Anti-MS4A4A antibodies and methods of use thereof

Abstract

The present disclosure generally relates to compositions comprising antibodies (eg, monoclonal antibodies, antibody fragments, etc.) that specifically bind to MS4A4A polypeptides (eg, mammalian MS4A4A or human MS4A4A), and the use of such compositions in preventing, reducing risk, or treating individuals in need thereof.

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This application claims the benefit of U.S. Provisional Application No. 62 / 624,600, filed January 31, 2018, and U.S. Provisional Application No. 62 / 783,096, filed December 20, 2018, each of which is hereby incorporated by reference in its entirety.

[0003] Submission of sequence listings as ASCII text files

[0004] The following content, submitted as an ASCII text file, is incorporated herein by reference in its entirety: Computer Readable Form (CRF) of the Sequence Listing (File Name: 735022002140SEQLIST.TXT, Record Date: January 31, 2019, Size: 146KB). Technical Field

[0005] The present disclosure relates to anti-MS4A4A antibodies and therapeutic uses of such antibodies. Background Art

[0006] The transmembrane 4 domain subfamily A (MS4A) gene cluster is present on chromosome 11q12 and contains eighteen genes. The MS4A gene family encodes membrane proteins that typically have a four-transmembrane topology (Ishibashi et al., 2001, Gene, 265:87-93; Liang and Tedder, 2001, Genomics, 72:119-127; Efthymiou and Goate, 2017, Molecular Neurodegeneration, 12:43). The transmembrane domains are interconnected by an intracellular loop and two extracellular loops, and both the N-terminus and C-terminus are located in the cytosol. Most MS4A proteins share amino acid sequence homology (20-30% similarity) with MS4A1 (CD20), and the highest degree of sequence identity is found in the first three transmembrane domains. Among different MS4A proteins, the highly conserved motifs within these transmembrane regions indicate that the transmembrane domain plays an important general role in the function of MS4A proteins. The regions that vary most among MS4A proteins occur within their N-terminal and C-terminal cytoplasmic domains and a putative second extracellular loop (Ishibashi et al., 2001, Gene, 265:87-93), suggesting that these regions confer unique functional properties.

[0007] Although there is such diversity, the MS4A domain has some common elements. For example, a notable feature conserved in the MS4A protein (except MS4A8B and MS4A12) is the conservation of two cysteine ​​residues in the second extracellular loop of the putative disulfide bond. The N-terminal and C-terminal domains of the MS4A protein are also rich in proline residues, but its functional significance still needs to be clarified (Hulett et al., 2001, Genomics, 72: 119-127). However, the proline-rich region is generally involved in various cellular processes, such as cytoskeleton rearrangement, initiation of transcription, signal transduction cascades and association with the SH3 domain as part of the adapter system, thereby facilitating protein-protein interactions (Kay et al., 2000, FASEB J, 14: 231-241).

[0008] The MS4A family of proteins is relatively undefined in function, with some important exceptions: MS4A1 (CD20) is expressed exclusively in B lymphocytes, where the protein functions in signaling through the B cell antigen receptor and calcium influx. CD20 is the target of immunotherapeutic antibodies that can be used to deplete pathogenic B cells in chronic lymphocytic leukemia, lymphoma, autoimmune diseases, and in solid organ transplantation. MS4A2 (FcεRβ) is the signaling subunit of the high-affinity IgE receptor (FcεRI) and the low-affinity IgG receptor (FcεRIII) on mast cells, with a key role in hypersensitivity and allergic reactions. MS4A2 is an ITAM domain protein that amplifies signals through a high-affinity IgE receptor complex of four proteins. MS4A3 (Htm4) is expressed on the intracellular membranes of lymphoid and myeloid cells and acts as an adaptor protein in cell cycle regulation.

[0009] Although most MS4A family members have not been characterized, reports indicate that MS4A proteins can act as chemical sensors and chemoreceptors for a variety of exogenous and endogenous ligands (including fatty acids, peptides, and sulfated steroids), and are involved in mediating calcium influx, regulating endocytosis, trafficking, and can serve as adaptors of signal transduction complexes (Cruse et al., 2015, Mol Biol Cell, 26:1711-1727; Greer et al., 2016, Cell, 165:1734-1748; Eon Kuek et al., 2016, Cell, 165:1734-1748; Koslowski et al., 2008, Cancer Res, 68:3458-3466; Bubien et al., 1993; J Cell Biol, 121:1121-1132).

[0010] Certain MS4A genes are genetically associated with a variety of conditions and diseases, particularly neurodegenerative conditions. For example, genome-wide significant association analysis has identified the MS4A gene cluster located on chromosome 11q12 as one of the most significant Alzheimer's disease loci. One gene of particular interest identified is MS4A4A (Lambert et al., 2013, Nat Genet, 45:1452-1458; Hollingworth et al., 2011, Nat Genet, 43:429-435; Naj et al., 2011, Nat Genet, 43:436-441).

[0011] Therefore, there is a need for therapies that target MS4A4A, including antibodies that specifically bind to MS4A4A, and / or therapies that can modulate (e.g., inhibit or reduce; activate or increase) the activity of MS4A4A, such as by reducing or increasing MS4A4A protein levels or activity, to treat various diseases, disorders, and conditions associated with MS4A4A activity.

[0012] All references cited herein, including patent applications and publications, are hereby incorporated by reference in their entirety. Summary of the Invention

[0013] The present disclosure generally relates to anti-MS4A4A antibodies and methods of using such antibodies. The methods provided herein can be used to prevent, reduce the risk of, or treat a subject suffering from a neurodegenerative disease, disorder, or condition. In some embodiments, the present disclosure provides a method of preventing, reducing the risk of, or treating a subject suffering from a neurodegenerative disease, disorder, or condition selected from the group consisting of Alzheimer's disease, late-onset Alzheimer's disease, dementia, and cognitive impairment, comprising administering a therapeutically effective amount of an anti-MS4A4A antibody to a subject in need thereof. In some embodiments, the present disclosure provides a method for preventing, reducing the risk of, or treating a subject suffering from a disease, disorder, or condition associated with increased expression or activity of MS4A4A, comprising administering a therapeutically effective amount of an anti-MS4A4A antibody to a subject in need thereof. In some embodiments, the present disclosure provides a method for preventing, reducing the risk of, or treating a subject having a disease, disorder, or condition associated with decreased expression or activity of MS4A4A, the method comprising administering a therapeutically effective amount of an anti-MS4A4A antibody to a subject in need thereof.

[0014] In certain embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to a discontinuous MS4A4A epitope. In certain embodiments that may be combined with any of the preceding embodiments, the discontinuous MS4A4A epitope comprises two or more peptides, three or more peptides, four or more peptides, five or more peptides, six or more peptides, seven or more peptides, eight or more peptides, nine or more peptides, or ten or more peptides. In certain embodiments that may be combined with any of the preceding embodiments, each peptide comprises five or more, six or more, seven or more, eight or more, nine or more, 10 or more, 11 or more, 12 or more, 13 or more, 14 or more, 15 or more, 16 or more, 17 or more, 18 or more, 19 or more, 20 or more, 21 or more, 22 or more, 23 or more, 24 or more, 25 or more, 26 or more, 27 or more, 28 or more, 29 or more, or 30 or more amino acid residues of the amino acid sequence of SEQ ID NO: 1, the amino acid sequence of SEQ ID NO: 2, or the amino acid sequence of SEQ ID NO: 3; or corresponds to the amino acid sequence of SEQ ID NO: 1, the amino acid sequence of SEQ ID NO: 2, or the amino acid sequence of SEQ ID NO: 3. In some embodiments, the present invention comprises at least one amino acid residue in the mammalian MS4A4A protein having the amino acid sequence of NO: 3. In some embodiments, the present invention comprises at least one amino acid residue in the mammalian MS4A4A protein having the amino acid sequence of NO: 3.

[0015] In certain embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to a conformational epitope of MS4A4A. In certain embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to a linear epitope of MS4A4A.

[0016] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to the extracellular domain 1 (ECL1) of MS4A4A. In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acids within the amino acid sequence CMASNTYGSNPIS (SEQ ID NO: 289) of SEQ ID NO: 1. In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to the extracellular domain 2 (ECL2) of MS4A4A. In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acids within the amino acid sequence SFHHPYCNYYGNSNNCHGTMS (SEQ ID NO: 290) of SEQ ID NO: 1.

[0017] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to a region or epitope in human MS4A4A comprising amino acid residues 155-177 of human MS4A4A of SEQ ID NO: 1. In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acids within amino acid residues 155-177 of human MS4A4A of SEQ ID NO: 1. In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to a region or epitope in human MS4A4A comprising the amino acid sequence LAFYSFHHPYCNYYG (SEQ ID NO: 296). In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acid residues within the amino acid sequence LAFYSFHHPYCNYYG (SEQ ID NO: 296). In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to a region or epitope in human MS4A4A comprising the amino acid sequence FYSFHHPYCNYYGNS (SEQ ID NO: 297). In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acid residues within the amino acid sequence FYSFHHPYCNYYGNS (SEQ ID NO: 297). In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to a region or epitope in human MS4A4A comprising the amino acid sequence SFHHPYCNYYGNSNN (SEQ ID NO: 298). In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acid residues within the amino acid sequence SFHHPYCNYYGNSNN (SEQ ID NO: 298). In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to a region or epitope in human MS4A4A comprising the amino acid sequence HHPYCNYYGNSNNCH (SEQ ID NO: 299). In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acid residues within the amino acid sequence HHPYCNYYGNSNNCH (SEQ ID NO: 299).In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to a region or epitope in human MS4A4A comprising the amino acid sequence PYCNYYGNSNNCHGT (SEQ ID NO: 300). In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acid residues within the amino acid sequence PYCNYYGNSNNCHGT (SEQ ID NO: 300).

[0018] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure compete for binding to MS4A4A with one or more reference anti-MS4A4A antibodies selected from the group consisting of: 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15, 4A-16, 4A-17, 4A-25, 4A-26, 4A-239, 4A-225, and 4A-220, and any combination thereof. In some embodiments, the anti-MS4A4A antibody comprises a V domain of an antibody selected from the group consisting of: H and V L (For example, as shown in Tables 4B and 9B below): 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15, 4A-16, 4A-17, 4A-18, 4A-19, 4A-20, 4A-21, 4A-23, 4A-24. 4. 4A-201, 4A-202, 4A-203, 4A-204, 4A-205, 4A-206, 4A-207, 4A-208, 4A-209, 4A-21 0, 4A-213, 4A-214, 4A-216, 4A-217, 4A-219, 4A-25, 4A-26, 4A-239, 4A-225 and 4A-220.

[0019] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to an epitope of human MS4A4A that is identical to or overlaps with an epitope of MS4A4A bound by at least one reference antibody selected from the group consisting of: 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15, 4A-16, 4A-17 , 4A-18, 4A-19, 4A-20, 4A-21, 4A-23, 4A-24, 4A-201, 4A-202, 4A-203, 4A-204, 4A-205, 4A-206, 4A-207, 4A-208, 4A-209, 4A-210, 4A-213, 4A-214, 4A-216, 4A-217, 4A-219, 4A-25, 4A-26, 4A-239, 4A-225, and 4A-220, and any combination thereof. In some embodiments, the anti-MS4A4A antibody comprises a V domain of an antibody selected from the group consisting of: H and V L (For example, as shown in Tables 4B and 9B below): 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15, 4A-16, 4A-17, 4A-18, 4A-19, 4A-20, 4A-21, 4A-23, 4A-24. 4. 4A-201, 4A-202, 4A-203, 4A-204, 4A-205, 4A-206, 4A-207, 4A-208, 4A-209, 4A-21 0, 4A-213, 4A-214, 4A-216, 4A-217, 4A-219, 4A-25, 4A-26, 4A-239, 4A-225 and 4A-220.

[0020] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to an epitope of MS4A4A that is substantially the same as the epitope bound by at least one reference antibody selected from the group consisting of: 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15, 4A-16, 4A-17, 4A-18, 4A-29, 4A-30, 4A-31, 4A-32, 4A-33, 4A-34, 4A-35, 4A-36, 4A-37, 4A-38, 4A-39, 4A-40, 4A-41, 4A-42, 4A-43, 4A-44, 4A-45, 4A-46, 4A-47, 4A-48, 4A-49, 4A-50, 4A-51, 4A-52, 4A-53, 4A-54, 4A-55, 4A-56, 4A-57, 4A-58, 4A-59, 4A-60, 4A-61, 4A-62, 4A-63, 4A-64, 4A-65 19, 4A-20, 4A-21, 4A-23, 4A-24, 4A-201, 4A-202, 4A-203, 4A-204, 4A-205, 4A-206, 4A-207, 4A-208, 4A-209, 4A-210, 4A-213, 4A-214, 4A-216, 4A-217, 4A-219, 4A-25, 4A-26, 4A-239, 4A-225, and 4A-220, and any combination thereof. In some embodiments, the anti-MS4A4A antibody comprises a V domain of an antibody selected from the group consisting of: H and V L (For example, as shown in Tables 4B and 9B below): 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15, 4A-16, 4A-17, 4A-18, 4A-19, 4A-20, 4A-21, 4A-23, 4A-24. 4. 4A-201, 4A-202, 4A-203, 4A-204, 4A-205, 4A-206, 4A-207, 4A-208, 4A-209, 4A-21 0, 4A-213, 4A-214, 4A-216, 4A-217, 4A-219, 4A-25, 4A-26, 4A-239, 4A-225 and 4A-220.

[0021] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to a linear epitope on MS4A4A. In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acids within amino acid residues 1-64 of human MS4A4A. In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acids within amino acid residues 65-85 of human MS4A4A. In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acids within amino acid residues 86-98 of human MS4A4A. In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acids within amino acid residues 99-119 of human MS4A4A. In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acids within amino acid residues 120-137 of human MS4A4A. In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acids within amino acid residues 138-158 of human MS4A4A. In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acids within amino acid residues 159-179 of human MS4A4A. In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acids within amino acid residues 180-200 of human MS4A4A. In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acids within amino acid residues 201-239 of human MS4A4A. In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure bind to extracellular domain 1 (ECL1) of MS4A4A. In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to extracellular domain 2 (ECL2) of MS4A4A. In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to the ECL1 domain and the ECL2 domain of MS4A4A.

[0022] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A of the present disclosure is an isolated antibody that binds to human MS4A4A, wherein the antibody comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises: HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 4-19; HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 20-37; and HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 38-55; and the light chain variable region comprises: HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 56-74; HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 75-85; and HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 86-100.

[0023] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A of the present disclosure is an isolated antibody that binds to human MS4A4A, wherein the antibody comprises a heavy chain variable region, wherein the heavy chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 101-120.

[0024] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A of the present disclosure is an isolated antibody that binds to human MS4A4A, wherein the antibody comprises a light chain variable region, wherein the light chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 121-139.

[0025] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A of the present disclosure is an isolated antibody that binds to human MS4A4A, wherein the antibody comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 101-120; and wherein the light chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 121-139.

[0026] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A of the present disclosure is an isolated antibody that binds to human MS4A4A, wherein the antibody comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises: HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 142-153; HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 154-167; and HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 168-182; and the light chain variable region comprises: HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 183-196; HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 197-209; and HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 210-224.

[0027] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A of the present disclosure is an isolated antibody that binds to human MS4A4A, wherein the antibody comprises a heavy chain variable region, wherein the heavy chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 225-239.

[0028] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A of the present disclosure is an isolated antibody that binds to human MS4A4A, wherein the antibody comprises a light chain variable region, wherein the light chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 240-254.

[0029] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A of the present disclosure is an isolated antibody that binds to human MS4A4A, wherein the antibody comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 225-239; and wherein the light chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 240-254.

[0030] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A of the present disclosure is an isolated antibody that binds to human MS4A4A, wherein the antibody comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises: HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 304-309; HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 310-315; and HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 316-321; and the light chain variable region comprises: HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 322-327; HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 328-333; and HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 334-339.

[0031] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A of the present disclosure is an isolated antibody that binds to human MS4A4A, wherein the antibody comprises a heavy chain variable region, wherein the heavy chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 340-345.

[0032] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A of the present disclosure is an isolated antibody that binds to human MS4A4A, wherein the antibody comprises a light chain variable region, wherein the light chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 346-351.

[0033] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A of the present disclosure is an isolated antibody that binds to human MS4A4A, wherein the antibody comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 340-345; and wherein the light chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 346-351.

[0034] In certain embodiments that may be combined with any of the preceding embodiments, the MS4A4A protein is a mammalian or human protein. In certain embodiments that may be combined with any of the preceding embodiments, the MS4A4A protein is a wild-type protein. In certain embodiments that may be combined with any of the preceding embodiments, the MS4A4A protein is a naturally occurring variant. In certain embodiments that may be combined with any of the preceding embodiments, the antibody is an agonist antibody, wherein the antibody induces one or more MS4A4A activities. In certain embodiments that may be combined with any of the preceding embodiments, the isolated antibody induces, enhances, or maintains MS4A4A clusters on the surface of a cell. In certain embodiments that may be combined with any of the preceding embodiments, the antibody is an antagonist antibody, wherein the antibody inhibits or reduces one or more MS4A4A activities. In certain embodiments that may be combined with any of the preceding embodiments, the isolated antibody reduces or inhibits MS4A4A clusters on the surface of a cell.

[0035] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure reduce or inhibit the interaction or binding of MS4A4A and at least one MS4A4A ligand or binding partner.

[0036] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure increase or enhance the interaction or binding of MS4A4A and at least one MS4A4A ligand or binding partner.

[0037] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure increase soluble TREM2 levels. In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure increase membrane TREM2 levels. In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure increase soluble TREM2 levels and plasma membrane / cell surface TREM2 levels.

[0038] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure lower or reduce the level of M2 cell surface markers in myeloid cells (e.g., macrophages). In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure lower or reduce the cell surface levels of CD200R, Dectin-1, and / or CD163 in myeloid cells (e.g., macrophages).

[0039] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure enhance or increase the formation of signaling complexes. In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure enhance or increase the formation of signaling complexes associated with adaptor proteins encoding ITAMs. In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure enhance or increase the formation of inhibitory signaling complexes. In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure enhance or increase the formation of inhibitory signaling complexes associated with adaptor proteins encoding ITIMs.

[0040] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure inhibit (e.g., block) or reduce the formation of signaling complexes. In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure inhibit (e.g., block) or reduce the formation of signaling complexes associated with adaptor proteins encoding ITAMs. In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure inhibit (e.g., block) or reduce the formation of inhibitory signaling complexes. In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure inhibit (e.g., block) or reduce the formation of inhibitory signaling complexes associated with adaptor proteins encoding ITIMs.

[0041] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure inhibit (e.g., block) or reduce the formation of MS4A4A homo-oligomeric cell surface protein complexes by: a) reducing the effective level of MS4A4A available for MS4A4A homo-oligomeric complex formation; b) blocking one or more sites on MS4A4A required for MS4A4A homo-oligomeric complex formation; c) preventing one or more post-translational events on MS4A4A required for MS4A4A homo-oligomeric complex formation and / or correct processing and / or cellular localization of MS4A4A; d) inducing degradation of MS4A4A; e) altering the conformation of MS4A4A, or any combination thereof.

[0042] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure increase or enhance the formation of MS4A4A homo-oligomeric cell surface protein complexes by: a) increasing the effective level of MS4A4A available for MS4A4A homo-oligomeric complex formation; b) stabilizing one or more of the sites on MS4A4A required for MS4A4A homo-oligomeric complex formation; c) maintaining cell surface expression of MS4A4A to allow homo-oligomeric complex formation and / or correct processing and / or maintaining correct cellular localization of MS4A4A; d) reducing degradation of MS4A4A; e) altering the conformation of MS4A4A, or any combination thereof.

[0043] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure inhibit (e.g., block) or reduce the formation of MS4A4A hetero-oligomeric cell surface protein complexes with one or more signaling adaptor proteins by: a) reducing the effective level of MS4A4A available for MS4A4A hetero-oligomeric complex formation; b) blocking one or more sites on MS4A4A required for MS4A4A hetero-oligomeric complex formation; c) preventing one or more post-translational events on MS4A4A required for MS4A4A hetero-oligomeric complex formation and / or correct processing and / or cellular localization of MS4A4A; d) inducing degradation of MS4A4A; e) altering the conformation of MS4A4A, or any combination thereof.

[0044] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure increase or enhance the formation of MS4A4A hetero-oligomeric cell surface protein complexes with one or more signaling adaptor proteins by: a) increasing the effective level of MS4A4A available for MS4A4A hetero-oligomeric complex formation; b) stabilizing one or more sites on MS4A4A required for MS4A4A hetero-oligomeric complex formation; c) maintaining cell surface expression of MS4A4A to allow MS4A4A hetero-oligomeric complex formation and / or correct processing and / or maintaining correct cellular localization of MS4A4A; d) reducing degradation of MS4A4A; e) altering the conformation of MS4A4A, or any combination thereof.

[0045] In certain embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibody is a human antibody, a humanized antibody, a bispecific antibody, a monoclonal antibody, a multivalent antibody, a conjugated antibody, or a chimeric antibody.

[0046] In certain embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibody is a bispecific antibody that recognizes a first antigen and a second antigen. In certain embodiments that may be combined with any of the preceding embodiments, the first antigen is MS4A4A and the second antigen is an antigen that facilitates transport across the blood-brain barrier. In certain embodiments that may be combined with any of the preceding embodiments, the second antigen is selected from the group consisting of MS4A4A, transferrin receptor (TR), insulin receptor (HIR), insulin-like growth factor receptor (IGFR), low-density lipoprotein receptor-related proteins 1 and 2 (LPR-1 and 2), diphtheria toxin receptor, CRM 197, llama single domain antibody, TMEM30 (A), protein transduction domain, TAT, Syn-B, penetrant peptide, polyarginine peptide, angiopep, basic immunoglobulin (basigin), Glut1 and CD98hc and ANG1005.

[0047] In some embodiments that may be combined with any of the preceding embodiments, the antibody is a monoclonal antibody. In some embodiments that may be combined with any of the preceding embodiments, the antibody is a human antibody. In some embodiments that may be combined with any of the preceding embodiments, the antibody is a humanized antibody. In some embodiments that may be combined with any of the preceding embodiments, the antibody is a bispecific antibody. In some embodiments that may be combined with any of the preceding embodiments, the antibody is a multivalent antibody. In some embodiments that may be combined with any of the preceding embodiments, the antibody is a chimeric antibody. In some embodiments that may be combined with any of the preceding embodiments, the antibody is of the IgG, IgM, or IgA class. In some embodiments, the antibody is of the IgG class and has an IgG1, IgG2, or IgG4 isotype. In certain embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibody is an antibody fragment that binds to an epitope comprising amino acid residues on human MS4A4A or mammalian MS4A4A protein. In certain embodiments that may be combined with any of the preceding embodiments, the fragment is a Fab, Fab', Fab'-SH, F(ab')2, Fv, or scFv fragment.

[0048] Other aspects of the present disclosure relate to an isolated (e.g., monoclonal) anti-MS4A4A antibody, wherein the anti-MS4A4A antibody comprises at least one, two, three, four, five, or six HVRs of an antibody selected from the group consisting of: 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15, 4A-16, 4A-17, 4A-18, 4A-19, 4A-20, 4A-21, 4A-23, 4A-24, 4A-201, 4A-202, 4A-203, 4A-204, 4A-205, 4A-206, 4A-207, 4A -208, 4A-209, 4A-210, 4A-213, 4A-214, 4A-216, 4A-217, 4A-219, 4A-25, 4A-26, 4A-239, 4A-225 and 4A-220. In some embodiments, an anti-MS4A4A antibody comprises six HVRs of an antibody selected from the group consisting of: 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15, 4A-16, 4A-17, 4A-18, 4A-19, 4A-20, 4A-21, 4A-22, 4A-23, 4A-24, 4A-25, 4A-26, 4A-27, 4A-28, 4A-29, 4A-30, 4A-31, 4A-32, 4A-33, 4A-34, 4A-35, 4A-36, 4A-37, 4A-38, 4A-39, 4A-40, 4A-41, 4A-42 In some embodiments, the anti-MS4A4A antibody comprises a V domain of an antibody selected from the group consisting of: H and V L: 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15, 4A-16, 4A-17, 4A-18, 4A-19, 4A-20, 4A-21, 4A-23, 4A-24, 4A-201 , 4A-202, 4A-203, 4A-204, 4A-205, 4A-206, 4A-207, 4A-208, 4A-209, 4A-210, 4A-213, 4A-214, 4A-216, 4A-217, 4A-219, 4A-25, 4A-26, 4A-239, 4A-225, and 4A-220. In some embodiments, the anti-MS4A4A antibody comprises a V domain of an antibody selected from the group consisting of: H and V L (For example, as shown in Tables 4B and 9B below): 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15, 4A-16, 4A-17, 4A-18, 4A-19, 4A-20, 4A-21, 4A-23, 4A-24. 4. 4A-201, 4A-202, 4A-203, 4A-204, 4A-205, 4A-206, 4A-207, 4A-208, 4A-209, 4A-21 0, 4A-213, 4A-214, 4A-216, 4A-217, 4A-219, 4A-25, 4A-26, 4A-239, 4A-225 and 4A-220.

[0049] Other aspects of the present disclosure relate to an isolated nucleic acid comprising a nucleic acid sequence encoding an anti-MS4A4A antibody according to any of the preceding embodiments. Other aspects of the present disclosure relate to a vector comprising a nucleic acid according to any of the preceding embodiments. Other aspects of the present disclosure relate to an isolated host cell comprising a vector according to any of the preceding embodiments. Other aspects of the present disclosure relate to a method of producing an anti-MS4A4A antibody, the method comprising culturing the host cell according to any of the preceding embodiments, thereby producing the anti-MS4A4A antibody. In certain embodiments, the method further comprises recovering the anti-MS4A4A antibody produced by the host cell. Other aspects of the present disclosure relate to an isolated anti-MS4A4A antibody produced by the method according to any of the preceding embodiments. Other aspects of the present disclosure relate to a pharmaceutical composition comprising the anti-MS4A4A antibody according to any of the preceding embodiments and a pharmaceutically acceptable carrier.

[0050] In certain embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibody specifically binds to human MS4A4A, mouse MS4A4A, cynomolgus monkey MS4A4A, or a combination thereof.

[0051] Some aspects of the present disclosure provide an isolated antibody that binds to human MS4A4A, wherein the antibody increases soluble TREM2 levels, increases plasma membrane or cell surface TREM2 levels, or increases soluble TREM2 and increases plasma membrane or cell surface TREM2 levels in myeloid cells. In some embodiments, the antibody increases soluble TREM2 levels by at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or at least 100% compared to a control antibody of the same isotype. In some embodiments, the antibody increases plasma membrane or cell surface TREM2 levels by at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 100%, at least 105%, at least 110%, at least 115%, at least 120%, at least 125%, at least 130%, at least 135%, at least 140%, at least 145%, or at least 150% compared to a control antibody of the same isotype. In some embodiments, the antibody reduces expression of an M2 cell surface receptor / marker on myeloid cells.

[0052] Other aspects of the present disclosure provide isolated antibodies that bind to human MS4A4A, wherein the antibodies reduce expression of M2 cell surface receptors / markers on myeloid cells. In some embodiments, the antibodies reduce expression of one or more of CD200R, Dectin-1, CD163, CD14, and SIRPα on myeloid cells.

[0053] In some embodiments of any of the foregoing aspects and embodiments of the present disclosure, the antibody increases the viability of macrophages.

[0054] Other aspects of the present disclosure provide an isolated anti-MS4A4A antibody, wherein the anti-MS4A4A antibody comprises at least one, two, three, four, five, or six HVRs of an antibody selected from the group consisting of: 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15, 4A-16, 4A-17, 4A-18, 18. 4A-19, 4A-20, 4A-21, 4A-23, 4A-24, 4A-201, 4A-202, 4A-203, 4A-204, 4A-205, 4A-206, 4A-207, 4A- 208, 4A-209, 4A-210, 4A-213, 4A-214, 4A-216, 4A-217, 4A-219, 4A-25, 4A-26, 4A-239, 4A-225 and 4A-220.

