Anti-MAGE-A4 T cell receptor

Abstract

The present invention relates to T cell receptors (TCRs), in particular TCRs specific for peptides derived from melanoma-associated antigen 4 (MAGE-A4). The invention also relates to antigen-binding portions of said TCRs, compositions comprising said TCRs or portions, fusion proteins comprising said TCRs or portions, nucleic acids and vectors encoding said TCRs or portions, cells comprising said TCRs or portions, and methods of treatment involving said TCRs or portions.

Description

[Technical Field]

[0001] The present invention relates to T cell receptors (TCRs), in particular TCRs specific for peptides derived from melanoma-associated antigen 4 (MAGE-A4). The invention also relates to antigen-binding portions of said TCRs, compositions comprising said TCRs or portions, fusion proteins comprising said TCRs or portions, nucleic acids and vectors encoding said TCRs or portions, cells comprising said TCRs or portions, and methods of treatment involving said TCRs or portions. [Background technology]

[0002] MAGE-A4 is a cancer-testis antigen that is expressed in testicular tissue and by some types of cancer cells, allowing MAGE-A4 expression to be used as a marker for some cancerous cells.

[0003] MAGE-A4 expression by certain cells can be recognized by T cells expressing TCRs that are specific for peptides derived from MAGE-A4 when presented on MHC molecules. TCRs with this property are reported in WO2021 / 236548A1 and elsewhere.

[0004] Other documents disclosing MAGE-A antigen-binding proteins include WO2021 / 023658A1 and WO2020 / 257288A2.

[0005] Such TCRs are useful for a variety of purposes, including several types of immunotherapy. TCRs with different properties are needed for the development of such therapies. Thus, there is a need for additional MAGE-A4-specific TCRs. [Prior art documents] [Patent documents]

[0006] [Patent Document 1] WO2021 / 236548A1 [Patent Document 2] WO2021 / 023658A1

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Patent document 4

Non-licensed literature

[0007]

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Summary of the Invention

[0009] In one embodiment, the CDR3a comprises SEQ ID NO: 49 or SEQ ID NO: 53, but not SEQ ID NO: 7, and the TCR-b variable region, or a portion thereof, comprises CDR3b comprising any one of SEQ ID NOs: 14 and 57 to 66.

[0010] In another embodiment, the TCR-a variable region, or a portion thereof, comprises a CDR3a comprising any one of SEQ ID NOs: 7, 26 to 55, 140, and 141, or optionally any one of SEQ ID NOs: 7 and 26 to 55, and a CDR3b comprising SEQ ID NO: 62 or SEQ ID NO: 64, wherein CDR3b does not comprise SEQ ID NO: 14.

[0011] The TCR-a variable region, or a portion thereof, may comprise a CDR1a comprising any one of SEQ ID NOs: 5 and 15 to 17. The TCR-a variable region, or a portion thereof, may comprise a CDR2a comprising any one of SEQ ID NOs: 6 and 18 to 25. The TCR-b variable region, or a portion thereof, may comprise a CDR1b comprising SEQ ID NO: 12. The TCR-b variable region, or a portion thereof, may comprise a CDR2b comprising SEQ ID NO: 13 or SEQ ID NO: 56.

[0012] In another aspect, there is provided a TCR, or antigen-binding portion thereof, capable of specifically binding to the peptide set forth in SEQ ID NO: 122 presented on an MHC class I molecule, comprising a TCR-a variable region, or a portion thereof, and a TCR-b variable region, or a portion thereof, wherein the TCR-a variable region, or a portion thereof, comprises CDR3a and the TCR-b variable region, or a portion thereof, comprises CDR3b, wherein the CDR3a comprises any one of the CDR3a sequences listed in Table 1 and the CDR3b comprises any one of the CDR3b sequences listed in Table 2, and wherein the CDR3a and CDR3b are associated with the same TCR ID listed in Table 1 and Table 2.

[0013] In another aspect, a TCR, or antigen-binding portion thereof, capable of specifically binding to the peptide set forth in SEQ ID NO: 122 presented on an MHC class I molecule, comprising a TCR-a variable region, or portion thereof, and a TCR-b variable region, or portion thereof, wherein the TCR-a variable region, or portion thereof, comprises CDR1a, CDR2a, and CDR3a, and the TCR-b variable region, or portion thereof, comprises CDR1b, CDR2b, and CDR3b, wherein CDR1a comprises any one of the CDR1a sequences listed in Table 1, CDR2a comprises any one of the CDR2a sequences listed in Table 1, CDR3a comprises any one of the CDR3a sequences listed in Table 1, CDR1b comprises any one of the CDR1b sequences listed in Table 2, CDR2b comprises any one of the CDR2b sequences listed in Table 2, and CDR3b comprises any one of the CDR1b sequences listed in Table 3.

[0023] Provided is a TCR, or antigen-binding portion thereof, comprising any one of the CDR3b sequences set forth in Table 2, wherein the CDR1a, CDR2a, CDR3a, CDR1b, CDR2b, and CDR3b are all associated with the same TCR ID (i.e., derived from the same clone) listed in Table 1 and Table 2.

[0014] The TCR-a variable region may comprise a sequence at least 70%, 80%, 90%, 95% or 100% identical to SEQ ID NO: 71, a sequence at least 70%, 80%, 90%, 95% or 100% identical to SEQ ID NO: 72, a sequence at least 70%, 80%, 90%, 95% or 100% identical to SEQ ID NO: 73, and a sequence at least 70%, 80%, 90%, 95% or 100% identical to SEQ ID NO: 74, and / or The R-b variable region may comprise a sequence at least 70%, 80%, 90%, 95%, or 100% identical to SEQ ID NO: 75, a sequence at least 70%, 80%, 90%, 95%, or 100% identical to any one of SEQ ID NOs: 76-79, a sequence at least 70%, 80%, 90%, 95%, or 100% identical to SEQ ID NO: 80, and a sequence at least 70%, 80%, 90%, 95%, or 100% identical to SEQ ID NO: 81.

[0015] The TCR-a variable region may comprise a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 3, 82 to 116, 142, and 143, or a portion thereof, and / or the TCR-b variable region may comprise a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 10, 117 to 121, and 144, or a portion thereof. The TCR-a variable region may comprise a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 3 and 82 to 116, or a portion thereof, and / or the TCR-b variable region may comprise a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 10 and 117 to 121, or a portion thereof.

[0016] In another aspect, a TCR capable of specifically binding to the peptide set forth in SEQ ID NO: 122 presented on an MHC class I molecule, or an antigen-binding portion thereof, comprising a TCR-a variable region comprising a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99% or 100% identical to, or a portion of, any one of SEQ ID NOs: 3, 82 to 116, 142, and 143, and a TCR-b variable region comprising a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99% or 100% identical to, or a portion of, any one of SEQ ID NOs: 10, 117 to 121, and 144, wherein the TCR does not comprise both SEQ ID NO: 3 and SEQ ID NO: 10, and / or the TCR-a variable region sequence and the TCR-b variable region sequence are the same TCRs listed in Table 3 and Table 4. TCRs, or antigen-binding portions thereof, associated with IDs are provided. In examples, the CDR3a sequence is unchanged and the CDR3b sequence is unchanged, where the CDR3a sequence is defined in Table 3 and the CDR3b sequence is defined in Table 4, and / or the CDR1a, CDR2a, and CDR3a sequences are unchanged and the CDR1b, CDR2b, and CDR3b sequences are unchanged, where the CDR sequences are defined in Tables 3 and 4, and / or the relevant mutations listed in Table 6 are retained.

[0017] In one aspect, a nucleic acid molecule is provided that encodes: (a) a TCRa variable region amino acid sequence; (b) a TCRb variable region amino acid sequence; or (c) the amino acid sequences of both the TCRa variable region and the TCRb variable region of any TCR or antigen-binding portion disclosed herein.

[0018] In one aspect, a vector is provided that includes any nucleic acid or nucleic acids disclosed herein.

[0019] In one aspect, a cell is provided that comprises any TCR, or antigen-binding portion thereof, any nucleic acid or nucleic acids disclosed herein, or any vector disclosed herein.

[0020] In certain aspects, a pharmaceutical composition is provided comprising any TCR, or antigen-binding portion thereof, any nucleic acid or nucleic acids disclosed herein, or any vector disclosed herein, or any cell disclosed herein.

