275] In one embodiment, the antibodies described herein are produced in stable mammalian cells by a method comprising transfecting at least one stable mammalian cell with a nucleic acid encoding the antibody and expressing the nucleic acid in at least one mammalian cell at a predetermined ratio. In some embodiments, the predetermined ratio of nucleic acid is determined in transient transfection experiments to determine the relative ratio of input nucleic acid that results in the highest percentage of antibody in the expression product.

[0276] In some embodiments, there is a method of producing an antibody in a stable mammalian cell as described herein, wherein the expression product of at least one stable mammalian cell comprises a greater percentage of the desired glycosylated antibody compared to a monomeric heavy or light chain polypeptide or other antibody.

[0277] In some embodiments, there is a method of producing a glycosylated antibody in the stable mammalian cells described herein, the method comprising identifying and purifying the desired glycosylated antibody. In some embodiments, the identification is by one or both of liquid chromatography and mass spectrometry.

[0278] Optionally, the antibody can be purified or isolated after expression. The protein may be isolated or purified by a variety of methods known to those skilled in the art. Standard purification methods include chromatographic techniques such as ion exchange, hydrophobic interaction, affinity, size or gel filtration, and reverse phase, which are carried out at atmospheric or high pressure using systems such as FPLC and HPLC. Purification methods also include electrophoretic techniques, immunological techniques, precipitation techniques, dialysis techniques, and chromatofocusing techniques. Ultrafiltration and diafiltration techniques combined with protein concentration are also useful. As is well known in the art, various natural proteins bind to Fc and antibodies, and these proteins may find use in the present invention for the purification of antibodies. For example, bacterial protein A and G bind to the Fc region. Similarly, bacterial protein L binds to the Fab region of some antibodies. Purification can often be enabled by a specific fusion partner. For example, an antibody may be purified using glutathione resin if a GST fusion is used, Ni +2 affinity chromatography, or if a FLAG tag is used, it may be purified using an anti-FLAG antibody. For general guidance on suitable purification techniques, see, for example, Protein Purification: Principles and Practice, 3 rd Ed., Scopes, Springer-Verlag, NY, 1994, which is incorporated herein by reference in its entirety. The degree of purification required will vary depending on the use of the antibody. In some cases, purification may not be necessary.

[0279] In certain embodiments, the antibody is purified using anion exchange chromatography including, but not limited to, chromatography on Q-Sepharose, DEAE-Sepharose, Poros HQ, Poros DEAF, Toyopearl Q, Toyopearl QAE, Toyopearl DEAE, Resource / Source Q and DEAE, Fractogel Q and DEAE columns.

[0280] In certain embodiments, the proteins described herein are purified using cation exchange chromatography including, but not limited to, SP-Sepharose, CM-Sepharose, Poros HS, Poros CM, Toyopearl SP, Toyopearl CM, Resource / Source S and CM, Fractogel S and CM columns, and equivalents and the like thereof.

[0281] Furthermore, the antibodies described herein can be chemically synthesized using techniques known in the art (see, e.g., Creighton, 1983, Proteins: Structures and Molecular Principles, W.H. Freeman & Co., N.Y and Hunkapiller et al., Nature, 310:105-111 (1984)). For example, polypeptides corresponding to fragments of a polypeptide can be synthesized by use of a peptide synthesizer. Furthermore, non-classical amino acids or chemical amino acid analogs can be introduced, as substitutions or additions to the polypeptide sequence, as needed. Non-classical amino acids generally include, but are not limited to, designer amino acids such as D-isomers of common amino acids like 2,4-diaminobutyric acid, α-aminoisobutyric acid, 4-aminobutyric acid, Abu, 2-aminobutyric acid, g-Abu, e-Ahx, 6-aminohexanoic acid, Aib, 2-aminoisobutyric acid, 3-aminopropionic acid, ornithine, norleucine, norvaline, hydroxyproline, sarcosine, citrulline, homocitrulline, cysteic acid, t-butylglycine, t-butylalanine, phenylglycine, cyclohexylalanine, alanine, fluoroamino acids, methylamino acids, C-methylamino acids, N-methylamino acids, and amino acid analogs. Furthermore, the amino acids can be D (right-handed) or L (left-handed).

[0282] Method of Use In one aspect, the present application provides a method of contacting non-stimulatory bone marrow cells with an anti-TREM2 antibody such as a human antibody, thereby resulting in inactivation of the non-stimulatory bone marrow cells.

[0283] In another aspect, the present application provides a method of contacting non-stimulatory bone marrow cells with an anti-TREM2 mouse antibody, thereby resulting in the inactivation of the non-stimulatory bone marrow cells.

[0284] In some embodiments, the non-stimulatory cells are one or more of DC1 cells and TAM cells.

[0285] In some embodiments, the present application provides a method of inactivating non-stimulatory bone marrow cells, comprising contacting the non-stimulatory bone marrow cells with a TREM2 antibody, thereby killing the non-stimulatory bone marrow cells. Inactivation refers to rendering the cells partially or completely non-functional. In some embodiments, inactivation of the non-stimulatory bone marrow cells results in inducing growth arrest in the cells. In some embodiments, inactivation of the non-stimulatory bone marrow cells results in apoptosis of the cells. In some embodiments, inactivation of the non-stimulatory cells results in lysis of the cells, for example, by complement-dependent cytotoxicity (CDC) or antibody-dependent cell-mediated cytotoxicity (ADCC). In some embodiments, inactivation of the non-stimulatory bone marrow cells results in necrosis of the cells. In some embodiments, inactivation of the non-stimulatory bone marrow cells results in inducing growth arrest in the cells. In some embodiments, inactivation of the non-stimulatory bone marrow cells results in inactivation of the cells. In some embodiments, inactivation of the non-stimulatory bone marrow cells results in neutralization of the activity of the TREM2 protein within the cells. In some embodiments, inactivation of the non-stimulatory bone marrow cells results in reduction of cell proliferation. In some embodiments, inactivation of the non-stimulatory bone marrow cells results in differentiation of the cells. In some embodiments, inactivation of the non-stimulatory bone marrow cells results in a decrease in the ability of the cells to act as inhibitory antigen-presenting cells or an increase in the ability of the cells to act as activated antigen-presenting cells. In some embodiments, inactivation of the non-stimulatory bone marrow cells results in abnormal localization of the cells within the tumor tissue or tumor microenvironment (TME). In some embodiments, inactivation of the non-stimulatory bone marrow cells results in alteration of the spatial organization of the cells within the tumor tissue or tumor microenvironment. In some embodiments, inactivation of the non-stimulatory bone marrow cells results in alteration of the temporal expression of the cells within the tumor tissue or TME. In some embodiments, the method further comprises removing the non-stimulatory bone marrow cells.

[0286] In any and all aspects of inactivating the non-stimulatory bone marrow cells described herein, any increase or decrease or change in the aspect(s) of property(ies) or function(s) is as compared to cells not in contact with the anti-TREM2 antibody.

[0287] In another aspect, the present application provides a method of contacting non-stimulatory bone marrow cells with an anti-TREM2 antibody, which results in the modulation of the function of the non-stimulatory bone marrow cells. The modulation can be any one or more of the following. In some embodiments, the non-stimulatory cells are one or more of DC1 cells, TAM1 cells, and TAM2 cells. In some embodiments, the modulation of the function results in the inactivation of the non-stimulatory bone marrow cells. In some embodiments, the modulation of the function of the non-stimulatory bone marrow cells results in an increase in the ability of the cells to stimulate both naive and activated CD8+ T cells, for example, by increasing the ability of the non-stimulatory cells to cross-present tumor antigens on MHC I molecules to naive CD8+ T cells. In some embodiments, the modulation includes increasing the T cell-stimulating function of the non-stimulatory bone marrow cells, which can include, for example, inducing the ability of the cells to induce T cell receptor (TCR) signaling, T cell proliferation, or T cell cytokine production. In one embodiment, the survival of the non-stimulatory cells is decreased or the proliferation of the non-stimulatory cells is decreased. In one embodiment, the ratio of stimulatory bone marrow cells to non-stimulatory bone marrow cells is increased.

[0288] In any and all aspects of decreasing the function of the non-stimulatory bone marrow cells described herein, any increase or decrease or change in the aspect(s) of property(ies) or function(s) is as compared to cells not in contact with the TREM2 antibody.

[0289] In some embodiments, the present application provides a method of killing (also referred to as inducing cell death) non-stimulatory bone marrow cells, comprising contacting the non-stimulatory bone marrow cells with an anti-TREM2 antibody, thereby killing the non-stimulatory bone marrow cells. In some embodiments, the killing is increased compared to non-stimulatory bone marrow cells not contacted with the anti-TREM2 antibody. In some embodiments, the contacting induces apoptosis of the non-stimulatory bone marrow cells. In some embodiments, the contacting induces apoptosis of the non-stimulatory bone marrow cells. In some embodiments, the non-stimulatory bone marrow cells are in a population of immune cells comprising non-stimulatory bone marrow cells and stimulatory bone marrow cells. In some embodiments, the method further comprises removing the non-stimulatory bone marrow cells. In some embodiments, 10% to 80% of the cells are killed. In some embodiments, at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, or 80% of the cells are killed.

[0290] In some embodiments, the present application provides a method of increasing the ratio of stimulatory bone marrow cells to non-stimulatory bone marrow cells in a population of immune cells comprising stimulatory bone marrow cells and non-stimulatory bone marrow cells, comprising contacting the population of immune cells with an anti-TREM2 antibody. In some embodiments, the ratio is increased compared to a population of cells not contacted with the anti-TREM2 antibody. In some embodiments, the ratio of DC2 cells to DC1 cells is increased. In some embodiments, the ratio of DC2 cells to TAM1 cells is increased. In some embodiments, the ratio of DC2 cells to TAM2 cells is increased. In some embodiments, the ratio of DC2 cells to TAM1 + TAM2 cells is increased. In some embodiments, the ratio of DC2 cells to TAM1 + DC1 cells is increased. In some embodiments, the ratio of DC2 cells to DC1 + TAM2 cells is increased. In some embodiments, the ratio of DC2 cells to DC1 + TAM1 + TAM2 cells is increased. In some embodiments, at least the ratio is increased by 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100%.

[0291] In some embodiments, the ratio of stimulatory bone marrow cells to non-stimulatory bone marrow cells before contact is in the range of 0.001:1 to 0.1:1. In some embodiments, the ratio of stimulatory bone marrow cells to non-stimulatory bone marrow cells after contact is in the range of 0.1:1 to 100:1.