[0055] Other aspects of the present disclosure provide an isolated antibody that binds to human MS4A4A, wherein the antibody comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises: (a) an HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 4-19, or an amino acid sequence with at least about 95% homology to SEQ ID NOs: 4-19, an amino acid sequence selected from the group consisting of SEQ ID NOs: 142-153, or an amino acid sequence with at least about 95% homology to SEQ ID NOs: 142-153, or an amino acid sequence selected from the group consisting of SEQ ID NOs: 304-309, or an amino acid sequence with at least about 95% homology to SEQ ID NOs: 304-309; and (b) an HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 20-37, or an amino acid sequence with at least about 95% homology to SEQ ID NOs: 20-37, an amino acid sequence selected from the group consisting of SEQ ID NOs: 154-167, or an amino acid sequence with at least about 95% homology to SEQ ID NOs: NOs: 154-167, or an amino acid sequence at least about 95% homologous to the group consisting of SEQ ID NOs: 310-315; and (c) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 38-55, or an amino acid sequence at least about 95% homologous to SEQ ID NOs: 38-55, an amino acid sequence selected from the group consisting of SEQ ID NOs: 168-182, or an amino acid sequence at least about 95% homologous to SEQ ID NOs: 168-182, or an amino acid sequence selected from the group consisting of SEQ ID NOs: 316-321, or an amino acid sequence at least about 95% homologous to SEQ ID NOs: 316-321; and wherein the light chain variable region comprises: (d) HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 56-74, or an amino acid sequence at least about 95% homologous to SEQ ID NOs: an amino acid sequence selected from the group consisting of SEQ ID NOs: 56-74 or an amino acid sequence having at least about 95% homology to SEQ ID NOs: 183-196, or an amino acid sequence selected from the group consisting of SEQ ID NOs: 322-327 or an amino acid sequence having at least about 95% homology to SEQ ID NOs: 322-327;(e) HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 75-85, or an amino acid sequence at least about 95% homologous to SEQ ID NOs: 75-85, an amino acid sequence selected from the group consisting of SEQ ID NOs: 197-209, or an amino acid sequence at least about 95% homologous to SEQ ID NOs: 197-209, or an amino acid sequence selected from the group consisting of SEQ ID NOs: 328-333, or an amino acid sequence at least about 95% homologous to SEQ ID NOs: 328-333; and (f) HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 86-100, or an amino acid sequence at least about 95% homologous to SEQ ID NOs: 86-100, an amino acid sequence selected from the group consisting of SEQ ID NOs: 210-224, or an amino acid sequence at least about 95% homologous to SEQ ID NOs: 210-224, or an amino acid sequence selected from the group consisting of SEQ ID NOs: The amino acid sequence of the group consisting of SEQ ID NOs: 334-339 or an amino acid sequence having at least about 95% homology to SEQ ID NOs: 334-339.

[0056] In some embodiments of any of the aforementioned aspects and embodiments of the present disclosure, the heavy chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 101-120, or an amino acid sequence that is at least 90% identical to SEQ ID NOs: 101-120, an amino acid sequence selected from the group consisting of SEQ ID NOs: 225-239, or an amino acid sequence that is at least 90% identical to SEQ ID NOs: 225-239, or an amino acid sequence selected from the group consisting of SEQ ID NOs: 340-345, or an amino acid sequence that is at least 90% identical to SEQ ID NOs: 340-345. In some embodiments of any of the aforementioned aspects and embodiments of the present disclosure, the light chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 121-139, or an amino acid sequence that is at least 90% identical to SEQ ID NOs: 121-139, an amino acid sequence selected from the group consisting of SEQ ID NOs: 240-254, or an amino acid sequence that is at least 90% identical to SEQ ID NOs: 240-254, or an amino acid sequence selected from the group consisting of SEQ ID NOs: 346-351, or an amino acid sequence that is at least 90% identical to SEQ ID NOs: 346-351.

[0057] Some aspects of the present disclosure provide an isolated antibody that binds to human MS4A4A, wherein the antibody comprises a heavy chain variable region and a light chain variable region, wherein: (a) the heavy chain variable region comprises HVR-H1 comprising the amino acid sequence of SEQ ID NO: 15, or an amino acid sequence at least about 95% homologous to SEQ ID NO: 15, HVR-H2 comprising the amino acid sequence of SEQ ID NO: 32, or an amino acid sequence at least about 95% homologous to SEQ ID NO: 32, and HVR-H3 comprising the amino acid sequence of SEQ ID NO: 50, or an amino acid sequence at least about 95% homologous to SEQ ID NO: 50, and the light chain variable region comprises HVR-L1 comprising the amino acid sequence of SEQ ID NO: 69, or an amino acid sequence at least about 95% homologous to SEQ ID NO: 69, HVR-L2 comprising the amino acid sequence of SEQ ID NO: 82, or an amino acid sequence at least about 95% homologous to SEQ ID NO: 82, and HVR-L3 comprising the amino acid sequence of SEQ ID NO: NO:96, or an amino acid sequence at least about 95% homologous to SEQ ID NO:96; (b) the heavy chain variable region comprises HVR-H1 comprising the amino acid sequence of SEQ ID NO:18, or an amino acid sequence at least about 95% homologous to SEQ ID NO:18, HVR-H2 comprising the amino acid sequence of SEQ ID NO:35, or an amino acid sequence at least about 95% homologous to SEQ ID NO:35, and HVR-H3 comprising the amino acid sequence of SEQ ID NO:53, or an amino acid sequence at least about 95% homologous to SEQ ID NO:53, and the light chain variable region comprises HVR-L1 comprising the amino acid sequence of SEQ ID NO:72, or an amino acid sequence at least about 95% homologous to SEQ ID NO:72, HVR-L2 comprising the amino acid sequence of SEQ ID NO:77, or an amino acid sequence at least about 95% homologous to SEQ ID NO:77, and HVR-L3 comprising the amino acid sequence of SEQ ID NO: The amino acid sequence of SEQ ID NO:99 or an amino acid sequence having at least about 95% homology to SEQ ID NO:99;or (c) the heavy chain variable region comprises HVR-H1 comprising the amino acid sequence of SEQ ID NO: 143, or an amino acid sequence at least about 95% homologous to SEQ ID NO: 143, HVR-H2 comprising the amino acid sequence of SEQ ID NO: 155, or an amino acid sequence at least about 95% homologous to SEQ ID NO: 155, and HVR-H3 comprising the amino acid sequence of SEQ ID NO: 169, or an amino acid sequence at least about 95% homologous to SEQ ID NO: 169, and the light chain variable region comprises HVR-L1 comprising the amino acid sequence of SEQ ID NO: 184, or an amino acid sequence at least about 95% homologous to SEQ ID NO: 184, HVR-L2 comprising the amino acid sequence of SEQ ID NO: 198, or an amino acid sequence at least about 95% homologous to SEQ ID NO: 198, and HVR-L3 comprising the amino acid sequence of SEQ ID NO: 211, or an amino acid sequence at least about 95% homologous to SEQ ID NO: NO:211 has an amino acid sequence with at least about 95% homology.

[0058] In some embodiments of any of the foregoing aspects and embodiments of the present disclosure, the antibody binds to cynomolgus monkey MS4A4A.

[0059] In some aspects of the disclosure, the antibody competes for binding to human MS4A4A with one or more of the antibodies of any of the above embodiments. In some aspects, the antibody binds to substantially the same or overlapping epitope of MS4A4A as the antibody of any of the above embodiments.

[0060] In some embodiments of any of the foregoing aspects and embodiments of the present disclosure, the antibody binds to extracellular domain 1 of human MS4A4A. In some embodiments, the antibody binds to extracellular domain 2 of human MS4A4A. In some embodiments, the antibody binds to one or more amino acids within amino acid residues 155-177 of human MS4A4A of SEQ ID NO: 1. In some embodiments, the antibody binds to one or more amino acid residues selected from the group consisting of Y92, H161, Y164, N166, N170, T177, and M178 of SEQ ID NO: 1, or one or more amino acid residues on a mammalian MS4A4A protein corresponding to amino acid residues selected from the group consisting of Y92, H161, Y164, N166, N170, T177, and M178 of SEQ ID NO: 1. In some embodiments, the antibody reduces or inhibits the interaction or binding of MS4A4A with at least one MS4A4A ligand or binding partner. In some embodiments, the antibodies increase or enhance the interaction or binding of MS4A4A with at least one MS4A4A ligand or binding partner. In some embodiments, the antibodies reduce the decline in cognitive and behavioral function and improve cognitive and behavioral function.

[0061] In some embodiments of any of the above aspects and embodiments of the present disclosure, the antibody is a murine antibody, a human antibody, a humanized antibody, a bispecific antibody, a monoclonal antibody, a multivalent antibody, a conjugated antibody, or a chimeric antibody. In some embodiments, the antibody belongs to the IgG class, the IgM class, or the IgA class. In some embodiments, the antibody belongs to the IgG class and has an IgG1, IgG2, or IgG4 isotype. In some embodiments, the antibody is an antibody fragment. In some embodiments, the fragment is a Fab, Fab', Fab'-SH, F(ab')2, Fv, or scFv fragment.In some embodiments, the antibody further comprises: (a) an antigen that facilitates transport across the blood-brain barrier; (b) an antigen that facilitates transport across the blood-brain barrier, the antigen being selected from the group consisting of transferrin receptor (TR), insulin receptor (HIR), insulin-like growth factor receptor (IGFR), low-density lipoprotein receptor-related proteins 1 and 2 (LPR-1 and 2), diphtheria toxin receptor, CRM197, llama single domain antibody, TMEM 30 (A), protein transduction domain, TAT, Syn-B, transmembrane peptide, polyarginine peptide, vascular peptide and ANG1005; (c) a pathogen, the pathogen being selected from the group consisting of a pathogenic peptide or protein or a pathogenic nucleic acid, wherein the pathogenic nucleic acid is an antisense GGCCCC (G2C4) repeat expansion RNA, the pathogenic protein being selected from the group consisting of amyloid beta, oligomeric amyloid beta, amyloid beta plaques, amyloid precursor protein or fragments thereof, tau, IAPP, α-synuclein, Synuclein, TDP-43, FUS protein, C9orf72 (chromosome 9 open reading frame 72), c9RAN protein, prion protein, PrPSc, huntingtin protein, calcitonin, superoxide dismutase, ataxia, ataxia 1, ataxia 2, ataxia 3, ataxia 7, ataxia 8, ataxia 10, Lewy body, atrial natriuretic factor, islet amyloid polypeptide, insulin, apolipoprotein AI, serum amyloid A, medin, prolactin, transthyretin proteoglycans, lysozyme, β2-microglobulin, gelsolin, corneal epithelial protein, cystatin, immunoglobulin light chain AL, S-IBM protein, repeat-associated non-ATG (RAN) translation products, dipeptide repeat (DPR) peptides, glycine-alanine (GA) repeat peptides, glycine-proline (GP) repeat peptides, glycine-arginine (GR) repeat peptides, proline-alanine (PA) repeat peptides, ubiquitin, and proline-arginine (PR) repeat peptides; (d) expressed on immune cells (e) a ligand and / or protein, wherein the ligand and / or protein is selected from the group consisting of: CD40, OX40, ICOS, CD28, CD137 / 4-1BB, CD27, GITR, PD-L1, CTLA-4, PD-L2, PD-1, B7-H3, B7-H4, HVEM, BTLA, KIR, GAL9, TIM3, A2AR, LAG-3 and phosphatidylserine; and (e) a protein, lipid, polysaccharide or glycolipid expressed on one or more tumor cells.

[0062] Other aspects of the present disclosure relate to an isolated nucleic acid comprising a nucleic acid sequence encoding an antibody according to any of the above embodiments. Other aspects of the present disclosure relate to a vector comprising a nucleic acid according to the present disclosure. Other aspects of the present disclosure relate to an isolated host cell comprising a vector according to the present disclosure. Other aspects of the present disclosure relate to a method of producing an antibody that binds to human MS4A4A, the method comprising culturing the cells according to the present disclosure, thereby producing the antibody. In some embodiments, the method further comprises recovering the antibody produced by the cells. Other aspects of the present disclosure relate to a pharmaceutical composition comprising an antibody according to the present disclosure and a pharmaceutically acceptable carrier.

[0063] Other aspects of the present disclosure relate to a method for preventing, reducing the risk of, or treating a disease, disorder, or injury selected from the group consisting of Alzheimer's disease, late-onset Alzheimer's disease, and cognitive impairment, comprising administering a therapeutically effective amount of an antibody of the present disclosure to a subject in need thereof. Other aspects of the present disclosure relate to a method for preventing, reducing the risk of, or treating a disease, disorder, condition, or injury caused by or associated with overexpression or increased activity of MS4A4A, comprising administering a therapeutically effective amount of an antibody of the present disclosure to a subject in need thereof.

[0064] It should be understood that one, some or all of the properties of the various embodiments described herein can be combined to form other embodiments of the present disclosure. These and other aspects of the present disclosure will be apparent to those skilled in the art. These and other embodiments of the present disclosure are further described by the following specific embodiments. BRIEF DESCRIPTION OF THE DRAWINGS

[0065] The patent or application file contains at least one drawing executed in color. Copies of this patent or patent application publication with color drawing(s) will be provided by the U.S. Patent and Trademark Office upon request and payment of the necessary fee.

[0066] Figure 1A Shown are representative FACS plots of HEK293 cells transiently transfected with human MS4A4A (clear trace) and untransfected cells (hatched trace) stained with hybridoma supernatant containing anti-MS4A4A antibodies of the present disclosure. Figure 1B Shown are representative FACS plots of HEK293 cells transiently transfected with cynomolgus monkey MS4A4A (clear trace) and untransfected cells (hatched trace) stained with hybridoma supernatant containing anti-MS4A4A antibodies of the present disclosure. Figure 1CShown are representative FACS plots of HEK293 cells transiently transfected with human MS4A4A (clear trace) and untransfected cells (hatched trace) stained with anti-MS4A4A antibody 5C12 (BioLegend). Figure 1D Shown are representative FACS plots of HEK293 cells transiently transfected with cynomolgus MS4A4A (clear trace) and untransfected cells (hatched trace) stained with anti-MS4A4A antibody 5C12.

[0067] FIG2 shows 300.19 cells transfected with human MS4A4A and stained with anti-MS4A4A antibody 5C12 (Biolegend) ( Figure 2A ), K562 cells ( Figure 2B ) and U937 cells ( Figure 2C ) FAC analysis; 300.19 cells transfected with cynomolgus monkey MS4A4A and stained with hybridoma supernatant containing anti-MS4A4A of the present disclosure ( Figure 2D ), K562 cells ( Figure 2E ) and U937 cells ( Figure 2F 2 , the red trace on the left corresponds to untransfected cells, and the blue trace on the right corresponds to transfected cells.

[0068] Figure 3 shows FACS analysis of anti-MS4A4A antibodies of the present disclosure binding to untransfected 300.19 cells (grey shading), 300.19 cells transfected with human MS4A4A (blue trace), and 300.19 cells transfected with cynomolgus monkey MS4A4A (orange trace). Figure 3A Shown is a table representing anti-MS4A4A antibodies of the present disclosure. Figure 3B FACS plots are shown and correspond to Figure 3A Each anti-MS4A4A antibody of the present disclosure is shown.

[0069] Figure 4 shows FACS analysis of anti-MS4A4A antibodies of the present disclosure binding to untransfected U937 cells (grey shading) or to U937 cells transfected with human MS4A4A (blue trace). Figure 4A Shown is a table representing anti-MS4A4A antibodies of the present disclosure. Figure 4B FACS plots are shown and correspond to each anti-MS4A4A antibody of the present disclosure as shown in the table on the left side of the figure. DETAILED DESCRIPTION

[0070] The present disclosure relates to anti-MS4A4A antibodies (eg, monoclonal antibodies); and methods of making and using such antibodies; pharmaceutical compositions comprising such antibodies; nucleic acids encoding such antibodies; and host cells comprising nucleic acids encoding such antibodies.

[0071] The techniques and procedures described or referenced herein are well understood and generally adapted by those skilled in the art using conventional methods, e.g., widely employed methods such as those described in Sambrook et al., Molecular Cloning: A Laboratory Manual, 3rd ed. (2001) Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY; Current Protocols in Molecular Biology (FM Ausubel et al., eds., (2003); Monoclonal Antibodies: A Practical Approach (P. Shepherd and C. Dean, eds., Oxford University Press, 2000).

[0072] I. Definition

[0073] Unless otherwise indicated, the terms "MS4A4A" or "MS4A4A polypeptide" are used interchangeably herein to refer to any native MS4A4A from any vertebrate source, including mammals, such as primates (e.g., humans and cynomolgus monkeys) and rodents (e.g., mice and rats). In some embodiments, the term encompasses both wild-type sequences and naturally occurring variant sequences (e.g., splice variants or allelic variants). In some embodiments, the term encompasses "full-length," unprocessed MS4A4A as well as any form of MS4A4A resulting from processing in cells. In some embodiments, MS4A4A is human MS4A4A. In some embodiments, the amino acid sequence of an exemplary MS4A4A is Uniprot Accession No. Q96JQ5 as of December 1, 2001. In some embodiments, the amino acid sequence of an exemplary human MS4A4A is SEQ ID NO: 1.

[0074] The terms "anti-MS4A4A antibody," "antibody that binds to MS4A4A," and "antibody that specifically binds to MS4A4A" refer to an antibody that is capable of binding to MS4A4A with sufficient affinity to permit use as a diagnostic and / or therapeutic agent targeting MS4A4A. In one embodiment, the extent of binding of the anti-MS4A4A antibody to an unrelated, non-MS4A4A polypeptide is less than about 10% of the binding of the antibody to MS4A4A, as measured, for example, by radioimmunoassay (RIA). In certain embodiments, the antibody binds to MS4A4A with an affinity of <1 μM, <100 nM, <10 nM, <1 nM, <0.1 nM, <0.01 nM, or <0.001 nM (e.g., 10 -8 M or smaller, for example, from 10 -8 M to 10 -13 M, for example, from 10 -9 M to 10 -13 In certain embodiments, an anti-MS4A4A antibody binds to an epitope of MS4A4A that is conserved between MS4A4As of different species.

[0075] With respect to the binding of an antibody to a target molecule, the term "specifically binds to" a specific polypeptide or an epitope on a specific polypeptide target or "specifically binds to" a specific polypeptide or an epitope on a specific polypeptide target or "has specificity" for a specific polypeptide or an epitope on a specific polypeptide target means binding that is measurably different from non-specific interactions. Specific binding can be measured, for example, by measuring the binding of a molecule compared to the binding of a control molecule. For example, specific binding can be measured by competition with a control molecule similar to the target (e.g., an excess of unlabeled target). In this case, if the binding of the labeled target to the probe is competitively inhibited by an excess of unlabeled target, specific binding is indicated. As used herein, the term "specifically binds to" a specific polypeptide or an epitope on a specific polypeptide target or "specifically binds to" a specific polypeptide or an epitope on a specific polypeptide target or "has specificity" for a specific polypeptide or an epitope on a specific polypeptide target can be measured, for example, by measuring the binding of the molecule to the target with a KD of about 10 -4 M or smaller, 10 -5 M or smaller, 10 -6 M or smaller, 10 -7 M or smaller, 10 -8 M or smaller, 10 -9 M or smaller, 10 -10 M or smaller, 10 -11 M or smaller, 10 -12 M or smaller or KD in 10 -4 M to 10 -6 M or 10 -6 M to 10-10 M or 10 -7 M to 10 -9 The term "specific binding" refers to the binding of a molecule to a specific polypeptide or epitope on a specific polypeptide without substantially binding to any other polypeptide or polypeptide epitope.

[0076] The term "immunoglobulin" (Ig) is used interchangeably herein with "antibody." The term "antibody" herein is used in the broadest sense and specifically encompasses monoclonal antibodies, polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies) (including those formed from at least two intact antibodies), and antibody fragments, so long as they exhibit the desired biological activity.

[0077] "Native antibodies" are usually heterotetrameric glycoproteins of about 150,000 daltons composed of two identical light (L) chains and two identical heavy (H) chains. Each light chain is linked to a heavy chain by one covalent disulfide bond, while the number of disulfide linkages varies between the heavy chains of different immunoglobulin isotypes. Each heavy and light chain also has regularly spaced intrachain disulfide bridges. Each heavy chain has a variable domain (V) at one end. H ), followed by several constant domains. Each light chain has a variable domain at one end (V L ), with a constant domain at the other end; the constant domain of the light chain is aligned with the first constant domain of the heavy chain, and the variable domain of the light chain is aligned with the variable domain of the heavy chain. Specific amino acid residues are believed to form an interface between the light and heavy chain variable domains.

[0078] For the structure and properties of different classes of antibodies, see, e.g., Basic and Clinical Immunology, 8th edition, Daniel P. Stites, Abba I. Terr, and Tristram G. Parslow (eds.), Appleton & Lange, Norwalk, CT, 1994, page 71 and Chapter 6.

[0079] Light chains from any vertebrate species can be classified into one of two clearly distinct types, called kappa ("κ") and lambda ("λ"), based on the amino acid sequence of the constant domain of the light chain. Immunoglobulins can be classified into different classes, or isotypes, based on the amino acid sequence of the constant domain of their heavy chains (CH). There are five classes of immunoglobulins: IgA, IgD, IgE, IgG, and IgM, which have heavy chains called alpha ("α"), delta ("δ"), epsilon ("ε"), gamma ("γ"), and muon ("μ"), respectively. The gamma and alpha classes can be further divided into subclasses (isotypes) based on relatively minor differences in CH sequence and function, for example, humans express the following subclasses: IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2. The subunit structures and three-dimensional configurations of different classes of immunoglobulins are well known and generally described in, for example, Abbas et al., Cellular and Molecular Immunology, 4th ed. (WB Saunders Co., 2000).

[0080] The "variable region" or "variable domain" of an antibody (such as the anti-MS4A4A antibodies of the present disclosure) refers to the amino-terminal domain of the heavy or light chain of the antibody. The variable domains of the heavy and light chains may be referred to as "V H ” and “V L These domains are generally the most variable parts of an antibody (relative to other antibodies of the same class) and contain the antigen-binding site.

[0081] The term "variable" refers to the fact that certain segments of the variable domain differ extensively in sequence among antibodies (such as the anti-MS4A4A antibodies of the present disclosure). The variable domain mediates antigen binding and defines the specificity of a particular antibody for its particular antigen. However, variability is not evenly distributed throughout the variable domain. Instead, in both the light and heavy chain variable domains, variability is concentrated in three segments called hypervariable regions (HVRs). The more highly conserved portions of the variable domain are called framework regions (FRs). The variable domains of native heavy and light chains each contain four FR regions, which mostly adopt a β-sheet configuration, connected by three HVRs, which form loops connecting (and in some cases forming part of) the β-sheet structure. The HVRs in each chain are held together in close proximity by the FR regions and, together with the HVRs of the other chain, contribute to the formation of the antibody's antigen-binding site (see Kabat et al., Sequences of Immunological Interest, 5th ed., National Institute of Health, Bethesda, MD (1991)). The constant domains are not involved directly in binding an antibody to an antigen, but exhibit various effector functions, such as participation of the antibody in antibody-dependent cellular toxicity.

[0082] As used herein, the term "monoclonal antibody" refers to an antibody (such as the monoclonal anti-MS4A4A antibody of the present disclosure) obtained from a population of substantially homogeneous antibodies, i.e., the individual antibodies comprising the population are identical except for possible naturally occurring mutations and / or post-translational modifications (e.g., isomerization, amidation, etc.) that may be present in minor amounts. Monoclonal antibodies are highly specific (directed against a single antigenic site). In contrast to polyclonal antibody preparations, which typically include different antibodies directed against different determinants (epitopes), each monoclonal antibody is directed against a single determinant on the antigen. In addition to specificity, an advantage of monoclonal antibodies is that they are synthesized by hybridoma cultures and are not contaminated by other immunoglobulins. The modifier "monoclonal" indicates the characteristic of the antibody as being obtained from a substantially homogeneous population of antibodies and is not to be construed as requiring production of the antibody by any particular method. For example, the monoclonal antibodies used in accordance with the present disclosure can be prepared by a variety of techniques, including, for example, hybridoma methods, recombinant DNA methods, and techniques for producing human or human-like antibodies in animals that have some or all of the human immunoglobulin loci or genes encoding human immunoglobulin sequences.

[0083] The terms "full-length antibody," "intact antibody," or "complete antibody" are used interchangeably to refer to an antibody (such as the anti-MS4A4A antibodies of the present disclosure) in its substantially intact form, as compared to an antibody fragment. Specifically, complete antibodies include those having heavy and light chains comprising an Fc region. The constant domains may be native sequence constant domains (e.g., human native sequence constant domains) or amino acid sequence variants thereof. In some cases, an intact antibody may possess one or more effector functions.

[0084] "Antibody fragment" refers to a molecule other than an intact antibody that includes a portion of an intact antibody that binds to the antigen to which the intact antibody binds. Examples of antibody fragments include Fab, Fab', F(ab')2, and Fv fragments; diabodies; linear antibodies (see U.S. Patent 5,641,870, Example 2; Zapata et al., Protein Eng. 8(10):1057-1062 (1995)); single-chain antibody molecules formed from antibody fragments; and multispecific antibodies.

[0085] Papain digestion of antibodies (such as the anti-MS4A4A antibodies of the present disclosure) produces two identical antigen-binding fragments called "Fab" fragments and a residual "Fc" fragment (a name reflecting the ability to crystallize readily). The Fab fragment consists of an entire light chain and the variable region domain (V) of the heavy chain. H ) and the first constant domain of a heavy chain (C H 1) Composition. Each Fab fragment is monovalent with respect to antigen binding, i.e., it has a single antigen binding site. Pepsin treatment of an antibody produces a single large F(ab')2 fragment that roughly corresponds to two disulfide-linked Fab fragments with different antigen binding activities and is still able to cross-link antigen. Fab' fragments differ from Fab fragments in that the C H The carboxyl terminus of the constant domain has several additional residues, including one or more cysteines from the antibody hinge region. Fab'-SH is the designation herein for Fab' in which one or more cysteine ​​residues of the constant domains bear a free thiol group. F(ab')2 antibody fragments originally were produced as pairs of Fab' fragments that have hinge cysteines between them. Other chemical conjugates of antibody fragments are also known.

[0086] The Fc fragment comprises the carboxyl terminal portions of two heavy chains held together by disulfide bonds.The effector functions of an antibody are determined by the sequence of the Fc region, which is also recognized by Fc receptors (FcR) present on certain cell types.

[0087] "Functional fragments" of antibodies (such as the anti-MS4A4A antibodies of the present disclosure) include a portion of an intact antibody, generally including the antigen-binding or variable region of the intact antibody, or the Fc region of an antibody that retains or has modified FcR binding ability. Examples of antibody fragments include linear antibodies, single-chain antibody molecules, and multispecific antibodies formed from antibody fragments.

[0088] The term "double body" refers to the use of V H domain and V L Small antibody fragments prepared by constructing sFv fragments (see the preceding paragraph) with short linkers (about 5-10 residues) between the variable domains to allow interchain, rather than intrachain, pairing of the variable domains, thereby generating bivalent fragments, i.e., fragments with two antigen-binding sites. Bispecific diabodies are heterodimers of two "cross-linked" sFv fragments, in which the V domains of the two antibodies are bound together. H domain and V L Domains are present on different polypeptide chains.

[0089] As used herein, "chimeric antibody" refers to an antibody (immunoglobulin) (such as the chimeric anti-MS4A4A antibody of the present disclosure) in which a portion of the heavy chain and / or light chain is identical or homologous to a corresponding sequence in an antibody derived from a particular species or belonging to a particular antibody class or subclass, while the remainder of one or more chains is identical or homologous to a corresponding sequence in an antibody derived from another species or belonging to another antibody class or subclass, as well as fragments of such antibodies, so long as they exhibit the desired biological activity. The chimeric antibodies contemplated herein include An antibody, wherein the antigen binding region of the antibody is derived from, for example, an antibody generated by immunizing a macaque monkey with an antigen of interest. As used herein, "humanized antibody" is used as a subset of "chimeric antibody."

[0090] "Humanized" forms of non-human (e.g., murine) antibodies (such as humanized forms of the anti-MS4A4A antibodies of the present disclosure) are chimeric antibodies comprising amino acid residues from non-human HVRs and amino acid residues from human FRs. In certain embodiments, a humanized antibody will comprise substantially all of at least one, and typically two, variable domains, in which all or substantially all of the HVRs (e.g., CDRs) correspond to those of a non-human antibody and all or substantially all of the FRs correspond to those of a human antibody. A humanized antibody optionally may comprise at least a portion of an antibody constant region derived from a human antibody. A "humanized form" of an antibody (e.g., a non-human antibody) refers to an antibody that has undergone humanization.

[0091] A "human antibody" is an antibody having an amino acid sequence corresponding to an antibody produced by a human (such as the anti-MS4A4A antibodies of the present disclosure) and / or has been prepared using any of the techniques for making human antibodies as disclosed herein. This definition of a human antibody specifically excludes humanized antibodies that contain non-human antigen-binding residues. Human antibodies can be produced using a variety of techniques known in the art, including phage display libraries and yeast display libraries. Human antibodies can be produced by administering an antigen to a transgenic animal that has been modified to produce such antibodies in response to an antigenic challenge, but whose endogenous loci have been disabled (e.g., an immunized xenomouse), as well as by human B-cell hybridoma technology.