[0021] In one aspect, there is provided any TCR, or antigen-binding portion thereof, any nucleic acid or nucleic acids disclosed herein, any vector disclosed herein, any cell disclosed herein, or any pharmaceutical composition disclosed herein for use as a medicament.

[0022] In one aspect, any TCR, or antigen-binding portion thereof, any nucleic acid or nucleic acids disclosed herein, any vector disclosed herein, any cell disclosed herein, or any pharmaceutical composition disclosed herein is provided for use in a method for treating, preventing, or ameliorating cancer. In an example, the cancer comprises cells that express MAGE-A4. In an example, the cancer is non-small cell lung cancer (NSCLC), ovarian cancer, gastroesophageal cancer, head and neck synovial sarcoma, or urothelial carcinoma. [Brief explanation of the drawings]

[0023] [Figure 1] FIG. 1 provides a graph showing the calculation of Kd data in Example 6 for selected TCR clones. [Figure 2] FIG. 2 provides a graph showing the calculation of EC50 data in Example 6 for selected TCR clones. [Figure 3] FIG. 3 provides a graph showing the response to NCI-H1703 for selected TCR clones, as discussed in Example 6. [Figure 4]FIG. 4 provides a graph showing the response to NCI-H1703 for selected TCR clones, as discussed in Example 7. DETAILED DESCRIPTION OF THE INVENTION

[0024] The present inventors provide herein a TCR, in particular a MAGE-A4-specific TCR, which is capable of specifically binding to a peptide derived from the MAGE-A4 antigen when presented by an appropriate MHC molecule. Also provided is an antigen-binding portion of the TCR.

[0025] Thus, in one aspect there is provided a TCR, or an antigen binding portion thereof, capable of specifically binding to a peptide of sequence KVLEHVVRV (SEQ ID NO: 122) presented on an MHC class I molecule, comprising: TCR-alpha (TCR-a) variable region, or a portion thereof, and TCR-beta (TCR-b) variable region, or a portion thereof Including, the TCR-a variable region, or a portion thereof, comprises a complementarity determining region 3 alpha (CDR3a) comprising SEQ ID NO:49 or SEQ ID NO:53, wherein the CDR3a does not comprise SEQ ID NO:7; and / or the TCR-b variable region, or a portion thereof, comprises a complementarity determining region 3 beta (CDR3b) comprising SEQ ID NO: 62 or SEQ ID NO: 64, wherein CDR3b does not comprise SEQ ID NO: 14; TCRs, or antigen-binding portions thereof, are provided.

[0026] Thus, in one example, a TCR, or antigen-binding portion thereof, capable of specifically binding to a peptide of sequence KVLEHVVRV (SEQ ID NO: 122) presented on an MHC class I molecule, TCR-a variable region, or a portion thereof, and TCR-b variable region, or a portion thereof Including, the TCR-a variable region, or a portion thereof, comprises a CDR3a comprising SEQ ID NO: 49 or SEQ ID NO: 53, wherein the CDR3a does not comprise SEQ ID NO: 7; TCRs, or antigen-binding portions thereof, are provided.

[0027] In one embodiment, the TCR-a variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 49, wherein the CDR3a does not comprise SEQ ID NO: 7, and the TCR-b variable region comprises any CDR3b disclosed herein, for example, a CDR3b comprising or according to any one of SEQ ID NOs: 14 and 57-66.

[0028] In another embodiment, the TCR-a variable region, or portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 53, wherein the CDR3a does not comprise SEQ ID NO: 54, and the TCR-b variable region comprises any CDR3b disclosed herein, for example, a CDR3b comprising or according to any one of SEQ ID NOs: 14 and 57-66.

[0029] Furthermore, a TCR, or an antigen-binding portion thereof, capable of specifically binding to a peptide of the sequence KVLEHVVRV (SEQ ID NO: 122) presented on an MHC class I molecule, TCR-a variable region, or a portion thereof, and TCR-b variable region, or a portion thereof Including, the TCR-b variable region, or a portion thereof, comprises a CDR3b comprising SEQ ID NO: 62 or SEQ ID NO: 64, wherein the CDR3b does not comprise SEQ ID NO: 14; TCRs, or antigen-binding portions thereof, are provided.

[0030] In certain embodiments, the TCR-a variable region, or a portion thereof, comprises any CDR3a disclosed herein, and the TCR-b variable region, or a portion thereof, comprises a CDR3b comprising SEQ ID NO: 62, wherein the CDR3b does not comprise SEQ ID NO: 14. In some examples, the CDR3a comprises any one of SEQ ID NOs: 7, 26-55, 140, and 141, or optionally any one of SEQ ID NOs: 7 and 26-55.

[0031] In certain embodiments, the TCR-a variable region, or a portion thereof, comprises any CDR3a disclosed herein, and the TCR-b variable region, or a portion thereof, comprises a CDR3b comprising SEQ ID NO: 64, wherein the CDR3b does not comprise SEQ ID NO: 65. In some examples, the CDR3a comprises any one of SEQ ID NOs: 7, 26-55, 140, and 141, or optionally any one of SEQ ID NOs: 7 and 26-55.

[0032] The TCR, or antigen binding portion thereof, may comprise a CDR3a comprising or in accordance with SEQ ID NO: 49, as follows: AVGSX1X2X3NYQLX4, wherein X1 is any amino acid, X2 is any amino acid, X3 is any amino acid, and X4 is any amino acid). In some examples, CDR3a does not comprise AVGSMDSNYQLI (SEQ ID NO: 7). In some examples, the amino acids X1 through X4 may be any naturally occurring or standard amino acids. In examples, X1 is M, N, R, V, or W, or a conservative substitution for any of the foregoing residues; X2 is D, N, or Q, or a conservative substitution for any of the foregoing residues; X3 is D, E, M, R, S, T, or V, or a conservative substitution for any of the foregoing residues; and / or X4 is I or V, or a conservative substitution for any of the foregoing residues. In examples, X1 is M, N, R, or W, or a conservative substitution for any of the foregoing residues; X2 is D or Q, or a conservative substitution for any of the foregoing residues; X3 is D, E, M, R, S, or V, or a conservative substitution for any of the foregoing residues; and / or X4 is I or V, or a conservative substitution for any of the foregoing residues. In examples, X1 is M, N, R, V, or W; X2 is D, N, or Q; X3 is D, E, M, R, S, T, or V; and X4 is I or V. In examples, X1 is M, N, R, or W; X2 is D or Q; X3 is D, E, M, R, S, or V; and X4 is I or V. In other examples, X1 is N, X2 is Q, and X3 and X4 are any amino acid. In some examples, X1 is N, X2 is Q, X3 is any amino acid, and X4 is I. In particular examples, X1 is N, X2 is Q, and X3 is D, E, M, R, S, or V, or a conservative substitution for any of the foregoing residues, and X4 is I. In other examples, X1 is V, X2 is N, and X3 is T; or X1 is V, X2 is N, X3 is T, and X4 is I.

[0033] A TCR, or antigen-binding portion thereof, may comprise a CDR3a comprising or in accordance with SEQ ID NO: 50, as follows: AVGSNQX1NYQLI, where X1 is any amino acid. In some examples, the amino acid of X1 may be any naturally occurring or standard amino acid. In examples, X1 is D, E, M, R, S, or V, or a conservative substitution for any of the foregoing residues.

[0034] As discussed in the Examples, residues 4, 5, 6, 8, and 9 of CDR3a (numbered according to SEQ ID NO:7-AVGSMDSNYQLI) are important for interacting with the peptide-HLA complex. Thus, a TCR, or antigen-binding portion thereof, may comprise a CDR3a comprising SEQ ID NO:53, as follows: SX1X2X3NY, where X1 is M, N, R, V, or W, or a conservative substitution thereof; X2 is D, N, or Q, or a conservative substitution thereof; and X3 is any amino acid). In some examples, X1 is M, N, R, or W or a conservative substitution thereof, and X2 is D or Q or a conservative substitution thereof. In some examples, X1 is M, N, R, V, or W, and X2 is D, N, or Q. In some examples, X1 is M, N, R, or W, and X2 is D or Q. In some examples, CDR3a does not comprise SMDSNY (SEQ ID NO: 54). In some examples, X3 may be any naturally occurring or standard amino acid. In examples, X3 is D, E, M, R, S, T, or V, or a conservative substitution. In examples, X3 is D, E, M, R, S, or V, or a conservative substitution. In particular examples, X1 is M, N, R, V, or W; X2 is D, N, or Q; and X3 is D, E, M, R, S, T, or V. In another specific example, X1 is M, N, R, or W; X2 is D, Q, or S; and X3 is D, E, M, R, S, or V. In another example, X1 is N, X2 is Q, and X3 is any amino acid. In a specific example, X1 is N, X2 is Q, and X3 is D, E, M, R, S, or V, or a conservative substitution for any of the foregoing residues. In another example, X1 is V, X2 is N, and X3 is T. In some examples, CDR3a comprises 1, 2, 3, 4, or 5 additional residues at the N-terminus and / or C-terminus. The additional residues may be contiguous with the sequence set forth in SEQ ID NO: 3, i.e., CDR3a may comprise a longer sequence derived from the parent clone.

