Anti-SIRP-beta1 antibodies and methods of use thereof

Abstract

The present disclosure relates generally to compositions comprising antibodies (e.g., monoclonal antibodies, antibody fragments, etc.) that specifically bind to a SIRP [beta] 1 polypeptide (e.g., human SIRP [beta] 1), and the use of such compositions in preventing, reducing risk, or treating an individual in need thereof.

Description

Cross-reference to related applications

[0001] This application is a divisional application of the invention application with the application date of June 28, 2019, the Chinese application number of 201980040821.9, and the invention name of "Anti-SIRP-β1 Antibodies and Methods of Using Them". This application claims priority to U.S. Provisional Application No. 62 / 691,913 filed on June 29, 2018, which is hereby incorporated by reference in its entirety for any purpose. Field of the Invention

[0002] The present disclosure relates to anti-SIRPβ1 antibodies and therapeutic uses of such antibodies. Background of the Invention

[0001] Signal regulatory protein β (SIRPβ) belongs to the SIRP family of transmembrane receptors, which are expressed in myeloid cell lineages (including monocytes, macrophages, granulocytes, and dendritic cells) and in neuronal cells. Proteins of the SIRP family are characterized by an extracellular region containing two membrane-proximal IgC domains and one distal IgV domain. Unlike the SIRPα receptor, the SIRPβ protein contains a short cytoplasmic domain that lacks cytoplasmic sequence motifs that can recruit protein tyrosine phosphatases SHP-2 and SHP-1. SIRPβ1 isoform 1 associates with the adaptor protein DAP12 containing a single cytoplasmic immunoreceptor tyrosine-based activation (ITAM) motif. See, e.g., Dietrich et al., 2000, J Immunol 164:9-12; Liu et al., 2005, J Biol Chem 280:36132-36140. SIRPβ has been shown to be a microglial regulator of phagocytosis in Alzheimer's disease. See, eg, Gaikwad et al., 2009, Am J Pathol 175:2528-2539.

[0002] Thus, there is a need for therapeutic anti-SIRPβ1 antibodies to treat diseases, disorders, and conditions associated with SIRPβ1 activity. SUMMARY OF THE INVENTION

[0003] In some embodiments, an isolated antibody that binds to human SIRPβ1 is provided, wherein the antibody has at least one, at least two, at least three, at least four, or at least five properties selected from the following: a) binds to human SIRPβ1 isoform 1, but not to human SIRPα; b) binds to human SIRPβ1 isoform 1, but not to human SIRPγ; c) binds to human SIRPβ1 isoform 1, but not to human SIRPβ1 isoform 3; d) binds to human SIRPβ1 isoform 1, but not to mouse SIRPβ1; e) binds to human SIRPβ1 isoform 1, but not to cynomolgus monkey SIRPβ1 isoform 1; f) stimulating SIRPβ1 activity on CD14-positive monocytes in vitro and / or in vivo; g) inducing or enhancing the respiratory burst in immune cells, such as neutrophils and / or monocytes, in vitro and / or in vivo; h) inducing or increasing IL-8 expression in monocytes in vitro and / or in vivo; i) inducing or increasing TNFα expression in macrophages and / or dendritic cells in vitro and / or in vivo; j) inducing neutrophil-mediated phagocytosis, such as phagocytosis of tumor cells, in vitro and / or in vivo; k) enhancing neutrophil-mediated tumor cell clearance in vivo; 1) increasing TREM2 expression on macrophages in vitro and / or in vivo; m) increasing the viability of macrophages in vitro and / or in vivo when alone and / or in combination with an agonistic anti-TREM2 antibody; and n) Improving the survival of dendritic cells in vitro and / or in vivo.

[0004] In some embodiments, the antibody binds to human SIRPβ1 isoform 1, but does not bind to human SIRPα or human SIRPβ1 isoform 3. In some embodiments, the antibody binds to human SIRPβ1 isoform 1, but does not bind to human SIRPα, SIRPγ, or human SIRPβ1 isoform 3. In some embodiments, the antibody has at least one, at least two, or at least three properties selected from the following: a) stimulating SIRPβ1 activity on CD14-positive monocytes in vitro and / or in vivo; b) inducing or enhancing the respiratory burst in immune cells, such as neutrophils and / or monocytes, in vitro and / or in vivo; c) inducing or increasing IL-8 expression in monocytes in vitro and / or in vivo; d) inducing or increasing TNFα expression in macrophages and / or dendritic cells in vitro and / or in vivo; e) inducing neutrophil-mediated phagocytosis, such as phagocytosis of tumor cells, in vitro and / or in vivo; f) enhancing neutrophil-mediated tumor cell clearance in vivo; g) increasing TREM2 expression on macrophages in vitro and / or in vivo; h) increasing the viability of macrophages in vitro and / or in vivo when alone and / or in combination with an agonistic anti-TREM2 antibody; and i) Improving the survival of dendritic cells in vitro and / or in vivo.

[0005] In some embodiments, an isolated antibody that binds to human SIRPβ1 is provided, wherein the antibody comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises (a) HVR-H1 comprising the HVR-H1 amino acid sequence shown in Table 4 and / or Table 9; (b) HVR-H2 comprising the HVR-H2 amino acid sequence shown in Table 4 and / or Table 9; (c) HVR-H3 comprising the HVR-H3 amino acid sequence shown in Table 4 and / or Table 9; the light chain variable region comprises (d) HVR-L1 comprising the HVR-L1 amino acid sequence shown in Table 3 and / or Table 8; (e) HVR-L2 comprising the HVR-L2 amino acid sequence shown in Table 3 and / or Table 8; and (f) HVR-L3 comprising the HVR-L3 amino acid sequence shown in Table 3 and / or Table 8. In some embodiments, the heavy chain variable region comprises one, two, three or four framework regions selected from VH FR1, VH FR2, VH FR3, and VH FR4 shown in Table 6. In some embodiments, the light chain variable region comprises one, two, three or four framework regions selected from VL FR1, VL FR2, VL FR3, and VL FR4 shown in Table 5.

[0006] In some embodiments, the antibody comprises SB-1, SB-2, SB-3,SB-4,SB-5,SB-6,SB-7,SB-8,SB-9,SB-10,SB-11,SB-12,SB-13,SB-14,SB-15, SB-16,SB-17,SB-18,SB-19,SB-20,SB-21,SB-22,SB-23,SB-24,SB-25,SB-26,SB-27, SB-28,SB-29,SB-30,SB-31,SB-32,SB-33,SB-34,SB-35,SB-36,SB-37,SB-38,SB-39, SB-40,SB-41,SB-42,SB-43,SB-44,SB-45,SB-46,SB-47,SB-48,SB-49,SB-50,SB-1-2, SB-1-3,SB-1-4,SB-1-5,SB-2-7,SB-2-8,SB-2-9,SB-2-10,SB-2-11,SB-8-13,SB-8-14, HVR-H1, HVR-H2, HVR-H3, HVR-L1, HVR-L2, and HVR-L3 of antibodies of SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21. In some embodiments, the antibody comprises a heavy chain variable region comprising a sequence selected from SEQ ID NO:268,270,272,274,276,279,281,283,285,287,289,291,293,295,297,299,301,303 ,305,307,309,311,313,315,317,319,321,323,325,327,329,331,333,335,337,339,34 1,343,345,347,349,351,353,355,357,359,361,363,365,366,367,368,369,371,372,373,374,375,376,377,378,379,380,381, or 382. In some embodiments, the antibody comprises an amino acid sequence selected from SEQ ID NO:268,270,272,274,276,279,281,283,285,287,289,291,293,295,297,299, 301,303,305,307,309,311,313,315,317,319,321,323,325,327,329,331,333, or 382.In some embodiments, the antibody comprises a light chain variable region comprising a sequence selected from SEQ ID or 370. An amino acid sequence that is at least 90%, at least 95%, at least 97%, or at least 99% identical to the sequence of NO: 267, 269, 271, 273, 275, 277, 278, 280, 282, 284, 286, 288, 290, 292, 294, 296, 298, 300, 302, 304, 306, 308, 310, 312, 314, 316, 318, 320, 322, 324, 326, 328, 330, 332, 334, 336, 338, 340, 342, 344, 346, 348, 350, 352, 354, 356, 358, 360, 362, 364, or 370. In some embodiments, the antibody comprises a light chain variable region selected from SEQ ID NO: 267, 269, 271, 273, 275, 277, 278, 280, 282, 284, 286, 288, 290, 292, 294, 296, 298, 300, 302, 304, 306, 308, 310, 312, 314, 316, 318, 320, 322, 324, 326, 328, 330, 332, 334, 336, 338, 340, 342, 344, 346, 348, 350, 352, 354, 356, 358, 360, 362, 364, or 370.In some embodiments, the antibody comprises an antibody selected from the group consisting of SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-10, SB-11, SB-12, SB-13, SB-14, SB-15, SB-16, SB-17, SB-18, SB-19, SB-20, SB-21, SB-22, SB-23, SB-24, SB-25, SB-26, SB-27, SB-28, SB-29, SB-30, SB-31, SB-32, SB-33, SB-34, SB-35, SB-36, SB-37, SB-38, SB The heavy chain variable regions of the antibodies of the invention are at least 90%, at least 95%, at least 97%, or at least 99% identical to those of the antibodies of the invention. The heavy chain variable region, and is selected from the group consisting of SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-10, SB-11, SB-12, SB-13, SB-14, SB-15, SB-16, SB-17, SB-18, SB-19, SB- 20,SB-21,SB-22,SB-23,SB-24,SB-25,SB-26,SB-27,SB-28,SB-29,SB-30,SB-31,SB-32,SB-33,SB-34,SB-35,SB-36,SB-37,SB-38,SB-39,SB SB-40, SB-41, SB-42, SB-43, SB-44, SB-45, SB-46, SB-47, SB-48, SB-49, SB-50, SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21 have a light chain variable region that is at least 90%, at least 95%, at least 97%, or at least 99% identical to the light chain variable region of the antibody.In some embodiments, the antibody comprises an antibody selected from the group consisting of SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-10, SB-11, SB-12, SB-13, SB-14, SB-15, SB-16, SB-17, SB-18, SB-19, SB-20, SB-21, SB-22, SB-23, SB-24, SB-25, SB-26, SB-27, SB-28, SB-29, SB-30, SB-31, SB-32, SB-33, SB-34, SB-35, SB-36, SB-37 , SB-38, SB-39, SB-40, SB-41, SB-42, SB-43, SB-44, SB-45, SB-46, SB-47, SB-48, SB-49, SB-50, SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21.

[0007] In some embodiments, the antibody comprises SB-1, SB-2, SB-3,SB-4,SB-5,SB-6,SB-7,SB-8,SB-9,SB-14,SB-28,SB-39,SB-40,SB-49,SB-1-2, SB-1-3,SB-1-4,SB-1-5,SB-2-7,SB-2-8,SB-2-9,SB-2-10,SB-2-11,SB-8-13,SB-8-14, HVR-H1, HVR-H2, HVR-H3, HVR-L1, HVR-L2, and HVR-L3 of the antibodies SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21. In some embodiments, the antibody comprises a heavy chain variable region that is at least 90%, at least 95%, at least 97%, or at least 99% identical to a heavy chain variable region of an antibody selected from SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-14, SB-28, SB-39, SB-40, SB-49, SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21. In some embodiments, the antibody comprises the heavy chain variable region of an antibody selected from SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-14, SB-28, SB-39, SB-40, SB-49, SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21. In some embodiments, the antibody comprises a light chain variable region that is at least 90%, at least 95%, at least 97%, or at least 99% identical to a light chain variable region of an antibody selected from SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-14, SB-28, SB-39, SB-40, SB-49, SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21.In some embodiments, the antibody comprises a light chain variable region of an antibody selected from SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-14, SB-28, SB-39, SB-40, SB-49, SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21. In some embodiments, the antibody comprises a heavy chain variable region that is at least 90%, at least 95%, at least 97%, or at least 99% identical to an antibody selected from SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-14, SB-28, SB-39, SB-40, SB-49, SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21. The invention further comprises a heavy chain variable region of the antibody selected from the group consisting of SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-14, SB-28, SB-39, SB-40, SB-49, SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21. In some embodiments, the antibody comprises the heavy chain variable region and the light chain variable region of an antibody selected from SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-14, SB-28, SB-39, SB-40, SB-49, SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21.

[0008] In some embodiments, the antibody comprises an SB-1-2, SB-1-3, SB-1-4,SB-1-5,SB-2-7,SB-2-8,SB-2-9,SB-2-10,SB-2-11,SB-8-13,SB-8-14,SB-8-15, In some embodiments, the antibody comprises a heavy chain variable region that is at least 90%, at least 95%, at least 97%, or at least 99% identical to a heavy chain variable region of an antibody selected from the group consisting of SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21. In some embodiments, the antibody comprises the heavy chain variable region of an antibody selected from SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21. In some embodiments, the antibody comprises a light chain variable region that is at least 90%, at least 95%, at least 97%, or at least 99% identical to a light chain variable region of an antibody selected from SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21. In some embodiments, the antibody comprises the light chain variable region of an antibody selected from SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21.In some embodiments, the antibody comprises a heavy chain variable region that is at least 90%, at least 95%, at least 97%, or at least 99% identical to an antibody selected from SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21. and a light chain variable region that is at least 90%, at least 95%, at least 97%, or at least 99% identical to a light chain variable region of an antibody selected from the group consisting of SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21. In some embodiments, the antibody comprises the heavy chain variable region and the light chain variable region of an antibody selected from SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21.

[0009] In some embodiments, the antibody comprises (a) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 80, 228, or 229; (b) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 101, 239, 239, 240, or 241; (c) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 125; (d) an HVR-L1 comprising the amino acid sequence of SEQ ID NO: 383; (e) an HVR-L2 comprising the amino acid sequence of SEQ ID NO: 16; and (f) an HVR-L3 comprising the amino acid sequence of SEQ ID NO: 31. In some embodiments, the antibody comprises a heavy chain variable region that is at least 90%, at least 95%, at least 97%, or at least 99% identical to the amino acid sequence of SEQ ID NO: 366, 367, 368, or 369. In some embodiments, the antibody comprises a light chain variable region that is at least 90%, at least 95%, at least 97%, or at least 99% identical to the amino acid sequence of SEQ ID NO: 267. In some embodiments, the antibody comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 366, 367, 368, or 369. In some embodiments, the antibody comprises a light chain variable region comprising the amino acid sequence of SEQ ID NO: 267.

[0010] In some embodiments, the antibody comprises (a) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 81, 99, 230, 231, or 232; (b) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 102, 242, 243, 244, or 245; (c) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 126 or 253; (d) an HVR-L1 comprising the amino acid sequence of SEQ ID NO: 383; (e) an HVR-L2 comprising the amino acid sequence of SEQ ID NO: 16; and (f) an HVR-L3 comprising the amino acid sequence of SEQ ID NO: 32. In some embodiments, the antibody comprises a heavy chain variable region that is at least 90%, at least 95%, at least 97%, or at least 99% identical to the amino acid sequence of SEQ ID NO: 371, 372, 373, 374, or 375. In some embodiments, the antibody comprises a light chain variable region that is at least 90%, at least 95%, at least 97%, or at least 99% identical to the amino acid sequence of SEQ ID NO: 269 or 370. In some embodiments, the antibody comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 371, 372, 373, 374, or 375. In some embodiments, the antibody comprises a light chain variable region comprising the amino acid sequence of SEQ ID NO: 269 or 370. In some embodiments, the antibody comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 371, 372, or 373 and a light chain variable region comprising the amino acid sequence of SEQ ID NO: 370, or the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 374 or 375 and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 269.

[0011] In some embodiments, the antibody comprises (a) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 85, 233, or 234; (b) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 107, 246, 247, or 248; (c) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 131 or 254; (d) an HVR-L1 comprising the amino acid sequence of SEQ ID NO: 383; (e) an HVR-L2 comprising the amino acid sequence of SEQ ID NO: 19; and (f) an HVR-L3 comprising the amino acid sequence of SEQ ID NO: 37. In some embodiments, the antibody comprises a heavy chain variable region that is at least 90%, at least 95%, at least 97%, or at least 99% identical to the amino acid sequence of SEQ ID NO: 376, 377, or 378. In some embodiments, the antibody comprises a light chain variable region that is at least 90%, at least 95%, at least 97%, or at least 99% identical to the amino acid sequence of SEQ ID NO: 280. In some embodiments, the antibody comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 376, 377, or 378. In some embodiments, the antibody comprises a light chain variable region comprising the amino acid sequence of SEQ ID NO: 280.

[0012] In some embodiments, the antibody comprises (a) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 97, 235, 236, or 237; (b) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 121, 249, 250, 251, or 252; (c) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 163; (d) an HVR-L1 comprising the amino acid sequence of SEQ ID NO: 2; (e) an HVR-L2 comprising the amino acid sequence of SEQ ID NO: 22; and (f) an HVR-L3 comprising the amino acid sequence of SEQ ID NO: 69. In some embodiments, the antibody comprises a heavy chain variable region that is at least 90%, at least 95%, at least 97%, or at least 99% identical to the amino acid sequence of SEQ ID NO: 379, 380, 381, or 382. In some embodiments, the antibody comprises a light chain variable region that is at least 90%, at least 95%, at least 97%, or at least 99% identical to the amino acid sequence of SEQ ID NO: 344. In some embodiments, the antibody comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 379, 380, 381, or 382. In some embodiments, the antibody comprises a light chain variable region comprising the amino acid sequence of SEQ ID NO: 344.

[0013] In some embodiments, the antibody comprises (a) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 229; (b) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 239; (c) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 125; (d) an HVR-L1 comprising the amino acid sequence of SEQ ID NO: 383; (e) an HVR-L2 comprising the amino acid sequence of SEQ ID NO: 16; and (f) an HVR-L3 comprising the amino acid sequence of SEQ ID NO: 31. In some embodiments, the antibody comprises a heavy chain variable region that is at least 90%, at least 95%, at least 97%, or at least 99% identical to the amino acid sequence of SEQ ID NO: 367. In some embodiments, the antibody comprises a light chain variable region that is at least 90%, at least 95%, at least 97%, or at least 99% identical to the amino acid sequence of SEQ ID NO: 267. In some embodiments, the antibody comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 367. In some embodiments, the antibody comprises a light chain variable region comprising the amino acid sequence of SEQ ID NO: 267.

[0014] In some embodiments, the antibody comprises (a) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 231; (b) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 243; (c) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 126; (d) an HVR-L1 comprising the amino acid sequence of SEQ ID NO: 383; (e) an HVR-L2 comprising the amino acid sequence of SEQ ID NO: 16; and (f) an HVR-L3 comprising the amino acid sequence of SEQ ID NO: 32. In some embodiments, the antibody comprises a heavy chain variable region that is at least 90%, at least 95%, at least 97%, or at least 99% identical to the amino acid sequence of SEQ ID NO: 372. In some embodiments, the antibody comprises a light chain variable region that is at least 90%, at least 95%, at least 97%, or at least 99% identical to the amino acid sequence of SEQ ID NO: 370. In some embodiments, the antibody comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 372. In some embodiments, the antibody comprises a light chain variable region comprising the amino acid sequence of SEQ ID NO: 270.

[0015] In some embodiments, the antibody comprises (a) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 233; (b) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 246; (c) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 131; (d) an HVR-L1 comprising the amino acid sequence of SEQ ID NO: 383; (e) an HVR-L2 comprising the amino acid sequence of SEQ ID NO: 19; and (f) an HVR-L3 comprising the amino acid sequence of SEQ ID NO: 37. In some embodiments, the antibody comprises a heavy chain variable region that is at least 90%, at least 95%, at least 97%, or at least 99% identical to the amino acid sequence of SEQ ID NO: 376. In some embodiments, the antibody comprises a light chain variable region that is at least 90%, at least 95%, at least 97%, or at least 99% identical to the amino acid sequence of SEQ ID NO: 280. In some embodiments, the antibody comprises a heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 376. In some embodiments, the antibody comprises a light chain variable region comprising the amino acid sequence of SEQ ID NO: 280.

[0016] In some embodiments, the antibody is a monoclonal antibody. In some embodiments, the antibody is of the IgG class, IgM class, or IgA class. In some embodiments, the antibody is of the IgG class and has an IgG1, IgG2, or IgG4 isotype. In some embodiments, the antibody has an IgG1 isotype. In some embodiments, the antibody comprises an E430G substitution and a P331S substitution according to EU numbering.

[0017] 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.

[0018] In some embodiments, the antibody has an affinity (K for human SIRPβ1 isoform 1) of 0.1 nM to 50 nM, or 0.5 nM to 10 nM, or 0.5 nM to 5 nM. D In some embodiments, affinity is measured using ForteBio System measured.

[0019] In some embodiments, the antibody recognizes a first and a second antigen, wherein the first antigen is SIRPβ1 and the second antigen is: (a) Antigens that promote transport across the blood-brain barrier; (b) an antigen that promotes 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), and diphtheria toxin receptor; (c) a pathogenic agent 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, and the pathogenic protein is selected from amyloid β, oligomeric amyloid β, amyloid β plaques, amyloid precursor protein or fragments thereof, Tau, IAPP, α-synuclein, TDP-43, FUS protein, C9orf72 (chromosome 9 open reading frame 72), c9RAN protein, prion protein, PrPSc, huntingtin protein, calcitonin, superoxide dismutase, ataxin, ataxin 1, ataxin 2, ataxin 3, ataxin 7, ataxin 8, ataxia-10, Lewy body, atrial natriuretic factor, islet amyloid polypeptide, insulin, apolipoprotein AI, serum amyloid A, medin, prolactin, transthyretin, lysozyme, β2-microglobulin, gelsolin, corneal epithelial protein, cystatin, immunoglobulin 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 group; (d) a ligand and / or protein expressed on an immune cell, 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.

[0020] In some embodiments, an isolated antibody that binds to human SIRPβ1 is provided, wherein the antibody competes with one or more antibodies provided herein for binding to human SIRPβ1 isoform 1. In some embodiments, an isolated antibody that binds to human SIRPβ1 is provided, wherein the antibody binds to substantially the same or overlapping epitope of SIRPβ1 isoform 1 as one or more antibodies provided herein.

[0021] In some embodiments, an isolated nucleic acid is provided, comprising a nucleic acid sequence encoding an anti-SIRPβ1 antibody provided herein. In some embodiments, a vector is provided, comprising the nucleic acid. In some embodiments, an isolated host cell is provided, comprising the vector. In some embodiments, an isolated host cell is provided, which expresses the anti-SIRPβ1 antibody provided herein. In some embodiments, a method for producing an antibody that binds to human SIRPβ1 is provided, comprising culturing a host cell expressing the anti-SIRPβ1 antibody provided herein, so that the antibody is produced. In some embodiments, the method further comprises recovering the antibody produced by the cell.

[0022] In some embodiments, a pharmaceutical composition is provided, comprising an anti-SIRPβ1 antibody provided herein and a pharmaceutically acceptable carrier.

[0023] In some embodiments, a method of treating cancer is provided, comprising administering a therapeutically effective amount of an anti-SIRPβ1 antibody provided herein to an individual in need thereof. In some embodiments, the cancer is selected from sarcoma, bladder cancer, brain cancer, breast cancer, colon cancer, rectal cancer, endometrial cancer, kidney cancer, renal pelvic cancer, leukemia, lung cancer, small cell lung cancer, melanoma, lymphoma, pancreatic cancer, prostate cancer, ovarian cancer, and fibrosarcoma, glioblastoma multiforme, renal clear cell carcinoma, adrenocortical carcinoma, bladder urothelial carcinoma, diffuse large B-cell lymphoma, lung adenocarcinoma, pancreatic adenocarcinoma, renal cell carcinoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, indolent B-cell lymphoma, aggressive B-cell lymphoma, T-cell lymphoma, acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), chronic lymphocytic leukemia (CLL), CLL, chronic myeloid leukemia (CML), multiple myeloma, myelodysplastic syndrome, myeloproliferative neoplasms, invasive breast carcinoma, cervical squamous cell carcinoma, endocervical adenocarcinoma, bile duct carcinoma, colon adenocarcinoma, diffuse large B-cell lymphoma, esophageal carcinoma, head and neck squamous cell carcinoma, renal chromophobe cell carcinoma, renal papillary cell carcinoma, low-grade glioma, hepatocellular carcinoma, lung squamous cell carcinoma, mesothelioma, ovarian serous cystadenocarcinoma, pancreatic adenocarcinoma, pheochromocytoma and paraganglioma, prostate adenocarcinoma, rectal adenocarcinoma, skin melanoma, gastric adenocarcinoma, testicular germ cell tumor, thyroid carcinoma, thymoma, endometrial carcinoma, uterine carcinosarcoma, and uveal melanoma.

[0024] In some embodiments, the method of treatment further comprises administering a therapeutic agent that inhibits or excites PD1, PDL1, CD40, OX40, ICOS, CD28, CD137 / 4-1BB, CD27, GITR, CTLA4, PD-L2, B7-H3, B7-H4, HVEM, LIGHT, BTLA, CD30, TIGIT, VISTA, KIR, GAL9, TIM1, TIM3, TIM4, A2AR, LAG3, DR-5, CD2, CD5, CD39, or CD73. In some embodiments, the method of treatment further comprises administering to the individual at least one antibody that specifically binds to an inhibitory checkpoint molecule, and / or one or more standard or investigational anti-cancer therapies.

[0025] In some embodiments, the method of treatment further comprises administering at least one antibody that specifically binds to an inhibitory checkpoint molecule in combination with the anti-SIRPA antibody. In some embodiments, the at least one antibody that specifically binds to an inhibitory checkpoint molecule is selected from anti-PD-L1 antibodies, anti-CTLA4 antibodies, anti-PD-L2 antibodies, anti-PD-1 antibodies, anti-B7-H3 antibodies, anti-B7-H4 antibodies, and anti-HVEM antibodies, anti-B and T lymphocyte attenuator (BTLA) antibodies, anti-KIR antibodies, anti-GAL9 antibodies, anti-TIM-1 antibodies, anti-TIM3 antibodies, anti-TIM-4 antibodies, anti-A2AR antibodies, anti-CD 39 antibody, anti-CD73 antibody, anti-LAG-3 antibody, anti-phosphatidylserine antibody, anti-CD27 antibody, anti-CD30 antibody, anti-TNFα antibody, anti-CD33 antibody, anti-Siglec-5 antibody, anti-Siglec-7 antibody, anti-Siglec-9 antibody, anti-Siglec-11 antibody, antagonist anti-TREM1 antibody, antagonist anti-TREM2 antibody, anti-TIGIT antibody, anti-VISTA antibody, anti-CD2 antibody, anti-CD5 antibody, and any combination thereof.

[0026] In some embodiments, the treatment method further comprises administering one or more standard or investigational anti-cancer therapies selected from radiation therapy, cytotoxic chemotherapy, targeted therapy, imatinib therapy, trastuzumab therapy, etanercept therapy, adoptive cell transfer (ACT) therapy, chimeric antigen receptor T cell transfer (CAR-T) therapy, vaccine therapy, and cytokine therapy.

[0027] In some embodiments, the method of treatment further comprises administering to the individual at least one antibody that specifically binds to an inhibitory cytokine. In some embodiments, the at least one antibody that specifically binds to an inhibitory cytokine is selected from an anti-CCL2 antibody, an anti-CSF-1 antibody, an anti-IL-2 antibody, and any combination thereof.

[0028] In some embodiments, the method of treatment further comprises administering to the individual at least one agonist antibody that specifically binds to a stimulatory checkpoint protein. In some embodiments, the at least one agonist antibody that specifically binds to a stimulatory checkpoint protein is selected from agonist anti-CD40 antibodies, agonist anti-OX40 antibodies, agonist anti-ICOS antibodies, agonist anti-CD28 antibodies, agonist anti-TREM1 antibodies, agonist anti-TREM2 antibodies, agonist anti-CD137 / 4-1BB antibodies, agonist anti-CD27 antibodies, agonist anti-glucocorticoid-induced TNFR-related protein GITR antibodies, agonist anti-CD30 antibodies, agonist anti-BTLA antibodies, agonist anti-HVEM antibodies, agonist anti-CD2 antibodies, agonist anti-CD5 antibodies, and any combination thereof.

[0029] In some embodiments, the method of treatment further comprises administering at least one stimulatory cytokine to the individual. In some embodiments, the at least one stimulatory cytokine is selected from IFN-α4, IFN-β, IL-1β, TNF-α, IL-6, IL-8, CRP, IL-20 family members, LIF, IFN-γ, OSM, CNTF, GM-CSF, IL-11, IL-12, IL-15, IL-17, IL-18, IL-23, CXCL10, IL-33, MCP-1, MIP-1-β, and any combination thereof.

[0030] In some embodiments, the use of the anti-SIRPβ1 antibody provided herein for preparing a medicament is provided. In some embodiments, the medicament is used to treat cancer. In some embodiments, the anti-SIRPβ1 antibody provided herein is provided for use in treating cancer.

[0031] In some embodiments, a method of treating a neurodegenerative disease or condition is provided, comprising administering to an individual in need thereof a therapeutically effective amount of an anti-SIRPβ1 antibody provided herein. In some embodiments, the neurodegenerative disease or condition is selected from dementia, frontotemporal dementia, progressive supranuclear palsy, Alzheimer's disease, vascular dementia, Nasu-Hakola disease, cognitive deficits, memory loss, seizures, retinal dystrophy, amyotrophic lateral sclerosis, traumatic brain injury, spinal cord injury, stroke, Parkinson's disease, acute disseminated encephalomyelitis, retinal degeneration, age-related macular degeneration, glaucoma, multiple sclerosis.

[0032] In some embodiments, the use of the anti-SIRPβ1 antibodies provided herein for the preparation of a medicament for treating a neurodegenerative disease or disorder is provided. In some embodiments, the neurodegenerative disease or disorder is selected from dementia, frontotemporal dementia, progressive supranuclear palsy, Alzheimer's disease, vascular dementia, Nasu-Hakola disease, cognitive deficits, memory loss, epileptic seizures, retinal dystrophy, amyotrophic lateral sclerosis, traumatic brain injury, spinal cord injury, stroke, Parkinson's disease, acute disseminated encephalomyelitis, retinal degeneration, age-related macular degeneration, glaucoma, multiple sclerosis. In some embodiments, the anti-SIRPβ1 antibodies provided herein are provided for use in treating a neurodegenerative disease or disorder. In some embodiments, the neurodegenerative disease or disorder is selected from dementia, frontotemporal dementia, progressive supranuclear palsy, Alzheimer's disease, vascular dementia, Nasu-Hakola disease, cognitive deficits, memory loss, seizures, retinal dystrophy, amyotrophic lateral sclerosis, traumatic brain injury, spinal cord injury, stroke, Parkinson's disease, acute disseminated encephalomyelitis, retinal degeneration, age-related macular degeneration, glaucoma, multiple sclerosis.

[0033] It is to be understood that one, some, or all of the features of the various embodiments described herein may be combined to form other embodiments of the present invention. These and other aspects of the present invention will become apparent to those skilled in the art. The following detailed description further describes these and other embodiments of the present invention. BRIEF DESCRIPTION OF THE DRAWINGS

[0034] Figure 1A Shown is an alignment of the amino acid sequences of SIRPβ1 isoform 1 (SEQ ID NO: 1; Uniprot Accession No. 000241) and SIRPβ1 isoform 3 (SEQ ID NO: 384; Uniprot Accession No. Q5TFQ8). Figure 1B show Alignment of the amino acid sequences of SIRPβ1 isoform 1 (SEQ ID NO: 1) and SIRPα (SEQ ID NO: 385; Uniprot accession number P78324), which showed high homology within the extracellular domain.

[0035] Figure 2 Shown is an alignment of the amino acid sequences of human SIRPβ1 isoform 1 (SEQ ID NO: 1) and mouse SIRPβ1 (SEQ ID NO: 386; Uniprot Accession No. Q8BFX8).

[0036] Figure 3A-3BInduction of human SIRPβ1-dependent luciferase expression in a cell-based reporter assay is shown. Cells were stimulated with plate-bound anti-SIRPβ1 antibody (3A) or anti-SIRPα (SA-9C2) (3B) or human IgG1 isotype control. Results are expressed as fold relative to background. Background levels were set to 1 on the y-axis.

