Antibodies against signal-regulatory protein α and methods of use

1. An isolated antibody that binds an extracellular domain of a human SIRP-α polypeptide, wherein the antibody comprises:

(a) a heavy chain comprising a heavy chain variable (VH) domain that comprises the amino acid sequence of SEQ ID NO:26; and

(b) a light chain comprising a light chain variable (VL) domain that comprises an amino acid sequence according to the formula SYELTQPPSVSVSPGQTARITCSGGSYSSYYYAWYQQKPGQAPVTLIYSDDKRPSNIPER FSGSSSGTTVTLTISGVQAEDEADYYCGGYDQSSYTNPFGX 1 GTX 2 X 3 TVL (SEQ ID NO:71), wherein X 1 is G or T; X 2 is K, Q, or R; and X 3 is L or V, and wherein the VL domain does not comprise the sequence of SEQ ID NO:25.

2. The antibody of claim 1 , wherein the VL domain comprises an amino acid sequence selected from the group consisting of SEQ ID NOs:39-41.

3. The antibody of claim 1 , wherein the light chain further comprises a light chain constant (CL) domain sequence that comprises an amino acid sequence according to the formula GQPKANPTVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADGSPVKAGVETTKPS KQSX 4 X 5 KYAASSYLSLTPEQWKSHRSYSCQVTHEGSTVEKTVAPTECS (SEQ ID NO:72), wherein X 4 X 5 is ND, DN, DS, or SD.

4. The antibody of claim 1 , wherein the light chain further comprises a light chain constant (CL) domain sequence that comprises an amino acid sequence selected from the group consisting of SEQ ID NOs:43-46.

5. The antibody of claim 1 , wherein the light chain further comprises a kappa light chain constant (CL) domain.

6. The antibody of claim 5 , wherein the light chain comprises the amino acid sequence of SEQ ID NO:36.

7. The antibody of claim 1 , wherein the light chain further comprises a lambda light chain constant (CL) domain.

8. The antibody of claim 7 , wherein the light chain comprises the amino acid sequence of SEQ ID NO:37 or SEQ ID NO:38.

9. The antibody of claim 1 , wherein the light chain comprises an amino acid sequence selected from the group consisting of SEQ ID NOs:48-57.

10. The antibody of claim 1 , wherein the antibody is a scFv-Fc antibody.

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

12. The antibody of claim 1 , wherein the heavy chain comprises a heavy chain constant domain that comprises an Fc region.

13. The antibody of claim 12 , wherein the Fc region is a human Fc region selected from the group consisting of an IgG1 Fc region, an IgG2 Fc region, and an IgG4 Fc region.

14. The antibody of claim 12 , wherein the Fc region is a human IgG1 Fc region.

15. The antibody of claim 12 , wherein the Fc region is a human IgG1 Fc region comprising L234A, L235A, and G237A substitutions, amino acid position numbering according to EU.

16. The antibody of claim 15 , wherein the Fc region further comprises an N297A substitution, amino acid position numbering according to EU.

17. The antibody of claim 12 , wherein the Fc region is a human IgG2 Fc region.

18. The antibody of claim 12 , wherein the Fc region is a human IgG2 Fc region comprising A330S and P331S substitutions, amino acid position numbering according to EU.

19. The antibody of claim 17 , wherein the Fc region comprises an N297A substitution, amino acid position numbering according to EU.

20. The antibody of claim 12 , wherein the Fc region is a human IgG4 Fc region, and wherein the heavy chain comprises an S228P substitution, amino acid position numbering according to EU.

21. The antibody of claim 12 , wherein the heavy chain constant domain comprises an amino acid sequence selected from the group consisting of SEQ ID NOs:31-35.

22. The antibody of claim 12 , wherein the heavy chain constant domain comprises an amino acid sequence selected from the group consisting of SEQ ID NOs:33, 34, and 137.