[0035] A TCR, or antigen-binding portion thereof, may comprise a CDR3a comprising or in accordance with SEQ ID NO: 52, as follows: SNQX1NY, where X1 is any amino acid. In some examples, the amino acid of X1 may be any naturally occurring or standard amino acid. In examples, X1 is D, E, M, R, S, or V, or a conservative substitution for any of the foregoing residues.

[0036] The TCR, or antigen binding portion thereof, may comprise a CDR3a comprising or in accordance with SEQ ID NO: 55, as follows: AVGSX1X2X3NYQLX4, wherein X1 is M, N, R, or W; X2 is D or Q; X3 is D, E, M, R, S, or V; X4 is I or V. In some embodiments, CDR3a does not comprise AVGSMDSNYQLI (SEQ ID NO: 7).

[0037] The TCR, or antigen-binding portion thereof, may comprise any CDR3a sequence set out in Table 1. The TCR, or antigen-binding portion thereof, may comprise a CDR3a comprising or according to any one of SEQ ID NOs: 26 to 48, 140 and 141. The TCR, or antigen-binding portion thereof, may comprise a CDR3a comprising or according to any one of SEQ ID NOs: 7, 26 to 55, 140 and 141.

[0038] The TCR, or antigen binding portion thereof, may comprise a CDR3b comprising or in accordance with SEQ ID NO: 62, as follows: ASSQERTX1PYNEQF, wherein X1 is any amino acid). In some embodiments, CDR3b does not comprise ASSQERTWPYNEQF (SEQ ID NO: 14). In some examples, the amino acid at X1 may be any naturally occurring or standard amino acid. In examples, X1 is A, Q, or W, or a conservative substitution for any of the foregoing residues. In certain examples, X1 is A, Q, or W.

[0039] The TCR, or antigen-binding portion thereof, may comprise a CDR3b comprising or in accordance with SEQ ID NO: 63, as follows: ASSQERTX1PYNEQF, where X1 is any amino acid different from W. In examples, X1 is A or Q, or a conservative substitution for any of the foregoing residues. In some examples, X1 is A or Q.

[0040] As discussed in the Examples, residues 5 through 10 of CDR3b (numbered according to SEQ ID NO: 14-ASSQERTWPYNEQF) are important for interacting with peptide-HLA complexes. Thus, a TCR, or antigen-binding portion thereof, may comprise a CDR3b comprising SEQ ID NO: 64: ERTX1PY (wherein X1 is A, Q, or W, or a conservative substitution thereof). In one example, X1 is A, Q, or W. In some embodiments, CDR3b does not comprise ERTWPY (SEQ ID NO: 65). In some examples, CDR3b comprises 1, 2, 3, 4, 5, 6, or 7 additional residues at the N-terminus and / or C-terminus. The additional residues may be contiguous with the sequence set forth in SEQ ID NO: 10, i.e., the CDR3b may comprise a longer sequence derived from the parent clone.

[0041] The TCR, or antigen-binding portion thereof, may comprise a CDR3b comprising or in accordance with SEQ ID NO: 66, as follows: ASSQERTX1PYNEQF, where X1 is A, Q, or W. In some embodiments, the CDR3b does not comprise ASSQERTWPYNEQF (SEQ ID NO: 14).

[0042] The TCR, or antigen-binding portion thereof, may comprise any CDR3b sequence set out in Table 2. The TCR, or antigen-binding portion thereof, may comprise a CDR3b comprising or according to any one of SEQ ID NOs: 57 to 61. The TCR, or antigen-binding portion thereof, may comprise a CDR3b comprising or according to any one of SEQ ID NOs: 14 and 57 to 66.

[0043] The TCR, or antigen-binding portion thereof, may comprise any CDR1a sequence set out in Table 1. The TCR, or antigen-binding portion thereof, may comprise a CDR1a comprising or according to any one of SEQ ID NOs: 15 to 17. The TCR, or antigen-binding portion thereof, may comprise a CDR1a comprising or according to any one of SEQ ID NOs: 5 and 15 to 17.

[0044] The TCR, or antigen-binding portion thereof, may comprise CDR2a. Specifically, the TCRa variable chain, or antigen-binding portion thereof, may comprise CDR2a. The CDR2a may comprise or be in accordance with SEQ ID NO: 24, as follows: LX1X2SGEV, where: X1 is any amino acid, X2 is any amino acid). In some embodiments, CDR2a does not comprise LVKSGEV (SEQ ID NO: 6). In some examples, the amino acids of X1-X2 may be any naturally occurring or standard amino acids. In examples, X1 is E, G, P, or V, or a conservative substitution for any of the foregoing residues, and / or X2 is K, L, or Q, or a conservative substitution for any of the foregoing residues. In some examples, X1 is E, G, P, or V, and X2 is K, L, or Q.

[0045] The TCR, or antigen-binding portion thereof, may comprise CDR2a. Specifically, the TCRa variable chain, or antigen-binding portion thereof, may comprise CDR2a. The CDR2a may comprise or be in accordance with SEQ ID NO: 25, as follows: LX1KSGEV, wherein: X1 is any amino acid). In some embodiments, CDR2a does not comprise LVKSGEV (SEQ ID NO: 6). In some examples, the amino acid of X1 may be any naturally occurring or standard amino acid. In examples, X1 is E, G, P, or V, or a conservative substitution for any of the foregoing residues. In certain examples, X1 is E, G, P, or V.

[0046] The TCR, or antigen-binding portion thereof, may comprise any CDR2a sequence listed in Table 1. The TCR, or antigen-binding portion thereof, may comprise a CDR2a comprising or according to any one of SEQ ID NOs: 18 to 23. The TCR, or antigen-binding portion thereof, may comprise a CDR2a comprising or according to any one of SEQ ID NOs: 6 and 18 to 25.

[0047] The TCR, or antigen-binding portion thereof, may comprise a CDR1b sequence as set out in Table 2. The TCR, or antigen-binding portion thereof, may comprise a CDR1b comprising or in accordance with SEQ ID NO: 12.

[0048] The TCR, or antigen-binding portion thereof, may comprise any of the CDR2b sequences set out in Table 2. The TCR, or antigen-binding portion thereof, may comprise a CDR2b comprising or according to SEQ ID NO: 56. The TCR, or antigen-binding portion thereof, may comprise a CDR2b comprising or according to SEQ ID NO: 13 or SEQ ID NO: 56.

[0049] In some embodiments, a TCR, or an antigen-binding portion thereof, TCR-a variable region, or an antigen-binding portion thereof, and TCR-b variable region, or an antigen-binding portion thereof Including, the TCR-a variable region, or antigen-binding portion thereof, comprises a CDR3a comprising or in accordance with SEQ ID NO: 49; the TCR-b variable region, or antigen-binding portion thereof, comprises a CDR3b comprising or in accordance with SEQ ID NO: 62; CDR3a does not contain AVGSMDSNYQLI (SEQ ID NO: 7) and / or CDR3b does not contain ASSQERTWPYNEQF (SEQ ID NO: 14), TCRs, or antigen-binding portions thereof, are provided.

[0050] In some embodiments, a TCR, or an antigen-binding portion thereof, TCR-a variable region, or a portion thereof, and TCR-b variable region, or a portion thereof Including, the TCR-a variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 50; The TCR-b variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 66; TCRs, or antigen-binding portions thereof, are provided.

[0051] In some embodiments, a TCR, or an antigen-binding portion thereof, TCR-a variable region, or a portion thereof, and TCR-b variable region, or a portion thereof Including, the TCR-a variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 53 (SX1X2X3NY, where X1 is V, X2 is N and X3 is T); The TCR-b variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 66; TCRs, or antigen-binding portions thereof, are provided.