[0092] The terms "hypervariable region," "HVR," or "HV," when used herein, refer to regions of an antibody variable domain (such as the variable domains of the anti-MS4A4A antibodies of the present disclosure) whose sequences are hypervariable and / or form structurally defined loops. Generally, an antibody comprises six HVRs: H There are three (H1, H2, H3), and V L There are three of these six HVRs (L1, L2, and L3). In native antibodies, H3 and L3 show the greatest diversity of the six HVRs, and H3 in particular is believed to play a unique role in conferring fine specificity to antibodies. Naturally occurring camelid antibodies composed solely of heavy chains are functional and stable in the absence of light chains.

[0093] Many HVR descriptions are in use and are encompassed herein. In some embodiments, HVR can be a Kabat complementary determining region (CDR) based on sequence variability, and is the most commonly used (Kabat et al., supra). In some embodiments, HVR can be a Chothia CDR. And Chothia refers to the position of the structural loop (Chothia and Lesk J. Mol. Biol. 196:901-917 (1987)). In some embodiments, HVR can be an AbM HVR. AbM HVR represents a compromise between Kabat CDR and Chothia structural loops, and is used by the AbM antibody modeling software of Oxford Molecular. In some embodiments, HVR can be a "contact" HVR. "Contact" HVR is based on the analysis of available complex crystal structures. The residue annotations from each of these HVRs are as follows.

[0094]

[0095] An HVR may comprise an "extended HVR" as follows: 24-36 or 24-34 (L1), 46-56 or 50-56 (L2), and 89-97 or 89-96 (L3) in VL and 26-35 (H1), 50-65 or 49-65 (preferred embodiments) (H2), and 93-102, 94-102, or 95-102 (H3) in VH. For each of these extended HVR definitions, the variable domain residues are numbered according to Kabat et al. (supra).

[0096] "Framework" or "FR" residues are those variable domain residues other than the HVR residues as herein defined.

[0097] As used herein, an "acceptor human framework" is a construct comprising a V derived from a human immunoglobulin framework or a human consensus framework. L or V H The acceptor human framework "derived from" a human immunoglobulin framework or a human consensus framework may comprise the same amino acid sequence thereof, or it may comprise pre-existing amino acid sequence changes. In some embodiments, the number of pre-existing amino acid changes is 10 or less, 9 or less, 8 or less, 7 or less, 6 or less, 5 or less, 4 or less, 3 or less, or 2 or less. If pre-existing amino acid changes are present in VH, preferably those changes occur in only three, two, or one of positions 71H, 73H, and 78H; for example, the amino acid residues at those positions may be 71A, 73T, and / or 78A. In one embodiment, the VL acceptor human framework is identical in sequence to the V L The framework sequences are identical to human immunoglobulin framework sequences or human consensus framework sequences.

[0098] The "human consensus framework" is a L or V H The framework sequences are selected to represent the most common amino acid residues. L or V H The sequence is selected from a subset of variable domain sequences. Generally, the subset of sequences is that described by Kabat et al., Sequences of Proteins of Immunological Interest, 5th ed., Public Health Service, National Institutes of Health, Bethesda, MD (1991). Examples include: L , the subgroup may be subgroup κI, κII, κIII or κIV as described by Kabat et al. (supra). H, the subgroup can be subgroup I, subgroup II or subgroup III as described by Kabat et al. (supra).

[0099] An "amino acid modification" at a designated position of, for example, an anti-MS4A4A antibody of the present disclosure refers to a substitution or deletion of the designated residue, or an insertion of at least one amino acid residue adjacent to the designated residue. An insertion "adjacent" to a designated residue means an insertion within one to two residues of the designated residue. The insertion can be at the N-terminus or C-terminus of the designated residue. Preferred amino acid modifications herein are substitutions.

[0100] "Affinity matured" antibodies (such as the affinity matured anti-MS4A4A antibodies of the present disclosure) are antibodies that have one or more alterations in one or more HVRs of the antibody that result in an improvement in the affinity of the antibody for the antigen compared to a parent antibody that does not have one or more of those alterations. In one embodiment, the affinity matured antibody has nanomolar or even picomolar affinities for the target antigen. Affinity matured antibodies are produced by procedures known in the art. For example, Marks et al., Bio / Technology 10:779-783 (1992) describe affinity matured antibodies by V H domain and V L Affinity maturation by domain shuffling. Random mutagenesis of HVR and / or framework residues is described, for example, in Barbas et al., Proc Nat. Acad. Sci. USA 91:3809-3813 (1994); Schier et al., Gene 169:147-155 (1995); Yelton et al., J. Immunol. 155:1994-2004 (1995); Jackson et al., J. Immunol. 154(7):3310-9 (1995); and Hawkins et al., J. Mol. Biol. 226:889-896 (1992).

[0101] "Fv" is the smallest antibody fragment that contains complete antigen recognition and binding sites. The fragment consists of a dimer of a heavy chain and a light chain variable region domain that are tightly and non-covalently associated. Six hypervariable loops (3 loops each from the H and L chains) are produced by the folding of these two domains. These six hypervariable loops contribute to the amino acid residues for antigen binding and confer antigen binding specificity to the antibody. However, even a single variable domain (or only half of an Fv comprising three HVRs specific for an antigen) has the ability to recognize and bind antigens, although the affinity is less than that of the entire binding site.

[0102] "Single-chain Fv", also abbreviated as "sFv" or "scFv", is an antibody fragment comprising the VH and VL antibody domains linked into a single polypeptide chain. Preferably, the sFv polypeptide also comprises a VH and V L A polypeptide linker between the domains that enables the sFv to form the desired structure for antigen binding.

[0103] Antibody "effector functions" refer to those biological activities attributable to the Fc region (a native sequence Fc region or amino acid sequence variant Fc region) of an antibody, and vary with the antibody isotype.

[0104] The term "Fc region" herein is used to define the C-terminal region of an immunoglobulin heavy chain, including native sequence Fc regions and variant Fc regions. Although the boundaries of the Fc region of an immunoglobulin heavy chain may vary, the human IgG heavy chain Fc region is typically defined as the sequence segment from the Cys226 position or from the amino acid residue at Pro230 to their carboxyl termini. The C-terminal lysine (residue 447 according to the EU numbering system) in the Fc region can be removed, for example, during the production or purification of the antibody or by recombinant engineering of the nucleic acid encoding the heavy chain of the antibody. Therefore, the composition of a complete antibody can include antibody groups in which all K447 residues are removed, antibody groups in which K447 residues are not removed, and antibody groups with and without a mixture of antibodies containing K447 residues. Native sequence Fc regions suitable for antibodies of the present disclosure include human IgG1, IgG2, IgG3, and IgG4.

[0105] A "native sequence Fc region" comprises an amino acid sequence identical to the amino acid sequence of an Fc region found in nature. Native sequence human Fc regions include native sequence human IgG1 Fc regions (non-A and A allotypes); native sequence human IgG2 Fc regions; native sequence human IgG3 Fc regions; and native sequence human IgG4 Fc regions, as well as naturally occurring variants thereof.

[0106] A "variant Fc region" comprises an amino acid sequence that differs from a native sequence Fc region by virtue of at least one amino acid modification (preferably one or more amino acid substitutions). Preferably, the variant Fc region has at least one amino acid substitution in the native sequence Fc region or in the Fc region of the parent polypeptide, e.g., from about one to about ten amino acid substitutions, and preferably from about one to about five amino acid substitutions, compared to the native sequence Fc region or the Fc region of the parent polypeptide. The variant Fc region herein preferably has at least about 80% homology to the native sequence Fc region and / or to the Fc region of the parent polypeptide, most preferably at least about 90% homology thereto, more preferably at least about 95% homology thereto.

[0107] "Fc receptor" or "FcR" describes a receptor that is bound to the Fc region of an antibody. Preferred FcRs are native sequence human FcRs. In addition, preferred FcRs are FcRs that bind to IgG antibodies (gamma receptors), and include receptors of FcγRI, FcγRII, and FcγRIII subclasses, including allelic variants and alternative splicing forms of these receptors, FcγRII receptors include FcγRIIA ("activating receptors") and FcγRIIB ("inhibitory receptors"), which have similar amino acid sequences, the main difference of which is their cytoplasmic domains. Activating receptor FcγRIIA contains an immunoreceptor tyrosine-based activation motif ("ITAM") in its cytoplasmic domain. Inhibitory receptor FcγRIIB contains an immunoreceptor tyrosine-based inhibition motif ("ITIM") in its cytoplasmic domain. Other FcRs (including those identified in the future) are also encompassed within the term "FcR" herein. FcR can also increase the serum half-life of antibodies.

[0108] As used herein, "percent (%) amino acid sequence identity" and "homology" with respect to peptide, polypeptide or antibody sequences refers to the percentage of amino acid residues in a candidate sequence that are identical with the amino acid residues in a particular peptide or polypeptide sequence, after aligning the sequences and introducing gaps (if necessary) to achieve the maximum percent sequence identity, and not considering any conservative substitutions as part of the sequence identity. Alignment for determining percent amino acid sequence identity can be achieved in various ways that are within the skill in the art, for example, using publicly available computer software such as BLAST, BLAST-2, ALIGN or MEGALIGN. TM (DNASTAR) software. Those skilled in the art can determine appropriate parameters for measuring alignment, including any algorithms known in the art needed to achieve maximal alignment over the full length of the sequences being compared.

[0109] When used in the context of antibodies that compete for the same epitope (e.g., neutralizing antibodies), the term "competition" refers to competition between antibodies as determined by an assay in which the antibody being tested prevents or inhibits (e.g., reduces) specific binding of a reference molecule (e.g., a ligand or reference antibody) to a common antigen (e.g., MS4A4A or a fragment thereof). Various types of competitive binding assays can be used to determine whether an antibody competes with another antibody, for example: solid phase direct or indirect radioimmunoassay (RIA), solid phase direct or indirect enzyme immunoassay (EIA), sandwich competition assays (see, e.g., Stahli et al., 1983, Methods in Enzymology 9:242-253); solid phase direct biotin-avidin EIA (see, e.g., Kirkland et al., 1986, J. Immunol. 137:3614-3619), solid phase direct label assays, solid phase direct label sandwich assays (see, e.g., Harlow and Lane, 1988, Antibodies, A Laboratory Manual, Cold Spring Harbor Laboratory Press, 1992). Press); solid phase direct labeling RIA using I-125 labeling (see, for example, Morel et al., 1988, Molec. Immunol. 25: 7-15); solid phase direct biotin-avidin EIA (see, for example, Cheung et al., 1990, Virology 176: 546-552); and direct labeling RIA (Moldenhauer et al., 1990, Scand. J. Immunol. 32: 77-82). Typically, this assay involves the use of purified antigens bound to a solid surface or cells carrying any of these unlabeled test antibodies and labeled reference antibodies. Competitive inhibition is measured by measuring the amount of labeling bound to a solid surface or cells in the presence of the test antibody. Typically, the test antibody is present in excess. Antibodies identified by competition assays (competing antibodies) include antibodies that bind to the same epitope as the reference antibody and antibodies that bind to adjacent epitopes that are close enough to the epitope bound to the reference antibody to cause steric hindrance. Additional details on methods for determining competitive binding are provided in the Examples herein. Typically, when a competing antibody is present in excess, it will inhibit (e.g., reduce) specific binding of a reference antibody to a common antigen by at least 20%, 30%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 95%, 97.5%, and / or nearly 100%.

[0110] As used herein, an "interaction" between an MS4A4A polypeptide and a second polypeptide encompasses, but is not limited to, protein-protein interactions, physical interactions, chemical interactions, binding, covalent binding, and ionic binding. As used herein, an antibody "inhibits" an "interaction" between two polypeptides when the antibody disrupts, reduces, or completely eliminates the interaction between the two polypeptides. An antibody to a polypeptide of the present disclosure "inhibits" an "interaction" between two polypeptides when the antibody to the polypeptide binds to one of the two polypeptides. In some embodiments, the interaction can be inhibited by at least about any of 20%, 30%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 95%, 97.5%, and / or nearly 100%.

[0111] The term "epitope" includes any determinant that can be bound by an antibody. An epitope is a region of an antigen that is bound by an antibody targeting the antigen, and when the antigen is a polypeptide, includes specific amino acids that directly contact the antibody. Most commonly, the epitope is located on a polypeptide, but in some cases, it may be located on other types of molecules (such as nucleic acids). An epitope determinant can include chemically active surface groups of a molecule, such as amino acids, sugar side chains, phosphoryl or sulfonyl groups, and can have specific three-dimensional structural characteristics and / or specific charge characteristics. Typically, an antibody specific for a particular target antigen will preferentially identify an epitope on the target antigen in a complex mixture of polypeptides and / or macromolecules.

[0112] An "agonistic" antibody or "activating" antibody is an antibody that induces (eg, increases) one or more activities or functions of the antigen upon binding of the antibody to the antigen.

[0113] An "antagonist" antibody or "blocking" antibody or "inhibitory" antibody is an antibody that, after the antibody binds to the antigen, reduces, inhibits, and / or eliminates (e.g., reduces) the binding of the antigen to one or more ligands, and / or reduces, inhibits, and / or eliminates (e.g., reduces) one or more activities or functions of the antigen after the antibody binds to the antigen. In some embodiments, the antagonist, blocking, or inhibitory antibody substantially or completely inhibits the binding of the antigen to one or more ligands and / or one or more activities or functions of the antigen.

[0114] An "isolated" antibody (such as the isolated anti-MS4A4A antibodies of the present disclosure) is one that has been identified, separated, and / or recovered from a component of the antibody's production environment (e.g., natural or recombinant). Preferably, the isolated antibody is free from all other contaminating components of the antibody's production environment. Contaminating components of the antibody's production environment, such as those produced from recombinant transfected cells, are substances that typically interfere with the research, diagnostic, or therapeutic use of the antibody and may include enzymes, hormones, and other proteinaceous or non-proteinaceous solutes. In preferred embodiments, the antibody will be purified to: (1) greater than 95%, in some embodiments, greater than 99%, by weight of the antibody as determined by, for example, the Lowry method; (2) to a degree sufficient to obtain at least 15 N-terminal residues or internal amino acid sequence as determined by use of a spinning cup sequenator, or (3) to homogeneity as determined by SDS-PAGE under non-reducing or reducing conditions using Coomassie blue or, preferably, silver stain. Isolated antibodies include antibodies in situ within recombinant T cells because at least one component of the antibody's natural environment will be absent. Ordinarily, however, isolated polypeptide or antibody will be prepared by at least one purification step.

[0115] An "isolated" nucleic acid molecule encoding an antibody (such as the anti-MS4A4A antibodies of the present disclosure) is one that has been identified and separated from at least one contaminating nucleic acid molecule, typically associated with the environment in which the isolated nucleic acid molecule is produced. Preferably, the isolated nucleic acid is free of all components associated with the environment in which it is produced. Isolated nucleic acid molecules encoding the polypeptides and antibodies herein are in a form that is different from the form or context in which the nucleic acid molecule is found in nature. Thus, isolated nucleic acid molecules are distinct from nucleic acids encoding the polypeptides and antibodies herein that are naturally present in cells.

[0116] As used herein, the term "vector" is intended to refer to a nucleic acid molecule capable of transporting another nucleic acid that is connected. One type of vector is a "plasmid," which refers to a circular double-stranded DNA to which additional DNA segments can be connected. Another type of vector is a phage vector. Another type of vector is a viral vector, in which additional DNA segments can be connected to a viral genome. Some vectors can replicate autonomously (e.g., bacterial vectors and additional mammalian vectors with bacterial replication origins) in the host cells they introduce. Other vectors (e.g., non-additional mammalian vectors) can be integrated into the genome of the host cell when introduced into the host cell, thereby replicating together with the host genome. In addition, some vectors can guide the expression of genes operably connected thereto. These vectors are referred to herein as "recombinant expression vectors," or simply as "expression vectors." Typically, expression vectors useful in recombinant DNA technology are typically in the form of plasmids. In this specification sheets, "plasmid" and "vector" can be used interchangeably because plasmids are the most commonly used vector forms.

[0117] "Polynucleotide" or "nucleic acid," as used interchangeably herein, refers to a polymer of nucleotides of any length, and includes DNA and RNA. The nucleotides may be deoxyribonucleotides, ribonucleotides, modified nucleotides or bases, and / or their analogs, or any substrate that can be incorporated into a polymer by DNA or RNA polymerase or by a synthetic reaction.

[0118] "Host cell" includes a single cell or cell culture that can be or has been a recipient of one or more vectors for incorporating a polynucleotide insert. Host cells include the progeny of a single host cell, and the progeny are not necessarily completely identical (in morphology or genomic DNA complement) to the original parent cell due to natural, accidental, or deliberate mutation. Host cells include cells transfected in vivo with one or more polynucleotides of the present disclosure.

[0119] As used herein, "carrier" includes pharmaceutically acceptable carriers, excipients, or stabilizers that are non-toxic to the cells or mammals to which they are exposed at the dosages and concentrations employed.

[0120] As used herein, the term "preventing" includes providing protection against the occurrence or recurrence of a particular disease, disorder, or condition in an individual. An individual may be predisposed to, susceptible to, or at risk for a particular disease, disorder, or condition, but has not yet been diagnosed as having the disease, disorder, or condition.

[0121] As used herein, an individual who is "at risk" of developing a particular disease, disorder, or condition may or may not have detectable symptoms of the disease or disease, and may or may not have displayed detectable symptoms of the disease or disease prior to the treatment methods described herein. As is known in the art, "at risk" means that an individual has one or more risk factors, which are measurable parameters associated with the development of a particular disease, disorder, or condition. An individual who has one or more of these risk factors has a higher probability of developing a particular disease, disorder, or condition than an individual who does not have one or more of these risk factors.

[0122] As used herein, the term "treatment" refers to a clinical intervention designed to alter the natural course of a disease in a treated individual during the course of clinical pathology. Desirable therapeutic effects include a reduction in the rate of progression, an amelioration or alleviation of the pathological state, and a alleviation or improvement in the prognosis of a particular disease, disorder, or condition. For example, a subject is successfully "treated" if one or more symptoms associated with a particular disease, disorder, or condition are alleviated or eliminated.

[0123] An "effective amount" refers to an amount that is effective to achieve the desired therapeutic or preventive outcome at least at the necessary dosage and for the required time period. An effective amount can be provided in one or more administrations. The effective amount herein can vary depending on various factors, such as the individual's disease state, age, sex, and weight, as well as the ability of the treatment to elicit the desired response in the individual. An effective amount is also an amount in which the beneficial effects of the treatment outweigh any toxic or deleterious effects of the treatment. For preventive uses, beneficial or desired outcomes include outcomes such as eliminating or reducing risk, reducing severity, or delaying the onset of a disease, including biochemical, histological, and / or behavioral symptoms of the disease, complications of the disease, and intermediate pathological phenotypes that occur during the development of the disease. For therapeutic uses, beneficial or desired outcomes include clinical outcomes such as reducing one or more symptoms caused by the disease, improving the quality of life of patients with the disease, reducing the dosage of other drugs required to treat the disease, or enhancing the effect of another drug, such as by targeting, delaying the progression of the disease, and / or prolonging survival. An effective amount of a drug, compound, or pharmaceutical composition is an amount sufficient to achieve a preventive or therapeutic treatment, either directly or indirectly. As understood in a clinical context, an effective amount of a drug, compound, or pharmaceutical composition may or may not be achieved in combination with another drug, compound, or pharmaceutical composition. Thus, an "effective amount" may be considered in the context of administering one or more therapeutic agents, and a single agent may be considered to be administered in an effective amount if it can achieve or has achieved the desired result in combination with one or more other agents.

[0124] For treatment, prevention, or risk reduction purposes, a "subject" refers to any animal classified as a mammal, including humans, livestock and farm animals, zoo animals, sports animals, or pets, such as dogs, horses, rabbits, cows, pigs, hamsters, gerbils, mice, mink, rats, cats, etc. In some embodiments, the subject is a human.

[0125] As used herein, administration "in conjunction with" another compound or composition includes simultaneous administration and / or administration at different times. Administration in conjunction also encompasses administration as a co-formulation or as a separate composition, including administration at different dosing frequencies or intervals, and using the same route of administration or different routes of administration. In some embodiments, administration in conjunction is administration as part of the same treatment regimen.

[0126] As used herein, the term "about" refers to the common error range of the corresponding value that is readily known to those skilled in the art. Reference herein to "about" a value or parameter includes (and describes) embodiments involving the value or parameter itself.

[0127] As used herein and in the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. For example, a reference to an "antibody" refers to one or more antibodies, such as molar amounts, and includes equivalents thereof known to those skilled in the art, and so forth.

[0128] It should be understood that aspects and embodiments of the present disclosure described herein include "comprising aspects and embodiments," "consisting of," and "consisting essentially of" aspects and embodiments.

[0129] I. Anti-MS4A4A Antibodies

[0130] Anti-MS4A4A antibodies are provided herein. The provided antibodies can be used, for example, for the diagnosis or treatment of MS4A4A-related diseases.

[0131] In one aspect, the present disclosure provides isolated (e.g., monoclonal) antibodies that bind to an epitope within the MS4A4A proteins of the present disclosure. The MS4A4A proteins of the present disclosure include, but are not limited to, mammalian MS4A4A proteins, human MS4A4A proteins, mouse MS4A4A proteins, and cynomolgus monkey MS4A4A proteins. The MS4A4A proteins of the present disclosure include naturally occurring variants of MS4A4A.

[0132] Human MS4A4A is a 239-amino acid protein that encodes a membrane glycoprotein. The amino acid sequence of human MS4A4A is shown in SEQ ID NO: 1:

[0133]

[0134] In addition, the amino acid sequence of mouse MS4A4A is shown in SEQ ID NO: 2:

[0135]

[0136] In addition, the amino acid sequence of cynomolgus monkey (cyno) MS4A4A is shown in SEQ ID NO: 3:

[0137]

[0138] In addition, the amino acid sequence of human MS4A6A is shown in SEQ ID NO: 291:

[0139]

[0140] In some embodiments, MS4A4A is expressed in cells. In some embodiments, MS4A4A is expressed in bone marrow cells. In some embodiments, MS4A4A is expressed in brain cells. In some embodiments, MS4A4A is expressed in astrocytes, including but not limited to mature astrocytes. In some embodiments, MS4A4A is expressed in oligodendrocytes. In some embodiments, MS4A4A is expressed in microglia. In some embodiments, MS4A4A is expressed in immune cells, including but not limited to macrophages, eosinophils, mast cells, dendritic cells, natural killer cells, neutrophils, and T cells. In some embodiments, MS4A4A is expressed in olfactory cells. In some embodiments, MS4A4A is expressed on the surface of cells.

[0141] The MS4A4A protein of the present disclosure comprises several domains, including but not limited to the cytoplasmic domain (amino acid residues 1-64 of human MS4A4A; see SEQ ID NO: 1); transmembrane domain (amino acid residues 65-85 of human MS4A4A); extracellular domain (extracellular domain 1; ECL1), corresponding to amino acid residues 86-98 of human MS4A4A; transmembrane domain (amino acid residues 99-119 of human MS4A4A); cytoplasmic domain (amino acid residues 120-137 of human MS4A4A); transmembrane domain (amino acid residues 138-158 of human MS4A4A); extracellular domain (extracellular domain 2; ECL2), corresponding to amino acid residues 159-179; transmembrane domain (amino acid residues 180-200 of human MS4A4A); and cytoplasmic domain (amino acid residues 201-239 of human MS4A4A). In addition, the MS4A4A protein of the present disclosure is expressed in many tissues and cells, including but not limited to the brain, neurons, glial cells, endothelial cells, pericytes, pericytes, etc.

[0142] MS4A4A binding partners and adaptor molecules

[0143] Also provided herein are methods for screening for anti-MS4A4A antibodies that bind to MS4A4A and block the interaction between MS4A4A and one or more MS4A4A ligands or binding partners. In some embodiments, peptide libraries can be synthesized in which the MS4A4A protein is cleaved into contiguous 15-mer and 25-mer peptides separated by one amino acid residue, and the peptide libraries are subsequently spotted onto filters. The ability of the MS4A4A ligand or binding partner to interact with the MS4A4A peptide or multiple peptides in the presence or absence of anti-MS4A4A antibodies can then be tested by SPOT binding assays (e.g., Frank, R and Overwin, H (1996) Methods. Mol. Biol. 66, 149-169; Reineke, U et al., (2002) J. Immunol. Methods 267, 13-26; and Andersen, OS et al., (2010) J. BIOLOGICAL CHEMISTRY 285, 12210-12222). In some embodiments, the cellulose support can be prepared as an N-modified cellulose-aminohydroxypropyl ether membrane, and all synthetic rounds begin with spot definition by 9-fluorenylmethoxycarbonylalanine-pentafluorophenyl ester (generating an alanine linker between the peptide and the membrane). For example, the automated linear synthesis performed by the stepwise addition of different amino acids is protected by 9-fluorenyl-methoxycarbonyl at the N-terminus, and the growing peptide chain is subjected to appropriate side chain protection. In some embodiments, the pattern of deprotection, activation and coupling is continued until a 16-mer peptide is produced, thereby producing a uniformly distributed array of covalently anchored peptides and cellulose supports at the C-terminus, and the N-terminus is a free end (Scharn, D et al., (2000) J. Comb. Chem. 2, 361-369). In some embodiments, the removal of side protecting groups can be carried out in two steps. First, the membrane can be treated with 90% trifluoroacetic acid (dissolved in dichloromethane, containing 3% triisobutylsilane and 2% H2O); secondly, the membrane is treated with, for example, 60% trifluoroacetic acid (dissolved in dichloromethane, containing 3% triisobutylsilane and 2% H2O). In order to remove trifluoroacetate, the membrane can be washed several times with H2O, ethanol, Tris-buffered saline and ethanol, and then the membrane is dried. Finally, membranes were blocked in blocking buffer swelled in Tris-buffered saline (pH 8.0) and supplemented with 5% sucrose for 2 h before preparation of the predetermined peptide library for ligand binding analysis.In some embodiments, for binding studies of cellulose-binding peptides, the membrane-bound library can be incubated with a combination of S peptides and polyhistidine-tagged ligands in the presence or absence of anti-MS4A4A antibodies, for example, overnight in blocking buffer at 4°C, followed by a second incubation with 1 mg / ml HRP-conjugated S protein in blocking buffer, but for 3 hours at room temperature. Subsequently, the membrane can be washed with Tris-buffered saline (e.g., three times for 10 minutes), and quantitative characterization of bound ligand can be performed using an UptiLight chemiluminescent substrate and a Lumilmager instrument to provide spot signal intensity (Boehringer light units). Alternatively, detection of bound ligand can be performed by immunochemical assay using an antibody against the histidine tag and an HRP-conjugated anti-mouse secondary antibody. Incubation can be performed using standard Western blotting procedures and spot detection.

[0144] Also provided herein are methods of screening for anti-MS4A4A antibodies that block the interaction (eg, binding) of MS4A4A with one or more MS4A4A ligands or binding partners.

[0145] In some embodiments, the interaction between MS4A4A and an MS4A4A ligand or binding partner can be characterized using surface plasmon resonance analysis (e.g., Skeldal et al., 2012 J Biol Chem., 287:43798; and Andersen et al., 201, J Biol Chem, 285, 12210-12222). Direct binding of an MS4A4A ligand or binding partner to immobilized MS4A4A can be determined, for example, on a Biacore 2000 instrument (Biacore, Sweden) using CaHBS as a standard running buffer (10 mM HEPES pH 7.4, 140 mM NaCl, 2 mM CaCl2, 1 mM EGTA, and 0.005% Tween 20) in the presence or absence of a blocking anti-MS4A4A antibody. In some embodiments, a biosensor chip from Biacore (CM5) can be activated using the NHS / EDC method and then coated with MS4A4A to 79 fmol / mm 2of protein density and used for affinity measurement of binding partners. The preparation of biosensor surfaces using pro-MS4A4A will follow the same procedure. After each cycle of the ligand binding experiment, the regeneration of the flow cell can be performed by two 10 μl buffer (10mM glycine-HCl pH4.0, 500mM NaCl, 20mM EDTA and 0.005% Tween20) pulses and a single injection of 0.001% SDS. Fitting of sensorgrams for affinity estimation can be performed, for example, by using BIAevaluation version 3.1. Following a similar protocol, the immobilization of HisS-NGFpro or HisS-BDNFpro on a CM5 biosensor chip can also be performed using an NHS / EDC coupling kit to obtain a similar surface density of immobilized protein (~300fmol / mm 2 Biosensor chips immobilized with MS4A4A ligands or binding partners can also be used to examine the binding of MS4A4A in the absence or presence of competing MS4A4A antibodies.