23. The antibody of claim 12 , wherein the heavy chain constant domain comprises an amino acid sequence selected from the group consisting of SEQ ID NOs:132-139.

24. The antibody of claim 1 , wherein the heavy chain comprises an amino acid sequence selected from the group consisting of SEQ ID NOs:58-62.

25. The antibody of claim 1 , wherein the heavy chain comprises an amino acid sequence selected from the group consisting of SEQ ID NOs:60, 61, and 129.

26. The antibody of claim 1 , wherein:

(a) the heavy chain comprises the amino acid sequence of SEQ ID NO:62, and the light chain comprises the amino acid sequence of SEQ ID NO:52;

(b) the heavy chain comprises the amino acid sequence of SEQ ID NO:62, and the light chain comprises the amino acid sequence of SEQ ID NO:53;

(c) the heavy chain comprises the amino acid sequence of SEQ ID NO:62, and the light chain comprises the amino acid sequence of SEQ ID NO:54;

(d) the heavy chain comprises the amino acid sequence of SEQ ID NO:62, and the light chain comprises the amino acid sequence of SEQ ID NO:55;

(e) the heavy chain comprises the amino acid sequence of SEQ ID NO:62, and the light chain comprises the amino acid sequence of SEQ ID NO:56; or (f) the heavy chain comprises the amino acid sequence of SEQ ID NO:62, and the light chain comprises the amino acid sequence of SEQ ID NO:57.

27. The antibody of claim 1 , wherein:

(a) the heavy chain comprises the amino acid sequence of SEQ ID NO:58, and the light chain comprises the amino acid sequence of SEQ ID NO:52;

(b) the heavy chain comprises the amino acid sequence of SEQ ID NO:59, and the light chain comprises the amino acid sequence of SEQ ID NO:52;

(c) the heavy chain comprises the amino acid sequence of SEQ ID NO:60, and the light chain comprises the amino acid sequence of SEQ ID NO:52;

(d) the heavy chain comprises the amino acid sequence of SEQ ID NO:61, and the light chain comprises the amino acid sequence of SEQ ID NO:52;

(e) the heavy chain comprises the amino acid sequence of SEQ ID NO:58, and the light chain comprises the amino acid sequence of SEQ ID NO:53;

(f) the heavy chain comprises the amino acid sequence of SEQ ID NO:59, and the light chain comprises the amino acid sequence of SEQ ID NO:53;

(g) the heavy chain comprises the amino acid sequence of SEQ ID NO:60, and the light chain comprises the amino acid sequence of SEQ ID NO:53;

(h) the heavy chain comprises the amino acid sequence of SEQ ID NO:61, and the light chain comprises the amino acid sequence of SEQ ID NO:53;

(i) the heavy chain comprises the amino acid sequence of SEQ ID NO:58, and the light chain comprises the amino acid sequence of SEQ ID NO:54;

(j) the heavy chain comprises the amino acid sequence of SEQ ID NO:59, and the light chain comprises the amino acid sequence of SEQ ID NO:54;

(k) the heavy chain comprises the amino acid sequence of SEQ ID NO:60, and the light chain comprises the amino acid sequence of SEQ ID NO:54;

(l) the heavy chain comprises the amino acid sequence of SEQ ID NO:61, and the light chain comprises the amino acid sequence of SEQ ID NO:54;

(m) the heavy chain comprises the amino acid sequence of SEQ ID NO:58, and the light chain comprises the amino acid sequence of SEQ ID NO:55;

(n) the heavy chain comprises the amino acid sequence of SEQ ID NO:59, and the light chain comprises the amino acid sequence of SEQ ID NO:55;

(o) the heavy chain comprises the amino acid sequence of SEQ ID NO:60, and the light chain comprises the amino acid sequence of SEQ ID NO:55;

(p) the heavy chain comprises the amino acid sequence of SEQ ID NO:61, and the light chain comprises the amino acid sequence of SEQ ID NO:55;