[0052] In some embodiments, a TCR, or an antigen-binding portion thereof, TCR-a variable region, or a portion thereof, and TCR-b variable region, or a portion thereof Including, the TCR-a variable region, or a portion thereof, comprises a CDR3a comprising or according to any one of SEQ ID NOs: 7, 26 to 55, 140 and 141, or optionally any one of SEQ ID NOs: 7 and 26 to 55, the TCR-b variable region, or a portion thereof, comprises a CDR1b comprising or according to SEQ ID NO: 12, a CDR2b comprising or according to SEQ ID NO: 13 or 56, and a CDR3b comprising or according to any one of SEQ ID NOs: 14 and 57 to 66; CDR3a does not contain SEQ ID NO: 7, CDR2b does not contain SEQ ID NO: 13, or CDR3b does not contain SEQ ID NO: 14; TCRs, or antigen-binding portions thereof, are provided.

[0053] In some embodiments, a TCR, or an antigen-binding portion thereof, TCR-a variable region, or a portion thereof, and TCR-b variable region, or a portion thereof Including, the TCR-a variable region, or a portion thereof, comprises a CDR1a comprising or according to any one of SEQ ID NOs: 5 and 15 to 17, a CDR2a comprising or according to any one of SEQ ID NOs: 6 and 18 to 25, and a CDR3a comprising or according to any one of SEQ ID NOs: 7, 26 to 55, 140, and 141, or optionally any one of SEQ ID NOs: 7 and 26 to 55, the TCR-b variable region, or a portion thereof, comprises a CDR3b comprising or according to any one of SEQ ID NOs: 14 and 57 to 66; CDR1a does not contain SEQ ID NO:5, CDR2a does not contain SEQ ID NO:6, CDR3a does not contain SEQ ID NO:7, or CDR3b does not contain SEQ ID NO:14; TCRs, or antigen-binding portions thereof, are provided.

[0054] In some embodiments, a TCR, or an antigen-binding portion thereof, TCR-a variable region, or a portion thereof, and TCR-b variable region, or a portion thereof Including, the TCR-a variable region, or a portion thereof, comprises a CDR1a comprising or according to any one of SEQ ID NOs: 5 and 15 to 17, a CDR2a comprising or according to any one of SEQ ID NOs: 6 and 18 to 25, and a CDR3a comprising or according to any one of SEQ ID NOs: 7, 26 to 55, 140, and 141, or optionally any one of SEQ ID NOs: 7 and 26 to 55, the TCR-b variable region, or a portion thereof, comprises a CDR1b comprising or according to SEQ ID NO: 12, a CDR2b comprising or according to SEQ ID NO: 13 or 56, and a CDR3b comprising or according to any one of SEQ ID NOs: 14 and 57 to 66; CDR1a does not contain SEQ ID NO:5, CDR2a does not contain SEQ ID NO:6, CDR3a does not contain SEQ ID NO:7, CDR2b does not contain SEQ ID NO:13, or CDR3b does not contain SEQ ID NO:14; TCRs, or antigen-binding portions thereof, are provided.

[0055] In some embodiments, a TCR, or an antigen-binding portion thereof, TCR-a variable region, or a portion thereof, and TCR-b variable region, or a portion thereof Including, the TCR-a variable region, or a portion thereof, comprises a CDR1a comprising or according to any one of SEQ ID NOs: 5 and 15 to 17, a CDR2a comprising or according to any one of SEQ ID NOs: 6 and 18 to 23, and a CDR3a comprising or according to any one of SEQ ID NOs: 7, 26 to 48, 140, and 141, or optionally any one of SEQ ID NOs: 7 and 26 to 48, the TCR-b variable region, or a portion thereof, comprises a CDR1b comprising or according to SEQ ID NO: 12, a CDR2b comprising or according to SEQ ID NO: 13 or 56, and a CDR3b comprising or according to any one of SEQ ID NOs: 14 and 57 to 61; CDR1a does not contain SEQ ID NO:5, CDR2a does not contain SEQ ID NO:6, CDR3b does not contain SEQ ID NO:7, CDR2b does not contain SEQ ID NO:13, or CDR3b does not contain SEQ ID NO:14; TCRs, or antigen-binding portions thereof, are provided. In some embodiments, the CDRs are associated with the same TCR ID in Table 1 and Table 2, i.e., are derived from the same clone.

[0056] [Table 1A]

[0057] [Table 1B]

[0058] In some embodiments, a TCRa variable region, or a portion thereof, may comprise one or more of the CDR sequences listed in the table above. For example, one of the CDR3a sequences. In another example, a TCRa variable region, or a portion thereof, may comprise one of the CDR1a sequences, one of the CDR2a sequences, or one of the CDR3a sequences.

[0059] In some embodiments, the TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a set forth in Table 1. In some embodiments, the TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a set forth in the same row of Table 1 above (i.e., derived from the same clone).

[0060] [Table 2A]

[0061] [Table 2B]

[0062] In some embodiments, the TCRb variable region, or a portion thereof, may comprise one or more of the CDR sequences listed in the table above. For example, one of the CDR3b sequences. In another example, the TCRb variable region, or a portion thereof, may comprise one of the CDR1b sequences, one of the CDR2b sequences, or one of the CDR3b sequences.

[0063] In some embodiments, the TCRb variable region comprises CDR1b, CDR2b, and CDR3b set forth in Table 2. In some embodiments, the TCRb variable region comprises CDR1b, CDR2b, and CDR3b set forth in the same row of Table 2 above (i.e., from the same clone).

[0064] The TCR, or antigen-binding portion thereof, may comprise a CDR3a comprising any one of SEQ ID NOs: 7, 26 to 48, 140, and 141, or optionally any one of SEQ ID NOs: 7 and 26 to 48, a CDR1b comprising SEQ ID NO: 12, a CDR2b comprising SEQ ID NO: 13 or SEQ ID NO: 56, and a CDR3b comprising any one of SEQ ID NOs: 14 and 57 to 61. The CDR3a, CDR1b, CDR2b, and CDR3b may be associated with the same TCR ID in Table 1 and Table 2 (i.e., derived from the same clone).

[0065] The TCR, or antigen-binding portion thereof, may comprise CDR1a comprising any one of SEQ ID NOs: 5, 15 to 17, CDR2a comprising any one of SEQ ID NOs: 6 and 18 to 23, CDR3a comprising any one of SEQ ID NOs: 7, 26 to 48, 140 and 141, or optionally any one of SEQ ID NOs: 7 and 26 to 48, and CDR3b comprising any one of SEQ ID NOs: 14 and 57 to 61. CDR1a, CDR2a, CDR3a and CDR3b may be associated with the same TCR ID in Table 1 and Table 2 (i.e., derived from the same clone).

[0066] The TCR, or antigen-binding portion thereof, may comprise CDR1a comprising any one of SEQ ID NOs: 5, 15 to 17, CDR2a comprising any one of SEQ ID NOs: 6 and 18 to 23, CDR3a comprising any one of SEQ ID NOs: 7, 26 to 48, 140 and 141, or optionally any one of SEQ ID NOs: 7 and 26 to 48, CDR1b comprising SEQ ID NO: 12, CDR2b comprising SEQ ID NO: 13 or SEQ ID NO: 56, and CDR3b comprising any one of SEQ ID NOs: 14 and 57 to 61. CDR1a, CDR2a, CDR3a, CDR1b, CDR2b and CDR3b may be associated with the same TCR ID in Table 1 and Table 2 (i.e., derived from the same clone).

