C-kit binding proteins, chimeric antigen receptors, and uses thereof

Single-domain antibodies and CAR-expressing immune cells address the lack of specificity and efficacy in existing therapies for c-Kit-related diseases, enhancing treatment and diagnosis through targeted binding and therapeutic conjugates.

HK40135201APending Publication Date: 2026-07-17TENEOBIO INC
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Patent Information

Authority / Receiving Office
HK · HK
Patent Type
Applications
Current Assignee / Owner
TENEOBIO INC
Filing Date
2026-06-08
Publication Date
2026-07-17

AI Technical Summary

Technical Problem

Current therapies for diseases associated with c-Kit expression lack specificity and efficacy, particularly in targeting c-Kit and its binding proteins, necessitating improved therapeutic and diagnostic agents.

Method used

Development of single-domain antibodies, anti-c-Kit antibodies and antibody fragments, antibody-drug conjugates, and chimeric antigen receptors (CARs) that specifically bind to c-Kit and its binding proteins, along with immune cells expressing CARs, to enhance treatment and diagnostic capabilities.

Benefits of technology

These agents provide targeted and effective treatment and diagnostic options for diseases related to c-Kit expression, improving therapeutic outcomes and diagnostic precision.

✦ Generated by Eureka AI based on patent content.

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Abstract

Single domain antibodies which specifically bind to c‑Kit and c‑Kit binding proteins, anti-c‑Kit antibodies and antibody fragments thereof, antibody‑drug conjugates, diagnostic agents, and chimeric antigen receptors (CARs) comprising the same are disclosed, along with CAR-expressing immune cells. Also disclosed are pharmaceutical compositions comprising any of the foregoing, uses of any of the foregoing in the treatment and / or diagnosis and / or monitoring of a disease associated with c-Kit expression, and uses of any of the foregoing in a stem cell transplantation preconditioning regimen.
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Description

Abstract This invention discloses single-domain antibodies that specifically bind to c-Kit and its binding proteins, anti-c-Kit antibodies and antibody fragments thereof, antibody-drug conjugates, diagnostic reagents, and chimeric antigen receptors (CARs) comprising the above substances, as well as immune cells expressing CARs. This invention also discloses pharmaceutical compositions comprising any of the above substances, the use of any of the above substances in the treatment and / or diagnosis and / or monitoring of diseases associated with c-Kit expression, and the use of any of the above substances in stem cell transplantation pretreatment protocols.

Claims

What is claimed is:

1. A single domain antibody which specifically binds to c-Kit, wherein the single domain antibody comprises a heavy chain variable (VH) region in which the full set of VH CDRs 1, 2, and 3 (combined) has at least 95% sequence identity to the VH CDRs 1, 2, and 3 of any one of SEQ ID NOs: 14-18.

2. A single domain antibody which specifically binds to c-Kit, wherein the single domain antibody comprises a heavy chain variable (VH) region comprising:(a) (i) a VH complementarity determining region one (CDR1) comprising a sequence having at most two amino acid modifications relative to SEQ ID NO: 1, SEQ ID NO: 2, or SEQ ID NO: 3;(ii) a VH CDR2 comprising a sequence having at most two amino acid modifications relative to SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, or SEQ ID NO: 7; and(iii) a VH CDR3 comprising a sequence having at most two amino acid modifications relative to SEQ ID NO: 8, SEQ ID NO: 9, or SEQ ID NO: 10; or(b) a VH CDR1 comprising a sequence having at most two amino acid modifications relative to SEQ ID NO: 11; a VH CDR2 comprising a sequence having at most two amino acid modifications relative to SEQ ID NO: 12; and a VH CDR3 comprising a sequence having at most two amino acid modifications relative to SEQ ID NO: 13.

3. A single domain antibody which specifically binds to c-Kit, wherein the single domain antibody comprises a heavy chain variable (VH) region comprising:(i) a VH complementarity determining region one (CDR1) comprising the sequenceG Xi T X2V X4Y A (SEQ ID NO: 53), wherein Xi is F or L; X2 is F or I; X3 is D or S; and X4 is S or T;(ii) a VH CDR2 comprising the sequenceI S X5Xe G X7X8T (SEQ ID NO: 71), wherein X5 is V or T, Xe is R or G; X7 is G or S; and Xs is S or R; and(iii) a VH CDR3 comprising the sequenceA T G Y D X9 S G X10 Y Y G G F D Y (SEQ ID NO: 54), wherein X9 is S or P; and X10 is Y or H.