(q) the heavy chain comprises the amino acid sequence of SEQ ID NO:58, and the light chain comprises the amino acid sequence of SEQ ID NO:56;

(r) the heavy chain comprises the amino acid sequence of SEQ ID NO:59, and the light chain comprises the amino acid sequence of SEQ ID NO:56;

(s) the heavy chain comprises the amino acid sequence of SEQ ID NO:60, and the light chain comprises the amino acid sequence of SEQ ID NO:56;

(t) the heavy chain comprises the amino acid sequence of SEQ ID NO:61, and the light chain comprises the amino acid sequence of SEQ ID NO:56;

(u) the heavy chain comprises the amino acid sequence of SEQ ID NO:58, and the light chain comprises the amino acid sequence of SEQ ID NO:57;

(v) the heavy chain comprises the amino acid sequence of SEQ ID NO:59, and the light chain comprises the amino acid sequence of SEQ ID NO:57;

(w) the heavy chain comprises the amino acid sequence of SEQ ID NO:60, and the light chain comprises the amino acid sequence of SEQ ID NO:57; or

(x) the heavy chain comprises the amino acid sequence of SEQ ID NO:61, and the light chain comprises the amino acid sequence of SEQ ID NO:57.

28. The antibody of claim 1 , wherein:

(a) the heavy chain comprises the amino acid sequence of SEQ ID NO:60, and the light chain comprises the amino acid sequence of SEQ ID NO:55;

(b) the heavy chain comprises the amino acid sequence of SEQ ID NO:61, and the light chain comprises the amino acid sequence of SEQ ID NO:55;

(c) the heavy chain comprises the amino acid sequence of SEQ ID NO:129, and the light chain comprises the amino acid sequence of SEQ ID NO:55;

(d) the heavy chain comprises the amino acid sequence of SEQ ID NO:124, and the light chain comprises the amino acid sequence of SEQ ID NO:52; or

(e) the heavy chain comprises the amino acid sequence of SEQ ID NO:124, and the light chain comprises the amino acid sequence of SEQ ID NO:55.

29. The antibody of claim 1 , wherein the antibody enhances phagocytosis by a macrophage expressing a human SIRP-α polypeptide.

30. The antibody of claim 1 , wherein the antibody enhances activation of a dendritic cell expressing a human SIRP-α polypeptide.

31. The antibody of claim 1 , wherein the antibody inhibits in vivo growth of a tumor that expresses CD47.

32. The antibody of claim 1 , wherein the antibody is conjugated to an agent.

33. The antibody of claim 1 , wherein the antibody is a bispecific antibody.

34. The antibody of claim 33 , wherein the antibody comprises a first antigen binding domain that binds an extracellular domain of a human SIRP-α polypeptide and a second antigen binding domain that binds an antigen expressed by a cancer cell.

35. The antibody of claim 34 , wherein the antigen expressed by the cancer cell is selected from the group consisting of CD19, CD20, CD22, CD30, CD33, CD38, CD52, CD56, CD70, CD74, CD79b, CD123, CD138, CS1/SLAMF7, Trop-2, 5T4, EphA4, BCMA, PD-L1, PTK7, STEAP1, Endothelin B Receptor, mesothelin, ENPP3, SLC44A4, GNMB, nectin 4, NaPi2b, LIV-1A, Guanylyl cyclase C, DLL3, EGFR, HER2, VEGF, VEGFR, integrin αVβ3, integrin α5β1, MET, IGF1R, TRAILR1, TRAILR2, RANKL, FAP, Tenascin, Le y , EpCAM, CEA, gpA33, PSMA, TAG72, a mucin, CAIX, EPHA3, folate receptor α, GD2, and GD3.

36. The antibody of claim 33 , wherein the antibody comprises a first antigen binding domain that binds an extracellular domain of a human SIRP-α polypeptide and a second antigen binding domain that binds an antigen expressed by an immune cell.