[0146] In some embodiments, the interaction between MS4A4A and MS4A4A ligands and binding partners can be characterized using sedimentation assays (e.g., Andersen et al., 2010, J Biol Chem, 285, 12210-12222). For example, the expressed intracellular or extracellular domain of MS4A4A can be incubated with labeled MS4A4A ligands or binding partners in the absence or presence of MS4A4A blocking antibodies and precipitated using 100 μl of glutathione (GSH)-agarose beads (Amersham Biosciences, catalog number 17-0756-01). The amount of applied receptor domain can be determined by precipitation using Talon beads as a control. Bound proteins can be separated by SDS-PAGE analysis and visualized by standard Western blot analysis using anti-histidine antibodies.

[0147] In some embodiments, the interaction between MS4A4A and MS4A4A ligands and binding partners can be characterized using cellulose binding proteins (e.g., Andersen et al., 2010, J Biol Chem, 285, 12210-12222). For example, the membrane-bound protein can be incubated with another MS4A4A ligand or binding partner; incubation in blocking buffer at 4°C overnight, followed by a second incubation with 1 μg / ml HRP-conjugated S protein in blocking buffer, but at room temperature for 3 hours. Subsequently, the membrane can be washed three times with Tris-buffered saline for 10 minutes, and then quantitative characterization of bound ligand can be performed using UptiLight chemiluminescent substrate and a Lumilmager instrument to provide spot signal intensity (Boehringer light units). Alternatively, detection of bound ligand can be performed by immunochemical assay using an antibody against the histidine tag and an HRP-conjugated anti-mouse secondary antibody. Incubation can be followed by standard Western blot analysis and spot detection.

[0148] In some embodiments, the interaction between MS4A4A and its ligands and binding partners can be characterized using proximity ligation assays (e.g., Gustafsen et al., 2013 The Journal of Neuroscience, 33: 64-71). For example, cells expressing or exposed to MS4A4A and its ligands or binding partners can be subjected to proximity ligation assays (PLA) (Duolink 11) using a primary antibody against MS4A4A and an antibody against the binding partner, followed by incubation with a secondary antibody conjugated to an oligonucleotide that hybridizes to the subsequently added ring-forming oligonucleotide and initiates rolling circle amplification when the antigen is localized within 40 nM. Amplified DNA can be visualized by adding a complementary fluorescently labeled oligonucleotide.

[0149] In some embodiments, alkaline phosphatase-labeled ligands can be used in cell binding assays to characterize the interaction between MS4A4A and MS4A4A ligands and binding partners (e.g., Hu et al., 2005, J. Neurosci. 25, 5298-5304; Fournier et al., 2001, Nature 409, 341-346; Lauren et al., 2009, Nature 457, 1128-1132; and Hu et al., 2010, Neuron 68, 654-667). For example, alkaline phosphatase (AP)-labeled ligands can be prepared to assess binding to MS4A4A on transfected cells or primary neurons. To detect the binding of AP-labeled ligand to cells expressing MS4A4A, the culture can be washed with, for example, Hanks' balanced salt solution (HBH) containing 20 mM sodium HEPES, pH 7.05, and 1 mg / ml bovine serum albumin (BSA). The plate can then be incubated with AP-labeled ligand in the presence or absence of MS4A4A blocking antibodies, for example, in HBH at 23°C for 2 hours. AP-bound ligand can be detected and quantified according to methods well known in the art.

[0150] In certain embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibody further inhibits the interaction between MS4A4A and its ligands, signaling proteins, or binding proteins by: a) reducing the effective level of MS4A4A available for interaction with one or more ligands or binding proteins; b) blocking one or more sites on MS4A4A required for interaction with one or more ligands or binding proteins; c) preventing one or more post-translational events on MS4A4A required for interaction with one or more ligands or binding proteins and / or correcting the processing and / or subcellular localization of MS4A4A; d) inducing degradation of MS4A4A; e) altering the conformation of MS4A4A, or any combination thereof. In certain embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibody specifically binds to human MS4A4A, mouse MS4A4A, cynomolgus monkey MS4A4A, or a combination thereof. In certain embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibody is a human antibody, a humanized antibody, a bispecific antibody, a monoclonal antibody, a multivalent antibody, a conjugated antibody, or a chimeric antibody. In certain embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibody is a bispecific antibody that recognizes a first antigen and a second antigen. In certain embodiments that may be combined with any of the preceding embodiments, the first antigen is MS4A4A and the second antigen is an antigen that facilitates transport across the blood-brain barrier. In certain embodiments that may be combined with any of the preceding embodiments, the second antigen is selected from the group consisting of: MS4A4A, transferrin receptor (TR), insulin receptor (HIR), insulin-like growth factor receptor (IGFR), low-density lipoprotein receptor-related proteins 1 and 2 (LPR-1 and 2), diphtheria toxin receptor, CRM197, llama single domain antibody, TMEM 30(A), protein transduction domain, TAT, Syn-B, penetratin, polyarginine peptide, angiopep, basic immunoglobulin (basigin), Glut1 and CD98hc, and ANG1005.

[0151] Signal transduction adaptor proteins are, but are not limited to, auxiliary proteins for other proteins associated with signal transduction pathways. Adaptor proteins contain various protein binding modules or motifs that connect protein binding partners together, helping to generate larger signal transduction complexes. Adaptor proteins typically contain several domains within their structure that allow for specific interactions with one or more other specific proteins. For example, the Src homology 2 (SH2) domain recognizes a specific amino acid residue sequence within a protein containing a phosphotyrosine residue. The interaction between adaptor proteins and other signal transduction proteins allows for a variety of specific and coordinated protein-protein interactions to occur within the cell during and in relation to signal transduction.

[0152] An immunoreceptor tyrosine-based activation motif (ITAM) is a conserved sequence of four amino acid residues that is repeated twice in the cytoplasmic tail of certain cell surface proteins of the immune system. In the C-terminus of certain proteins, the consensus motif is YxxI / Lx (6-12) YxxI / L (SEQ ID NO: 355). The two repeat sequences are usually separated by between 6 and 12 amino acid residues (YxxL / Ix (6-12) YxxL / I (SEQ ID NO: 355). ITAMs are important for signal transduction; tyrosine residues within these motifs are phosphorylated upon interaction between receptor molecules and their ligands and form docking sites for other proteins involved in signal transduction (Barrow and Trowsdale, 2006, Eur J Immunol, 36: 1646-1653).

[0153] The immunoreceptor tyrosine-based inhibitory motif (ITIM) is a conserved sequence of amino acids (S / I / V / LxYxxI / V / L (SEQ ID NO: 352)) found in the cytoplasmic tails of certain inhibitory receptors of the immune system. Upon interaction between ITIM-containing inhibitory receptors and their ligands, their ITIM motifs are phosphorylated by enzymes of the Src kinase family, enabling them to recruit other enzymes, such as various phosphotyrosine phosphatases, thereby reducing the activation of molecules involved in signal transduction (Barrow and Trowsdale, 2006, Eur J Immunol, 36: 1646-1653).

[0154] In some embodiments, the ITAM motifs of some receptors are positively charged, such as lysine or arginine. These positively charged amino acid residues mediate the association of the negatively charged transmembrane amino acid residues of the adapter protein of the corresponding coding ITAM. After ligand recognition and receptor clustering, the tyrosine amino acid residue is phosphorylated by Src family protein tyrosine kinase (PTK). The ITAM motifs of dual phosphorylation serve as the docking site of the series connection SH2 domains of Syk family PTK (such as ZAP-70 or Syk). Syk family PTK makes a series of intracellular substrate phosphorylation, causes the formation of near membrane end support, thus causes the raising of important effector molecules (such as phospholipase Cc (PLCc)), thus causes calcium signal conduction and Ras activation, and described Ras activation stimulates ERK pathway and cell activation.

[0155] Studies support the importance of MS4A proteins in forming signaling complexes with other cell surface membrane proteins that regulate or replicate downstream biochemical signals. Although in most cases, binding partners have not been clearly identified for MS4A proteins other than MS4A1 and MS4A2, predictive protein analysis has shown that Src homology 2 (SH2) and SH3 domain binding sites are commonly found in the N-terminal and C-terminal regions of MS4A proteins, which can serve as docking platforms for other signaling molecules (Dinkel et al., 2012, Nucl Acids Res, 40: DD242-D251). The SH domain preferentially binds to proline-rich sequences, which are also commonly found in the cytoplasmic tail of MS4A proteins (Liang and Tedder, 2001, Genomics, 72: 119-127; Kay et al., 2000, FASEB J, 14: 231-241). This feature of MS4A proteins provides a solid basis for their close involvement in protein-protein interactions and for their association with each other to form signaling complexes.

[0156] Thus, in some embodiments, the anti-MS4A4A antibodies of the present disclosure enhance or increase the formation of signaling complexes. In some embodiments, the anti-MS4A4A antibodies of the present disclosure enhance or increase the formation of signaling complexes associated with adaptor proteins encoding ITAMs. In some embodiments, the anti-MS4A4A antibodies of the present disclosure enhance or increase the formation of inhibitory signaling complexes. In some embodiments, the anti-MS4A4A antibodies of the present disclosure enhance or increase the formation of inhibitory signaling complexes associated with adaptor proteins encoding ITIMs.

[0157] In some embodiments, the anti-MS4A4A antibodies of the present disclosure inhibit (e.g., block) or reduce the formation of signaling complexes. In some embodiments, the anti-MS4A4A antibodies of the present disclosure inhibit (e.g., block) or reduce the formation of signaling complexes associated with adaptor proteins encoding ITAMs. In some embodiments, the anti-MS4A4A antibodies of the present disclosure inhibit (e.g., block) or reduce the formation of inhibitory signaling complexes. In some embodiments, the anti-MS4A4A antibodies of the present disclosure inhibit (e.g., block) or reduce the formation of inhibitory signaling complexes associated with adaptor proteins encoding ITIMs.

[0158] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure inhibit (e.g., block) or reduce the formation of MS4A4A homo-oligomeric cell surface protein complexes by: a) reducing the effective level of MS4A4A available for MS4A4A homo-oligomeric complex formation; b) blocking one or more sites on MS4A4A required for MS4A4A homo-oligomeric complex formation; c) preventing one or more post-translational events on MS4A4A required for MS4A4A homo-oligomeric complex formation and / or correct processing and / or cellular localization of MS4A4A; d) inducing degradation of MS4A4A; e) altering the conformation of MS4A4A, or any combination thereof.

[0159] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure increase or enhance the formation of MS4A4A homo-oligomeric cell surface protein complexes by: a) increasing the effective level of MS4A4A available for MS4A4A homo-oligomeric complex formation; b) stabilizing one or more of the sites on MS4A4A required for MS4A4A homo-oligomeric complex formation; c) maintaining cell surface expression of MS4A4A to allow homo-oligomeric complex formation and / or correct processing and / or maintaining correct cellular localization of MS4A4A; d) reducing degradation of MS4A4A; e) altering the conformation of MS4A4A, or any combination thereof.

[0160] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure inhibit (e.g., block) or reduce the formation of MS4A4A hetero-oligomeric cell surface protein complexes with one or more signaling adaptor proteins by: a) reducing the effective level of MS4A4A available for MS4A4A hetero-oligomeric complex formation; b) blocking one or more sites on MS4A4A required for MS4A4A hetero-oligomeric complex formation; c) preventing one or more post-translational events on MS4A4A required for MS4A4A hetero-oligomeric complex formation and / or correct processing and / or cellular localization of MS4A4A; d) inducing degradation of MS4A4A; e) altering the conformation of MS4A4A, or any combination thereof.

[0161] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure increase or enhance the formation of MS4A4A hetero-oligomeric cell surface protein complexes with one or more signaling adaptor proteins by: a) increasing the effective level of MS4A4A available for MS4A4A hetero-oligomeric complex formation; b) stabilizing one or more sites on MS4A4A required for MS4A4A hetero-oligomeric complex formation; c) maintaining cell surface expression of MS4A4A to allow MS4A4A hetero-oligomeric complex formation and / or correct processing and / or maintaining correct cellular localization of MS4A4A; d) reducing degradation of MS4A4A; e) altering the conformation of MS4A4A, or any combination thereof.

[0162] In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure increase or enhance MS4A4A receptor clustering. In some embodiments that may be combined with any of the preceding embodiments, the anti-MS4A4A antibodies of the present disclosure inhibit or reduce MS4A4A receptor clustering. In some embodiments, the increase or enhancement or inhibition or reduction of MS4A4A receptor clustering is in myeloid cells.

[0163] Function of MS4A4A in macrophages and microglia

[0164] Macrophages and myeloid cells of the central nervous system (CNS) have inherent plasticity in phenotype and function. In vitro macrophages can be divided into M1 macrophages and M2 macrophages, which have different phagocytic and inflammatory potentials, phenotypes, and activities. For example, in peripheral organs, macrophages with an M1 phenotype are considered to have pro-inflammatory and anti-microbial phenotypes and functions, while macrophages with an M2 phenotype are considered to be in a more homeostatic state with an anti-inflammatory phenotype and functions.

[0165] Microglia associated with healthy homeostatic conditions express more M2 markers (e.g., CD200R, CD163, and CD115) on the cell surface compared to M1 markers (e.g., CD16, MHC class II, CD86) (Ginhoux and Prinz, 2015, Cold Spring Harb Perspect Biol, 7: a020537). However, in both mouse models of Alzheimer's disease and human Alzheimer's disease, disease-associated microglia (DAMs) are in a proinflammatory or activated state. Disease-associated microglia in a proinflammatory or activated state are considered beneficial by playing an active role in reducing the pathological processes associated with Alzheimer's disease and other neurodegenerative diseases.

[0166] In vitro, MS4A4A expression in M2 macrophages is elevated, and it has been shown that MS4A4A is a novel cell surface marker for M2 macrophages. In addition, MS4A4A has also been shown to regulate the cell surface transport of cKit on mast cells, which indicates the role of MS4A4A in regulating mast cell degranulation and survival (Cruse et al., 2015, Molecular Biol Cell, 26: 1711-1727). In summary, these reported findings indicate that targeting MS4A4A can affect the recycling, expression and / or degradation of various macrophage surface receptors associated with M1 and M2 macrophage phenotypes, thereby affecting their function and activity.

[0167] As described herein, the anti-MS4A4A antibodies of the present disclosure affect the expression of M2 macrophage surface markers. Specifically, the anti-MS4A4A antibodies of the present disclosure reduce the cell surface expression of CD200R, Dectin-1, and CD163 in macrophages, suggesting that the anti-MS4A4A antibodies modulate macrophage polarization, function, and / or activity by reducing the expression of M2 macrophage surface receptors. The anti-MS4A4A antibodies of the present disclosure reduce M2 macrophage surface receptors, suggesting that the anti-MS4A4A antibodies of the present disclosure can effectively alter the physiological state of microglia to a more protective phenotype, such as a more proinflammatory or activated state. Therefore, the anti-MS4A4A antibodies of the present disclosure can be used to treat Alzheimer's disease and other neurodegenerative diseases, in part, by altering the phenotype of macrophages and microglia to a proinflammatory and activated state.

[0168] MS4A4A and TREM2 expression

[0169] Neurodegenerative diseases are characterized, in part, by defects in the immune function of the central nervous system (CNS). For example, it is believed that a reduction in the viability and function of the CNS myeloid cell compartment (including but not limited to microglia) contributes to susceptibility to neurodegenerative diseases such as Alzheimer's disease. Pharmacological interventions that enhance the viability and / or function of myeloid cells would provide effective treatments to improve the onset, severity, or progression of such neurodegenerative diseases and conditions.

[0170] Triggering receptor expressed on myeloid cells 2 (TREM2) is an immunoglobulin-like receptor that is primarily expressed on myeloid cells (such as macrophages, dendritic cells, monocytes, Langerhans skin cells, Kupffer cells, osteoclasts, and microglia). During experimental autoimmune encephalomyelitis and Alzheimer's disease, TREM2 is highly expressed on microglia and infiltrating macrophages in the CNS (Piccio et al., 2007, Eur J Immunol, 37: 1290-1301; Wang, 2015, Cell, 160: 1061-1071). The TREM2 pathway is considered a key regulator of CNS myeloid cell viability and function.

[0171] Data from human genetic studies indicate a close genetic linkage between MS4A4A and TREM2 and a predisposition to Alzheimer's disease (Piccio et al., Acta Neuropathol, 131:925-9330). Specifically, the MS4A4A allele, which is protective against Alzheimer's disease, is associated with elevated sTREM2 levels in the cerebrospinal fluid of patients.

[0172] As described herein, the anti-MS4A4A antibodies of the present disclosure increase cellular ATP levels in macrophages, indicating that the anti-MS4A4A antibodies are effective in increasing, maintaining, or enhancing cell (e.g., macrophage, myeloid cell) viability and function. Additionally, compared to the previously reported commercially available anti-MS4A4A antibody 5C12, which reduced sTREM2 levels in cultured human macrophage supernatants, the anti-MS4A4A antibodies of the present disclosure increased sTREM2 and mTREM2 levels in macrophages (Deming et al., 2018, bioRxiv, doi:dx doi org / 10.1101 / 352179). Since increased protective alleles of MS4A4A against Alzheimer's disease are associated with increased sTREM2 levels, the results provided herein suggest that the anti-MS4A4A antibodies of the present disclosure mimic or replicate the protective phenotype in neurodegenerative diseases and disorders, such as Alzheimer's disease, by increasing sTREM2 and mTREM2 levels.

[0173] As described herein, the anti-MS4A4A antibodies of the present disclosure increase sTREM2 levels, increase mTREM2 levels, or increase both sTREM2 and mTREM2 levels in myeloid cells. In some embodiments, the anti-MS4A4A antibodies increase soluble TREM2 levels by at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or at least 100% compared to a control antibody of the same isotype. In some embodiments, the anti-MS4A4A antibody increases plasma membrane or cell surface TREM2 levels by at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 100%, at least 105%, at least 110%, at least 115%, at least 120%, at least 125%, at least 130%, at least 135%, at least 140%, at least 145%, or at least 150% compared to a control antibody of the same isotype. TREM2 levels in bone marrow cells can be measured by any assay known to those of skill in the art. For example, Example 41 of the present disclosure describes an assay for measuring modulation of soluble and plasma membrane or cell surface TREM2 in human primary macrophages when treated with purified anti-MS4A4A antibodies. Exemplary methods for producing purified antibodies that are free of endotoxin or have low levels of endotoxin are described in Example 41 herein. After incubation of macrophages with purified anti-MS4A4A antibodies, supernatants were collected and sTREM2 levels were determined using the Meso Scale Discovery (MSD) assay. Alternatively, macrophages were collected after incubation and mTREM2 levels were determined by flow cytometry using anti-TREM2 antibodies conjugated to allophycocyanin or a similar fluorophore.

[0174] Agonist antibodies

[0175] The anti-MS4A4A antibodies disclosed herein generally bind to one or more MS4A4A proteins expressed in cells. One class of antibodies is agonistic antibodies. For example, MS4A4A receptors may need to cluster on the cell surface to transduce signals. Thus, agonistic antibodies may have unique characteristics to stimulate, for example, MS4A4A receptors. For example, they may have the correct epitope specificity compatible with receptor activation, as well as the ability to induce or maintain receptor clustering on the cell surface.

[0176] In vivo, antibodies can cluster receptors through a variety of potential mechanisms. Some isotypes of human antibodies (such as IgG2) have an intrinsic ability to cluster receptors or maintain receptors in a clustered configuration due to their unique structure, thereby activating receptors (such as MS4A4A) without binding to Fc receptors (e.g., White et al., (2015) Cancer Cell 27, 138-148).

[0177] Other antibodies cluster receptors (e.g., MS4A4A) by binding to Fcg receptors on neighboring cells. The binding of the constant IgG Fc portion of the antibody to the Fcg receptors leads to aggregation of the antibodies, which in turn aggregate the receptors bound to them through their variable regions (Chu et al. (2008) Mol Immunol, 45: 3926-3933; and Wilson et al. (2011) Cancer Cell 19, 101-113). Binding to inhibitory Fcg receptors FcgR (FcgRIIB) that does not induce cytokine secretion, oxidative burst, increased phagocytosis, and antibody-dependent cell-mediated cytotoxicity (ADCC) is generally a preferred way to cluster antibodies in vivo, because binding to FcgRIIB is not associated with adverse immune reactions.

[0178] Other mechanisms can also be used to cluster receptors (e.g., MS4A4A). For example, as described above, cross-linked antibody fragments (e.g., Fab fragments) can be used to cluster receptors (e.g., MS4A4A) in a manner similar to antibodies with Fc regions that bind to Fcg receptors. Without wishing to be bound by theory, it is believed that if cross-linked antibody fragments (e.g., Fab fragments) induce receptor clustering on the cell surface and bind to the appropriate epitope on the target (e.g., MS4A4A), they can act as agonistic antibodies.

[0179] Thus, in some embodiments, antibodies that bind to the MS4A4A protein may include agonist antibodies that bind to MS4A4A due to their epitope specificity and activate one or more MS4A4A activities. Without wishing to be bound by theory, such antibodies may bind to a ligand binding site on a target antigen (e.g., MS4A4A) and mimic the effects of a natural ligand, or stimulate target antigen transduction signaling by binding to one or more domains that are not ligand binding sites. Such antibodies do not interfere with ligand binding and may act additively or synergistically with the natural ligand.

[0180] In some embodiments, the anti-MS4A4A antibodies of the present disclosure are agonist antibodies that induce or increase one or more MS4A4A activities. In some embodiments, the anti-MS4A4A antibodies induce or increase one or more activities of an MS4A4A protein expressed in a cell. In some embodiments, the anti-MS4A4A antibodies of the present disclosure are antagonist antibodies that reduce or inhibit one or more MS4A4A activities. In some embodiments, the anti-MS4A4A antibodies reduce or inhibit one or more activities of an MS4A4A protein expressed in a cell.

[0181] Inert antibodies

[0182] Another class of antibodies disclosed herein includes inert antibodies. As used herein, "inert" antibodies refer to antibodies that specifically bind to their target antigen but do not modulate (e.g., reduce / inhibit or activate / induce) the function of the antigen. For example, with respect to MS4A4A, inert antibodies do not modulate ligand binding and / or MS4A4A activity. Without wishing to be bound by theory, it is believed that antibodies that do not have the ability to cluster MS4A4A on the cell surface may be inert antibodies, even if they have epitope specificity compatible with receptor activation.

[0183] In some embodiments, antibodies that bind to the MS4A4A protein may include antibodies that bind to MS4A4A but do not modulate protein function due to their epitope specificity. Such functionally inert antibodies can be used as cargo for the transport of toxins, as described for the CD33 antibody gemtuzumab zogamicin (trade name Mylotarg), which is conjugated to a cytotoxic agent from the calicheamicin class and used to target and kill acute myeloid leukemia tumors (Naito et al., (2000), Leukemia, 14, 1436-1443; Ricart (2011) Clin Cancer Res 17; 6417-6436; Hamann et al., (2002) Journal: Bioconjugate Chemistry, 13, 47-58; and Beitz et al., (2001) Clin Cancer Res 7; 1490-6.). Thus, in some embodiments, the antibodies of the present disclosure are inert antibodies that bind MS4A4A but are unable to induce one or more MS4A4A activities (eg, an MS4A4A activity described herein).

[0184] Antagonistic antibodies

[0185] The third class of antibodies disclosed herein includes antagonistic antibodies. In some embodiments, antibodies that bind to the MS4A4A protein may include antagonistic antibodies that, in the presence of a ligand, bind to MS4A4A and inhibit one or more MS4A4A activities by preventing interaction between MS4A4A and one or more of its ligands, or by preventing transduction of MS4A4A signals into the cytoplasm. In some embodiments, the antagonistic antibodies disclosed herein may have the epitope specificity of the agonistic antibodies disclosed herein, but have an Fc domain that is incapable of binding to Fcg receptors and, therefore, is incapable of, for example, clustering of MS4A4A receptors.

[0186] In some embodiments, the antibodies of the present disclosure are antagonistic antibodies. In some embodiments, the antagonistic antibodies inhibit one or more MS4A4A activities. In some embodiments, the antagonistic antibodies reduce the activity of one or more MS4A4A-dependent genes. In some embodiments, the antagonistic antibodies inhibit the interaction between MS4A4A and one or more MS4A4A ligands. In some embodiments, the antagonistic antibodies inhibit MS4A4A signaling. In some embodiments, the antagonistic antibodies inhibit the interaction between MS4A4A and one or more MS4A4A ligands and inhibit MS4A4A signaling.

[0187] In some embodiments, antibody cross-linking is necessary for agonist antibody function. Antibody cross-linking can occur by binding to a secondary antibody in vitro or binding to an Fc receptor in vivo. For example, antagonist antibodies can be converted into agonist antibodies via biotin / streptavidin cross-linking or in vitro secondary antibody binding (see, for example, Gravestein et al., 1996, J. Exp. Med. 184: 675-685; Gravestein et al., 1994, International Immunol, 7: 551-557). Agonist antibodies can exert their activity by mimicking the biological activity of receptor ligands or by enhancing receptor aggregation to activate receptor signaling. In some embodiments, the absence of antibody cross-linking is necessary for antagonist activity. Antagonist antibodies can exert their activity by blocking receptor-ligand interactions.

[0188] calcium influx

[0189] Cell lines transfected with MS4A1 showed increased calcium conductance across the plasma membrane, suggesting that MS4A1 acts as an important channel for regulating calcium homeostasis. (Parolini et al., 2012, Int J Biochem Cell Biol, 44: 2095-2105; Li et al., 2003, J Biol Chem, 278: 42427-42434) Therefore, MS4A proteins participate in the control of intracellular free calcium concentration by regulating (e.g., increasing) calcium influx and / or by mobilizing calcium from intracellular reservoirs (Ishibashi et al., 2001, Gene, 264: 87-93). Due to the conservation of protein structure in the MS4A family, other MS4A proteins (e.g., MS4A4A and MS4A6A) may share overlapping calcium regulatory functions (Ma et al., 2015, Mol Neurobiol, 51: 1240-1248). Thus, in some embodiments, the anti-MS4A4A antibodies of the present disclosure modulate calcium levels, calcium influx, and / or calcium mobilization from intracellular stores in a cell. In some embodiments, the anti-MS44A antibodies of the present disclosure increase calcium influx in a cell and / or increase calcium mobilization in a cell from intracellular stores. In some embodiments, the anti-MS4A4A antibodies of the present disclosure decrease calcium influx in a cell and / or decrease calcium mobilization in a cell from intracellular stores.

[0190] In addition, members of the MS4A protein family are chemoreceptors expressed in olfactory sensory neurons. The results support a model in which MS4A receptors bind inhaled odors and induce calcium influx into olfactory sensory neurons, supporting a role for MS4A family members in regulating calcium influx in various cells (Greer et al., 2016, Cell, 165:1734-1748).

[0191] A. Exemplary Antibodies and Certain Other Antibody Embodiments

[0192] In some embodiments, provided herein are anti-MS4A4A antibodies comprising at least one, two, three, four, five, or six HVRs selected from the group consisting of: (a) HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, and 19, or an amino acid having at least about 95% homology to an amino acid selected from the group consisting of SEQ ID NO: 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, and 19; (b) HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 47, 48, 49, 50, 51, 52, 53, 54, and 55, or an amino acid having at least about 95% homology to an amino acid selected from the group consisting of SEQ ID NO: 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, and 37; (c) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, and 55, or an amino acid having at least about 95% homology to an amino acid selected from the group consisting of SEQ ID NO: 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, and 37; NO:38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, and 55; (d) HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO:56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, and 74, or an amino acid having at least about 95% homology to an amino acid selected from the group consisting of SEQ ID NO:56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, and 74; (e) HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: NO: 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, and 85, or an amino acid having at least about 95% homology to an amino acid selected from the group consisting of SEQ ID NO: 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, and 85;and (f) HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, and 100, or an amino acid having at least about 95% homology to an amino acid selected from the group consisting of SEQ ID NOs: 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, and 100.;

[0193] In some embodiments, provided herein are anti-MS4A4A antibodies comprising at least one, two, three, four, five, or six HVRs selected from the group consisting of: (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 4; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 20; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: 38; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO: 56; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO: 75; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO: 86; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 5; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 21; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: 39; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO: NO:57; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:76; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:87; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:6; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:22; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:40; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:58; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:76; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:88; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:7; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:23; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: NO:41; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:59; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:77; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:89; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:8; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:24; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:42; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:60; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:78;and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:90; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:9; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:25; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:43; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:61; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:76; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:91; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:10; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:26; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:44; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:62; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO: NO:79; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:92; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:7; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:27; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:45; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:63; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:77; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:89; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:7; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:28; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:46; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO: NO:64; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:77; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:89; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:11; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:29; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:47; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:65; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:80; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:93;(a) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 7; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 27; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: 45; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO: 63; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO: 77; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO: 89; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 12; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 27; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: 45; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO: 66; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO: 77; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO: NO:89; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:13; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:30; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:48; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:67; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:76; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:94; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:7; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:27; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:45; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:63; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO: NO:77; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:89; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:14; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:31; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:49; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:68; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:81; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:95; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:15;(b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:32; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:50; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:69; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:82; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:96; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:16; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:33; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:51; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:70; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:77; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:97; NO:17; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:34; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:52; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:71; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:83; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:98; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:18; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:35; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:53; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:72; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:77; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO: ID NO:99; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:19; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:36; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:54; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:73; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:84; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:100; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:7; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:37;(c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: 55; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO: 74; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO: 85; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO: 89.