[0067] In one aspect, the TCR, or antigen binding portion thereof, (i) a TCRa variable region, or a portion thereof, comprising CDR3a and a TCRb variable region, or a portion thereof, comprising CDR3b; or (ii) a TCRa variable region, or a portion thereof, including CDR3a, and a TCRb variable region, or a portion thereof, including CDR1b, CDR2b, and CDR3b; or (iii) a TCRa variable region, or a portion thereof, including CDR1a, CDR2a, and CDR3a, and a TCRb variable region, or a portion thereof, including CDR3b; or (iv) a TCRa variable region, or a portion thereof, including CDR1a, CDR2a, and CDR3a, and a TCRb variable region, or a portion thereof, including CDR1b, CDR2b, and CDR3b. may include Here, the CDR sequences are clones KVL-01-018, KVL-01-019, KVL-01-020, KVL-01-022, KVL-01-026, KVL-01-034, KVL-01-036, KVL-01-039, KVL-01-040, KVL-01-041, KVL-01-042, KVL-01-043, KVL-01-048, KVL-01-049、KVL-01-050、KVL-01-052、KVL-01-056、KVL-01-060、KVL-01-061、KVL-01-068、KVL-01-069、KVL-01-070、KVL-01-071、KVL-01-073、KVL-01-074、KVL-01-075、KVL-01-076、KVL-0 1-077、KVL-01-078、KVL-01-079、KVL-01-083、KVL-01-084、KVL-01-088、KVL-01-089、KVL-01-090、KVL-01-094、KVL-01-097、KVL-01-105、KVL-01-108、KVL-01-110、KVL-01-111、KVL-01-11 2、KVL-01-117、KVL-01-119、KVL-01-123、KVL-01-124、KVL-01-125、KVL-01-126、KVL-01-128、 One of KVL-01-129、KVL-01-145、KVL-01-149、KVL-01-151、or KVL-01-153 is a set of CDR sequence derived from it.

[0068] The TCR, or antigen-binding portion thereof, may comprise a CDR3a comprising any one of SEQ ID NOs: 7, 26-55, 140, and 141, or optionally any one of SEQ ID NOs: 7 and 26-55, where CDR3a has been grafted into a human TCRa variable region framework, and CDR3b has been grafted into a human TCRb variable region framework. The TCR, or antigen-binding portion thereof, may comprise a CDR3a comprising any one of SEQ ID NOs: 7, 26-55, 140, and 141, or optionally any one of SEQ ID NOs: 7 and 26-55, where CDR1b has been grafted into a human TCRa variable region framework, and CDR2b has been grafted into a human TCRb variable region framework. The TCR, or antigen-binding portion thereof, may comprise CDR1a comprising any one of SEQ ID NOs: 5 and 15-17, CDR2a comprising any one of SEQ ID NOs: 6 and 18-25, CDR3a comprising any one of SEQ ID NOs: 7, 26-55, 140, and 141, or optionally any one of SEQ ID NOs: 7 and 26-55, and CDR3b comprising any one of SEQ ID NOs: 14 and 57-66, wherein CDR1a, CDR2a, and CDR3a are grafted into a human TCRb variable region framework and CDR3b is grafted into a human TCRb variable region framework. The TCR, or antigen-binding portion thereof, may comprise CDR1a comprising any one of SEQ ID NOs: 5 and 15-17, CDR2a comprising any one of SEQ ID NOs: 6 and 18-25, CDR3a comprising any one of SEQ ID NOs: 7, 26-55, 140, and 141, or optionally any one of SEQ ID NOs: 7 and 26-55, CDR1b comprising SEQ ID NO: 12, CDR2b comprising SEQ ID NO: 13 or SEQ ID NO: 56, and CDR3b comprising any one of SEQ ID NOs: 14 and 57-66, wherein CDR1a, CDR2a, and CDR3a are grafted into a human TCRb variable region framework, and CDR1b, CDR2b, and CDR3b are grafted into a human TCRb variable region framework.In any such embodiment, the TCR, or antigen binding portion thereof, does not comprise all six of SEQ ID NOs: 5, 6, 7, 12, 13, and 14. In any such embodiment, the CDRs may be associated with the same TCR ID, i.e., a set of CDRs derived from the same clone.

[0069] Any of the CDR sequences disclosed herein may comprise one, two, or three additional residues at the N-terminus and / or C-terminus. Any of the CDR sequences disclosed herein may comprise one to ten additional residues at the N-terminus and / or C-terminus. The additional residues may be contiguous with the CDR listed in the relevant sequence in Table 3 or Table 4, i.e., the CDR may comprise a longer sequence derived from the parent clone. In some examples, the CDR grafted onto the human TCR framework may be that listed in the CDRs listed in Table 1 and Table 2 and may comprise one, two, or three additional residues at the N-terminus and / or C-terminus, where the additional residues are contiguous residues of the relevant sequence in Table 3 or Table 4. In any such embodiment, the grafted CDRs may be associated with the same TCR ID, i.e., may be derived from the same clone.

[0070] The TCRa variable region may comprise a first framework region (FR1a) of the sequence: GQQVMQIPQYQHVQEGEDFTTYCNSS (SEQ ID NO: 71). The TCRa variable region may comprise a second framework region (FR2a) of the sequence: IQWYKQRPGGHPVFLIQ (SEQ ID NO: 72). The TCRa variable region may comprise a third framework region (FR3a) of the sequence: KKQKRLTFQFGEAKKNSSLHITATQTTDVGTYF (SEQ ID NO: 73). The TCRa variable region may comprise a fourth framework region (FR4a) of the sequence: GAGTKLIX1KP (SEQ ID NO: 74), where X1 is I or V.

[0071] The TCRa variable region may comprise a first framework region that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 71. The TCRa variable region may comprise a second framework region that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 72. The TCRa variable region may comprise a third framework region that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 73. The TCRa variable region may comprise a fourth framework region that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 74.

[0072] The TCRb variable region may comprise a first framework region (FR1b) of the sequence: NAGVTQTPKFRVLKTGQSMTLLCAQD (SEQ ID NO: 75). The TCRb variable region may comprise a second framework region (FR2b) of the sequence: X1YWYRQDPX2MGLRLIHY (SEQ ID NO: 76) (wherein X1 is M or L and X2 is D or G). The TCRb variable region may comprise a third framework region (FR3b) of the sequence: TX1KGEVPDGYNVSRLKKQNFLLGLESAAPSQTSVYF (SEQ ID NO: 80) (wherein X1 is A or T). The TCRb variable region may comprise a fourth framework region (FR4b) of the sequence: GPGTRLTVL (SEQ ID NO: 81).

[0073] In some embodiments, the FR2b sequence may include or be MYWYRQDPDMGLRLIHY (SEQ ID NO: 77), MYWYRQDPGMGLRLIHY (SEQ ID NO: 78), or LYWYRQDPGMGLRLIHY (SEQ ID NO: 79).

[0074] The TCRb variable region may comprise a first framework region that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 75. The TCRb variable region may comprise a second framework region that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any of SEQ ID NOs: 76-79. The TCRb variable region may comprise a third framework region that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 80. The TCRb variable region may comprise a fourth framework region that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 81.

[0075] Any of SEQ ID NOs: 71-81 may contain 1, 2, 3, or 4 conservative substitutions of the depicted sequence. Any of SEQ ID NOs: 71-81 may carry the mutations listed in Table 6.

[0076] In one aspect, a TCR, or an antigen-binding portion thereof, TCR-a variable region, or an antigen-binding portion thereof, and TCR-b variable region, or an antigen-binding portion thereof Including, the TCR-a variable region comprises, in order, a sequence at least 70%, 80%, 90%, 95% or 100% identical to SEQ ID NO: 71, CDR1a, a sequence at least 70%, 80%, 90%, 95% or 100% identical to SEQ ID NO: 72, CDR2a, a sequence at least 70%, 80%, 90%, 95% or 100% identical to SEQ ID NO: 73, CDR3a, and a sequence at least 70%, 80%, 90%, 95% or 100% identical to SEQ ID NO: 74; CDR3a comprises or is in accordance with any one of SEQ ID NOs: 7, 26 to 55, 140, and 141, or optionally any one of SEQ ID NOs: 7 and 26 to 55, the TCR-b variable region comprises, in order, a sequence at least 70%, 80%, 90%, 95%, or 100% identical to SEQ ID NO: 75, CDR1b, a sequence at least 70%, 80%, 90%, 95%, or 100% identical to any one of SEQ ID NOs: 76 to 79, CDR2b, a sequence at least 70%, 80%, 90%, 95%, or 100% identical to SEQ ID NO: 80, CDR3b, and a sequence at least 70%, 80%, 90%, 95%, or 100% identical to SEQ ID NO: 81; CDR1b comprises or is according to SEQ ID NO: 12, CDR2b comprises or is according to SEQ ID NO: 13 or SEQ ID NO: 56, CDR3b comprises or is in accordance with any one of SEQ ID NOs: 14 and 57 to 66; TCRs, or antigen-binding portions thereof, are provided.