37. The antibody of claim 36 , wherein the antigen expressed by the immune cell is selected from the group consisting of BDCA2, BDCA4, ILT7, LILRB1, LILRB2, LILRB3, LILRB4, LILRB5, Siglec-3, Siglec-7, Siglec-9, Siglec-15, FGL-1, CD200, CD200R, CSF-1R, CD40, CD40L, CD163, CD206, DEC205, CD47, CD123, arginase, IDO, TDO, AhR, EP2, COX-2, CCR2, CCR-7, CXCR1, CX3CR1, CXCR2, CXCR3, CXCR4, CXCR7, TGF-βRI, TGF-βRII, c-Kit, CD244, L-selectin/CD62L, CD11b, CD11c, CD68, 41BB, CTLA4, PD1, PD-L1, PD-L2, TIM-3, BTLA, VISTA, LAG-3, CD28, OX40, GITR, CD137, CD27, HVEM, CCR4, CD25, CD103, KIrg1, Nrp1, CD278, Gpr83, TIGIT, CD154, CD160, TNFR2, PVRIG, DNAM, and ICOS.

38. The antibody of claim 33 , wherein the antibody comprises a first antigen binding domain that binds an extracellular domain of a human SIRP-α polypeptide and a second antigen binding domain that binds an antigen expressed by a natural killer (NK) cell.

39. The antibody of claim 38 , wherein the antigen expressed by the NK cell is selected from the group consisting of NKR-P1A, CD94, KLRG1, KIR2DL5A, KIR2DL5B, KIR2DL1, KIR2DL2, KIR2DL3, KIR2DS2, KIR2DS3, KIR2DS4, KIR2DS5, KIR3DS1, KIR2DS1, CD94, NKG2D, CD160, CD16, NKp46, NKp30, NKp44, DNAM1, CRTAM, CD27, NTB-A, PSGL1, CD96, CD100, NKp80, SLAMF7, and CD244.

40. A method of treating or delaying progression of cancer in an individual, the method comprising administering to the individual an effective amount of the antibody of claim 1 .

41. The method of claim 40 , further comprising administering to the individual an effective amount of a second antibody that binds an antigen expressed by the cancer.

42. The method of claim 41 , wherein the antigen expressed by the cancer is selected from the group consisting of CD19, CD20, CD22, CD30, CD33, CD38, CD52, CD56, CD70, CD74, CD79b, CD123, CD138, CS1/SLAMF7, Trop-2, 5T4, EphA4, BCMA, PTK7, STEAP1, Endothelin B Receptor, mesothelin, ENPP3, SLC44A4, GNMB, nectin 4, NaPi2b, LIV-1A, Guanylyl cyclase C, DLL3, EGFR, HER2, VEGF, VEGFR, integrin αVβ3, integrin α5β1, MET, IGF1R, TRAILR1, TRAILR2, RANKL, FAP, Tenascin, Le y , EpCAM, CEA, gpA33, PSMA, TAG72, a mucin, CAIX, EPHA3, folate receptor α, GD2, and GD3.

43. The method of claim 40 , further comprising administering to the individual an effective amount of an immunotherapeutic agent.

44. The method of claim 43 , wherein the immunotherapeutic agent comprises a second antibody, vaccine, oncolytic virus, adoptive cell therapy, cytokine, or small molecule agent.

45. The method of claim 44 , wherein the second antibody binds to an antigen selected from the group consisting of BDCA2, BDCA4, ILT7, LILRB1, LILRB2, LILRB3, LILRB4, LILRB5, Siglec-3, Siglec-7, Siglec-9, Siglec-15, FGL-1, CD200, CD200R, CSF-1R, CD40, CD40L, CD163, CD206, DEC205, CD47, CD123, arginase, IDO, TDO, AhR, EP2, COX-2, CCR2, CCR-7, CXCR1, CX3CR1, CXCR2, CXCR3, CXCR4, CXCR7, TGF-β RI, TGF-β RII, c-Kit, CD244, L-selectin/CD62L, CD11b, CD11c, CD68, 41BB, CTLA4, PD1, PD-L1, PD-L2, TIM-3, BTLA, VISTA, LAG-3, CD28, OX40, GITR, CD137, CD27, HVEM, CCR4, CD25, CD103, KIrg1, Nrp1, CD278, Gpr83, TIGIT, CD154, CD160, TNFR2, PVRIG, DNAM, and ICOS.