[0194] In some embodiments, an anti-MS4A4A antibody of the present disclosure comprises HVR-H1 comprising the amino acid sequence of SEQ ID NO: 15, HVR-H2 comprising the amino acid sequence of SEQ ID NO: 32, HVR-H3 comprising the amino acid sequence of SEQ ID NO: 50, HVR-L1 comprising the amino acid sequence of SEQ ID NO: 69, HVR-L2 comprising the amino acid sequence of SEQ ID NO: 82, and HVR-L3 comprising the amino acid sequence of SEQ ID NO: 96.

[0195] In some embodiments, an anti-MS4A4A antibody of the present disclosure comprises HVR-H1 comprising the amino acid sequence of SEQ ID NO: 18, HVR-H2 comprising the amino acid sequence of SEQ ID NO: 35, HVR-H3 comprising the amino acid sequence of SEQ ID NO: 53, HVR-L1 comprising the amino acid sequence of SEQ ID NO: 72, HVR-L2 comprising the amino acid sequence of SEQ ID NO: 77, and HVR-L3 comprising the amino acid sequence of SEQ ID NO: 99.

[0196] In some embodiments, provided herein are anti-MS4A4A antibodies comprising at least one, at least two, or all three V selected from the group consisting of: HHVR sequences: (a) HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, and 19, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, and 19; (b) HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, and 37, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: NO: 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, and 37; and (c) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, and 55, or an amino acid sequence at least about 95% homologous to an amino acid sequence selected from the group consisting of SEQ ID NO: 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, and 55.

[0197] In some embodiments, provided herein are anti-MS4A4A antibodies comprising at least one, at least two, or all three V selected from the group consisting of: LHVR sequences: (a) HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, and 74, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, and 74; (e) HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: NO: 75, 76, 77, 78, 79, 80, 81, 82, 83, 84 and 85, or an amino acid sequence at least about 95% homologous to an amino acid sequence selected from the group consisting of SEQ ID NO: 75, 76, 77, 78, 79, 80, 81, 82, 83, 84 and 85; and (f) HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99 and 100, or an amino acid sequence at least about 95% homologous to an amino acid sequence selected from the group consisting of SEQ ID NO: 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99 and 100.

[0198] In some embodiments, provided herein are anti-MS4A4A antibodies comprising (a) V H A domain comprising at least one, at least two or all three V HHVR sequences: (i) HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, and 19, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, and 19, (ii) HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, and 37, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: NO: 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, and 37, and (iii) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, and 55, or an amino acid sequence at least about 95% homologous to an amino acid sequence selected from the group consisting of SEQ ID NO: 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, and 55, and (b) V L A domain comprising at least one, at least two or all three V LHVR sequences: (i) HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, and 74, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, and 74, (ii) HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: NO: 75, 76, 77, 78, 79, 80, 81, 82, 83, 84 and 85, or an amino acid sequence at least about 95% homologous to an amino acid sequence selected from the group consisting of SEQ ID NO: 75, 76, 77, 78, 79, 80, 81, 82, 83, 84 and 85, and (iii) HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99 and 100, or an amino acid sequence at least about 95% homologous to an amino acid sequence selected from the group consisting of SEQ ID NO: 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99 and 100. NO: 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99 and 100. In some embodiments, provided herein are anti-MS4A4A antibodies comprising (a) V H, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, and 37, or an amino acid sequence that is at least about 95% homologous to an amino acid sequence selected from the group consisting of SEQ ID NO: 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, and 19, (ii) HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, and 37, or an amino acid sequence that is at least about 95% homologous to an amino acid sequence selected from the group consisting of SEQ ID NO: 2 NO: 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, and 37, and (iii) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, and 55, or an amino acid sequence at least about 95% homologous to an amino acid sequence selected from the group consisting of SEQ ID NO: 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, and 55, and (b) V L78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, and 94, or an amino acid sequence at least about 95% homologous to an amino acid sequence selected from the group consisting of SEQ ID NOs: 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 90, 91, 92, 93, and 94, (ii) an HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, and 85, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NOs: NO: 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, and 85, or an amino acid sequence at least about 95% homologous to an amino acid sequence selected from the group consisting of SEQ ID NO: 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, and 85, and (iii) an HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, and 100, or an amino acid sequence at least about 95% homologous to an amino acid sequence selected from the group consisting of SEQ ID NO: 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, and 100.

[0199] In some embodiments, provided herein are anti-MS4A4A antibodies comprising at least one, two, three, four, five, or six HVRs selected from the group consisting of: (a) HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, and 153, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NOs: 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, and 153; (b) HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: NO: 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166 and 167, or an amino acid sequence at least about 95% homologous to an amino acid sequence selected from the group consisting of SEQ ID NO: 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166 and 167; (c) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166 and 167; NO: 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181 and 182, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181 and 182; (d) HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181 and 182; NO: 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195 and 196, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195 and 196;(e) HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 81, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, and 209, or an amino acid sequence at least about 95% homologous to an amino acid sequence selected from the group consisting of SEQ ID NO: 81, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, and 209; and (f) HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: NO: 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, and 224, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, and 224.;

[0200] In some embodiments, provided herein are anti-MS4A4A antibodies comprising at least one, two, three, four, five, or six HVRs selected from the group consisting of: (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 142; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 154; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: 168; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO: 183; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO: 197; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO: 210; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 143; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 155; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: NO:169; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:184; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:198; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:211; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:144; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:156; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:170; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:185; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:199; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:212; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:144; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:156 NO:157; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:171; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:186; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:81; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:213; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:145; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:158; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:172; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:186;(e) HVR-L2 comprising the amino acid sequence of SEQ ID NO: 81; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO: 214; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 146; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 159; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: 173; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO: 187; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO: 200; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO: 215; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 147; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 160; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: NO:174; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:188; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:201; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:216; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:148; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:161; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:175; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:189; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:202; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:217; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:149; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: NO:162; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:176; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:190; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:203; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:218; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:150; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:163; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:177; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:191;(e) HVR-L2 comprising the amino acid sequence of SEQ ID NO: 204; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO: 219; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 151; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 164; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: 178; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO: 192; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO: 205; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO: 220; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 147; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 160; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: NO:179; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:193; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:206; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:221; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:152; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:165; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:180; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:194; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:207; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:222; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:153; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: NO:166; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:181; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:195; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:208; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:223; and (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:151; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:167; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:182; and (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:196;(e) HVR-L2 comprising the amino acid sequence of SEQ ID NO: 209; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO: 224.

[0201] In some embodiments, an anti-MS4A4A antibody of the present disclosure comprises HVR-H1 comprising the amino acid sequence of SEQ ID NO: 143, HVR-H2 comprising the amino acid sequence of SEQ ID NO: 155, HVR-H3 comprising the amino acid sequence of SEQ ID NO: 169, HVR-L1 comprising the amino acid sequence of SEQ ID NO: 184, HVR-L2 comprising the amino acid sequence of SEQ ID NO: 198, and HVR-L3 comprising the amino acid sequence of SEQ ID NO: 211.

[0202] In some embodiments, provided herein are anti-MS4A4A antibodies comprising at least one, at least two, or all three V selected from the group consisting of: H HVR sequences: (a) HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, and 153, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, and 153; (b) HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: NO: 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166 and 167, or an amino acid sequence at least about 95% homologous to an amino acid sequence selected from the group consisting of SEQ ID NO: 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166 and 167; and (c) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166 and 167; ID NO: 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181 and 182, or an amino acid sequence at least about 95% homologous to an amino acid sequence selected from the group consisting of SEQ ID NO: 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181 and 182.

[0203] In some embodiments, provided herein are anti-MS4A4A antibodies comprising at least one, at least two, or all three V selected from the group consisting of: L HVR sequences: (a) HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, and 196, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, and 196; (e) HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: NO: 81, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208 and 209, or an amino acid sequence at least about 95% homologous to an amino acid sequence selected from the group consisting of SEQ ID NO: 81, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208 and 209; and (f) HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223 and 224, or an amino acid sequence at least about 95% homologous to an amino acid sequence selected from the group consisting of SEQ ID NO: IDNO: 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223 and 224.

[0204] In some embodiments, provided herein are anti-MS4A4A antibodies comprising (a) V H A domain comprising at least one, at least two or all three V HHVR sequences: (i) HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, and 153, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, and 153, (ii) HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: NO: 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166 and 167, or an amino acid sequence at least about 95% homologous to an amino acid sequence selected from the group consisting of SEQ ID NO: 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166 and 167, and (iii) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166 and 167. NO: 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181 and 182, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181 and 182, and (b) V L A domain comprising at least one, at least two or all three V LHVR sequences: (i) HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, and 196, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, and 196, (ii) HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, and 224, or an amino acid sequence at least about 95% homologous to an amino acid sequence selected from the group consisting of SEQ ID NO: 81, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, and 209, and (iii) an HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, and 224, or an amino acid sequence at least about 95% homologous to an amino acid sequence selected from the group consisting of SEQ ID NO: 81, 197, 198, 199, 200, 201, 202, ID NO:210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223 and 224.

[0205] In some embodiments, provided herein are anti-MS4A4A antibodies comprising (a) V HA domain comprising (i) an HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, and 153, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, and 153, (ii) an HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: NO: 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166 and 167, or an amino acid sequence at least about 95% homologous to an amino acid sequence selected from the group consisting of SEQ ID NO: 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166 and 167, and (iii) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166 and 167. NO: 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181 and 182, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181 and 182, and (b) V LA domain comprising (i) an HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, and 196, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, and 196, (ii) an HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: NO: 81, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208 and 209, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 81, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208 and 209, and (iii) HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 81, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208 and 209, and NO: 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223 and 224, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223 and 224.

[0206] In some embodiments, provided herein are anti-MS4A4A antibodies comprising at least one, two, three, four, five, or six HVRs selected from the group consisting of: (a) HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, and 153, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NOs: 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, and 153; (b) HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: NO: 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166 and 167, or an amino acid sequence at least about 95% homologous to an amino acid sequence selected from the group consisting of SEQ ID NO: 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166 and 167; (c) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166 and 167; NO: 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181 and 182, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181 and 182; (d) HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181 and 182; NO: 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195 and 196, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195 and 196;(e) HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 81, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, and 209, or an amino acid sequence at least about 95% homologous to an amino acid sequence selected from the group consisting of SEQ ID NO: 81, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, and 209; and (f) HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: NO: 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, and 224, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, and 224.;

[0207] In some embodiments, provided herein are anti-MS4A4A antibodies comprising at least one, two, three, four, five, or six HVRs selected from the group consisting of: (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 304; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 310; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: 316; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO: 322; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO: 328; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO: 334; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 305; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 311; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: NO:317; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:323; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:329; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:335; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:306; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:312; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:318; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:324; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:330; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:336; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:307; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: NO:313; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:319; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:325; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO:331; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO:337; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO:308; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO:314; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO:320; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO:326;(e) HVR-L2 comprising the amino acid sequence of SEQ ID NO: 332; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO: 338; (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 309; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 315; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: 321; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO: 327; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO: 333; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO: 339.

[0208] In some embodiments, provided herein are anti-MS4A4A antibodies comprising at least one, at least two, or all three V selected from the group consisting of: H HVR sequences: (a) HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 304, 305, 306, 307, 308, and 309, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NOs: 304, 305, 306, 307, 308, and 309; (b) HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 310, 311, 312, 313, 314, and 315, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NOs: 310, 311, 312, 313, 314, and 315; and (c) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: NO: 316, 317, 318, 319, 320 and 321, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 316, 317, 318, 319, 320 and 321.

[0209] In some embodiments, provided herein are anti-MS4A4A antibodies comprising at least one, at least two, or all three V selected from the group consisting of: LHVR sequences: (a) HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 322, 323, 324, 325, 326, and 327, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NOs: 322, 323, 324, 325, 326, and 327; (e) HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 328, 329, 330, 331, 332, and 333, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NOs: 328, 329, 330, 331, 332, and 333; and (f) HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: NO: 334, 335, 336, 337, 338 and 339, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 334, 335, 336, 337, 338 and 339.

[0210] In some embodiments, provided herein are anti-MS4A4A antibodies comprising (a) V H A domain comprising at least one, at least two or all three V H HVR sequences: (i) HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 304, 305, 306, 307, 308, and 309, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NOs: 304, 305, 306, 307, 308, and 309, (ii) HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 310, 311, 312, 313, 314, and 315, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NOs: 310, 311, 312, 313, 314, and 315, and (iii) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: NO: 316, 317, 318, 319, 320 and 321, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 316, 317, 318, 319, 320 and 321, and (b) V LA domain comprising at least one, at least two or all three V L HVR sequences: (i) HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 322, 323, 324, 325, 326, and 327, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NOs: 322, 323, 324, 325, 326, and 327, (ii) HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 328, 329, 330, 331, 332, and 333, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NOs: 328, 329, 330, 331, 332, and 333, and (iii) HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 328, 329, 330, 331, 332, and 333. NO: 334, 335, 336, 337, 338 and 339, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 334, 335, 336, 337, 338 and 339.

[0211] In some embodiments, provided herein are anti-MS4A4A antibodies comprising (a) V H A domain comprising: (i) HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 304, 305, 306, 307, 308, and 309, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NOs: 304, 305, 306, 307, 308, and 309, (ii) HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 310, 311, 312, 313, 314, and 315, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NOs: 310, 311, 312, 313, 314, and 315, and (iii) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: NO: 316, 317, 318, 319, 320 and 321, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 316, 317, 318, 319, 320 and 321, and (b) V LA domain comprising: (i) an HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 322, 323, 324, 325, 326, and 327, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NOs: 322, 323, 324, 325, 326, and 327, (ii) an HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 328, 329, 330, 331, 332, and 333, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NOs: 328, 329, 330, 331, 332, and 333, and (iii) an HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: NO: 334, 335, 336, 337, 338 and 339, or an amino acid sequence having at least about 95% homology to an amino acid sequence selected from the group consisting of SEQ ID NO: 334, 335, 336, 337, 338 and 339.

[0212] In another aspect, an anti-MS4A4A antibody comprises a heavy chain variable domain (V H ) sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NO: 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119 and 120. In certain embodiments, the V domain sequence has at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NO: 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119 and 120. HSequences containing substitutions (e.g., conservative substitutions), insertions, or deletions relative to a reference sequence include SEQ ID NOs: 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, and 120, but anti-MS4A4A antibodies comprising such sequences retain the ability to bind to MS4A4A. In certain embodiments, a total of 1 to 10 amino acids are substituted, inserted, and / or deleted in SEQ ID NOs: 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, or 120. In certain embodiments, a total of 1 to 5 amino acids are substituted, inserted, and / or deleted in SEQ ID NO: 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, or 120. In certain embodiments, the substitutions, insertions, or deletions occur in regions outside of the HVRs (i.e., in the FRs). Optionally, the anti-MS4A4A antibody comprises the V sequences of SEQ ID NO: 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, or 120. H Sequence, including post-translational modifications of the sequence. In a specific embodiment, V H comprising one, two or three HVRs selected from the group consisting of: (a) HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18 and 19, (b) HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36 and 37, and (c) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54 and 55.

[0213] In another aspect, an anti-MS4A4A antibody is provided, wherein the antibody comprises a light chain variable domain (V L), said light chain variable domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to an amino acid selected from the group consisting of SEQ ID NO: 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138 and 139. In certain embodiments, the V domain has at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity to an amino acid sequence selected from the group consisting of L The sequences contain substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence: SEQ ID NO: 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, and 139, but anti-MS4A4A antibodies comprising the sequences retain the ability to bind to MS4A4A. In some embodiments, a total of 1 to 10 amino acids are substituted, inserted, and / or deleted in SEQ ID NO: 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, or 139. In certain embodiments, a total of 1 to 5 amino acids are substituted, inserted, and / or deleted in SEQ ID NO: 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, or 139. In certain embodiments, the substitutions, insertions, or deletions occur in regions outside of the HVRs (i.e., in the FRs). Optionally, the anti-MS4A4A antibody comprises the V sequences of SEQ ID NO: 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, or 139. L Sequence, including post-translational modifications of the sequence. In a specific embodiment, V Lcomprising one, two or three HVRs selected from the group consisting of: (a) HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73 and 74, (b) HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 75, 76, 77, 78, 79, 80, 81, 82, 83, 84 and 85, and (c) HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99 and 100.

[0214] In some embodiments, an anti-MS4A4A antibody is provided, wherein the antibody comprises V as in any of the embodiments provided above. H and V in any of the embodiments provided above L In some embodiments, provided herein are anti-MS4A4A antibodies, wherein the antibody comprises V as in any of the embodiments provided above. H and V in any of the embodiments provided above L In one embodiment, the antibody comprises V sequences of SEQ ID NOs: 101-120 and 121-139, respectively. H and V L sequences, including post-translational modifications of those sequences.

[0215] In some embodiments, provided herein are anti-MS4A4A antibodies comprising a heavy chain variable domain (V H ) and light chain variable domain (V L ), where V H and V L Selected from the group consisting of: V comprising the amino acid sequence of SEQ ID NO: 101 H and V comprising the amino acid sequence of SEQ ID NO: 121 L ; V comprising the amino acid sequence of SEQ ID NO: 102 H and V comprising the amino acid sequence of SEQ ID NO: 122 L ; V comprising the amino acid sequence of SEQ ID NO: 103 H and V comprising the amino acid sequence of SEQ ID NO: 123 L ; V comprising the amino acid sequence of SEQ ID NO: 104 Hand V comprising the amino acid sequence of SEQ ID NO: 124 L ; V comprising the amino acid sequence of SEQ ID NO: 105 H and V comprising the amino acid sequence of SEQ ID NO: 125 L ; V comprising the amino acid sequence of SEQ ID NO: 106 H and V comprising the amino acid sequence of SEQ ID NO: 126 L ; V comprising the amino acid sequence of SEQ ID NO: 107 H and V comprising the amino acid sequence of SEQ ID NO: 127 L ; V comprising the amino acid sequence of SEQ ID NO: 108 H and V comprising the amino acid sequence of SEQ ID NO: 128 L ; V comprising the amino acid sequence of SEQ ID NO: 109 H and V comprising the amino acid sequence of SEQ ID NO: 129 L ; V comprising the amino acid sequence of SEQ ID NO: 110 H and V comprising the amino acid sequence of SEQ ID NO: 130 L ; V comprising the amino acid sequence of SEQ ID NO: 111 H and V comprising the amino acid sequence of SEQ ID NO: 131 L ; V comprising the amino acid sequence of SEQ ID NO: 112 H and V comprising the amino acid sequence of SEQ ID NO: 132 L ; V comprising the amino acid sequence of SEQ ID NO: 113 H and V comprising the amino acid sequence of SEQ ID NO: 128 L ; V comprising the amino acid sequence of SEQ ID NO: 114 H and V comprising the amino acid sequence of SEQ ID NO: 133 L ; V comprising the amino acid sequence of SEQ ID NO: 115 H and V comprising the amino acid sequence of SEQ ID NO: 134 L ; V comprising the amino acid sequence of SEQ ID NO: 116 H and V comprising the amino acid sequence of SEQ ID NO: 135 L ; V comprising the amino acid sequence of SEQ ID NO: 117 H and V comprising the amino acid sequence of SEQ ID NO: 136 L; V comprising the amino acid sequence of SEQ ID NO: 118 H and V comprising the amino acid sequence of SEQ ID NO: 137 L ; V comprising the amino acid sequence of SEQ ID NO: 119 H and V comprising the amino acid sequence of SEQ ID NO: 138 L ; and V comprising the amino acid sequence of SEQ ID NO: 120 H and V comprising the amino acid sequence of SEQ ID NO: 139 L .

[0216] In some embodiments, the present disclosure provides an anti-MS4A4A antibody, wherein the antibody comprises a heavy chain variable domain and a light chain variable domain, wherein the heavy chain variable domain comprises the amino acid sequence of SEQ ID NO: 115 and the light chain variable domain comprises the amino acid sequence of SEQ ID NO: 134.

[0217] In some embodiments, the present disclosure provides an anti-MS4A4A antibody, wherein the antibody comprises a heavy chain variable domain and a light chain variable domain, wherein the heavy chain variable domain comprises the amino acid sequence of SEQ ID NO: 118, and the light chain variable domain comprises the amino acid sequence of SEQ ID NO: 137.

[0218] In another aspect, an anti-MS4A4A antibody comprises a heavy chain variable domain (V H ) sequence, said heavy chain variable domain sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NO: 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238 and 239. In certain embodiments, the V domain sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity to an amino acid sequence selected from the group consisting of SEQ ID NO: 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238 and 239. HThe sequence contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence: SEQ ID NO: 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, and 239, but an anti-MS4A4A antibody comprising the sequence retains the ability to bind to MS4A4A. In certain embodiments, a total of 1 to 10 amino acids are substituted, inserted, and / or deleted in SEQ ID NO: 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, or 239. In certain embodiments, a total of 1 to 10 amino acids are substituted, inserted, and / or deleted in SEQ ID NO: In certain embodiments, the anti-MS4A4A antibody comprises SEQ ID NO: 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238 or 239, wherein a total of 1 to 5 amino acids are substituted, inserted and / or deleted. In certain embodiments, the substitutions, insertions or deletions occur in regions outside the HVRs (i.e., in the FRs). Optionally, the anti-MS4A4A antibody comprises SEQ ID NO: 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238 or 239. H Sequence, including post-translational modifications of the sequence, in a specific embodiment, V H comprising one, two or three HVRs selected from the group consisting of: (a) HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152 and 153, (b) HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166 and 167; and (c) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181 and 182.

[0219] In another aspect, an anti-MS4A4A antibody is provided, wherein the antibody comprises a light chain variable domain (V L), said light chain variable domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to an amino acid selected from the group consisting of SEQ ID NO: 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253 and 254. In certain embodiments, the V domain has at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity to an amino acid sequence selected from the group consisting of L The sequences contain substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence: SEQ ID NO: 240, 241, 242, 243, 244, 245, 246, 247, 232, 248, 249, 250, 251, 252, 253, and 254, but anti-MS4A4A antibodies comprising the sequences retain the ability to bind to MS4A4A. In some embodiments, a total of 1 to 10 amino acids are substituted, inserted, and / or deleted in SEQ ID NO: 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, or 254. In certain embodiments, a total of 1 to 5 amino acids are substituted, inserted, and / or deleted in SEQ ID NO: 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, or 254. In certain embodiments, the substitutions, insertions, or deletions occur in regions outside of the HVRs (i.e., in the FRs). Optionally, the anti-MS4A4A antibody comprises the V sequences of SEQ ID NO: 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, or 254. L Sequence, including post-translational modifications of the sequence. In a specific embodiment, V Lcomprising one, two or three HVRs selected from the group consisting of: (a) HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195 and 196, (b) HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 81, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208 and 209, and (c) HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: NO: 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223 and 224.

[0220] In some embodiments, an anti-MS4A4A antibody is provided, wherein the antibody comprises V as in any of the embodiments provided above. H and V in any of the embodiments provided above L In some embodiments, provided herein are anti-MS4A4A antibodies, wherein the antibody comprises V as in any of the embodiments provided above. H and V in any of the embodiments provided above L In one embodiment, the antibody comprises V sequences of SEQ ID NOs: 225-239 and 240-254, respectively. H and V L sequences, including post-translational modifications of those sequences.

[0221] In some embodiments, provided herein are anti-MS4A4A antibodies comprising a heavy chain variable domain (V H ) and light chain variable domain (V L ), where V H and V L Selected from the group consisting of: V comprising the amino acid sequence of SEQ ID NO: 225 H and V comprising the amino acid sequence of SEQ ID NO: 240 L ; V comprising the amino acid sequence of SEQ ID NO: 226 H and V comprising the amino acid sequence of SEQ ID NO: 241 L ; V comprising the amino acid sequence of SEQ ID NO: 227 H and V comprising the amino acid sequence of SEQ ID NO: 242 L; V comprising the amino acid sequence of SEQ ID NO: 228 H and V comprising the amino acid sequence of SEQ ID NO: 243 L ; V comprising the amino acid sequence of SEQ ID NO: 229 H and V comprising the amino acid sequence of SEQ ID NO: 244 L ; V comprising the amino acid sequence of SEQ ID NO: 230 H and V comprising the amino acid sequence of SEQ ID NO: 245 L ; V comprising the amino acid sequence of SEQ ID NO: 231 H and V comprising the amino acid sequence of SEQ ID NO: 246 L ; V comprising the amino acid sequence of SEQ ID NO: 232 H and V comprising the amino acid sequence of SEQ ID NO: 247 L ; V comprising the amino acid sequence of SEQ ID NO: 233 H and V comprising the amino acid sequence of SEQ ID NO: 248 L ; V comprising the amino acid sequence of SEQ ID NO: 234 H and V comprising the amino acid sequence of SEQ ID NO: 249 L ; V comprising the amino acid sequence of SEQ ID NO: 235 H and V comprising the amino acid sequence of SEQ ID NO: 250 L ; V comprising the amino acid sequence of SEQ ID NO: 236 H and V comprising the amino acid sequence of SEQ ID NO: 251 L ; V comprising the amino acid sequence of SEQ ID NO: 237 H and V comprising the amino acid sequence of SEQ ID NO: 252 L ; V comprising the amino acid sequence of SEQ ID NO: 238 H and V comprising the amino acid sequence of SEQ ID NO: 253 L ; and V comprising the amino acid sequence of SEQ ID NO: 239 H and V comprising the amino acid sequence of SEQ ID NO: 254 L .

[0222] In some embodiments, the present disclosure provides an anti-MS4A4A antibody, wherein the antibody comprises a heavy chain variable domain and a light chain variable domain, wherein the heavy chain variable domain comprises the amino acid sequence of SEQ ID NO: 226, and the light chain variable domain comprises the amino acid sequence of SEQ ID NO: 241.

[0223] In another aspect, an anti-MS4A4A antibody comprises a heavy chain variable domain (V H ) sequence, said heavy chain variable domain sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 340, 341, 342, 343, 344 and 345. In certain embodiments, the V domain sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity to an amino acid sequence selected from the group consisting of H Sequences containing substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence: SEQ ID NOs: 340, 341, 342, 343, 344, and 345, but anti-MS4A4A antibodies comprising such sequences retain the ability to bind to MS4A4A. In certain embodiments, a total of 1 to 10 amino acids are substituted, inserted, and / or deleted in SEQ ID NOs: 340, 341, 342, 343, 344, and 345. In certain embodiments, a total of 1 to 5 amino acids are substituted, inserted, and / or deleted in SEQ ID NOs: 340, 341, 342, 343, 344, and 345. In certain embodiments, a total of 1 to 5 amino acids are substituted, inserted, and / or deleted in SEQ ID NOs: 340, 341, 342, 343, 344, and 345. In certain embodiments, substitutions, insertions, or deletions occur in regions outside of the HVRs (i.e., in the FRs). Optionally, the anti-MS4A4A antibody comprises V sequences of SEQ ID NOs: 340, 341, 342, 343, 344, and 345. H Sequence, including post-translational modifications of the sequence. In a specific embodiment, V Hcomprising one, two or three HVRs selected from the group consisting of: (a) HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 304, 305, 306, 307, 308 and 309, (b) HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 310, 311, 312, 313, 314 and 315, and (c) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 316, 317, 318, 319, 320 and 321.

[0224] In another aspect, an anti-MS4A4A antibody is provided, wherein the antibody comprises a light chain variable domain (V L ), said light chain variable domain having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to an amino acid selected from the group consisting of SEQ ID NOs: 346, 347, 348, 349, 350 and 351. In certain embodiments, the V domain has at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity to an amino acid sequence selected from the group consisting of L The sequences contain substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence: SEQ ID NO: 346, 347, 348, 349, 350, and 351, but the anti-MS4A4A antibodies comprising the sequences retain the ability to bind to MS4A4A. In some embodiments, a total of 1 to 10 amino acids are substituted, inserted, and / or deleted in SEQ ID NO: 346, 347, 348, 349, 350, or 351. In certain embodiments, a total of 1 to 5 amino acids are substituted, inserted, and / or deleted in SEQ ID NO: 346, 347, 348, 349, 350, or 351. In certain embodiments, the substitutions, insertions, or deletions occur in regions outside of the HVRs (i.e., in the FRs). Optionally, the anti-MS4A4A antibodies comprise the V sequences of SEQ ID NO: 346, 347, 348, 349, 350, or 351. L Sequence, including post-translational modifications of the sequence. In a specific embodiment, V Lcomprising one, two or three HVRs selected from the group consisting of: (a) HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 322, 323, 324, 325, 326 and 327, (b) HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 334, 335, 336, 337, 338 and 339, and (c) HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 334, 335, 336, 337, 338 and 339.