[0292] In some embodiments, the number of non-stimulatory bone marrow cells is reduced. In some embodiments, the stimulatory bone marrow cells are DC2 cells. In some embodiments, the non-stimulatory bone marrow cells are killed, for example, by necrosis or apoptosis. In some embodiments, the non-stimulatory bone marrow cells are induced to cause growth arrest. In some embodiments, the non-stimulatory bone marrow cells no longer proliferate. In some embodiments, the spatial localization of the non-stimulatory bone marrow cells is altered and the ratio increases in a specific region of the TME. In some embodiments, the transient expression of the non-stimulatory bone marrow cells is altered and the ratio increases during a specific time during tumor development.

[0293] In some embodiments, the contact is in vitro. In some embodiments, the contact is in vivo. In some specific embodiments, the contact is in human in vivo. In some embodiments, the contact is caused by administering an anti-TREM2 antibody. In some embodiments, the individual to whom the antibody (e.g., human) is administered has cancer.

[0294] In another aspect, the present invention provides a method of treating an immune-related condition (e.g., cancer) in an individual, comprising administering to the individual an effective amount of a composition comprising an anti-TREM2 antibody. In another aspect, the present invention provides a method of enhancing an immune response in an individual, comprising administering to the individual an effective amount of a composition comprising an anti-TREM2 antibody. In some embodiments, these methods further comprise other combination therapies, such as PDL blockade therapy, anti-PD-1 antibody, anti-PD-L1 antibody, anti-PD-L2 antibody, CTLA4 blockade therapy, anti-CTLA-4 antibody, systemic checkpoint blockade therapy in which inhibitory molecules of T cells are blocked, adoptive T cell therapy, CAR T cell therapy, dendritic cells, or other cell therapies, and are provided in combination with conventional chemotherapy.

[0295] In some embodiments, the method further includes determining the expression level of TREM2 protein in a biological sample derived from an individual. In some embodiments, the biological sample includes, but is not limited to, body fluids, tissue samples, organ samples, urine, feces, blood, saliva, CSF, and any combination thereof. In some embodiments, the biological sample is derived from tumor tissue. In some embodiments, the expression level includes the mRNA expression level of mRNA encoding the TREM2 protein. In some embodiments, the expression level of the TREM2 protein includes the protein expression level of NSM. In some embodiments, the expression level of the TREM2 protein is detected in the sample using a method selected from the group consisting of FACS, Western blot, ELISA, immunoprecipitation, immunohistochemistry, immunofluorescence, radioimmunoassay, dot blotting, immunoassay, HPLC, surface plasmon resonance, optical spectroscopy, mass spectrometry, HPLC, qPCR, RT-qPCR, multiplex qPCR or RT-qPCR, RNA-seq, microarray analysis, SAGE, MassARRAY technology, and FISH, and combinations thereof.

[0296] In another aspect, the present application provides a method for determining the presence or absence of general non-stimulatory myeloid cells, or for determining the presence or absence of specific non-stimulatory myeloid cells (e.g., DC1 cells, TAM1 cells, and / or TAM2 cells), including contacting a population of cells containing non-stimulatory myeloid cells with an anti-TREM2 antibody and quantifying the number of non-stimulatory myeloid cells. In another aspect, the present application provides a method for determining the presence or absence of non-stimulatory myeloid cells, including contacting a population of immune cells containing non-stimulatory myeloid cells and stimulatory myeloid cells with an anti-TREM2 antibody, detecting a complex or moiety indicative of antibody binding to the cells, and optionally quantifying the number of non-stimulatory myeloid cells in the population. In another aspect, there is provided a method for determining the relative ratio of non-stimulatory myeloid cells to stimulatory myeloid cells, including contacting a population of immune cells containing non-stimulatory myeloid cells and stimulatory myeloid cells with an anti-TREM2 antibody; quantifying the number of stimulatory myeloid cells and non-stimulatory myeloid cells; and determining the relative ratio of non-stimulatory myeloid cells to stimulatory myeloid cells.

[0297] In the embodiments described herein for detection and / or quantification, the anti-TREM2 antibody binds to the TREM2 protein but does not necessarily have to affect a biological response such as ADCC, although it may have an impact on the biological response.

[0298] In another aspect, the present invention provides a method for identifying an individual who may respond to immunotherapy (e.g., using an anti-TREM2 antibody) for the treatment of an immune-related condition (e.g., cancer), the method comprising detecting the expression level of TREM2 protein in a biological sample derived from the individual and determining whether the individual may respond to immunotherapy based on the expression level of the TREM2 protein. An elevated level of TREM2 protein in an individual compared to that of a healthy individual indicates that the individual may respond to immunotherapy. In some embodiments, these methods may also be used to diagnose an immune-related condition (e.g., cancer) in an individual, and an elevated level of TREM2 protein in an individual compared to that of a healthy individual based on the expression level of the TREM2 protein indicates that the individual has cancer. In some embodiments, the expression level comprises the mRNA expression level of mRNA encoding the TREM2 protein. In other embodiments, the expression level of the TREM2 protein comprises the protein expression level of the TREM2 protein. In some embodiments, the expression level of the TREM2 protein is detected in the sample using a method selected from the group consisting of FACS, Western blot, ELISA, immunoprecipitation, immunohistochemistry, immunofluorescence, radioimmunoassay, dot blotting, immunodetection methods, HPLC, surface plasmon resonance, optical spectroscopy, mass spectrometry, HPLC, qPCR, RT-qPCR, multiplex qPCR or RT-qPCR, RNA-seq, microarray analysis, SAGE, MassARRAY technology, and FISH, and combinations thereof. In these embodiments, the anti-TREM2 antibody binds to the TREM2 protein but does not necessarily affect biological responses such as ADCC. In some embodiments, the biological sample is derived from tumor tissue. In some embodiments, the biological sample includes, but is not limited to, body fluids, tissue samples, organ samples, urine, feces, blood, saliva, CSF, and any combination thereof.

[0299] Methods for enhancing an immune response to a tumor in a subject, or methods for enhancing the effectiveness of an immunotherapy treatment, are also disclosed herein. Generally, a treatment that increases the abundance of SDCs will improve the subject's outcome, such as the recurrence-free survival period, and will enhance the effectiveness of a cancer immunotherapy treatment. The treatment can increase the relative or absolute abundance of SDC cells in the subject's tumor. The treatment can decrease the relative or absolute abundance of NSM cells in the subject's tumor.

[0300] Exemplary methods of general treatment strategies include increasing the number of SDCs by systemic introduction of Flt3L. Another method is treatment of the subject's autologous bone marrow or blood cells with Flt3L while simultaneously blocking CSF1. For example, expression of SDC transcription factors such as IRF8, Mycl1, or BATF3 or ZBTB46 in bone marrow or blood progenitor cell populations by retrovirus may also be used to promote the emergence of SDCs. Another treatment strategy includes systematically eliminating NSM cells while selectively sparing SDCs. This can result in an overall favorable change in the ratio of these populations. Elimination of NSM cells can be achieved by any means including administration of an antibody against the TREM2 surface protein (systemic or local to the tumor).

[0301] In some embodiments, the SDC-enhancing treatment is applied as a therapeutic treatment that enables the subject's natural immune system to better control or eradicate cancer. In another embodiment, the SDC-enhancing treatment of the invention is applied in combination with a therapeutic treatment such as an immunotherapy treatment (such application is performed before, simultaneously with, or after the immunotherapy treatment), and the SDC-enhancing treatment acts as an adjunct or adjuvant treatment to increase the effectiveness of the therapeutic treatment.

[0302] Route of Administration In some embodiments, the methods provided herein are useful for treating an immune-related condition in an individual. In one embodiment, the individual is a human and the antibody is a TREM2 antibody. In another embodiment, the individual is a mouse and the antibody is a TREM2 antibody.

[0303] In some embodiments, for in vivo administration of the anti-TREM2 antibodies described herein, the normal dosage can vary from about 10 ng / kg to about 100 mg / kg of the body weight of an individual per day, preferably from about 1 mg / kg / day to 10 mg / kg / day, depending on the route of administration. For repeated administration over several days or more, depending on the severity of the disease or disorder to be treated, the treatment is continued until the desired suppression of symptoms is achieved. An exemplary dosing regimen includes administering an initial dose of about 2 mg / kg of the anti-TREM2 antibody, followed by a maintenance dose of about 1 mg / kg weekly, administered biweekly. Other dosing regimens may be useful depending on the pattern of pharmacokinetic decline the physician wishes to achieve. For example, administering to an individual once to 21 times a week is contemplated herein. In certain embodiments, administrations ranging from about 3 μg / kg to about 2 mg / kg (e.g., about 3 μg / kg, about 10 μg / kg, about 30 μg / kg, about 100 μg / kg, about 300 μg / kg, about 1 mg / kg, and about 2 mg / kg) may be used. In certain embodiments, the frequency of administration is three times a day, twice a day, once a day, once every two days, once a week, once every two weeks, once every four weeks, once every five weeks, once every six weeks, once every seven weeks, once every eight weeks, once every nine weeks, once every ten weeks, or once a month, once every two months, once every three months, or more. The progress of the treatment is readily monitored by conventional techniques and assays. The dosing regimen, including the anti-TREM2 antibody administered, can vary over time, independent of the dose used.

[0304] In some embodiments, the methods provided herein (e.g., methods of enhancing an immune response or methods that result in inactivation of non-stimulatory bone marrow cells) are useful for the treatment of cancer, and the individual to whom the anti-TREM2 antibody or anti-TREM2 antibodies are administered has cancer.

[0305] Any suitable cancer can be treated with the antibodies provided herein. The cancer can be any cancer tumor, adenocarcinoma, soft tissue, sarcoma, teratoma, melanoma, leukemia, Hodgkin's lymphoma, non-Hodgkin's lymphoma, or brain cancer known in the medical field. In some embodiments, the cancer is a solid cancer. In some embodiments, the cancer is a liquid cancer. In some embodiments, the cancer is immune evasive. In some embodiments, the cancer is immune responsive. In some embodiments, the cancer is melanoma, kidney, hepatobiliary duct, head and neck squamous cell carcinoma (HNSC), pancreas, colon, bladder, glioblastoma, prostate, lung, breast (mammary gland), ovary, stomach, kidney, bladder, esophagus, kidney, melanoma, leukemia, lymphoma, or mesothelioma. In some embodiments, the cancer is colon cancer, pancreatic cancer, or breast cancer.

[0306] In some embodiments, the immune-related condition is an immune-related condition associated with the expression of the TREM2 protein on (human) non-stimulatory myeloid cells or the expression of a homolog of the TREM2 protein in non-human species. In some embodiments, the immune-related condition is an immune-related condition associated with overexpression of the TREM2 protein in non-stimulatory myeloid cells compared to stimulatory myeloid cells. In some embodiments, the overexpression of TREM2 mRNA or the TREM2 protein is at least about 2-fold, 5-fold, 10-fold, 25-fold, 50-fold, or 100-fold higher compared to stimulatory myeloid cells.