4. A single domain antibody which specifically binds to c-Kit, wherein the single domain antibody comprises a heavy chain variable (VH) region comprising:(i) a VH complementarity determining region one (CDR1) comprising a sequence chosen from SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3;(ii) a VH CDR2 comprising a sequence chosen from SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, and SEQ ID NO: 7; and(iii) a VH CDR3 comprising a sequence chosen from SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10.

5. A single domain antibody which specifically binds to c-Kit, wherein the single domain antibody comprises a heavy chain variable (VH) region comprising:(a) a VH CDR1, a VH CDR2, and a VH CDR3 comprising the sequences of SEQ ID NOs: 1, 4, and 8, respectively;(b) a VH CDR1, a VH CDR2, and a VH CDR3 comprising the sequences of SEQ ID NOs: 2, 5, and 9, respectively;(c) a VH CDR1, a VH CDR2, and a VH CDR3 comprising the sequences of SEQ ID NOs: 3, 6, and 8, respectively;(d) a VH CDR1, a VH CDR2, and a VH CDR3 comprising the sequences of SEQ ID NOs: 2, 7, and 10, respectively; or(e) a VH CDR1, a VH CDR2, and a VH CDR3 comprising the sequences of SEQ ID NOs: 11, 12, and 13, respectively.

6. A single domain antibody which specifically binds to c-Kit, wherein the single domain antibody comprises a heavy chain variable (VH) region comprising the VH CDR1, VH CDR2, and VH CDR3 of any one of SEQ ID NOs: 14-18.

7. The single domain antibody according to any one of claims 1 to 6, wherein the VH CDR1, VH CDR2, and VH CDR3 sequences are present in a human VH framework.

8. A single domain antibody which specifically binds to c-Kit, wherein the single domain antibody comprises a heavy chain variable (VH) region having at least 95% sequence identity to any one of SEQ ID NOs: 14-18.

9. A single domain antibody which specifically binds to c-Kit, wherein the single domain antibody comprises a heavy chain variable (VH) region chosen from SEQ ID NOs: 14-18.

10. The single domain antibody according to any one of claims 1 to 9, wherein the single domain antibody specifically binds to human c-Kit.

11. The single domain antibody according to any one of claims 1 to 10, wherein the single domain antibody binds to human c-Kit with a KD of from about 10'9M to about 10'6M.

12. The single domain antibody according to any one of claims 1 to 11, wherein the single domain antibody is an isolated single domain antibody.

13. A c-Kit binding protein comprising the single domain antibody according to any one of claims 1 to 11.

14. The c-Kit binding protein according to claim 13, wherein the c-Kit binding protein specifically binds to human c-Kit.

15. The c-Kit binding protein according to claim 13 or 14, wherein the c-Kit binding protein further specifically binds to CD3.

16. The c-Kit binding protein according to any one of claims 13 to 15, wherein the c-Kit binding protein further specifically binds to human CD3.

17. The c-Kit binding protein according to any one of claims 13 to 16, wherein the c-Kit binding protein is an isolated monoclonal antibody.

18. An antibody-drug conjugate comprising the single domain antibody according to any one of claims 1 to 11.

19. An anti-c-Kit antibody comprising the single domain antibody according to any one of claims 1 to 11.

20. The anti-c-Kit antibody according to claim 19, wherein the anti-c-Kit antibody is multi-specific.

21. The anti-c-Kit antibody according to claim 19 or 20, wherein the anti-c-Kit antibody further specifically binds to a tumor-specific antigen other than c-Kit.

22. The anti-c-Kit antibody according to any one of claims 19 to 21, wherein the anti-c-Kit antibody is bispecific.

23. The anti-c-Kit antibody according to any one of claims 19 to 22, wherein the anti-c-Kit antibody further specifically binds to CD3.

24. The anti-c-Kit antibody according to any one of claims 19 to 23, wherein the anti-c-Kit antibody further specifically binds to human CD3.

25. The anti-c-Kit antibody according to any one of claims 19 to 24, wherein the anti-c-Kit antibody further comprises a CD3-binding heavy chain variable (VH) region.

26. The anti-c-Kit antibody according to any one of claims 19 to 25, wherein the anti-c-Kit antibody is of the IgGl or IgG4 subtype.

27. The anti-c-Kit antibody according to any one of claims 19 to 26, wherein the anti-c-Kit antibody further comprises a CD3-binding heavy chain variable (VH) region that is paired with a light chain variable (VL) region.