[0225] In some embodiments, an anti-MS4A4A antibody is provided, wherein the antibody comprises V as in any of the embodiments provided above. H and V in any of the embodiments provided above L In some embodiments, provided herein are anti-MS4A4A antibodies, wherein the antibody comprises V as in any of the embodiments provided above. H and V in any of the embodiments provided above L In one embodiment, the antibody comprises V sequences of SEQ ID NOs: 340-345 and 346-351, respectively. H and V L sequences, including post-translational modifications of those sequences.

[0226] In some embodiments, provided herein are anti-MS4A4A antibodies comprising a heavy chain variable domain (V H ) and light chain variable domain (V L ), where V H and V L Selected from the group consisting of: V comprising the amino acid sequence of SEQ ID NO: 340 H and V comprising the amino acid sequence of SEQ ID NO: 346 L ; V comprising the amino acid sequence of SEQ ID NO: 341 H and V comprising the amino acid sequence of SEQ ID NO: 347 L ; V comprising the amino acid sequence of SEQ ID NO: 342 H and V comprising the amino acid sequence of SEQ ID NO: 348 L ; V comprising the amino acid sequence of SEQ ID NO: 343 H and V comprising the amino acid sequence of SEQ ID NO: 349 L ; V comprising the amino acid sequence of SEQ ID NO: 344H and V comprising the amino acid sequence of SEQ ID NO: 350 L ; and V comprising the amino acid sequence of SEQ ID NO: 345 H and V comprising the amino acid sequence of SEQ ID NO: 351 L .

[0227] In some embodiments, an anti-MS4A4A antibody of the present disclosure competitively inhibits the binding of at least one reference antibody selected from the group consisting of: 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15, 4A-16, 4A-17, 4A-18, 4A-19, 4A-20, 4A- 21. 4A-23, 4A-24, 4A-201, 4A-202, 4A-203, 4A-204, 4A-205, 4A-206, 4A-207, 4A-208, 4A-20 9. 4A-210, 4A-213, 4A-214, 4A-216, 4A-217, 4A-219, 4A-25, 4A-26, 4A-239, 4A-225 and 4A-220.

[0228] In some embodiments, when an anti-MS4A4A antibody of the present disclosure reduces binding to MS4A4A by one or more of the following: 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15, 4A-16, 4A-17, 4A-18, 4A-19, 4A-20, 4A-21, 4A-22, 4A-23, 4A-24, 4A-25, 4A-26, 4A-27, 4A-28, 4A-29, 4A-30, 4A-31, 4A-32, 4A-33, 4A-34, 4A-35, 4A-36, 4A-37, 4A-38, 4A-39, 4A-40, 4A-41, 4A-42 0, 4A-21, 4A-23, 4A-24, 4A-201, 4A-202, 4A-203, 4A-204, 4A-205, 4A-206, 4A-207, 4A-208, 4A-209, 4A-210, 4A-213, 4A-214, 4A-216, 4A-217, 4A-219, 4A-25, 4A-26, 4A-239, 4A-225 and 4A-220 and any combination thereof The anti-MS4A4A antibody binds to one or more of the group consisting of 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15, 4A-16, 4A-17, 4A-18, 4A-19, 4A-20, 4A-21, 4A-23, 4A-24, 4A-25, 4A-26, 4A-27, 4A-28, 4A-30, 4A-31, 4A-32, 4A-33, 4A-34, 4A-35, 4A-36, 4A-37, 4A-38, 4A-39, 4A-40, 4A-41, 4A-42, 4A-43, 4A-44, 4A-45, 4A-46, 4A-47, 4A-48, 4A-49, 4A-50, 4A-51, 4A-52, 4A-53, 4A-54, 4A-55, 4A-56, 4A-57, 4A-58, 4A-225, 4A-226, 4A-227, 4A-228, 4A-239, 4A-230, 4A-231, 4A-232, 4A-233, 4A-234, 4A-235, 4A-236, 4A-237, 4A-238, 4A-239, 4A-240, 4A-241, 4A-242, 4A-243, 4A-244, 4A-245, 4A-246, 4A-247, 4A-248, 4A-249, 4A-250, 4A-251, 4A-252, 4A-253, 4A-254, 4A-256, 4A-257, 4A-258, 4A-259, 4A-260, 4A-261, 4A-262, 4A-263, 4A-264, 4A-265, 4A-266, 4A-267, 4A-268, 4A-269, 4A-270, 4A-271, 4A-272, 4A-273, 4A-274, 4A-275, 4A-276, 4A-277, 4A-278, 4A-279, 4A-280, 4A-281, 4A-282, 4A-283

[0229] In some embodiments, when an anti-MS4A4A antibody of the present disclosure reduces binding to MS4A4A by one or more of the following: 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15, 4A-16, 4A-17, 4A-18, 4A-19, 4A-20, 4A-21, 4A-22, 4A-23, 4A-24, 4A-25, 4A-26, 4A-27, 4A-28, 4A-30, 4A-31, 4A-32, 4A-33, 4A-34, 4A-35, 4A-36, 4A-37, 4A-38, 4A-39, 4A-40, 4A-41, 4A-42, 4A-43, 4A-44, 4A-45, 4A-46, 4A-47, 4A-48, 4A-49, 4A-50, 4A-51, 4A-52, 4A-53, 4A-54, 4A-55 3, 4A-24, 4A-201, 4A-202, 4A-203, 4A-204, 4A-205, 4A-206, 4A-207, 4A-208, 4A-209, 4A-210, 4A-213, 4A-214, 4A-216, 4A-217, 4A-219, 4A-25, 4A-26, 4A-239, 4A-225, and 4A-220, and any combination thereof, that binds to MS4A4A by at least 50%, at least 55%, or both. , at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95% or 100%, when the anti-MS4A4A antibody is combined with one or more of the group consisting of 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15, 4A-16, 4A-17, 4A-18, 4A-19, 4A-20, 4A-21, 4A-22, 4A-23, 4A-24, 4A-25, 4A-26, 4A-27, 4A-28, 4A-30, 4A-31, 4A-32, 4A-33, 4A-34, 4A-35, 4A-36, 4A-37, 4A-38, 4A-39, 4A-40, 4A-41, 4A-42, 4A-43, 4A-44, 4A-45, 4A-46, 4A-47, 4A-48, 4A-49, 4A-50, 4A-51, 4A-52, 4A-53, 4A-54, 4A-55, 4A-56, 4A-57, 4A-58, 4A-24, 4A-201, 4A-202, 4A-203, 4A-204, 4A-205, 4A-206, 4A-207, 4A-208, 4A-209, 4A-210, 4A-213, 4A-214, 4A-216, 4A-217, 4A-219, 4A-25, 4A-26, 4A-239, 4A-225, and 4A-220, and any combination thereof, compete for binding to MS4A4A. In some embodiments, the anti-MS4A4A antibodies of the present disclosure reduce one or more of the following: 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15, 4A-16, 4A-17, 4A-18, 4A-19, 4A-20, 4A-21, 4A-22, 4A-23, 4A-24, 4A-25, 4A-26, 4A-27, 4A-28, 4A-29, 4A-30, 4A-31, 4A-32, 4A-33, 4A-34, 4A-35, 4A-36, 4A-37, 4A-38, 4A-39, 4A-40, 4A-41, 4A-42, 4A-43, 4A-44, 4A-45, 4A-46, 4A-47, 4A-48, 4A-49, 4A-50, 4A-51, 4A-52, 4A-53, 4A-54, 4A-55, 4A-56, 4A-57, 4A-58, -21, 4A-23, 4A-24, 4A-201, 4A-202, 4A-203, 4A-204, 4A-205, 4A-206, 4A-207, 4A-20 8. 4A-209, 4A-210, 4A-213, 4A-214, 4A-216, 4A-217, 4A-219, 4A-25, 4A-26, 4A-239,Binding of 4A-225 and 4A-220 and any combination thereof to MS4A4A by 100% means that the anti-MS4A4A antibody substantially completely blocks one or more of the antibodies selected from 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15, 4A-16, 4A-17, 4A-18, 4A-19, 4A-20, 4A-21, 4A-22, 4A-23, 4A-24, 4A-25, 4A-26, 4A-27, 4A-28, 4A-29, 4A-30, 4A-31, 4A-32, 4A-33, 4A-34, 4A-35 1, 4A-23, 4A-24, 4A-201, 4A-202, 4A-203, 4A-204, 4A-205, 4A-206, 4A-207, 4A-208, 4A-209, 4A-210, 4A-213, 4A-214, 4A-216, 4A-217, 4A-219, 4A-25, 4A-26, 4A-239, 4A-225, and 4A-220, and any combination thereof, to MS4A4A. In some embodiments, an anti-MS4A4A antibody and one or more of the group consisting of 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15, 4A-16, 4A-17, 4A-18, 4A-19, 4A-20, 4A-21, 4A-23, 4A-24, 4A-25, 4A-26, 4A-27, 4A-28, 4A-30, 4A-31, 4A-32, 4A-33, 4A-34, 4A-35, 4A-36, 4A-37, 4A-38, 4A-39, 4A-40, 4A-41, 4A-42, 4A-43, 4A-44, 4A-45, 4A-46, 4A-47, 4A-48, 4A-49, 4A-50, 4A-51, 4A-52, 4A-53, 4A-54, 4A-55, 4A-56, 4A-57, 4A-58, 4A-59, 4A-60, 4A-61, 4A-62, 4A-63, 4A-64, 4A-65 The antibodies of 4A-202, 4A-203, 4A-204, 4A-205, 4A-206, 4A-207, 4A-208, 4A-209, 4A-210, 4A-213, 4A-214, 4A-216, 4A-217, 4A-219, 4A-25, 4A-26, 4A-239, 4A-225, and 4A-220, and any combination thereof, are present in amounts corresponding to a 10:1 ratio, a 9:1 Ratio, 8:1 Ratio, 7:1 Ratio, 6:1 Ratio, 5:1 Ratio, 4:1 Ratio, 3:1 Ratio, 2:1 Ratio, 1:1 Ratio, 0.75:1 Ratio, 0.5:1 Ratio, 0.25:1 Ratio, 0.1:1 Ratio, 0.075:1 Ratio, 0.050:1 Ratio, 0.025:1 Ratio, 0.01:1 Ratio, 0.0075: Ratio, 0.0050:1 Ratio, 0.0025:1 Ratio, 0.00 1: ratio, 0.00075:1 ratio, 0.00050:1 ratio, 0.00025:1 ratio, 0.0001: ratio, 1:10 ratio, 1:9 ratio, 1:8 ratio, 1:7 ratio, 1:6 ratio, 1:5 ratio, 1:4 ratio, 1:3 ratio, 1:2 ratio, 1:0.75 ratio, 1:0.5 ratio, 1:0.25 ratio, 1:0.1 ratio, 1:0.075 ratio, 1:0.050 ratio,1:0.025 ratio, 1:0.01 ratio, 1:0.0075 ratio, 1:0.0050 ratio, 1:0.0025 ratio, 1:0.001 ratio, 1:0.00075 ratio, 1:0.00050 ratio, 1:0.00025 ratio or 1:0.0001 ratio of an anti-MS4A4A antibody to one or more of 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15 , 4A-16, 4A-17, 4A-18, 4A-19, 4A-20, 4A-21, 4A-23, 4A-24, 4A-201, 4A-202, 4A-203, 4A-204, 4A-205, 4A-206, 4A-207, 4A-208, 4A-209, 4A-210, 4A-213, 4A-214, 4A-216, 4A-217, 4A-219, 4A-25, 4A-26, 4A-239, 4A-225, and 4A-220, and any combination thereof. In some embodiments, the invention relates to one or more of the following: 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15, 4A-16, 4A-17, 4A-18, 4A-19, 4A-20, 4A-21, 4A-23, 4A-24, 4A-201, 4A-202, 4A-203, 4A-2 In some embodiments, the anti-MS4A4A antibody is present in an excess ranging from about 1.5-fold to 100-fold, or greater than 100-fold, compared to the amount of the antibody against 4A-04, 4A-205, 4A-206, 4A-207, 4A-208, 4A-209, 4A-210, 4A-213, 4A-214, 4A-216, 4A-217, 4A-219, 4A-25, 4A-26, 4A-239, 4A-225, and 4A-220, and any combination thereof. In some embodiments, the invention relates to one or more of the following: 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15, 4A-16, 4A-17, 4A-18, 4A-19, 4A-20, 4A-21, 4A- 23. 4A-24, 4A-201, 4A-202, 4A-203, 4A-204, 4A-205, 4A-206, 4A-207, 4A-208, 4 A-209, 4A-210, 4A-213, 4A-214, 4A-216, 4A-217, 4A-219, 4A-25, 4A-26, 4A-239,The anti-MS4A4A antibody is present in about a 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, 10-fold, 15-fold, 20-fold, 25-fold, 30-fold, 35-fold, 40-fold, 45-fold, 50-fold, 55-fold, 60-fold, 65-fold, 70-fold, 75-fold, 80-fold, 85-fold, 90-fold, 95-fold, or 100-fold excess compared to the amount of the reference antibody, 4A-225, and 4A-220, and any combination thereof.

[0230] In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to an epitope of human MS4A4A that is distinct from at least one of the epitopes selected from 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15, 4A-16, 4A-17, 4A-18, 4A-19, 4A-20, 4A-21, 4A-23, , 4A-24, 4A-201, 4A-202, 4A-203, 4A-204, 4A-205, 4A-206, 4A-207, 4A-208, 4A-209, 4A-210, 4A-213, 4A-214, 4A-216, 4A-217, 4A-219, 4A-25, 4A-26, 4A-239, 4A-225, and 4A-220 reference antibodies bind to the same or overlapping epitopes of MS4A4A.

[0231] In some embodiments, an anti-MS4A4A antibody of the present disclosure binds to an epitope of MS4A4A that is substantially the same as an epitope bound by at least one reference antibody selected from the group consisting of: 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15, 4A-16, 4A-17, 4A-18, 4A-19, 4A-20, 4A-21, 4A-22, 4A-23, 4A-24, 4A-25, 4A-26, 4A-27, 4A-28, 4A-29, 4A-30, 4A-31, 4A-32, 4A-33, 4A-34, 4A-35, 4A-36, 4A-37, 4A-38, 4A-39, 4A-40, 4A-41, 4A-42, 4A-43, 4A-44, 4A-45, 4A-46, 4A-47, 4A-48, 4A-49, 4A-50, 4A-51, 4A-52, 4A-53, 4A-54, 4A-55, 4A-56, 4A-57, 4A-58, 4A-59, 4A-60, 4A-61, 4A-62, 4A-63, 4A-64, 4A-65 Detailed exemplary methods for locating epitopes to which antibodies bind are provided in Morris (1996) "Epitope Mapping Protocols", in Methods in Molecular Biology, Vol. 66 (Humana Press, Totowa, NJ).

[0232] In some embodiments, the anti-MS4A4A antibodies of the present disclosure compete for binding to MS4A4A with one or more reference antibodies selected from 4A-1, 4A-2, 4A-3, 4A-4, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-14, 4A-15, 4A-16, 4A-19, 4A-20, 4A-21, and 4A-23, and any combination thereof. In some embodiments, the anti-MS4A4A antibodies of the present disclosure compete for binding to MS4A4A with one or more reference antibodies selected from 4A-5, 4A-12, 4A-13, 4A-17, 4A-18, 4A-22, and 4A-24, and any combination thereof.

[0233] In some embodiments, the anti-MS4A4A antibodies of the present disclosure competitively inhibit binding of at least one reference antibody selected from 4A-1, 4A-2, 4A-3, 4A-4, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-14, 4A-15, 4A-16, 4A-19, 4A-20, 4A-21, and 4A-23, and any combination thereof, to MS4A4A. In some embodiments, the anti-MS4A4A antibodies of the present disclosure competitively inhibit binding of at least one reference antibody selected from 4A-5, 4A-12, 4A-13, 4A-17, 4A-18, 4A-22, and 4A-24, and any combination thereof, to MS4A4A.

[0234] In some embodiments, the anti-MS4A4A antibodies of the present disclosure have an epitope on MS4A4A for binding to MS4A4A that is identical or overlapping with at least one reference antibody selected from 4A-1, 4A-2, 4A-3, 4A-4, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-14, 4A-15, 4A-16, 4A-19, 4A-20, 4A-21, and 4A-23, and any combination thereof. In some embodiments, the anti-MS4A4A antibodies of the present disclosure have an epitope on MS4A4A for binding to MS4A4A that is identical or overlapping with at least one reference antibody selected from 4A-5, 4A-12, 4A-13, 4A-17, 4A-18, 4A-22, and 4A-24, and any combination thereof.

[0235] In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acids within amino acid residues 1-64 of human MS4A4A (SEQ ID NO: 1). In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acids within amino acid residues 65-85 of human MS4A4A (SEQ ID NO: 1). In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acids within amino acid residues 86-98 of human MS4A4A (SEQ ID NO: 1). In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acids within amino acid residues 99-119 of human MS4A4A. In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acids within amino acid residues 120-137 of human MS4A4A. In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acids within amino acid residues 138-158 of human MS4A4A. In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acids within amino acid residues 159-179 of human MS4A4A. In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acids within amino acid residues 180-200 of human MS4A4A. In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acids within amino acid residues 201-239 of human MS4A4A. In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to extracellular domain 1 (ECL1) of MS4A4A. In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acids within the amino acid sequence CMASNTYGSNPIS (SEQ ID NO: 289) of SEQ ID NO: 1. In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to extracellular domain 2 (ECL2) of MS4A4A. In some embodiments, an anti-MS4A4A antibody of the present disclosure binds to one or more amino acids within the amino acid sequence SFHHPYCNYYGNSNNCHGTMS (SEQ ID NO: 290) of SEQ ID NO: 1.

[0236] In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to a region or epitope in human MS4A4A comprising amino acid residues 155-177 of human MS4A4A of SEQ ID NO: 1. In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acids within amino acid residues 155-177 of human MS4A4A of SEQ ID NO: 1. In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to a region or epitope in human MS4A4A comprising the amino acid sequence LAFYSFHHPYCNYYG (SEQ ID NO: 296). In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acid residues within the amino acid sequence LAFYSFHHPYCNYYG (SEQ ID NO: 296). In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to a region or epitope in human MS4A4A comprising the amino acid sequence FYSFHHPYCNYYGNS (SEQ ID NO: 297). In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acid residues within the amino acid sequence FYSFHHPYCNYYGNS (SEQ ID NO: 297). In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to a region or epitope in human MS4A4A comprising the amino acid sequence SFHHPYCNYYGNSNN (SEQ ID NO: 298). In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acid residues within the amino acid sequence SFHHPYCNYYGNSNN (SEQ ID NO: 298). In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to a region or epitope in human MS4A4A comprising the amino acid sequence HHPYCNYYGNSNNCH (SEQ ID NO: 299). In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acid residues within the amino acid sequence HHPYCNYYGNSNNCH (SEQ ID NO: 299). In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to a region or epitope in human MS4A4A comprising the amino acid sequence PYCNYYGNSNNCHGT (SEQ ID NO: 300). In some embodiments, the anti-MS4A4A antibodies of the present disclosure bind to one or more amino acid residues within the amino acid sequence PYCNYYGNSNNCHGT (SEQ ID NO: 300).

[0237] Any suitable competition assay or MS4A4A binding assay known in the art (such as BIAcore analysis, ELISA assay, or flow cytometry) can be used to determine whether an anti-MS4A4A antibody binds to one or more of the antibodies selected from 4A-2, 4A-3, 4A-4, 4A-5, 4A-6, 4A-7, 4A-8, 4A-9, 4A-10, 4A-11, 4A-12, 4A-13, 4A-14, 4A-15, 4A-16, 4A-17, 4A-18, 4A-19, 4A-20, 4A- 4A-21, 4A-23, 4A-24, 4A-201, 4A-202, 4A-203, 4A-204, 4A-205, 4A-206, 4A-207, 4A-208, 4A-209, 4A-210, 4A-213, 4A-214, 4A-216, 4A-217, 4A-219, 4A-25, 4A-26, 4A-239, 4A-225, and 4A-220, and any combination thereof, compete for binding to (or competitively inhibit such binding of) MS4A4A. In one exemplary competition assay, immobilized MS4A4A or cells expressing MS4A4A on their cell surface are incubated in a solution containing a first labeled antibody (e.g., human or non-human primate) that binds to MS4A4A and a second unlabeled antibody whose ability to compete with the first antibody for binding to MS4A4A is being tested. The second antibody can be present in hybridoma supernatant. As a control, immobilized MS4A4A or cells expressing MS4A4A are incubated in a solution containing the first labeled antibody but not the second unlabeled antibody. Following incubation under conditions permissive for binding of the first antibody to MS4A4A, excess unbound antibody is removed, and the amount of label associated with the immobilized MS4A4A or cells expressing MS4A4A is measured. If the amount of label associated with the immobilized MS4A4A or cells expressing MS4A4A is substantially reduced in the test sample relative to the control sample, this indicates that the second antibody is competing with the first antibody for binding to MS4A4A. See, Harlow and Lane (1988) Antibodies: A Laboratory Manual Chapter 14 (Cold Spring Harbor Laboratory, Cold Spring Harbor, NY).

[0238] Also provided herein are anti-MS4A4A antibodies that competitively inhibit binding of and / or compete with binding of an anti-MS4A4A antibody comprising (a) V HA domain comprising (i) HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, and 19, (ii) HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, and 37, and (iii) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, and 55, and (b) V L In some embodiments, the antibody comprises an HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, and 74, (ii) an HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, and 85, and (iii) an HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, and 100. In some embodiments, the antibody comprises V domains selected from SEQ ID NOs: 101-120 and SEQ ID NOs: 121-139, respectively. H and V L sequence.

[0239] Provided herein are anti-MS4A4A antibodies that bind to an epitope of human MS4A4A that is identical to or overlaps with an epitope bound by an anti-MS4A4A antibody comprising (a) V HA domain comprising (i) HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, and 19, (ii) HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, and 37, and (iii) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, and 55, and (b) V L In some embodiments, the antibody comprises an HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, and 74, (ii) an HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, and 85, and (iii) an HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, and 100. In some embodiments, the antibody comprises V domains selected from SEQ ID NOs: 101-120 and SEQ ID NOs: 121-139, respectively. H and V L In some embodiments, the epitope of human MS4A4A is the same as the epitope bound by the anti-MS4A4A antibody.

[0240] Also provided herein are anti-MS4A4A antibodies that competitively inhibit binding of and / or compete with binding of an anti-MS4A4A antibody comprising (a) V HA domain comprising (i) HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, and 153, (ii) HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, and 167, and (iii) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, and 182, and (b) V L A domain comprising (i) an HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, and 196, (ii) an HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 81, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, and 209, and (iii) an HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, and 224. In some embodiments, the antibody comprises V sequences of SEQ ID NOs: 225-239 and SEQ ID NOs: 240-254, respectively. H and V L sequence.

[0241] Provided herein are anti-MS4A4A antibodies that bind to an epitope of human MS4A4A that is identical to or overlaps with an epitope bound by an anti-MS4A4A antibody comprising (a) V HA domain comprising (i) HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, and 153, (ii) HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, and 167, and (iii) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, and 182, and (b) V L A domain comprising (i) an HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, and 196, (ii) an HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 81, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, and 209, and (iii) an HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, and 224. In some embodiments, the antibody comprises V sequences of SEQ ID NOs: 225-239 and SEQ ID NOs: 240-254, respectively. H and V L In some embodiments, the epitope of human MS4A4A is the same as the epitope bound by the anti-MS4A4A antibody.

[0242] Also provided herein are anti-MS4A4A antibodies that competitively inhibit binding of and / or compete with binding of an anti-MS4A4A antibody comprising (a) V HA domain comprising (i) HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 304, 305, 306, 307, 308, and 309, (ii) HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 310, 311, 312, 313, 314, and 315, and (iii) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 316, 317, 318, 319, 320, and 321, and (b) V L A domain comprising (i) HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 322, 323, 324, 325, 326, and 327, (ii) HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 328, 329, 330, 331, 332, and 333, and (iii) HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 334, 335, 336, 337, 338, and 339. In some embodiments, the antibody comprises V sequences of SEQ ID NOs: 340-345 and SEQ ID NOs: 346-351, respectively. H and V L sequence.

[0243] Provided herein are anti-MS4A4A antibodies that bind to an epitope of human MS4A4A that is identical to or overlaps with an epitope bound by an anti-MS4A4A antibody comprising (a) V H A domain comprising (i) HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 304, 305, 306, 307, 308, and 309, (ii) HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 310, 311, 312, 313, 314, and 315, and (iii) HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 316, 317, 318, 319, 320, and 321, and (b) V LA domain comprising (i) HVR-L1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 322, 323, 324, 325, 326, and 327, (ii) HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 328, 329, 330, 331, 332, and 333, and (iii) HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 334, 335, 336, 337, 338, and 339. In some embodiments, the antibody comprises V sequences of SEQ ID NOs: 340-345 and SEQ ID NOs: 346-351, respectively. H and V L In some embodiments, the epitope of human MS4A4A is the same as the epitope bound by the anti-MS4A4A antibody.

[0244] In some embodiments, the anti-MS4A4A antibody according to any of the above embodiments is a monoclonal antibody, including a humanized antibody and / or a human antibody. In some embodiments, the anti-MS4A4A antibody is an antibody fragment, such as an Fv, Fab, Fab', scFv, diabody, or F(ab')2 fragment. In some embodiments, the anti-MS4A4A antibody is a substantially full-length antibody, such as an IgG1 antibody, an IgG2a antibody, or another antibody class or isotype as defined herein.

[0245] In some embodiments, an anti-MS4A4A antibody according to any of the above embodiments may include, alone or in combination, any of the features described in Sections 1-7 below:

[0246] (1) Anti-MS4A4A antibody binding affinity

[0247] In some embodiments of any of the antibodies provided herein, the antibody has a dissociation constant (Kd) of <1 μM, <100 nM, <10 nM, <1 nM, <0.1 nM, <0.01 nM, or <0.001 nM (e.g., 10 -8 M or smaller, such as 10 -8 M to 10 - 13 M, for example 10 -9 M to 10 -13M). The dissociation constant can be determined by any analytical technique, including any biochemical or biophysical technique, such as ELISA, surface plasmon resonance (SPR), biolayer interferometry (see, e.g., by ForteBio's Octet system), isothermal titration calorimetry (ITC), differential scanning calorimetry (DSC), circular dichroism (CD), stopped-flow analysis, and colorimetric or fluorescent protein melting analysis. In one embodiment, Kd is measured by a radiolabeled antigen binding assay (RIA). In a certain embodiment, an RIA is performed using a Fab version of the antibody of interest and its antigen, e.g., as described in Chen et al. J. Mol. Biol. 293:865-881 (1999). In some embodiments, Kd is measured using a BIACORE surface plasmon resonance assay, such as an assay performed by a BIACORE-2000 or BIACORE-3000 (BIAcore, Inc., Piscataway, NJ) with an immobilized antigen CM5 chip at approximately 10 response units (RU) at 25°C. In some embodiments, Kd is determined using a monovalent antibody (e.g., Fab) or a full-length antibody. D In some embodiments, K is determined using a monovalent form of the full-length antibody. D .

[0248] (2) Antibody fragments

[0249] In some embodiments of any of the antibodies provided herein, the antibody is an antibody fragment. Antibody fragments include, but are not limited to, Fab, Fab', Fab'-SH, F(ab')2, Fv and scFv fragments, and other fragments described below. For a review of certain antibody fragments, see Hudson et al. Nat. Med. 9:129-134 (2003). For a review of scFv fragments, see, for example, WO 93 / 16185; and U.S. Patent Nos. 5571894 and 5587458. For a discussion of Fab and F(ab')2 fragments comprising salvage receptor binding epitope residues and having increased in vivo half-life, see U.S. Patent No. 5869046.

[0250] Diabodies are antibody fragments with two antigen-binding sites that can be bivalent or bispecific. See, e.g., EP404097; WO 1993 / 01161; Hudson et al. Nat. Med. 9:129-134 (2003). Tribodies and tetrabodies are also described in Hudson et al., Nat. Med. 9:129-134 (2003). Single-domain antibodies are antibody fragments that contain all or a portion of the heavy chain variable domain or all or a portion of the light chain variable domain of an antibody. In certain embodiments, a single-domain antibody is a human single-domain antibody (see, e.g., U.S. Patent No. 6,248,516).

[0251] As described herein, antibody fragments can be prepared by a variety of techniques, including, but not limited to, proteolytic digestion of intact antibodies and production by recombinant host cells (eg, E. coli or phage).

