Compositions and methods for TCR reprogramming using fusion proteins
Provided herein are T-cell receptor (TCR) fusion proteins (TFPs), T-cells engineered to express one or more TFPs, and methods of use thereof for the treatment of diseases, including cancer.
1. A pharmaceutical composition comprising
(I) a T cell from a human subject, wherein the T cell comprises a recombinant nucleic acid molecule encoding a T cell receptor (TCR) fusion protein (TFP) comprising
(a) a TCR subunit comprising a full-length CD3 epsilon sequence or a full-length CD3 gamma sequence; and
(b) a mammalian scFv or single domain antibody comprising an anti-B-cell maturation antigen (BCMA) binding domain; and
(II) a pharmaceutically acceptable carrier;
wherein the TCR subunit and the anti-BCMA binding domain are operatively linked;
wherein the TFP functionally interacts with an endogenous TCR when expressed in the T cell; and
wherein the T cell exhibits increased cytotoxicity to a cell expressing an antigen that specifically interacts with the anti-BCMA binding domain compared to a T cell not containing the TFP.
2. The pharmaceutical composition of claim 1 , wherein the anti-BCMA binding domain is connected to an extracellular domain of the TCR subunit by a linker.
3. The pharmaceutical composition of claim 2 , wherein the linker comprises (G 4 S) n , wherein G is glycine, S is serine, and n is an integer from 1 to 4.
4. The pharmaceutical composition of claim 1 , wherein the anti-BCMA binding domain comprises
(i) a light chain (LC) CDR1, LC CDR2 and LC CDR3 sequence of SEQ ID NO: 37, SEQ ID NO: 39 and SEQ ID NO: 41, respectively;
(ii) a heavy chain (HC) CDR1, HC CDR2 and HC CDR3 sequence of SEQ ID NO: 43, SEQ ID NO: 45 and SEQ ID NO: 47, respectively; or
(iii) a combination thereof.
5. The pharmaceutical composition of claim 1 , wherein the pharmaceutical composition is substantially free of serum.
6. The pharmaceutical composition of claim 1 , wherein the TFP comprises the scFv.
7. The pharmaceutical composition of claim 1 , wherein the TFP comprises the single domain antibody.
8. The pharmaceutical composition of claim 7 , wherein the single domain antibody is a V H domain.
9. The pharmaceutical composition of claim 1 , wherein in the presence of a human cell expressing an antigen that specifically interacts with the anti-BCMA binding domain the T cell has greater than or more efficient cytotoxic activity than a T cell comprising a nucleic acid encoding a chimeric antigen receptor (CAR) comprising the anti-BCMA binding domain operatively linked to at a CD28 extracellular domain, a CD28 transmembrane domain, a CD28 intracellular domain, and a CD3 zeta intracellular domain.
10. The pharmaceutical composition of claim 1 , wherein the TFP molecule functionally interacts with an endogenous TCR complex, at least one endogenous TCR polypeptide, or a combination thereof when expressed in the T cell.
11. The pharmaceutical composition of claim 1 , wherein the T cell is a primary T cell.
12. The pharmaceutical composition of claim 4 , wherein in the presence of a human cell expressing BCMA production of IL-2 by the T cell is lower than production of the IL-2 by a T cell comprising a nucleic acid encoding a chimeric antigen receptor (CAR) comprising the anti-BCMA binding domain operatively linked to a CD28 extracellular domain, a CD28 transmembrane domain, a CD28 intracellular domain, and a CD3 zeta intracellular domain.
13. The pharmaceutical composition of claim 1 , wherein the T cell is a human CD8+ T cell or a human CD4+ T cell.
14. The pharmaceutical composition of claim 1 , wherein the TCR subunit comprises a sequence as set forth in SEQ ID NO: 56.
15. The pharmaceutical composition of claim 1 , wherein the TCR subunit comprises a sequence as set forth in SEQ ID NO: 57.
16. The pharmaceutical composition of claim 1 , wherein production of IFNγ by the T cell is increased in the presence of a cell expressing compared to a T cell not containing the TFP.
17. The pharmaceutical composition of claim 1 , wherein the T cell is a population of human CD8+ or CD4+ T cells, wherein an individual T cell of the population comprises at least two TFP molecules, or at least two T cells of the population collectively comprise at least two TFP molecules; wherein the at least two TFP molecules comprise a mammalian anti-BCMA binding domain, a TCR extracellular domain, a TCR transmembrane domain, and a TCR intracellular domain; and wherein at least one of the at least two TFP molecules functionally interacts with an endogenous TCR complex, at least one endogenous TCR polypeptide, or a combination thereof.
18. The pharmaceutical composition of claim 1 , wherein the single TCR subunit comprises an intracellular domain derived only from CD3 gamma.
19. The pharmaceutical composition of claim 1 , wherein the TCR subunit comprises an intracellular domain derived only from CD3 epsilon.
20. The pharmaceutical composition of claim 1 , wherein the TFP lacks a heterologous stimulatory domain.
21. The pharmaceutical composition of claim 1 , wherein the TFP lacks a costimulatory domain.
22. The pharmaceutical composition of claim 1 , wherein in the presence of a human cell expressing BCMA production of a pro-inflammatory cytokine by the T cell is lower compared to production of the pro-inflammatory cytokine by a T cell comprising a nucleic acid encoding a CAR comprising the anti-BCMA binding domain operatively linked to a CD28 extracellular domain, a CD28 transmembrane domain, a CD28 intracellular domain, and a CD3 zeta intracellular domain.
23. The pharmaceutical composition of claim 21 , wherein the pro-inflammatory cytokine is TNFα.
24. The pharmaceutical composition of claim 21 , wherein the pro-inflammatory cytokine is IL-2.
25. The pharmaceutical composition of claim 21 , wherein the pro-inflammatory cytokine is GM-CSF.
26. A pharmaceutical composition comprising
(I) a T cell from a human subject, wherein the T cell comprises a recombinant nucleic acid molecule encoding a T cell receptor (TCR) fusion protein (TFP) comprising
(a) a TCR subunit comprising a full-length CD3 epsilon sequence or a full-length CD3 gamma sequence; and
(b) a mammalian scFv or single domain antibody comprising an anti-BCMA binding domain; and
(II) a pharmaceutically acceptable carrier;
wherein the TCR subunit and the anti-BCMA binding domain are operatively linked;
wherein the TFP functionally interacts with an endogenous TCR when expressed in the T cell;
wherein the TFP lacks a costimulatory domain and lacks a heterologous stimulatory domain; and
wherein the T cell exhibits increased cytotoxicity to a cell expressing an antigen that specifically interacts with the anti-BCMA binding domain compared to a T cell not containing the TFP.
27. The pharmaceutical composition of claim 26 , wherein in the presence of a human cell expressing BCMA production of a pro-inflammatory cytokine by the T cell is lower compared to production of the pro-inflammatory cytokine by a T cell comprising a nucleic acid encoding a CAR comprising the anti-BCMA binding domain operatively linked to a CD28 extracellular domain, a CD28 transmembrane domain, a CD28 intracellular domain, and a CD3 zeta intracellular domain.
28. The pharmaceutical composition of claim 27 , wherein the pro-inflammatory cytokine is selected from the group consisting of TNFα, GM-CSF, IL-2 and combinations thereof.
29. The pharmaceutical composition of claim 26 , wherein the TCR subunit comprises a sequence as set forth in SEQ ID NO: 56.
30. The pharmaceutical composition of claim 26 , wherein the TCR subunit comprises a sequence as set forth in SEQ ID NO: 57.