[0307] In some embodiments, the treatment enhances the immune response in the subject. In some embodiments, the enhanced immune response is an adaptive immune response. In some embodiments, the enhanced immune response is a innate immune response.

[0308] In some embodiments, the antibody is administered intravenously, intramuscularly, subcutaneously, locally, orally, transdermally, intraperitoneally, intraorbitally, by transplantation, by inhalation, into the subarachnoid space, into the cerebral ventricle, or intranasally. An effective amount of the anti-TREM2 antibody may be administered for the treatment of cancer. The appropriate dosage of the anti-TREM2 antibody may be determined based on the type of cancer to be treated, the type of anti-TREM2 antibody, the severity and course of the cancer, the clinical condition of the individual, the clinical history of the individual, and the response to the treatment, as well as the judgment of the attending physician.

[0309] Combination therapy In some embodiments, the antibodies provided herein are administered with at least one additional therapeutic agent. Any suitable additional therapeutic agent may be administered with the antibodies provided herein. In some embodiments, the immunotherapy is selected from checkpoint inhibitors; T cell checkpoint inhibitors; anti-PD1 antibodies; anti-PDL1 antibodies; anti-CTLA4 antibodies; adoptive T cell therapy; CAR-T cell therapy; dendritic cell vaccines; monocyte vaccines; antigen-binding proteins that bind to both T cells and antigen-presenting cells; BiTE bispecific antigen-binding proteins; toll-like receptor ligands; cytokines; cytotoxic therapies; chemotherapy; cytostatic agents; radiation therapy; small molecule inhibitors; small molecule agonists; immunomodulatory agents; and epigenetic regulators, combinations thereof.

[0310] In some embodiments, the additional therapeutic agent is an antibody. In some embodiments, the additional therapeutic agent is an antibody that binds to a protein(s) on the surface of tumor cells.

[0311] For the treatment of cancer, the anti-TREM2 antibody may be combined with one or more antibodies that inhibit immune checkpoint proteins. Immune checkpoint proteins displayed on the surface of tumor cells are of particular interest. The immune checkpoint receptors most actively studied in the context of clinical cancer immunotherapy, cytotoxic T lymphocyte-associated antigen 4 (CTLA4, also known as CD152), and programmed cell death protein 1 (PD1, also known as CD279), are both inhibitory receptors. The clinical activity of antibodies that block either of these receptors implies that they can enhance anti-tumor immunity at multiple levels and that combination strategies can be rationally designed based on mechanistic considerations and preclinical models.

[0312] The two ligands of PD-1 are PD-1 ligand 1 (PD-L1, also known as B7-H1 and CD274) and PD-L2 (also known as B7-DC and CD273). PD-L1 is expressed on cancer cells and inhibits the activation / function of T cells through binding to the receptor PD-1 on T cells. Inhibitors that block the interaction of PD-1 with its cognate ligands PD-L1 and PD-L2 on cancer cells can result in increased activation and function of T cells and prevent cancer cells from evading the immune system.

[0313] In some embodiments, the immunotherapy is an agent that interferes with the binding of PD-1 and PD-L1 or PD-L2. In some embodiments, the immunotherapy is an anti-PD1 antibody. In some embodiments, the immunotherapy is an anti-PD-L1 antibody. In some embodiments, the immunotherapy is an anti-PD-L2 antibody.

[0314] Various PD-1, PD-L1, and PD-L2 antibodies are known in the art. In some embodiments, the additional therapeutic agent is at least one of atezolizumab (PD-L1), avelumab (PD-L1), durvalumab (PD-L1), nivolumab (PD-1), pembrolizumab (PD-1), semaprimab (PD-1), ipilimumab (CTLA-4), tremelimumab (CTLA-4), or any combination thereof.

[0315] The additional therapeutic agent can be administered by any suitable means. In some embodiments, the antibodies provided herein and the additional therapeutic agent are included in the same pharmaceutical composition. In some embodiments, the antibodies provided herein and the additional therapeutic agent are included in different pharmaceutical compositions.

[0316] In embodiments where the antibodies provided herein and the additional therapeutic agent are included in different pharmaceutical compositions, administration of the antibody can be performed before, simultaneously with, and / or after administration of the additional therapeutic agent. In some embodiments, administration of the antibodies provided herein and the additional therapeutic agent are performed within about 1 month of each other. In some embodiments, administration of the antibodies provided herein and the additional therapeutic agent are performed within about 1 week of each other. In some embodiments, administration of the antibodies provided herein and the additional therapeutic agent are performed within about 1 day of each other. In some embodiments, administration of the antibodies provided herein and the additional therapeutic agent are performed within about 12 hours of each other. In some embodiments, administration of the antibodies provided herein and the additional therapeutic agent are performed within about 1 hour of each other.

[0317] Kits and Manufactured Articles This application provides a kit comprising any one or more of the antibody compositions described herein. In some embodiments, the kit further contains a component selected from any of a secondary antibody, immunohistochemistry assay reagents, pharmaceutically acceptable excipients, and instructions for use, and any combination thereof. In one particular embodiment, the kit comprises a pharmaceutical composition comprising any one or more of the antibody compositions described herein together with one or more pharmaceutically acceptable excipients.

[0318] This application also provides a manufactured article comprising any one of the antibody compositions or kits described herein. Examples of manufactured articles include vials (including sealed vials).

Examples

[0319] The following are examples of specific embodiments for carrying out the present invention. The examples are provided for illustrative purposes only and are in no way intended to limit the scope of the present invention. Efforts have been made to ensure accuracy with respect to the numerical values used (e.g., amounts, temperatures, etc.), but some experimental errors and deviations are to be expected.

[0320] The practice of the present invention will employ, unless otherwise indicated, conventional methods of protein chemistry, biochemistry, recombinant DNA techniques, and pharmacology within the skill of the art. Such techniques are fully explained in the literature. For example, see T.E. Creighton, Proteins: Structures and Molecular Properties (W.H. Freeman and Company, 1993); A.L. Lehninger, Biochemistry (Worth Publishers, Inc., current addition); Sambrook, et al., Molecular Cloning: A Laboratory Manual (2nd Edition, 1989); Methods In Enzymology (S. Colowick and N. Kaplan eds., Academic Press, Inc.); Remington’s Pharmaceutical Sciences, 18th Edition (Easton, Pennsylvania: Mack Publishing Company, 1990); Carey and Sundberg Advanced Organic Chemistry 3 rd Ed. (Plenum Press) Vols A and B (1992).

[0321] Example 1: Humanization of an anti-TREM2 antibody. Humanization of clone #237920 Monoclonal rat IgG specific for mouse and human TREM2 2B Clone #237920 (R&D Systems catalog number MAB17291) was used for sequencing and humanization. Briefly, the disulfide bonds of the antibody were reduced with dithiothreitol (DTT), and the free sulfhydryl groups were alkylated with iodoacetamide. The alkylated antibody was digested with a sequencing-grade endoprotease, purified using a spin column, and the sequence was determined by LC-MS / MS analysis. The sequence is shown below. TIFF2025109867000006.tif69166

[0322] The VH and VL sequences were compared with a library of known human germline sequences on the NCBI website (http: / / www.ncbi.nlm.nih.gov / igblast / ; Ye, J. et al. Nucleic Acids Research 41: W34-W40 (2013)). The databases used were the IMGT human VH gene (F+ORF, 273 germline sequences) and the IMGT human VL kappa gene (F+ORF, 74 germline sequences).

[0323] For 237920 VH, the human germline IGHV3-23 (allele 1) was selected as the acceptor sequence, and the human heavy chain IGHJ4 (allele 1) joining region (J gene) was selected from the human joining region sequences curated by IMGT® the international ImMunoGeneTics information system® www.imgt.org (founder and director: Marie-Paule Lefranc, Montpellier, France).

[0324] In 237920VL, the human germline IGKV1-39 (allele 1) was selected as the acceptor sequence, and the human light chain IGKJ2 (allele 1) binding region (J gene) was selected from the human binding region sequences curated by IMGT® the international ImMunoGeneTics information system® at www.imgt.org (founder and director: Marie-Paule Lefranc, Montpellier, France).

[0325] The CDRs were defined according to the AbM definition (for a table comparing CDR definitions, see the website of Dr. Andrew C.R. Martin at www.bioinf.org.uk / abs / ). For example, in order to optimize the binding of the humanized antibody, changes in the positions of the human germline frameworks (i.e., the non-CDR residues of VH and VL) to the corresponding parental mouse sequences were used.

[0326] Table 1A shows the VL, VH, and complete heavy and light chain sequences of the humanized version of the generated mAb 237920. 37017 is the parental humanized clone from which other humanized versions were created by additional mutations. Table 1B shows the CDR sequences.

[0327] (Table 1A) TIFF2025109867000007.tif245170TIFF2025109867000008.tif25170

[0328] (Table 1B) CDRs of the humanized antibody TIFF2025109867000009.tif52128

[0329] Alignment of the frameworks of the humanized antibody (in the order of appearance, SEQ ID NOs: 7, 3, 5, 21, 6, 23, and 24 respectively) TIFF2025109867000010.tif117158

[0330] In the CDRs within the VL domain, Asn28, Asn31, Asn32, and Asn53 have a low potential for deamidation based on sequence and conformation. Asn93 has a low to moderate potential for deamidation and may exhibit low levels of this post-translational modification. In the VH domain, Asn31 has a low potential for deamidation based on sequence and conformation. In CDR-H2, Asn53 has a moderate potential for deamidation that prevents post-translational modification, and Asn53 can be modified to Gln, Ser, or Ala, and the maintenance of binding was experimentally determined. In CDR-H3, Trp100 is exposed to the solvent and may have the potential for oxidation, especially under stress conditions.

[0331] In-solution endoprotease digestion For mAb sequencing analysis, in-solution endoprotease digestion of monoclonal antibody (mAb) was performed. 50 μg of the antibody was reduced with DTT, alkylated using iodoacetamide, acetone precipitated, and reconstituted in water at a concentration of 1 μg / μL. According to the manufacturer's instructions, five individual enzyme digestions: Asp-N, chymotrypsin, elastase, trypsin, and pepsin were used to perform in-solution digestion of the antibody sample. Next, the sample was lyophilized, resuspended in 0.1% TFA, and purified using a C18 Zip-Tip. Next, the sample was dried by vacuum centrifugation and stored frozen until mass spectrometry.

[0332] Mass spectrometry Intact mass measurement Samples of mAb were denatured, reduced, and acidified. Next, the proteins were analyzed using an Agilent 1100 HPLC connected to a Waters QToF Ultima Global mass spectrometer (LC-ESI-TOF MS). The appropriate LC-MS spectra were processed (merged, subtracted, smoothed, deconvoluted) using Waters MassLynx4.1 software.