[0037] Figure 4A The expression patterns of two DAP12-related receptors (SIRPβ1 and TREM1) on human monocytes are shown. The shaded histograms represent background fluorescence from isotype-stained cells. The black outline histograms represent receptor expression levels of cells stained with target-specific antibodies. Figure 4B The expression of SIRPβ1 on M1 and M2 polarized macrophages from 2 healthy donors is shown. The shaded histogram represents the background fluorescence from isotype-stained cells. The black outline histogram represents the SIRPβ1 expression level on M2 macrophages, while the dashed histogram represents the SIRPβ1 expression level on M1 macrophages. Figure 4C Shown is the expression of SIRPβ1 and TREM1 on human monocyte-derived dendritic cells (DCs) with or without LPS stimulation. The shaded histograms represent background fluorescence from isotype-stained cells. The black outline histograms represent receptor expression levels of target-specific antibody-stained cells.

[0038] Figure 5A Delineation of the RNA expression pattern of SIRPβ1 isoform 1 in various tissues from human donors. Figure 5B Delineation of the RNA expression pattern of SIRPβ1 isoform 3 in various tissues from human donors. Peak expression of SIRPβ1 isoform 3 occurs in the brain, with limited expression in peripheral tissues.

[0039] Fig. 6A Shown is the SIRPβ1-mediated respiratory burst from primary human neutrophils obtained from two different donors. Figure 6B SIRPβ1-mediated respiratory burst (top panels) or IL-8 release (bottom panels) from primary human monocytes are shown. Figure 6C SIRPβ1-mediated TNFα cytokine release from primary human macrophages or dendritic cells (DCs) is shown.

[0040] Figure 7 Antitumor activity of primary human neutrophils stimulated with plate-bound anti-SIRPβ1 antibody is shown. Luminescence values ​​are presented on a relative scale, with the signal of Raji-Luc co-cultured with neutrophils in the absence of opsonizing antibodies set to 1 on the y-axis.

[0041] Figure 8Induction of human SIRPβ1-dependent luciferase expression by affinity matured antibodies in a cell-based reporter assay is shown. Results are presented as raw luminescence values. Background level was set to 10,000 on the y-axis.

[0042] Fig. 9 SIRPβ1-mediated TNFα cytokine release from primary human dendritic cells (DCs) is shown.

[0043] Figures 10A-10B The cross-reactivity of anti-SIRPβ1 antibodies to various SIRPβ1 antigens was shown. Soluble human SIRPβ1-Fc isoform 1 (O00241) or isoform 3 (Q5TFQ8) or cynomolgus monkey SIRPβ1-Fc (XM005568541) was coated onto plates and incubated with increasing concentrations of parental and affinity matured forms of anti-SIRPβ1 antibodies SB-1, SB-1-2, SB-2, SB-2-7, SB-8, SB-8-13, SB-40, and SB-40-21. All anti-SIRPβ1 antibodies only bound to human SIRPβ1-Fc isoform 1 antigen.

[0044] Fig.11A Upregulation of TREM2 expression on monocyte-derived human macrophages from three different healthy donors by plate-bound full-length anti-SIRPβ1 antibodies SB-1-3, SB-2-8, SB-8-13, SB-8-15, and SB-40-20. Baseline TREM2 expression was determined from cells incubated on human IgG1 isotype control antibody. TREM2 was detected with an anti-TREM2 antibody (clone ADI-22) conjugated to a DyLight650 fluorophore. Fig. 11B The viability of monocyte-derived human macrophages cultured on plate-bound full-length anti-SIRPβ1 antibodies SB-1-3, SB-2-8, SB-8-13, SB-8-15, and SB-40-20 is shown. Viable cells were quantified by measuring luminescence after addition of Cell Titer Glo substrate. Cells cultured on human IgG1 isotype control or in the absence of plate-bound antibodies (no antibody) established baseline viability of cells.

[0045] Fig.12 Shown is the viability of monocyte-derived macrophages from two different healthy donors cultured on increasing concentrations of plate-bound full-length anti-SIRPβ1 antibodies SB-1-3 and SB-2-8 or human IgG1 isotype control. Viable cells were quantified by measuring luminescence following addition of Cell Titer Glo substrate.

[0046] Fig.13Anti-SIRPβ1 antibodies were shown to enhance the agonistic activity of anti-TREM2 antibodies in a macrophage viability assay. Monocyte-derived macrophages were cultured on plate-bound full-length SB-1-3, SB-2-8, or huIgG1 isotype controls in the presence or absence of soluble anti-TREM2 antibodies. Viable cells were quantified by measuring luminescence values ​​after adding Cell Titer Glo substrate.

[0047] Fig.14A Shown is the viability of bone marrow-derived macrophages obtained from human SIRPβ1 BAC transgenic mice cultured on plate-bound full-length anti-SIRPβ1 antibodies SB-1-3, SB-2-8, and SB-8-13 or human IgG1 isotype control. Fig. 14B The viability of bone marrow-derived dendritic cells obtained from human SIRPβ1 BAC transgenic mice cultured on plate-bound full-length anti-SIRPβ1 antibodies SB-1-3, SB-2-8, and SB-8-13 or human IgG1 isotype control is shown. Viable cells were quantified by measuring luminescence values ​​after adding Cell TiterGlo substrate. Cells cultured on human IgG1 isotype control or in the absence of plate-bound antibodies (no antibody) established the baseline viability of the cells.

[0048] Figures 15A-15C The cross-reactivity of anti-SIRPβ1 antibody to different receptors of the SIRP family was shown. Fig.15A and Fig. 15B Titration curves of various anti-SIRP receptor antibodies binding to BW5147.G.1.4 cells overexpressing recombinant human SIRPβ1 or recombinant human SIRPα are shown respectively. Fig. 15C Titration curves showing antibody binding to Jurkat cells, an immortalized human T cell line that endogenously expresses SIRPγ. Positive control antibodies were anti-SIRPα / β clone 18D5, anti-SIRPα / β / γ clone KWAR23, and anti-SIRPγ clone LSB2.20. DETAILED DESCRIPTION OF THE INVENTION

[0049] The present disclosure relates to anti-SIRPβ1 antibodies (eg, monoclonal antibodies); 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.

[0050] The techniques and procedures described or referred to herein are generally well understood and commonly employed by those skilled in the art using conventional methods, such as, for example, widely utilized methods such as those described in Sambrook et al., Molecular Cloning: A Laboratory Manual 3d edition (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).

[0051] All references cited herein, including patent applications and publications, are hereby incorporated by reference in their entirety. I. Definitions

[0052] The terms "signal regulatory protein β1", "SIRPβ1", and "SIRPβ1 polypeptide" are used interchangeably herein and refer herein to any native SIRPβ1 from any vertebrate source, including mammals, such as primates (e.g., humans and cynomolgus monkeys) and rodents (e.g., mice and rats), unless otherwise indicated. In some embodiments, the term encompasses both wild-type sequences and naturally occurring variant sequences, such as splice variants or allelic variants. In some embodiments, the term encompasses "full-length", unprocessed SIRPβ1 as well as any form of SIRPβ1 derived from processing in cells. In some embodiments, SIRPβ1 is human SIRPβ1. In some embodiments, the amino acid sequence of an exemplary SIRPβ1 isoform 1 is Uniprot Accession No. O00241 of February 28, 2018. In some embodiments, the amino acid sequence of an exemplary human SIRPβ1 is SEQ ID NO: 1. In some embodiments, the amino acid sequence of an exemplary SIRPβ1 isoform 3 is Uniprot Accession No. Q5TFQ8 on January 31, 2018. In some embodiments, the amino acid sequence of an exemplary human SIRPβ1 isoform 3 is SEQ ID NO: 384. Unless otherwise specifically indicated, "SIRPβ1" as used herein refers to SIRPβ1 isoform 1.

[0053] The terms "anti-SIRPβ1 antibody", "antibody that binds to SIRPβ1", and "antibody that specifically binds to SIRPβ1" refer to antibodies that are capable of binding to SIRPβ1 with sufficient affinity such that the antibody is useful as a diagnostic and / or therapeutic agent in targeting SIRPβ1. In one embodiment, the extent of binding of an anti-SIRPβ1 antibody to an unrelated, non-SIRPβ1 polypeptide is less than about 10% of the binding of the antibody to SIRPβ1, as measured, for example, by radioimmunoassay (RIA). In certain embodiments, an antibody that binds to SIRPβ1 has a binding 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, e.g. 10 -8 M to 10 -13 M, for example 10 -9 M to 10 -13 In certain embodiments, the anti-SIRPβ1 antibody binds to a SIRPβ1 epitope that is conserved among SIRPβ1 polypeptides from different species.

[0054] With respect to the binding of an antibody to a target molecule, the term "specifically binds" or "is specific for" a particular polypeptide or an epitope on a particular polypeptide target means that binding can be measured differently from non-specific interactions. Specific binding can be measured, for example, by measuring the binding of a molecule compared to binding to a control molecule. For example, specific binding can be measured by competition with a control molecule similar to the target, such as 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" or "is specific for" a particular polypeptide or an epitope on a particular polypeptide target can be measured, for example, by measuring the binding of a molecule with about 10 -4 M or lower, 10 -5 M or lower, 10 -6 M or lower, 10 -7 M or lower, 10 -8 M or lower, 10 -9 M or lower, 10 -10 M or lower, 10 -11 M or lower, 10 -12 M or lower either KD or 10 -4 M to 10 -6 M or 10 -6 M to 10 -10 M or 10 -7 M to 10 -9A molecule exhibits a KD for a target in the range of 1.5 M. As will be appreciated by the skilled artisan, affinity and KD values ​​are inversely correlated. High affinity for an antigen is measured by a low KD value. In one embodiment, the term "specific binding" refers to a binding in which a molecule binds to a specific polypeptide or an epitope on a specific polypeptide without substantially binding to any other polypeptide or polypeptide epitope.

[0055] The term "immunoglobulin" (Ig) is used interchangeably with "antibody" herein. The term "antibody" is used herein in the broadest sense and specifically covers 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.

[0056] "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 linkages. Each heavy chain has a variable domain (V H ), followed by some constant domains. Each light chain has a variable domain at one end (V L ) and a constant domain at its other end; the constant domain of the light chain is aligned with the first constant domain of the heavy chain, while the light chain variable domain 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.

[0057] 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, p. 71 and Chapter 6.

[0058] The light chains from any vertebrate species can be assigned to one of two distinct types, called kappa ("κ") and lambda ("λ"), based on the amino acid sequence of their constant domains. Immunoglobulins can be assigned to different classes, or isotypes, based on the amino acid sequence of their heavy chain constant domains (CH). There are five classes of immunoglobulins: IgA, IgD, IgE, IgG, and IgM, with heavy chains called alpha ("α"), delta ("δ"), epsilon ("ε"), gamma ("γ"), and muon ("μ"), respectively. Based on minor differences in CH sequence and function, the gamma and alpha classes can be further divided into subclasses (isotypes), for example humans express the following subclasses: IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2. The subunit structures and three-dimensional conformations of the different classes of immunoglobulins are well known and generally described in, for example, Abbas et al., Cellular and Molecular Immunology, 4 th ed.(WB Saunders Co.,2000).

[0059] The "variable region" or "variable domain" of an antibody (such as the anti-SIRPβ1 antibody of the present disclosure) refers to the amino terminal domain of the antibody heavy chain or light chain. The heavy chain variable domain and the light chain variable domain may be referred to as "V H ” and “V L These domains are generally the most variable parts of the antibody (relative to other antibodies of the same class) and contain the antigen binding site.

[0060] The term "variable" refers to the fact that certain segments in the variable domain differ widely in sequence between antibodies (such as the anti-SIRPβ1 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 across the entire span of the variable domain. Instead, it is concentrated in three segments called hypervariable regions (HVRs) in both the light chain and heavy chain variable domains. The more highly conserved parts of the variable domain are called framework regions (FRs). The variable domains of native heavy and light chains each contain four FR regions, most of which adopt a β-pleated sheet conformation, connected by three HVRs that form loops and in some cases form part of the β-pleated 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 antigen binding site of the antibody (see Kabat et al., Sequences of Immunological Interest, 5th Edition, National Institute of Health, Bethesda, MD. (1991)). The constant domains are not directly involved in the binding of the antibody to the antigen, but exhibit various effector functions, such as participation of the antibody in antibody-dependent cellular cytotoxicity.

[0061] The term "monoclonal antibody" as used herein refers to an antibody (such as the monoclonal anti-SIRPβ1 antibody of the present disclosure) obtained from a population of substantially homogeneous antibodies, i.e., the individual antibodies constituting 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 very small amounts. Monoclonal antibodies are highly specific, being directed against a single antigenic site. Unlike typical polyclonal antibody preparations that contain different antibodies directed against different determinants (epitopes), each monoclonal antibody is directed against a single determinant on the antigen. In addition to their specificity, the 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 that the antibody is obtained from a substantially homogeneous population of antibodies and should not be interpreted as requiring the antibody to be produced by any particular method. For example, the monoclonal antibodies to be used in accordance with the present invention can be produced by a variety of techniques, including, for example, hybridoma methods, recombinant DNA methods, and techniques for producing human or human-like antibodies in animals having part or all of the human immunoglobulin loci or genes encoding human immunoglobulin sequences.

[0062] The terms "full-length antibody", "intact antibody" and "whole antibody" are used interchangeably and refer to antibodies (such as the anti-SIRPβ1 antibodies of the present disclosure) in a substantially complete form, as opposed to antibody fragments. Specifically, intact antibodies include those having heavy and light chains (including an Fc region). The constant domain may be a native sequence constant domain (e.g., a human native sequence constant domain) or an amino acid sequence variant thereof. In some cases, an intact antibody may have one or more effector functions.

[0063] "Antibody fragment" refers to a molecule other than an intact antibody that comprises 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. Pat. No. 5,641,870, Example 2; Zapata et al., Protein Eng. 8(10):1057-1062 (1995)); single-chain antibody molecules; and multispecific antibodies formed from antibody fragments.

[0064] Digestion of antibodies (such as the anti-SIRPβ1 antibodies of the present disclosure) with papain produces two identical antigen-binding fragments, called "Fab" fragments, and a residual "Fc" fragment, the name of which reflects its ability to crystallize readily. The Fab fragment consists of an intact light chain and the variable region of the heavy chain (V H ) and the first constant domain of the heavy chain (C H1) Composition. Each Fab fragment is monovalent for antigen binding, that is, it has one antigen binding site. Pepsin treatment of antibodies produces a larger F(ab')2 fragment, which is roughly equivalent to two Fab fragments linked by disulfide bonds, with different antigen binding activities and still able to cross-link antigens. H The Fab fragment differs from the Fab fragment by the addition of a few residues at the carboxyl terminus of the 1 domain, including one or more cysteines from the antibody hinge region. Fab'-SH is the designation herein for Fab' in which the cysteine ​​residues of the constant domains bear free thiol groups. F(ab')2 antibody fragments were originally generated as pairs of Fab' fragments with hinge cysteines between them. Other chemical couplings of antibody fragments are also known.

[0065] The Fc fragment contains the carboxyl-terminal portions of two heavy chains held together by disulfide bonds. The effector functions of an antibody are determined by sequences in the Fc region, which is also recognized by Fc receptors (FcRs) found on certain types of cells.

[0066] The "functional fragment" of the antibody of the present invention (such as the anti-SIRPβ1 antibody of the present disclosure) comprises a portion of the intact antibody, generally including the antigen binding or variable region of the intact antibody or the Fc region of the antibody that retains or has improved FcR binding ability. Examples of antibody fragments include linear antibodies, single-chain antibody molecules, and multispecific antibodies formed from antibody fragments.

[0067] The term "diabody" refers to a H and V L Small antibody fragments prepared by constructing sFv fragments (see previous paragraph) using short linkers (about 5-10 residues) between domains. Due to the short linker, the variable domains achieve inter-chain rather than intra-chain pairing, resulting in a bivalent fragment, that is, a fragment with two antigen-binding sites. Bispecific diabodies are heterodimers of two "crossed" sFv fragments, in which the V H and V L Domains are present on different polypeptide chains.

[0068] As used herein, "chimeric antibodies" refer to antibodies (immunoglobulins), such as the chimeric anti-SIRPβ1 antibodies of the present disclosure, in which a portion of the heavy chain and / or light chain is identical or homologous to the corresponding sequence in an antibody derived from a particular species or belonging to a particular antibody class or subclass, and the remainder of the chain is identical or homologous to the corresponding sequence in an antibody derived from another species or belonging to another antibody class or subclass, as well as fragments of such antibodies, as long as they exhibit the desired biological activity. Chimeric antibodies of interest herein include "primatized" antibodies, in which the antigen-binding region of the antibody is derived from an antibody generated by immunizing a macaque monkey with, for example, an antigen of interest. As used herein, "humanized antibodies" are a subset of "chimeric antibodies".

[0069] A "humanized" form of a non-human (e.g., mouse) antibody, such as a humanized form of an anti-SIRPβ1 antibody of the present disclosure, is a chimeric antibody comprising amino acid residues from a non-human HVR and amino acid residues from a human FR. In certain embodiments, a humanized antibody will comprise substantially the entirety of at least one and typically two variable domains, wherein all or substantially all HVRs (e.g., CDRs) correspond to those of a non-human antibody, and all or substantially all FRs correspond to those of a human antibody. A humanized antibody may optionally comprise at least a portion of an antibody constant region derived from a human antibody. An antibody, such as a "humanized form" of a non-human antibody, refers to an antibody that has undergone humanization.

[0070] "Human antibody" refers to an antibody that has an amino acid sequence corresponding to the amino acid sequence of an antibody produced by a human and / or is produced using any of the techniques disclosed herein for producing human antibodies, such as the anti-SIRPβ1 antibodies of the present disclosure. This definition of human antibodies explicitly excludes humanized antibodies comprising 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-based platform technologies. Human antibodies can be prepared by administering an antigen to a transgenic animal, such as an immunized xenogeneic mouse, that has been modified to produce human antibodies in response to antigenic stimulation but whose endogenous loci have been incapable of producing human antibodies, or by human B-cell hybridoma technology.

[0071] The term "hypervariable region" or "HVR" as used herein refers to regions of an antibody variable domain (such as the variable domain of the anti-SIRPβ1 antibody of the present disclosure) that are highly variable in sequence and / or form structurally defined loops. Typically, antibodies contain six HVRs: three in VH (H1, H2, H3) and three in VL (L1, L2, L3). In natural antibodies, H3 and L3 display the greatest diversity of the six HVRs, and H3 in particular is believed to play a unique role in conferring exquisite specificity to antibodies. Naturally occurring camelid antibodies composed only of heavy chains are functional and stable in the absence of light chains.

[0072] Many HVR descriptions are used and covered herein. In some embodiments, the HVR may be a Kabat complementarity determining region (CDR), which is based on sequence variability and is the most commonly used (Kabat et al., supra). In some embodiments, the HVR may be a Chothia CDR. Chothia refers instead to the position of the structural loop (Chothia and Lesk, J. Mol. Biol. 196: 901-917 (1987)). In some embodiments, the HVR may be an AbM HVR. The AbM HVR represents a compromise between the Kabat CDR and the Chothia structural loop, and is used by Oxford Molecular's AbM antibody modeling software. In some embodiments, the HVR may be a "contact" HVR. The "contact" HVR is based on analysis of available complex crystal structures. The residues of each of these HVRs are recorded below.

[0073] HVRs may include "extended HVRs" 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 (a preferred embodiment) (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.

[0074] "Framework" or "FR" residues are those variable domain residues other than the HVR residues as herein defined.

[0075] As used herein, an "acceptor human framework" is a human immunoglobulin framework or a human consensus framework comprising a V L or V H The amino acid sequence of the framework. An acceptor human framework or human consensus framework "derived" from a human immunoglobulin 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. In cases where there are pre-existing amino acid changes in VH, preferably those changes are present at 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 VH. LThe human immunoglobulin framework sequences or human consensus framework sequences are identical.

[0076] "Human common framework" refers to human immunoglobulin V L or V H The framework sequence contains the most common amino acid residues in the framework sequence selection. L or V H The sequence selection is from a subgroup of variable domain sequences. Typically, the sequence subgroup is a subgroup such as that in Kabat et al., Sequences of Proteins of Immunological Interest, 5th ed., Public Health Service, National Institutes of Health, Bethesda, MD (1991). Examples include for V L , the subgroup may be subgroup κI, κII, κIII or κIV, as in Kabat et al., supra. In addition, for V H , the subgroup can be subgroup I, subgroup II, or subgroup III, as in Kabat et al., supra.

[0077] "Amino acid modification" at a specified position (e.g., of an anti-SIRPβ1 antibody of the present disclosure) refers to substitution or deletion of the specified residue, or insertion of at least one amino acid residue near the specified residue. Insertion "near" the specified residue means insertion within one to two residues thereof. The insertion may be at the N-terminus or C-terminus of the specified residue. The preferred amino acid modification herein is substitution.

[0078] An "affinity matured" antibody (such as the affinity matured anti-SIRPβ1 antibody disclosed herein) refers to an antibody having one or more changes 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 without these changes. In one embodiment, the affinity matured antibody has a nanomolar or even picomolar affinity for the target antigen. Affinity matured antibodies can be generated by procedures known in the art. For example, Marks et al., Bio / Technology 10:779-783 (1992) describe the use of V H and V LAffinity maturation by domain shuffling. Random mutagenesis of HVR and / or framework residues is described in, for example, 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); Hawkins et al., J. Mol. Biol. 226:889-896 (1992).

[0079] "Fv" is the smallest antibody fragment that contains a complete antigen recognition and binding site. This fragment consists of a dimer of one heavy-chain variable region and one light-chain variable region in tight, non-covalent association. From the folding of these two domains arise six hypervariable loops (3 loops each from the heavy and light chains) that contribute the amino acid residues for antigen binding and confer antigen-binding specificity to the antibody. However, even a single variable domain (or half of an Fv containing only three HVRs specific for an antigen) has the ability to recognize and bind antigen, although at a lower affinity than the entire binding site.

[0080] "Single-chain Fv", also abbreviated as "sFv" or "scFv", is a protein that contains V peptides linked into one polypeptide chain. H and V L Preferably, the sFv polypeptide is in the V H With V L The domains further include polypeptide linkers that enable the sFv to form the desired structure for binding to the antigen.

[0081] 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.

[0082] The term "Fc region" is used herein to define the C-terminal region of an immunoglobulin heavy chain, including a native sequence Fc region and a variant Fc region. Although the boundaries of an immunoglobulin heavy chain Fc region may vary, a human IgG heavy chain Fc region is generally defined as a segment from an amino acid residue at position Cys226 or Pro230 thereof to the carboxyl terminus. The C-terminal lysine (residue 447, according to the EU numbering system) of the Fc region may be eliminated, for example, during the production or purification of the antibody, or by recombinant engineering of a nucleic acid encoding the heavy chain of the antibody. Thus, a composition of a complete antibody may include an antibody population in which all K447 residues are eliminated, an antibody population in which none of the K447 residues are eliminated, and an antibody population in which antibodies with and without the K447 residue are mixed. Suitable native sequence Fc regions for use in the antibodies of the present disclosure include human IgG1, IgG2, IgG3, and IgG4.

[0083] 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.

[0084] A "variant Fc region" comprises an amino acid sequence that differs from the amino acid sequence of a native sequence Fc region by 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 compared to the native sequence Fc region or the Fc region of a parent polypeptide, such as from about one to about ten amino acid substitutions in the native sequence Fc region or the Fc region of a parent polypeptide, and preferably from about one to about five amino acid substitutions. The variant Fc region herein will preferably possess at least about 80% homology to the native sequence Fc region and / or the Fc region of a parent polypeptide, and most preferably at least about 90% homology thereto, and more preferably at least about 95% homology thereto.

[0085] "Fc receptor" or "FcR" describes a receptor that binds to the Fc region of an antibody. Preferred FcRs are native sequence human FcRs. In addition, preferred FcRs are FcRs (gamma receptors) that bind IgG antibodies, including receptors of the FcγRI, FcγRII, and FcγRIII subclasses, including allelic variants and alternatively spliced ​​forms of these receptors. FcγRII receptors include FcγRIIA ("activating receptors") and FcγRIIB ("inhibiting receptors"), which have similar amino acid sequences and differ primarily in their cytoplasmic domains. The activating receptor FcγRIIA contains an immunoreceptor tyrosine-based activation motif (ITAM) in its cytoplasmic domain. The inhibitory receptor FcγRIIB contains an immunoreceptor tyrosine-based inhibition motif (ITIM) in its cytoplasmic domain. The term "FcR" herein encompasses other FcRs, including those that will be identified in the future. FcRs can also extend the serum half-life of antibodies.

[0086] As used herein, "percent (%) amino acid sequence identity" and "homology" with respect to peptide, polypeptide or antibody sequences refer to the percentage of amino acid residues in a candidate sequence that are identical to 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 without considering any conservative substitutions as part of the sequence identity. Alignment for the purpose of determining percent amino acid sequence identity can be achieved in a variety of ways within the skill in the art, for example using publicly available computer software such as BLAST, BLAST-2, ALIGN or MEGALIGN™ (DNASTAR) software. One skilled in the art can determine appropriate parameters for measuring alignment, including any algorithm known in the art required to achieve maximum alignment over the full length of the sequences being compared.

[0087] When used in the context of antibodies that compete for the same epitope (e.g., neutralizing antibodies), the term "competition" means 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 a reference antibody) to a common antigen (e.g., SIRPβ1 or a fragment thereof). Numerous types of competitive binding assays can be used to determine whether antibodies compete with each other, for example: solid phase direct or indirect radioimmunoassay (RIA), solid phase direct or indirect enzyme immunoassay (EIA), sandwich competition assay (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 assay, solid phase direct label sandwich assay (see, e.g., Harlow and Lane, 1988, Antibodies, A Laboratory Manual, Cold Spring Harbor Laboratory Press, 1996 ... Harbor Press); solid phase direct labeling RIA using I-125 label (see, e.g., Morel et al., 1988, Molec. Immunol. 25: 7-15); solid phase direct biotin-avidin EIA (see, e.g., Cheung et al., 1990, Virology 176: 546-552); and direct labeling RIA (Moldenhauer et al., 1990, Scand. J. Immunol. 32: 77-82). Typically, such assays involve the use of purified antigen bound to a solid surface or cells carrying either of these, unlabeled test antibodies, and labeled reference antibodies. Competitive inhibition is measured by determining the amount of label bound to the 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 sufficiently close to the epitope bound by the reference antibody to cause steric hindrance. Additional details about 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 approach 100%.

[0088] As used herein, the "interaction" between a SIRPβ1 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 the interaction between two polypeptides" when the antibody destroys, reduces, or completely eliminates the interaction between the two polypeptides. An antibody of the present disclosure "inhibits the interaction between the two polypeptides" when the antibody binds to one of the two polypeptides. In some embodiments, the interaction can be inhibited by at least about 20%, 30%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 95%, 97.5%, and / or close to 100%.

[0089] The term "epitope" includes any determinant capable of being bound by an antibody. An epitope is a region of an antigen that is bound by an antibody targeting that antigen, and when the antigen is a polypeptide, includes specific amino acids that directly contact the antibody. Most commonly, epitopes are located on polypeptides, but in some cases can be located on other types of molecules, such as nucleic acids. Epitope determinants can include chemically active surface groupings of molecules such as amino acids, sugar side chains, phosphoryl or sulfonyl groups, and can have specific three-dimensional structural characteristics, and / or specific charge characteristics. In general, an antibody specific for a particular target antigen will preferentially recognize an epitope on the target antigen in a complex mixture of polypeptides and / or macromolecules.

[0090] An "agonistic" antibody or "activating" antibody is an antibody that induces (eg, increases) one or more activities or functions of the antigen after the antibody binds to the antigen.

[0091] An "antagonist" antibody or "blocking" antibody or "inhibitory" antibody is an antibody that reduces, inhibits, and / or eliminates (e.g., reduces) the binding of an antigen to one or more ligands after the antibody binds to the antigen, 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, an antagonist, blocking, or inhibitory antibody substantially or completely inhibits the binding of an antigen to one or more ligands and / or one or more activities or functions of the antigen.

[0092] An "isolated" antibody, such as an isolated anti-SIRPβ1 antibody of the present disclosure, is an antibody that has been identified, separated and / or recovered from a component of its production environment (e.g., natural or recombinant). Preferably, an isolated antibody is free from all other contaminant components from its production environment. Contaminant components from its production environment, such as those derived 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% by weight of the antibody, as determined by, for example, the Lowry method, and in some embodiments, greater than 99% by weight; (2) a degree sufficient to obtain at least 15 residues of N-terminal or internal amino acid sequence by use of a spinning cup sequenator, or (3) homogeneity, as determined by SDS-PAGE under non-reducing or reducing conditions using Coomassie blue or, preferably, silver stain. An isolated antibody includes the antibody in situ within a recombinant T cell, since at least one component of the antibody's natural environment will not be present. However, isolated polypeptide or antibody will generally be prepared by at least one purification step.

[0093] An "isolated" nucleic acid molecule encoding an antibody, such as an anti-SIRPβ1 antibody of the present disclosure, is a nucleic acid molecule that is identified and separated from at least one contaminant nucleic acid molecule that is normally associated with it in the environment in which it is produced. Preferably, the isolated nucleic acid is free of all components associated with the environment in which it is produced. An isolated nucleic acid molecule encoding the polypeptides and antibodies herein is in a form or setting that is different from the form or setting in which it is found in nature. Therefore, an isolated nucleic acid molecule is distinguished from the nucleic acid encoding the polypeptides and antibodies herein that is naturally present in a cell.

[0094] As used herein, the term "vector" is intended to refer to a nucleic acid molecule capable of transporting another nucleic acid connected thereto. 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 bacteriophage vector. Another type of vector is a viral vector, in which additional DNA segments can be connected to the viral genome. Certain vectors are capable of autonomous replication in host cells into which they are introduced (e.g., bacterial vectors and episomal mammalian vectors having bacterial replication origins). Other vectors (e.g., non-episomal mammalian vectors) can be integrated into the genome of the host cell after introduction into the host cell, and thus replicated together with the host genome. In addition, certain vectors are capable of directing the expression of genes operably connected thereto. Such vectors are referred to herein as "recombinant expression vectors" or simply "expression vectors". In general, expression vectors utilized in recombinant DNA technology are often in the form of plasmids. In this specification, "plasmid" and "vector" can be used interchangeably because plasmids are the most commonly used form of vectors.

[0095] As used interchangeably herein, "polynucleotide" or "nucleic acid" refers to polymers 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.

[0096] "Host cell" includes an individual cell or cell culture that can be or has been a recipient of a vector for incorporating a polynucleotide insert. Host cells include progeny of a single host cell, and progeny may not necessarily be completely identical (in morphology or in genomic DNA complement) to the original parent cell due to natural, accidental, or deliberate mutation. Host cells include cells transfected in vivo with a polynucleotide of the invention.

[0097] "Carrier" as used herein includes pharmaceutically acceptable carriers, excipients or stabilizers that are non-toxic to cells or mammals exposed thereto at the dosages and concentrations employed.

[0098] As used herein, the term "preventing" includes providing prevention with respect to the occurrence or recurrence of a particular disease, disorder, or condition in an individual. An individual may be susceptible or predisposed to a particular disease, disorder, or condition, or be at risk of developing such a disease, disorder, or condition, but has not yet been diagnosed with the disease, disorder, or condition.

[0099] As used herein, an individual who is "at risk" for developing a particular disease, disorder, or condition may or may not have detectable disease or disease symptoms, and may or may not exhibit detectable disease or disease symptoms prior to the treatment methods described herein. "At risk" means that the individual has one or more risk factors, which are measurable parameters associated with the occurrence of a particular disease, disorder, or condition, as known in the art. An individual with one or more of these risk factors has a higher probability of developing a particular disease, disorder, or condition than an individual without one or more of these risk factors.

[0100] As used herein, "treatment" refers to a clinical intervention designed to alter the natural course of the individual being treated during the course of clinical pathology. Desired effects of treatment include reduced progression rate, amelioration or alleviation of the pathological state, and regression or improved prognosis for a particular disease, disorder, or condition. For example, an individual is successfully "treated" if one or more symptoms associated with a particular disease, disorder, or condition are alleviated or eliminated.