[0252] (3) Chimeric and humanized antibodies

[0253] In some embodiments of any one of the antibodies provided herein, the antibody is a chimeric antibody. Certain chimeric antibodies are, for example, described in U.S. Patent No. 4,816,567. In one example, a chimeric antibody comprises a non-human variable region (e.g., a variable region derived from a mouse, rat, hamster, rabbit, or non-human primate (such as a monkey)) and a human constant region. In another example, a chimeric antibody is a "class switching" antibody, in which the class or subclass has been changed from the class or subclass of the parent antibody. Chimeric antibodies include their antigen-binding fragments.

[0254] In some embodiments of any one of the antibodies provided herein, the antibody is a humanized antibody. Generally, non-human antibodies are humanized to reduce immunogenicity to humans while retaining the specificity and affinity of the parent non-human antibody. In certain embodiments, humanized antibodies are substantially non-immunogenic in humans. In certain embodiments, the affinity of the humanized antibody to the target is substantially the same as that of an antibody from another species from which the humanized antibody is derived. See, for example, U.S. Patents 5530101, 5693761; 5693762; and 5585089. In certain embodiments, the amino acids of the antibody variable domains that can be modified without reducing the natural affinity of the antigen-binding domain and reducing immunogenicity are identified. See, for example, U.S. Patents 5766886 and 5869619. Generally speaking, a humanized antibody comprises one or more variable domains, wherein HVR (or portions thereof) are derived from non-human antibodies, and FR (or portions thereof) are derived from human antibody sequences. The humanized antibody optionally also comprises at least a portion of a human constant region. In some embodiments, some FR residues in a humanized antibody are substituted with corresponding residues from a non-human antibody (eg, the antibody from which the HVR residues are derived), eg, to restore or improve antibody specificity or affinity.

[0255] Humanized antibodies and methods for their preparation are reviewed, for example, in Almagro et al., Front. Biosci. 13:1619-1633 (2008), and further described, for example, in U.S. Patent Nos. 5821337, 7527791, 6982321, and 7087409. Human framework regions that can be used for humanization include, but are not limited to, framework regions selected using a "best fit" approach (see, for example, Sims et al., J. Immunol. 151:2296 (1993)); framework regions derived from the consensus sequence of human antibodies of a particular subgroup of light chain variable regions or heavy chain variable regions (see, for example, Carter et al., Proc. Natl. Acad. Sci. USA, 89:4285 (1992); and Presta et al., J. Immunol., 151:2623 (1993)). 993)); human mature (somatically mutated) framework regions or human germline framework regions (see, e.g., Almagro and Fransson, Front. Biosci. 13: 1619-1633 (2008)); and framework regions derived from screening FR libraries (see, e.g., Baca et al., J. Biol. Chem. 272: 10678-10684 (1997) and Rosok et al., J. Biol. Chem. 271: 22611-22618 (1996)).

[0256] (4) Human antibodies

[0257] In some embodiments of any of the antibodies provided herein, the antibody is a human antibody. Human antibodies can be produced using various techniques known in the art. Human antibodies are generally described in van Dijk et al., Curr. Opin. Pharmacol. 5: 368-74 (2001) and Lonberg Curr. Opin. Immunol. 20: 450-459 (2008).

[0258] Human antibodies can be prepared by administering an immunogen to a transgenic animal that has been modified to produce complete human antibodies or complete antibodies with human variable regions in response to antigenic challenges. Large fragments of the human Ig locus can be used to engineer mouse antibody production defective mouse strains in the hope that such mice can produce human antibodies in the absence of mouse antibodies. Large fragments of human Ig can retain larger variable gene diversity and the appropriate regulation of antibody production and expression. By utilizing the antibody diversification and selection mechanism of mice and the immune tolerance of human proteins, the human antibody library replicated in these mouse strains can produce high-affinity fully human antibodies against any antigen (including human antigens) of interest. Using hybridoma technology, antigen-specific human MAbs with desired specificity can be produced and selected. Some exemplary methods are described in U.S. Patent number 5545807, EP 546073 and EP 546073. See also, for example, describing XENOMOUSE TM US Patent Nos. 6,075,181 and 6,150,584; Description US Patent No. 5770429; describes KM US Patent No. 7,041,870, and describes The human variable regions of intact antibodies produced by such animals can be further modified, for example, by combining them with different human constant regions.

[0259] Human antibodies can also be prepared by hybridoma-based methods. Human myeloma and mouse-human heteromyeloma cell lines for the production of human monoclonal antibodies have been described. (See, for example, Kozbor. J. Immunol. 133: 3001 (1984) and Boemer et al., J. Immunol. 147: 86 (1991)). Human antibodies produced via human B cell hybridoma technology are also described in Li et al., Proc. Natl. Acad. Sci. USA, 1 03: 3557-3562 (2006). Additional methods include, for example, those described in U.S. Patent No. 7,189,826 (describing the production of monoclonal human IgM antibodies from hybridoma cell lines). The human hybridoma technique (trioma technique) is also described in Vollmers et al., Histology and Histopathology 20(3):927-937 (2005) and Vollmers et al., Methods and Findings in Experimental and Clinical Pharmacology 27(3):185-91 (2005). Human antibodies can also be produced by isolating Fv clone variable domain sequences selected from human-derived phage display libraries. Such variable domain sequences can then be combined with desired human constant domains. Techniques for selecting human antibodies from antibody libraries are described below.

[0260] In some embodiments of any of the antibodies provided herein, the antibodies are human antibodies isolated by in vitro methods and / or screening combinatorial libraries (to obtain antibodies with one or more desired activities). Suitable examples include, but are not limited to, phage display (CAT, Morphosys, Dyax, Biosite / Medarex, Xoma, Symphogen, Alexion (formerly known as Proliferon), Affimed), ribosome display (CAT), yeast display (Adimab), and the like. In certain phage display methods, VH and VL gene libraries are cloned separately by polymerase chain reaction (PCR) and random recombination is performed in phage libraries, which can then be screened for antigen binding phage, as described in Winter et al., Ann. Rev. Immunol. 12: 433-455 (1994). For example, many methods for generating phage display libraries and screening antibodies with desired binding characteristics from such libraries are known in the art. See also Sidhu et al., J. Mol. Biol. 338(2):299-310, 2004; Lee et al., J. Mol. Biol. 340(5):1073-1093, 2004; Fellouse Proc. Natl. Acad. Sci. USA 101(34):12467-12472 (2004); and Lee et al., J. Immunol. Methods 284(-2):119-132 (2004). Phages typically display antibody fragments as single-chain Fv (scFv) fragments or as Fab fragments. Libraries derived from immune sources provide high-affinity antibodies to the immunogen without the need to construct hybridomas. Alternatively, the initial library can be cloned (e.g., from humans) to provide a single source of antibodies against a wide range of non-self antigens and self antigens without any immunization, as described by Griffiths et al., EMBO J. 12: 725-734 (1993). Finally, the initial library can also be prepared synthetically by cloning unrearranged V gene segments from stem cells using PCR primers containing random sequences to encode highly variable HVR3 regions and achieving rearrangement in vitro, as described by Hoogenboom et al., J. Mol. Biol., 227; 381-388, 1992. Patent disclosures describing human antibody phage libraries include, for example, U.S. Patent No. 5,750,373 and U.S. Patent Publication Nos. 2007 / 0292936 and 2009 / 0002360. Antibodies isolated from human antibody libraries are considered to be human antibodies or human antibody fragments herein.

[0261] (5) Constant region including Fc region

[0262] In some embodiments of any one of the antibodies provided herein, the antibody comprises an Fc. In some embodiments, the Fc is a human IgG1, IgG2, IgG3, and / or IgG4 isotype. In some embodiments, the antibody belongs to the IgG class, the IgM class, or the IgA class.

[0263] In certain embodiments of any of the antibodies provided herein, the antibody has an IgG2 isotype. In some embodiments, the antibody contains a human IgG2 constant region. In some embodiments, the human IgG2 constant region comprises an Fc region. In some embodiments, the antibody induces one or more MS4A4A activities or is unrelated to binding to an Fc receptor. In some embodiments, the antibody binds to an inhibitory Fc receptor. In certain embodiments, the inhibitory Fc receptor is inhibitory Fc-gamma receptor IIB (FcγIIB).

[0264] In certain embodiments of any of the antibodies provided herein, the antibody has an IgG1 isotype. In some embodiments, the antibody contains a mouse IgG1 constant region. In some embodiments, the antibody contains a human IgG1 constant region. In some embodiments, the human IgG1 constant region comprises an Fc region. In some embodiments, the antibody binds to an inhibitory Fc receptor. In certain embodiments, the inhibitory Fc receptor is inhibitory Fc-gamma receptor IIB (FcγIIB).

[0265] In certain embodiments of any of the antibodies provided herein, the antibody has an IgG4 isotype. In some embodiments, the antibody contains a human IgG4 constant region. In some embodiments, the human IgG4 constant region comprises an Fc region. In some embodiments, the antibody binds to an inhibitory Fc receptor. In certain embodiments, the inhibitory Fc receptor is an inhibitory Fc-gamma receptor IIB (FcγIIB).

[0266] In certain embodiments of any one of the antibodies provided herein, the antibody has a hybrid IgG2 / 4 isotype. In some embodiments, the antibody comprises an amino acid sequence comprising amino acids 118 to 260 according to the EU numbering of human IgG2 and amino acids 261 to 447 according to the EU numbering of human IgG4 (WO 1997 / 11971; WO 2007 / 106585).

[0267] In some embodiments, the Fc region increases clustering but does not activate complement compared to a corresponding antibody comprising the Fc region that does not comprise the amino acid substitutions. In some embodiments, the antibody induces one or more activities of the target to which the antibody specifically binds. In some embodiments, the antibody binds to MS4A4A.

[0268] It is also desirable to modify the anti-MS4A4A antibodies of the present disclosure to modify effector function and / or increase the serum half-life of the antibody. For example, the Fc receptor binding sites on the constant region can be modified or mutated to remove or reduce binding affinity for certain Fc receptors (such as FcγRI, FcγRII and / or FcγRIII), thereby reducing antibody-dependent cell-mediated cytotoxicity. In some embodiments, effector function is impaired by removing N-glycosylation of the Fc region of the antibody (e.g., in the CH2 domain of IgG). In some embodiments, effector function is impaired by modifying regions such as 233-236, 297 and / or 327-331 of human IgG, as described in WO 99 / 58572 and Armour et al., Molecular Immunology 40:585-593 (2003); Reddy et al., J. Immunology 164:1925-1933 (2000). In other embodiments, modification of the anti-MS4A4A antibodies of the present disclosure to modify effector function to increase selectivity for detection of ITIM-containing FcgRIIb (CD32b) without activating humoral responses, increase clustering of MS4A4A antibodies on neighboring cells, including antibody-dependent cell-mediated cytotoxicity and antibody-dependent cellular phagocytosis, is also desirable.

[0269] For example, to increase the serum half-life of an antibody, a salvage receptor binding epitope can be incorporated into an antibody (particularly an antibody fragment), as described in U.S. Patent No. 5,739,277. As used herein, the term "salvage receptor binding epitope" refers to an epitope of the Fc region of an IgG molecule (e.g., IgG1, IgG2, IgG3, or IgG4) that is responsible for increasing the in vivo serum half-life of the IgG molecule. Other amino acid sequence modifications.

[0270] (6) Multispecific antibodies

[0271] Multispecific antibodies are antibodies that have binding specificity for at least two different epitopes, including those on the same or another polypeptide (e.g., one or more MS4A4A polypeptides disclosed herein). In some embodiments, multispecific antibodies can be bispecific antibodies. In some embodiments, multispecific antibodies can be trispecific antibodies. In some embodiments, multispecific antibodies can be tetraspecific antibodies. Such antibodies can be derived from full-length antibodies or antibody fragments (e.g., F(ab')2 bispecific antibodies). In some embodiments, the multispecific antibody comprises a first antigen-binding region that binds to a first site on MS4A4A and a second antigen-binding region that binds to a second site on MS4A4A. In a certain embodiment, the multispecific antibody comprises a first antigen-binding region that binds to MS4A4A and a second antigen-binding region that binds to a second polypeptide.

[0272] Provided herein are multispecific antibodies comprising a first antigen-binding region comprising six HVRs of an antibody described herein that binds to MS4A4A and a second antigen-binding region that binds to a second polypeptide. In some embodiments, the first antigen-binding region comprises six HVRs of an antibody described herein that binds to MS4A4A. H or V L .

[0273] In some embodiments of any of the multispecific antibodies, the second polypeptide is: a) an antigen that facilitates transport across the blood-brain barrier; (b) an antigen that facilitates transport across the blood-brain barrier selected from the group consisting of transferrin receptor (TR), insulin receptor (HIR), insulin-like growth factor receptor (IGFR), low-density lipoprotein receptor-related proteins 1 and 2 (LPR-1 and 2), diphtheria toxin receptor, CRM 197, llama single domain antibody, TMEM 30 (A), protein transduction domain, TAT, Syn-B, penetratin, polyarginine peptide, vascular peptide, and ANG1005; (c) a pathogenic protein selected from the group consisting of amyloid beta, oligomeric amyloid beta, amyloid beta plaques, amyloid precursor protein or fragments thereof, tau, IAPP, alpha-synuclein, TDP-43, FUS protein, C9orf72 (chromosome 9 open reading frame 72), c9RAN protein, prion protein, PrPSc, huntingtin, calcitonin, superoxide dismutase, ataxia, ataxia 1, ataxia 2, ataxia 3, ataxia 7, ataxia 8, ataxia 10, Lewy bodies, atrial natriuretic factor, islet amyloid polypeptide, insulin, apolipoprotein A1, serum amyloid A, medin, prolactin, transthyretin, lysozyme, β2-microglobulin, gelsolin, corneal epithelial protein, cystatin, immune globin light chain AL, S-IBM protein, repeat-associated non-ATG (RAN) translation product, dipeptide repeat (DPR) peptide, glycine-alanine (GA) repeat peptide, glycine-proline (GP) repeat peptide, glycine-arginine (GR) repeat peptide, proline-alanine (PA) repeat peptide, ubiquitin and proline-arginine (PR) repeat peptide; (d) ligands and / or proteins expressed on immune cells, wherein the ligands and / or proteins are selected from: CD40, OX40, ICOS, CD28, CD137 / 4-1BB, CD27, GITR, PD-L1, CTLA-4, PD-L2, PD-1, B7-H3, B7-H4, HVEM, BTLA, KIR, GAL9, TIM3, A2AR, LAG-3 and phosphatidylserine; and / or (e) proteins, lipids, polysaccharides or glycolipids expressed on one or more tumor cells and any combination thereof.

[0274] Many antigens that facilitate transport across the blood-brain barrier are known in the art (see, e.g., Gabathuler R. Neurobiol. Dis. 37:48-57 (2010)). Such second antigens include, but are not limited to, transferrin receptor (TR), insulin receptor (HIR), insulin-like growth factor receptor (IGFR), low-density lipoprotein receptor-related proteins 1 and 2 (LPR-1 and 2), diphtheria toxin receptor (including CRM 197 (a non-toxic mutant of diphtheria toxin)), llama single domain antibodies (such as TMEM 30 (A) (Flippase)), protein transduction domains (such as TAT), Syn-B or transmembrane peptides, polyarginine or generally positively charged peptides, vascular peptides (such as ANG1005) (see, e.g., Gabathuler, 2010) and other cell surface proteins enriched on blood-brain barrier endothelial cells (see, e.g., Daneman et al., PLoS One 5 (10): e13741 (2010)).

[0275] The multivalent antibody may recognize the MS4A4A antigen, as well as, but not limited to, additional antigens, Aβ peptide antigens, or α-synuclein antigens, or tau protein antigens, or TDP-43 protein antigens, or prion protein antigens, or huntingtin protein antigens, or RAN, translation product antigens (including dipeptide repeats (DPR peptides) consisting of glycine-alanine (GA), glycine-proline (GP), glycine-arginine (GR), proline-alanine (PA), or proline-arginine (PR), insulin receptor, insulin-like growth factor receptor. Transferrin receptor or any other antigen that facilitates the transfer of antibodies across the blood-brain barrier. In some embodiments, the second polypeptide is transferrin. In some embodiments, the second polypeptide is tau. In some embodiments, the second polypeptide is Aβ. In some embodiments, the second polypeptide is TREM2. In some embodiments, the second polypeptide is α-synuclein.

[0276] A multivalent antibody comprises at least one polypeptide chain (and preferably two polypeptide chains), wherein one or more polypeptide chains comprise two or more variable domains. For example, one or more polypeptide chains may comprise VD1-(X1) n -VD2-(X2) n -Fc, wherein VD1 is the first variable domain, VD2 is the second variable domain, Fc is a polypeptide chain of the Fc region, X1 and X2 represent amino acids or polypeptides, and n is 0 or 1. Similarly, one or more polypeptide chains may comprise V H -C H 1-Flexible joint-V H -C H 1-Fc region chain; or V H -CH 1-V H -C H 1-FC region chain. The multivalent antibody herein preferably further comprises at least two (and preferably four) light chain variable domain polypeptides. The multivalent antibody herein may, for example, comprise from about two to about eight light chain variable domain polypeptides. The light chain variable domain polypeptides contemplated herein comprise a light chain variable domain and, optionally, further comprise a CL domain.

[0277] Techniques for preparing multispecific antibodies include, but are not limited to, recombinant co-expression of two immunoglobulin heavy chain-light chain pairs with different specificities (see Milstein and Cuello Nature 305:537 (1983), WO 93 / 08829 and Traunecker et al., EMBO J. 10:3655 (1991)), and "knob-in-hole" engineering (see, e.g., U.S. Patent No. 5,731,168). See also WO 2013 / 026833 (CrossMab). Multispecific antibodies can also be prepared by engineering electrostatic steering effects for preparing antibody Fc heterodimeric molecules (WO 2009 / 089004A1); cross-linking two or more antibodies (see, e.g., U.S. Patent No. 4,676,980); using leucine; using "diabody" technology to prepare bispecific antibody fragments (see, e.g., Hollinger et al., Proc. Natl. Acad. Sci. USA 90:6444-6448 (1993)); and using single-chain Fv (scFv) dimers (see, e.g., Gruber et al., J. Immunol. 152:5368 (1994)); and preparing trispecific antibodies as described, e.g., in Tutt et al., J. Immunol. 147:60 (1991).

[0278] Also included herein are engineered antibodies with three or more functional antigen-binding sites, including "octopus antibodies" (see, e.g., US 2006 / 0025576). Antibodies herein also include "dual-acting FAbs" or "DAFs," which contain antigen-binding sites that bind to multiple MS4A4A proteins (see, e.g., US 2008 / 0069820).

[0279] (7) Antibody variants

[0280] In some embodiments of any of the antibodies provided herein, amino acid sequence variants of the antibodies are contemplated. For example, it may be desirable to improve the binding affinity and / or other biological properties of the antibody.

[0281] (i) Substitution, insertion, and deletion variants

[0282] In some embodiments of any one of the antibodies provided herein, antibody variants with one or more amino acid substitutions are provided. Appropriate modifications can be introduced into the nucleotide sequence encoding the antibody, or amino acid sequence variants of the antibody can be prepared by peptide synthesis. Such modifications include, for example, deletions and / or insertions and / or substitutions of residues within the amino acid sequence of the antibody.

[0283] Table A: Amino Acid Substitutions

[0284]

[0285] Substantial modification of the biological properties of the antibody is achieved by selecting substitutions that differ significantly in their effect on maintaining: (a) the structure of the polypeptide backbone in the area of ​​substitution (e.g., sheet or helical conformation), (b) the charge or hydrophobicity of the molecule at the target site, or (c) the bulk of the side chain. Naturally occurring residues are divided into several groups based on their common side chain properties:

[0286] (1) Hydrophobicity: norleucine, Met, Ala, Val, Leu, Ile;

[0287] (2) Neutral hydrophilicity: Cys, Ser, Thr, Asn, Gln;

[0288] (3) Acidic: Asp, Glu;

[0289] (4) Basic: His, Lys, Arg;

[0290] (5) Residues that affect chain orientation: Gly, Pro; and

[0291] (6) Aromatic: Trp, Tyr, Phe.

[0292] For example, non-conservative substitutions may involve exchanging a member of one of these classes for a member of another class. For example, such substituted residues may be introduced into regions of a human antibody that are homologous to a non-human antibody, or into non-homologous regions of the molecule.

[0293] According to certain embodiments, when altering the polypeptides or antibodies described herein, the hydropathic index of amino acids can be considered. Based on the hydrophobicity and charge characteristics of the amino acids, each amino acid has been assigned a hydropathic index. They are: isoleucine (+4.5); valine (+4.2); leucine (+3.8); phenylalanine (+2.8); cysteine / cystine (+2.5); methionine (+1.9); alanine (+1.8); glycine (-0.4); threonine (-0.7); serine (-0.8); tryptophan (-0.9); tyrosine (-1.3); proline (-1.6); histidine (-3.2); glutamic acid (-3.5); glutamine (-3.5); aspartic acid (-3.5); asparagine (-3.5); lysine (-3.9); and arginine (-4.5).

[0294] The importance of the hydropathic amino acid index in conferring interactive biological function on proteins is understood in the art. Kyte et al., J. Mol. Biol., 157: 105-131 (1982). It is known that certain amino acids can be substituted for other amino acids with similar hydropathic indices or scores and still retain similar biological activity. When making changes based on the hydropathic index, in certain embodiments, substitutions of amino acids with a hydropathic index within ±2 are included. In certain embodiments, those amino acids within ±1 are included, and in certain embodiments, those amino acids within ±0.5 are included.

[0295] It is also understood in the art that similar amino acid substitutions can be made effectively based on hydrophilicity, particularly where the resulting biologically functional protein or peptide is intended for use in immunological embodiments, as is the case in the present case. In certain embodiments, the maximum local average hydrophilicity of a protein, as governed by the hydrophilicity of the protein's neighboring amino acids, correlates with the immunogenicity and antigenicity of the protein, i.e., the biological properties of the protein.

[0296] The following hydropathicity values ​​have been assigned to these amino acid residues: arginine (+3.0); lysine (+3.0 ± 1); aspartic acid (+3.0 ± 1); glutamic acid (+3.0 ± 1); serine (+0.3); asparagine (+0.2); glutamine (+0.2); glycine (0); threonine (-0.4); proline (-0.5 ± 1); alanine (-0.5); histidine (-0.5); cysteine ​​(-1.0); methionine (-1.3); valine (-1.5); leucine (-1.8); isoleucine (-1.8); tyrosine (-2.3); phenylalanine (-2.5) and tryptophan (-3.4). When making changes based on similar hydrophilicity values, in certain embodiments, substitutions of amino acids with hydrophilicity values ​​within ±2 are included, in certain embodiments, substitutions of amino acids within ±1 are included, and in certain embodiments, substitutions of amino acids within ±0.5 are included. Epitopes can also be identified from the primary amino acid sequence based on hydrophilicity. These regions are also referred to as "epitope core regions."

[0297] In certain embodiments, substitutions, insertions or deletions may occur within one or more HVRs, as long as these changes do not substantially reduce the ability of the antibody to bind to antigens. For example, conservative changes that do not substantially reduce binding affinity may be made in HVRs (e.g., conservative substitutions as provided herein). Such changes may be, for example, outside the antigen contact residues in HVRs. In certain embodiments of the variant VH and VL sequences provided above, each HVR is unchanged or contains no more than one, two or three amino acid substitutions.

[0298] Amino acid sequence insertions include amino and / or carboxyl terminal fusions ranging in length from one residue to polypeptides comprising one hundred or more residues, as well as intrasequence insertions of single or multiple amino acid residues. Examples of terminal insertions include antibodies with an N-terminal methionyl residue. Other insertion variants of the antibody molecule include fusions of the N- or C-terminus of the antibody with an enzyme (e.g., for ADEPT) or a polypeptide that extends the serum half-life of the antibody.

[0299] Any cysteine ​​residues not involved in maintaining the proper conformation of the antibody may also be substituted, usually with serine, to improve the oxidative stability of the molecule and prevent abnormal cross-linking. Conversely, one or more cysteine ​​bonds may be added to the antibody to improve its stability (especially where the antibody is an antibody fragment, such as an Fv fragment).

[0300] (ii) Glycosylation variants

[0301] In some embodiments of any one of the antibodies provided herein, the antibody is altered to increase or decrease the degree of glycosylation of the antibody. Addition or deletion of glycosylation sites of the antibody can be conveniently achieved by altering the amino acid sequence to create or remove one or more glycosylation sites.

[0302] Glycosylation of antibodies is usually N-connected or O-connected. N-connection refers to the attachment of the carbohydrate moiety to the side chain of asparagine residues. The tripeptide sequences asparagine-X-serine and asparagine-X-threonine are recognition sequences for enzymatic attachment of the carbohydrate moiety to the asparagine side chain, where X is any amino acid except proline. Therefore, the presence of any of these tripeptide sequences in the polypeptide creates a potential glycosylation site. O-linked glycosylation refers to the attachment of a sugar (i.e., one of N-acetylgalactosamine, galactose, or xylose) to a hydroxyamino acid (most commonly serine or threonine, but 5-hydroxyproline or 5-hydroxylysine can also be used).

[0303] The addition of glycosylation sites to the antibody is conveniently accomplished by altering the amino acid sequence such that the antibody contains one or more of the above-described tripeptide sequences (for N-linked glycosylation sites). The alteration can also be made by the addition of one or more serine or threonine residues to the sequence of the original antibody, or by replacement of one or more serine or threonine residues (for O-linked glycosylation sites).

[0304] In the case where the antibody comprises an Fc district, the carbohydrate connected thereto can be changed. The natural antibody produced by mammalian cells generally comprises the biantennary oligosaccharide of side chain, and the oligosaccharide is generally connected to the Asn297 (according to Kabat numbering) of the CH2 domain in the Fc district by N-connection. Oligosaccharide can include various carbohydrates, such as mannose, N-acetylglucosamine (GlcNAc), galactose and sialic acid, and the fucose of the GlcNAc in the " stem " of the biantennary oligosaccharide structure. In some embodiments, the oligosaccharide in the antibody of the present disclosure can be modified to produce antibody variants with some improved properties.

[0305] In one embodiment, antibody variants are provided that have a carbohydrate structure that lacks fucose attached (directly or indirectly) to the Fc region. See, e.g., U.S. Patent Publication Nos. 2003 / 0157108 and 2004 / 0093621. Examples of disclosures involving “defucosylated” or “fucose-deficient” antibody variants include: US 2003 / 0157108; US 2003 / 0115614; US 2002 / 0164328; US 2004 / 0093621; US ​​2004 / 0132140; US 2004 / 0110704; US 2004 / 0110282; US 2004 / 0109865; Okazaki et al., J. Mol. Biol. 336:1239-1249 (2004); Yamane-Ohnuki et al., Biotech. Bioeng. 87:614 (2004). Examples of cell lines capable of producing defucosylated antibodies include Led 3 CHO cells deficient in protein fucosylation (Ripka et al., Arch. Biochem. Biophys. 249:533-545 (1986); US 2003 / 0157108), and knockout cell lines, such as α-1,6-fucosyltransferase gene, FUT8, knockout CHO cells (see, e.g., Yamane-Ohnuki et al., Biotech. Bioeng. 87:614 (2004) and Kanda et al., Biotechnol. Bioeng. 94(4):680-688 (2006)).

[0306] (iii) Modified constant region

[0307] In some embodiments of any of the antibodies provided herein, the antibody Fc is an antibody Fc isotype and / or modification. In some embodiments, the antibody Fc isotype and / or modification is capable of binding to an Fcγ receptor.

[0308] In some embodiments of any of the antibodies provided herein, the modified antibody Fc is an IgG1 modified Fc. In some embodiments, the IgG1 modified Fc comprises one or more modifications. For example, in some embodiments, the IgG1 modified Fc comprises one or more amino acid substitutions (e.g., relative to a wild-type Fc region of the same isotype). In some embodiments, the one or more amino acid substitutions are selected from N297A (Bolt S et al., (1993) Eur J Immunol 23:403-411), D265A (Shields et al., (2001) RJ Biol. Chem. 276, 6591-6604), L234A, L235A (Hutchins et al., (1995) Proc Natl Acad Sci USA, 92: 11980-11984; Alegre et al., (1994) Transplantation 57: 1537-1543.31; Xu et al., (2000) Cell Immunol, 200: 16-26), G237A (Alegre et al., (1994) Transplantation 57: 1537-1543.31; Xu et al., (2000) Cell Immunol, 200: 16-26). Immunol, 200: 16-26), C226S, C229S, E233P, L234V, L234F, L235E (McEarchem et al., (2007) Blood, 109: 1185-1192), P331S (Sazinsky et al., (2008) Proc Natl Acad Sci USA 2008, 105: 20167-20172), S267E, L328F, A330L, M252Y, S254T and / or T256E, where amino acid positions are according to the EU numbering convention.