[0333] LC-MS / MS analysis The purified peptide was resuspended in 0.1% formic acid, and each half of the digest was analyzed on an Orbitrap analyzer (Q-Exactive, Thermo Fisher Scientific) equipped with a nano spray source and an EASY-NLC 1000 system (Thermo Fisher Scientific). The peptide was loaded onto a 50 cm (75 μm inner diameter) EASY-Spray column packed with PepMap® RSLC 2 μm C18 resin at a pressure of 800 bar. The peptide was eluted at a rate of 250 μL / min using a gradient set from 0% to 30% acetonitrile in 0.1% formic acid over 60 minutes. The peptide was introduced into the Q-Exactive mass spectrometer (Thermo Fisher Scientific) by a nanoelectrospray ionization source. The instrument method consisted of one MS full scan (400 - 1600 m / z) in the Orbitrap mass spectrometer with an automatic gain control (AGC) target of 1E6, a maximum ion injection time of 120 ms, and a resolution of 70000, followed by 10 data-dependent MS / MS scans with a resolution of 17500, an AGC target of 5E5, and a maximum ion time of 100 ms, and one microscan. The intensity threshold for triggering the MS / MS scan was set at a 1.0% underfill ratio. Fragmentation occurred in the HCD collision cell with a normalized collision energy set at 30. Dynamic exclusion was applied using an 8-second setting.

[0334] Table 2 summarizes the biophysical properties of the humanized clones. The molecular weight and extinction coefficient were estimated for the total contributing protein chains in the quaternary structure. By default, the calculation assumes an equal monomer contribution from each chain. The extinction coefficient is the predicted absorbance at 280 nm per mole of protein in units of M -1 cm -1 per cm. Potential post-translational modifications such as glycosylation, phosphorylation, and proteolysis are not considered in the estimation of the molecular weight or extinction coefficient.

[0335] (Table 2) TIFF2025109867000011.tif35157

[0336] Example 2: Production and Characterization of Anti-TREM2 Antibodies Production and Characterization of Antibodies Using standard methods, a standard protein expression vector was transfected into HEK293, and then the cells were grown for 7 days and harvested. In addition to HEK293, antibodies were also produced in 293 cells deficient in mammalian a1,6 fucosyltransferase (FUT8) by CRISPR / Cas9 editing (Alexander Weiss, University of Toronto). The pH of the supernatant was adjusted with 1 M Hepes pH 7.4, and sodium azide was added to prevent microbial growth. The protein was captured using KanCap A resin, washed with PBS containing 1 M sodium chloride and then with PBS, and then eluted with 50 mM citric acid pH 3.5, 100 mM NaCl. Immediately after elution, the solution was neutralized with 1 M Tris (pH 8) containing 0.5 M arginine. Biophysical characterization of the protein buffer-exchanged into PBS was performed using standard techniques. The protein was quantified at OD280, and the calculated extinction coefficient was used to determine the amount and concentration. Purity and approximate molecular weight were determined using reducing and non-reducing SDS-PAGE (Biorad standard Tris / glycine / SDS, 4 - 20%) or the Perkin Elmer GXII capillary electrophoresis system. The aggregation state was determined by HPLC using detection at 280 nm with a Sepax Zenix-C SEC-300, 3um, 300Å, 4.6 * 150 mm size exclusion column and PBS running buffer.

[0337] Measurement of Antibody Affinity Using Surface Plasmon Resonance (SPR) Human TREM2 His (Sino Biological, Beijing, People's Republic of China) immobilized directly on the chip by amine coupling or human or TREM2 human IgG1 Fc fusion protein (in-house SEC purified to greater than 95% purity) captured on Series S CM5 chips through anti-His capture was used to determine binding kinetics by surface plasmon resonance using a Biacore T200 (GE Healthcare, UK). Serial dilutions of the indicated antibodies were injected at 30 μl / min for 2 minutes. Next, PBS or system buffer was injected at 30 μl / min for 400 seconds to observe dissociation. The binding response was corrected by subtracting the response of the blank flow cell. In the kinetic analysis, a 1:1 Langmuir model of global fitting of the k on value and k off value was used. The K d value was determined from the ratio of k on and k off .

[0338] Table 3 shows the antibody binding affinities to human TREM2-His measured by SPR. (Table 3) TIFF2025109867000012.tif21154

[0339] Table 4 shows the antibody binding affinities to human TREM2-Fc measured by SPR. (Table 4) TIFF2025109867000013.tif32154

[0340] At low ligand density (RL = 500 RU), the binding kinetics of PI37017 to human TREM2-Fc did not result in a good fit. This data indicates that A at position 97 and K at position 98 (clone 37017) of the sequence of SEQ ID NO: 31 are rat IgG 2BDuring humanization of clone #237920, it is highly likely to cause substantial loss of human TREM2 binding. Mutations of these framework residues (A97T and K98R) result in an increase in human TREM2 binding by the humanized clone. See, for example, clone 37012.

[0341] Example 3: Cell Binding of Anti-TREM2 Antibodies Cell Binding (EC50 Measurement): 100,000 - 500,000 Expi293 parental cells or Expi293 cells overexpressing human or mouse TREM2 were plated in 96-well plates, and dead cells were stained with Zombie Near Infrared (Biolegend). The indicated non-conjugated antibody titrations were incubated with these cells over an 8 - 10-point range at a 1:3 dilution range within the range of 0 - 10 μg / ml. Depending on their isotype (hIgG1 or mIgG2a), these primary non-conjugated antibodies were detected with Alexa Fluor 647-conjugated anti-human Fc or anti-mouse Fc secondary antibodies (Jackson Immunoresearch). The Alexa Fluor 647 signal was measured by flow cytometry (BD Fortessa X-14, BD Biosciences). EC50 values were calculated by curve fitting the signal from the antibody binding to the overexpressing cells through the background fluorescence generated from HEK293 parental cells using Graphpad Prism (Graphpad Software).

[0342] This data shows that A at position 97 and K at position 98 of the sequence of SEQ ID NO: 31 (clone 37017) are rat IgG 2B During humanization of clone #237920, it is highly likely to cause substantial loss of human TREM2 binding. Mutations of these framework residues (A97T and K98R) result in an increase in human TREM2 binding by the humanized clone. See, for example, clone 37012.

[0343] Table 5 shows the maximum half-life of the saturation binding of the anti-TREM2 antibody to cell surface TREM2. (Table 5) TIFF2025109867000014.tif71128

[0344] Example 4: PI-7012 improves antitumor activity in combination with anti-PD-1 Materials and Methods CT26.WT (CRL-2638) cells were purchased from the American Type Culture Collection (ATCC). All antibodies for in vivo use were tested for endotoxin and proteins with less than 0.2 EU / mg were used. The amino acid sequence of the anti-mouse PD-1 antibody derived from clone RMP1-14 (Absolute Antibody Inc., catalog number Ab00813-7.1) was determined by mass spectrometry (LC-MS / MS). As described in the literature (Nimmerjahn and Ravetch 2005 Science 310:1510-1512 1 )), a single-point mutation [D265A] was introduced into the Fc region of the RMP1-14 antibody in the mouse IgG1 version to eliminate binding to FcgR. Mouse IgG1 [clone MOPC-21] and mouse IgG2a [clone C1.18.4] isotype controls were purchased from BioXCell. PI-7012 and Afuc-PI-7012 (having the CDR sequences of PI37012 and humanized in the mouse IgG2a format) were produced in Expi293 cells (Thermo Fisher Scientific) or 293 / FUT8 knockout cells (University of Toronto) in the mouse IgG2a format, respectively, and purified using MabSelect protein A resin (GE Life Sciences). The antibodies were eluted with 0.1 M citrate buffer (pH 3.0) and the buffer was exchanged before use.

[0345] All experimental procedures involving live animals were approved by Murigenics' Institutional Animal Care and Use Committees. Female BALB / c mice, 6 - 8 weeks old, were purchased from Taconic and allowed to acclimatize in the animal facility for 1 week before use. After thawing from liquid nitrogen stocks, CT26 cells were harvested by 3 - 7 passages and then used in in vivo experiments. The right flank of female Balb / C mice was shaved and prepared for injection the previous day. On the day of tumor inoculation, cells were harvested and used within 30 minutes. To establish subcutaneous tumors, 1x10 6 CT26 cells were transplanted and then mice were monitored for tumor growth. Tumor volume was calculated from caliper measurements of tumor dimensions using the formula (L×W2) / 2, where L is the longer measurement. When tumors reached an average size of 80 - 100 cubic mm, mice were randomized into treatment groups as shown in Table 6: (Table 6) TIFF2025109867000015.tif93132

[0346] Tumor volume and body weight were monitored twice a week and graphed for inter - group comparative analysis by one - way ANOVA. Mice were euthanized when tumor volume reached approximately 2000 cubic mm during the study, when body weight decreased by more than 15%, or for other health - related concerns.

[0347] Results The inventors determined whether the affinity of an mAb that binds to a specific FcgR via glycan engineering (i.e., by generating an afucosylated version of an anti-TREM2 mAb) can increase antitumor activity. In the CT26 tumor model, PI-7012 and afuc-PI-7012 were tested in combination with anti-PD-1. PI-7012 and afuc-PI-7012 showed similar levels of tumor growth inhibition (79% vs. 88% TGI). Treatment with afuc-PI-7012 resulted in a 30% cure rate. As shown in Figure 1A, afuc-PI-7012, when combined with anti-PD-1, increased antitumor activity more than PI-7012. The effect of afucosylation of PI-7012 on antitumor activity was more clearly seen in the analysis of individual mouse tumor volumes (Figures 1B and 1C). This indicates that afucosylation of the anti-TREM2 antibody provides a significant therapeutic advantage over the core-fucosylated antibody.

[0348] During the course of the study, no significant decrease in body weight was observed in any of the treatment groups (Figure 2). Body weight loss is commonly used as a surrogate marker for treatment-related toxicity. This data indicates that short-term or long-term treatment with anti-TREM2, either as a single agent or in combination with anti-PD-1, was well-tolerated and performed without any significant toxicity being observed.

[0349] Example 5: No apparent toxicity associated with anti-TREM2 therapy Materials and Methods Tissues (lung, liver, brain, kidney, and heart) from mice treated in the above examples were stored in 10% neutral buffered formalin for at least 24 hours, processed as per the protocol for histology, sectioned at 5 - 6 μm, and the sections were stained with hematoxylin and eosin. The stained slides were examined using a low magnification (40 - 100x) light microscope and images were obtained by Histo Wiz. Anti-CD68 antibody (AbD Serotec) was used to detect CD68 positive cells and 8 - 9 fields of 40X sections were quantified using a light microscope.

