Genetically modified immune cells targeting NY-ESO-1 and methods of use thereof

1. A modified T cell comprising:

(a) an exogenous T cell receptor (TCR) having affinity for NY-ESO-1 on a target cell, wherein the exogenous TCR comprises:

(i) a TCR α chain comprising a variable region having the amino acid sequence set forth in SEQ ID NO:2; and

(ii) a TCR β chain comprising a variable region having the amino acid sequence set forth in SEQ ID NO:7;

(b) an endogenous TCR α chain coding sequence comprising the nucleic acid sequence set forth in SEQ ID NO:128, wherein SEQ ID NO:128 comprises at least one nucleotide substitution, deletion, insertion, or insertion and deletion;

(c) an endogenous TCR β chain coding sequence comprising the nucleic acid sequence set forth in SEQ ID NO:129, wherein SEQ ID NO:129 comprises at least one nucleotide substitution, deletion, insertion, or insertion and deletion; and

(d) a switch receptor comprising:

(i) a first domain comprising at least a portion of the extracellular domain of a human PD-1 comprising an amino acid sequence set forth at positions 1-170 of SEQ ID NO: 134, wherein the human PD-1 is a variant PD-1 comprising an alanine to leucine substitution at amino acid position 132 of SEQ ID NO: 134;

(ii) a second domain comprising a switch receptor transmembrane domain; and

(iii) a third domain comprising the intracellular domain of CD28 or 4-1BB;

wherein the expression of the endogenous TCR α and β chain coding sequences are downregulated;

wherein the modified T cell exhibits increased resistance to a PD1 immunosuppressive signal or a non-PD1 immunosuppressive signal; enhanced tumor-infiltrating lymphocytes (TIL) infiltration and enhanced secretion of a cytokine selected from CD107, IFNγ, IL-2, or TNF; and

wherein the non-PD1 immunosuppressive signal is selected from the group consisting of TGFβ, adenosine, Indoleamine 2,3-dioxygenase (IDO), hypoxia, T regulatory cells (Tregs), and combinations thereof.

2. The modified T cell of claim 1 , wherein the non-PD1 immunosuppressive signal is T regulatory cells (Tregs).

3. The modified T cell of claim 2 , wherein the T cell comprises an endogenous PD1 coding sequence comprising the nucleic acid sequence set forth in SEQ ID NO:130, wherein SEQ ID NO:130 comprises at least one nucleotide substitution, deletion, insertion, or insertion and deletion.

4. The modified T cell of claim 2 , wherein

the switch receptor transmembrane domain comprises at least a portion of the transmembrane domain of CD28; and

the third domain comprises at least a portion of the intracellular domain of CD28.

5. The modified T cell of claim 2 , wherein

the switch receptor transmembrane domain comprises at least a portion of the transmembrane domain of CD8alpha; and

the third domain comprises at least a portion of the intracellular domain of 4-1BB.

6. The modified T cell of claim 2 , wherein

the switch receptor transmembrane domain comprising at least a portion of the transmembrane domain of CD8alpha; and

the third domain comprises at least a portion of the intracellular domain of CD28.

7. The modified T cell of claim 2 , wherein the switch receptor comprises the amino acid sequence set forth in any one of SEQ ID NOs: 136 or 138.

8. The modified T cell of claim 1 , wherein the modified T cell is an autologous T cell derived from a human.

9. A method of treating a cancer in a subject in need thereof, the method comprising administering a therapeutically effective composition comprising the modified T cell of claim 1 to the subject.

10. The method of claim 9 , further comprising administering to the subject a lymphodepleting chemotherapy.

11. The method of claim 10 , wherein the lymphodepleting chemotherapy comprises administering to the subject an effective amount of cyclophosphamide, and an effective amount of fludarabine.

12. The method of claim 9 , wherein the cancer is selected from the group consisting of multiple myeloma, melanoma, synovial sarcoma, and myxoid/round cell liposarcoma.