28. The anti-c-Kit antibody according to claim 25 or 27, wherein the CD3 -binding VH region comprises:(i) a VH complementarity determining region one (CDR1) comprising a sequence having at most two amino acid modifications relative to any one of SEQ ID NOs: 20-25;(ii) a VH CDR2 comprising a sequence having at most two amino acid modifications relative to SEQ ID NO: 26; and(iii) a VH CDR3 comprising a sequence having at most two amino acid modifications relative to any one of SEQ ID NOs: 27-30.

29. The anti-c-Kit antibody according to claim 28, wherein the CD3-binding VH CDR1 comprises a sequence chosen from SEQ ID NOs: 20-25.

30. The anti-c-Kit antibody according to claim 28 or 29, wherein the CD3-binding VH CDR2 comprises the sequence of SEQ ID NO: 26.

31. The anti-c-Kit antibody according to any one of claims 28 to 30, wherein the CD3-binding VH CDR3 comprises a sequence chosen from SEQ ID NOs: 27-30.

32. The anti-c-Kit antibody according to any one of claims 25 or 27 to 31, wherein the full set of VH CDRs 1, 2, and 3 (combined) in the CD3 -binding VH region has at least 95% sequence identity to the VH CDRs 1, 2, and 3 of any one of SEQ ID NOs: 31-48.

33. The anti-c-Kit antibody according to claim 25 or 27, wherein the CD3-binding VH region comprises:(i) a VH complementarity determining region one (CDR1) comprising the sequenceG F T F Xu X12 Y A (SEQ ID NO: 55), wherein Xu is D, A, or H; and X12 is D or N;(ii) a VH CDR2 comprising the sequence ISWNSGSI (SEQ ID NO: 26); and(iii) a VH CDR3 comprising the sequenceA K D S R G Y G X13 Y X14 X15 G G A Y (SEQ ID NO: 56), wherein X13 is D or S; X14 is R or S; and X15 is L or R.

34. The anti-c-Kit antibody according to claim 25 or 27, wherein the CD3-binding VH region comprises the VH CDR1, VH CDR2, and VH CDR3 of any one of SEQ IDNOs: 31-48.

35. The anti-c-Kit antibody according to any one of claims 25 to 34, wherein the VH CDR1, VH CDR2, and VH CDR3 sequences in the CD3-binding VH region are present in a human VH framework.

36. The anti-c-Kit antibody according to any one of claims 25 to 35, wherein the CD3 -binding VH region has at least 95% sequence identity to any one of SEQ ID NOs: 31-48.

37. The anti-c-Kit antibody according to any one of claims 25 to 36, wherein the CD3-binding VH region comprises:(a) a VH CDR1, a VH CDR2, and a VH CDR3 comprising the sequences of SEQ ID NOs: 20, 26, and 27, respectively;(b) a VH CDR1, a VH CDR2, and a VH CDR3 comprising the sequences of SEQ ID NOs: 20, 26, and 28, respectively;(c) a VH CDR1, a VH CDR2, and a VH CDR3 comprising the sequences of SEQ ID NOs: 20, 26, and 29, respectively;(d) a VH CDR1, a VH CDR2, and a VH CDR3 comprising the sequences of SEQ ID NOs: 21, 26, and 28, respectively;(e) a VH CDR1, a VH CDR2, and a VH CDR3 comprising the sequences of SEQ ID NOs: 22, 26, and 28, respectively;(f) a VH CDR1, a VH CDR2, and a VH CDR3 comprising the sequences of SEQ ID NOs: 23, 26, and 28, respectively;(g) a VH CDR1, a VH CDR2, and a VH CDR3 comprising the sequences of SEQ ID NOs: 24, 26, and 28, respectively;(h) a VH CDR1, a VH CDR2, and a VH CDR3 comprising the sequences of SEQ ID NOs: 20, 26, and 30, respectively;(i) a VH CDR1, a VH CDR2, and a VH CDR3 comprising the sequences of SEQ ID NOs: 25, 26, and 29, respectively; or(j) a VH CDR1, a VH CDR2, and a VH CDR3 comprising the sequences of SEQ ID NOs: 24, 26, and 29, respectively.

38. The anti-c-Kit antibody according to claim 37, wherein the CD3 -binding VH region comprises a VH CDR1, a VH CDR2, and a VH CDR3 comprising the sequences of SEQ ID NOs: 20, 26, and 27, respectively.

39. The anti-c-Kit antibody according to any one of claims 27 to 38, wherein the light chain variable region comprises the VL CDR1, VL CDR2, and VL CDR3 of SEQ ID NO: 52.

40. The anti-c-Kit antibody according to any one of claims 27 to 39, wherein the light chain variable region comprises a VL CDR1, a VL CDR2, and a VL CDR3 comprising the sequences of SEQ ID NOs: 49, 50, and 51, respectively.