[0101] "Effective amount" refers to an amount that effectively achieves the desired therapeutic or preventive result at least in the necessary dosage and time period. An effective amount can be provided in one or more administrations. The effective amount herein can vary with factors such as the disease state, age, sex, and weight of the individual, and 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 exceed any toxic or adverse effects of the treatment. For preventive use, beneficial or desired results include results such as eliminating or reducing risk, reducing severity, or delaying the onset of the disease, including biochemical, histological and / or behavioral symptoms of the disease, its complications, and intermediate pathological phenotypes presented during disease formation. For therapeutic use, beneficial or desired results include clinical results, such as reducing one or more symptoms from the disease, improving the quality of life of those subjects suffering from the disease, reducing the dose of other drugs required to treat the disease, enhancing the effect of another drug (such as via targeting), delaying the progression of the disease, and / or prolonging survival. The effective amount of a drug, compound, or pharmaceutical composition is an amount sufficient to directly or indirectly achieve preventive or therapeutic treatment. As understood in a clinical context, an effective amount of a drug, compound, or pharmaceutical composition can be achieved with or without another drug, compound, or pharmaceutical composition. Thus, an "effective amount" can be considered in the context of administering one or more therapeutic agents, and a single agent can be considered to be administered in an effective amount if, together with one or more other agents, the desired result can be achieved or realized.

[0102] "Subject" for purposes of treatment, prevention, or risk reduction refers to any animal classified as a mammal, including humans, domesticated and farm animals, and zoo, sport, or pet animals, such as dogs, horses, rabbits, cattle, pigs, Hamster, gerbil, mouse, ferret, rat, cat, etc. In some embodiments, the subject is a human.

[0103] 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 administration as separate compositions, including 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.

[0104] As used herein, the term "about" refers to the usual error range of the corresponding numerical value that is readily known to those skilled in the art. References herein to "about" numerical values ​​or parameters include (and describe) embodiments involving the numerical value or parameter itself.

[0105] As used herein and in the appended claims, the singular forms "a", "an", and "the" include plural references unless the context clearly dictates otherwise. For example, reference to an "antibody" is a reference to one to a plurality of antibodies, such as molar amounts, and includes equivalents thereof known to those skilled in the art, and so forth.

[0106] It is understood that various aspects and embodiments of the present disclosure described herein include "comprising," "consisting of," and "consisting essentially of" aspects and embodiments. I. Anti-SIRPβ1 Antibody

[0107] Provided herein are anti-SIRPβ1 antibodies. The provided antibodies are useful, for example, for the diagnosis or treatment of SIRPβ1-mediated disorders.

[0108] In some embodiments, the anti-SIRPβ1 antibodies of the present disclosure are agonists of SIRPβ1 activity. In some embodiments, anti-SIRPβ1 antibodies that bind to human SIRPβ1 isoform 1 but not to human SIRPα are provided. In some embodiments, anti-SIRPβ1 antibodies that bind to human SIRPβ1 isoform 1 but not to human SIRPγ are provided. In some embodiments, anti-SIRPβ1 antibodies that bind to human SIRPβ1 isoform 1 but not to human SIRPβ1 isoform 3 are provided. In some embodiments, anti-SIRPβ1 antibodies that bind to human SIRPβ1 isoform 1 but not to mouse SIRPβ1 and / or cynomolgus monkey SIRPβ1 are provided. In some embodiments, anti-SIRPβ1 antibodies of the present disclosure agonize SIRPβ1 activity on CD14-positive monocytes. In some embodiments, anti-SIRPβ1 antibodies of the present disclosure induce respiratory bursts in immune cells, such as neutrophils, monocytes, and / or macrophages. In some embodiments, anti-SIRPβ1 antibodies of the present disclosure induce or increase IL-8 expression in monocytes. In some embodiments, the anti-SIRPβ1 antibodies of the present disclosure induce or increase TNFα expression in macrophages and / or dendritic cells. In some embodiments, the anti-SIRPβ1 antibodies of the present disclosure induce neutrophil-mediated phagocytosis, such as phagocytosis of tumor cells. In some embodiments, the anti-SIRPβ1 antibodies of the present disclosure increase neutrophil-mediated tumor cell clearance. In some embodiments, the anti-SIRPβ1 antibodies of the present disclosure increase or enhance the anti-tumor properties or activity of neutrophils.

[0109] In some embodiments, the anti-SIRPβ1 antibodies of the present disclosure recruit immune cells. In some embodiments, the anti-SIRPβ1 antibodies of the present disclosure induce syk phosphorylation. In some embodiments, the anti-SIRPβ1 antibodies of the present disclosure induce syk phosphorylation when clustered by adjacent cells expressing Fcγ receptors.

[0110] In some embodiments, the SIRPβ1 antibodies of the present disclosure downregulate SIRPβ1 expression on the surface of a cell. In some embodiments, the SIRPβ1 antibodies of the present disclosure do not downregulate SIRPβ1 expression on the surface of a cell. In some embodiments, the SIRPβ1 antibodies of the present disclosure block ligand binding to SIRPβ1. In some embodiments, the SIRPβ1 antibodies of the present disclosure do not block ligand binding to SIRPβ1.

[0111] In some embodiments, an anti-SIRPβ1 antibody having one or more properties selected from the following is provided: a) binds to human SIRPβ1 isoform 1, but does not bind to human SIRPα; b) binds to human SIRPβ1 isoform 1, but not to human SIRPγ; c) binds to human SIRPβ1 isoform 1, but not to human SIRPβ1 isoform 3; d) binds to human SIRPβ1 isoform 1, but not to mouse SIRPβ1; e) binds to human SIRPβ1 isoform 1, but not to cynomolgus monkey SIRPβ1; f) stimulating SIRPβ1 activity on CD14-positive monocytes in vitro and / or in vivo; g) inducing or enhancing the respiratory burst in immune cells, such as neutrophils and / or monocytes, in vitro and / or in vivo; h) inducing or increasing IL-8 expression in monocytes in vitro and / or in vivo; i) inducing or increasing TNFα expression in macrophages and / or dendritic cells in vitro and / or in vivo; j) inducing or enhancing neutrophil-mediated phagocytosis, such as phagocytosis of tumor cells, in vitro and / or in vivo; k) inducing or enhancing neutrophil-mediated tumor cell clearance in vivo; 1) upregulating TREM2 expression on macrophages in vitro and / or in vivo; m) increasing the viability of macrophages in vitro and / or in vivo when alone and / or in combination with an agonistic anti-TREM2 antibody; and n) Improving the survival of dendritic cells in vitro and / or in vivo.

[0112] In some embodiments, an anti-SIRPβ1 antibody having the following properties is provided: a) binds to human SIRPβ1 isoform 1, but not to human SIRPα; b) binds to human SIRPβ1 isoform 1, but not to human SIRPγ; c) binds to human SIRPβ1 isoform 1, but not to human SIRPβ1 isoform 3; d) binds to human SIRPβ1 isoform 1, but not to mouse SIRPβ1; e) binds to human SIRPβ1 isoform 1, but not to cynomolgus monkey SIRPβ1; f) stimulating SIRPβ1 activity on CD14-positive monocytes in vitro and / or in vivo; g) inducing or enhancing the respiratory burst in immune cells, such as neutrophils and / or monocytes, in vitro and / or in vivo; h) inducing or increasing IL-8 expression in monocytes in vitro and / or in vivo; i) inducing or increasing TNFα expression in macrophages and / or dendritic cells in vitro and / or in vivo; j) inducing or enhancing neutrophil-mediated phagocytosis, such as phagocytosis of tumor cells, in vitro and / or in vivo; k) inducing or enhancing neutrophil-mediated tumor cell clearance in vivo; 1) upregulating TREM2 expression on macrophages in vitro and / or in vivo; m) increasing the viability of macrophages in vitro and / or in vivo when alone and / or in combination with an agonistic anti-TREM2 antibody; and n) Improving the survival of dendritic cells in vitro and / or in vivo. A. Exemplary Antibodies and Certain Other Antibody Embodiments

[0113] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising at least one, two, three, four, five, or six HVRs selected from the following: (a) HVR-H1 comprising the HVR-H1 amino acid sequence shown in Table 4 and / or Table 9; (b) HVR-H2 comprising the HVR-H2 amino acid sequence shown in Table 4 and / or Table 9; (c) HVR-H3 comprising the HVR-H3 amino acid sequence shown in Table 4 and / or Table 9; (d) HVR-L1 comprising the HVR-L1 amino acid sequence shown in Table 3 and / or Table 8; (e) HVR-L2 comprising the HVR-L2 amino acid sequence shown in Table 3 and / or Table 8; and (f) HVR-L3 comprising the HVR-L3 amino acid sequence shown in Table 3 and / or Table 8.

[0114] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising HVR-H1, HVR-H2, HVR-H3, HVR-L1, HVR-L2, and HVR-L3, wherein: (a) HVR-H1 comprises the HVR-H1 amino acid sequence shown in Table 4 and / or Table 9; (b) HVR-H2 comprises the HVR-H2 amino acid sequence shown in Table 4 and / or Table 9; (c) HVR-H3 comprises the HVR-H3 amino acid sequence shown in Table 4 and / or Table 9; (d) HVR-L1 comprises the HVR-L1 amino acid sequence shown in Table 3 and / or Table 8; (e) HVR-L2 comprises the HVR-L2 amino acid sequence shown in Table 3 and / or Table 8; and (f) HVR-L3 comprises the HVR-L3 amino acid sequence shown in Table 3 and / or Table 8.

[0115] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising an SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-10, SB-11, SB-12, SB-13, SB-14, SB-15, SB-16, SB-17, SB-18, SB-19, SB-20, SB-21, SB-22, SB-23, SB-24, SB-25, SB-26, SB-27, SB-28, SB-29, SB-30, SB-31, SB-32, SB-33, SB-34, SB-35, SB-36, SB-37, SB-38, SB-39, SB-40, SB-41, SB-42, SB-43, SB-44, SB-45, SB-46, SB-47, SB-48, SB-49, SB-50, SB-51, SB-52, SB-53, SB-54 ,SB-35,SB-36,SB-37,SB-38,SB-39,SB-40,SB-41,SB-42,SB-43,SB-44,SB-45,SB-46,SB-47,SB-48,SB-49,SB-50,SB-1-2,SB-1-3,SB-1-4,SB Six HVRs for antibodies to -1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21.

[0116] In some embodiments, provided herein are anti-SIRPβ1 antibodies comprising six HVRs of an antibody selected from SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-14, SB-28, SB-39, SB-40, SB-49, SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21 (shown in Tables 3, 4, 8, and 9).

[0117] In some embodiments, provided herein are anti-SIRPβ1 antibodies comprising six HVRs of an antibody selected from SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21 (shown in Tables 8 and 9).

[0118] In some embodiments, provided herein are anti-SIRPβ1 antibodies comprising six HVRs of an antibody selected from SB-1-3, SB-2-8, and SB-8-13 (shown in Tables 8 and 9).

[0119] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising (a) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 228; (b) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 238; (c) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 125; (d) an HVR-L1 comprising the amino acid sequence of SEQ ID NO: 383; (e) an HVR-L2 comprising the amino acid sequence of SEQ ID NO: 16; and (f) an HVR-L3 comprising the amino acid sequence of SEQ ID NO: 31.

[0120] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising (a) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 229; (b) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 239; (c) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 125; (d) an HVR-L1 comprising the amino acid sequence of SEQ ID NO: 383; (e) an HVR-L2 comprising the amino acid sequence of SEQ ID NO: 16; and (f) an HVR-L3 comprising the amino acid sequence of SEQ ID NO: 31.

[0121] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising (a) an HVR-H1 comprising the amino acid sequence of SEQ ID NO:229; (b) an HVR-H2 comprising the amino acid sequence of SEQ ID NO:240; (c) an HVR-H3 comprising the amino acid sequence of SEQ ID NO:125; (d) an HVR-L1 comprising the amino acid sequence of SEQ ID NO:383; (e) an HVR-L2 comprising the amino acid sequence of SEQ ID NO:16; and (f) an HVR-L3 comprising the amino acid sequence of SEQ ID NO:31.

[0122] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising (a) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 229; (b) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 241; (c) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 125; (d) an HVR-L1 comprising the amino acid sequence of SEQ ID NO: 383; (e) an HVR-L2 comprising the amino acid sequence of SEQ ID NO: 16; and (f) an HVR-L3 comprising the amino acid sequence of SEQ ID NO: 31.

[0123] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising (a) an HVR-H1 comprising an amino acid sequence of SEQ ID NO: 230; (b) an HVR-H2 comprising an amino acid sequence of SEQ ID NO: 242; (c) an HVR-H3 comprising an amino acid sequence of SEQ ID NO: 126; (d) an HVR-L1 comprising an amino acid sequence of SEQ ID NO: 383; (e) an HVR-L2 comprising an amino acid sequence of SEQ ID NO: 16; and (f) an HVR-L3 comprising an amino acid sequence of SEQ ID NO: 32.

[0124] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising (a) an HVR-H1 comprising the amino acid sequence of SEQ ID NO:231; (b) an HVR-H2 comprising the amino acid sequence of SEQ ID NO:243; (c) an HVR-H3 comprising the amino acid sequence of SEQ ID NO:126; (d) an HVR-L1 comprising the amino acid sequence of SEQ ID NO:383; (e) an HVR-L2 comprising the amino acid sequence of SEQ ID NO:16; and (f) an HVR-L3 comprising the amino acid sequence of SEQ ID NO:32.

[0125] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising (a) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 232; (b) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 244; (c) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 126; (d) an HVR-L1 comprising the amino acid sequence of SEQ ID NO: 383; (e) an HVR-L2 comprising the amino acid sequence of SEQ ID NO: 16; and (f) an HVR-L3 comprising the amino acid sequence of SEQ ID NO: 32.

[0126] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising (a) an HVR-H1 comprising the amino acid sequence of SEQ ID NO:99; (b) an HVR-H2 comprising the amino acid sequence of SEQ ID NO:245; (c) an HVR-H3 comprising the amino acid sequence of SEQ ID NO:126; (d) an HVR-L1 comprising the amino acid sequence of SEQ ID NO:383; (e) an HVR-L2 comprising the amino acid sequence of SEQ ID NO:16; and (f) an HVR-L3 comprising the amino acid sequence of SEQ ID NO:32.

[0127] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising (a) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 230; (b) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 242; (c) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 253; (d) an HVR-L1 comprising the amino acid sequence of SEQ ID NO: 383; (e) an HVR-L2 comprising the amino acid sequence of SEQ ID NO: 16; and (f) an HVR-L3 comprising the amino acid sequence of SEQ ID NO: 32.

[0128] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising (a) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 233; (b) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 246; (c) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 131; (d) an HVR-L1 comprising the amino acid sequence of SEQ ID NO: 383; (e) an HVR-L2 comprising the amino acid sequence of SEQ ID NO: 19; and (f) an HVR-L3 comprising the amino acid sequence of SEQ ID NO: 37.

[0129] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising (a) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 234; (b) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 247; (c) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 131; (d) an HVR-L1 comprising the amino acid sequence of SEQ ID NO: 383; (e) an HVR-L2 comprising the amino acid sequence of SEQ ID NO: 19; and (f) an HVR-L3 comprising the amino acid sequence of SEQ ID NO: 37.

[0130] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising (a) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 233; (b) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 248; (c) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 131; (d) an HVR-L1 comprising the amino acid sequence of SEQ ID NO: 383; (e) an HVR-L2 comprising the amino acid sequence of SEQ ID NO: 19; and (f) an HVR-L3 comprising the amino acid sequence of SEQ ID NO: 37.

[0131] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising (a) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 233; (b) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 246; (c) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 254; (d) an HVR-L1 comprising the amino acid sequence of SEQ ID NO: 383; (e) an HVR-L2 comprising the amino acid sequence of SEQ ID NO: 19; and (f) an HVR-L3 comprising the amino acid sequence of SEQ ID NO: 37.

[0132] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising (a) an HVR-H1 comprising the amino acid sequence of SEQ ID NO:235; (b) an HVR-H2 comprising the amino acid sequence of SEQ ID NO:249; (c) an HVR-H3 comprising the amino acid sequence of SEQ ID NO:163; (d) an HVR-L1 comprising the amino acid sequence of SEQ ID NO:2; (e) an HVR-L2 comprising the amino acid sequence of SEQ ID NO:22; and (f) an HVR-L3 comprising the amino acid sequence of SEQ ID NO:69.

[0133] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising (a) an HVR-H1 comprising the amino acid sequence of SEQ ID NO:236; (b) an HVR-H2 comprising the amino acid sequence of SEQ ID NO:250; (c) an HVR-H3 comprising the amino acid sequence of SEQ ID NO:163; (d) an HVR-L1 comprising the amino acid sequence of SEQ ID NO:2; (e) an HVR-L2 comprising the amino acid sequence of SEQ ID NO:22; and (f) an HVR-L3 comprising the amino acid sequence of SEQ ID NO:69.

[0134] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising (a) an HVR-H1 comprising the amino acid sequence of SEQ ID NO:237; (b) an HVR-H2 comprising the amino acid sequence of SEQ ID NO:251; (c) an HVR-H3 comprising the amino acid sequence of SEQ ID NO:163; (d) an HVR-L1 comprising the amino acid sequence of SEQ ID NO:2; (e) an HVR-L2 comprising the amino acid sequence of SEQ ID NO:22; and (f) an HVR-L3 comprising the amino acid sequence of SEQ ID NO:69.

[0135] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising (a) an HVR-H1 comprising the amino acid sequence of SEQ ID NO:236; (b) an HVR-H2 comprising the amino acid sequence of SEQ ID NO:252; (c) an HVR-H3 comprising the amino acid sequence of SEQ ID NO:163; (d) an HVR-L1 comprising the amino acid sequence of SEQ ID NO:2; (e) an HVR-L2 comprising the amino acid sequence of SEQ ID NO:22; and (f) an HVR-L3 comprising the amino acid sequence of SEQ ID NO:69.

[0136] In some embodiments, provided herein are anti-SIRPβ1 antibodies comprising at least one, at least two, or all three selected from the following V HHVR sequences: (a) HVR-H1 comprising the HVR-H1 amino acid sequence shown in Table 4 and / or Table 9; (b) HVR-H2 comprising the HVR-H2 amino acid sequence shown in Table 4 and / or Table 9; (c) HVR-H3 comprising the HVR-H3 amino acid sequence shown in Table 4 and / or Table 9. In some embodiments, the anti-SIRPβ1 antibody comprises an SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-10, SB-11, SB-12, SB-13, SB-14, SB-15, SB-16, SB-17, SB-18, SB-19, SB-20, SB-21, SB-22, SB-23, SB-24, SB-25, SB-26, SB-27, SB-28, SB-29, SB-30, SB-31, SB-32, SB-33, SB-34, SB-35, SB-36, SB-37, SB-38, SB-39, SB-40, SB-41, SB-42, SB-43, SB-44, SB-45, SB-46, SB-47, SB-48, SB-49, SB-50, SB-51, SB-52, SB-53, SB-54, SB-55, SB-56, SB-57, SB-58, SB-59, SB-60, SB-61, SB-62, SB-63, SB-64, SB-65 -37,SB-38,SB-39,SB-40,SB-41,SB-42,SB-43,SB-44,SB-45,SB-46,SB-47,SB-48,SB-49,SB-50,SB-1-2,SB-1-3,SB-1-4,SB-1-5,SB-2-7,SB- 2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21 antibodies to HVR-H1, HVR-H2, and HVR-H3. In some embodiments, the anti-SIRPβ1 antibody comprises HVR-H1, HVR-H2, and HVR-H3 of an antibody selected from SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-14, SB-28, SB-39, SB-40, SB-49, SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21 (shown in Tables 4 and 9).In some embodiments, the anti-SIRPβ1 antibody comprises HVR-H1, HVR-H2, and HVR-H3 of an antibody selected from SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21 (shown in Table 9). In some embodiments, the anti-SIRPβ1 antibody comprises HVR-H1, HVR-H2, and HVR-H3 of an antibody selected from SB-1-3, SB-2-8, and SB-8-13 (shown in Table 9). In some embodiments, the anti-SIRPβ1 antibody comprises (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 229; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 239; and (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: 125. In some embodiments, the anti-SIRPβ1 antibody comprises (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 231; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 243; and (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: 126. In some embodiments, the anti-SIRPβ1 antibody comprises (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 233; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 246; and (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: 131.

[0137] In some embodiments, provided herein are anti-SIRPβ1 antibodies comprising at least one, at least two, or all three selected from the following V LHVR sequences: (a) HVR-L1 comprising the HVR-L1 amino acid sequence shown in Table 3 and / or Table 8; (b) HVR-L2 comprising the HVR-L2 amino acid sequence shown in Table 3 and / or Table 8; (c) HVR-L3 comprising the HVR-L3 amino acid sequence shown in Table 3 and / or Table 8. In some embodiments, the anti-SIRPβ1 antibody comprises an SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-10, SB-11, SB-12, SB-13, SB-14, SB-15, SB-16, SB-17, SB-18, SB-19, SB-20, SB-21, SB-22, SB-23, SB-24, SB-25, SB-26, SB-27, SB-28, SB-29, SB-30, SB-31, SB-32, SB-33, SB-34, SB-35, SB-36, SB-37, SB-38, SB-39, SB-40, SB-41, SB-42, SB-43, SB-44, SB-45, SB-46, SB-47, SB-48, SB-49, SB-50, SB-51, SB-52, SB-53, SB-54, SB-55, SB-56, SB-57, SB-58, SB-59, SB-60, SB-61, SB-62, SB-63, SB-64, SB-65 -37,SB-38,SB-39,SB-40,SB-41,SB-42,SB-43,SB-44,SB-45,SB-46,SB-47,SB-48,SB-49,SB-50,SB-1-2,SB-1-3,SB-1-4,SB-1-5,SB-2-7,SB- 2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21 antibodies to HVR-L1, HVR, L2, and HVR-L3. In some embodiments, the anti-SIRPβ1 antibody comprises HVR-L1, HVR, L2, and HVR-L3 of an antibody selected from SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-14, SB-28, SB-39, SB-40, SB-49, SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21 (shown in Tables 3 and 8).In some embodiments, the anti-SIRPβ1 antibody comprises HVR-L1, HVR, L2, and HVR-L3 of an antibody selected from SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21 (shown in Table 8). In some embodiments, the anti-SIRPβ1 antibody comprises HVR-L1, HVR, L2, and HVR-L3 of an antibody selected from SB-1-3, SB-2-8, and SB-8-13 (shown in Table 8). In some embodiments, the anti-SIRPβ1 antibody comprises (a) HVR-L1 comprising the amino acid sequence of SEQ ID NO: 383; (b) HVR-L2 comprising the amino acid sequence of SEQ ID NO: 16; and (c) HVR-L3 comprising the amino acid sequence of SEQ ID NO: 31. In some embodiments, the anti-SIRPβ1 antibody comprises (a) HVR-L1 comprising the amino acid sequence of SEQ ID NO: 383; (b) HVR-L2 comprising the amino acid sequence of SEQ ID NO: 16; and (c) HVR-L3 comprising the amino acid sequence of SEQ ID NO: 32. In some embodiments, the anti-SIRPβ1 antibody comprises (a) HVR-L1 comprising the amino acid sequence of SEQ ID NO: 383; (b) HVR-L2 comprising the amino acid sequence of SEQ ID NO: 19; and (c) HVR-L3 comprising the amino acid sequence of SEQ ID NO: 37.

[0138] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising (a) V H A domain comprising at least one, at least two, or all three VH HVR sequences selected from the group consisting of: (i) an HVR-H1 comprising the HVR-H1 amino acid sequence shown in Table 4 and / or Table 9, (ii) an HVR-H2 comprising the HVR-H2 amino acid sequence shown in Table 4 and / or Table 9, and (iii) an HVR-H3 comprising the HVR-H3 amino acid sequence shown in Table 4 and / or Table 9; and (b) a V L A domain comprising at least one, at least two, or all three selected from the following V L HVR sequences: (i) HVR-L1 comprising the HVR-L1 amino acid sequence shown in Table 3 and / or Table 8, (ii) HVR-L2 comprising the HVR-L2 amino acid sequence shown in Table 3 and / or Table 8, and (iii) HVR-L3 comprising the HVR-L3 amino acid sequence shown in Table 3 and / or Table 8.

[0139] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising (a) V H A domain comprising (i) an HVR-H1 comprising the HVR-H1 amino acid sequence shown in Table 4 and / or Table 9, (ii) an HVR-H2 comprising the HVR-H2 amino acid sequence shown in Table 4 and / or Table 9, and (iii) an HVR-H3 comprising the HVR-H3 amino acid sequence shown in Table 4 and / or Table 9; and (b) a V L A domain comprising (i) an HVR-L1 comprising the HVR-L1 amino acid sequence shown in Table 3 and / or Table 8, (ii) an HVR-L2 comprising the HVR-L2 amino acid sequence shown in Table 3 and / or Table 8, and (iii) an HVR-L3 comprising the HVR-L3 amino acid sequence shown in Table 3 and / or Table 8.

[0140] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising a V region comprising HVR-H1, HVR-H2, and HVR-H3. H domain, and V containing HVR-L1, HVR-L2, and HVR-L3 L domain, wherein HVR-H1, HVR-H2, HVR-H3, HVR-L1, HVR-L2, and HVR-L3 are selected from the group consisting of SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-10, SB-11, SB-12, SB-13, SB-14, SB-15, SB-16, SB-17, SB-18, SB-19, SB-20, SB-21, SB-22, SB-23, SB-24, SB-25, SB-26, SB-27, SB-28, SB-29, SB-30, SB-31, SB-32, SB-33, SB-34, SB-35 B-33,SB-34,SB-35,SB-36,SB-37,SB-38,SB-39,SB-40,SB-41,SB-42,SB-43,SB-44,SB-45,SB-46,SB-47,SB-48,SB-49,SB-50,SB-1-2,SB-1-3,SB- HVR of antibodies to 1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21.

[0141] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising a V region comprising HVR-H1, HVR-H2, and HVR-H3. H domain, and V containing HVR-L1, HVR-L2, and HVR-L3 L domains, wherein HVR-H1, HVR-H2, HVR-H3, HVR-L1, HVR-L2, and HVR-L3 are HVRs of an antibody selected from SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-14, SB-28, SB-39, SB-40, SB-49, SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21 (shown in Tables 3, 4, 8, and 9).

[0142] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising a V region comprising HVR-H1, HVR-H2, and HVR-H3. H domain, and V containing HVR-L1, HVR-L2, and HVR-L3 L domains, wherein HVR-H1, HVR-H2, HVR-H3, HVR-L1, HVR-L2, and HVR-L3 are HVRs of an antibody selected from SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21 (shown in Tables 8 and 9).

[0143] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising a V region comprising HVR-H1, HVR-H2, and HVR-H3. H domain, and V containing HVR-L1, HVR-L2, and HVR-L3 L Domains, wherein HVR-H1, HVR-H2, HVR-H3, HVR-L1, HVR-L2, and HVR-L3 are HVRs of an antibody selected from SB-1-3, SB-2-8, and SB-8-13 (shown in Tables 8 and 9).

[0144] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising (a) V H A domain comprising (i) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 229; (ii) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 239; (iii) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 125; and (b) a V L A domain comprising (iv) HVR-L1 comprising the amino acid sequence of SEQ ID NO:383; (v) HVR-L2 comprising the amino acid sequence of SEQ ID NO:16; and (vi) HVR-L3 comprising the amino acid sequence of SEQ ID NO:31.

[0145] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising (a) V H A domain comprising (i) an HVR-H1 comprising the amino acid sequence of SEQ ID NO: 231; (ii) an HVR-H2 comprising the amino acid sequence of SEQ ID NO: 243; (iii) an HVR-H3 comprising the amino acid sequence of SEQ ID NO: 126; and (b) a V L A domain comprising (iv) HVR-L1 comprising the amino acid sequence of SEQ ID NO:383; (v) HVR-L2 comprising the amino acid sequence of SEQ ID NO:16; and (vi) HVR-L3 comprising the amino acid sequence of SEQ ID NO:32.

[0146] In some embodiments, provided herein is an anti-SIRPβ1 antibody comprising (a) V H A domain comprising (i) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 233; (ii) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 246; (iii) HVR-H3 comprising the amino acid sequence of SEQ ID NO: 131; and (b) V L A domain comprising (iv) HVR-L1 comprising the amino acid sequence of SEQ ID NO:383; (v) HVR-L2 comprising the amino acid sequence of SEQ ID NO:19; and (vi) HVR-L3 comprising the amino acid sequence of SEQ ID NO:37.

[0147] In another aspect, the anti-SIRPβ1 antibody comprises a polypeptide having a sequence similar to SEQ ID NO: 268, 270, 272, 274, 276, 279, 281, 283, 285, 287, 289, 291, 293, 295, 297, 299, 301, 303, 305, 307, 309, 311, 313, 315, 317, 319, 321, 323, 325, 327, 329, 331, 333, 335, 337, 339, 341, 343, 345, 347, 348 9, 351, 353, 355, 357, 359, 361, 363, 365, 366, 367, 368, 369, 371, 372, 373, 374, 375, 376, 377, 378, 379, 380, 381, or 382 having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity H ) sequence. In certain embodiments, or V having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity to an amino acid sequence of any of: HThe sequence contains substitutions (e.g., conservative substitutions), insertions, or deletions relative to the reference sequence, but the anti-SIRPβ1 antibody comprising the sequence retains the ability to bind to SIRPβ1. In certain embodiments, the sequence has been in SEQ ID NO: 268, 270, 272, 274, 276, 279, 281, 283, 285, 287, 289, 291, 293, 295, 297, 299, 301, 303, 305, 307, 309, 311, 313, 315, 317, 319, 321, 323, 325, 327, 329, 331, 333, 335, 337, 339,341,343,345,347,349,351,353,355,357,359,361,363,365,366,367,368,369,371,372,373,374,375,376,377,378,379,380,381, or 382 by substitution, insertion, and / or deletion of a total of 1 to 10 amino acids. NO:268,270,272,274,276,279,281,283,285,287,289,291,293,295,297,299,301, 303,305,307,309,311,313,315,317,319,321,323,325,327,329,331,333,335,337 , 339,341,343,345,347,349,351,353,355,357,359,361,363,365,366,367,368,369,371,372,373,374,375,376,377,378,379,380,381, or 382 by substitution, insertion and / or deletion of a total of 1 to 5 amino acids. In certain embodiments, the substitution, insertion, or deletion occurs in a region outside of the HVR (i.e., in the FR). Optionally, the anti-SIRPβ1 antibody comprises SEQ ID NO:268,270,272,274,276,279,281,283,285,287,289,291,293,295,297,299 ,301,303,305,307,309,311,313,315,317,319,321,323,325,327,329,331,3 33,335,337,339,341,343,345,347,349,351,353,355,357,359,361,363,365,366,367,368,369,371,372,373,374,375,376,377,378,379,380,381, or 382 of VH In a specific embodiment, V H Comprising one, two or three HVRs selected from the following: (a) HVR-H1 comprising the HVR-H1 amino acid sequence shown in Table 4 and / or Table 9; (b) HVR-H2 comprising the HVR-H2 amino acid sequence shown in Table 4 and / or Table 9; (c) HVR-H3 comprising the HVR-H3 amino acid sequence shown in Table 4 and / or Table 9.

