High affinity merkel cell polyomavirus T antigen-specific TCRS and uses thereof
The present disclosure provides binding proteins and TCRs with high affinity and specificity against Merkel cell polyomavirus T antigen epitopes or peptides, T cells expressing such high affinity Merkel cell polyomavirus T antigen specific TCRs, nucleic acids encoding the same, and compositions for use in treating Merkel cell carcinoma.
1. An expression vector comprising a polynucleotide encoding a binding protein, wherein the binding protein comprises:
(a) a T cell receptor (TCR) α-chain variable (Vα) domain having a CDR3 amino acid sequence of any one of SEQ ID NOS.:13, 44 and 355-364; and
(b) a TCR β-chain variable (Vβ) domain having a CDR3 amino acid sequence of any one of SEQ ID NOS.:14, 69 and 365-374.
2. The expression vector of claim 1 , wherein the binding protein is capable of specifically binding to a Merkel cell polyomavirus T antigen peptide:HLA complex on a cell surface independent of CD8 or in the absence of CD8.
3. The expression vector according to claim 1 , wherein the binding protein is capable of specifically binding to a KLLEIAPNC (SEQ ID NO.:17):human leukocyte antigen (HLA) complex or a KLLEIAPNA (SEQ ID NO.:37):human leukocyte antigen (HLA) complex with a K d less than or equal to about 10 −8 M.
4. The expression vector according to claim 1 , wherein the binding protein specifically binds to a KLLEIAPNC (SEQ ID NO.:17):HLA-A*02:01 complex or a KLLEIAPNA (SEQ ID NO.:37):HLA-A*02:01 complex.
5. The expression vector according to claim 1 , wherein the
Vα domain is at least about 90% identical to the Vα amino acid sequence set forth in
MDKILGASFLVLWLQLCWVSGQQKEKSDQQQVKQSPQSLIVQKGGISIINCAYE NTAFDYFPWYQQFPGKGPALLIAIRPDVSEKKEGRFTISFNKSAKQFSLHIMDSQP GDSATYFCAVPNTGNQFYFGTGTSLTVIP (SEQ ID NO.: 428); and/or
the Vβ domain is at least about 90% identical to the V β amino acid sequence set forth in
MGTRLLCWVVLGFLGTDHTGAGVSQSPRYKVAKRGQDVALRCDPISGHVSLFW YQQALGQGPEFLTYFQNEAQLDKSGLPSDRFFAERPEGSVSTLKIQRTQQEDSAV YLCASSLIAGLSYEQYFGPGTRLTVT (SEQ ID NO.: 429).
6. The expression vector according to claim 1 , wherein the
Vα domain comprises the Vα amino acid sequence set forth in
MDKILGASFLVLWLQLCWVSGQQKEKSDQQQVKQSPQSLIVQKGGISIINCAYE NTAFDYFPWYQQFPGKGPALLIAIRPDVSEKKEGRFTISFNKSAKQFSLHIMDSQP GDSATYFCAVPNTGNQFYFGTGTSLTVIP (SEQ ID NO.: 428); and/or
the Vβ domain comprises the Vβ amino acid sequence set forth in
MGTRLLCWVVLGFLGTDHTGAGVSQSPRYKVAKRGQDVALRCDPISGHVSLFW YQQALGQGPEFLTYFQNEAQLDKSGLPSDRFFAERPEGSVSTLKIQRTQQEDSAV YLCASSLIAGLSYEQYFGPGTRLTVT (SEQ ID NO.: 429).
7. The expression vector according claim 1 , wherein the binding protein further comprises a TCR α-chain constant domain (Cα) having at least 90% sequence identity to the amino acid sequence of SEQ ID NO.:2, and/or wherein the binding protein further comprises a TCR β-chain constant domain (Cβ) having at least 90% sequence identity to the amino acid sequence of SEQ ID NO.:4.
8. The expression vector according claim 1 , wherein:
the Vα domain comprises the Vα amino acid sequence set forth in
MDKILGASFLVLWLQLCWVSGQQKEKSDQQQVKQSPQSLIVQKGGISIINCAYENTAFD YFPWYQQFPGKGPALLIAIRPDVSEKKEGRFTISFNKSAKQFSLHIMDSQPGDSATYFCA VPNTGNQFYFGTGTSLTVIP (SEQ ID NO.: 428); and
the Vβ domain comprises the Vβ amino acid sequence set forth in
MGTRLLCWVVLGFLGTDHTGAGVSQSPRYKVAKRGQDVALRCDPISGHVSLFWYQQA LGQGPEFLTYFQNEAQLDKSGLPSDRFFAERPEGSVSTLKIQRTQQEDSAVYLCASSLIA GLSYEQYFGPGTRLTVT (SEQ ID NO.: 429); and
wherein the binding protein further comprises a TCR α-chain constant domain (Cα) comprising the amino acid sequence of SEQ ID NO.:2; and a TCR β-chain constant domain (Cβ) comprising the amino acid sequence of SEQ ID NO.:4,
wherein the Vα and the Cα are comprised in a TCRα chain and wherein the Vβ and the Cβ are comprised in a TCRβ chain.