[0077] In any such embodiment, the TCR, or antigen-binding portion thereof, does not comprise all six of SEQ ID NOs: 5, 6, 7, 12, 13, and 14. In any such embodiment, the CDRs may be associated with the same TCR ID, i.e., a set of CDRs derived from the same clone. In some embodiments, the framework regions have 95%, 97%, 98%, 99%, or 100% identity to the sequences listed. Any of SEQ ID NOs: 71-81 may possess the mutations listed in Table 6.

[0078] In one aspect, a TCR, or an antigen-binding portion thereof, TCR-a variable region, or an antigen-binding portion thereof, and TCR-b variable region, or an antigen-binding portion thereof Including, the TCR-a variable region comprises, in order, a sequence at least 70%, 80%, 90%, 95% or 100% identical to SEQ ID NO: 71, CDR1a, a sequence at least 70%, 80%, 90%, 95% or 100% identical to SEQ ID NO: 72, CDR2a, a sequence at least 70%, 80%, 90%, 95% or 100% identical to SEQ ID NO: 73, CDR3a, and a sequence at least 70%, 80%, 90%, 95% or 100% identical to SEQ ID NO: 74; CDR1a comprises or is according to any one of SEQ ID NOs: 5 and 15 to 17, CDR2a comprises or is in accordance with any one of SEQ ID NOs: 6 and 18 to 25, CDR3a comprises or is in accordance with any one of SEQ ID NOs: 7, 26 to 55, 140, and 141, or optionally any one of SEQ ID NOs: 7 and 26 to 55, the TCR-b variable region comprises, in order, a sequence at least 70%, 80%, 90%, 95%, or 100% identical to SEQ ID NO: 75, CDR1b, a sequence at least 70%, 80%, 90%, 95%, or 100% identical to any one of SEQ ID NOs: 76 to 79, CDR2b, a sequence at least 70%, 80%, 90%, 95%, or 100% identical to SEQ ID NO: 80, CDR3b, and a sequence at least 70%, 80%, 90%, 95%, or 100% identical to SEQ ID NO: 81; CDR3b comprises or is in accordance with any one of SEQ ID NOs: 14 and 57 to 66; TCRs, or antigen-binding portions thereof, are provided.

[0079] In any such embodiment, the TCR, or antigen-binding portion thereof, does not comprise all six of SEQ ID NOs: 5, 6, 7, 12, 13, and 14. In any such embodiment, the CDRs may be associated with the same TCR ID, i.e., a set of CDRs derived from the same clone. In some embodiments, the framework regions have 95%, 97%, 98%, 99%, or 100% identity to the sequences listed. Any of SEQ ID NOs: 71-81 may possess the mutations listed in Table 6.

[0080] In one aspect, a TCR, or an antigen-binding portion thereof, TCR-a variable region, or an antigen-binding portion thereof, and TCR-b variable region, or an antigen-binding portion thereof Including, the TCR-a variable region comprises, in order, a sequence at least 70%, 80%, 90%, 95% or 100% identical to SEQ ID NO: 71, CDR1a, a sequence at least 70%, 80%, 90%, 95% or 100% identical to SEQ ID NO: 72, CDR2a, a sequence at least 70%, 80%, 90%, 95% or 100% identical to SEQ ID NO: 73, CDR3a, and a sequence at least 70%, 80%, 90%, 95% or 100% identical to SEQ ID NO: 74; CDR1a comprises or is according to any one of SEQ ID NOs: 5 and 15 to 17, CDR2a comprises or is in accordance with any one of SEQ ID NOs: 6 and 18 to 25, CDR3a comprises or is in accordance with any one of SEQ ID NOs: 7, 26 to 55, 140, and 141, or optionally any one of SEQ ID NOs: 7 and 26 to 55, the TCR-b variable region comprises, in order, a sequence at least 70%, 80%, 90%, 95%, or 100% identical to SEQ ID NO: 75, CDR1b, a sequence at least 70%, 80%, 90%, 95%, or 100% identical to any one of SEQ ID NOs: 76 to 79, CDR2b, a sequence at least 70%, 80%, 90%, 95%, or 100% identical to SEQ ID NO: 80, CDR3b, and a sequence at least 70%, 80%, 90%, 95%, or 100% identical to SEQ ID NO: 81; CDR1b comprises or is according to SEQ ID NO: 12, CDR2b comprises or is according to SEQ ID NO: 13 or SEQ ID NO: 56, CDR3b comprises or is in accordance with any one of SEQ ID NOs: 14 and 57 to 66; TCRs, or antigen-binding portions thereof, are provided.

[0081] In any such embodiment, the TCR, or antigen-binding portion thereof, does not comprise all six of SEQ ID NOs: 5, 6, 7, 12, 13, and 14. In any such embodiment, the CDRs may be associated with the same TCR ID, i.e., a set of CDRs derived from the same clone. In some embodiments, the framework regions have 95%, 97%, 98%, 99%, or 100% identity to the sequences listed. Any of SEQ ID NOs: 71-81 may possess the mutations listed in Table 6.

[0082] [Table 3A]

[0083] [Table 3B]

[0084] In one embodiment, the TCR, or antigen-binding portion thereof, comprises an alpha chain variable region comprising or according to any one of the amino acid sequences set forth in Table 3.

[0085] In some embodiments, the TCR, or antigen binding portion thereof, comprises any one of SEQ ID NOs: 3, 82-116, 142, and 143. The TCR, or antigen binding portion thereof, may comprise a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 3, 82-116, 142, and 143. In some embodiments, the TCR, or antigen binding portion thereof, comprises a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 3, 82-116, 142, and 143, where CDR3a is unchanged. The TCR, or antigen-binding portion thereof, may comprise a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 3, 82-116, 142, and 143, in which CDR1a, CDR2a, and CDR3a are unchanged. The CDR sequences may be those set forth in Table 1.

[0086] The TCR, or antigen-binding portion thereof, may include any of the alpha chain mutations listed in Table 6. Where less than 100% sequence identity is exhibited, any of SEQ ID NOs: 82-116, 142, and 143 may retain the mutation associated with the related clone in Table 6.

[0087] As used herein, a CDR is said to be "invariant" if it is maintained without change in a sequence that is not 100% identical to the otherwise described variable region sequence.

[0088] The antigen-binding portion of the TCR may comprise a fragment of any one of SEQ ID NOs: 3, 82 to 116, 142, and 143. The antigen-binding portion of the TCR may comprise a fragment that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to the corresponding portion of any one of SEQ ID NOs: 3, 82 to 116, 142, and 143. The antigen-binding portion of the TCR may comprise a fragment that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to the corresponding portion of any one of SEQ ID NOs: 3, 82 to 116, 142, and 143, in which CDR3a is retained and unchanged. The antigen-binding portion of the TCR may comprise a fragment that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to the corresponding portion of any one of SEQ ID NOs: 3, 82-116, 142, and 143, in which CDR1a, CDR2a, and CDR3a are retained and unchanged. The CDR sequences may be those set out in Table 1.

[0089] The TCR may comprise an alpha chain variable region set forth in any one of SEQ ID NOs: 3, 82-116, 142, and 143, or an antigen-binding portion thereof. The TCR may comprise an alpha chain variable region set forth in a sequence that is at least 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 3, 82-116, 142, and 143, or an antigen-binding portion thereof. In some embodiments, the TCR comprises an alpha chain variable region set forth in a sequence that is at least 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 3, 82-116, 142, and 143, or an antigen-binding portion thereof, wherein CDR3a is unchanged. The TCR may comprise an alpha chain variable region, or an antigen binding portion thereof, set forth in a sequence that is at least 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 3, 82 to 116, 142 and 143, in which CDR1a, CDR2a and CDR3a are unchanged.The TCR may comprise an alpha chain variable region, or an antigen binding portion thereof, set forth in a sequence that is at least 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 3, 82 to 116, 142 and 143, in which CDR1a, CDR2a and CDR3a are unchanged.

[0090] [Table 4A]

[0091] [Table 4B]

[0092] In one embodiment, the TCR, or antigen-binding portion thereof, comprises a beta chain variable region comprising or according to any one of the amino acid sequences set forth in Table 4.

[0093] In some embodiments, the TCR, or antigen binding portion thereof, comprises any one of SEQ ID NOs: 10, 117-121, and 144. The TCR, or antigen binding portion thereof, may comprise a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 10, 117-121, and 144. In some embodiments, the TCR, or antigen binding portion thereof, comprises a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 10, 117-121, and 144, in which the CDR3b is unchanged. The TCR, or antigen-binding portion thereof, may comprise a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 10, 117-121, and 144, in which CDR1b, CDR2b, and CDR3b are unchanged. The CDR sequences may be those set forth in Table 2.