[0148] In another aspect, an anti-SIRPβ1 antibody is provided, wherein the antibody comprises a residue corresponding to SEQ ID NO: 267, 269, 271, 273, 275, 277, 278, 280, 282, 284, 286, 288, 290, 292, 294, 296, 298, 300, 302, 304, 306, 308, 310, 312, 314, 316, 318, 320, 322, 324, 326, 328, 330, 33 2,334,336,338,340,342,344,346,348,350,352,354,356,358,360,362,364, or 370 having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an amino acid sequence of L ). In certain embodiments, the same as SEQ ID NO: 267, 269, 271, 273, 275, 277, 278, 280, 282, 284, 286, 288, 290, 292, 294, 296, 298, 300, 302, 304, 306, 308, 310, 312, 314, 316, 318, 320, 322, 324, 326, 328, 330, 332, 334, 336, 338, 340, 342, 344, 346, 348, 350, 352, 354, 356, 358, 360, 362, 364, or 370 having an amino acid sequence at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to LThe sequence contains substitutions (eg, conservative substitutions), insertions, or deletions relative to the reference sequence, but the anti-SIRPβ1 antibody comprising the sequence retains the ability to bind to SIRPβ1. In some embodiments, a total of 1 to 10 amino acids have been substituted, inserted and / or deleted in SEQ ID NO: 267, 269, 271, 273, 275, 277, 278, 280, 282, 284, 286, 288, 290, 292, 294, 296, 298, 300, 302, 304, 306, 308, 310, 312, 314, 316, 318, 320, 322, 324, 326, 328, 330, 332, 334, 336, 338, 340, 342, 344, 346, 348, 350, 352, 354, 356, 358, 360, 362, 364, or 370. In certain embodiments, a total of 1 to 5 amino acids have been substituted, inserted and / or deleted in SEQ ID NO: 267, 269, 271, 273, 275, 277, 278, 280, 282, 284, 286, 288, 290, 292, 294, 296, 298, 300, 302, 304, 306, 308, 310, 312, 314, 316, 318, 320, 322, 324, 326, 328, 330, 332, 334, 336, 338, 340, 342, 344, 346, 348, 350, 352, 354, 356, 358, 360, 362, 364, or 370. In certain embodiments, substitutions, insertions, or deletions occur in regions outside of the HVRs (i.e., within the FRs). The SIRP β1 antibody comprises a V of SEQ ID NO: 267, 269, 271, 273, 275, 277, 278, 280, 282, 284, 286, 288, 290, 292, 294, 296, 298, 300, 302, 304, 306, 308, 310, 312, 314, 316, 318, 320, 322, 324, 326, 328, 330, 332, 334, 336, 338, 340, 342, 344, 346, 348, 350, 352, 354, 356, 358, 360, 362, 364, or 370. L In a specific embodiment, V LComprising one, two or three HVRs selected from the following: (a) HVR-L1 comprising the HVR-L1 amino acid sequence shown in Table 3 and / or Table 8; (b) HVR-L2 comprising the HVR-L2 amino acid sequence shown in Table 3 and / or Table 8; and (c) HVR-L3 comprising the HVR-L3 amino acid sequence shown in Table 3 and / or Table 8.

[0149] In some embodiments, an anti-SIRPβ1 antibody is provided, wherein the antibody comprises V in any of the embodiments provided above. H and V in any of the embodiments provided above L In some embodiments, provided herein is an anti-SIRPβ1 antibody, wherein the antibody comprises V in any of the embodiments provided above. H and V in any of the embodiments provided above L In one embodiment, the antibody comprises a member selected from the group consisting of SEQ ID NOs: 268, 270, 272, 274, 276, 279, 281, 283, 285, 287, 289, 291, 293, 295, 297, 299, 301, 303, 305, 307, 309, 311, 313, 315, 317, 319, 321, 323, 325, 327, 329, 331, 3 33,335,337,339,341,343,345,347,349,351,353,355,357,359,361,363,365,366,367,368,369,371,372,373,374,375,376,377,378,379,380,381, and 382 V H 340,342,344,346,348,350,352,354,356,358,360,362,364,370,371,372,373,374,376,377,378,380,382,384,386,38 ... Lsequences, including post-translational modifications of those sequences. In some embodiments, the antibody comprises an SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-10, SB-11, SB-12, SB-13, SB-14, SB-15, SB-16, SB-17, SB-18, SB-19, SB-20, SB-21, SB-22, SB-23, SB-24, SB-25, SB-26, SB-27, SB-28, SB-29, SB-30, SB-31, SB-32, SB-33, SB-34, SB-35, SB-36, SB-37,SB-38,SB-39,SB-40,SB-41,SB-42,SB-43,SB-44,SB-45,SB-46,SB-47,SB-48,SB-49,SB-50,SB-1-2,SB-1-3,SB-1-4,SB-1-5 , SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21 antibodies. H Sequence and V L In some embodiments, the antibody comprises a V sequence selected from the group consisting of SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-14, SB-28, SB-39, SB-40, SB-49, SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21. H Sequence and V L In some embodiments, the antibody comprises a V sequence selected from the group consisting of SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21. H Sequence and V L In some embodiments, the antibody comprises a V sequence of an antibody selected from SB-1-3, SB-2-8, and SB-8-13. H Sequence and VL In some embodiments, the antibody comprises V of SEQ ID NO: 367. H Sequence and V of SEQ ID NO:267 L In some embodiments, the antibody comprises V of SEQ ID NO: 372. H Sequence and V of SEQ ID NO:370 L In some embodiments, the antibody comprises V of SEQ ID NO: 376. H Sequence and V of SEQ ID NO: 280 L sequence.

[0150] In some embodiments, the anti-SIRPβ1 antibody is combined with a polypeptide comprising a polypeptide selected from the group consisting of SEQ ID NO: 268, 270, 272, 274, 276, 279, 281, 283, 285, 287, 289, 291, 293, 295, 297, 299, 301, 303, 305, 307, 309, 311, 313, 315, 317, 319, 321, 323, 325, 327, 329, 331, 3 33,335,337,339,341,343,345,347,349,351,353,355,357,359,361,363,365,366,367,368,369,371,372,373,374,375,376,377,378,379,380,381, and 382 V H A sequence and a V selected from the group consisting of SEQ ID NOs: 267, 269, 271, 273, 275, 277, 278, 280, 282, 284, 286, 288, 290, 292, 294, 296, 298, 300, 302, 304, 306, 308, 310, 312, 314, 316, 318, 320, 322, 324, 326, 328, 330, 332, 334, 336, 338, 340, 342, 344, 346, 348, 350, 352, 354, 356, 358, 360, 362, 364, and 370 LIn some embodiments, the anti-SIRPβ1 antibody competes for binding with an antibody selected from the group consisting of SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-10, SB-11, SB-12, SB-13, SB-14, SB-15, SB-16, SB-17, SB-18, SB-19, SB-20, SB-21, SB-22, SB-23, SB-24, SB-25, SB-26, SB-27, SB-28, SB-29, SB-30, SB-31, SB-32, SB-33, SB-34, SB-35, SB-36, SB-37, SB-38, SB-39, SB-40, SB-41, SB-42, SB-43, SB-44, SB-45, SB-46, SB-47, SB-48, SB-49, SB-50, SB-51, SB-52, SB-53, SB-54, SB-55, SB-56, SB-57, SB-58, SB-59, SB-60, SB-61, SB-62, SB-63, SB-64, SB-65 6,SB-37,SB-38,SB-39,SB-40,SB-41,SB-42,SB-43,SB-44,SB-45,SB-46,SB-47,SB-48,SB-49,SB-50,SB-1-2,SB-1-3,SB-1-4,SB-1-5 , SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21 antibodies compete for binding. In some embodiments, the anti-SIRPβ1 antibody competes for binding with an antibody selected from SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-14, SB-28, SB-39, SB-40, SB-49, SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21. In some embodiments, the anti-SIRPβ1 antibody competes for binding with an antibody selected from SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21. In some embodiments, the anti-SIRPβ1 antibody competes for binding with an antibody selected from SB-1-3, SB-2-8, and SB-8-13. In some embodiments, the anti-SIRPβ1 antibody competes for binding with a V comprising SEQ ID NO: 367. H Sequence and V of SEQ ID NO:267 LIn some embodiments, the anti-SIRPβ1 antibody competes with a V comprising SEQ ID NO: 372. H Sequence and V of SEQ ID NO:370 L In some embodiments, the anti-SIRPβ1 antibody competes with a V comprising SEQ ID NO: 376. H Sequence and V of SEQ ID NO: 280 L Sequences of antibodies competing for binding.

[0151] In some embodiments, the antibody binds to a polypeptide comprising a polypeptide selected from the group consisting of SEQ ID NO: 268, 270, 272, 274, 276, 279, 281, 283, 285, 287, 289, 291, 293, 295, 297, 299, 301, 303, 305, 307, 309, 311, 313, 315, 317, 319, 321, 323, 325, 327, 329, 331, 3 33,335,337,339,341,343,345,347,349,351,353,355,357,359,361,363,365,366,367,368,369,371,372,373,374,375,376,377,378,379,380,381, and 382 V H A sequence and a V selected from the group consisting of SEQ ID NOs: 267, 269, 271, 273, 275, 277, 278, 280, 282, 284, 286, 288, 290, 292, 294, 296, 298, 300, 302, 304, 306, 308, 310, 312, 314, 316, 318, 320, 322, 324, 326, 328, 330, 332, 334, 336, 338, 340, 342, 344, 346, 348, 350, 352, 354, 356, 358, 360, 362, 364, and 370 LIn some embodiments, the antibody binds to an epitope of human SIRP β1 that is identical to or overlaps with an epitope of an anti-SIRP β1 antibody of the sequence. In some embodiments, the antibody binds to an epitope selected from the group consisting of SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-10, SB-11, SB-12, SB-13, SB-14, SB-15, SB-16, SB-17, SB-18, SB-19, SB-20, SB-21, SB-22, SB-23, SB-24, SB-25, SB-26, SB-27, SB-28, SB-29, SB-30, SB-31, SB-32, SB-33, SB-34, SB-35, SB-36, SB-37, SB-38, The anti-SIRPβ1 antibodies of SB-39, SB-40, SB-41, SB-42, SB-43, SB-44, SB-45, SB-46, SB-47, SB-48, SB-49, SB-50, SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21 bind to the same or overlapping epitope of human SIRPβ1.

[0152] In some embodiments, the antibody binds to an epitope of human SIRP β1 that is the same as or overlaps with an epitope bound by an anti-SIRP β1 antibody selected from SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-14, SB-28, SB-39, SB-40, SB-49, SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21. In some embodiments, the antibody binds to a human SIRP β1 epitope that is identical or overlapping with an epitope bound by an anti-SIRP β1 antibody selected from SB-1-2, SB-1-3, SB-1-4, SB-1-5, SB-2-7, SB-2-8, SB-2-9, SB-2-10, SB-2-11, SB-8-13, SB-8-14, SB-8-15, SB-8-16, SB-40-18, SB-40-19, SB-40-20, and SB-40-21. In some embodiments, the antibody binds to a human SIRP β1 epitope that is identical or overlapping with an epitope bound by an anti-SIRP β1 antibody selected from SB-1-3, SB-2-8, and SB-8-13. In some embodiments, the anti-SIRP β1 antibody binds to a V comprising SEQ ID NO: 367. H Sequence and V of SEQ ID NO:267 L In some embodiments, the anti-SIRPβ1 antibody binds to a human SIRPβ1 epitope that is identical to or overlaps with a V sequence comprising SEQ ID NO: 372. H Sequence and V of SEQ ID NO:370 L In some embodiments, the anti-SIRPβ1 antibody binds to a human SIRPβ1 epitope that is identical to or overlaps with a V sequence comprising SEQ ID NO: 376. H Sequence and V of SEQ ID NO: 280 L In some embodiments, the epitope that the anti-SIRPβ1 antibody binds to is identical to or overlaps with a human SIRPβ1 epitope. In some embodiments, the human SIRPβ1 epitope is identical to the epitope that the anti-SIRPβ1 antibody binds to.

[0153] In some embodiments, the anti-SIRP β1 antibody binds to an epitope within amino acids 30 to 148 of SEQ ID NO: 1. In some embodiments, the anti-SIRP β1 antibody binds to an epitope within amino acids 30 to 136 of SEQ ID NO: 1. In some embodiments, the anti-SIRP β1 antibody binds to an epitope within amino acids 30 to 80 of SEQ ID NO: 1. In some embodiments, the anti-SIRP β1 antibody binds to an epitope within amino acids 40 to 90 of SEQ ID NO: 1. In some embodiments, the anti-SIRP β1 antibody binds to an epitope within amino acids 50 to 100 of SEQ ID NO: 1. In some embodiments, the anti-SIRP β1 antibody binds to an epitope within amino acids 60 to 110 of SEQ ID NO: 1. In some embodiments, the anti-SIRP β1 antibody binds to an epitope within amino acids 70 to 120 of SEQ ID NO: 1. In some embodiments, the anti-SIRP β1 antibody binds to an epitope within amino acids 80 to 130 of SEQ ID NO: 1. In some embodiments, the anti-SIRPβ1 antibody binds to an epitope within amino acids 90 to 140 of SEQ ID NO:1.

[0154] In some embodiments, the anti-SIRPβ1 antibody according to any of the above embodiments is a monoclonal antibody, including humanized and / or human antibodies. In some embodiments, the anti-SIRPβ1 antibody is an antibody fragment, such as Fv, Fab, Fab', scFv, diabody, or F(ab')2 fragment. In some embodiments, the anti-SIRPβ1 antibody is a substantially full-length antibody, such as an IgG1 antibody, an IgG2a antibody, or other antibody classes or isotypes as defined herein.

[0155] In some embodiments, the anti-SIRPβ1 antibody according to any of the above embodiments may incorporate any of the features described in Sections 1-7 below, alone or in combination: (1) Anti-SIRPβ1 antibody binding affinity

[0156] In some embodiments of any of the antibodies provided herein, the antibody has a p-value 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, e.g. 10 -8 M to 10 -13 M, for example 10 -9 M to 10 - 13 The dissociation constant (K D). The dissociation constant can be determined via any analytical technique, including any biochemical or biophysical technique, such as ELISA, surface plasmon resonance (SPR), biolayer interferometry (see, e.g., Octet System, ForteBio), isothermal titration calorimetry (ITC), differential scanning calorimetry (DSC), circular dichroism (CD), stopped-flow analysis, and colorimetric or fluorescent protein melting analysis. In one embodiment, K D is measured by a radiolabeled antigen binding assay (RIA). In some embodiments, the RIA is performed using a Fab version of the antibody of interest and its antigen, such as described in Chen et al., J. Mol. Biol. 293:865-881 (1999). In some embodiments, K D K is measured using a BIACORE surface plasmon resonance assay, e.g., an assay performed at 25°C with an immobilized antigen CM5 chip using a BIACORE-2000 or BIACORE-3000 (BIAcore, Inc., Piscataway, NJ). In some embodiments, K D Is using ForteBio Red384 system (ForteBio, Menlo Park, CA), e.g., as discussed in the Examples herein.

[0157] In some embodiments, K D is determined using a monovalent antibody (e.g., Fab) or a full-length antibody. D The assay was performed in a monovalent format using full-length antibodies. (2) Antibody fragments

[0158] 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. Pat. Nos. 5,571,894 and 5,587,458. For a discussion of Fab and F(ab')2 fragments that contain salvage receptor binding epitope residues and have extended in vivo half-lives, see U.S. Pat. No. 5,869,046.

[0159] Diabodies are antibody fragments with two antigen binding sites, which can be bivalent or bispecific. See, e.g., EP 404,097; WO 1993 / 01161; Hudson et al., Nat. Med. 9:129-134 (2003). Triabodies 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 part of the heavy chain variable domain, or all or part 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).

[0160] 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), as described herein. (3) Chimeric and humanized antibodies

[0161] In some embodiments of any of the antibodies provided herein, the antibody is a chimeric antibody. Certain chimeric antibodies are described, for example, 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-switched" antibody in which the class or subclass has been changed from that of the parent antibody. Chimeric antibodies include antigen-binding fragments thereof.

[0162] In some embodiments of any of the antibodies provided herein, the antibody is a humanized antibody. Typically, a non-human antibody is humanized to reduce immunogenicity to humans while retaining the specificity and affinity of the parent non-human antibody. In certain embodiments, a humanized antibody is substantially non-immunogenic in humans. In certain embodiments, a humanized antibody has substantially the same affinity for a target as an antibody from another species from which the humanized antibody is derived. See, e.g., U.S. Pat. Nos. 5,530,101, 5,693,761; 5,693,762; and 5,585,089. In certain embodiments, amino acids in an antibody variable domain that can be modified without reducing the natural affinity of the antigen binding domain while reducing its immunogenicity are identified. See, e.g., U.S. Pat. Nos. 5,766,886 and 5,869,619. Generally, a humanized antibody comprises one or more variable domains, wherein the HVR (or a portion thereof) is derived from a non-human antibody, and the FR (or a portion thereof) is derived from a human antibody sequence. Optionally, the humanized antibody 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), for example, to restore or improve antibody specificity or affinity.

[0163] Humanized antibodies and methods for their generation are reviewed in, for example, Almagro et al., Front. Biosci. 13: 1619-1633 (2008), and further described in, for example, U.S. Patent Nos. 5,821,337, 7,527,791, 6,982,321, and 7,087,409. Human framework regions that can be used for humanization include, but are not limited to, framework regions selected using the "best-fit" method (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 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)); 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 by 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)). (4) Human Antibodies

[0164] In some embodiments of any of the antibodies provided herein, the antibody is a human antibody. Human antibodies can be generated using a variety of techniques known in the art. Generally, human antibodies are described in van Dijk et al., Curr. Opin. Pharmacol. 5: 368-74 (2001) and Lonberg, Curr. Opin. Immunol. 20: 450-459 (2008).

[0165] Human antibodies can be prepared by administering immunogens to transgenic animals that have been modified to produce complete human antibodies or complete antibodies with human variable regions in response to antigenic challenge. Mouse strains deficient in mouse antibody production can be transformed with large fragments of the human Ig locus, with the expectation that such mice will produce human antibodies in the absence of mouse antibodies. Large human Ig fragments can retain a large variable gene diversity and appropriate regulation of antibody production and expression. By utilizing the mouse machinery for antibody diversification and selection and lack of immunological tolerance to human proteins, the full set of human antibodies reproduced in these mouse strains can produce high-affinity fully human antibodies against any antigen of interest, including human antigens. Using hybridoma technology, antigen-specific human monoclonal antibodies with desired specificity can be generated and selected. Certain exemplary methods are described in U.S. Patent Nos. 5,545,807, EP 546,073, and EP 546,073. See also, for example, U.S. Patent Nos. 6,075,181 and 6,150,584, which describe XENOMOUSETM technology; U.S. Patent No. 5,770,429, which describes Technology; U.S. Patent No. 7041870, which describes KM Technology; and U.S. Patent Application Publication No. US2007 / 0061900, which describes The human variable regions from intact antibodies generated by such animals can be further modified, for example, by combining with a different human constant region.

[0166] Human antibodies can also be generated by hybridoma-based methods. Human myeloma and mouse-human heteromyeloma cell lines for generating human monoclonal antibodies have been described (see, e.g., Kozbor, J. Immunol. 133: 3001 (1984) and Boerner et al., J. Immunol. 147: 86 (1991)). Human antibodies generated via human B cell hybridoma technology are also described in Li et al., Proc. Natl. Acad. Sci. USA 103: 3557-3562 (2006). Other methods include those described in, e.g., U.S. Patent No. 7,189,826 (which describes the generation of monoclonal human IgM antibodies from hybridoma cell lines). Human hybridoma technology (Trioma technology) is also recorded 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 generated as follows, that is, Fv clone variable domain sequences selected from human-derived phage display libraries are separated. Then, such variable domain sequences can be combined with desired human constant domains. The technology of selecting human antibodies from antibody libraries is described below.

[0100] In some embodiments of any of the antibodies provided herein, the antibody is a human antibody isolated by in vitro methods and / or screening of combinatorial libraries for antibodies having the desired activity or activities. Suitable examples include, but are not limited to, phage display (CAT, Morphosys, Dyax, Biosite / Medarex, Xoma, Symphogen, Alexion (formerly Proliferon), Affimed), ribosome display (CAT), yeast-based platform technology (Adimab), and the like. In certain phage display methods, repertoires of VH and VL genes are cloned separately by polymerase chain reaction (PCR) and randomly recombined in phage libraries, which can then be screened for antigen-binding phages, as described in Winter et al., Ann. Rev. Immunol. 12:433-455 (1994). For example, a variety of methods for generating phage display libraries and screening such libraries for antibodies having the desired binding characteristics 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 from immunized sources provide high-affinity antibodies to the immunogen without the need to construct hybridomas. Alternatively, a non-immune repertoire can be cloned (e.g., from humans) to provide a single source of antibodies against a large number of non-self and also self-antigens without any immunization, as described by Griffiths et al., EMBO J, 12: 725-734 (1993). Finally, non-immune libraries can also be generated synthetically by cloning non-rearranged V gene segments from stem cells and using PCR primers containing random sequences to encode highly variable HVR3 regions and achieve rearrangement in vitro, as described by Hoogenboom et al., J. Mol. Biol. 227: 381-388 (1992). Patent publications 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. (5) Constant region including Fc region

[0101] In some embodiments of any 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 is of the IgG class, the IgM class, or the IgA class.

[0102] 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 includes an Fc region. In some embodiments, the antibody induces one or more SIRPβ1 activities or is independent of binding to an Fc receptor. In some embodiments, the antibody binds to an inhibitory Fc receptor. In certain embodiments, the inhibitory Fc receptor is an inhibitory Fcγ receptor IIB (FcγIIB).

[0103] 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 includes 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 (Fc gamma IIB).

[0104] 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 includes 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 (Fc gamma IIB).

[0105] In certain embodiments of any 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 EU numbering of human IgG2 and amino acids 261-447 according to EU numbering of human IgG4 (WO 1997 / 11971; WO 2007 / 106585).

[0106] In some embodiments, the Fc region increases clustering but does not activate complement compared to a corresponding antibody comprising an Fc region that does not comprise the amino acid substitution. In some embodiments, the antibody induces one or more activities of a target specifically bound by the antibody. In some embodiments, the antibody binds to SIRPβ1.

[0107] It may also be desirable to modify the anti-SIRPβ1 antibodies of the present disclosure to modify the effector function of the antibody and / or increase serum half-life. For example, the Fc receptor binding site on the constant region may be modified or mutated to remove or reduce binding affinity for certain Fc receptors, such as FcγRI, FcγRII, and / or FcγRIII, to reduce antibody-dependent cell-mediated cytotoxicity. In some embodiments, the effector function is weakened by removing N-glycosylation of the Fc region of the antibody (e.g., in the CH2 domain of IgG). In some embodiments, the effector function is weakened 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, it may also be desirable to modify the anti-SIRPβ1 antibodies of the present disclosure to modify effector function to increase selectivity for targeting ITIM-containing FcγRIIb (CD32b) to increase clustering of SIRPβ1 antibodies on adjacent cells without activating humoral responses, including antibody-dependent cell-mediated cytotoxicity and antibody-dependent cellular phagocytosis.

[0108] In order to increase the serum half-life of the antibody, a salvage receptor binding epitope can be incorporated into the antibody (particularly an antibody fragment), as described, for example, in U.S. Pat. No. 5,739,277. As used herein, the term "salvage receptor binding epitope" refers to an epitope in 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. (6) Multispecific Antibodies

[0109] Multispecificity is an antibody with binding specificity for at least two different epitopes, including those on the same or another polypeptide (e.g., one or more SIRPβ1 polypeptides of the present disclosure). In some embodiments, the multispecific antibody is a bispecific antibody. In some embodiments, the multispecific antibody is a trispecific antibody. In some embodiments, the multispecific antibody is a tetraspecific antibody. 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 SIRPβ1 and comprises a second antigen binding region that binds to a second site on SIRPβ1. In some embodiments, the multispecific antibody comprises a first antigen binding region that binds to SIRPβ1 and a second antigen binding region that binds to a second polypeptide.

[0110] Provided herein are multispecific antibodies comprising a first antigen binding region that binds SIRPβ1, wherein the first antigen binding region comprises six HVRs of an antibody described herein, and a second antigen binding region that binds a second polypeptide. In some embodiments, the first antigen binding region comprises VVRs of an antibody described herein. H or V L .

[0111] In some embodiments of any of the multispecific antibodies, the second polypeptide is an antigen that promotes transport across the blood-brain barrier. In some embodiments, the antibodies herein are conjugated to a peptide that promotes transport across the blood-brain barrier. Numerous antigens and peptides that promote 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 and peptides 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 CRM197 (a non-toxic mutant of diphtheria toxin), TMEM 30 (A) (flipase), protein transduction domains, such as TAT, Syn-B, or penetratin, polyarginine or generally positively charged peptides, Angiopep 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)). In some embodiments, the second polypeptide is transferrin.

[0112] In some embodiments of any of the multispecific antibodies, the second polypeptide is a pathogenic protein selected from 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 protein, calcitonin, superoxide dismutase, ataxin, ataxin 1, ataxin 2, ataxin 3, ataxin 7, ataxin 8, ataxin 10, Lewy body , atrial natriuretic factor, islet amyloid polypeptide, insulin, apolipoprotein AI, serum amyloid A, medin, prolactin, transthyretin, lysozyme, β2 microglobulin, gelsolin, corneal epithelial protein, cystatin, immunoglobulin 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. 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 alpha-synuclein.

[0113] In some embodiments of any of the multispecific antibodies, the second polypeptide is a ligand and / or protein expressed on an immune cell, wherein the ligand and / or protein is 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.

[0114] In some embodiments of any of the multispecific antibodies, the second polypeptide is a protein, lipid, polysaccharide, or glycolipid, and any combination thereof, expressed on one or more tumor cells.

[0115] In some embodiments of any of the multispecific antibodies, the second polypeptide is an immunoglobulin-like receptor, such as TREM2. In some embodiments of any of the multispecific antibodies, the second polypeptide is an immunoglobulin-like receptor expressed on cells of the myeloid lineage.

[0116] The multivalent antibody contains at least one polypeptide chain (and preferably two polypeptide chains), wherein the one or more polypeptide chains contain two or more variable domains. For example, one or more polypeptide chains may contain 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 include V H -C H 1-Flexible joint-V H -C H 1-Fc region chain; or V H -C H 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 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.

[0117] Techniques for generating multispecific antibodies include, but are not limited to, recombinant co-expression of two pairs of 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-into-hole" engineering (see, e.g., U.S. Pat. No. 5,731,168). See also WO 2013 / 026833 (CrossMab). Multispecific antibodies can also be generated by engineering electrostatic manipulation effects for generating antibody Fc-heterodimer molecules (WO 2009 / 089004A1); cross-linking two or more antibodies (see, e.g., U.S. Pat. No. 4,676,980); using leucine; using "diabody" technology for generating 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).

[0118] Also included herein are engineered antibodies with three or more functional antigen binding sites, including "octopus antibodies" (see, e.g., US2006 / 0025576). Antibodies herein also include "dual-acting FAbs" or "DAFs" that contain antigen binding sites that bind to multiple SIRPβ1 (see, e.g., US2008 / 0069820). (7) Antibody variants

[0119] 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. (i) Substitution, insertion, and deletion variants

[0120] In some embodiments of any of the antibodies provided herein, antibody variants having one or more amino acid substitutions are provided. The amino acid sequence variants of the antibody can be prepared by introducing suitable modifications into the nucleotide sequence encoding the antibody or by peptide synthesis. Such modifications include, for example, residue deletions and / or insertions and / or substitutions within the antibody amino acid sequence. Table 1: Amino Acid Substitutions Initial residue Exemplary Alternatives Preferred alternative Ala(A) Val; Leu; Ile Val Arg(R) Lys; Gln; Asn Lys Asn(N) Gln; His; Asp; Lys; Arg Gln Asp(D) Glu; Asn Glu Cys(C) Ser; Ala Ser Gln(Q) Asn;Glu Asn Glu(E) Asp; Gln Asp Gly(G) Ala Ala His(H) Asn; Gln; Lys; Arg Arg Ile(I) Leu; Val; Met; Ala; Phe; Norleucine Leu Leu(L) Norleucine; Ile; Val; Met; Ala; Phe Ile Lys(K) Arg; Gln; Asn Arg Met(M) Leu; Phe; Ile Leu Phe(F) Leu; Val; Ile; Ala; Tyr Tyr Pro(P) Ala Ala Ser(S) Thr Thr Thr(T) Ser Ser Trp(W) Tyr; Phe Tyr Tyr(Y) Trp; Phe; Thr; Ser Phe Val(V) Ile; Leu; Met; Phe; Ala; norleucine Leu

[0121] 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 region of the substitution, for example as a 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 grouped based on common side chain properties: (1) Hydrophobic: norleucine, Met, Ala, Val, Leu, Ile; (2) Neutral hydrophilic: Cys, Ser, Thr, Asn, Gln; (3) Acidic: Asp, Glu; (4) Basic: His, Lys, Arg; (5) Residues that affect chain orientation: Gly, Pro; and (6) Aromatic: Trp, Tyr, Phe.

[0122] For example, non-conservative substitutions may involve exchanging a member of one of these classes for a member from another class. Such replacement residues may be introduced, for example, into regions of human antibodies homologous to non-human antibodies or into non-homologous regions of the molecule.

[0123] When making changes to the polypeptides or antibodies described herein, according to certain embodiments, the hydrophobicity index of the amino acids can be considered. Each amino acid has been assigned a hydrophobicity index based on its hydrophobicity and charge characteristics. 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).

[0124] The importance of the hydrophobic 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 replace other amino acids with similar hydrophobicity indexes or scores and still retain similar biological activity. When making changes based on the hydrophobicity index, in certain embodiments, replacement of amino acids whose hydrophobicity index is within ±2 is included. In certain embodiments, those within ±1 are included, and in certain embodiments, those within ±0.5 are included.

[0125] It is also understood in the art that similar amino acid substitutions can be made effectively based on hydrophobicity, particularly where the biologically functional protein or peptide created thereby is intended for use in immunological embodiments, as in the present case. In certain embodiments, the greatest local average hydrophilicity of a protein (as determined by the hydrophilicity of its neighboring amino acids) correlates with its immunogenicity and antigenicity, i.e., the biological properties of the protein.

[0126] The following hydrophilicity 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, substitution of amino acids whose hydrophilicity values ​​are within ±2 is included, in certain embodiments, those within ±1, and in certain embodiments, those within ±0.5. Epitopes can also be identified from the primary amino acid sequence based on hydrophilicity. These regions are also referred to as "epitope core regions".

[0127] In certain embodiments, substitutions, insertions, or deletions may occur within one or more HVRs, as long as such changes do not substantially reduce the ability of the antibody to bind to antigen. For example, conservative changes (e.g., conservative substitutions, as provided herein) may be made in HVRs that do not substantially reduce binding affinity. Such changes may, for example, be outside of antigen contact residues in HVRs. In certain embodiments of the variant VH and VL sequences provided above, each HVR is either unchanged or contains no more than 1, 2, or 3 amino acid substitutions.

[0128] Amino acid sequence insertions include amino and / or carboxyl terminal fusions of polypeptides ranging in length from 1 residue to 100 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 antibody molecules include fusions of the N or C termini of antibodies with enzymes (e.g., for ADEPT) or polypeptides that extend the serum half-life of the antibody.

[0129] Any cysteine ​​residues not involved in maintaining the correct conformation of the antibody may also be substituted, generally with serine, to improve the oxidative stability of the molecule and prevent aberrant cross-linking. Conversely, cysteine ​​bond(s) may be added to the antibody to improve its stability (particularly where the antibody is an antibody fragment, such as an Fv fragment). (ii) Glycosylation variants

[0130] In some embodiments of any of the antibodies provided herein, the antibody is altered to increase or decrease the degree of glycosylation of the antibody. Adding or deleting glycosylation sites to an antibody can be conveniently accomplished by altering the amino acid sequence so as to create or eliminate one or more glycosylation sites.

[0131] Glycosylation of antibodies is typically either N-linked or O-linked. N-linked refers to the attachment of a carbohydrate moiety to the side chain of an asparagine residue. The tripeptide sequences asparagine-X-serine and asparagine-X-threonine (where X is any amino acid except proline) are the recognition sequences for enzymatic attachment of a carbohydrate moiety to the side chain of asparagine. Thus, the presence of either of these tripeptide sequences in a polypeptide creates a potential glycosylation site. O-linked glycosylation refers to the attachment of one of the sugars N-acetylgalactosamine, galactose, or xylose to a hydroxyamino acid, most commonly serine or threonine, although 5-hydroxyproline or 5-hydroxylysine may also be used.

[0132] Addition of glycosylation sites to the antibody is conveniently accomplished by altering the amino acid sequence so that it contains one or more of the tripeptide sequences described above (for N-linked glycosylation sites). The alteration may also be made by adding or substituting one or more serine or threonine residues to the sequence of the original antibody (for O-linked glycosylation sites).

[0133] In the case where the antibody comprises an Fc region, the carbohydrate attached to the Fc region can be altered. Natural antibodies produced by mammalian cells typically contain branched, biantennary oligosaccharides, which are generally attached to the CH2 domain of the Fc region by an N-linkage at Asn297 according to Kabat numbering. Oligosaccharides can include various carbohydrates, such as mannose, N-acetylglucosamine (GlcNAc), galactose, and sialic acid, as well as fucose attached to the GlcNAc in the "backbone" of the biantennary oligosaccharide structure. In some embodiments, the oligosaccharides in the antibodies of the invention can be modified to create antibody variants with certain improved properties.