[0350] Results Overall morphological analysis by H&E staining of mouse tissues (lung, liver, heart, kidney, and brain) after treatment showed no morphological changes in mice treated with the combination of PI-7012, afuc-PI-7012, and anti-PD-1 compared to mice treated with the isotype control (Figure 3 shows the staining of lung tissue).

[0351] In addition to H&E staining, tissues were also stained for macrophages using anti-CD68. The intracellular marker CD68 is widely used in the literature as a reliable cytochemical marker for immunostaining monocyte / macrophages in inflamed tissues and tumors. In the lungs (Figure 4A) and other tissues analyzed (Figure 4B), no distinguishable changes in the number of CD68+ macrophages, indicating anti-TREM2-mediated depletion particularly in the TME, were observed in any of the treatment groups compared to the control.

[0352] Example 6: Limited TREM2 Expression in Healthy Mouse Tissues Materials and Methods All animal studies were approved by the Murigenics Animal Studies Committee. C57BL / 6J-Trem2 em2Adiuj / J (hereinafter referred to as TREM2KO) and control C57BL / 6J mice were from the Jackson Laboratory. Whole lungs, spleens, and bones were harvested and processed immediately for flow cytometry. In parallel, blood was collected by cardiac puncture. The tissues were processed into single cell suspensions using the Miltenyi MACS Tissue Dissociation Kit. Red blood cells were lysed using 1x Red Blood Cell Lysis Buffer (Biolegend). Prior to processing for cell surface staining, cells were stained with Fixable Viability Dye (ThermoFisher Scientific). Anti-mouse immunophenotype antibodies were diluted with FACS buffer (2% FBS, 2 mM EDTA, 1×PBS) with Fc block and stained on ice for 30 minutes. After staining, cells were washed twice with FACS buffer and then fixed with 2% paraformaldehyde in PBS for 15 minutes. All data were collected on an LSR Fortessa flow cytometer (BD) or an Attune flow cytometer (Thermo Fisher) and analyzed using FlowJo software. TREM2KO cell staining is shown as gray plots, and wild-type cell staining is shown as white-out plots.

[0353] Results TREM2 is expressed on activated macrophages, immature dendritic cells, osteoclasts, and microglia. 2,3 Cells expressing high levels of TREM2 are thought to be involved in immune surveillance, cell-cell interactions, clearance of tissue debris, and dampening of potential inflammatory responses. 4 The absence of TREM2 expression on these cells by gene knockdown or knockout impairs the ability of the cells to phagocytose debris and also increases the production of regulatory cytokines. 5 In a physiological setting, as seen in FACS plots, TREM2 expression in peripheral blood, spleen, liver, or lung is very low or undetectable (Figure 5). However, when lung or liver resident macrophages are isolated and stained for TREM2 as a pure cell population, TREM2 expression becomes detectable.

[0354] Example 7: TREM2 is mainly expressed on mouse TAM. Materials and Methods Tumor tissues were processed to isolate single-cell suspensions in a standard manner. Briefly, tumors were minced with a scalpel blade and digested in RPMI-1640 medium containing the enzyme of the Miltenyi MACS dissociation kit. The tumors were processed with GentleMACs according to the manufacturer's recommendation and incubated at 37 °C for approximately 40 minutes. The digestion mixture was quenched with PBS containing 2 mM EDTA and 2% fetal bovine serum. Next, the single-cell suspension was passed through a 70um filter, and then the cells were rinsed with FACS buffer. After centrifugation, the cell pellet was resuspended in FACS buffer and stained with an antibody cocktail 6 to identify tumor-associated macrophages and other immune cell populations. TREM2KO cell staining is shown as gray plots, and wild-type cell staining is shown as white plots.

[0355] Results T cells, B cells, NK cells, and other non-myeloid cell populations, as well as CD45-negative cells, do not express detectable TREM2 expression on the cell surface. However, a subset of myeloid cells, including tumor-associated macrophages (TAM) and myeloid-derived suppressor cells (MDSC), express TREM2 to varying degrees on the cell surface. Among the cell types that are positive for TREM2 in the tumor microenvironment, the density of receptor expression on TAM was significantly higher than that of other cell types, regardless of the origin of the tumor (CT26 and MC38 shown in Figure 6).

[0356] Example 8: Restricted TREM2 Expression in Human Peripheral Blood Leukocytes Materials and Methods Peripheral blood mononuclear cells (PBMC) and negatively selected CD14+ monocytes obtained from volunteer normal humans were provided by AllCells Inc. Standard protocol 5CD14+ monocytes were differentiated in vitro using RPMI-1640 complete medium supplemented with 2 mM L-glutamine, 100 μg streptomycin per ml, 100 U penicillin per ml, and 10% heat-inactivated FBS. CD14 + monocytes were cultured. To induce differentiation into macrophages, 50 ng / mL M-CSF was added to the medium. The medium was replenished every 2 - 3 days. After 7 days, macrophages were collected by pipetting and adherent cells were collected by subsequent trypsin treatment. Next, the cells were centrifuged and resuspended in RPMI-1640 supplemented with antibiotics, 2% FBS, recombinant human IFN-γ, and 100 ng / mL LPS. To evaluate cell surface staining of TREM2 in cell subsets, these macrophages were surface stained simultaneously with PBMC using a standard myeloid cocktail. Cells stained with control mAb are shown as gray plots. Cells stained with anti-TREM2 mAb are shown as white plots.

[0357] Results As shown in Figure 7, ex-vivo differentiated macrophages show significantly higher cell surface receptor density of TREM2 compared to the PBMC-based cell types evaluated. Similar to the observations reported in the literature, monocytes and some neutrophils express lower levels of TREM2.

[0358] Example 9: TREM2 is mainly expressed on human TAMs. Materials and Methods Human tumor tissues were obtained from the Cooperative Human Tissue Network (CHTN). Fresh human tumor tissues were dissociated into single-cell suspensions using the Miltenyi MACS separation kit and gentleMACS protocol. Single-cell suspensions of human tumor tissues were surface stained using a pre-validated multi-color FACS panel. All data were collected on an LSR Fortessa flow cytometer (BD) or an Attune flow cytometer (Thermo Fisher) and analyzed using FlowJo software. Numbers indicate the staining index of each population, which is defined as the anti-TREM2 staining minus the isotype control staining.

[0359] Results Within the tumor microenvironment, TREM2 expression was differentially expressed at high levels on TAMs compared to other cells (Figure 8), making it a translational relevant marker for TAMs. Representative histograms of TREM2 antibody staining (white) or isotype control staining (gray) in various cell populations of mucinous adenocarcinoma are shown. Collectively, this data supports the hypothesis that TREM2-targeted agents promote the depletion of specific TAMs with relatively little to no collateral effects on peripheral cells or other tissue-resident immune subsets.

[0360] Example 10: Antitumor Efficacy of Anti-TREM2 Antibody in Combination with Anti-PD-1 in Multiple Syngeneic Tumor Models Materials and Methods CT26.WT (CRL-2638), Py8119 (CRL-3278), 4T1 (CRL-2539), and EMT6 (CTL-2755) cells were purchased from the American Type Culture Collection (ATCC). Panc-02 cells were used from AJES Life Sciences (Stony Brook, NY). All antibodies used in vivo were proteins with less than 0.2 EU / mg. The amino acid sequence of the anti-mouse PD-1 antibody of clone RMP1-14 was determined by mass spectrometry (LC-MS / MS). To eliminate binding to FcgR, a single point mutation D265A was introduced into the Fc region of the RMP1-14 antibody. Mouse IgG1, clone MOPC-21, and mouse IgG2a, clone C1.18.4 were purchased from BioXCell as isotype controls. PI-7012 and afuc-PI-7012 (both as mouse IgG2a) were produced in Expi293 cells (Thermo Fisher Scientific) or 293 / FUT8 knockout cells, respectively, and then purified using MabSelect protein A resin (GE Life Sciences). The mAb was eluted with 0.1 M citrate buffer (pH 3.0), and the buffer was exchanged before use.

[0361] All experimental procedures involving live animals were approved by the Institutional Animal Care and Use Committees of Murigenics. Female BALB / c or C57BL / 6 mice (6 - 8 weeks old) were purchased from Taconic Farms or The Jackson Laboratory and allowed to acclimatize in the animal facility for 1 week before use. After thawing from liquid nitrogen stocks, tumor cells were harvested by 3 - 7 passages and then used in in vivo experiments. The right flank of female mice was shaved and prepared for injection the day before tumor cell inoculation. On the day of tumor inoculation, cells were harvested and used within 30 minutes. To establish subcutaneous tumors, 1x10 6 of CT26, EMT6, or Panc-02 cells, or 1x10 54T1 cells were transplanted into a suitable strain of mice, and then the animals were monitored for tumor growth. An equal volume of a single cell suspension of Py8119 cells was mixed with Matrigel (Corning catalog number 354248 or 354263), and then 2x10 6 cells per mouse were transplanted.

[0362] Tumor volume was calculated using Vernier caliper measurements of tumor dimensions and the formula (L×W2) / 2, where L is the longer measurement. When tumors reached an average size of 80 - 100 cubic mm, mice were randomized into treatment groups as shown in Table 7.

[0363] Tumor volume and body weight were monitored twice a week and graphed for between-group comparative analysis by one-way ANOVA. Mice were euthanized when tumor volume reached approximately 2,000 cubic mm or when body weight decreased by more than 15% during the study.

[0364] (Table 7) TIFF2025109867000016.tif35166

[0365] Results The results are summarized in Table 8. Tumor growth inhibition (%TGI) was determined at the end of the dosing period (t) using the formula: %TGI = (1 - {Tt / T0 / Ct / C0} / 1 - {C0 / Ct}) X 100, where Tt = the median tumor volume of the combination-treated group at time t, T0 = the median tumor volume of the combination-treated group at time 0, Ct = the median tumor volume of the isotype control group at time t, and C0 = the median tumor volume of the isotype-treated group at time 0 (before treatment initiation).

[0366] (Table 8) TIFF2025109867000017.tif63153

[0367] Figures 9A - F show the antitumor activity of anti - TREM2 PI - 7012 or afuc - PI7012 in combination with anti - PD - 1 in multiple syngeneic mouse tumor models. The anti - TREM2 mAb afuc - PI7012 in combination with anti - PD - 1 mAb confers significant antitumor activity in the Panc - 02 pancreatic tumor model. Figure 9A shows the mean + / - standard deviation of the mean tumor volume of 10 mice in each group. Figures 9B, 9C, 9D, and 9E show the tumor volume of individual animals in each treatment group over time. Figure 9F shows the statistical analysis of the group - mean tumor volume on day 32 after transplantation. The differences in tumor volume between groups were evaluated using statistical analysis available in Graph Pad Prism software. For the study data, one - way ANOVA was followed by Sidak's multiple - comparison test.