9. The expression vector according to claim 1 , wherein the binding protein is a T cell receptor (TCR), an antigen-binding fragment of a TCR, or a chimeric antigen receptor.
10. The expression vector according to claim 9 , wherein the TCR, the chimeric antigen receptor, or the antigen-binding fragment of the TCR is chimeric, humanized or human.
11. The expression vector according to claim 9 , wherein the antigen-binding fragment of the TCR comprises a single chain TCR (scTCR).
12. A composition comprising the expression vector according to claim 1 and a pharmaceutically acceptable carrier, diluent, or excipient.
13. The expression vector of claim 1 , wherein the polynucleotide is operably linked to an expression control sequence.
14. A genetically engineered host cell, comprising the expression vector of claim 1 and expressing the binding protein on its cell surface.
15. The genetically engineered host cell of claim 14 , wherein:
(a) the host cell is a hematopoietic progenitor cell or a human immune system cell;
(b) the host cell is an immune system cell selected from the group consisting of: a CD4+ T cell, a CD8+ T cell, a CD4−CD8− double negative T cell, a γδ T cell, a natural killer cell, and a dendritic cell;
(c) the host cell is a T cell; or
(d) the host cell is a T cell selected from the group consisting of: a naïve T cell, a central memory T cell, and an effector memory T cell.
16. An adoptive immunotherapy method for treating a subject having a Merkel cell carcinoma, comprising administering to the subject an effective amount of a genetically engineered host cell according to claim 15 .
17. A unit dose form comprising a genetically engineered host cell according to claim 15 .
18. An adoptive immunotherapy method for treating a subject having a Merkel cell carcinoma, comprising administering to the subject an effective amount of a genetically engineered host cell according to claim 14 .
19. A unit dose form comprising a genetically engineered host cell according to claim 14 .
20. A host cell comprising a heterologous polynucleotide encoding a binding protein that is capable of specifically binding to a Merkel cell polyomavirus T antigen peptide:HLA complex, wherein the binding protein comprises:
(a) a T cell receptor (TCR) α chain variable (Vα) domain having a CDR3 amino acid sequence of any one of SEQ ID NOS.:13, 44, and 355-364, and
(b) a TCR β chain variable (Vβ) domain having a CDR3 amino acid sequence of any one of SEQ ID NOS.:14, 69, and 365-374.
21. The host cell of claim 20 , wherein the host cell comprises a CD4+ T cell, a CD8+ T cell, a CD4−CD8− double negative T cell, a γδ T cell, a naïve T cell, a central memory T cell, an effector memory T cell, a natural killer cell, a dendritic cell, or any combination thereof.
22. A method of treating a Merkel cell carcinoma, comprising administering a therapeutically effective amount of the host cell according to claim 20 to a subject having or at risk of having Merkel cell carcinoma.
23. The host cell according to claim 20 , wherein the binding protein comprises a TCR.
24. The host cell according to claim 20 , wherein the host cell comprises a CD4+ T cell and further comprises a heterologous polynucleotide encoding a CD8 co-receptor molecule.
25. The host cell according to claim 20 , wherein the binding protein is capable of specifically binding to a Merkel cell polyomavirus T antigen peptide:HLA complex on a cell surface independent of CD8 or in the absence of CD8.
26. The host cell according to claim 20 , wherein the binding protein is capable of specifically binding to a KLLEIAPNC (SEQ ID NO.:17):human leukocyte antigen (HLA) complex and/or a KLLEIAPNA (SEQ ID NO.:37):human leukocyte antigen (HLA) complex, with a K d less than or equal to about 10 −8 M.
27. The host cell according to claim 20 , wherein the binding protein is capable of specifically binding to a KLLEIAPNC (SEQ ID NO.:17):HLA-A*02:01 complex and/or a KLLEIAPNA (SEQ ID NO.:37):HLA-A*02:01 complex.
28. The host cell according to claim 20 , wherein
the V α domain is at least about 90% identical to a V α domain amino acid sequence set forth in
MDKILGASFLVLWLQLCWVSGQQKEKSDQQQVKQSPQSLIVQKGGISIINCAYE NTAFDYFPWYQQFPGKGPALLIAIRPDVSEKKEGRFTISFNKSAKQFSLHIMDSQP GDSATYFCAVPNTGNQFYFGTGTSLTVIP (SEQ ID NO.: 428); and/or
the V β domain is at least about 90% identical to a V β domain amino acid sequence set forth in
MGTRLLCWVVLGFLGTDHTGAGVSQSPRYKVAKRGQDVALRCDPISGHVSLFW YQQALGQGPEFLTYFQNEAQLDKSGLPSDRFFAERPEGSVSTLKIQRTQQEDSAV YLCASSLIAGLSYEQYFGPGTRLTVT (SEQ ID NO.: 429).