[0134] 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 publications relating to "defucosylated" or "fucose-deficient" antibody variants include: US2003 / 0157108; US2003 / 0115614; US2002 / 0164328; US2004 / 0093621; US2004 / 0132140; US2004 / 0110704; US2004 / 0110282; US2004 / 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); Kanda et al., Biotechnol. Bioeng. 94(4):680-688 (2006)). (iii) Modified constant region

[0135] 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.

[0136] 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, 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 (McEarchern 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, wherein the amino acid positions are according to the EU numbering convention.

[0137] In some embodiments of any IgG1 modified Fc, the Fc comprises an N297A mutation according to EU numbering. In some embodiments of any IgG1 modified Fc, the Fc comprises a D265A and N297A mutation according to EU numbering. In some embodiments of any IgG1 modified Fc, the Fc comprises a 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 IgG1 modified Fc, the Fc comprises an L234A and G237A mutation according to EU numbering. In some embodiments of any IgG1 modified Fc, the Fc comprises an L234A, L235A, and G237A mutation according to EU numbering. In some embodiments of any IgG1 modified Fc, the Fc comprises one or more (including all) of P238D, L328E, E233, G237D, H268D, P271G and A330R mutations according to EU numbering. In some embodiments of any IgG1 modified Fc, the Fc comprises one or more of S267E / L328F mutations according to EU numbering. In some embodiments of any IgG1 modified Fc, the Fc comprises P238D, L328E, E233D, G237D, H268D, P271G and A330R mutations according to EU numbering. In some embodiments of any IgG1 modified Fc, the Fc comprises P238D, L328E, G237D, H268D, P271G and A330R mutations according to EU numbering. In some embodiments of any IgG1 modified Fc, the Fc comprises P238D, S267E, L328E, E233D, G237D, H268D, P271G, and A330R mutations according to EU numbering. In some embodiments of any IgG1 modified Fc, the Fc comprises P238D, S267E, L328E, G237D, H268D, P271G, and A330R mutations according to EU numbering. In some embodiments of any IgG1 modified Fc, the Fc comprises C226S, C229S, E233P, L234V, and L235A mutations according to EU numbering. In some embodiments of any IgG1 modified Fc, the Fc comprises L234F, L235E, and P331S mutations according to EU numbering. In some embodiments of any IgG1 modified Fc, the Fc comprises S267E and L328F mutations according to EU numbering. In some embodiments of any IgG1 modified Fc, the Fc comprises an S267E mutation according to EU numbering.In some embodiments of any of the IgG1 modified Fc, the Fc comprises replacing the constant heavy 1 (CH1) and hinge regions of IgG1, and kappa light chains with the CH1 and hinge regions of IgG2 (amino acids 118-230 of IgG2 according to EU numbering).

[0138] In some embodiments of any IgG1 modified Fc, the Fc comprises two or more amino acid substitutions that increase antibody clustering but do not activate complement compared to a corresponding antibody having an Fc region that does not include the two or more amino acid substitutions. Thus, in some embodiments of any IgG1 modified Fc, the IgG1 modified Fc is an antibody comprising an Fc region, wherein the antibody comprises an amino acid substitution at position E430G and one or more amino acid substitutions at a residue position in the Fc region selected from L234F, L235A, L235E, S267E, K322A, L328F, A330S, P331S, and any combination thereof according to EU numbering. In some embodiments, the IgG1 modified Fc comprises amino acid substitutions at positions E430G, L243A, L235A, and P331S according to EU numbering. In some embodiments, the IgG1 modified Fc comprises amino acid substitutions at positions E430G and P331S according to EU numbering. In some embodiments, the IgG1 modified Fc comprises amino acid substitutions at positions E430G and K322A according to the EU numbering. In some embodiments, the IgG1 modified Fc comprises amino acid substitutions at positions E430G, A330S, and P331S according to the EU numbering. In some embodiments, the IgG1 modified Fc comprises amino acid substitutions at positions E430G, K322A, A330S, and P331S according to the EU numbering. In some embodiments, the IgG1 modified Fc comprises amino acid substitutions at positions E430G, K322A, and A330S according to the EU numbering. In some embodiments, the IgG1 modified Fc comprises amino acid substitutions at positions E430G, K322A, and P331S according to the EU numbering.

[0139] In some embodiments of any IgG1 modified Fc, the IgG1 modified Fc herein can be further combined with 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 IgG1 modified Fc, the IgG1 modified Fc can further comprise one or more of A330L, A330S, L234F, L235E, and / or P331S according to the EU numbering convention. In some embodiments of any IgG1 modified Fc, the IgG1 modified Fc may further comprise one or more mutations to enhance the half-life of the antibody 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 IgG1 modified Fc, 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 the EU numbering convention.

[0140] Other aspects of the present disclosure relate to antibodies with modified constant regions (i.e., Fc regions). Antibodies that rely on binding to FcγR receptors to activate targeted receptors may lose their agonist activity if engineered to eliminate FcγR binding (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)). As such, it is believed that when the antibody has an Fc domain from the human IgG2 isotype (CH1 and hinge region) or another type of Fc domain that is capable of preferentially binding to the inhibitory FcγRIIB receptor, or a variant thereof, the anti-SIRPβ1 antibodies of the present disclosure with the correct epitope specificity can activate the target antigen with minimal adverse effects.

[0141] 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 IgG2 modified Fc, one or more amino acid substitutions are selected from V234A according to 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 (US2007 / 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):28 (2000); Journal1(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 IgG2 modified Fc, the Fc comprises amino acid substitutions at positions V234A and G237A according to EU numbering. In some embodiments of any IgG2 modified Fc, the Fc comprises amino acid substitutions at positions C219S or C220S according to EU numbering. In some embodiments of any IgG2 modified Fc, the Fc comprises amino acid substitutions at positions A330S and P331S according to EU numbering. In some embodiments of any of the IgG2 modified Fc, the Fc comprises amino acid substitutions at positions S267E and L328F according to EU numbering.

[0142] In some embodiments of any IgG2 modified Fc, 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 IgG2 modified Fc, the antibody has an IgG2 isotype and 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).

[0143] In some embodiments of any IgG2 modified Fc, the Fc comprises a C220S amino acid substitution according to the EU numbering convention. In some embodiments of any IgG2 modified Fc, the antibody has an IgG2 isotype and a kappa light chain constant domain comprising a C214S amino acid substitution according to the EU numbering convention.

[0144] In some embodiments of any IgG2 modified Fc, the Fc comprises a C219S amino acid substitution according to the EU numbering convention. In some embodiments of any IgG2 modified Fc, the antibody has an IgG2 isotype and a kappa light chain constant domain comprising a C214S amino acid substitution according to the EU numbering convention.

[0145] In some embodiments of any IgG2 modified Fc, 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 IgG2 modified Fc, the IgG2 isotype CH1 and hinge region comprise an amino acid sequence of 118-230 according to the EU numbering convention. In some embodiments of any IgG2 modified Fc, the antibody Fc region comprises an S267E amino acid substitution, an L328F amino acid substitution, or both, and / or an N297A or N297Q amino acid substitution according to the EU numbering convention.

[0146] In some embodiments of any IgG2 modified Fc, the Fc further comprises one or more amino acid substitutions at positions E430G, E430S, E430F, E430T, E345K, E345Q, E345R, E345Y, S440Y, and S440W according to the EU numbering method. In some embodiments of any IgG2 modified Fc, the Fc may further comprise one or more mutations to enhance the half-life of the antibody in human serum (e.g., one or more (including all) of the M252Y, S254T, and T256E mutations according to the EU numbering rules). In some embodiments of any IgG2 modified Fc, the Fc may further comprise A330S and P331S.

[0147] In some embodiments of any IgG2 modified Fc, 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 IgG2 modified Fc, the Fc comprises one or more amino acid substitutions at positions H268Q, V309L, A330S, and P331S according to EU numbering.

[0148] In some embodiments of any 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.

[0149] In certain embodiments of any IgG1 and / or IgG2 modified Fc, the Fc comprises one or more amino acid substitutions at a residue position 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 IgG1 and / or IgG2 modified Fc, the Fc comprises amino acid substitutions at positions E430G, L243A, L235A, and P331S according to EU numbering. In some embodiments of any IgG1 and / or IgG2 modified Fc, the Fc comprises amino acid substitutions at positions E430G and P331S according to EU numbering. In some embodiments of any IgG1 and / or IgG2 modified Fc, the Fc comprises amino acid substitutions at positions E430G and K322A according to the EU numbering. In some embodiments of any IgG1 and / or IgG2 modified Fc, the Fc comprises amino acid substitutions at positions E430G, A330S, and P331S according to the EU numbering. In some embodiments of any IgG1 and / or IgG2 modified Fc, the Fc comprises amino acid substitutions at positions E430G, K322A, A330S, and P331S according to the EU numbering. In some embodiments of any IgG1 and / or IgG2 modified Fc, the Fc comprises amino acid substitutions at positions E430G, K322A, and A330S according to the EU numbering. In some embodiments of any IgG1 and / or IgG2 modified Fc, the Fc comprises amino acid substitutions at positions E430G, K322A, and P331S according to the EU numbering. In some embodiments of any IgG1 and / or IgG2 modified Fc, the Fc comprises amino acid substitutions at positions S267E and L328F according to the EU numbering. In some embodiments of any IgG1 and / or IgG2 modified Fc, the Fc comprises amino acid substitutions at positions C127S according to the EU numbering. In some embodiments of any IgG1 and / or IgG2 modified Fc, the Fc comprises amino acid substitutions at positions E345R, E430G, and S440Y according to the EU numbering.

[0150] In some embodiments of any of the antibodies provided herein, the modified antibody Fc is an IgG4 modified Fc. In some embodiments, the IgG4 modified Fc comprises one or more modifications. For example, in some embodiments, the IgG4 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 IgG4 modified Fc, one or more amino acid substitutions 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 rules. In some embodiments of any IgG4 modified Fc, the Fc may further comprise L235A, G237A, and E318A according to EU numbering rules. In some embodiments of any IgG4 modified Fc, the Fc may further comprise S228P and L235E according to the EU numbering convention. In some embodiments of any IgG4 modified Fc, the IgG4 modified Fc may further comprise S267E and L328F according to the EU numbering convention.

[0151] In some embodiments of any IgG4 modified Fc, the IgG4 modified Fc can be combined with one or more mutations described in (Angal et al., Mol Immunol. 30: 105-108 (1993)) and / or (Peters et al., J Biol Chem. 287 (29): 24525-33 (2012)) according to the EU numbering convention to enhance antibody stabilization.

[0152] In some embodiments of any IgG4 modified Fc, the IgG4 modified Fc may further comprise one or more mutations to enhance the half-life of the antibody in human serum (e.g., one or more (including all) of the M252Y, S254T, and T256E mutations according to the EU numbering convention).

[0153] In some embodiments of any IgG4 modified Fc, the Fc comprises L235E according to EU numbering. In certain embodiments of any IgG4 modified Fc, the Fc comprises one or more amino acid substitutions at a residue position 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 IgG4 modified Fc, the Fc comprises amino acid substitutions at positions E430G, L243A, L235A, and P331S according to EU numbering. In some embodiments of any IgG4 modified Fc, the Fc comprises amino acid substitutions at positions E430G and P331S according to EU numbering. In some embodiments of any IgG4 modified Fc, the Fc comprises amino acid substitutions at positions E430G and K322A according to EU numbering. In some embodiments of any IgG4 modified Fc, the Fc comprises an amino acid substitution at position E430 according to the EU numbering. In some embodiments of any IgG4 modified Fc, the Fc region comprises amino acid substitutions at positions E430G and K322A according to the EU numbering. In some embodiments of any IgG4 modified Fc, the Fc comprises amino acid substitutions at positions S267E and L328F according to the EU numbering. In some embodiments of any IgG4 modified Fc, the Fc comprises amino acid substitutions at position C127S according to the EU numbering. In some embodiments of any IgG4 modified Fc, the Fc comprises amino acid substitutions at positions E345R, E430G, and S440Y according to the EU numbering. (8) Other Antibody Modifications

[0154] In some embodiments of any antibody, the antibody is a derivative. The term "derivative" refers to a molecule that includes chemical modifications other than insertion, deletion, or substitution of amino acids (or nucleic acids). In certain embodiments, the derivative comprises a covalent modification, including but not limited to chemical bonds with polymers, lipids, or other organic or inorganic moieties. In certain embodiments, the chemically modified antigen binding protein may have a longer circulating half-life than the antigen binding protein that has not been chemically modified. In certain embodiments, the chemically modified antigen binding protein may have improved targeting capabilities to desired cells, tissues, and / or organs. In some embodiments, the derivative antigen binding protein is covalently modified to include 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 (either 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.

[0155] In certain embodiments, the derivatives are covalently modified with polyethylene glycol (PEG) subunits. In certain embodiments, one or more water-soluble polymers are bonded at one or more specific positions of the derivative, such as at the amino terminus. 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. Certain such methods are discussed, for example, in U.S. Patent No. 6,133,426, which is hereby incorporated by reference for any purpose.

[0156] Peptide analogs are often used in the pharmaceutical industry as non-peptide drugs with properties similar to those of the template peptide. These types of non-peptide compounds are called peptide mimetics ("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 often developed with the aid of computer molecular modeling. Peptide mimetics that are structurally similar to therapeutically useful peptides can be used to produce similar therapeutic or preventive effects. In general, a peptide mimetic is structurally similar to an exemplary polypeptide (i.e., a polypeptide having biochemical properties or pharmacological activity), such as a human antibody, but one or more peptide linkages are optionally replaced by methods known in the art with linkages selected from the group consisting of: -CH2NH-, -CH2S-, -CH2-CH2-, -CH=CH- (cis and trans), -COCH2-, -CH(OH)CH2-, and -CH2SO-. In certain embodiments, systematic replacement of one or more amino acids of a consensus sequence with D-amino acids of the same type (e.g., D-lysine in place of L-lysine) can be used 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.

[0157] Drug conjugation involves coupling a biologically active cytotoxic (anticancer) payload or drug to an antibody that specifically targets certain tumor markers (e.g., ideally, a polypeptide found only in or on tumor cells). The antibodies track these proteins in the body 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 takes up or internalizes the antibody along with the cytotoxin. After ADC internalization, the cytotoxic drug is released and kills the cancer. Because of this targeting, the drug ideally has lower side effects than other chemotherapeutic agents and gives a wider therapeutic window. The art discloses and knows the technology of conjugating antibodies (see, e.g., Jane de Lartigue OncLive July 5, 2012; ADC Review on antibody-drug conjugates; and Ducry et al., Bioconjugate Chemistry 21(1):5-13 (2010)). II. Antibody Activity

[0158] In some embodiments, anti-SIRPβ1 antibodies that bind to human SIRPβ1 isoform 1 but not to human SIRPα are provided. In some embodiments, anti-SIRPβ1 antibodies that bind to human SIRPβ1 isoform 1 but not to human SIRPγ are provided. In some embodiments, anti-SIRPβ1 antibodies that bind to human SIRPβ1 isoform 1 but not to human SIRPβ1 isoform 3 are provided. In some embodiments, anti-SIRPβ1 antibodies that bind to human SIRPβ1 isoform 1 but not to mouse SIRPβ1 are provided. In some embodiments, anti-SIRPβ1 antibodies that bind to human SIRPβ1 isoform 1 but not to cynomolgus monkey SIRPβ1 are provided. In some embodiments, anti-SIRPβ1 antibodies that bind to human SIRPβ1 isoform 1 but not to cynomolgus monkey SIRPβ1 are provided. In some embodiments, anti-SIRPβ1 antibodies that bind to human SIRPβ1 isoform 1 but not to cynomolgus monkey SIRPβ1 isoform 1 are provided.

[0159] In some embodiments, ForteBio can be used The binding of anti-SIRPβ1 antibodies to antigen is determined using the Red384 system (ForteBio, Menlo Park, CA), for example as described in Example 1. Antibody binding can be determined using, for example, Fab fragments of the antibody (for monovalent binding) or full-length antibodies such as IgG (for divalent binding, or avidity). An exemplary binding assay for a full-length antibody using the Red384 system is as follows. The antibody is loaded onto an AHQ sensor. The loaded antibody is then exposed to the antigen, and the dissociation rate is measured in an assay buffer at multiple time intervals (such as 3 minutes). The binding model can then be used to fit the kinetic data in the data analysis software provided with the system. For antibody Fab fragment affinity measurements, in some embodiments, the antigen-Fc can be loaded onto the AHQ sensor and then exposed to the antibody Fab fragment. The dissociation rate is measured as above, and the data is analyzed using the software provided with the system.

[0160] In some embodiments, anti-SIRPβ1 antibodies are provided that agonize SIRPβ1 activity on CD14-positive monocytes in vitro and / or in vivo. In some embodiments, anti-SIRPβ1 antibodies are provided that agonize SIRPβ1 activity on CD14-positive monocytes in vitro. In some embodiments, anti-SIRPβ1 antibodies are provided that agonize SIRPβ1 activity on CD14-positive monocytes in vivo. In some embodiments, anti-SIRPβ1 antibodies agonize SIRPβ1 activity on CD14-positive monocytes in vitro in an assay in which the anti-SIRPβ1 antibodies are immobilized on a solid support, such as a culture plate, or bound by an anti-IgG secondary antibody, or bound by an Fcγ receptor on an auxiliary cell. In some embodiments, anti-SIRPβ1 antibodies are provided that agonize SIRPβ1 activity on CD14-positive monocytes in vivo. A non-limiting exemplary assay for determining whether an anti-SIRPβ1 antibody agonizes SIRPβ1 activity on CD14-positive monocytes in vitro is described in Examples 3 and 4. For example, isolated monocytes are starved for a period of time, such as 4 hours, and then incubated with anti-SIRPβ1 antibodies, for example in the presence of anti-IgG antibodies. After incubation, the cells are lysed and the phosphorylation of one or more of Syk, ERK, AKT, SIRPβ1, and / or DAP12 is measured, for example using anti-phosphotyrosine and / or anti-phosphoserine antibodies. An increase in the phosphorylation of one or more of Syk, ERK, AKT, SIRPβ1, and / or DAP12 indicates an increase in SIRPβ1 activity (i.e., an agonist of SIRPβ1 activity). As another example, monocytes can be incubated with anti-SRPβ1 antibodies in the presence of auxiliary cells expressing Fcγ receptors, such as B cells. After incubation, the cells are lysed and the phosphorylation of one or more of Syk, ERK, AKT, SIRPβ1, and / or DAP12 is determined as above. An increase in phosphorylation of one or more of Syk, ERK, AKT, SIRPβ1, and / or DAP12 indicates an increase in SIRPβ1 activity (i.e., the anti-SIRPβ1 antibody is an agonist of SIRPβ1 activity). A non-limiting exemplary assay for determining whether the anti-SIRPβ1 antibody agonizes SIRPβ1 activity on CD14-positive monocytes in vivo comprises administering the anti-SIRPβ1 antibody to human SIRPβ1 BAC transgenic C57BL / 6 mice, and isolating CD14-positive monocytes, for example, by FACS. The monocytes are then lysed and the phosphorylation of one or more of Syk, ERK, AKT, SIRPβ1, and / or DAP12 is measured as described above. An increase in phosphorylation of one or more of Syk, ERK, AKT, SIRPβ1, and / or DAP12 indicates an increase in SIRPβ1 activity (i.e., an agonist of SIRPβ1 activity).

[0161] In some embodiments, anti-SIRPβ1 antibodies are provided that induce or increase the respiratory burst in immune cells, such as neutrophils and / or monocytes, in vitro and / or in vivo. In some embodiments, anti-SIRPβ1 antibodies are provided that induce or increase the respiratory burst in immune cells, such as neutrophils and / or monocytes, in vitro. In some embodiments, anti-SIRPβ1 antibodies that induce or increase the respiratory burst in immune cells, such as neutrophils and / or monocytes, in vivo are provided. Non-limiting exemplary in vitro assays for determining whether anti-SIRPβ1 antibodies induce or increase the respiratory burst in immune cells, such as neutrophils and / or monocytes, are described in Examples 5 and 6. For example, primary neutrophils are contacted with anti-SIRPβ1 antibodies and the generation of reactive oxygen species (ROS) is detected using universal oxidative stress indicators, such as the fluorescent dye CM-H2DCFDA. Alternatively, an assay can be performed using an anti-SIRPβ1 antibody that is immobilized on a solid support, such as a culture plate, or bound by an anti-IgG secondary antibody, or bound by an Fcγ receptor on an auxiliary cell.

[0162] In some embodiments, anti-SIRPβ1 antibodies are provided that induce or increase IL-8 expression in monocytes in vitro and / or in vivo. In some embodiments, anti-SIRPβ1 antibodies are provided that induce or increase IL-8 expression in monocytes in vitro. In some embodiments, anti-SIRPβ1 antibodies are provided that induce or increase IL-8 expression in monocytes in vivo. A non-limiting exemplary in vitro assay for determining whether anti-SIRPβ1 antibodies induce or increase IL-8 expression in monocytes is described in Example 5. For example, primary monocytes can be stimulated with an anti-SIRPβ1 antibody that is immobilized on a solid support, such as a culture plate, or bound to an anti-IgG secondary antibody, or bound to an Fcγ receptor on an auxiliary cell, for example overnight. The supernatant can be collected to measure IL-8 release. A non-limiting exemplary assay for determining whether anti-SIRPβ1 antibodies induce or increase IL-8 expression in monocytes in vivo comprises administering anti-SIRPβ1 antibodies to human SIRPβ1 BAC transgenic C57BL / 6 mice and obtaining one or more blood samples. Serum concentrations of IL-8 can be determined using commercial assays, such as the Duoset ELISA kit (R&D Systems).

[0163] In some embodiments, anti-SIRPβ1 antibodies are provided that induce or increase TNFα expression in macrophages and / or dendritic cells in vitro and / or in vivo. In some embodiments, anti-SIRPβ1 antibodies are provided that induce or increase TNFα expression in macrophages and / or dendritic cells in vitro. In some embodiments, anti-SIRPβ1 antibodies are provided that induce or increase TNFα expression in macrophages and / or dendritic cells in vivo. A non-limiting exemplary in vitro assay for determining whether an anti-SIRPβ1 antibody induces or increases TNFα expression is described in Example 6. For example, monocyte-derived macrophages or dendritic cells can be stimulated with LPS in the presence of an anti-SIRPβ1 antibody that is immobilized on a solid support, such as a culture plate, or bound by an anti-IgG secondary antibody, or bound by an Fcγ receptor on an accessory cell, for example, overnight. The supernatant can be collected to determine TNFα release. A non-limiting exemplary assay for determining whether an anti-SIRPβ1 antibody induces or increases TNFα expression in macrophages and / or dendritic cells in vivo comprises administering an anti-SIRPβ1 antibody to human SIRPβ1 BAC transgenic C57BL / 6 mice along with LPS (such as 5 μg LPS / mouse). The mice are then sacrificed, for example, by CO2 asphyxiation and the peritoneal fluid is recovered and clarified by centrifugation. TNF-α concentrations can be determined using commercial assays, such as the Duoset ELISA kit (R&D Systems).

[0164] In some embodiments, anti-SIRPβ1 antibodies are provided that induce or increase neutrophil-mediated phagocytosis, such as phagocytosis of tumor cells, in vitro and / or in vivo. In some embodiments, anti-SIRPβ1 antibodies are provided that induce or increase neutrophil-mediated phagocytosis, such as phagocytosis of tumor cells in vitro. In some embodiments, anti-SIRPβ1 antibodies are provided that induce or increase neutrophil-mediated phagocytosis, such as phagocytosis of tumor cells in vivo. In some embodiments, anti-SIRPβ1 antibodies that induce or increase neutrophil-mediated phagocytosis, such as phagocytosis of tumor cells in vivo are provided. In some embodiments, anti-SIRPβ1 antibodies that induce or increase neutrophil-mediated tumor cell clearance in vivo are provided. A non-limiting exemplary in vitro assay for determining whether an anti-SIRPβ1 antibody induces or increases neutrophil-mediated phagocytosis is described in Example 7. For example, primary neutrophils are contacted with an anti-SIRPβ1 antibody that is immobilized on a solid support, such as a culture plate, or bound to an anti-IgG secondary antibody, or bound to an Fcγ receptor on an auxiliary cell. Cancer cells engineered to express luciferase, such as Raji B cell lymphoma cells, are co-cultured with neutrophils and immobilized anti-SIRPβ1 antibodies in the presence of opsonic antibodies, such as anti-CD20 antibodies. Viable Raji cells are quantified by measuring luciferase activity. A reduction in viable Raji cells in the presence of anti-SIRPβ1 antibodies compared to IgG control antibodies indicates that the anti-SIRPβ1 antibodies induce or enhance neutrophil-mediated phagocytosis. A non-limiting exemplary assay to determine whether anti-SIRPβ1 antibodies induce or enhance neutrophil-mediated tumor cell clearance in vivo comprises intravenously injecting B16F10 melanoma cells expressing GFP into human SIRPβ1 BAC transgenic mice. The mice are then treated with anti-SIRPβ1 antibodies together with opsonic anti-gp75 antibodies. Peripheral blood is collected, red blood cells are lysed, and white blood cells are resuspended in FACS buffer. Neutrophils are stained with anti-CD11b and anti-Ly6G antibodies and analyzed by flow cytometry for green fluorescent signals acquired from ingested tumor cells. An increase in green fluorescence signal in neutrophils compared to the same experiment with an isotype-matched control antibody indicates that the anti-SIRPβ1 antibody induces or enhances neutrophil-mediated phagocytosis.

[0165] Another non-limiting exemplary assay to determine whether anti-SIRPβ1 antibodies induce or enhance neutrophil-mediated tumor cell clearance in vivo uses a lung metastasis model, such as the following. Fc receptor γ chain-deficient C57BL / 6 mice lacking expression of activating FcγRs are intravenously injected with B16F10 melanoma cells. Bone marrow neutrophils isolated from human SIRPβ1 BAC transgenic mice are then intravenously injected with opsonic anti-gp75 antibodies in combination with anti-SIRPβ1 antibodies. After a period of time, the mice are euthanized and the lungs are harvested and fixed. The number of metastatic tumor nodules with dark pigmentation is visually counted. A reduction in the number of metastatic tumor nodules compared to the same experiment with an isotype-matched control antibody indicates that the anti-SIRPB1 antibody induces or enhances neutrophil-mediated tumor cell clearance in vivo.

[0166] In some embodiments, anti-SIRPβ1 antibodies that increase TREM2 expression on macrophages in vitro and / or in vivo are provided. In some embodiments, anti-SIRPβ1 antibodies that increase TREM2 expression on macrophages in vitro are provided. In some embodiments, anti-SIRPβ1 antibodies that increase TREM2 expression on macrophages in vivo are provided. A non-limiting exemplary in vitro assay for determining whether anti-SIRPβ1 antibodies increase TREM2 expression on macrophages is described in Example 16. For example, monocyte-derived macrophages differentiated in culture with M-CSF are incubated with anti-SIRPβ1 antibodies that are immobilized on a solid support, such as a culture plate, or bound to an anti-IgG secondary antibody, or bound to an Fcγ receptor on an accessory cell. TREM2 expression on macrophages is analyzed, for example, by FACS analysis. A non-limiting exemplary assay for determining whether anti-SIRPβ1 antibodies increase TREM2 expression on macrophages in vivo is as follows. Human SIRPβ1 BAC transgenic C57BL / 6 mice are challenged with aged sterile thioglycolate medium injected into the peritoneum. After challenge, mice are treated with anti-SIRPβ1 antibodies. Mice are then sacrificed and peritoneal macrophages are harvested after euthanasia, for example by flushing the peritoneal cavity with PBS. Macrophages are defined as CD11b+Ly6C-F4 / 80+ cells and can be analyzed by FACS to determine TREM2 expression levels.

[0167] In some embodiments, anti-SIRPβ1 antibodies are provided that increase the viability of macrophages in vitro and / or in vivo when alone and / or in combination with agonistic anti-TREM2 antibodies. In some embodiments, anti-SIRPβ1 antibodies are provided that increase the viability of macrophages in vitro when alone and / or in combination with agonistic anti-TREM2 antibodies. In some embodiments, anti-SIRPβ1 antibodies are provided that increase the viability of macrophages in vivo when alone and / or in combination with agonistic anti-TREM2 antibodies. In some embodiments, anti-SIRPβ1 antibodies are provided that increase the viability of dendritic cells in vitro and / or in vivo. In some embodiments, anti-SIRPβ1 antibodies are provided that increase the viability of dendritic cells in vitro. In some embodiments, anti-SIRPβ1 antibodies are provided that increase the viability of dendritic cells in vivo. Non-limiting exemplary in vitro assays for determining whether anti-SIRPβ1 antibodies increase the viability of macrophages in vitro when alone or in combination with agonistic anti-TREM2 antibodies are described in Examples 17, 18, and 19. For example, monocyte-derived macrophages differentiated in culture with M-CSF are incubated with anti-SIRPβ1 antibodies immobilized on a solid support, such as a culture plate, or bound by an anti-IgG secondary antibody, or bound by an Fcγ receptor on an accessory cell, and incubated with or without an anti-TREM2 antibody. Cell viability is measured, for example, using the Cell Titer Glo kit (Promega), a reagent that generates a luminescent signal relative to the ATP concentration in the sample. A similar assay can be used to determine whether anti-SIRPβ1 antibodies increase the viability of bone marrow-derived macrophages or dendritic cells in vitro. For this assay, bone marrow cells are cultured with M-CSF (for differentiating macrophages) or GM-CSF (for differentiating dendritic cells). Macrophages or dendritic cells are incubated with anti-SIRPβ1 antibodies immobilized on a solid support, such as a culture plate, or bound by an anti-IgG secondary antibody, or bound by an Fcγ receptor on an accessory cell, and incubated with or without an anti-TREM2 antibody. As above, cell viability is measured, for example, using the Cell Titer Glo kit (Promega), a reagent that generates a luminescent signal relative to the ATP concentration in the sample. A non-limiting exemplary assay to determine whether anti-SIRPβ1 antibodies increase the viability of macrophages in vivo is as follows. Due to the elimination of M-CSF-mediated survival signals, prolonged treatment with blocking antibodies to CSF1R depletes peritoneal and tissue-resident F4 / 80+ macrophages. For this assay, anti-CSF1R antibodies are administered to human SIRPβ1 BAC transgenic C57BL / 6 mice along with anti-SIRPβ1 antibodies. After treatment, mice are sacrificed, for example, by CO2 asphyxiation, and peripheral blood is collected by cardiac puncture. Peritoneal macrophages are collected by lavage with PBS.Macrophage populations were enumerated by FACS by gating on CD11b+Ly6C-F4 / 80+ cells in blood and peritoneum. III. Nucleic Acids, Vectors, and Host Cells

[0168] Recombinant methods and compositions can be used to produce the anti-SIRPβ1 antibodies of the present disclosure, for example, as described in U.S. Patent No. 4,816,567. In some embodiments, isolated nucleic acids having nucleotide sequences encoding any of the anti-SIRPβ1 antibodies of the present disclosure are provided. Such nucleic acids may encode V sequences comprising the anti-SIRPβ1 antibodies. 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., via transduction): (1) a V-encoding region encoding an antibody; L The amino acid sequence of the antibody V H or (2) a vector containing a nucleic acid encoding an antibody V L The first vector comprises a nucleic acid encoding an amino acid sequence of an antibody V H In some embodiments, the host cell is a eukaryotic cell, such as a Chinese hamster ovary (CHO) cell or a lymphoid cell (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.

[0169] Methods for producing anti-SIRPβ1 antibodies of the present disclosure are provided. In some embodiments, the method comprises culturing a host cell of the present disclosure comprising a nucleic acid encoding an anti-SIRPβ1 antibody under conditions suitable for antibody expression. In some embodiments, the antibody is subsequently recovered from the host cell (or host cell culture medium).

[0170] In order to recombinantly produce the anti-SIRPβ1 antibodies disclosed herein, nucleic acids encoding anti-SIRPβ1 antibodies are isolated and inserted into one or more vectors for further cloning and / or expression in host cells. Such nucleic acids can be easily isolated and sequenced using conventional procedures (e.g., by using oligonucleotide probes that can specifically bind to genes encoding the heavy and light chains of antibodies).

