Treatment of a canine CD20 positive disease or condition using a canine CD20-specific chimeric antigen receptor
The present invention relates to compositions and methods for the treatment of a canine CD20 positive disease or condition using a canine CD20-specific chimeric antigen receptor. One aspect includes a modified canine T cells and pharmaceutical compositions comprising the modified cells for adoptive cell therapy and treating a disease or condition associated with enhanced immunity in canine.
1. An isolated nucleic acid sequence encoding a chimeric antigen receptor (CAR) comprising a canine CD20 antigen binding domain, a transmembrane domain, a costimulatory signaling region, and a CD3 zeta signaling domain, wherein the canine CD20 antigen binding domain comprises an anti-CD20 single chain variable fragment (scFv) comprising SEQ ID NO: 2.
2. The isolated nucleic acid sequence of claim 1 , wherein the isolated nucleic acid sequence comprises a sequence selected from the group consisting of SEQ ID NOs: 3-6.
3. The isolated nucleic acid sequence of claim 1 , wherein the canine CD20 antigen binding domain binds to CD20 expressed by tumor cells and/or tumor vasculature.
4. The isolated nucleic acid sequence of claim 3 , wherein the tumor cells are from a cancer selected from the group consisting of lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, lymph nodes cancer, bone marrow cancer, liver cancer, spleen cancer, ovarian cancer, renal cell carcinoma, bladder cancer, kidney cancer, testicular cancer, prostate cancer, breast cancer, colon cancer, pancreatic cancer, lung cancer, stomach cancer, eye cancer, skin cancer and any combination thereof.
5. The isolated nucleic acid sequence of claim 4 , wherein the cancer is Hodgkin's lymphoma or non-Hodgkin's lymphoma.
6. The isolated nucleic acid sequence of claim 1 , wherein the costimulatory signaling region comprises an intracellular domain of a costimulatory molecule selected from the group consisting of CD8, CD27, CD28, 4-1BB, OX40, CD30, CD40, PD-1, ICOS, lymphocyte function-associated antigen-1 (LFA-1), CD2, CD7, LIGHT, NKG2C, B7-H3, a ligand that specifically binds with CD83, and any combination thereof.
7. The isolated nucleic acid sequence of claim 1 , wherein the nucleic acid sequence is selected from the group consisting of a DNA and an mRNA.
8. The isolated nucleic acid sequence of claim 1 , wherein the nucleic acid sequence encoding the CAR is codon optimized to favor an increase in gene expression, translation efficiency and/or protein expression.
9. A vector comprising the isolated nucleic acid sequence of claim 1 .
10. A chimeric antigen receptor (CAR) comprising a canine CD20 antigen binding domain, a transmembrane domain, a costimulatory signaling region, and a CD3 zeta signaling domain, wherein the canine CD20 antigen binding domain comprises an anti-CD20 single-chain variable fragment (scFv) comprising SEQ ID NO: 2.
11. The CAR of claim 10 , wherein the canine CD20 antigen binding domain binds to CD20 expressed by tumor cells and/or tumor vasculature.
12. The CAR of claim 11 , wherein the tumor cells are from a cancer selected from the group consisting of lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, lymph nodes cancer, bone marrow cancer, liver cancer, spleen cancer, ovarian cancer, renal cell carcinoma, bladder cancer, kidney cancer, testicular cancer, prostate cancer, breast cancer, colon cancer, pancreatic cancer, lung cancer, stomach cancer, eye cancer, skin cancer and any combination thereof.
13. The CAR of claim 12 , wherein the cancer is Hodgkin's lymphoma or non-Hodgkin's lymphoma.
14. The CAR of claim 10 , wherein the costimulatory signaling region comprises an intracellular domain of a costimulatory molecule selected from the group consisting of CD8, CD27, CD28, 4-IBB, OX40, CD30, CD40, PD-1, ICOS, lymphocyte function-associated antigen-1 (LFA-1), CD2, CD7, LIGHT, NKG2C, B7-H3, a ligand that specifically binds with CD83, and any combination thereof.
15. The CAR of claim 14 , wherein the costimulatory signaling region comprises an intracellular domain of CD28, and further wherein the CD28 is canine CD28.
16. The CAR of claim 14 , wherein the costimulatory signaling region comprises an intracellular domain of 4-1BB, and further wherein the 4-1BB is canine 4-1BB.