[0094] The TCR, or antigen-binding portion thereof, may contain any of the beta chain mutations listed in Table 6. If exhibiting less than 100% sequence identity, any of SEQ ID NOs: 117-121, and 144 may retain the mutation associated with the related clone in Table 6.

[0095] The antigen-binding portion of the TCR may comprise a fragment of any one of SEQ ID NOs: 10, 117-121, and 144. The antigen-binding portion of the TCR may comprise a fragment that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to the corresponding portion of any one of SEQ ID NOs: 10, 117-121, and 144. The antigen-binding portion of the TCR may comprise a fragment that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to the corresponding portion of any one of SEQ ID NOs: 10, 117-121, and 144, in which the CDR3b is retained and unchanged. The antigen-binding portion of the TCR may comprise a fragment that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to the corresponding portion of any one of SEQ ID NOs: 10, 117-121, and 144, in which CDR1b, CDR2b, and CDR3b are retained and unchanged. The CDR sequences may be those set out in Table 2.

[0096] The TCR may comprise a beta chain variable region set forth in any one of SEQ ID NOs: 10, 117-121, and 144, or an antigen-binding portion thereof. The TCR may comprise a beta chain variable region set forth in a sequence that is at least 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 10, 117-121, and 144, or an antigen-binding portion thereof. In some embodiments, the TCR comprises a beta chain variable region set forth in a sequence that is at least 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 10, 117-121, and 144, or an antigen-binding portion thereof, wherein CDR3b is unchanged. The TCR may comprise a beta chain variable region, or an antigen-binding portion thereof, set forth in a sequence that is at least 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 10, 117-121, and 144, in which CDR1b, CDR2b, and CDR3b are unchanged.

[0097] The TCR may comprise a sequence set out in Table 3 and a sequence set out in Table 4. The sequence from Table 3 and the sequence from Table 4 may be of the same TCR ID, i.e. may be from the same clone. Antigen-binding portions of said TCRs and said sequences are also provided.

[0098] The TCR may comprise an alpha chain variable region set forth in any one of SEQ ID NOs: 3, 82-116, 142, and 143 and a beta chain variable region set forth in any one of SEQ ID NOs: 10, 117-121, and 144; or optionally an alpha chain variable region set forth in any one of SEQ ID NOs: 3 and 82-116 and a beta chain variable region set forth in any one of SEQ ID NOs: 10 and 117-121. The alpha chain variable region and the beta chain variable region may be of the same TCR ID, i.e., derived from the same clone. An antigen-binding portion of the TCR is also provided. The TCR is not of the sequence set forth in SEQ ID NO: 3 or SEQ ID NO: 10.

[0099] The TCR may comprise an alpha chain variable region set forth in a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 3, 82-116, 142, and 143, and a beta chain variable region set forth in a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 10, 117-121, and 144. The TCR may comprise an alpha chain variable region set forth in a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 3, 82 to 116, 142 and 143 and a beta chain variable region set forth in a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 10, 117 to 121 and 144, wherein the alpha chain variable region and the beta chain variable region have the same TCR ID in Table 3 and Table 4. The TCR may comprise an alpha chain variable region set forth in a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 3, 82 to 116, 142 and 143, and a beta chain variable region set forth in a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 10, 117 to 121 and 144, in which CDR3a and CDR3b are unchanged. The TCR may comprise an alpha chain variable region set forth in a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 3, 82 to 116, 142 and 143 and a beta chain variable region set forth in a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 10, 117 to 121 and 144, wherein the alpha chain variable region and the beta chain variable region have the same TCR ID in Table 3 and Table 4, and CDR3a and CDR3b are unchanged.The TCR may comprise an alpha chain variable region set forth in a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 3, 82-116, 142, and 143, and a beta chain variable region set forth in a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 10, 117-121, and 144, in which CDR3a, CDR1b, CDR2b, and / or CDR3b are unchanged. The TCR may comprise an alpha chain variable region set forth in a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 3, 82 to 116, 142 and 143 and a beta chain variable region set forth in a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 10, 117 to 121 and 144, wherein the alpha chain variable region and the beta chain variable region have the same TCR ID in Table 3 and Table 4, and wherein CDR3a, CDR1b, CDR2b and / or CDR3b are unchanged. The TCR may comprise an alpha chain variable region set forth in a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 3, 82 to 116, 142, and 143, and a beta chain variable region set forth in a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 10, 117 to 121, and 144, in which CDR1a, CDR2a, CDR3a, and / or CDR3b are unchanged.The TCR may comprise an alpha chain variable region set forth in a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 3, 82 to 116, 142 and 143 and a beta chain variable region set forth in a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 10, 117 to 121 and 144, wherein the alpha chain variable region and the beta chain variable region have the same TCR ID in Table 3 and Table 4, and CDR1a, CDR2a, CDR3a and / or CDR3b are unchanged. The TCR may comprise an alpha chain variable region set forth in a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 3, 82 to 116, 142, and 143, and a beta chain variable region set forth in a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 10, 117 to 121, and 144, wherein CDR1a, CDR2a, CDR3a, CDR1b, CDR2b, and / or CDR3b are unchanged. The TCR may comprise an alpha chain variable region set forth in a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 3, 82-116, 142, and 143, and a beta chain variable region set forth in a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 10, 117-121, and 144, wherein the alpha chain variable region and beta chain variable region have the same TCR ID in Table 3 and Table 4, and CDR1a, CDR2a, CDR3a, CDR1b, CDR2b, and / or CDR3b are unchanged. Full antigen-binding portions of the TCRs are also provided. The TCR does not comprise both SEQ ID NO: 3 and SEQ ID NO: 10. In any of the descriptions in this paragraph, the combination may be SEQ ID NO: 3 and one of 82-116 with SEQ ID NO: 10 and one of 117-121.

[0100] The TCR, or antigen-binding portion thereof, may include any of the alpha and / or beta chain mutations listed in Table 6. In some instances, the alpha and beta chain mutations may be from the same row in Table 6. Where less than 100% sequence identity is exhibited, any of SEQ ID NOs: 82-121 and 142-144 may retain mutations associated with the related clone in Table 6.

[0101] The TCR may comprise an alpha chain variable region set forth in a sequence that is at least 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 3, 82-116, 142, and 143, and a beta chain variable region set forth in a sequence that is at least 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 10, 117-121, and 144. The TCR may comprise an alpha chain variable region set forth in a sequence that is at least 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 3, 82 to 116, 142 and 143 and a beta chain variable region set forth in a sequence that is at least 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 10, 117 to 121 and 144, wherein the alpha chain variable region and the beta chain variable region have the same TCR ID in Table 3 and Table 4. The TCR may comprise an alpha chain variable region set forth in a sequence that is at least 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 3, 82 to 116, 142 and 143, and a beta chain variable region set forth in a sequence that is at least 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 10, 117 to 121 and 144, in which CDR3a and CDR3b are unchanged. The TCR may comprise an alpha chain variable region set forth in a sequence that is at least 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 3, 82 to 116, 142 and 143 and a beta chain variable region set forth in a sequence that is at least 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 10, 117 to 121 and 144, wherein the alpha chain variable region and the beta chain variable region have the same TCR ID in Table 3 and Table 4, and CDR3a and CDR3b are unchanged.The TCR may comprise an alpha chain variable region set forth in a sequence that is at least 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 3, 82 to 116, 142, and 143, and a beta chain variable region set forth in a sequence that is at least 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 10, 117 to 121, and 144, in which CDR3a, CDR1b, CDR2b, and / or CDR3b are unchanged. The TCR may comprise an alpha chain variable region set forth in a sequence that is at least 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 3, 82 to 116, 142 and 143 and a beta chain variable region set forth in a sequence that is at least 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 10, 117 to 121 and 144, wherein the alpha chain variable region and beta chain variable region have the same TCR ID in Table 3 and Table 4, and wherein CDR3a, CDR1b, CDR2b and / or CDR3b are unchanged. The TCR may comprise an alpha chain variable region set forth in a sequence that is at least 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 3, 82 to 116, 142, and 143, and a beta chain variable region set forth in a sequence that is at least 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 10, 117 to 121, and 144, in which CDR1a, CDR2a, CDR3a, and / or CDR3b are unchanged. The TCR may comprise an alpha chain variable region set forth in a sequence that is at least 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 3, 82 to 116, 142 and 143 and a beta chain variable region set forth in a sequence that is at least 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 10, 117 to 121 and 144, wherein the alpha chain variable region and the beta chain variable region have the same TCR ID in Table 3 and Table 4, and CDR1a, CDR2a, CDR3a and / or CDR3b are unchanged.The TCR may comprise an alpha chain variable region set forth in a sequence that is at least 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 3, 82 to 116, 142 and 143, and a beta chain variable region set forth in a sequence that is at least 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 10, 117 to 121 and 144, wherein CDR1a, CDR2a, CDR3a, CDR1b, CDR2b and / or CDR3b are unchanged. The TCR may comprise an alpha chain variable region set forth in a sequence at least 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 3, 82-116, 142, and 143, and a beta chain variable region set forth in a sequence at least 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 10, 117-121, and 144, wherein the alpha chain variable region and beta chain variable region have the same TCR ID in Table 3 and Table 4, and CDR1a, CDR2a, CDR3a, CDR1b, CDR2b, and / or CDR3b are unchanged. Antigen-binding portions of the TCR are also provided. The TCR does not comprise both SEQ ID NO: 3 and SEQ ID NO: 10. In any of the descriptions in this paragraph, the combination may be one of SEQ ID NOs: 3 and 82-116 with one of SEQ ID NOs: 10 and 117-121.

