Chimeric antigen receptor compositions and methods for treating MUC1* diseases
Disclosed herein are chimeric antigen receptors (CARs) that target MUC1*. In some embodiments, the CARs have been optimized to reduce T cell exhaustion.
1. A chimeric antigen receptor (CAR) comprising:
(a) a MUC1* binding single chain antibody domain comprising:
(i) heavy chain (HC) complementarity determining regions (CDRs) comprising:
a HC-CDR1 comprising SEQ ID NO: 18,
a HC-CDR2 comprising SEQ ID NO: 19, and
a HC-CDR3 comprising SEQ ID NO: 20; and
(ii) light chain (LC) CDRs comprising:
a LC-CDR1 comprising SEQ ID NO: 21,
a LC-CDR2 comprising SEQ ID NO: 22, and
a LC-CDR3 comprising SEQ ID NO: 23;
(b) a hinge region comprising SEQ ID NO: 40;
(c) a transmembrane domain comprising SEQ ID NO: 41; and
(d) a signaling domain comprising SEQ ID NO: 44.
2. The CAR of claim 1 , further comprising a costimulatory domain comprising SEQ ID NO: 42.
3. The CAR of claim 2 , wherein the MUC1* binding single chain antibody domain comprises a heavy chain variable domain that comprises an amino acid sequence that has at least 90% identity to SEQ ID NO: 1, a linker, and a light chain variable domain that comprises an amino acid sequence that has at least 90% identity to SEQ ID NO: 2.
4. The CAR of claim 3 , wherein the linker comprises an amino acid sequence of SEQ ID NO: 24, SEQ ID NO: 26, or SEQ ID NO: 50.
5. The CAR of claim 3 , wherein the linker comprises SEQ ID NO: 24.
6. The CAR of claim 3 , wherein the linker comprises any one of SEQ ID NOs: 26, 28, 30, 32, 34, 36, or 38.
7. The CAR of claim 1 , wherein the MUC1* binding single chain antibody domain comprises SEQ ID NO: 39.
8. The CAR of claim 1 , wherein the hinge region and the transmembrane domain together comprise SEQ ID NO: 3.
9. The CAR of claim 2 , wherein a cytoplasmic domain comprising the costimulatory domain and the signaling domain comprises SEQ ID NO: 4.
10. The CAR of claim 1 , wherein the CAR comprises a sequence having at least 95% identity to SEQ ID NO: 48.
11. The CAR of claim 1 , wherein the CAR consists of SEQ ID NO: 48.
12. An immune cell comprising the CAR of claim 1 .
13. A vector comprising a nucleic acid sequence encoding the CAR of claim 1 .
14. An immune cell comprising the vector of claim 13 .
15. A method of treating a MUC1* positive cancer in an individual, comprising administering to the individual an engineered immune cell expressing a CAR comprising:
(a) a MUC1* binding single chain antibody domain comprising:
(i) heavy chain (HC) complementarity determining regions (CDRs) comprising:
a HC-CDR1 comprising SEQ ID NO: 18,
a HC-CDR2 comprising SEQ ID NO: 19, and
a HC-CDR3 comprising SEQ ID NO: 20; and
(ii) light chain (LC) CDRs comprising:
a LC-CDR1 comprising SEQ ID NO: 21,
a LC-CDR2 comprising SEQ ID NO: 22, and
a LC-CDR3 comprising SEQ ID NO: 23;
(b) a hinge region comprising SEQ ID NO: 40;
(c) a transmembrane domain comprising SEQ ID NO: 41;
(d) a costimulatory domain comprising SEQ ID NO: 42; and
(e) a signaling domain comprising SEQ ID NO: 44.
16. The method of claim 15 , wherein the MUC1* positive cancer comprises a solid tumor.
17. The method of claim 15 , wherein the MUC1* positive cancer is breast cancer.
18. The method of claim 15 , wherein the MUC1* positive cancer is lung cancer.
19. The method of claim 15 , wherein the MUC1* positive cancer is pancreatic cancer.
20. The method of claim 15 , wherein a section of a tumor of the MUC1* positive cancer has low MUC1* expression characterized by an anti-MUC1* H-score of 100 or less.
21. The method of claim 15 , wherein a cell of the MUC1* positive cancer reacts with a MUC1* antibody in an immunohistochemistry assay.
22. The method of claim 15 , wherein a cell of the MUC1* positive cancer reacts with a MUC1* antibody in an enzyme linked immunosorbent assay (ELISA).
23. The method of claim 15 , wherein a cell of the MUC1* positive cancer reacts with a MUC1* antibody in flow cytometry assay.
24. The method of claim 15 , wherein the method reduces tumor recurrence compared to treatment with an immune cell comprising an otherwise identical CAR wherein the signaling domain comprises SEQ ID NO: 43 instead of SEQ ID NO: 44.
25. The method of claim 15 , wherein the engineered immune cell is a T cell.
26. The method of claim 25 , wherein the T cell remains active after 6 or more days of stimulation with MUC1* or a synthetic MUC1* peptide comprising SEQ ID NO: 49.
27. The method of claim 25 , wherein the T cell is derived from a healthy donor.
28. The method of claim 25 , wherein the T cell is derived from an individual with a MUC1* positive cancer.
29. The method of claim 15 , wherein the MUC1* binding single chain antibody domain comprises a heavy chain variable domain that comprises an amino acid sequence that has at least 90% identity to SEQ ID NO: 1, a linker, and a light chain variable domain that comprises an amino acid sequence that has at least 90% identity to SEQ ID NO: 2.
30. The CAR of claim 29 , wherein the linker comprises an amino acid sequence of SEQ ID NO: 24, SEQ ID NO: 26, or SEQ ID NO: 50.
31. The method of claim 29 , wherein the linker comprises SEQ ID NO: 24.
32. The method of claim 29 , wherein the linker comprises any one of SEQ ID NOs: 26, 28, 30, 32, 34, 36, or 38.
33. The method of claim 15 , wherein the MUC1* binding single chain antibody domain comprises SEQ ID NO: 39.
34. The method of claim 15 , wherein the hinge region and the transmembrane domain together comprise SEQ ID NO: 3.
35. The method of claim 15 , wherein a cytoplasmic domain comprising the costimulatory domain and the signaling domain comprises SEQ ID NO: 4.
36. The method of claim 15 , wherein the CAR comprises a sequence having at least 95% identity to SEQ ID NO: 48.
37. The method of claim 15 , wherein the CAR consists of SEQ ID NO: 48.
38. A method of killing a MUC1* positive cancer cell comprising contacting the cell with an immune cell expressing a CAR comprising:
(a) a MUC1* binding single chain antibody domain comprising:
(i) heavy chain (HC) complementarity determining regions (CDRs) comprising:
a HC-CDR1 comprising SEQ ID NO: 18,
a HC-CDR2 comprising SEQ ID NO: 19, and
a HC-CDR3 comprising SEQ ID NO: 20, and
(ii) light chain (LC) CDRs comprising:
a LC-CDR1 comprising SEQ ID NO: 21,
a LC-CDR2 comprising SEQ ID NO: 22, and
a LC-CDR3 comprising SEQ ID NO: 23;
(b) a hinge region comprising SEQ ID NO: 40;
(c) a transmembrane domain comprising SEQ ID NO: 41;
(d) a costimulatory domain comprising SEQ ID NO: 42; and
(e) a signaling domain comprising SEQ ID NO: 44.
39. The method of claim 38 , wherein the contacting is carried out in vitro.