[0309] In some embodiments of any one of the IgG1 modified Fc, the Fc comprises an N297A mutation according to EU numbering. In some embodiments of any one of the IgG1 modified Fc, the Fc comprises an D265A and N297A mutation according to EU numbering. In some embodiments of any one of the IgG1 modified Fc, the Fc comprises an D270A mutation according to EU numbering. In some embodiments, the IgG1 modified Fc comprises an L234A and L235A mutation according to EU numbering. In some embodiments of any one of the IgG1 modified Fc, the Fc comprises an L234A and G237A mutation according to EU numbering. In some embodiments of any one of the IgG1 modified Fc, the Fc comprises an L234A, L235A, and G237A mutation according to EU numbering. In some embodiments of any one of the Fc modified by IgG1, Fc comprises one or more (including all mutations) in P238D, L328E, E233, G237D, H268D, P271G and A330R mutations according to EU numbering. In some embodiments of any one of the Fc modified by IgG1, Fc comprises one or more (including all mutations) in S267E / L328F mutations according to EU numbering. In some embodiments of any one of the Fc modified by IgG1, Fc comprises one or more in P238D, L328E, E233D, G237D, H268D, P271G and A330R mutations according to EU numbering. In some embodiments of any one of the Fc modified by IgG1, Fc comprises one or more in P238D, L328E, E233D, G237D, H268D, P271G and A330R mutations according to EU numbering. In some embodiments of any one of the Fc modified by IgG1, Fc comprises P238D, S267E, L328E, E233D, G237D, H268D, P271G and A330R mutations according to EU numbering. In some embodiments of any one of the Fc modified by IgG1, Fc comprises P238D, S267E, L328E, G237D, H268D, P271G and A330R mutations according to EU numbering. In some embodiments of any one of the Fc modified by IgG1, Fc comprises C226S, C229S, E233P, L234V and L235A mutations according to EU numbering. In some embodiments of any one of the Fc modified by IgG1, Fc comprises L234F, L235E and P331S mutations according to EU numbering. In some embodiments of any of the IgG1 modified Fc's, the Fc comprises S267E and L328F mutations according to EU numbering. In some embodiments of any of the IgG1 modified Fc's, the Fc comprises an S267E mutation according to EU numbering.In some embodiments of any of the IgG1 modified Fcs, the Fc comprises replacement of the constant heavy chain 1 (CH1) and hinge region with CH1 and replacement of the hinge region of IgG2 (amino acids 118-230 of IgG2 according to EU numbering) with a kappa light chain.

[0310] In some embodiments of any one of the Fc modified by IgG1, compared with the corresponding antibody with Fc district (not comprising two or more amino acid replacements), Fc includes two or more amino acid replacements, and the amino acid replacement increases antibody clustering but does not activate complement. Therefore, in some embodiments of any one of the Fc modified by IgG1, the Fc modified by IgG1 is an antibody comprising Fc district, wherein the antibody is included in the amino acid replacement at position E430G and one or more amino acid replacements in the Fc district, the residue position of the amino acid replacement is selected from: according to EU numbering L234F, L235A, L235E, S267E, K322A, L328F, A330S, P331S and any combination thereof. In some embodiments, the Fc modified by IgG1 includes the amino acid replacement at position E430G, L243A, L235A and P331S according to EU numbering. In some embodiments, the Fc modified by IgG1 includes the amino acid replacement at position E430G and P331S according to EU numbering. In some embodiments, the IgG1 modified Fc comprises amino acid replacements at positions E430G and K322A according to EU numbering. In some embodiments, the IgG1 modified Fc comprises amino acid replacements at positions E430G, A330S, and P331S according to EU numbering. In some embodiments, the IgG1 modified Fc comprises amino acid replacements at positions E430G, K322A, A330S, and P331S according to EU numbering. In some embodiments, the IgG1 modified Fc comprises amino acid replacements at positions E430G, K322A, A330S, and P331S according to EU numbering. In some embodiments, the IgG1 modified Fc comprises amino acid replacements at positions E430G, K322A, and A330S according to EU numbering. In some embodiments, the IgG1 modified Fc comprises amino acid replacements at positions E430G, K322A, and P331S according to EU numbering.

[0311] In some embodiments of any of the IgG1 modified Fcs, the IgG1 modified Fc may further comprise an A330L mutation according to the EU numbering convention (Lazar et al., Proc Natl Acad Sci USA, 103:4005-4010 (2006)), or one or more of L234F, L235E, and / or P331S mutations (Sazinsky et al., Proc Natl Acad Sci USA, 105:20167-20172 (2008)) to eliminate complement activation. In some embodiments of any of the IgG1 modified Fcs, the IgG1 modified Fc may further comprise one or more of A330L, A330S, L234F, L235E, and / or P331S according to EU numbering convention. In some embodiments of any of the IgG1 modified Fcs, the IgG1 modified Fc may further comprise one or more mutations to enhance antibody half-life in human serum (e.g., one or more (including all) of the M252Y, S254T, and T256E mutations according to the EU numbering convention). In some embodiments of any of the IgG1 modified Fcs, the IgG1 modified Fc may further comprise one or more of E430G, E430S, E430F, E430T, E345K, E345Q, E345R, E345Y, S440Y, and / or S440W according to EU numbering.

[0312] Other aspects of the present disclosure relate to antibodies with modified constant regions (i.e., Fc regions). If the antibody is engineered to eliminate FcgR binding, the antibody that activates the targeted receptor by binding to the FcgR receptor may lose agonist activity (see, e.g., Wilson et al., Cancer Cell 19: 101-113 (2011); Armour et al., Immunology 40: 585-593 (2003); and White et al., Cancer Cell 27: 138-148 (2015)). Therefore, it is believed that when the antibody has an Fc domain CH1 and hinge region from a human IgG2 isotype or another type of Fc domain that can preferentially bind to inhibitory FcgRIIB r receptors or their variants, the anti-MS4A4A antibodies of the present disclosure with the correct epitope specificity can activate the target antigen with minimal adverse reactions.

[0313] In some embodiments of any of the antibodies provided herein, the modified antibody Fc is an IgG2 modified Fc. In some embodiments, the IgG2 modified Fc comprises one or more modifications. For example, in some embodiments, the IgG2 modified Fc comprises one or more amino acid substitutions (e.g., relative to a wild-type Fc region of the same isotype). In some embodiments of any of the IgG2 modified Fcs, one or more amino acid substitutions are selected from V234A according to the EU numbering convention (Alegre et al., Transplantation 57:1537-1543 (1994); Xu et al., Cell Immunol, 200:16-26 (2000)); G237A (Cole et al., Transplantation, 68:563-571 (1999)); H268Q, V309L, A330S, P331S (US 2007 / 0148167; Armour et al., Eur J Immunol 29:2613-2624 (1999); Armour et al., The Haematology Journal 1(Suppl. 1):27 (2000); Armour et al., The Haematology Journal 1(Suppl 1):27(2000)), C219S and / or C220S (White et al., Cancer Cell 27, 138-148(2015)); S267E, L328F (Chu et al., Mol Immunol, 45:3926-3933(2008)); and M252Y, S254T and / or T256E. In some embodiments of any of the IgG2 modified Fcs, the Fc comprises amino acid replacements at positions V234A and G237A according to EU numbering. In some embodiments of any of the IgG2 modified Fcs, the Fc comprises amino acid replacements at positions C219S or C220S according to EU numbering. In some embodiments of any of the IgG2 modified Fcs, the Fc comprises amino acid replacements at positions A330S and P331S according to EU numbering. In some embodiments of any of the IgG2 modified Fcs, the Fc comprises amino acid substitutions at positions S267E and L328F according to EU numbering.

[0314] In some embodiments of any of the IgG2 modified Fcs, the Fc comprises a C127S amino acid substitution according to the EU numbering convention (White et al., (2015) Cancer Cell 27, 138-148; Lightle et al., Protein Sci. 19:753-762 (2010); and WO 2008 / 079246). In some embodiments of any of the IgG2 modified Fcs, the antibody has an IgG2 isotype having a kappa light chain constant domain comprising a C214S amino acid substitution according to the EU numbering convention (White et al., Cancer Cell 27: 138-148 (2015); Lightle et al., Protein Sci. 19:753-762 (2010); and WO 2008 / 079246).

[0315] In some embodiments of any of the IgG2 modified Fcs, the Fc comprises a C220S amino acid substitution according to the EU numbering convention. In some embodiments of any of the IgG2 modified Fcs, the antibody has an IgG2 isotype having a kappa light chain constant domain comprising a C214S amino acid substitution according to the EU numbering convention.

[0316] In some embodiments of any of the IgG2 modified Fc's, the Fc comprises a C219S amino acid substitution according to the EU numbering convention. In some embodiments of any of the IgG2 modified Fc's, the antibody has an IgG2 isotype having a kappa light chain constant domain comprising a C214S amino acid substitution according to the EU numbering convention.

[0317] In some embodiments of any one of the IgG2 modified Fcs, the Fc comprises an IgG2 isotype heavy chain constant domain 1 (CH1) and a hinge region (White et al., Cancer Cell 27: 138-148 (2015)). In certain embodiments of any one of the IgG2 modified Fcs, the IgG2 isotype CH1 and hinge region comprise an amino acid sequence of 118-230 according to EU numbering. In some embodiments of any one of the IgG2 modified Fcs, the antibody Fc region comprises an S267E amino acid replacement, an L328F amino acid replacement, or both, and / or an N297A or N297Q amino acid replacement according to the EU numbering convention.

[0318] In some embodiments of any of the IgG2 modified Fcs, the Fc further comprises one or more amino acid replacements at positions E430G, E430S, E430F, E430T, E345K, E345Q, E345R, E345Y, S440Y, and S440W according to EU numbering. In some embodiments of any of the IgG2 modified Fcs, the Fc may further comprise one or more mutations to enhance antibody half-life in human serum (e.g., one or more (including all) of the M252Y, S254T, and T256E mutations according to EU numbering convention). In some embodiments of any of the IgG2 modified Fcs, the Fc may further comprise A330S and P331S.

[0319] In some embodiments of any of the IgG2 modified Fcs, the Fc is an IgG2 / 4 hybrid Fc. In some embodiments, the IgG2 / 4 hybrid Fc comprises IgG2 aa 118 to 260 and IgG4 aa 261 to 447. In some embodiments of any of the IgG2 modified Fcs, the Fc comprises one or more amino acid substitutions at positions H268Q, V309L, A330S, and P331S according to EU numbering.

[0320] In some embodiments of either of the IgG1 and / or IgG2 modified Fc, the Fc comprises one or more additional amino acid substitutions selected from A330L, L234F, L235E, or P331S according to EU numbering; and any combination thereof.

[0321] In certain embodiments of any one of the IgG1 and / or IgG2 modified Fcs, the Fc comprises one or more amino acid replacements at residue positions selected from C127S, L234A, L234F, L235A, L235E, S267E, K322A, L328F, A330S, P331S, E345R, E430G, S440Y, and any combination thereof, according to EU numbering. In some embodiments of any one of the IgG1 and / or IgG2 modified Fcs, the Fc comprises amino acid replacements at positions E430G, L243A, L235A, and P331S, according to EU numbering. In some embodiments of any one of the IgG1 and / or IgG2 modified Fcs, the Fc comprises amino acid replacements at positions E430G and P331S, according to EU numbering. In some embodiments of any one of the Fc modified by IgG1 and / or IgG2, the Fc comprises amino acid replacements at positions E430G and K322A according to EU numbering. In some embodiments of any one of the Fc modified by IgG1 and / or IgG2, the Fc comprises amino acid replacements at positions E430G, A330S, and P331S according to EU numbering. In some embodiments of any one of the Fc modified by IgG1 and / or IgG2, the Fc comprises amino acid replacements at positions E430G, K322A, A330S, and P331S according to EU numbering. In some embodiments of any one of the Fc modified by IgG1 and / or IgG2, the Fc comprises amino acid replacements at positions E430G, K322A, A330S, and P331S according to EU numbering. In some embodiments of any one of the Fc modified by IgG1 and / or IgG2, the Fc comprises amino acid replacements at positions E430G, K322A, and A330S according to EU numbering. In some embodiments of any one of the Fc modified by IgG1 and / or IgG2, the Fc comprises amino acid replacements at positions E430G, K322A, and P331S according to EU numbering. In some embodiments of any of the IgG1 and / or IgG2 modified Fc, the Fc comprises amino acid replacements at positions S267E and L328F according to EU numbering. In some embodiments of any of the IgG1 and / or IgG2 modified Fc, the Fc comprises amino acid replacements at position C127S according to EU numbering. In some embodiments of any of the IgG1 and / or IgG2 modified Fc, the Fc comprises amino acid replacements at positions E345R, E430G, and S440Y according to EU numbering.

[0322] In some embodiments of any one of the antibodies provided herein, modified antibody Fc is the Fc modified by IgG4. In some embodiments, the Fc modified by IgG4 includes one or more modifications. For example, in some embodiments, the Fc modified by IgG4 includes one or more amino acid replacements (for example, relative to the wild-type Fc district of the same isotype). In some embodiments of any one of the Fc modified by IgG4, one or more amino acid replacements are selected from L235A, G237A, S229P, L236E (Reddy et al., J Immunol 164:1925-1933 (2000)), S267E, E318A, L328F, M252Y, S254T and / or T256E according to EU numbering convention. In some embodiments of any one of the Fc modified by IgG4, Fc can additionally include L235A, G237A and E318A according to EU numbering convention. In some embodiments of any of the IgG4 modified Fcs, the Fc may further comprise S228P and L235E according to the EU numbering convention. In some embodiments of any of the IgG4 modified Fcs, the IgG4 modified Fc may further comprise S267E and L328F according to the EU numbering convention.

[0323] In some embodiments of any of the IgG4 modified Fcs, the IgG4 modified Fc comprises can be combined with an S228P mutation according to the EU numbering convention (Angal et al., Mol Immunol. 30: 105-108 (1993)) and / or with one or more mutations described in (Peters et al., J Biol Chem. 287(29): 24525-33 (2012)) to enhance antibody stability.

[0324] In some embodiments of any of the IgG4 modified Fcs, the IgG4 modified Fc may further comprise one or more mutations to enhance antibody half-life in human serum (e.g., one or more (including all) of the M252Y, S254T, and T256E mutations according to the EU numbering convention).

[0325] In some embodiments of any one of the Fc modified by IgG4, Fc comprises L235E according to EU numbering. In certain embodiments of any one of the Fc modified by IgG4, Fc comprises one or more amino acid replacements at residue positions selected from C127S, F234A, L235A, L235E, S267E, K322A, L328F, E345R, E430G, S440Y and any combination thereof according to EU numbering. In some embodiments of any one of the Fc modified by IgG4, Fc comprises amino acid replacements at position E430G, L243A, L235A and P331S according to EU numbering. In some embodiments of any one of the Fc modified by IgG4, Fc comprises amino acid replacements at position E430G and P331S according to EU numbering. In some embodiments of any one of the Fc modified by IgG4, Fc comprises amino acid replacements at position E430G and P331S according to EU numbering. In some embodiments of any of the IgG4 modified Fc's, the Fc comprises an amino acid replacement at position E430 according to EU numbering. In some embodiments of any of the IgG4 modified Fc's, the Fc region comprises an amino acid replacement at positions E430G and K322A according to EU numbering. In some embodiments of any of the IgG4 modified Fc's, the Fc comprises an amino acid replacement at positions S267E and L328F according to EU numbering. In some embodiments of any of the IgG4 modified Fc's, the Fc comprises an amino acid replacement at position C127S according to EU numbering. In some embodiments of any of the IgG4 modified Fc's, the Fc comprises an amino acid replacement at positions E345R, E430G, and S440Y according to EU numbering.

[0326] (8) Other antibody modifications

[0327] In some embodiments of any one of the antibodies, the antibody is a derivative. The term "derivative" refers to a chemically modified molecule comprising an insertion, deletion or replacement of an amino acid (or nucleic acid). In certain embodiments, the derivative comprises covalent modification, including but not limited to chemical bonding with polymers, lipids or other organic or inorganic moieties. In certain embodiments, the chemically modified antigen-binding proteins can have a longer circulation half-life than the antigen-binding proteins without chemical modification. In certain embodiments, the chemically modified antigen-binding proteins can have improved targeting capabilities for desired cells, tissues and / or organs. In some embodiments, the derivative antigen-binding proteins are covalently modified to comprise one or more water-soluble polymer attachments, including but not limited to polyethylene glycol, polyoxyethylene glycol or polypropylene glycol. See, for example, U.S. Patent Nos. 4,640,835, 4,496,689, 4,301,144, 4,670,417, 4,791,192 and 4,179,337. In certain embodiments, the derivative antigen binding protein comprises one or more polymers, including but not limited to monomethoxy-polyethylene glycol, dextran, cellulose, copolymers of ethylene glycol / propylene glycol, carboxymethyl cellulose, polyvinyl pyrrolidone, poly-1,3-dioxolane, poly-1,3,6-trioxane, ethylene / maleic anhydride copolymers, polyamino acids (homopolymers or random copolymers), poly-(N-vinyl pyrrolidone)-polyethylene glycol, propylene glycol homopolymers, polypropylene oxide / ethylene oxide copolymers, polyoxyethylated polyols (e.g., glycerol) and polyvinyl alcohol, and mixtures of such polymers.

[0328] In certain embodiments, derivatives are covalently modified by polyethylene glycol (PEG) subunits. In certain embodiments, one or more water-soluble polymers are bonded at one or more specific positions of the derivative (e.g., at the amino terminal). In certain embodiments, one or more water-soluble polymers are randomly attached to one or more side chains of the derivative. In certain embodiments, PEG is used to improve the therapeutic ability of antigen-binding proteins. In certain embodiments, PEG is used to improve the therapeutic ability of humanized antibodies. Some such methods are discussed, for example, in U.S. Patent number 6,133,426, which is hereby incorporated by reference for any purpose.

[0329] Peptide analogs are commonly used in the pharmaceutical industry as non-peptide drugs whose properties are similar to those of the template peptide. These types of non-peptide compounds are called "peptide mimetics or peptidomimetics". Fauchere, J. Adv. Drug Res., 15: 29 (1986); and Evans et al., J. Med. Chem., 30: 1229 (1987), which are incorporated herein by reference for any purpose. Such compounds are typically developed using computerized molecular modeling. Peptide mimetics that are structurally similar to therapeutically used peptides can be used to produce similar therapeutic or preventive effects. Typically, mimetics are structurally similar to an exemplary polypeptide (i.e., a polypeptide having biochemical properties or pharmacological activity), such as a human antibody, but have one or more peptide bonds that are optionally replaced by a bond selected from the group consisting of: -CH2NH-, -CH2S-, -CH2-CH2-, -CH-CH- (cis and trans), -COCH2-, -CH(OH)CH2-, and -CH2SO-, by methods well known in the art. Systematic substitution of one or more amino acids of a consensus sequence with a D-amino acid of the same type (e.g., D-lysine for L-lysine) can be used in certain embodiments to generate more stable peptides. In addition, constrained peptides comprising a consensus sequence or substantially identical consensus sequence variations can be generated by methods known in the art (Rizo and Gierasch Ann. Rev. Biochem., 61:387 (1992), incorporated herein by reference for any purpose); for example, by adding internal cysteine ​​residues capable of forming intramolecular disulfide bridges that cyclize the peptide.

[0330] Drug conjugation involves coupling a biologically active cytotoxic (anticancer) payload or drug to an antibody that specifically targets a tumor marker (e.g., a polypeptide that ideally is present only in or on tumor cells). The antibodies track these proteins in vivo and attach themselves to the surface of cancer cells. The biochemical reaction between the antibody and the target protein (antigen) triggers a signal in the tumor cell, which then absorbs or internalizes the antibody along with the cytotoxin. After the ADC is internalized, the cytotoxic drug is released and kills the cancer. Due to this targeting, the drug ideally has fewer side effects and provides a wider therapeutic window than other chemotherapeutic agents. The technology of conjugating antibodies has been disclosed and is known in the art (see, for example, Jane de Lartigue OncLive July 5, 2012; ADC Review on antibody-drug conjugates; and Ducry et al. Bioconjugate Chemistry 21(1):5-13 (2010).

[0331] II. Nucleic Acids, Vectors, and Host Cells

[0332] The anti-MS4A4A antibodies of the present disclosure can be produced using recombinant methods and compositions, for example, as described in U.S. Patent No. 4,816,567. In some embodiments, isolated nucleic acids having a nucleotide sequence encoding any of the anti-MS4A4A antibodies of the present disclosure are provided. Such nucleic acids can encode a V sequence comprising an anti-MS4A4A antibody. L The amino acid sequence and / or comprising V H In some embodiments, one or more vectors (e.g., expression vectors) comprising such nucleic acids are provided. In some embodiments, host cells comprising such nucleic acids are also provided. In some embodiments, the host cell comprises (e.g., has been transduced with): (1) a vector comprising a nucleic acid encoding a V sequence comprising an antibody; L The amino acid sequence of the antibody V H (2) a first vector comprising a V encoding an antibody L A nucleic acid having an amino acid sequence of H In some embodiments, the host cell is a eukaryotic cell, such as a Chinese hamster ovary (CHO) cell or a lymphocyte (e.g., a Y0, NS0, Sp20 cell). The host cells of the present disclosure also include, but are not limited to, isolated cells, cells cultured in vitro, and cells cultured in vitro.

[0333] Methods for preparing the anti-MS4A4A antibodies of the present disclosure are provided. In some embodiments, the methods comprise culturing a host cell of the present disclosure comprising a nucleic acid encoding an anti-MS4A4A antibody under conditions suitable for antibody expression. In some embodiments, the antibody is subsequently recovered from the host cell (or host cell culture medium).

[0334] For recombinant production of the anti-MS4A4A antibodies of the present disclosure, nucleic acid encoding the anti-MS4A4A antibody is isolated and inserted into one or more vectors for further cloning and / or expression in a host cell. Such nucleic acid can be readily isolated and sequenced using conventional procedures (e.g., by using oligonucleotide probes capable of binding specifically to genes encoding the heavy and light chains of the antibody).

[0335] Suitable vectors comprising a nucleic acid sequence encoding any of the anti-MS4A4A antibodies disclosed herein, or a cell surface expressed fragment thereof, including an antibody, include, but are not limited to, cloning vectors and expression vectors. Suitable cloning vectors can be constructed according to standard techniques or can be selected from a large number of cloning vectors available in the art. Although the cloning vector selected can vary depending on the host cell to be used, useful cloning vectors are generally capable of self-replication, can have a single target for a specific restriction endonuclease, and / or can carry a gene for a marker that can be used to select clones containing the vector. Suitable examples include plasmids and bacterial viruses, such as pUC18, pUC19, Bluescript (e.g., pBS SK+) and its derivatives, mpl8, mpl9, pBR322, pMB9, ColE1, pCR1, RP4, phage DNA, and shuttle vectors such as pSA3 and pAT28. These and many other cloning vectors are available from commercial suppliers such as BioRad, Strategene, and Invitrogen.

[0336] Suitable host cells for cloning or expressing antibody encoding vectors include prokaryotic or eukaryotic cells. For example, the anti-MS4A4A antibodies of the present disclosure can be produced in bacteria, especially when glycosylation and Fc effector functions are not desired. Regarding the expression of antibody fragments and polypeptides in bacteria (e.g., U.S. Patent Nos. 5,648,237, 5,789,199, and 5,840,523). After expression, the antibodies can be isolated from the bacterial cytoplasm as a soluble fraction and can be further purified.

[0337] In addition to prokaryotes, eukaryotic microorganisms (such as filamentous fungi or yeast) are suitable cloning or expression hosts for antibody-encoding vectors, including fungal and yeast strains whose glycosylation pathways have been "humanized" to produce antibodies with partial or full human glycosylation patterns (e.g., Gerngross Nat. Biotech. 22: 1409-1414 (2004); and Li et al., Nat. Biotech. 24: 210-215 (2006)).

[0338] Suitable host cells for expressing glycosylated antibodies can also be derived from multicellular organisms (invertebrates and vertebrates). Examples of invertebrate cells include plants and insect cells. Many baculovirus strains have been identified that can be used in conjunction with insect cells, particularly for transfection of Spodoptera frugiperda cells. Plant cell cultures can also be used as hosts (e.g., U.S. Patent Nos. 5,959,177, 6,040,498, 6,420,548, 7,125,978, and 6,417,429, which describe PLANTIBODIES for producing antibodies in transgenic plants). TM technology).

[0339] Vertebrate cells can also be used as hosts. For example, mammalian cell lines suitable for growth in suspension can be useful. Other examples of useful mammalian host cell lines are monkey kidney CV1 lines (COS-7) transformed by SV40; human embryonic kidney cell lines (293 or 293 cells, such as Graham et al. J. Gen Virol. 36: 59 (1977)); baby hamster kidney cells (BHK); mouse Sertoli cells (TM4 cells, such as Mather, Biol. Reprod. 23: 243-251 (1980)); monkey kidney cells (CV1); African green monkey kidney cells (VERO-76); human cervical cancer cells (HELA); canine 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, for example, in Mather et al. Annals NY. Acad. Sci. 383:44-68 (1982); MRC 5 cells; and FS4 cells. Other useful mammalian host cell lines include 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 certain mammalian host cell lines suitable for antibody production, see, for example, Yazaki and Wu, Methods in Molecular Biology, Vol. 248 (BKC Lo, ed., Humana Press, Totowa, NJ), pp. 255-268 (2003).

[0340] III. Pharmaceutical Compositions / Formulations

[0341] Provided herein are pharmaceutical compositions and / or pharmaceutical formulations comprising an anti-MS4A4A antibody of the present disclosure and a pharmaceutically acceptable carrier.

[0342] In some embodiments, the pharmaceutically acceptable carrier is preferably non-toxic to the recipient at the dosage and concentration used. The antibodies described herein can be formulated as preparations in solid, semi-solid, liquid or gaseous form. Examples of such preparations include, but are not limited to, tablets, capsules, powders, granules, ointments, solutions, suppositories, injections, inhalants, gels, microspheres and aerosols. Depending on the desired preparation, the pharmaceutically acceptable carrier may include a pharmaceutically acceptable non-toxic carrier of a diluent, which is a vehicle commonly used in the preparation of pharmaceutical compositions for animal or human administration. In certain embodiments, the pharmaceutical composition may include a formulation material for changing, maintaining or preserving, for example, the pH, osmotic pressure molarity, viscosity, transparency, color, isotonicity, odor, sterility, stability, dissolution or release rate, adsorption or penetration of the composition.

[0343] In certain embodiments, pharmaceutically acceptable carriers include, but are not limited to, amino acids (such as glycine, glutamine, asparagine, arginine, or lysine); antimicrobial agents; antioxidants (such as ascorbic acid, sodium sulfite, or sodium bisulfite); buffers (such as borates, bicarbonates, Tris...

Claims

1. An isolated antibody that binds to human MS4A4A, wherein the antibody comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises HVR-H1 of SEQ ID NO: 18, HVR-H2 of SEQ ID NO: 35, and HVR-H3 of SEQ ID NO: 53, and the light chain variable region comprises HVR-L1 of SEQ ID NO: 72, HVR-L2 of SEQ ID NO: 77, and HVR-L3 of SEQ ID NO:

99.

2. The antibody of claim 1, wherein the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 118, and the light chain variable region comprises the amino acid sequence of SEQ ID NO:

137.

3. The antibody of claim 1, wherein the antibody is a monoclonal antibody.

4. The antibody of claim 3, wherein the monoclonal antibody is a murine antibody, a human antibody, a multivalent antibody, or a chimeric antibody.

5. The antibody of claim 3, wherein the antibody is a humanized antibody.

6. The antibody of any one of claims 1 to 5, wherein the antibody is of the IgG class, the IgM class or the IgA class.

7. The antibody of claim 6, wherein the antibody is of the IgG class and has an IgG1, IgG2 or IgG4 isotype.

8. The antibody of any one of claims 1-5, wherein the antibody is an antibody fragment.

9. The antibody of claim 8, wherein the fragment is a Fab, Fab', Fab'-SH, F(ab')2, Fv or scFv fragment.

10. An isolated nucleic acid comprising a nucleic acid sequence encoding the antibody of any one of claims 1-9. A vector comprising the nucleic acid according to claim 10 .

12. An isolated host cell comprising the vector of claim 11.

13. A method for producing an antibody that binds to human MS4A4A, the method comprising culturing the host cell of claim 12, thereby producing the antibody.

14. The method of claim 13, further comprising recovering the antibody produced by the cell.

15. A pharmaceutical composition comprising the antibody according to any one of claims 1 to 9 and a pharmaceutically acceptable carrier.

16. Use of an antibody as described in any one of claims 1 to 9 in the preparation of a medicament for preventing, reducing the risk of, or treating an individual suffering from a disease, disorder, or injury selected from the group consisting of Alzheimer's disease and cognitive impairment.

17. Use of the antibody of any one of claims 1 to 9 in the preparation of a medicament for preventing late-onset Alzheimer's disease, reducing the risk of late-onset Alzheimer's disease, or treating an individual suffering from late-onset Alzheimer's disease.

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