Reducing immune tolerance induced by PD-L1
The present disclosure relates to compositions and methods for reducing immune tolerance associated with CAR T cell therapy. Embodiments of the present disclosure include isolated nucleic acid sequence comprising a nucleic acid sequence that encodes modified programmed cell death protein 1 (PD-1) and a nucleic acid sequence that encodes chimeric antigen receptor (CAR).
1. A pharmaceutical composition comprising human T cells, wherein the human T cells comprising:
a first nucleic acid sequence that encodes a modified programmed cell death protein 1 (PD-1) that reduces an inhibitory effect of programmed death ligand 1 (PD-L1) on the human T cells, the modified PD-1 lacking a functional PD-1 intracellular domain for PD-1 signal transduction, wherein the modified PD-1 is a soluble receptor or comprises a mutation at amino acid 223 and/or 248; and
a second nucleic acid sequence that encodes a chimeric antigen receptor (CAR) comprising an extracellular domain that recognizes a tumor antigen of a target cell, a transmembrane domain, and an intracellular domain comprising a CD3-zeta signaling domain and a signaling domain of a costimulatory molecule; and
wherein the modified PD-1 and the CAR are expressed as gene products that are separate polypeptides.
2. The pharmaceutical composition of claim 1 , wherein the modified PD-1 interferes with a pathway between PD-1 of a human T cell of the human cells and PD-L1 of a certain cell.
3. The pharmaceutical composition of claim 1 , wherein the modified PD-1 comprises a PD-1 extracellular domain.
4. The pharmaceutical composition of claim 1 , wherein the modified PD-1 comprises:
a modified PD-1 intracellular domain comprising a substitution at amino acid 223 and/or 224; and
the amino acid sequence of SEQ ID NO: 9 or the amino acid sequence of SEQ ID NO: 10, or a combination thereof.
5. The pharmaceutical composition of claim 1 , wherein the modified PD-1 is a soluble receptor comprising a PD-1 extracellular domain that binds to PD-L1 of a certain cell.
6. The pharmaceutical composition of claim 1 , wherein the tumor antigen is CD19.
7. The pharmaceutical composition of claim 1 , wherein the costimulatory molecule comprises 4-1 BB or CD28, or a combination thereof.
8. The pharmaceutical composition of claim 3 , wherein an intracellular domain of the modified PD-1 comprises the amino acid sequence of SEQ ID NO: 12.
9. The pharmaceutical composition of claim 3 , wherein an intracellular domain of the modified PD-1 comprises the amino acid sequence of SEQ ID NO: 13.
10. The pharmaceutical composition of claim 3 , wherein an intracellular domain of the modified PD-1 comprises the amino acid sequence of SEQ ID NO: 14.
11. A method of reducing an inhibitory effect of PD-L1 on human T cells, the method comprising:
introducing a first nucleic acid sequence and a second nucleic acid sequence into the human T cells, wherein:
a first nucleic acid sequence encodes a modified PD-1 that reduces the inhibitory effect of PD-L1 on the human T cells, the modified PD-1 lacking a functional PD-1 intracellular domain for PD-1 signal transduction, wherein the modified PD-1 is a soluble receptor or comprises a mutation at amino acid 223 and/or 248; and
a second nucleic acid sequence encodes a CAR comprising an extracellular domain that recognizes a tumor antigen of a target cell, a transmembrane domain, and an intracellular domain comprising a CD3-zeta signaling domain and a signaling domain of a costimulatory molecule;
and wherein the modified PD-1 and the CAR are expressed as gene products that are separate.
12. The method of claim 11 , wherein the modified PD-1 interferes with a pathway between PD-1 of a human T cell of the human cells and PD-L1 of a certain cell.
13. The method of claim 11 , wherein the modified PD-1 comprises a PD-1 extracellular domain.
14. The method of claim 11 , wherein the modified PD-1 comprises:
a modified PD-1 intracellular domain comprising a substitution at amino acid 223 and/or 224; and
the amino acid sequence of SEQ ID NO: 9 or the amino acid sequence of SEQ ID NO: 10, or a combination thereof.
15. The method of claim 11 , wherein the modified PD-1 is a soluble receptor comprising a PD-1 extracellular domain that binds to PD-L1 of a certain cell.
16. The method of claim 11 , wherein the tumor antigen is CD19.
17. The method of claim 11 , wherein the costimulatory molecule comprises 4-1 BB or CD28, or a combination thereof.
18. The method of claim 13 , wherein an intracellular domain of the modified PD-1 comprises the amino acid sequence of SEQ ID NO: 12.
19. The method of claim 13 , wherein an intracellular domain of the modified PD-1 comprises the amino acid sequence of SEQ ID NO: 13.
20. The method of claim 13 , wherein an intracellular domain of the modified PD-1 comprises the amino acid sequence of SEQ ID NO: 14.
21. The pharmaceutical composition of claim 1 , wherein the first nucleic acid sequence comprises SEQ ID NO: 4.
22. The method of claim 11 , wherein the first nucleic acid sequence comprises sequence of SEQ ID NO: 4.
23. The method of claim 11 , wherein the modified PD-1 comprises a truncated PD-1 that does not include an intracellular domain.
24. The pharmaceutical composition of claim 1 , wherein the PD-1 is a soluble receptor.
25. The pharmaceutical composition of claim 1 , wherein the PD-1 comprises a mutation at amino acid 223 and/or 248.
26. The method of claim 11 , wherein the PD-1 is a soluble receptor.
27. The method of claim 11 , wherein the PD-1 comprises a mutation at amino acid 223 and/or 248.