[0171] Suitable vectors containing nucleic acid sequences encoding any anti-SIRPβ1 antibody of the present disclosure, or polypeptides (including antibodies) described herein, or cell surface expression fragments thereof 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 wide variety of cloning vectors available in the art. Although the selected cloning vector may vary according to the intended host cell, useful cloning vectors generally have the ability to replicate themselves, may have a single target for a specific restriction endonuclease, and / or may carry genes for markers 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, mp18, mp19, pBR322, pMB9, ColE1, pCR1, RP4, phage DNA, and shuttle vectors, such as pSA3 and pAT28. These and many other cloning vectors are available from suppliers such as BioRad, Strategene, and Invitrogen.

[0172] Suitable host cells for cloning or expressing antibody encoding vectors include prokaryotic or eukaryotic cells. For example, the anti-SIRPβ1 antibodies disclosed herein can be produced in bacteria, especially when glycosylation and Fc effector functions are not required. For expressing antibody fragments and polypeptides in bacteria (e.g., U.S. Pat. Nos. 5,648,237, 5,789,199, and 5,840,523. After expression, the antibody can be isolated from the bacterial cell mass in a soluble fraction and can be further purified.

[0173] In addition to prokaryotes, eukaryotic microorganisms such as filamentous fungi or yeast are also suitable cloning or expression hosts for antibody-encoding vectors, including fungi and yeast strains whose glycosylation pathways have been "humanized" to result in the production of antibodies with partially or fully human glycosylation patterns (e.g., Gerngross, Nat. Biotech. 22:1409-1414 (2004); and Li et al., Nat. Biotech. 24:210-215 (2006)).

[0174] Host cells suitable for expressing glycosylated antibodies can also be derived from multicellular organisms (invertebrates and vertebrates). Examples of invertebrate cells include plant and insect cells. Many baculovirus strains have been identified that can be used with insect cells, particularly for transfecting Spodoptera frugiperda cells. Plant cell cultures can also be used as hosts (e.g., U.S. Pat. Nos. 5,959,177, 6,040,498, 6,420,548, 7,125,978 and 6,417,429, which describe the PLANTIBODIES™ technology for producing antibodies in transgenic plants).

[0175] Vertebrate cells can also be used as hosts. For example, mammalian cell lines adapted to grow in suspension can be useful. Other examples of useful mammalian host cell lines are monkey kidney CV1 line (COS-7) transformed by SV40; human embryonic kidney line (293 or 293 cells, as described, for example, in Graham et al., J. Gen Virol. 36:59 (1977)); baby hamster kidney cells (BHK); mouse Sertoli cells (TM4 cells, as described, for example, in Mather, Biol. Reprod. 23:243-251 (1980)); monkey kidney cells (CV1); African green monkey kidney cells (VERO-76); human cervical carcinoma 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 N.Y. 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, e.g., Yazaki and Wu, Methods in Molecular Biology, Vol. 248 (BKCLo, ed., Humana Press, Totowa, NJ), pp. 255-268 (2003). IV. Pharmaceutical Compositions / Formulations

[0176] Provided herein are pharmaceutical compositions and / or pharmaceutical formulations comprising an anti-SIRPβ1 antibody of the present disclosure and a pharmaceutically acceptable carrier.

[0177] In some embodiments, the pharmaceutically acceptable carrier is preferably non-toxic to the recipient at the dose and concentration used. The antibodies described herein can be formulated into solid, semi-solid, liquid or gaseous preparations. Examples of such formulations include, but are not limited to, tablets, capsules, powders, granules, ointments, solutions, suppositories, injections, inhalants, gels, microspheres, and aerosols. Depending on the desired formulation, the pharmaceutically acceptable carrier may include a pharmaceutically acceptable non-toxic carrier diluent, which is a medium often used to prepare pharmaceutical compositions for animal or human administration. In certain embodiments, the pharmaceutical composition may include a formulation material for changing, maintaining or maintaining, for example, the pH, osmoticity, viscosity, clarity, color, isotonicity, odor, sterility, stability, dissolution or release, absorption or penetration rate of the composition.

[0178] 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-HCl, citrates, phosphates or other organic acids); bulking agents (such as mannitol or glycine); chelating agents (such as ethylenediaminetetraacetic acid (EDTA)); complexing agents (such as caffeine, polyvinyl pyrrolidone, β-cyclodextrin or hydroxypropyl β-cyclodextrin); fillers; monosaccharides; disaccharides; and other carbohydrates (such as glucose, mannose or dextrin); proteins (such as serum albumin, gelatin or immunoglobulins); colorants, flavorings and diluents; emulsifiers; hydrophilic polymers (such as polyvinylpyrrolidone); vinyl pyrrolidone); low molecular weight polypeptides; salt-forming counterions (such as sodium); preservatives (such as benzalkonium chloride, benzoic acid, salicylic acid, thimerosal, phenylethyl alcohol, methylparaben, propylparaben, chlorhexidine, sorbic acid or hydrogen peroxide); solvents (such as glycerol, propylene glycol or polyethylene glycol); sugar alcohols (such as mannitol or sorbitol); suspending agents; surfactants or wetting agents (such as Pluronics, PEG, sorbitan esters, polysorbates such as polysorbate 20, polysorbate 80, Triton, tromethamine, lecithin, cholesterol, tyloxapol); stability enhancers (such as sucrose or sorbitol); tonicity enhancers (such as alkali metal halides, preferably sodium or potassium chloride, mannitol sorbitol); delivery vehicles; diluents; excipients and / or pharmaceutical adjuvants. Additional examples of formulations suitable for various types of administration can be found in Remington: The Science and Practice of Pharmacy, Pharmaceutical Press 22nd ed. (2013). For a brief review of methods for drug delivery, see Langer, Science 249: 1527-1533 (1990).

[0179] Formulations suitable for parenteral administration include aqueous and non-aqueous isotonic sterile injection solutions which may contain antioxidants, buffers, bacteriostats, and solutes to render the formulation isotonic with the blood of the intended recipient, and aqueous and non-aqueous sterile suspensions which may include suspending agents, solubilizers, thickening agents, stabilizers, and preservatives.

[0180] The formulation can be optimized for retention and stability in the brain or central nervous system. When the agent is administered into the cranial compartment, it is desirable that the agent remain in the compartment rather than diffuse or otherwise cross the blood-brain barrier. Stabilization techniques include cross-linking, polymerizing, or linking to groups such as polyethylene glycol, polyacrylamide, neutral protein carriers, etc. to achieve an increase in molecular weight.

[0181] Other strategies for increasing retention include embedding antibodies (such as the anti-SIRPβ1 antibodies of the present disclosure) in biodegradable or bioerodible implants. The release rate of the therapeutically active agent is controlled by the transport rate through the polymer matrix and the biodegradation of the implant. The implant can be a particle, a sheet, a patch, a plaque, a fiber, a microcapsule, etc., and can be any size or shape compatible with the selected insertion site. The biodegradable polymer composition that can be used can be an organic ester or ether, which produces physiologically acceptable degradation products when degraded, including monomers. Anhydrides, amides, orthoesters, etc. can be used, either independently or in combination with other monomers. The polymer will be a condensation polymer. The polymer can be cross-linked or non-cross-linked. Of particular interest are polymers of hydroxyaliphatic carboxylic acids (homopolymers or copolymers) and polysaccharides. Included among the polyesters of interest are polymers of D-lactic acid, L-lactic acid, racemic lactic acid, glycolic acid, polycaprolactone, and combinations thereof. Among the polysaccharides of interest are calcium alginate and functionalized cellulose, particularly carboxymethyl cellulose esters, characterized by being insoluble in water, having a molecular weight of about 5 kD to 500 kD, etc. Biodegradable hydrogels may also be employed in the implants of the present invention. Hydrogels are typically copolymer materials characterized by being able to absorb liquids. V. Therapeutic Uses

[0182] As disclosed herein, the anti-SIRPβ1 antibodies of the present disclosure can be used to prevent, reduce risk, or treat diseases and disorders. In various embodiments, the anti-SIRPβ1 antibodies provided herein have one or more activities selected from the following: stimulating SIRPβ1 activity on CD14-positive monocytes in vitro and / or in vivo; inducing immune cells, such as respiratory bursts in neutrophils and / or monocytes in vitro and / or in vivo; inducing IL-8 expression in monocytes in vitro and / or in vivo; inducing TNFα expression in macrophages and / or dendritic cells in vitro and / or in vivo; inducing neutrophil-mediated phagocytosis, such as phagocytosis of tumor cells in vitro and / or in vivo; improving neutrophil-mediated tumor cell clearance in vivo; upregulating TREM2 expression on macrophages in vitro and / or in vivo; improving the viability of macrophages in vitro and / or in vivo alone and / or in combination with agonistic anti-TREM2 antibodies; and improving the viability of dendritic cells in vitro and / or in vivo.

[0183] In various embodiments, the anti-SIRPβ1 antibodies provided herein can be used to prevent, reduce the risk, or treat dementia, frontotemporal dementia, Alzheimer's disease, vascular dementia, mixed dementia, Creutzfeldt-Jakob disease, normal pressure hydrocephalus, amyotrophic lateral sclerosis, Huntington's disease, Tau disease, Nasu-Hakola disease, stroke, acute trauma, chronic trauma, lupus, acute and chronic colitis, wound healing, Crohn's disease, inflammatory bowel disease, ulcerative colitis, obesity, malaria, essential tremor, central nervous system lupus, Behemoth disease, Chet's disease, Parkinson's disease, dementia with Lewy bodies, multiple system atrophy, Shy-Drager syndrome, progressive supranuclear palsy, corticobasal ganglionic degeneration, acute disseminated encephalomyelitis, granulomatous disorders, sarcoidosis, diseases of aging, seizures, spinal cord injury, traumatic brain injury, age-related macular degeneration, glaucoma, retinitis pigmentosa, retinal degeneration, respiratory tract infection, sepsis, eye infection, systemic infection, lupus, arthritis, multiple sclerosis, low bone density, osteoporosis, osteogenesis, osteosclerotic disease, Paget's disease of bone, and / or cancer. In some such embodiments, the anti-SIRPβ1 antibody is an agonistic antibody.

[0184] In some embodiments, provided herein are methods of preventing, reducing the risk of, or treating a disorder having dementia, frontotemporal dementia, Alzheimer's disease, vascular dementia, mixed dementia, Creutzfeldt-Jakob disease, normal pressure hydrocephalus, amyotrophic lateral sclerosis, Huntington's disease, tau diseases, Nasu-Hakola disease, stroke, acute trauma, chronic trauma, lupus, acute and chronic colitis, wound healing, Crohn's disease, inflammatory bowel disease, ulcerative colitis, obesity, malaria, essential tremor, central nervous system lupus, Behcet's disease, Parkinson's disease, dementia with Lewy bodies, multiple system atrophy, A method of treating an individual with Schiele-Derrickson syndrome, progressive supranuclear palsy, corticobasal ganglionic degeneration, acute disseminated encephalomyelitis, granulomatous disorders, sarcoidosis, diseases of aging, epileptic seizures, spinal cord injury, traumatic brain injury, age-related macular degeneration, glaucoma, retinitis pigmentosa, retinal degeneration, respiratory tract infection, sepsis, eye infection, systemic infection, lupus, arthritis, multiple sclerosis, low bone density, osteoporosis, osteogenesis, osteosclerotic disease, Paget's disease of bone, and / or cancer by administering to the individual a therapeutically effective amount of an anti-SIRPβ1 antibody of the present disclosure. In some such embodiments, the anti-SIRPβ1 antibody is an agonist that (e.g., induces or increases) SIRPβ1 activity.

[0185] As disclosed herein, the anti-SIRPβ1 antibodies of the present disclosure can also be used to induce and / or promote the survival of innate immune cells. In some embodiments, the present disclosure provides methods of inducing or promoting the survival of innate immune cells in an individual in need thereof by administering to the individual a therapeutically effective amount of an agonistic anti-SIRPβ1 antibody of the present disclosure.

[0186] As disclosed herein, the anti-SIRPβ1 antibodies of the present disclosure can also be used to induce and / or promote wound healing, such as after injury. In some embodiments, wound healing can be colonic wound repair after injury. In some embodiments, the present disclosure provides a method of inducing or promoting wound healing in an individual in need thereof by administering to the individual a therapeutically effective amount of an agonistic anti-SIRPβ1 antibody of the present disclosure.

[0187] In some embodiments, the subject or individual is a mammal. Mammals include, but are not limited to, domesticated animals (e.g., cattle, sheep, cats, dogs, and horses), primates (e.g., humans and non-human primates, such as monkeys), rabbits, and rodents (e.g., mice and rats). In some embodiments, the subject or individual is a human.

[0188] The antibodies (and any additional therapeutic agents) provided herein can be administered by any suitable means, including parenteral, intrapulmonary, intranasal, intralesional administration, intracerebrospinal, intracranial, intraspinal, intrasynovial, intrathecal, oral, topical, or inhalation routes. Parenteral infusions include intramuscular, intravenous administration (as a push or by continuous infusion over a period of time), intraarterial, intraarticular, intraperitoneal, or subcutaneous administration. In some embodiments, administration is intravenous administration. In some embodiments, administration is subcutaneous. Dosage administration can be by any suitable route, for example by injection, such as intravenous or subcutaneous injection, depending in part on whether the administration is short-lived or long-term. Various dosage administration schedules are contemplated herein, including but not limited to multiple administrations at single or multiple time points, push administration, and pulse infusions.

[0189] The antibodies provided herein will be formulated, dosed, and administered in a manner consistent with good medical practice. Factors considered in this context include the specific condition being treated, the specific mammal being treated, the clinical condition of the individual patient, the cause of the condition, the delivery site of the agent, the method of administration, the schedule of administration, and other factors known to medical practitioners. The antibody need not but is optionally formulated with one or more agents currently used to prevent or treat the condition in question. The effective amount of such other agents depends on the amount of antibodies present in the formulation, the type of condition or treatment, and other factors discussed above. These are generally used in the same dosage and administration route as described herein, or about 1 to 99% of the dosage described herein, or any dosage and any route determined to be suitable empirically / clinically.

[0190] For the prevention or treatment of disease, the appropriate dosage of the antibodies of the invention (when used alone or in combination with one or more other additional therapeutic agents) will depend on the type of disease to be treated, the type of antibody, the severity and course of the disease, whether the antibody is administered for preventive or therapeutic purposes, previous therapy, the patient's clinical history and response to the antibody, and the discretion of the attending physician. The antibody is suitably administered to the patient at one time or over a series of treatments.

[0191] Depending on the type and severity of the disease, about 1 μg / kg to 15 mg / kg (e.g., 0.1 mg / kg-10 mg / kg) of antibody may be an initial candidate dose for administration to a patient, for example, either by one or more separate administrations, or by continuous infusion. A typical daily dosage range may be about 1 μg / kg to 100 mg / kg or more, depending on the factors mentioned above. For repeated administration over several days or longer, depending on the condition, treatment generally continues until the desired disease symptom suppression occurs. An exemplary dose of the antibody will be in the range of about 0.05 mg / kg to about 10 mg / kg. Thus, one or more doses of about 0.5 mg / kg, 2.0 mg / kg, 4.0 mg / kg, or 10 mg / kg (or any combination thereof) may be administered to the patient. Such doses may be administered intermittently, for example, weekly or every three weeks (e.g., so that the patient receives about 2 to about 20, or, for example, about 6 doses of the antibody). In certain embodiments, the dosing frequency is three times a day, twice a day, once a day, every other day, once a week, once every two weeks, once every four weeks, once every five weeks, once every six weeks, once every seven weeks, once every eight weeks, once every nine weeks, once every ten weeks, or once a month, once every two months, once every three months, or longer. An initial higher loading dose may be administered, followed by one or more lower doses. However, other dosage regimens may be useful. The progress of this therapy is easily monitored by conventional techniques and assays. dementia

[0192] Dementia is a nonspecific syndrome (i.e., a collection of signs and symptoms) characterized by a severe loss of overall cognitive ability in previously unimpaired people, beyond what might be expected based on normal aging. Dementia can be static, as a result of unique global brain damage. Or, dementia can be progressive, resulting from damage or disease in the body, leading to a long-term decline. Although dementia is much more common in older populations, it can also occur before the age of 65. Cognitive areas affected by dementia include, but are not limited to, memory, attention span, language, and problem solving. Generally, symptoms must be present for at least six months before an individual is diagnosed with dementia.

[0193] Exemplary forms of dementia include, but are not limited to, frontotemporal dementia, Alzheimer's disease, vascular dementia, semantic dementia, and dementia with Lewy bodies.

[0194] In some embodiments, administration of an anti-SIRPβ1 antibody of the present disclosure can prevent, reduce the risk of, and / or treat dementia. In some embodiments, administration of an anti-SIRPβ1 antibody can induce one or more SIRPβ1 activities in an individual with dementia. Frontotemporal dementia

[0195] Frontotemporal dementia (FTD) is a condition caused by the gradual deterioration of the frontal lobe of the brain. Over time, degeneration may progress to the temporal lobe. Second only to Alzheimer's disease (AD) in prevalence, FTD accounts for 20% of cases of Alzheimer's disease. Clinical features of FTD include memory deficits, abnormal behavior, personality changes, and language impairment (Cruts, M. & Van Broeckhoven, C., Trends Genet. 24: 186-194 (2008); Neary, D. et al., Neurology 51: 1546-1554 (1998); Ratnavalli, E., Brayne, C., Dawson, K. & Hodges, JR, Neurology 58: 1615-1621 (2002)).

[0196] A significant portion of FTD cases are inherited in an autosomal dominant manner, but even within a family, symptoms can span the range of FTD with behavioral disorders, to primary progressive aphasia, to cortical-basal ganglionic degeneration. Like most neurodegenerative diseases, FTD can be characterized by the pathological presence of specific protein aggregates in the diseased brain. Historically, the first description of FTD identified the presence of intraneuronal accumulation of hyperphosphorylated Tau protein in neurofibrillary tangles or Pick bodies. The identification of mutations in genes encoding Tau protein in several families supports the causative role of microtubule-associated protein Tau (Hutton, M. et al., Nature 393: 702-705 (1998)). However, most FTD brains show no accumulation of hyperphosphorylated Tau, but do exhibit immunoreactivity to ubiquitin (Ub) and TAR DNA binding protein (TDP43) (Neumann, M. et al., Arch. Neurol. 64: 1388-1394 (2007)). Those FTD cases with Ub inclusions (FTD-U) were mostly shown to carry mutations in the progranulin gene.

[0197] In some embodiments, administration of an anti-SIRPβ1 antibody of the present disclosure can prevent, reduce the risk of, and / or treat FTD. In some embodiments, administration of an anti-SIRPβ1 antibody can induce one or more SIRPβ1 activities in an individual with FTD. Alzheimer's disease

[0198] Alzheimer's disease (AD) is the most common form of dementia. The disease has no cure, worsens as it progresses, and ultimately leads to death. Most commonly, AD is diagnosed in people over the age of 65. However, the less prevalent early-onset Alzheimer's can occur much earlier.

[0199] Common symptoms of Alzheimer's disease include behavioral symptoms, such as difficulty remembering recent events; cognitive symptoms, confusion, irritability and aggression, mood swings, trouble speaking, and long-term memory loss. As the disease progresses, physical function is lost, eventually leading to death. Alzheimer's disease develops for an unknown and variable amount of time before becoming fully apparent, and it can progress for years without being diagnosed.

[0200] In some embodiments, administration of an anti-SIRPβ1 antibody of the present disclosure can prevent, reduce the risk of, and / or treat Alzheimer's disease. In some embodiments, administration of an anti-SIRPβ1 antibody can induce one or more SIRPβ1 activities in an individual with Alzheimer's disease. Nasu-Hakola disease

[0201] Nasu-Hakola disease (NHD), alternatively known as polycystic lipomembranous skeletal dysplasia and sclerosing leukoencephalopathy (PLOSL), is a rare hereditary leukodystrophy characterized by progressive precocious dementia associated with recurrent fractures due to polycystic bone lesions of the lower or upper extremities. The NHD disease process generally progresses into four stages: latent, skeletal, early neurological, and late neurological. After normal development during childhood (latent stage), NHD begins to manifest during adolescence or young adulthood (typical age of onset 20-30 years) with pain in the hands, wrists, ankles, and feet. Patients then begin to suffer from recurrent fractures due to polycystic bone and osteoporotic lesions in the bones of the extremities (skeletal stage). During the third or fourth decade of life (early neurological stage), patients present with personality changes (e.g., euphoria, difficulty concentrating, loss of judgment, and social inhibition) characteristic of a prominent frontal lobe syndrome. Patients also typically suffer from progressive memory impairment. Seizures are also frequently observed. In the final (late neurological stage), patients develop severe dementia, become unable to speak and move, and usually die by age 50.

[0202] In some embodiments, administration of the anti-SIRPβ1 antibodies of the present disclosure can prevent, reduce the risk of, and / or treat Nasu-Hakola disease (NHD). In some embodiments, administration of anti-SIRPβ1 antibodies can induce one or more SIRPβ1 activities in individuals with NHD. Parkinson's disease

[0203] Parkinson's disease, which may be called idiopathic or primary parkinsonism, hypokinetic rigidity syndrome (HRS), or parkinsonism, is a neurodegenerative brain disorder that affects the control of the motor system. The progressive death of dopamine-producing cells in the brain causes the main symptoms of Parkinson's disease. Most commonly, Parkinson's disease is diagnosed in people over the age of 50. Parkinson's disease is idiopathic (has an unknown cause) in most people. However, genetic factors also play a role in the disease.

[0204] Symptoms of Parkinson's disease include, but are not limited to, tremors in the hands, arms, legs, jaw, and face, muscle stiffness in the limbs and trunk, slow movements (bradykinesia), postural instability, difficulty walking, neuropsychiatric problems, changes in speech or behavior, depression, anxiety, pain, psychosis, dementia, hallucinations, and sleep problems.

[0205] In some embodiments, administration of the anti-SIRPβ1 antibodies of the present disclosure can prevent, reduce the risk of, and / or treat Parkinson's disease. In some embodiments, administration of anti-SIRPβ1 antibodies can induce one or more SIRPβ1 activities in individuals with Parkinson's disease. Amyotrophic lateral sclerosis

[0206] As used herein, amyotrophic lateral sclerosis (ALS) or motor neuron disease or Lou Gehrig's disease are used interchangeably to refer to a debilitating disease of varying etiology characterized by rapidly progressive weakness, muscle wasting and fasciculations, muscle spasticity, difficulty speaking (dysarthria), difficulty swallowing (dysphagia), and difficulty breathing (dyspnea).

[0207] Progranulin has been shown to play a role in ALS (Schymick, JC et al., (2007) J Neurol Neurosurg Psychiatry, 78: 754-6) and to provide protection against damage caused by proteins that cause ALS, such as TDP-43 (Laird, AS et al., (2010). PLoS ONE 5: e13368). Pro-NGF has also been shown to induce p75-mediated oligodendrocyte and corticospinal neuron death after spinal cord injury (Beatty et al., Neuron (2002), 36, pp. 375-386; Giehl et al., Proc. Natl. Acad. Sci USA (2004), 101, pp 6226-30).

[0208] In some embodiments, administration of an anti-SIRPβ1 antibody of the present disclosure can prevent, reduce the risk of, and / or treat ALS. In some embodiments, administration of an anti-SIRPβ1 antibody can induce one or more SIRPβ1 activities in an individual with ALS. Huntington's disease

[0209] Huntington's disease (HD) is an inherited neurodegenerative disease caused by an autosomal dominant mutation in the huntingtin gene (HTT). The expansion of the cytosine-adenine-guanine (CAG) triplet repeat in the huntingtin gene results in the production of a mutant form of the huntingtin protein (Htt) encoded by the gene. This mutant huntingtin protein (mHtt) is toxic and contributes to neuronal death. Symptoms of Huntington's disease most commonly appear between the ages of 35 and 44, although they can appear at any age.

[0210] Symptoms of Huntington's disease include, but are not limited to, problems with motor control, cramps, random movements (chorea), abnormal eye movements, impaired balance, seizures, difficulty chewing, difficulty swallowing, cognitive problems, language changes, memory loss, difficulty thinking, insomnia, fatigue, dementia, personality changes, depression, anxiety, and obsessive-compulsive behaviors.

[0211] In some embodiments, administration of the anti-SIRPβ1 antibodies of the present disclosure can prevent, reduce risk, and / or treat Huntington's disease (HD). In some embodiments, administration of anti-SIRPβ1 antibodies can induce one or more SIRPβ1 activities in individuals with HD. Tauopathies

[0212] Tauopathies, or tauopathies, are a class of neurodegenerative diseases caused by the aggregation of the microtubule-associated protein Tau in the brain. Alzheimer's disease (AD) is the best-known tauopathic disease and involves the accumulation of Tau protein in the form of insoluble neurofibrillary tangles (NFTs) within neurons. Other tauopathic diseases and conditions include progressive supranuclear palsy, dementia pugilistica (chronic traumatic encephalopathy), frontotemporal dementia and parkinsonism linked to chromosome 17, Lytico-Bodig disease (Guam Parkinsonism-dementia complex), tangle-predominant dementias, gangliogliomas and gangliocytomas, meningioangiomatosis, subacute sclerosing panencephalitis, lead encephalopathy, tuberous sclerosis, Hartmann-Strauss disease, lipofuscinosis, Pick's disease, corticobasal degeneration, argyrophilic grain disease (AGD), Huntington's disease, frontotemporal dementia, and frontotemporal lobar degeneration.

[0213] In some embodiments, administration of the anti-SIRPβ1 antibodies of the present disclosure can prevent, reduce the risk of, and / or treat tauopathy. In some embodiments, administration of anti-SIRPβ1 antibodies can induce one or more SIRPβ1 activities in an individual with a tauopathy. Multiple sclerosis

[0214] Multiple sclerosis (MS) may also be called disseminated cerebral sclerosis or disseminated encephalomyelitis. MS is an inflammatory disease in which the fatty myelin sheaths around the axons of the brain and spinal cord are destroyed, leading to demyelination and scarring, as well as a range of signs and symptoms. MS affects the ability of nerve cells in the brain and spinal cord to communicate with each other effectively. Nerve cells communicate by sending electrical signals called action potentials along long fibers called axons, which are contained in an insulating material called myelin. In MS, the body's own immune system attacks and destroys myelin. When myelin is lost, the axons can no longer conduct signals effectively. MS attacks usually occur in young adults and are more common in women.

[0215] Symptoms of MS include, but are not limited to, changes in sensation, such as loss of sensitivity or tingling; tingling or numbness, such as hypoesthesia and paresthesia; muscle weakness; clonus; muscle spasms; difficulty moving; difficulty with coordination and balance, such as ataxia; speech problems, such as dysarthria, or swallowing problems, such as dysphagia; vision problems, such as nystagmus, optic neuritis, including phosphenes, and diplopia; fatigue; acute or chronic pain; and bladder and bowel difficulties; varying degrees of cognitive impairment; mood symptoms of depression or mood swings; Uthof's phenomenon, which is an exacerbation of existing symptoms caused by exposure to higher than usual ambient temperatures; and Lhermitte's sign, which is a sensation of electricity running down the back when the neck is bent.

[0216] In some embodiments, administration of an anti-SIRPβ1 antibody of the present disclosure can prevent, reduce the risk of, and / or treat multiple sclerosis. In some embodiments, administration of an anti-SIRPβ1 antibody can induce one or more SIRPβ1 activities in an individual with multiple sclerosis. cancer

[0217] Still other aspects of the present disclosure provide methods for preventing, reducing risk, or treating an individual with cancer, comprising administering to the individual a therapeutically effective amount of an isolated anti-SIRPβ1 antibody of the present disclosure. Any isolated antibody of the present disclosure can be used in these methods. In some embodiments, the isolated antibody is an agonist antibody of the present disclosure.

[0218] The tumor microenvironment is known to contain a heterogeneous immune infiltrate, including T lymphocytes, macrophages, and cells of the myeloid / granulocyte lineage. In particular, the presence of M2 macrophages in tumors is associated with a poor prognosis. Therapies that reduce the number of these cells in tumors (such as CSF1R blockers) have shown beneficial effects in preclinical models and early-stage clinical studies. A pioneering preclinical study has also shown synergy between drugs targeting tumor-associated macrophages (e.g., CSF1 / CSF1R blocking antibodies) and checkpoint blocking antibodies targeting T cells, indicating that manipulation of these two cell types shows efficacy in tumor models where individual therapies are less effective (Zhu Y., Cancer Res. 2014 Sep 15; 74 (18): 5057-69). Without wishing to be bound by any particular theory, it is believed that inducing SIRPβ1 signaling in tumor-associated macrophages and / or neutrophils can inhibit the containment of immune responses in the tumor microenvironment, leading to a therapeutic anti-tumor immune response.

[0219] In certain embodiments, cancers to be prevented or treated by the methods of the present disclosure include, but are not limited to, squamous cell carcinomas (e.g., epithelial squamous cell carcinomas), lung cancer, small cell lung cancer, non-small cell lung cancer, squamous non-small cell lung cancer (NSCLC), adenocarcinomas of the lung, squamous carcinomas of the lung, non-squamous NSCLC, gliomas, peritoneal cancer, hepatocellular carcinoma, gastrointestinal cancer, gastric cancer or gastric cancer (including gastrointestinal cancer and gastrointestinal stromal cancer), renal cancer (e.g., clear cell carcinoma), ovarian cancer, liver cancer, colorectal cancer, endometrial cancer, renal cancer (e.g., renal cell carcinoma (RCC)), prostate cancer (e.g., hormone-refractory prostate adenocarcinoma), thyroid cancer, neuroblastoma, pancreatic cancer, glioblastoma (glioblastoma multiforme), cervical cancer, stomach cancer, bladder cancer, liver cancer, breast cancer, colon cancer, and head and neck cancer (or carcinoma), gastric cancer, germ cell tumors, pediatric sarcoma, sinonasal natural killer, melanoma (e.g., metastatic malignant melanoma, such as skin or intraocular malignant melanoma), bone cancer, skin cancer, uterine cancer, anal area cancer, testicular cancer, fallopian tube cancer, endometrial cancer, cervical cancer, vaginal cancer, vulvar cancer, esophageal cancer, small intestine cancer, endocrine system cancer, parathyroid cancer, adrenal cancer, soft tissue sarcoma, urethral cancer, penile cancer, solid tumors of children, ureteral cancer, renal pelvic cancer, central nervous system neoplasms (CNS), prostate cancer, CNS lymphoma, myelodysplastic syndrome, colorectal neoplasms, solid tumors, tumor angiogenesis, spinal tumors, brain stem gliomas, pituitary adenomas, Kaposi's sarcoma, epidermoid carcinomas, squamous cell carcinomas, T-cell lymphomas, environmentally induced cancers (including those induced by asbestos), virus-related cancers (e.g., human papillomavirus (HPV)-related tumors), and hematological malignancies derived from either of the two major blood cell lineages, the myeloid cell lineage (which generates granulocytes, erythrocytes, platelets, macrophages, and mast cells) or the lymphoid cell lineage (which generates B, T, NK, and plasma cells), such as all types of leukemias, lymphomas, and myelomas, e.g., acute, Chronic, lymphocytic and / or myeloid leukemias, such as acute leukemia (ALL), acute myeloid leukemia (AML), chronic lymphocytic leukemia (CLL), and chronic myeloid leukemia (CML), undifferentiated AML (M0), myeloblastic leukemia (M1), myeloblastic leukemia (M2; with cell maturation), promyelocytic leukemia (M3 or M3 variant [M3V]), myelomonocytic leukemia (M4 or M4 variant with eosinophilia [M4E]), monocytic leukemia (M5), erythroleukemia (M6), megakaryoblastic leukemia (M7), isolated granulocytic sarcoma, and chloroma;Lymphomas, such as Hodgkin's lymphoma (HL), non-Hodgkin's lymphoma (NHL), indolent lymphoma, large B-cell diffuse lymphoma, B-cell hematologic malignancies, such as B-cell lymphoma, T-cell lymphoma, lymphoplasmacytoid lymphoma, monocytoid B-cell lymphoma, mucosa-associated lymphoid tissue (MALT) lymphoma, anaplastic (e.g., Ki 1+) large cell lymphoma, adult T-cell lymphoma / leukemia, mantle cell lymphoma, angioimmunoblastic T-cell lymphoma, angiocentric lymphoma, intestinal T-cell lymphoma, primary mediastinal B-cell lymphoma, precursor T-lymphoblastic lymphoma, T-lymphoblastic; and lymphoma / leukemia (T-Lbly / T-ALL), peripheral T-cell lymphoma, lymphoblastic lymphoma, post-transplant lymphoproliferative disorder, true histiocytic lymphoma, primary central nervous system systemic lymphoma, primary effusion lymphoma, lymphoblastic lymphoma (LBL), hematopoietic neoplasms of the lymphoid lineage, acute lymphoblastic leukemia, diffuse large B-cell lymphoma, Burkitt's lymphoma, follicular lymphoma, diffuse histiocytic lymphoma (DHL), immunoblastic large cell lymphoma, precursor B-lymphoblastic lymphoma, cutaneous T-cell lymphoma (CTLC) (also called mycosis fungoides or Sezary syndrome), and Waldenstrom's macroglobulinemia lymphoplasmacytic lymphoma (LPL); myelomas, such as IgG myeloma, light chain myeloma, non-secretory myeloma, sultry myeloma (also called indolent myeloma), solitary plasmacytoma, and multiple myeloma, chronic lymphocytic leukemia (CLL), hairy cell lymphoma; hematopoietic tumors of myeloid lineage, tumors of mesenchymal origin, including fibrosarcoma and rhabdomyosarcoma; seminoma, teratoma, central and peripheral nerve tumors, including astrocytoma, Schwann cell tumor; Tumors of mesenchymal origin, including fibrosarcoma, rhabdomyosarcoma, and osteosarcoma; and other tumors, including melanoma, xeroderma pigmentosum, keratoacanthoma, seminoma, thyroid follicular carcinoma and teratocarcinoma, hematopoietic tumors of lymphoid lineage, such as T-cell and B-cell tumors, including but not limited to T-cell disorders, such as T-prolymphocytic leukemia (T-PLL), including small cell and gyrus cell types; large granular lymphocytic leukemia (LGL), preferably T-cell type; a / d T-NHL heptasyllabic lymphoma; peripheral / post-thymic T-cell lymphoma (polymorphic and immunoblastic subtypes); angiocentric (nasal) T-cell lymphoma; head and neck cancer, renal cancer, rectal cancer, thyroid cancer; acute myeloid lymphoma, and any combination of the cancers. The anti-SIRPβ1 antibodies disclosed herein can also be used to treat metastatic cancer. ;

[0220] In some embodiments, the cancer is selected from sarcoma, bladder cancer, brain cancer, breast cancer, colon cancer, rectal cancer, endometrial cancer, kidney cancer, renal pelvic cancer, leukemia, lung cancer, small cell lung cancer, melanoma, lymphoma, pancreatic cancer, prostate cancer, ovarian cancer, and fibrosarcoma. In some embodiments, the cancer is triple negative breast cancer. In some embodiments, the cancer can be an early stage cancer or an advanced stage cancer. In some embodiments, the cancer can be a primary tumor. In some embodiments, the cancer can be a metastatic tumor derived from any of the above types of cancer at a second site.