41. The anti-c-Kit antibody according to any one of claims 27 to 40, wherein the VL CDR1, VL CDR2, and VL CDR3 sequences are present in a human VH framework.

42. The anti-c-Kit antibody according to any one of claims 27 to 41, wherein the light chain variable region has at least 95% sequence identity to SEQ ID NO: 52.

43. The anti-c-Kit antibody according to any one of claims 19 to 42, wherein the anti-c-Kit antibody specifically binds to human c-Kit.

44. The anti-c-Kit antibody according to any one of claims 19 to 43, wherein the anti-c-Kit antibody is an isolated antibody.

45. An antibody fragment that specifically binds to c-Kit, wherein the antibody fragment comprises a fragment of the anti-c-Kit antibody according to any one of claims 19 to 44.

46. The antibody fragment according to claim 45, wherein the antibody fragment is a three-chain antibody-like molecule.

47. A chimeric antigen receptor (CAR)-expressing immune cell comprising a CAR that comprises an extracellular antigen-binding domain which specifically binds to c-Kit, wherein the extracellular antigen-binding domain comprises the single domain antibody according to any one of claims 1 to 11.

48. The CAR-expressing immune cell according to claim 47, wherein the single domain antibody comprises a heavy chain variable (VH) region comprising a VH CDR1, a VH CDR2, and a VH CDR3 comprising the sequences of SEQ ID NOs: 2, 5, and 9, respectively.

49. The CAR-expressing immune cell according to claim 47 or claim 48, wherein the single domain antibody comprises a heavy chain variable (VH) region of SEQ ID NO: 15.

50. The CAR-expressing immune cell according to claim 47, wherein the single domain antibody comprises a heavy chain variable (VH) region comprising a VH CDR1, a VH CDR2, and a VH CDR3 comprising the sequences of SEQ ID NOs: 11, 12, and 13, respectively.

51. The CAR-expressing immune cell according to claim 47 or claim 50, wherein the single domain antibody comprises a heavy chain variable (VH) region of SEQ ID NO: 18.

52. The CAR-expressing immune cell according to any one of claims 47 to 51, wherein the CAR further comprises a transmembrane domain and an intracellular signaling domain.

53. The CAR-expressing immune cell according to claim 52, wherein the CAR further comprises a hinge between the extracellular antigen binding domain and the transmembrane domain.

54. The CAR-expressing immune cell according to any one of claims 47 to 53, wherein the immune cell is chosen from T cells and natural killer (NK) cells.

55. A pharmaceutical composition comprising at least one c-Kit binding protein, antibody-drug conjugate, anti-c-Kit antibody, antibody fragment, or CAR-expressing immune cell according to any one of claims 13 to 54 and a pharmaceutically acceptable excipient.

56. A polynucleotide encoding: the single domain antibody according to any one of claims 1 to 12; or a chimeric antigen receptor comprising an extracellular antigen-binding domain which specifically binds to c-Kit, wherein the extracellular antigen-binding domain comprises the single domain antibody according to any one of claims 1 to 11.

57. A composition comprising one or more polynucleotide(s) encoding the c-Kit binding protein, anti-c-Kit antibody, or antibody fragment according to any one of claims 13 to 17 or 19 to 46.

58. A recombinant expression vector comprising the polynucleotide or composition according to claim 56 or claim 57.I l l59. A host cell comprising the recombinant expression vector according to claim 58.

60. A method of treating a disease associated with c-Kit expression in a subject in need thereof, comprising administering to the subject at least one c-Kit binding protein, antibodydrug conjugate, anti-c-Kit antibody, antibody fragment, or CAR-expressing immune cell according to any one of claims 13 to 54.

61. The method according to claim 60, wherein the disease associated with c-Kit expression is a cancer.

62. The method according to claim 60 or claim 61, wherein the disease associated with c- Kit expression is chosen from small cell lung cancer (SCLC), gastrointestinal stromal tumors (GISTs), melanoma, and acute myeloid leukemia (AML).

63. A method of preconditioning a subject prior to a stem cell transplant, comprising administering to the subject at least one c-Kit binding protein, antibody-drug conjugate, anti-c-Kit antibody, antibody fragment, or CAR expressing immune cell according to any one of claims 13 to 54.

64. The method according to claim 63, wherein the subject is suffering from a condition in which a stem cell transplant is considered to be beneficial.

65. The method according to claim 63 or claim 64, wherein the subject is suffering from myelodysplastic syndrome or leukemia.

66. The method according to any one of claims 63 to 65, wherein the stem cell transplant is an autologous stem cell transplant (ASCT).