[0368] As seen in Figures 9A and 9D, subcutaneous Panc - 02 tumors do not respond to anti - PD - 1 mAb monotherapy or anti - TREM2 mAb afuc - PI - 7012 therapy alone. However, combination treatment of Panc - 02 tumor - bearing animals with anti - TREM2 mAb afuc - PI - 7012 and anti - PD - 1 mAb resulted in significant tumor growth inhibition.

[0369] Combined therapeutic strategies for myeloid modulation, with reversal of immune checkpoint - mediated CD8 T - cell exhaustion, were tested in multiple syngeneic tumor models. As shown in Table 8, the combination of anti - TREM2 and anti - PD - 1 mAbs resulted in significant tumor growth inhibition and complete regression in some of the tested tumor models. It is important to note that these syngeneic models were grown in the background of two different mouse strains (the typical Th - 1 C57BL / 6 and Th - 2 BALB / c strains), and these backgrounds are known to have significant differences in the composition of the immune infiltrates of tumors grown in these strains in vivo.

[0370] Example 11: Induce long - term antitumor immune memory in mice responsive to combination treatment with anti - TREM2 mAb and anti - PD - 1 mAb. Materials and Methods BALB / c mice without tumors from previous studies after treatment with the anti-TREM2 mAb and anti-PD-1 mAb described in Example 9 were rechallenged 3 months later with 1 × 10 6 CT26 tumor cells. Tumor volume was measured for 25 days after transplantation. An equal number of CT26 cells were administered to age-matched naive mice, and tumor growth was followed during the study period. No additional treatment was provided to the mice during the study period.

[0371] Results Mice in which CT26 tumors had been cured after treatment with the combination of the anti-TREM2 mAb afuc-PI-7012 and the anti-PD-1 mAb established an effective anti-tumor memory response (Figure 10). Cured mice were able to reject the growth of new tumors without additional treatment, indicating long-term immune memory against the original transplanted tumors. This form of long-term immune memory utilizes the maintenance of an active CD8+ effector memory response.

[0372] References TIFF2025109867000018.tif78162

[0373] Although the present invention has been shown and described with respect to preferred embodiments and various alternative embodiments, those skilled in the art will understand that various changes in form and detail may be made herein without departing from the spirit and scope of the invention.

[0374] All references, issued patents, and patent applications cited within the text of this specification are hereby incorporated by reference in their entirety for all purposes.

[0375] Sequences TIFF2025109867000019.tif217170TIFF2025109867000020.tif234170TIFF2025109867000021.tif185170