29. The host cell according to claim 28 , wherein
the V α domain comprises an amino acid sequence set forth in
MDKILGASFLVLWLQLCWVSGQQKEKSDQQQVKQSPQSLIVQKGGISIINCAYE NTAFDYFPWYQQFPGKGPALLIAIRPDVSEKKEGRFTISFNKSAKQFSLHIMDSQP GDSATYFCAVPNTGNQFYFGTGTSLTVIP (SEQ ID NO.: 428); and/or
the V β domain comprises an amino acid sequence set forth in
MGTRLLCWVVLGFLGTDHTGAGVSQSPRYKVAKRGQDVALRCDPISGHVSLFW YQQALGQGPEFLTYFQNEAQLDKSGLPSDRFFAERPEGSVSTLKIQRTQQEDSAV YLCASSLIAGLSYEQYFGPGTRLTVT (SEQ ID NO.: 429).
30. The host cell according to claim 29 , wherein the binding protein further comprises a TCR α-chain constant domain having at least 90% sequence identity to the amino acid sequence set forth in SEQ ID NO.:2, and/or wherein the binding protein further comprises a TCR β-chain constant domain having at least 90% sequence identity to the amino acid sequence set forth in SEQ ID NO.:4.
31. The host cell according to claim 30 , wherein the binding protein comprises a TCR comprising:
the V α domain amino acid sequence set forth in
MDKILGASFLVLWLQLCWVSGQQKEKSDQQQVKQSPQSLIVQKGGISIINCAYENTAFD YFPWYQQFPGKGPALLIAIRPDVSEKKEGRFTISFNKSAKQFSLHIMDSQPGDSATYFCA VPNTGNQFYFGTGTSLTVIP (SEQ ID NO.: 428);
the α-chain constant domain amino acid sequence set forth in SEQ ID NO.:2;
the V β domain amino acid sequence set forth in
MGTRLLCWVVLGFLGTDHTGAGVSQSPRYKVAKRGQDVALRCDPISGHVSLFWYQQA LGQGPEFLTYFQNEAQLDKSGLPSDRFFAERPEGSVSTLKIQRTQQEDSAVYLCASSLIA GLSYEQYFGPGTRLTVT (SEQ ID NO.: 429); and
the β-chain constant domain amino acid sequence set forth in SEQ ID NO.:4.
32. The host cell according to claim 20 , wherein the host cell comprises:
(i) a CD4+ T cell;
(ii) a CD8+ T cell;
(iii) a CD8+ CD62L+ T cell; or
(iv) any combination of (i)-(iii).
33. The host cell according to claim 20 , wherein the host cell is a hematopoietic progenitor cell or a human immune system cell.
34. An isolated polynucleotide encoding a binding protein, wherein the binding protein comprises:
(a) a T cell receptor (TCR) α-chain variable (Vα) domain having a CDR3 amino acid sequence of any one of SEQ ID NOS.:13, 44 and 355-364; and
(b) a TCR β-chain variable (Vβ) domain having a CDR3 amino acid sequence of any one of SEQ ID NOS.:14, 69 and 365-374,
wherein the polynucleotide comprises a Vα domain-encoding polynucleotide having at least 80% identity to the polynucleotide sequence of any one of SEQ ID NOs.:5, 6, 375-384 and 395-404, and a Vβ domain-encoding polynucleotide having at least 80% identity to the polynucleotide sequence of any one of SEQ ID NOs.: 9, 10, 385-394, and 405-414.
35. The isolated polynucleotide according to claim 34 , wherein the TCR Vβ-encoding polynucleotide comprises or consists of the polynucleotide sequence set forth in SEQ ID NO.:405 and the TCR Vα-encoding the polynucleotide comprises or consists of the polynucleotide sequence set forth in SEQ ID NO.:395.
36. The isolated polynucleotide according to claim 35 , wherein the binding protein further comprises a TCR beta chain constant domain (TCR Cβ) and/or a TCR alpha chain constant domain (TCR Cα),
wherein the TCR Cβ-encoding polynucleotide comprises or consists of the polynucleotide sequence set forth in SEQ ID NO.:415 and the TCR Cα-encoding polynucleotide comprises or consists of the polynucleotide sequence set forth in SEQ ID NO.:8.
37. The isolated polynucleotide according to claim 35 , comprising the polynucleotide sequence set forth in SEQ ID NO.:417.