[0221] In some embodiments, the cancer is selected from glioblastoma multiforme; renal clear cell carcinoma; adrenocortical carcinoma; bladder urothelial carcinoma; diffuse large B-cell lymphoma; lung adenocarcinoma; pancreatic adenocarcinoma, renal cell carcinoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, indolent B-cell lymphoma, aggressive B-cell lymphoma, T-cell lymphoma, acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), chronic lymphocytic leukemia (CLL), chronic myeloid leukemia (CML), multiple myeloma, myelodysplastic syndrome signs, myeloproliferative neoplasms, invasive breast carcinoma, cervical squamous cell carcinoma, endocervical adenocarcinoma, bile duct carcinoma, colon adenocarcinoma, diffuse large B-cell lymphoma, esophageal carcinoma, head and neck squamous cell carcinoma, renal chromophobe cell carcinoma, renal papillary cell carcinoma, low-grade glioma, hepatocellular carcinoma, lung squamous cell carcinoma, mesothelioma, ovarian serous cystadenocarcinoma, pancreatic adenocarcinoma, pheochromocytoma and paraganglioma, prostate adenocarcinoma, rectal adenocarcinoma, skin melanoma, gastric adenocarcinoma, testicular germ cell tumor, thyroid carcinoma, thymoma, endometrial carcinoma, uterine carcinosarcoma, and uveal melanoma.

[0222] In some embodiments, the present disclosure provides methods of preventing, reducing risk, or treating an individual having cancer by administering to the individual a therapeutically effective amount of an anti-SIRPβ1 antibody of the present disclosure.

[0223] In some embodiments, the present disclosure provides methods of preventing, reducing risk, or treating an individual having cancer, wherein the individual is refractory to checkpoint inhibitor therapy, by administering to the individual a therapeutically effective amount of an anti-SIRPβ1 antibody of the present disclosure.

[0224] In some embodiments, the anti-SIRPβ1 antibodies of the present disclosure can be administered in combination with a therapeutic agent that acts as a checkpoint inhibitor. In some embodiments, the method further comprises administering to the individual at least one antibody that specifically binds to an inhibitory immune checkpoint molecule, and / or another standard or investigative anti-cancer therapy. In some embodiments, the inhibitory checkpoint molecule is selected from PD1, PDL1, CD40, OX40, ICOS, CD28, CD137 / 4-1BB, CD27, GITR, CTLA4, PD-L2, B7-H3, B7-H4, HVEM, LIGHT, BTLA, CD30, TIGIT, VISTA, KIR, GAL9, TIM1, TIM3, TIM4, A2AR, LAG3, DR-5, CD2, CD5, CD39, and CD73. In typical embodiments, the therapeutic agent is an antibody to a checkpoint inhibitor selected from PD1, PDL1, CD40, OX40, ICOS, CD28, CD137 / 4-1BB, CD27, GITR, CTLA4, PD-L2, B7-H3, B7-H4, HVEM, LIGHT, BTLA, CD30, TIGIT, VISTA, KIR, GAL9, TIM1, TIM3, TIM4, A2AR, LAG3, DR-5, CD2, CD5, CD39, or CD73. In some embodiments, at least one antibody that specifically binds to an inhibitory checkpoint molecule is administered in combination with an anti-SIRPβ1 antibody of the present disclosure. In some embodiments, a combination of antibodies to checkpoint inhibitors is administered in combination with an anti-SIRPβ1 antibody of the present invention.

[0225] In some embodiments, the anti-SIRPβ1 antibodies of the present disclosure can be administered in combination with at least one agonistic antibody that specifically binds to a stimulatory checkpoint protein, such as an agonistic anti-CD40 antibody, an agonistic anti-OX40 antibody, an agonistic anti-ICOS antibody, an agonistic anti-CD28 antibody, an agonistic anti-TREM1 antibody, an agonistic anti-TREM2 antibody, an agonistic anti-CD137 / 4-1BB antibody, an agonistic anti-CD27 antibody, an agonistic anti-glucocorticoid-induced TNFR-related protein GITR antibody, an agonistic anti-CD30 antibody, an agonistic anti-BTLA antibody, an agonistic anti-HVEM antibody, an agonistic anti-CD2 antibody, an agonistic anti-CD5 antibody, and any combination thereof.

[0226] In some embodiments, the method further comprises administering to the individual at least one antibody that specifically binds to an inhibitory immune checkpoint molecule, and / or another standard or investigative anti-cancer therapy. In some embodiments, at least one antibody that specifically binds to an inhibitory checkpoint molecule is administered in combination with an anti-SIRPβ1 antibody of the present disclosure. In some embodiments, at least one antibody that specifically binds to an inhibitory checkpoint molecule is selected from anti-PD-L1 antibodies, anti-CTLA4 antibodies, anti-PD-L2 antibodies, anti-PD-1 antibodies, anti-B7-H3 antibodies, anti-B7-H4 antibodies, and anti-HVEM antibodies, anti-B and T lymphocyte attenuators (BTLA) antibodies, anti-KIR antibodies, anti-GAL9 antibodies, anti-TIM3 antibodies, anti-A2AR antibodies, anti-LAG-3 antibodies, anti-phosphatidylserine antibodies, anti-CD27 antibodies, and any combination thereof. In some embodiments, standard or investigative anti-cancer therapy is selected from radiation therapy, cytotoxic chemotherapy, targeted therapy, imatinib Trastuzumab One or more of adoptive cell transfer (ACT), chimeric antigen receptor T cell transfer (CAR-T), vaccine therapy, and cytokine therapy. In some embodiments, the method further comprises administering to the individual at least one antibody that specifically binds to an inhibitory cytokine. In some embodiments, at least one antibody that specifically binds to an inhibitory cytokine is administered in combination with an anti-SIRPβ1 antibody of the present disclosure. In some embodiments, at least one antibody that specifically binds to an inhibitory cytokine is selected from an anti-CCL2 antibody, an anti-CSF-1 antibody, an anti-IL-2 antibody, and any combination thereof.

[0227] In some embodiments, the method further comprises administering to the individual at least one agonistic antibody that specifically binds to a stimulatory immune checkpoint protein. In some embodiments, at least one agonistic antibody that specifically binds to a stimulatory checkpoint protein is administered in combination with an anti-SIRPβ1 antibody of the present disclosure. In some embodiments, at least one agonistic antibody that specifically binds to a stimulatory checkpoint protein is selected from agonistic anti-CD40 antibodies, agonistic anti-OX40 antibodies, agonistic anti-ICOS antibodies, agonistic anti-CD28 antibodies, agonistic anti-CD137 / 4-1BB antibodies, agonistic anti-CD27 antibodies, agonistic anti-glucocorticoid-induced TNFR-related protein GITR antibodies, and any combination thereof.

[0228] In some embodiments, the anti-SIRPβ1 antibodies of the present invention are administered in combination with radiation therapy and / or chemotherapeutic agents. Chemotherapeutic agents include, for example, the following groups: antimetabolites / anticancer agents, such as pyrimidine analogs (5-fluorouracil, floxuridine, capecitabine, gemcitabine, and cytarabine) and purine analogs, folic acid antagonists and related inhibitors (methotrexate, pemetrexed, mercaptopurine, thioguanine, pentostatin, and 2-chlorodeoxyadenosine (cladribine)); antiproliferative / antimitotic agents, including natural products, such as vinca alkaloids (vinblastine, vincristine, and vinorelbine), microtubule disrupting agents, such as taxanes (paclitaxel, docetaxel), vincristine, vinblastine, Nocodazole, epothilones, eribulin, and navelbine; epipodophyllotoxins (etoposide, teniposide); DNA damaging agents (actinomycin, amsacrine, anthracyclines, bleomycin, busulfan, camptothecin, carboplatin, chlorambucil, cisplatin, cyclophosphamide, Cytoxan, dactinomycin, daunorubicin, doxorubicin, epirubicin, hexamethylmelamine, oxaliplatin, ifosfamide, melphalan, mechlorethamine, mitomycin, mitoxantrone, nitrosoureas, plicamycin, procarbazine, taxol, taxotere, temozolomide, teniposide, trimethylenethiophosphoramide, and etoposide (VP 16)); DNA methyltransferase inhibitors (azacytidine); antibiotics such as dactinomycin (actinomycin D), daunorubicin, doxorubicin (adriamycin), idarubicin, anthracyclines, mitoxantrone, bleomycin, plicamycin (mithramycin), and mitomycin; enzymes (L-asparaginase, which systemically metabolizes L-asparagine and deprives cells that do not have the ability to synthesize their own asparagine); antiplatelet agents; antiproliferative / antimitotic alkylating agents such as nitrogen mustards (mechlorethamine, cyclophosphamide and analogs, melphalan, chlorambucil), ethyleneimines and methylmelamines (hexamethylmelamine and thiotepa), alkyl sulfonates / esters (busulfan), nitrosoureas (carmustine (BCNU) and analogs, streptozocin), triazines (dacarbazine (DTI) C)); antiproliferative / antimitotic antimetabolites, such as folic acid analogs (methotrexate); platinum coordination complexes (cisplatin, carboplatin), procarbazine, hydroxyurea, mitotane, aminoglutethimide; hormones, hormone analogs (estrogens, tamoxifen, goserelin, bicalutamide, nilutamide) and aromatase inhibitors (letrozole, anastrozole); anticoagulants (heparin, synthetic heparin salts and other inhibitors of thrombin); fibrinolytic agents (such as tissue plasminogen activator, streptokinase and urokinase), aspirin, dipyridamole, ticlopidine, clopidogrel, abciximab; antimigratory agents; antisecretory agents (brefeldin); immunosuppressive drugs (cyclosporine, tacrolimus (FK-506), sirolimus (rapamycin), azathioprine, mycophenolate mofetil);Antiangiogenic compounds (TNP470, genistein, pomalidomide) and growth factor inhibitors (vascular endothelial growth factor (VEGF) inhibitors such as ziv-aflibercept; fibroblast growth factor (FGF) inhibitors); inhibitor of apoptosis protein (IAP) antagonists (birinapant); histone deacetylase (HDAC) inhibitors (vorinostat, romidepsin, chidamide, panobinostat, mocetinostat, abexinostat, belinostat, entinostat, resminostat, givinostat, quisinostat, SB939); proteasome inhibitors (ixazomib); angiotensin receptor blockers; nitric oxide donors; antisense oligonucleotides; antibodies (trastuzumab, panitumumab, pertuzumab, cetuximab, adalimumab, golimumab, infliximab, rituximab, ocrelizumab, ofatumumab, obinutuzumab, alemtuzumab, abciximab, atlizumab, daclizumab, denosumab, efalizumab, elotuzumab, rovizumab, rulizumab, ustekinumab, visilizumab, gemtuzumab ozogamicin, brentuximab vedotin); chimeric antigen receptors; cell cycle inhibitors (flavopiridol, roscovitine, bryostatin-1) and differentiation inducers (tretinoin); mTOR inhibitors, topoisomerase inhibitors (doxorubicin (adriamycin), amsacrine, camptothecin, daunorubicin, dactinomycin, teniposide, epirubicin, etoposide, idarubicin, irinotecan (CPT-11) and mitoxantrone, topotecan, irinotecan), corticosteroids (cortisone, dexamethasone, hydrocortisone, methylprednisolone, prednisone, and prednisolone); PARP inhibitors (niraparib, olaparib); focal adhesion kinase (FAK) inhibitors; (defactinib (VS-6063), VS-4718, VS-6062, GSK2256098); growth factor signaling kinase inhibitors (cediranib, galunisertib, rociletinib, vandetanib, afatinib, EGF816, AZD4547); c-Met inhibitors (capmatinib, INC280); ALK inhibitors (seritinib, crizotinib); mitochondrial dysfunction inducers, toxins, such as cholera toxin, ricin, Pseudomonas exotoxin, Bordetella pertussis adenylate cyclase toxin, or diphtheria toxin, and caspase activators; and chromatin disruptors. In some embodiments, the chemotherapeutic agent is a B-Raf inhibitor, a MEK inhibitor, a VEGF inhibitor, a VEGFR inhibitor, a tyrosine kinase inhibitor, an anti-mitotic agent, or any combination thereof.

[0229] In some embodiments, the anti-SIRPβ1 antibodies of the present disclosure are administered in combination with adoptive cell transfer (ACT) therapy, chimeric antigen receptor T cell transfer (CAR-T) therapy, vaccine therapy, and / or cytokine therapy.

[0230] In some embodiments, an anti-SIRPβ1 antibody of the present disclosure is administered in combination with at least one antibody that specifically binds to an inhibitory cytokine, for example, an inhibitory cytokine, such as an anti-CCL2 antibody, an anti-CSF-1 antibody, or an anti-IL-2 antibody.

[0231] In some embodiments, the anti-SIRPβ1 antibodies of the present disclosure are administered in combination with at least one stimulatory cytokine. In some embodiments that may be combined with any of the preceding embodiments, at least one stimulatory cytokine is selected from IFN-α4, IFN-β, IL-1β, TNF-α, IL-6, IL-8, CRP, IL-20 family members, LIF, IFN-γ, OSM, CNTF, GM-CSF, IL-11, IL-12, IL-15, IL-17, IL-18, IL-23, CXCL10, IL-33, MCP-1, MIP-1-β, and any combination thereof. VI. Diagnostic Uses

[0232] In some embodiments of any antibody, any anti-SIRPβ1 antibody provided herein is useful for detecting the presence of SIRPβ1 in a sample or individual. As used herein, the term "detection" encompasses quantitative or qualitative detection. Provided herein are methods of using the disclosed antibodies for diagnostic purposes, such as detecting SIRPβ1 in an individual or in a tissue sample derived from an individual. In some embodiments, the individual is a human.

[0233] The detection method may involve quantifying the antigen-bound antibody. Detection of antibodies in biological samples can be performed using any method known in the art, including immunofluorescence microscopy, immunocytochemistry, immunohistochemistry, ELISA, FACS analysis, immunoprecipitation, or micropositron emission tomography. In certain embodiments, the antibody is radiolabeled, for example with 18 F, which is then detected using micro-positron emission tomography analysis. Antibody binding can also be quantified in patients by non-invasive techniques such as positron emission tomography (PET), X-ray computed tomography, single photon emission computed tomography (SPECT), computed tomography (CT), and computed axial tomography (CAT). VII. Products

[0234] Provided herein are articles (e.g., kits) comprising an anti-SIRPβ1 antibody described herein. The article may include one or more containers containing an antibody described herein. The container may be any suitable packaging, including but not limited to vials, bottles, cans, flexible packaging (e.g., sealed Mylar or plastic bags), etc. The container may be a unit dose, bulk (e.g., multi-dose packaging), or a subunit dose.

[0235] In some embodiments, the kit may further include a second agent. In some embodiments, the second agent is a pharmaceutically acceptable buffer or diluent, including but not limited to, such as bacteriostatic water for injection (BWFI), phosphate buffered saline, Ringer's solution and dextrose solution. In some embodiments, the second agent is a pharmaceutically active agent. In some embodiments, the second agent is a pharmaceutically active agent described herein.

[0236] In some embodiments of any article of manufacture, the article of manufacture further includes instructions for use according to the methods disclosed herein. The instructions generally include information about the dosage, dosing schedule, and route of administration for the intended treatment. In some embodiments, these instructions contain instructions for administering the isolated antibodies of the present disclosure (e.g., anti-SIRPβ1 antibodies described herein) according to any method disclosed herein to prevent, reduce risk, or treat an individual with a disease, disorder, or injury selected from the following: dementia, frontotemporal dementia, Alzheimer's disease, vascular dementia, mixed dementia, Creutzfeldt-Jakob disease, normal pressure hydrocephalus, amyotrophic lateral sclerosis, Huntington's disease, Tau disease, Nasu-Hakola disease, stroke, acute trauma, chronic trauma, lupus, acute and chronic colitis, rheumatoid arthritis, wound healing, Crohn's disease, inflammatory bowel disease , ulcerative colitis, obesity, malaria, essential tremor, lupus of the central nervous system, Behcet's disease, Parkinson's disease, dementia with Lewy bodies, multiple system atrophy, Shy-Drager syndrome, progressive supranuclear palsy, corticobasal ganglionic degeneration, acute disseminated encephalomyelitis, granulomatous disorders, sarcoidosis, diseases of aging, seizures, spinal cord injury, traumatic brain injury, age-related macular degeneration, glaucoma, retinitis pigmentosa, retinal degeneration, respiratory tract infection, sepsis, eye infection, systemic infection, lupus, arthritis, multiple sclerosis, low bone density, osteoporosis, osteogenesis, osteosclerotic disease, Paget's disease of bone, and cancer. In some embodiments, the instructions include instructions for use of the anti-SIRPβ1 antibody and the second agent (e.g., a second pharmaceutically active agent).

[0237] The present disclosure will be more fully understood by reference to the following examples. However, they should not be interpreted as limiting the scope of the present disclosure. All references throughout the present disclosure are hereby expressly incorporated by reference. Example

[0238] The following examples are provided by way of illustration only and not by way of limitation. Those skilled in the art will readily recognize that a variety of noncritical parameters can be changed or modified to produce essentially similar results. Example 1: Generation, identification, and characterization of agonistic anti-SIRPβ1 antibodies

[0239] The amino acid sequence of human SIRPβ1 isoform 1 protein (SIRPβ1) is set forth below in SEQ ID NO: 1. Human SIRPβ1 contains a signal peptide located at amino acid residues 1-29 of SEQ ID NO: 1; an extracellular immunoglobulin-like variable (IgV) domain located at amino acid residues 30-136 of SEQ ID NO: 1; two immunoglobulin-like constant (IgC) domains located at amino acids 147-246 and 253-347 of SEQ ID NO: 1; a transmembrane domain located at amino acid residues 372-392 of SEQ ID NO: 1; and an intracellular domain located at amino acid residues 393-398 of SEQ ID NO: 1. Human SIRPβ1 amino acid sequence (SEQ ID NO: 1):

[0240] The human SIRPβ1 amino acid sequence contains a lysine residue (aa380) within the transmembrane domain that interacts with an aspartate in DAP12, a key adaptor protein that transduces signals from SIRPβ1, as well as TREM1, TREM2, and other related IgV family members. BLAST analysis of human SIRPβ1 identified multiple isoforms that may be derived from alternative splicing. Among these transcript variants, SIRPβ1 isoform 3 shares the greatest sequence identity with SIRPβ1 isoform 1 ( Figure 1A Human SIRPβ1 is also related to human SIRPα. An alignment of the amino acid sequences of human SIRPβ1 and human SIRPα was generated by bidirectional BLAST ( Figure 1B The extracellular regions of the two receptors share 85% sequence identity. In contrast, human SIRPβ1 displays poor homology to mouse SIRPβ1 ( Figure 2 ), suggesting that the SIRP loci in the two species experience different selective pressures.

[0241] Antibodies that bind to the extracellular domain of human SIRPβ1, particularly the extracellular IgV domain (amino acid residues 30-136 of SEQ ID NO: 1), were generated using mouse hybridoma technology, phage display technology, and yeast-based platform technology. Antibodies were screened for their ability to bind to cells expressing SIRPβ1 and their ability to activate SIRPβ1 signaling, activity, and function in vivo in cells and in intact animals as described in Examples 2-20 below. For example, agonistic anti-SIRPβ1 antibodies targeting the IgV domain (amino acid residues 30-136) can be generated. Based on the similarity to SIRPα, the IgV domain of SIRPβ1 is predicted to bind to unknown ligands and, via receptor multimerization, lead to activation. Generation of His-tagged and Fc-conjugated human SIRPα and human SIRPβ1 IgV domains

[0242] For mammalian expression of human SIRPα and human SIRPβ1 IgV domain (also referred to as "domain 1") antigens (SEQ ID NOs: 387 and 388, respectively), cDNA-based synthetic genes were cloned into mammalian expression vectors, followed by transient transfection and expression in HEK293 / T cells. The constructs included a heterologous signal peptide and a C-terminal human IgG1 Fc and / or His tag. Briefly, the cells were complexed with a transfection reagent, followed by exposure to HEK293 / T cells for 1 hour, followed by dilution with culture medium to a final density of 4×10 6 Cells / mL were transfected with expression vectors containing the antigen of interest. The cells were then cultured with fresh feed medium every 48 hours for 7 days. After 7 days, the supernatant was collected after centrifugation and purified using Ni-Sepharose and in some cases SEC column chromatography to achieve >95% non-aggregated monomer content. SIRPα and SIRPβ1 monomer antigens were prepared by fragmenting SIRPα / β1 Fc fusion antigens with a modified hinge region (Lynaugh et al., MAbs. 2013 Oct; 5(5): 641-45) with FabRICATOR (IdeS) protease (Genovis, catalog number A2-FR2-1000), followed by protein A affinity purification to remove undigested Fc fusion protein and SEC to remove aggregated monomers. Library screening for anti-SIRPβ1 antibodies

[0243] Eight unimmunized (pre-immunization) human synthetic yeast libraries, each containing approximately 10 9 Diversity. Ten parallel selections were performed, 8 non-immunized libraries for human SIRPβ1-Fc fusion antigen selection and 2 pools of 8 libraries for human SIRPβ1 monomer selection. For the first two rounds of selection, magnetic bead sorting using the Miltenyi MACs system was performed essentially as described (Siegel et al., J Immunol Methods 2004 Mar; 286(1-2): 141-53). Briefly, yeast cells (approximately 10 9 cells / library, total density about 10 10) were incubated with 3 ml of 10 nM biotinylated SIRPβ1-Fc fusion antigen or 100 nM biotinylated SIRPβ1 monomer antigen in PBS containing 0.1% BSA in FACS wash buffer at room temperature for 15 minutes. After washing once with 50 ml of ice-cold wash buffer, the cell pellet was resuspended in 40 mL of wash buffer, and 500 μL of streptavidin microbeads (Miltenyi Biotec, Bergisch Gladbach, Germany, catalog number 130-048-101) were added to the yeast and incubated at 4°C for 15 minutes. Next, the yeast cells were pelleted, resuspended in 5 mL of wash buffer, and loaded onto a MACS LS column (Miltenyi Biotec, Bergisch Gladbach, Germany, catalog number 130-042-401). After loading 5 mL, the column was washed 3 times with 3 ml of FACS wash buffer. The column was then removed from the magnetic field and the yeast was eluted with 5 mL of growth medium and then grown overnight. The following four rounds of sorting were performed using flow cytometry.

[0244] For the first round of FACS selection, make approximately 1x10 8 Yeast were pelleted, washed three times with wash buffer, and incubated with 10 nM biotinylated SIRPβ1-Fc fusion antigen or 100 nM biotinylated SIRPβ1 monomer antigen at room temperature for 10 minutes. The yeast were then washed twice and stained with either a 1:100 dilution of goat anti-human F(ab')2κ-FITC (Southern Biotech, Birmingham, Alabama, catalog number 2062-02) and a 1:500 dilution of streptavidin-Alexa Fluor 633 (LifeTechnologies, Grand Island, NY, catalog number S21375) or a 1:50 dilution of Extravidin-phycoerythrin (Sigma-Aldrich, St Louis, catalog number E4011) at 4°C for 15 minutes. After washing twice with ice-cold wash buffer, the cell pellet was resuspended in 0.4 mL wash buffer and transferred to a sorting tube with a filter cap. Sorting was performed using a FACS ARIA sorter (BD Biosciences) and sorting gates were determined to select only SIRPβ1 binding clones. In the following round of selection, approximately 2×10 7The yeast were incubated with a polyspecific reactive reagent for negative selection to reduce polyspecific binders (Xu et al., PEDS. 2013 Oct; 26(10): 663-70), and binders against a control protein, a His-tagged human SIRPα monomer. The next round was labeled with 10 nM human SIRPβ1-Fc fusion antigen and 100 nM human SIRPβ1 monomer antigen. After the last round of sorting, yeast cells were plated and individual colonies were picked for characterization. The last round was labeled with 100 nM and 10 nM human SIRPβ1 monomer antigen.

[0245] The heavy chains from the second and fourth rounds of FACS sorting selection output were used to prepare the light chain diversified library for additional selections. For these selections, the first round of selection utilized Miltenyi MACs beads and was labeled with 10 nM human SIRPβ1-Fc fusion antigen. This was followed by four rounds of FACS sorting. The first round used 100 nM human SIRPβ1 monomer antigen. The second round of FACS was negative sorting to reduce binders to the binding reagent, multispecific binders, and binders to the control protein human SIRPα His-tagged monomer. The final two rounds utilized human SIRPβ1 monomer titration (100 nM, 10 nM, and 1 nM) to select the highest affinity binder, 100 nM human SIRPα monomer, and competition with the control AM4-5 antibody that binds to the SIRPβ1 IgV domain to assess competitor representation in the enriched population (see U.S. Patent Application Publication No. US2014 / 0242095). After the final round of sorting, the yeast cells were plated and individual colonies were picked for characterization. Antibody IgG and Fab production and purification

[0246] Yeast clones were grown to saturation and then induced at 30°C for 48 hours with shaking. After induction, yeast cells were pelleted and supernatant was harvested for purification. IgG was purified using a protein A column and eluted with acetic acid pH 2.0. Fab fragments were generated by papain digestion and purified on CaptureSelect IgG-CH1 affinity matrix (Life Technologies, catalog number 1943200250). Combination experiment

[0247] By using ForteBio The dissociation constant (K) of anti-SIRPβ1 antibody was measured using the Red384 system (ForteBio, Menlo Park, CA). D) to determine their affinity, generally as previously described (Estep et al., MAbs. 2013 Mar-Apr; 5(2): 270-8). Briefly, Octet affinity measurements were performed as follows, i.e., IgG was loaded online onto the AHQ sensor. The sensor was equilibrated offline in assay buffer for 30 minutes and then monitored online for 60 seconds to establish a baseline. For affinity binding measurements, the IgG-loaded sensor was exposed to 100 nM antigen (human SIRPα domain 1 (D1) Fc fusion or SIRPβ1 domain 1 (D1) Fc fusion) for 3 minutes and then transferred to assay buffer for 3 minutes for dissociation rate measurements. Additional affinity binding was determined by loading biotinylated SIRPβ1 monomers onto the SA sensor and exposing to 100 nM IgG in solution. Monovalent binding measurements were obtained by loading human SIRPα- or SIRPβ1-Fc fusion antigens onto the AHQ sensor, followed by exposure to 100 nM anti-SIRPβ1 antibody Fab. Additional monovalent measurements were performed by loading biotinylated human SIRPα or SIRPβ1 monomers onto the SA sensor, followed by exposure to 100 nM Fab in solution. Kinetic data were fitted using a 1:1 binding model in the data analysis software provided by ForteBio (ForteBio Data Analysis Software 7.0). Epitope binning

[0248] The standard sandwich-type binning assay was performed at ForteBio Epitope binning of anti-SIRPβ1 antibodies was performed on the Red384 system (ForteBio, Menlo Park, CA). Control anti-target IgG was loaded onto the AHQ sensor and unoccupied Fc binding sites on the sensor were blocked with an irrelevant human IgG1 antibody. The sensor was then exposed to 100 nM target antigen followed by a secondary anti-target antibody. Data were processed using ForteBio Data Analysis Software 7.0. Additional binding of the secondary antibody after antigen association indicates an unoccupied epitope (non-competitor), while no binding indicates epitope blocking (competitor). This process was repeated for two reference antibodies that bind to the SIRPβ1 IgV domain: (i) AM4-5, which defines bin 1; and (ii) SB-17, which defines bin 2.

[0249] The final anti-SIRPβ1 antibody set was selected based on antigen binding affinity. Antibodies that were positive for binding to human SIRPβ1 were tested for cross-reactivity with human SIRPα. The bin class of each antibody is listed in Table 2 below. In Table 2, "ND" refers to antibodies with undetermined bin class; "NB" refers to antibodies with no binding to the indicated antigen; "PF" refers to antibodies whose antigen binding kinetics showed a poor fit to a 1:1 binding model. No detectable binding to mouse SIRPβ1D1-Fc, human SIRPαD1-Fc, or mouse SIRPαD1-Fc was observed for any of the anti-SIRPβ1 antibodies. Table 2: Biochemical characterization of anti-SIRPβ1 antibodies Antibody heavy and light chain variable domain sequences

[0250] Using standard techniques, the amino acid sequences encoding the light chain variable domain and heavy chain variable domain of the antibody were determined. The EU or Kabat sequences of the antibodies are listed in Tables 3-6, as follows. The EU or Kabat light chain HVR sequences of the antibodies are listed in Table 3. The EU or Kabat heavy chain HVR sequences of the antibodies are listed in Table 4. The EU or Kabat light chain framework (FR) sequences of the antibodies are listed in Table 5. The EU or Kabat heavy chain framework (FR) sequences of the antibodies are listed in Table 6. Table 3: EU or Kabat light chain HVR sequences of anti-SIRPβ1 antibodies Table 4: EU or Kabat heavy chain HVR sequences of anti-SIRPβ1 antibodies Table 5: EU or Kabat light chain framework sequences of anti-SIRPβ1 antibodies Table 6: EU or Kabat heavy chain framework sequences of anti-SIRPβ1 antibodies Characterization of SIRPβ1 antibody binding

[0251] Initial characterization of anti-SIRPβ1 antibodies involved determining their ability to bind to cell lines expressing human or mouse SIRPβ1. Cells were harvested and plated in 96-well plates at 10 6 / ml distribution, washed, and incubated on ice for 0.5 hours in 100 μl FACS buffer containing 1 μg / ml anti-SIRPβ1 antibody. The cells were then washed twice and incubated on ice for 30 minutes in 100 μl FACS buffer containing 0.5 μg / ml PE-conjugated secondary antibody. The cells were washed twice in cold FACS buffer and acquired on a BD FACSCanto. Data analysis and calculation of mean fluorescence intensity (MFI) values ​​or % positive cells were performed using FlowJo (TreeStar) software version 10.0.7.