13. A modified T cell comprising:

(a) an exogenous T cell receptor (TCR) having affinity for NY-ESO-1 on a target cell, wherein the exogenous TCR comprises:

(i) a TCR α chain comprising a variable region having the amino acid sequence set forth in SEQ ID NO:2; and

(ii) a TCR β chain comprising a variable region having the amino acid sequence set forth in SEQ ID NO:7;

(b) an endogenous TCR α chain coding sequence comprising the nucleic acid sequence set forth in SEQ ID NO:128, wherein SEQ ID NO:128 comprises at least one nucleotide substitution, deletion, insertion, or insertion and deletion;

(c) an endogenous TCR β chain coding sequence comprising the nucleic acid sequence set forth in SEQ ID NO:129, wherein SEQ ID NO:129 comprises at least one nucleotide substitution, deletion, insertion, or insertion and deletion;

(d) an endogenous PD1 coding sequence comprising the nucleic acid sequence set forth in SEQ ID NO:130, wherein SEQ ID NO:130 comprises at least one nucleotide substitution, deletion, insertion, or insertion and deletion; and

(e) a switch receptor comprising:

(i) a first domain comprising at least a portion of the extracellular domain of a human PD-1 comprising an amino acid sequence set forth at positions 1-170 of SEQ ID NO: 134, wherein the human PD1 is a variant of human PD-1 comprising an alanine to leucine substitution at amino acid position 132 of SEQ ID NO: 134;

(ii) a second domain comprising a switch receptor transmembrane domain; and

(iii) a third domain comprising the intracellular domain of CD28 or 4-1BB;

wherein the expression of the endogenous TCR α chain, TCR β chain, and PD1 coding sequences are downregulated;

wherein the modified T cell exhibits increased resistance to a PD1 immunosuppressive signal or a non-PD1 immunosuppressive signal; enhanced tumor-infiltrating lymphocytes (TIL) infiltration and enhanced secretion of a cytokine selected from CD107, IFNγ, IL-2, or TNF; and

wherein the non-PD1 immunosuppressive signal is selected from the group consisting of TGFβ, adenosine, Indoleamine 2,3-dioxygenase (IDO), hypoxia, T regulatory cells (Tregs), and combinations thereof.

14. The modified T cell of claim 13 , wherein the modified T cell is an autologous T cell derived from a human.

15. The modified T cell of claim 13 , wherein the non-PD1 immunosuppressive signal is T regulatory cells (Tregs).

16. A method of treating a cancer in a subject in need thereof, the method comprising administering a therapeutically effective composition comprising the modified T cell of claim 13 to the subject, wherein the cancer is selected from the group consisting of multiple myeloma, melanoma, synovial sarcoma, and myxoid/round cell liposarcoma.

17. A modified T cell comprising:

(a) an exogenous T cell receptor (TCR) having affinity for NY-ESO-1 on a target cell, wherein the exogenous TCR comprises:

(i) a TCR α chain comprising a variable region having the amino acid sequence set forth in SEQ ID NO:2; and

(ii) a TCR β chain comprising a variable region having the amino acid sequence set forth in SEQ ID NO:7;

(b) an endogenous TCR α chain coding sequence comprising the nucleic acid sequence set forth in SEQ ID NO:128, wherein SEQ ID NO:128 comprises at least one nucleotide substitution, deletion, insertion, or insertion and deletion;

(c) an endogenous TCR β chain coding sequence comprising the nucleic acid sequence set forth in SEQ ID NO:129, wherein SEQ ID NO:129 comprises at least one nucleotide substitution, deletion, insertion, or insertion and deletion; and

(d) a switch receptor comprising the amino acid sequence set forth in any one of SEQ ID NOs: 136 or 138,

wherein the expression of the endogenous TCR α chain coding sequence and the expression of the endogenous TCR β chain coding sequence are downregulated;

wherein the modified T cell exhibits increased resistance to a PD1 immunosuppressive signal or a non-PD1 immunosuppressive signal; enhanced tumor-infiltrating lymphocytes (TIL) infiltration; and enhanced secretion of a cytokine selected from CD107, IFNγ, IL-2, or TNF; and

wherein the non-PD1 immunosuppressive signal is selected from the group consisting of TGFβ, adenosine, Indoleamine 2,3-dioxygenase (IDO), hypoxia, T regulatory cells (Tregs), and combinations thereof.

18. The modified T cell of claim 17 , wherein the modified T cell is an autologous T cell derived from a human.

19. The modified T cell of claim 17 , wherein the non-PD1 immunosuppressive signal is T regulatory cells (Tregs).

20. A method of treating a cancer in a subject in need thereof, the method comprising administering a therapeutically effective composition comprising the modified T cell of claim 17 to the subject, wherein the cancer is selected from the group consisting of multiple myeloma, melanoma, synovial sarcoma, and myxoid/round cell liposarcoma.