46. The method of claim 45 , wherein the second antibody binds to PD-1.

47. The method of claim 45 , wherein the second antibody binds to PD-L1.

48. The method of claim 40 , further comprising administering to the individual an effective amount of a second antibody that binds an antigen expressed by a natural killer (NK) cell.

49. The method of claim 48 , wherein the antigen expressed by the NK cell is selected from the group consisting of NKR-P1A, CD94, KLRG1, KIR2DL5A, KIR2DL5B, KIR2DL1, KIR2DL2, KIR2DL3, KIR2DS2, KIR2DS3, KIR2DS4, KIR2DS5, KIR3DS1, KIR2DS1, CD94, NKG2D, CD160, CD16, NKp46, NKp30, NKp44, DNAM1, CRTAM, CD27, NTB-A, PSGL1, CD96, CD100, NKp80, SLAMF7, and CD244.

50. The method of claim 40 , further comprising administering to the individual an effective amount of a chemotherapeutic agent or small molecule anti-cancer agent.

51. The method of claim 40 , further comprising administering to the individual an effective amount of a targeted small molecule inhibitor.

52. The method of claim 51 , wherein the targeted small molecule inhibitor is a VEGFR and/or PDGFR inhibitor, EGFR inhibitor, ALK inhibitor, CDK4/6 inhibitor, PARP inhibitor, mTOR inhibitor, KRAS inhibitor, TRK inhibitor, BCL2 inhibitor, IDH inhibitor, PI3K inhibitor, DNA damage response (DDR) inhibitor, or hypomethylation agent.

53. The method of claim 35 , wherein the mucin is Mucin 1 or Mucin 16.

54. The method of claim 35 , wherein the EGFR is EGFRvIII.

55. The method of claim 34 , wherein the antigen expressed by the cancer cell is an WIC/peptide complex comprising a peptide from NY-ESO-1/LAGE, SSX-2, a MAGE family protein, gp100/pmel17, Melan-A/MART-1, a tyrosinase, CEA, PSA, TAG-72, immature laminin receptor, MOK/RAGE-1, WT-1, SAP-1, BING-4, EpCAM, MUC1, PRAME, survivin, BRCA1, BRCA2, CDK4, CML66, MART-2, p53, Ras, β-catenin, TGF-βRII, HPV E6, or HPV E7.

56. The method of claim 55 , wherein the MAGE family protein is MAGE-A3.

57. The method of claim 55 , wherein the tyrosinase is gp75/TRP1 or TRP2.

58. The method of claim 42 , wherein the mucin is Mucin 1 or Mucin 16 .

59. The method of claim 42 , wherein the EGFR is EGFRvIII.

60. The method of claim 36 , wherein the antigen expressed by the cancer cell is an MHC/peptide complex comprising a peptide from NY-ESO-1/LAGE, SSX-2, a MAGE family protein, gp100/pmel17, Melan-A/MART-1, a tyrosinase, CEA, PSA, TAG-72, immature laminin receptor, MOK/RAGE-1, WT-1, SAP-1, BING-4, EpCAM, MUC1, PRAME, survivin, BRCA1, BRCA2, CDK4, CML66, MART-2, p53, Ras, β-catenin, TGF-βRII, HPV E6, or HPV E7.

61. The method of claim 60 , wherein the MAGE family protein is MAGE-A3.

62. The method of claim 60 , wherein the tyrosinase is gp75/TRP1 or TRP2.