17. The CAR of claim 14 , wherein the CD3 zeta signaling domain is a canine CD3 zeta signaling domain.
18. The CAR of claim 10 , wherein the CAR is encoded by a codon optimized sequence to favor an increase in gene expression, translation efficiency and/or protein expression.
19. The CAR of claim 10 , further comprising a canine CD8α leader sequence.
20. The CAR of claim 10 , further comprising a canine CD8α hinge.
21. The CAR of claim 10 , wherein the transmembrane domain is selected from a canine CD8a transmembrane domain and a canine CD28 transmembrane domain.
22. A cell comprising the CAR of claim 10 .
23. A chimeric antigen receptor (CAR) comprising a canine CD20 antigen binding domain, a transmembrane domain, a costimulatory signaling region, and a CD3 zeta signaling domain, wherein the CAR is encoded by a sequence selected from the group consisting of SEQ ID NOs: 3-6.
24. A modified cell comprising a nucleic acid sequence encoding a chimeric antigen receptor (CAR) comprising a canine CD20 antigen binding domain, a transmembrane domain, a costimulatory signaling region, and a CD3 zeta signaling domain, wherein the canine CD20 antigen binding domain comprises an anti-CD20 single-chain variable fragment (scFv) comprising SEQ ID NO: 2.
25. The modified cell of claim 24 , wherein the canine CD20 antigen binding domain binds to CD20 expressed by tumor cells and/or tumor vasculature.
26. The modified cell of claim 25 , wherein the tumor cells are from a cancer selected from the group consisting of lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, lymph nodes cancer, bone marrow cancer, liver cancer, spleen cancer, ovarian cancer, renal cell carcinoma, bladder cancer, kidney cancer, testicular cancer, prostate cancer, breast cancer, colon cancer, pancreatic cancer, lung cancer, stomach cancer, eye cancer, skin cancer and any combination thereof.
27. The modified cell of claim 26 , wherein the cancer is Hodgkin's lymphoma or non-Hodgkin's lymphoma.
28. The modified cell of claim 24 , wherein the cell is selected from the group consisting of a T cell, a natural killer (NK) cell, a cytotoxic T lymphocyte (CTL), and a regulatory T cell.
29. The modified cell of claim 24 , wherein the nucleic acid sequence is selected from the group consisting of a DNA and an mRNA.
30. The modified cell of claim 24 , wherein the nucleic acid sequence is introduced into the cell by at least one procedure selected from the group consisting of electroporation, usage of a lentivirus, usage of a retrovirus and a chemical-based transfection.
31. A composition comprising the modified cell of claim 24 .
32. A pharmaceutical composition comprising the modified cell of claim 24 and a pharmaceutically acceptable carrier.
33. A method for stimulating a T cell-mediated immune response in a canine, the method comprising administering to the canine an effective amount of a modified cell that expresses a chimeric antigen receptor comprising a canine CD20 antigen binding domain, a transmembrane domain, a costimulatory signaling region, and a CD3 zeta signaling domain, wherein the canine CD20 antigen binding domain comprises an anti-CD20 single-chain variable fragment (scFv) comprising SEQ ID NO: 2.
34. A method of treating a canine with disease or condition, the method comprising administering to the canine a modified canine T cell that expresses a chimeric antigen receptor comprising a canine CD20 antigen binding domain, a transmembrane domain, a costimulatory signaling region, and a CD3 zeta signaling domain, wherein the canine CD20 antigen binding domain comprises an anti-CD20 single-chain variable fragment (scFv) comprising SEQ ID NO: 2, and wherein the disease or condition is a CD20+cancer.
35. The method of claim 34 , wherein the cancer is selected from the group consisting of lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, lymph nodes cancer, bone marrow cancer, liver cancer, spleen cancer, ovarian cancer, renal cell carcinoma, bladder cancer, kidney cancer, testicular cancer, prostate cancer, breast cancer, colon cancer, pancreatic cancer, lung cancer, stomach cancer, eye cancer, skin cancer and any combination thereof.
36. The method of claim 34 , wherein the modified canine T cell is autologous to the subject.
37. The method of claim 34 , further comprising administering an antitumor vaccine to the canine.
38. The method of claim 37 , wherein the modified canine T cell and the antitumor vaccine are co-administered to the canine.