[0376] Sequence information SEQUENCE LISTING <110> PIONYR IMMUNOTHERAPEUTICS, INC. <120> ANTI-TREM2 ANTIBODIES AND RELATED METHODS <150> US 62 / 648,089 <151> 2018-03-26 <150> US 62 / 597,827 <151> 2017-12-12 <160> 34 <170> PatentIn version 3.5 <210> 1 <211> 119 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 1 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Tyr 20 25 30 Tyr Met Ala Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Leu Thr Asn Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Thr Arg Glu Trp Ala Gly Ser Gly Tyr Phe Asp Tyr Trp Gly Gln Gly 100 105 110 Thr Leu Val Thr Val Ser Ser 115 <210> 2 <211> 107 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 2 Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr Cys Lys Ala Ser Gln Asn Val Gly Asn Asn 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Tyr Tyr Thr Ser Asn Arg Phe Thr Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Arg Ile Tyr Asn Ser Pro Trp 85 90 95 Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 <210> 3 <211> 119 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 3 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Tyr 20 25 30 Tyr Met Ala Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Leu Thr Asn Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Thr Arg Glu Trp Ala Gly Ser Gly Tyr Phe Asp Tyr Trp Gly Gln Gly 100 105 110 Thr Leu Val Thr Val Ser Ser 115 <210> 4 <211> 107 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 4 Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Met Thr Cys Lys Ala Ser Gln Asn Val Gly Asn Asn 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Leu 35 40 45 Tyr Tyr Thr Ser Asn Arg Phe Thr Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Val Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Arg Ile Tyr Asn Ser Pro Trp 85 90 95 Thr Phe Gly Gln Gly Thr Lys Leu Glu Leu Lys 100 105 <210> 5 <211> 119 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 5 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Tyr 20 25 30 Tyr Met Ala Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Ser Leu Thr Asn Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Leu Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Thr Arg Glu Trp Ala Gly Ser Gly Tyr Phe Asp Tyr Trp Gly Gln Gly 100 105 110 Thr Leu Val Thr Val Ser Ser 115 <210> 6 <211> 107 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 6 Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr Cys Lys Ala Ser Gln Asn Val Gly Asn Asn 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Tyr Tyr Thr Ser Asn Arg Phe Thr Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Arg Ile Tyr Asn Ser Pro Trp 85 90 95 Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 <210> 7 <211> 119 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 7 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Tyr 20 25 30 Tyr Met Ala Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Leu Thr Asn Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Glu Trp Ala Gly Ser Gly Tyr Phe Asp Tyr Trp Gly Gln Gly 100 105 110 Thr Leu Val Thr Val Ser Ser 115 <210> 8 <211> 107 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 8 Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr Cys Lys Ala Ser Gln Asn Val Gly Asn Asn 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Tyr Tyr Thr Ser Asn Arg Phe Thr Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Arg Ile Tyr Asn Ser Pro Trp 85 90 95 Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 <210> 9 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 9 Phe Ser Asn Tyr Tyr Met Ala 1 5 <210> 10 <211> 10 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 10 Ser Leu Thr Asn Ser Gly Gly Ser Thr Tyr 1 5 10 <210> 11 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 11 Glu Trp Ala Gly Ser Gly Tyr 1 5 <210> 12 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 12 Asn Val Gly Asn Asn Leu Ala 1 5 <210> 13 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 13 Tyr Thr Ser Asn Arg Phe Thr 1 5 <210> 14 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic peptide <400> 14 Arg Ile Tyr Asn Ser Pro Trp 1 5 <210> 15 <211> 230 <212> PRT <213> Homo sapiens <400> 15 Met Glu Pro Leu Arg Leu Leu Ile Leu Leu Phe Val Thr Glu Leu Ser 1 5 10 15 Gly Ala His Asn Thr Thr Val Phe Gln Gly Val Ala Gly Gln Ser Leu 20 25 30 Gln Val Ser Cys Pro Tyr Asp Ser Met Lys His Trp Gly Arg Arg Lys 35 40 45 Ala Trp Cys Arg Gln Leu Gly Glu Lys Gly Pro Cys Gln Arg Val Val 50 55 60 Ser Thr His Asn Leu Trp Leu Leu Ser Phe Leu Arg Arg Trp Asn Gly 65 70 75 80 Ser Thr Ala Ile Thr Asp Asp Thr Leu Gly Gly Thr Leu Thr Ile Thr 85 90 95 Leu Arg Asn Leu Gln Pro His Asp Ala Gly Leu Tyr Gln Cys Gln Ser 100 105 110 Leu His Gly Ser Glu Ala Asp Thr Leu Arg Lys Val Leu Val Glu Val 115 120 125 Leu Ala Asp Pro Leu Asp His Arg Asp Ala Gly Asp Leu Trp Phe Pro 130 135 140 Gly Glu Ser Glu Ser Phe Glu Asp Ala His Val Glu His Ser Ile Ser 145 150 155 160 Arg Ser Leu Leu Glu Gly Glu Ile Pro Phe Pro Pro Thr Ser Ile Leu 165 170 175 Leu Leu Leu Ala Cys Ile Phe Leu Ile Lys Ile Leu Ala Ala Ser Ala 180 185 190 Leu Trp Ala Ala Ala Trp His Gly Gln Lys Pro Gly Thr His Pro Pro 195 200 205 Ser Glu Leu Asp Cys Gly His Asp Pro Gly Tyr Gln Leu Gln Thr Leu 210 215 220 Pro Gly Leu Arg Asp Thr 225 230 <210> 16 <211> 693 <212> DNA <213> Homo sapiens <400> 16 atggagcctc tccggctgct catcttactc tttgtcacag agctgtccgg agcccacaac 60 accacagtgt tccagggcgt ggcgggccag tccctgcagg tgtcttgccc ctatgactcc 120 atgaagcact gggggaggcg caaggcctgg tgccgccagc tgggagagaa gggcccatgc 180 cagcgtgtgg tcagcacgca caacttgtgg ctgctgtcct tcctgaggag gtggaatggg 240 agcacagcca tcacagacga taccctgggt ggcactctca ccattacgct gcggaatcta 300 caaccccatg atgcgggtct ctaccagtgc cagagcctcc atggcagtga ggctgacacc 360 ctcaggaagg tcctggtgga ggtgctggca gaccccctgg atcaccggga tgctggagat 420 ctctggttcc ccggggagtc tgagagcttc gaggatgccc atgtggagca cagcatctcc 480 aggagcctct tggaaggaga aatccccttc ccacccactt ccatccttct cctcctggcc 540 tgcatctttc tcatcaagat tctagcagcc agcgccctct gggctgcagc ctggcatgga 600 cagaagccag ggacacatcc acccagtgaa ctggactgtg gccatgaccc agggtatcag 660 ctccaaactc tgccagggct gagagacacg tga 693 <210> 17 <211> 227 <212> PRT <213> Mus musculus <400> 17 Met Gly Pro Leu His Gln Phe Leu Leu Leu Leu Ile Thr Ala Leu Ser 1 5 10 15 Gln Ala Leu Asn Thr Thr Val Leu Gln Gly Met Ala Gly Gln Ser Leu 20 25 30 Arg Val Ser Cys Thr Tyr Asp Ala Leu Lys His Trp Gly Arg Arg Lys 35 40 45 Ala Trp Cys Arg Gln Leu Gly Glu Glu Gly Pro Cys Gln Arg Val Val 50 55 60 Ser Thr His Gly Val Trp Leu Leu Ala Phe Leu Lys Lys Arg Asn Gly 65 70 75 80 Ser Thr Val Ile Ala Asp Asp Thr Leu Ala Gly Thr Val Thr Ile Thr 85 90 95 Leu Lys Asn Leu Gln Ala Gly Asp Ala Gly Leu Tyr Gln Cys Gln Ser 100 105 110 Leu Arg Gly Arg Glu Ala Glu Val Leu Gln Lys Val Leu Val Glu Val 115 120 125 Leu Glu Asp Pro Leu Asp Asp Gln Asp Ala Gly Asp Leu Trp Val Pro 130 135 140 Glu Glu Ser Ser Ser Phe Glu Gly Ala Gln Val Glu His Ser Thr Ser 145 150 155 160 Arg Asn Gln Glu Thr Ser Phe Pro Pro Thr Ser Ile Leu Leu Leu Leu 165 170 175 Ala Cys Val Leu Leu Ser Lys Phe Leu Ala Ala Ser Ile Leu Trp Ala 180 185 190 Val Ala Arg Gly Arg Gln Lys Pro Gly Thr Pro Val Val Arg Gly Leu 195 200 205 Asp Cys Gly Gln Asp Ala Gly His Gln Leu Gln Ile Leu Thr Gly Pro 210 215 220 Gly Gly Thr 225 <210> 18 <211> 119 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 18 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Tyr 20 25 30 Tyr Met Ala Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Leu Thr Asn Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Glu Trp Ala Gly Ser Gly Tyr Phe Asp Tyr Trp Gly Gln Gly 100 105 110 Thr Leu Val Thr Val Ser Ser 115 <210> 19 <211> 119 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 19 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Tyr 20 25 30 Tyr Met Ala Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Leu Thr Asn Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Thr Arg Glu Trp Ala Gly Ser Gly Tyr Phe Asp Tyr Trp Gly Gln Gly 100 105 110 Thr Leu Val Thr Val Ser Ser 115 <210> 20 <211> 119 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 20 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Tyr 20 25 30 Tyr Met Ala Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Ser Leu Thr Asn Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Leu Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Thr Arg Glu Trp Ala Gly Ser Gly Tyr Phe Asp Tyr Trp Gly Gln Gly 100 105 110 Thr Leu Val Thr Val Ser Ser 115 <210> 21 <211> 109 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 21 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser 100 105 <210> 22 <211> 107 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 22 Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr Cys Lys Ala Ser Gln Asn Val Gly Asn Asn 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Tyr Tyr Thr Ser Asn Arg Phe Thr Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Arg Ile Tyr Asn Ser Pro Trp 85 90 95 Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 <210> 23 <211> 107 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 23 Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Met Thr Cys Lys Ala Ser Gln Asn Val Gly Asn Asn 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Leu 35 40 45 Tyr Tyr Thr Ser Asn Arg Phe Thr Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Val Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Arg Ile Tyr Asn Ser Pro Trp 85 90 95 Thr Phe Gly Gln Gly Thr Lys Leu Glu Leu Lys 100 105 <210> 24 <211> 106 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 24 Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Ser Ile Ser Ser Tyr 20 25 30 Leu Asn Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Tyr Ala Ala Ser Ser Leu Gln Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Ser Tyr Ser Thr Pro Pro 85 90 95 Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 <210> 25 <211> 449 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 25 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Tyr 20 25 30 Tyr Met Ala Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Leu Thr Asn Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Thr Arg Glu Trp Ala Gly Ser Gly Tyr Phe Asp Tyr Trp Gly Gln Gly 100 105 110 Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe 115 120 125 Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu 130 135 140 Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp 145 150 155 160 Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu 165 170 175 Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser 180 185 190 Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro 195 200 205 Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys Asp Lys 210 215 220 Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro 225 230 235 240 Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser 245 250 255 Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp 260 265 270 Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn 275 280 285 Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val 290 295 300 Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu 305 310 315 320 Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys 325 330 335 Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr 340 345 350 Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr 355 360 365 Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu 370 375 380 Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu 385 390 395 400 Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys 405 410 415 Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu 420 425 430 Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly 435 440 445 Lys <210> 26 <211> 214 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 26 Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr Cys Lys Ala Ser Gln Asn Val Gly Asn Asn 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Tyr Tyr Thr Ser Asn Arg Phe Thr Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Arg Ile Tyr Asn Ser Pro Trp 85 90 95 Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys Arg Thr Val Ala Ala 100 105 110 Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly 115 120 125 Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala 130 135 140 Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln 145 150 155 160 Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser 165 170 175 Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr 180 185 190 Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser 195 200 205 Phe Asn Arg Gly Glu Cys 210 <210> 27 <211> 449 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 27 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Tyr 20 25 30 Tyr Met Ala Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Leu Thr Asn Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Thr Arg Glu Trp Ala Gly Ser Gly Tyr Phe Asp Tyr Trp Gly Gln Gly 100 105 110 Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe 115 120 125 Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu 130 135 140 Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp 145 150 155 160 Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu 165 170 175 Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser 180 185 190 Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro 195 200 205 Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys Asp Lys 210 215 220 Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro 225 230 235 240 Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser 245 250 255 Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp 260 265 270 Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn 275 280 285 Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val 290 295 300 Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu 305 310 315 320 Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys 325 330 335 Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr 340 345 350 Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr 355 360 365 Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu 370 375 380 Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu 385 390 395 400 Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys 405 410 415 Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu 420 425 430 Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly 435 440 445 Lys <210> 28 <211> 214 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 28 Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Met Thr Cys Lys Ala Ser Gln Asn Val Gly Asn Asn 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Leu 35 40 45 Tyr Tyr Thr Ser Asn Arg Phe Thr Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Val Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Arg Ile Tyr Asn Ser Pro Trp 85 90 95 Thr Phe Gly Gln Gly Thr Lys Leu Glu Leu Lys Arg Thr Val Ala Ala 100 105 110 Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly 115 120 125 Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala 130 135 140 Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln 145 150 155 160 Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser 165 170 175 Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr 180 185 190 Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser 195 200 205 Phe Asn Arg Gly Glu Cys 210 <210> 29 <211> 449 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 29 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Tyr 20 25 30 Tyr Met Ala Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Ser Leu Thr Asn Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Leu Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Thr Arg Glu Trp Ala Gly Ser Gly Tyr Phe Asp Tyr Trp Gly Gln Gly 100 105 110 Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe 115 120 125 Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu 130 135 140 Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp 145 150 155 160 Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu 165 170 175 Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser 180 185 190 Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro 195 200 205 Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys Asp Lys 210 215 220 Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro 225 230 235 240 Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser 245 250 255 Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp 260 265 270 Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn 275 280 285 Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val 290 295 300 Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu 305 310 315 320 Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys 325 330 335 Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr 340 345 350 Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr 355 360 365 Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu 370 375 380 Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu 385 390 395 400 Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys 405 410 415 Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu 420 425 430 Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly 435 440 445 Lys <210> 30 <211> 214 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 30 Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr Cys Lys Ala Ser Gln Asn Val Gly Asn Asn 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Tyr Tyr Thr Ser Asn Arg Phe Thr Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Arg Ile Tyr Asn Ser Pro Trp 85 90 95 Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys Arg Thr Val Ala Ala 100 105 110 Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly 115 120 125 Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala 130 135 140 Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln 145 150 155 160 Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser 165 170 175 Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr 180 185 190 Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser 195 200 205 Phe Asn Arg Gly Glu Cys 210 <210> 31 <211> 449 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 31 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Tyr 20 25 30 Tyr Met Ala Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Leu Thr Asn Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Glu Trp Ala Gly Ser Gly Tyr Phe Asp Tyr Trp Gly Gln Gly 100 105 110 Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe 115 120 125 Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu 130 135 140 Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp 145 150 155 160 Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu 165 170 175 Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser 180 185 190 Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro 195 200 205 Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys Asp Lys 210 215 220 Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro 225 230 235 240 Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser 245 250 255 Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp 260 265 270 Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn 275 280 285 Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val 290 295 300 Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu 305 310 315 320 Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys 325 330 335 Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr 340 345 350 Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr 355 360 365 Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu 370 375 380 Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu 385 390 395 400 Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys 405 410 415 Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu 420 425 430 Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly 435 440 445 Lys <210> 32 <211> 214 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 32 Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr Cys Lys Ala Ser Gln Asn Val Gly Asn Asn 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Tyr Tyr Thr Ser Asn Arg Phe Thr Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Arg Ile Tyr Asn Ser Pro Trp 85 90 95 Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys Arg Thr Val Ala Ala 100 105 110 Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly 115 120 125 Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala 130 135 140 Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln 145 150 155 160 Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser 165 170 175 Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr 180 185 190 Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser 195 200 205 Phe Asn Arg Gly Glu Cys 210 <210> 33 <211> 451 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 33 Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Arg 1 5 10 15 Ser Leu Lys Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Asn Tyr 20 25 30 Tyr Met Ala Trp Val Arg Gln Ala Pro Thr Lys Gly Leu Glu Trp Val 35 40 45 Ala Ser Leu Thr Asn Ser Gly Gly Ser Thr Tyr Tyr Arg Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Leu Ser Arg Asp Asn Ala Lys Ser Thr Leu Tyr 65 70 75 80 Leu Gln Met Asp Ser Leu Arg Ser Glu Asp Thr Ala Thr Tyr Tyr Cys 85 90 95 Thr Arg Glu Trp Ala Gly Ser Gly Tyr Phe Asp Tyr Trp Gly Gln Gly 100 105 110 Val Met Val Thr Val Ser Ser Ala Gln Thr Thr Ala Pro Ser Val Tyr 115 120 125 Pro Leu Ala Pro Gly Cys Gly Asp Thr Thr Ser Ser Thr Val Thr Leu 130 135 140 Gly Cys Leu Val Lys Gly Tyr Phe Pro Glu Pro Val Thr Val Thr Trp 145 150 155 160 Asn Ser Gly Ala Leu Ser Ser Asp Val His Thr Phe Pro Ala Val Leu 165 170 175 Gln Ser Gly Leu Tyr Thr Leu Thr Ser Ser Val Thr Ser Ser Thr Trp 180 185 190 Pro Ser Gln Thr Val Thr Cys Asn Val Ala His Pro Ala Ser Ser Thr 195 200 205 Lys Val Asp Lys Lys Val Glu Arg Arg Asp Gly Gly Ile Gly His Lys 210 215 220 Cys Pro Thr Cys Pro Thr Cys His Lys Cys Pro Val Pro Glu Leu Leu 225 230 235 240 Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Ile Leu 245 250 255 Leu Leu Ser Gln Asn Ala Lys Val Thr Cys Val Val Val Asp Val Ser 260 265 270 Glu Glu Glu Pro Asp Val Gln Phe Ser Trp Phe Val Asn Asn Val Glu 275 280 285 Val His Thr Ala Gln Thr Gln Pro Arg Glu Glu Gln Tyr Asn Ser Thr 290 295 300 Phe Arg Val Val Ser Ala Leu Pro Leu Gln His Gln Asp Trp Met Ser 305 310 315 320 Gly Lys Glu Phe Lys Cys Lys Val Asn Asn Lys Ala Leu Pro Ser Pro 325 330 335 Ile Glu Lys Thr Leu Ser Lys Pro Lys Gly Leu Val Arg Lys Pro Gln 340 345 350 Val Tyr Val Met Gly Pro Pro Thr Glu Gln Leu Thr Glu Gln Thr Val 355 360 365 Ser Leu Thr Cys Leu Thr Ser Gly Phe Leu Pro Asn Asp Ile Gly Val 370 375 380 Glu Trp Thr Ser Asn Gly His Ile Glu Lys Asn Tyr Lys Asn Thr Glu 385 390 395 400 Pro Val Met Asp Ser Asp Gly Ser Phe Phe Met Tyr Ser Lys Leu Asn 405 410 415 Val Glu Arg Ser Arg Trp Asp Ser Arg Ala Pro Phe Val Cys Ser Val 420 425 430 Val His Glu Gly Leu His Asn His His Val Glu Lys Ser Leu Ser Arg 435 440 445 Pro Pro Gly 450 <210> 34 <211> 214 <212> PRT <213> Artificial Sequence <220> <223> Description of Artificial Sequence: Synthetic polypeptide <400> 34 Asn Ile Val Met Thr Gln Ser Pro Lys Ser Met Ser Leu Ser Val Gly 1 5 10 15 Asp Arg Val Thr Met Asn Cys Lys Ala Ser Gln Asn Val Gly Asn Asn 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ser Pro Lys Leu Leu Leu 35 40 45 Tyr Tyr Thr Ser Asn Arg Phe Thr Gly Val Pro Asp Arg Phe Thr Gly 50 55 60 Gly Gly Tyr Gly Thr Asp Phe Thr Leu Thr Ile Asn Ser Val Gln Ala 65 70 75 80 Glu Asp Ala Ala Phe Tyr Tyr Cys Gln Arg Ile Tyr Asn Ser Pro Trp 85 90 95 Thr Phe Gly Gly Gly Thr Lys Leu Glu Leu Lys Arg Ala Asp Ala Ala 100 105 110 Pro Thr Val Ser Ile Phe Pro Pro Ser Thr Glu Gln Leu Ala Thr Gly 115 120 125 Gly Ala Ser Val Val Cys Leu Met Asn Asn Phe Tyr Pro Arg Asp Ile 130 135 140 Ser Val Lys Trp Lys Ile Asp Gly Thr Glu Arg Arg Asp Gly Val Leu 145 150 155 160 Asp Ser Val Thr Asp Gln Asp Ser Lys Asp Ser Thr Tyr Ser Met Ser 165 170 175 Ser Thr Leu Ser Leu Thr Lys Ala Asp Tyr Glu Ser His Asn Leu Tyr 180 185 190 Thr Cys Glu Val Val His Lys Thr Ser Ser Ser Pro Val Val Lys Ser 195 200 205 Phe Asn Arg Asn Glu Cys 210