[0252] Table 7 shows the mean fluorescence intensity (MFI) values ​​of anti-SIRPβ1 antibodies binding to Chinese hamster ovary (CHO) cell lines expressing low levels of recombinant human SIRPβ1. The human IgG1 isotype control established a background fluorescence signal set to 1. Of the 50 anti-SIRPβ1 antibody clones tested, 32 clones bound to cells at an MFI ≥ 2 times above background. As a negative control, anti-SIRPβ1 antibodies were also screened for surface binding to CHO cells overexpressing recombinant mouse SIRPβ1. As expected, none of the tested antibodies bound to mouse SIRPβ1. Given the high sequence similarity between receptors of the SIRP family, anti-SIRPβ1 antibodies were also screened for cross-reactivity to human and mouse SIRPα. In the cell binding assay, none of the anti-SIRPβ1 antibodies bound to cells overexpressing human or mouse SIRPα. Table 7: Cell binding characterization of anti-SIRPβ1 antibodies

[0253] In addition, the anti-SIRPβ1 antibodies were screened for antigen specificity by using a reporter cell line expressing the luciferase gene under the control of the NFAT (nuclear factor of activated T cells) promoter. The cell line BW5147.G.1.4 (1.4 μg / mL) derived from mouse thymic lymphoma T lymphocytes was infected with Cignal Lenti NFAT-luciferase virus (Qiagen). TIB48 TM ), generating BWZ / NFAT-luciferase reporter cells. Subsequently, cells were transduced with a lentivirus expressing a human SIRPα-DAP12 chimera (in which the intracellular ITIM motif of SIRPα was replaced with the intracellular ITAM motif of DAP12) or with two lentiviruses expressing human SIRPβ1 and human DAP12. Test antibodies and human IgG1 isotype controls were adsorbed to 96-well plates at 10 μg / mL. After washing, 10 5 NFAT-luciferase reporter cells expressing huSIRPα / DAP12 chimeras (BWZ-huSIRPα) or co-expressing huSIRPβ1 and DAP12 (BWZ-huSIRPβ1) were plated and incubated overnight at 37°C. Luciferase activity was measured by adding OneGlo reagent (Promega) to each well and incubating the samples on a plate shaker at room temperature for 3 minutes. GEN5 TM 2.04 software using BioTek Synergy TM The luminescent signal is quantified by a microplate reader. Figure 3A As shown in Figure 2, 48 of 50 anti-SIRPβ1 clones induced luciferase expression at >5 fold above background in BWZ-huSIRPβ1 reporter cells. In contrast, anti-SIRPβ1 antibodies failed to induce luciferase expression when BWZ-huSIRPα reporter cells were added to the antibody-coated wells ( Figure 3B ). In conclusion, only reporter cells expressing human SIRPβ1 induced luciferase expression in the presence of immobilized anti-SIRPβ1 antibodies, as measured by luminescent signals. These results establish that most anti-SIRPβ1 antibodies capable of binding to membrane-bound antigens exhibit specificity against the target antigen and do not cross-react with other SIRP receptors. Example 2: SIRPβ1 expression profile on myeloid cells

[0254] The SIRP family includes several transmembrane glycoproteins expressed primarily in the myeloid cell compartment. The expression pattern of SIRPβ1 was validated on primary human cells isolated from healthy human donors. Human primary monocytes were isolated from heparinized human peripheral blood obtained from two healthy donors (Blood Centers of the Pacific) using RosetteSep Human Monocyte Enrichment Mix (STEMCELL Technologies) according to the manufacturer's protocol. Both SIRPβ1 and TREM1 are preferentially expressed in CD14 高 Expression on monocytes. Monocytes were plated in RPMI (Invitrogen) containing 10% fetal bovine serum (Hyclone) and 50ng / ml M-CSF or GM-CSF (Peprotech) to induce differentiation of M2-like or M1-like macrophages, respectively. After 5-6 days, macrophages were harvested by scraping cells attached to plastic. Alternatively, monocytes were plated in RPMI medium containing 10% fetal bovine serum (Hyclone) and 20ng / ml IL-4 and GM-CSF (Peprotech) to induce differentiation of immature dendritic cells. After 6-7 days, dendritic cells were harvested by scraping cells attached to plastic.

[0255] like Figure 4A As shown in , CD14 expression defines a subset of primary human monocytes. Classical and intermediate monocytes express high levels of CD14, while nonclassical monocytes lack CD14 expression. Like TREM1, SIRPβ1 is abundantly expressed in CD14+ monocytes. Monocytes isolated from peripheral blood as described above were cultured with M-CSF or GM-CSF for 5 days to generate M2 or M1 macrophages, respectively. SIRPβ1 expression decreases upon differentiation of primary monocytes into macrophages, with M1-like macrophages expressing higher levels of the receptor relative to M2-like macrophages ( Figure 4B ). Similarly, SIRPβ1 expression is reduced during differentiation of primary monocytes into immature dendritic cells. LPS-induced dendritic cell maturation further downregulates SIRPβ1 expression, in contrast to the increased expression observed for TREM1 ( Figure 4C ). Unlike TREM1, LPS treatment downregulated SIRPβ1 expression on DCs. Peak expression of SIRPβ1 occurred in the blood, spleen, and lung. Observation of SIRPβ1 expression patterns from RNA-seq datasets (from multiple tissue samples collected from healthy donors) confirmed that SIRPβ1 transcript levels were predominant in the blood, consistent with high expression of the receptor in circulating monocytes and neutrophils ( Figure 5AHowever, RNA-seq data revealed that the expression pattern of SIRPβ1 isoform 3 transcripts is different from that of SIRPβ1 isoform 1; SIRPβ1 isoform 3 is mostly expressed in the brain rather than in peripheral tissues ( Figure 5B ). (Data were obtained from the Genotype-Tissue Expression (GTEx) database.) Example 3: SIRPβ1 antibody induces Syk phosphorylation

[0256] Spleen tyrosine kinase (Syk) is an intracellular signaling molecule that functions downstream of SIRPβ1 by phosphorylating several substrates, thereby driving the formation of signaling complexes leading to cell activation and inflammatory processes. The ability of agonistic SIRPβ1 antibodies to induce Syk activation was determined by culturing mouse monocytes and measuring the phosphorylation state of the Syk protein in cell extracts. In these experiments, a secondary antibody was used to crosslink the anti-SIRPβ1 antibody on the cells to induce intracellular signaling.

[0257] Bone marrow derived mononuclear cells (BMDM) from wild-type (WT) mice, human SIRPβ1 BAC transgenic mice, and mice lacking functional Fc receptor common γ chain gene expression (FcgR KO; REF: Takai T, 1994, Cell, 76 (3): 519-29) were starved in 1% serum RPMI for 4 hours, then removed from tissue culture dishes with PBS-EDTA, washed with PBS, and counted. Cells were coated with full-length SIRPβ1 antibody or huIgG1 isotype control for 15 minutes on ice. After washing with cold PBS, cells were incubated at 37°C in the presence of goat anti-human IgG for the indicated time periods. After stimulation, cells were lysed with lysis buffer (1% v / v NP-40%, 50 mM Tris-HCl (pH 8.0), 150 mM NaCl, 1 mM EDTA, 1.5 mM MgCl2, 10% glycerol, plus protease and phosphatase inhibitors), followed by centrifugation at 16,000 g for 10 minutes at 4°C to remove insoluble material. The lysate was then immunoprecipitated with anti-Syk antibody (N-19 for BMDM or 4D10 for human DC, Santa Cruz Biotechnology). The precipitated proteins were fractionated by SDS-PAGE, transferred to PVDF membranes and probed with anti-phosphotyrosine antibodies (4G10, Millipore). To confirm that all substrates were adequately immunoprecipitated, immunoblots were re-probed with anti-Syk antibody (Abcam, for BMDM) or anti-Syk (Novus Biological, for human DC). Visualization was performed using an enhanced chemiluminescence (ECL) system (GE healthcare) as described (eg, Peng et al., (2010) Sci Signal., 3(122):ra38). These studies provide support that the anti-SIRPβ1 antibodies of the present invention induce syk phosphorylation. Example 4: When expressing Fc γ SIRPβ1 antibody induces Syk phosphorylation when the receptor is clustered in adjacent cells

[0258] Activation of spleen tyrosine kinase (Syk) is promoted by cross-linking of two or more SIRPβ1 receptors by antibodies, thereby promoting the formation of signaling complexes, leading to cell activation and inflammatory processes. In vivo cross-linking is mediated by adjacent cells expressing Fc receptors (FcRs), such as B cells and other leukocytes (White AL, Cancer Immunol Immunother, 62: 941-948, 2013; Wilson NS, Cancer Cell 19: 101-113, 2011; Bartholomaeus P, J Immunol, 192: 2091-2098, 2014). In these experiments, helper cells expressing Fcγ receptors (i.e., B cells) were used to cross-link anti-SIRPβ1 antibodies to induce intracellular signaling.

[0259] The ability of Fc receptors to induce Syk activation via antibody clustering was determined by culturing mouse monocytes in the presence of cells expressing Fc receptors and measuring the phosphorylation state of Syk protein in cell extracts. Bone marrow-derived mononuclear cells (BMDM) from wild-type (WT) mice and human SIRPβ1 BAC transgenic mice were starved in 1% serum RPMI for 4 hours, then removed from tissue culture dishes with PBS-EDTA, washed with PBS, and counted. Cells were coated with full-length SIRPβ1 antibody or huIgG1 isotype control on ice for 15 minutes. After washing with cold PBS, cells were incubated at 37°C for 5 minutes with glutaraldehyde-fixed Fc receptor-expressing cells previously prepared as follows. Briefly, cells expressing Fc receptors were fixed with MACS microbeads (CD19 + B cells were isolated from mouse spleen using B cell isolation kit (Miltenyi Biotec) according to the manufacturer's protocol, or P815 cell line overexpressing FcR2b and FcR3. 6Cells / ml were fixed with 0.05% glutaraldehyde for 1 min at room temperature, the reaction was stopped with 1 μM glycine, and then the cells were washed thoroughly with PBS. After stimulation, cells were lysed with lysis buffer (1% v / v NP-40%, 50 mM Tris-HCl (pH 8.0), 150 mM NaCl, 1 mM EDTA, 1.5 mM MgCl2, 10% glycerol, plus protease and phosphatase inhibitors), followed by centrifugation at 16,000 g for 10 min at 4°C to remove insoluble material. Lysates were then immunoprecipitated with anti-Syk antibody (N-19 for BMDM or 4D10 for human DC, Santa Cruz Biotechnology). Precipitated proteins were fractionated by SDS-PAGE, transferred to PVDF membranes and probed with anti-phosphotyrosine antibody (4G10, Millipore). To confirm that all substrates were adequately immunoprecipitated, immunoblots were re-probed with anti-Syk antibody (Abcam, for BMDM) or anti-Syk (Novus Biological, for human DC). Visualization was performed using an enhanced chemiluminescence (ECL) system (GE healthcare) as described (e.g., Peng et al., (2010) Sci Signal., 3(122):ra38). These studies provide support that the anti-SIRPβ1 antibodies of the present invention induce syk phosphorylation when clustered by adjacent cells expressing Fcγ receptors. Example 5: SIRPβ1 antibody induces respiratory burst in immune cells

[0260] The agonistic function of the SIRPβ1 antibodies of the present disclosure was evaluated in primary human innate immune cells, such as monocytes and neutrophils.

[0261] Anti-SIRPβ1 and isotype control antibodies were diluted in serum-free RPMI medium and mixed with 100,000 neutrophils at a concentration of 10 μg / ml on a 96-well plate. Primary neutrophils were isolated from peripheral blood obtained on the same day using the EasySepTM Direct Human Neutrophil Isolation Kit (STEMCELL) according to the manufacturer's instructions. To detect the generation of reactive oxygen species (ROS), cells were labeled with 2 μM fluorescent dye CM-H2DCFDA. Cells were stimulated with soluble full-length human IgG1 isotype control or anti-SIRPβ1 antibodies SB-1, -2, -3, -4, -5, -6, -7, -8, -9, -11, -14, -15, -17, -27, -28, -31, -39, -40, -41, -45, -46, and -49. After antibody-mediated stimulation in the presence of CM-H2DCFDA for 1 hour at 37°C, relative fluorescence units in cells were measured at an excitation wavelength of 495 nm and an emission wavelength of 530 nm. The specific fluorescence index of stimulated cells was obtained by subtracting the background fluorescence of labeled cells incubated in medium alone and / or with an isotype control antibody (huIgG1). TM Microplate reader using GEN5 TM The plate was read using 2.04 software.

[0262] exist Fig. 6A Primary human neutrophils from two healthy donors were stimulated with 10 μg / mL soluble full-length anti-SIRPβ1 antibody or human IgG1 isotype control and labeled with 2 μM CM-H2DCFDA for 1 hour at 37°C to monitor SIRPβ1-mediated ROS generation. SIRPβ1 antibodies SB-1, -2, -3, and -5 demonstrated high agonism in solution. In contrast, SIRPβ1 antibodies SB-15, -17, -27, -28, -31, -39, -40, -41, -45, -46, and -49 weakly activated SIRPβ1-mediated respiratory burst in solution.

[0263] Cells were left untreated or stimulated with soluble full-length human IgG1 isotype control or anti-SIRPβ1 antibodies SB-1, -2, -3, -4, -5, -6, -7, -8, -9, -11, -12, -13, -14, -15, -17, -23, -26, -27, -28, -31, -33, -34, -35, -36, -37, -39, -40, -41, -45, -46, and -49. Figure 6BIn the present study, primary human monocytes were stimulated with 20 μg / mL soluble full-length anti-SIRPβ1 antibody or human IgG1 isotype control and labeled with 2 μM CM-H2DCFDA for 1 hour at 37°C to monitor SIRPβ1-mediated ROS generation and IL-8 production. SIRPβ1 antibodies SB-2, -3, -4, -5, -6, -7, -28, and -49 demonstrated high agonism in solution. In contrast, SIRPβ1 antibodies SB-14, -15, -17, -27, -41, -45, and -46 weakly activated SIRPβ1-mediated respiratory burst when in solution. In addition, primary monocytes were stimulated with anti-SIRPβ1 antibody adsorbed to 96-well plates at 10 μg / mL overnight at 37°C. Supernatant fractions were then collected and assayed for IL-8 release. SIRPβ1 antibodies SB-1, -2, -3, -7, -8, -9, -14, 28, and -49 demonstrated high agonism as plate-bound antibodies. In contrast, SIRPβ1 antibodies SB-4, -12, -23, -26, -33, -34, -35, -36, and -37 weakly activated SIRPβ1-mediated cytokine release. Example 6: SIRPβ1 increases the secretion of inflammatory cytokines from macrophages

[0264] Published literature describes bone marrow derived macrophages (BMDM) or primary peritoneal macrophages that possess altered TLR responses when DAP12 is deficient. To determine whether the SIRPβ1 antibodies of the present disclosure induce changes in inflammatory cytokine production, primary human monocyte-derived macrophages and dendritic cells were cultured with non-saturating levels of TLR stimulants in combination with plate-bound test antibodies and cytokine levels were measured after 24 hours. To generate monocyte-derived macrophages and dendritic cells, human primary monocytes were isolated from heparinized human blood (Blood Centers of the Pacific) using RosetteSep human monocyte enrichment mixture (STEMCELL Technologies) according to the manufacturer's protocol. Monocytes were seeded in RPMI (Invitrogen) containing 10% fetal bovine serum (Hyclone) and 50ng / ml M-CSF to induce macrophage differentiation. After 5-6 days, macrophages were harvested by scraping cells attached to plastic. Alternatively, monocytes were seeded in RPMI medium containing 10% fetal bovine serum (Hyclone) and 20 ng / ml IL-4 and GM-CSF (Peprotech) to induce differentiation of immature dendritic cells. After 6-7 days, dendritic cells were harvested by scraping cells attached to plastic. 5Macrophages or dendritic cells were plated on 96-well plates coated with the indicated antibodies at 10 cells / well and incubated for 24 hours at 37° C. Cells were co-stimulated with the TLR4 agonist LPS (Salmonella abortus equi).

[0265] like Figure 6C As shown in , monocyte-derived macrophages and dendritic cells were stimulated with 0.5 ng / mL LPS overnight at 37°C in the presence of anti-SIRPβ1 antibodies (SB-1, -2, -3, -4, -5, -6, -7, -8, -9, -11, -12, -14, -15, -16, -17, -18, -19, -20, -21, -28, -32, -40, and -49) adsorbed to 96-well plates at 10 μg / mL. The supernatant fractions were then collected and TNFα release was determined. As a negative control, DCs were stimulated with LPS in the presence of plate-bound anti-SIRPα antibody SA-56-90 (which suppresses TNFα release). In all experiments measuring respiratory burst, the generation of reactive oxygen species (ROS) was monitored by labeling cells with 2 μM fluorescent indicator CM-H2DCFDA. SIRPβ1 antibodies SB-1, -2, -6, -8, and -49 demonstrated high agonism as plate-bound antibodies. In contrast, most of the remaining SIRPβ1 antibodies weakly activated SIRPβ1-mediated cytokine release. Similarly, plate-bound SB-1 and SB-40, but not SB-32, enhanced LPS-induced TNFα release from monocyte-derived dendritic cells. In contrast, LPS-induced TNFα release was reduced in dendritic cells stimulated with plate-bound anti-SIRPα antibody SA-90, confirming that agonistic anti-SIRPβ1 antibodies activated cells, whereas agonistic SIRPα antibodies inhibited cell activity. Example 7: Anti-tumor activity of neutrophils stimulated by SIRPβ1

[0266] Although neutrophils are efficient phagocytic cells for antibody- or complement-opsonized target cells, in the context of the tumor microenvironment, neutrophils generally contribute toward tumor progression, invasion, and angiogenesis. However, recent publications suggest that tumor-associated neutrophils retain the potential to polarize toward an anti-tumor phenotype like other myeloid cells. Since neutrophils express high levels of SIRPβ1, anti-SIRPβ1 antibodies were evaluated for their ability to induce neutrophil-mediated tumor cell clearance in vitro. Primary neutrophils were isolated from peripheral blood of healthy donors obtained on the same day using the EasySepTM Direct Human Neutrophil Isolation Kit (STEMCELL) according to the manufacturer's instructions. The isolated human neutrophils were then added to a 96-well plate previously coated with 10 μg / mL anti-SIRPβ1 antibody or isotype control. Subsequently, Raji B-cell lymphoma cells engineered to stably express luciferase were mixed with neutrophils at a 1:1 ratio in the presence or absence of opsonic antibodies (anti-CD20 human IgG1). The co-cultured cells were incubated overnight at 37°C and viable Raji cells were quantified by measuring luciferase activity after adding OneGlo reagent (Promega) and incubating the samples on a plate shaker for 3 minutes at room temperature. TM 2.04 software for BioTek Synergy TM The luminescent signal was detected by a microplate reader.

[0267] like Figure 7 As shown in , human neutrophils failed to eliminate Raji cells when co-cultured in the presence of immobilized isotype control antibodies. Addition of anti-CD20 huIgG1 did not restore tumor cell clearance with unstimulated neutrophils. Similarly, neutrophils stimulated with immobilized anti-SIRPβ1 antibodies failed to significantly eliminate Raji cells. However, SIRPβ1-stimulated neutrophils eliminated anti-CD20-opsonized Raji cells. Agonistic anti-SIRPβ1 antibodies SB-1, SB-2, SB-3, and SB-4 reduced the luminescent signal from viable Raji cells by approximately 50% compared to isotype control-treated neutrophils. Anti-SIRPβ1 antibodies SB-28 and SB-49 exhibited weak activity against optimized Raji cells. These results suggest that agonistic anti-SIRPβ1 antibodies enhance the anti-tumor properties of neutrophils. Example 8: Affinity maturation of anti-SIRPβ1 antibodies

[0268] Five anti-SIRPβ1 antibodies, SB-1, SB-2, SB-8, and SB-40 (referred to as "parental" antibodies) with various physical and functional properties were subjected to affinity maturation. Briefly, a diverse antibody library was created in yeast for each starting parental antibody. Diversity was created by combining the parental heavy chain CDR-H3 and light chain (LC) with existing genetically diverse heavy chains (HC) in the CDR-H1 and CDR-H2 regions using standard molecular cloning techniques (referred to as "H1 / H2" optimization). This produced six libraries, roughly 10 5 1:755-768 for selecting antibodies with improved affinity. The selection pressure used to screen the library included human SIRPα and SIRPβ1 antigen equilibrium titration, parent antibody Fab competition kinetics, and the use of multispecific agents to select (as described in, for example, WO 2014 / 179363; Xu et al., Protein Eng Des Sel, 26(10):663-670). The antibodies were then visualized and selected using FACS flow cytometry using standard techniques (see, for example, Chao et al., Nature Protocols, 2006; 1:755-768). The desired population was then brought forward to additional rounds of selection. After 6 rounds of enrichment, the yeast cells were plated to obtain single antibody isolates, which were then generated and characterized as described in Example 1. 17 antibodies with improved affinity were thus obtained from 4 of the 5 starting parent antibodies. Antibody IgG and Fab production and purification

[0269] Yeast colonies were grown to saturation and then induced for 48 hours with shaking at 30° C. After induction, yeast cells were pelleted and supernatants were harvested for purification.

[0270] Purification was performed using a protein A column and immunoglobulins were eluted with acetic acid pH 2.0. Fab fragments were generated by papain digestion and purified on CaptureSelect IgG-CH1 affinity matrix (Life Technologies). Affinity determination

[0271] By using ForteBio and Meso Scale Discovery (MSD) instruments to measure K D The affinity of anti-SIRPβ1 antibody was determined by PCR. The results were generally as described previously (Estep et al., MAbs. 2013 Mar-Apr; 5(2): 270-8) and carried out at room temperature. Affinity measurement. Briefly, the following was performed Affinity measurements, i.e., IgG was loaded online onto the AHQ sensor. The sensor was equilibrated offline in assay buffer for 30 minutes and then monitored online for 60 seconds to establish a baseline. For affinity binding measurements, the IgG-loaded sensor was exposed to 100 nM antigen (human SIRPα or SIRPβ1 Fc fusion) for 3 minutes and then transferred to assay buffer for 3 minutes for dissociation rate measurements. Additional affinity binding was determined by loading biotinylated SIRPβ1 monomer onto the SA sensor and exposing it to 100 nM IgG in solution. Monovalent binding measurements were obtained by loading human SIRPβ1 Fc fusion antigen onto the AHQ sensor, followed by exposure to 100 nM anti-SIRPβ1 antibody Fab. Additional monovalent measurements were performed by loading biotinylated human SIRPβ1 monomer onto the SA sensor, followed by exposure to 100 nM Fab in solution. Kinetic data were fitted using a 1:1 binding model in the data analysis software provided by ForteBio.

[0272] For MSD-SET K D For measurement, solution equilibrium titration (SET) was performed in PBS + 0.1% IgG-free BSA (PBSF) with recombinant human SIRPβ1, kept constant at 100 pM and incubated with 3- to 5-fold serial dilutions of the antibody, starting at around 50 nM. Antibodies (20 nM in PBS) were coated onto standard binding MSD-ECL plates at 4°C overnight or at room temperature for 30 minutes. The plates were then blocked with 1% BSA for 30 minutes with shaking at 700 rpm, followed by three washes with wash buffer (PBSF + 0.05% Tween20). SET samples were applied and incubated on the plates for 150 seconds with shaking at 700 rpm, followed by one wash. Antigen captured on the plates was detected with 250 ng / ml thio-tagged streptavidin in PBSF by incubation on the plates for 3 minutes. The plates were washed three times with wash buffer and then read on an MSD Sector Imager2400 instrument using 1x read buffer T with surfactant. Percent free antigen was plotted as a function of titrated antibody in Prism and a quadratic equation was fitted to extract the KD. To improve throughput, a liquid handling robot was used throughout the MSD-SET experiments, including SET sample preparation.

[0273] Cellular binding affinity measurements were performed at 4°C using BWZ reporter cells expressing human SIRPα or SIRPβ1. Briefly, cells were harvested, washed in PBS and incubated with increasing concentrations of anti-SIRPβ1 antibodies or isotype controls. Antibodies were diluted in FACS buffer (PBS + 2% FBS). After incubation on ice for 30 minutes, cells were washed twice in FACS buffer and incubated on ice for 30 minutes with PE-conjugated anti-human secondary antibodies (BD Biosciences). Cells were then washed twice in 200 μl FACS buffer and subsequently analyzed on a FACS Canto screener (BD). Apparent K was determined by nonlinear curve fitting (Modified OneSiteTotal, Graph Pad Prism) D value. Anti-SIRPβ1 antibody selection

[0274] Affinity matured anti-SIRPβ1 antibody clones showing improved affinity compared to the respective parental antibodies were further characterized.After initial screening of all affinity matured antibody clones, clones of each parental antibody were selected for further analysis. Antibody heavy and light chain variable domain sequences

[0275] The amino acid sequences encoding the light chain variable and heavy chain variable domains of the affinity matured antibodies were determined using standard techniques. The Kabat light chain HVR sequences of the affinity matured antibodies are listed in Table 8. The Kabat heavy chain HVR sequences of the antibodies are listed in Table 9. The Kabat heavy chain framework (FR) sequences of the antibodies are listed in Table 10. The Kabat light chain framework (FR) sequences of the antibodies are listed in Table 11. Table 8: Kabat light chain HVR sequences of affinity matured anti-SIRPβ1 antibodies (p) indicates the parent antibody sequence Table 9: Ka...

Claims

1. An isolated antibody that binds to human SIRPβ1, wherein the antibody has at least one, at least two, at least three, at least four, or at least five properties selected from the following: a) binds to human SIRPβ1 isoform 1, but not to human SIRPα; b) binds to human SIRPβ1 isoform 1, but not to human SIRPγ; c) binds to human SIRPβ1 isoform 1, but not to human SIRPβ1 isoform 3; d) binds to human SIRPβ1 isoform 1, but not to mouse SIRPβ1; e) binds to human SIRPβ1 isoform 1, but not to cynomolgus monkey SIRPβ1 isoform 1; f) stimulating SIRPβ1 activity on CD14-positive monocytes in vitro and / or in vivo; g) inducing or enhancing the respiratory burst in immune cells, such as neutrophils and / or monocytes, in vitro and / or in vivo; h) inducing or increasing IL-8 expression in monocytes in vitro and / or in vivo; i) inducing or increasing TNFα expression in macrophages and / or dendritic cells in vitro and / or in vivo; j) inducing neutrophil-mediated phagocytosis, such as phagocytosis of tumor cells, in vitro and / or in vivo; k) enhancing neutrophil-mediated tumor cell clearance in vivo; 1) increasing TREM2 expression on macrophages in vitro and / or in vivo; m) increasing the viability of macrophages in vitro and / or in vivo when alone and / or in combination with an agonistic anti-TREM2 antibody; and n) Improving the survival of dendritic cells in vitro and / or in vivo.

2. The isolated antibody of claim 1, wherein the antibody binds to human SIRPβ1 isoform 1 but does not bind to human SIRPα or human SIRPβ1 isoform 3; or wherein the antibody binds to human SIRPβ1 isoform 1 but does not bind to human SIRPα, SIRPγ, or human SIRPβ1 isoform 3.

3. The isolated antibody of claim 1 or claim 2, wherein the antibody has at least one, at least two, or at least three properties selected from the group consisting of: a) stimulating SIRPβ1 activity on CD14-positive monocytes in vitro and / or in vivo; b) inducing or enhancing the respiratory burst in immune cells, such as neutrophils and / or monocytes, in vitro and / or in vivo; c) inducing or increasing IL-8 expression in monocytes in vitro and / or in vivo; d) inducing or increasing TNFα expression in macrophages and / or dendritic cells in vitro and / or in vivo; e) inducing neutrophil-mediated phagocytosis, such as phagocytosis of tumor cells, in vitro and / or in vivo; f) enhancing neutrophil-mediated tumor cell clearance in vivo; g) increasing TREM2 expression on macrophages in vitro and / or in vivo; h) increasing the viability of macrophages in vitro and / or in vivo when alone and / or in combination with an agonistic anti-TREM2 antibody; and i) Improving the survival of dendritic cells in vitro and / or in vivo.

4. An isolated antibody that binds to human SIRPβ1, wherein the antibody comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region and the light chain variable region comprise (a) HVR-H1 comprising the HVR-H1 amino acid sequence shown in Table 4 and / or Table 9; (b) HVR-H2 comprising the HVR-H2 amino acid sequence shown in Table 4 and / or Table 9; (c) HVR-H3 comprising the HVR-H3 amino acid sequence shown in Table 4 and / or Table 9; (d) HVR-L1 comprising the HVR-L1 amino acid sequence shown in Table 3 and / or Table 8; (e) HVR-L2 comprising the HVR-L2 amino acid sequence shown in Table 3 and / or Table 8; and (f) HVR-L3 comprising the HVR-L3 amino acid sequence shown in Table 3 and / or Table 8.

5. The antibody of claim 4, wherein the heavy chain variable region comprises one, two, three or four framework regions selected from VH FR1, VH FR2, VH FR3, and VH FR4 shown in Table 6.

6. The antibody of claim 4 or claim 5, wherein the light chain variable region comprises one, two, three or four framework regions selected from VL FR1, VL FR2, VL FR3, and VL FR4 shown in Table 5.

7. The antibody of any one of claims 1-6, wherein the antibody comprises SB-1, SB-2, SB-3, SB-4, SB-5, SB-6, SB-7, SB-8, SB-9, SB-10, SB-11, SB-12, SB-13, SB-14, SB-15, SB-16, SB-17, SB-18, SB-19, SB-20, SB-21, SB-22, SB-23, SB-24, SB-25, SB-26, SB-27, SB-28, SB-29, SB-30, SB-31, SB-32, SB-33, SB-34, SB-35, SB-36, SB-37, SB-38, SB-39, SB-40, SB-41, SB-42, SB-43, SB-44, SB-45, SB-46, SB-47, SB-48, SB-49, SB-50, SB-51, SB-52, SB-53, SB-54, SB-55, SB-56, SB-57, SB-58 -38,SB-39,SB-40,SB-41,SB-42,SB-43,SB-44,SB-45,SB-46,SB-47,SB-48,SB-49,SB-50,SB-1-2,SB-1-3,SB-1-4,SB-1-5,SB-2-7,SB-2-8,SB-2-9,SB- Antibodies to HVR-H1, HVR-H2, HVR-H3, HVR-L1, HVR-L2, and HVR-L3.

8. The antibody of any one of claims 1-7, wherein the antibody comprises a heavy chain variable region comprising a residue selected from the group consisting of SEQ ID NO: 268, 270, 272, 274, 276, 279, 281, 283, 285, 287, 289, 291, 293, 295, 297, 299, 301, 303, 305, 307, 309, 311, 313, 315, 317, 319, 321, 323, 325, 327, 329, 331, 333, 335, 337, 339, 341 1,343,345,347,349,351,353,355,357,359,361,363,365,366,367,368,369,371,372,373,374,375,376,377,378,379,380,381, or 382 is an amino acid sequence that is at least 90%, at least 95%, at least 97%, or at least 99% identical to the sequence.

9. The antibody of any one of claims 1-8, wherein the antibody comprises a moiety selected from the group consisting of SEQ ID NO: 268, 270, 272, 274, 276, 279, 281, 283, 285, 287, 289, 291, 293, 295, 297, 299, 301, 303, 305, 307, 309, 311, 313, 315, 317, 319, 321, 323, 325, 327, 329, 331, 333, or 382.

10. The antibody of any one of claims 1 to 9, wherein the antibody comprises a light chain variable region comprising a or 370. An amino acid sequence that is at least 90%, at least 95%, at least 97%, or at least 99% identical to the sequence of NO: 267, 269, 271, 273, 275, 277, 278, 280, 282, 284, 286, 288, 290, 292, 294, 296, 298, 300, 302, 304, 306, 308, 310, 312, 314, 316, 318, 320, 322, 324, 326, 328, 330, 332, 334, 336, 338, 340, 342, 344, 346, 348, 350, 352, 354, 356, 358, 360, 362, 364, or 370.

Images