Claims

**Claim 1** A pharmaceutical composition for treating a cancer of a subject in need thereof, a. the subject is determined to have myeloid cells positive for Triggering Receptor Expressed on Myeloid Cells 2 (TREM2+) present in the cancer, and b. the pharmaceutical composition comprises an effective amount of an isolated antibody that binds to human TREM2 (SEQ ID NO: 15), the antibody comprising a heavy chain comprising a variable heavy chain (VH) sequence comprising three heavy chain CDR sequences of CDR-H1, CDR-H2, and CDR-H3, and a light chain comprising a variable light chain (VL) sequence comprising three light chain CDR sequences of CDR-L1, CDR-L2, and CDR-L3, i. CDR-H1 comprises the sequence set forth in SEQ ID NO: 9, ii. CDR-H2 comprises the sequence set forth in SEQ ID NO: 10, iii. CDR-H3 comprises the sequence set forth in SEQ ID NO: 11, iv. CDR-L1 comprises the sequence set forth in SEQ ID NO: 12, v. CDR-L2 comprises the sequence set forth in SEQ ID NO: 13, and vi. CDR-L3 comprises the sequence set forth in SEQ ID NO: 14, the pharmaceutical composition. **Claim 2** the subject will have previously received, be receiving simultaneously, or will receive subsequently an immunotherapy, the immunotherapy being at least one of a checkpoint inhibitor; a T cell checkpoint inhibitor; an anti-PD1 antibody; an anti-PDL1 antibody; an anti-CTLA4 antibody; adoptive T cell therapy; CAR-T cell therapy; dendritic cell vaccine; monocyte vaccine; an antigen-binding protein that binds to both T cells and antigen-presenting cells; a BiTE bispecific antigen-binding protein; a toll-like receptor ligand; a cytokine; a cytotoxic therapy; chemotherapy; radiotherapy; a small molecule inhibitor; a small molecule agonist; an immunomodulatory agent; and an epigenetic regulator, the pharmaceutical composition according to claim 1. **Claim 3** the pharmaceutical composition according to claim 2, wherein the immunotherapy is an anti-PD1 antibody, an anti-PDL1 antibody, or an anti-CTLA4 antibody. **Claim 4** the pharmaceutical composition according to claim 1, wherein the cancer is a solid cancer or a liquid cancer. **Claim 5** the pharmaceutical composition according to claim 1, wherein the cancer is selected from the group consisting of colon, breast, melanoma, kidney, hepatobiliary tract, head and neck squamous cell carcinoma (HNSC), pancreas, bladder, glioblastoma, prostate, lung, ovary, stomach, kidney, bladder, esophagus, kidney, melanoma, and mesothelioma. **Claim 6** A pharmaceutical composition for killing, inactivating, or depleting target TREM2+ myeloid cells, a. the subject has been determined to have Triggering Receptor Expressed on Myeloid Cells 2 positive (TREM2+) myeloid cells present in the cancer, and b. the pharmaceutical composition comprises an effective amount of an isolated antibody that binds to human TREM2 (SEQ ID NO: 15), the antibody comprising a heavy chain comprising a variable heavy chain (VH) sequence comprising three heavy chain CDR sequences of CDR-H1, CDR-H2, and CDR-H3, and a light chain comprising a variable light chain (VL) sequence comprising three light chain CDR sequences of CDR-L1, CDR-L2, and CDR-L3, i. CDR-H1 comprises the sequence set forth in SEQ ID NO: 9, ii. CDR-H2 comprises the sequence set forth in SEQ ID NO: 10, iii. CDR-H3 comprises the sequence set forth in SEQ ID NO: 11, iv. CDR-L1 comprises the sequence set forth in SEQ ID NO: 12, v. CDR-L2 comprises the sequence set forth in SEQ ID NO: 13, and vi. CDR-L3 comprises the sequence set forth in SEQ ID NO: 14, the pharmaceutical composition.

7. The pharmaceutical composition according to claim 6, wherein the antibody has at least one of antibody-dependent cell-mediated cytotoxicity (ADCC) activity, complement-dependent cytotoxicity (CDC) activity, and antibody-mediated cell phagocytosis (ADCP) activity.

8. The pharmaceutical composition according to claim 6, wherein the TREM2+ myeloid cells comprise at least one of dendritic cells, tumor-associated macrophages (TAM), neutrophils, or monocytes.

9. The pharmaceutical composition according to claim 6, wherein the subject has a tumor and the TREM2+ myeloid cells are within the tumor.

10. The pharmaceutical composition according to claim 6, wherein the subject has previously received, is receiving simultaneously, or will receive immunotherapy thereafter.

11. The pharmaceutical composition according to claim 6, wherein the immunotherapy is at least one of a checkpoint inhibitor; a T cell checkpoint inhibitor; an anti-PD1 antibody; an anti-PDL1 antibody; an anti-CTLA4 antibody; adoptive T cell therapy; CAR-T cell therapy; dendritic cell vaccine; monocyte vaccine; an antigen-binding protein that binds to both T cells and antigen-presenting cells; a BiTE bispecific antigen-binding protein; a toll-like receptor ligand; a cytokine; a cytotoxic therapy; chemotherapy; radiotherapy; a small molecule inhibitor; a small molecule agonist; an immunomodulatory agent; and an epigenetic regulator.

12. The pharmaceutical composition according to claim 6, wherein the immunotherapy is an anti-PD1 antibody, an anti-PDL1 antibody, or an anti-CTLA4 antibody.

13. The pharmaceutical composition according to claim 1, wherein the antibody comprises a human Fc region.

14. The pharmaceutical composition according to claim 1, wherein the VH chain sequence comprises the VH sequence shown in SEQ ID NO: 1, and the VL chain sequence comprises the VL sequence shown in SEQ ID NO:

2.

15. The pharmaceutical composition according to claim 1, wherein the VH chain sequence comprises the VH sequence shown in SEQ ID NO: 1, 3, or 5, and the VL chain sequence comprises the VL sequence shown in SEQ ID NO: 2, 4, or 6.

16. The pharmaceutical composition according to claim 1, wherein the heavy chain comprises the heavy chain sequence shown in SEQ ID NO: 25, and the light chain comprises the light chain sequence shown in SEQ ID NO:

26.

17. The pharmaceutical composition according to claim 6, wherein the antibody comprises a human Fc region.

18. The pharmaceutical composition according to claim 6, wherein the VH chain sequence comprises the VH sequence shown in SEQ ID NO: 1, and the VL chain sequence comprises the VL sequence shown in SEQ ID NO:

2.

19. The pharmaceutical composition according to claim 6, wherein the VH chain sequence comprises the VH sequence shown in SEQ ID NO: 1, 3, or 5, and the VL chain sequence comprises the VL sequence shown in SEQ ID NO: 2, 4, or 6.

20. The pharmaceutical composition according to claim 6, wherein the heavy chain comprises the heavy chain sequence shown in SEQ ID NO: 25, and the light chain comprises the light chain sequence shown in SEQ ID NO:

26.

21. The pharmaceutical composition according to claim 1, wherein the antibody is afucosylated, the VH chain sequence comprises the VH sequence shown in SEQ ID NO: 1, and the VL chain sequence comprises the VL sequence shown in SEQ ID NO:

2.

22. The pharmaceutical composition according to claim 13, wherein the antibody is afucosylated, the human Fc is wild-type human IgG1 Fc, the VH chain sequence comprises the VH sequence shown in SEQ ID NO: 1, and the VL chain sequence comprises the VL sequence shown in SEQ ID NO:

2.

23. The pharmaceutical composition according to claim 1, wherein the antibody is afucosylated, the heavy chain comprises the heavy chain sequence shown in SEQ ID NO: 25, and the light chain comprises the light chain sequence shown in SEQ ID NO:

26.

24. The pharmaceutical composition according to claim 6, wherein the antibody is afucosylated, the VH chain sequence comprises the VH sequence shown in SEQ ID NO: 1, and the VL chain sequence comprises the VL sequence shown in SEQ ID NO:

2.

25. The pharmaceutical composition according to claim 6, wherein the antibody is afucosylated, the heavy chain comprises the heavy chain sequence shown in SEQ ID NO: 25, and the light chain comprises the light chain sequence shown in SEQ ID NO:

26.

26. The pharmaceutical composition according to claim 17, wherein the antibody is afucosylated, the human Fc is wild-type human IgG1 Fc, the VH chain sequence comprises the VH sequence shown in SEQ ID NO: 1, and the VL chain sequence comprises the VL sequence shown in SEQ ID NO: 2.

Images