[0102] The TCR, or antigen portion thereof, is capable of specifically binding to a peptide of the sequence KVLEHVVRV (SEQ ID NO: 122) presented by MHC class I molecules. MHC class I molecules are HLA-A, particularly HLA-A * 0201. Thus, the TCR, or antigen-binding portion, may be HLA-A * It is capable of specifically binding to the peptide of sequence KVLEHVVRV (SEQ ID NO: 122) presented by 0201.

[0103] It is known in the art that TCRs specific for peptides presented by a particular HLA subtype can also recognize target peptides presented in other HLA subtypes, e.g., related or similar subtypes.

[0104] Thus, the TCRs, or antigen-binding portions thereof, disclosed herein are capable of binding to HLA-A * 0201. This is also applicable to all TCR and antigen-binding moieties described herein.

[0105] In one embodiment, HLA-A * 0201, or an antigen-binding portion thereof, capable of specifically binding to the peptide set forth in SEQ ID NO: 122, presented by (i) a TCRa variable region, or a portion thereof, comprising CDR3a and a TCRb variable region, or a portion thereof, comprising CDR3b; or (ii) a TCRb variable region, or a portion thereof, including CDR1b, CDR2b, and CDR3b, and a TCRb variable region, or a portion thereof, including CDR3a, CDR1b, CDR2b, and CDR3b; or (iii) a TCRa variable region, or a portion thereof, including CDR1a, CDR2a, and CDR3a, and a TCRb variable region, or a portion thereof, including CDR3b; or (iv) a TCRa variable region, or a portion thereof, including CDR1a, CDR2a, and CDR3a, and a TCRb variable region, or a portion thereof, including CDR1b, CDR2b, and CDR3b. Including, CDR sequences are clones KVL-01-018, KVL-01-019, KVL-01-020, KVL-01-022, KVL-01-026, KVL-01-034, K VL-01-036, KVL-01-039, KVL-01-040, KVL-01-041, KVL-01-042, KVL-01-043, KVL-01-048, KVL -01-049, KVL-01-050, KVL-01-052, KVL-01-056, KVL-01-060, KVL-01-061, KVL-01-068, KVL-0 1-069, KVL-01-070, KVL-01-071, KVL-01-073, KVL-01-074, KVL-01-075, KVL-01-076, KVL-01- 077, KVL-01-078, KVL-01-079, KVL-01-083, KVL-01-084, KVL-01-088, KVL-01-089, KVL-01-0 90, KVL-01-094, KVL-01-097, KVL-01-105, KVL-01-108, KVL-01-110, KVL-01-111, KVL-01-112 , KVL-01-117, KVL-01-119, KVL-01-123, KVL-01-124, KVL-01-125, KVL-01-126, KVL-01-128, KVL-01-129, KVL-01-145, KVL-01-149, KVL-01-151, or KVL-01-153; Optionally, the TCRa variable region is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical, or 95%, 97%, 98%, 99%, or 100% identical, to the TCRa variable region of the clone from which the set of CDR sequences is derived; Optionally, a TCR, or antigen-binding portion thereof, is provided in which the TCRb variable region is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical, or 95%, 97%, 98%, 99%, or 100% identical, to the TCRb variable region of the clone from which the set of CDR sequences is derived. Optionally, the mutations listed in Table 6 may be retained.

[0106] The TCRs disclosed herein can specifically bind to target peptides presented on MHC class I molecules. As used herein, "specifically binds" means that the TCR can bind to a peptide-MHCI complex, where the affinity of binding is influenced by the sequence of the peptide. The TCR does not bind as strongly to peptide-MHCI, where the peptide is a scrambled peptide presented by the same MHC class I. The TCR can specifically bind to more than one peptide, for example, if minor or conservative changes are made to the target peptide.

[0107] In embodiments, the TCR, or antigen-binding portion thereof, has low cross-reactivity to a panel of peptides similar to KVLEHVVRV (SEQ ID NO: 122). In particular, the TCR, or antigen-binding portion thereof, may have low cross-reactivity to naturally occurring peptides within the human peptidome.

[0108] In some embodiments, the TCR, or antigen-binding portion thereof, has a K of less than or equal to 200 μM, 175 μM, 143 μM, 100 μM, 75 μM, 50 μM, 25 μM, 20 μM, 15 μM, 10 μM, 5 μM, or 2 μM. d K d can be calculated by surface plasmon resonance (SPR), where the target can be a peptide-HLA monomer. The peptide-HLA monomer is immobilized on a sensor chip (e.g., by using biotin and streptavidin). The equilibrium binding constant can be determined by flowing serial dilutions of a test TCR at 10 μl / min for 60 seconds over a flow cell loaded with 50-600 response units of peptide-HLA complexes. The specific response can be determined by subtracting the nonspecific response of a control flow cell (e.g., immobilized with streptavidin only) from the response of the flow cell loaded with the relevant peptide-HLA. K can be determined by nonlinear curve fitting and a binding isotherm equivalent to the Langmuir adsorption isotherm for gas absorption onto a surface. dFurther details are provided in the Examples section herein. Using the methods described in the Examples, K values ​​of the embodiments herein can be determined. d can be determined.

[0109] In some embodiments, the TCR, or antigen-binding portion thereof, has an EC of less than or equal to 500,000 μM, 10,000 μM, 5000 μM, 2500 μM, 2000 μM, 1500 μM, 1161 μM, 1000 μM, 500 μM, 400 μM, 300 μM, 200 μM, 150 μM, 100 μM, 75 μM, 50 μM, 40 μM, 30 μM, 25 μM, 20 μM, 15 μM, or 10 μM. 50 The EC was determined by transduction of the TCR or portion into PBMCs and subsequent measurement of reactivity in an IFNγ ELISPOT assay against antigen-presenting cells (APCs) pulsed with different concentrations of a peptide having the sequence KVLEHVVRV (SEQ ID NO: 122). 50 can be calculated. For more information, see Example 6.

[0110] In some embodiments, T cells or NK cells expressing a TCR, or an antigen-binding portion thereof, respond to a cell line expressing a peptide having the sequence KVLEHVVRV (SEQ ID NO: 122) in the correct MHC. For example, cell lines NCI-H1703 or C33A. In some cases, the T cells or NK cells are more responsive to C33A cells than reference T cells or NK cells expressing the TCR described in Example 1 as the parent / wild-type TCR. In some embodiments, the cells are 1.5-fold, 2-fold, 3-fold, 4-fold, 5-fold, 10-fold, 11-fold, or 12-fold more responsive to C33A cells compared to reference cells expressing the parent TCR. In some embodiments, the cells are 1.1-fold, 1.2-fold, 1.3-fold, 1.5-fold, 1.75-fold, 2-fold, 2.2-fold, or 2.3-fold more responsive to NCI-H1703 cells compared to reference cells expressing the parent TCR.

[0111] As used herein, the term "T cell receptor" or "TCR" refers to a class of molecules that are members of the immunoglobulin superfamily. A functional TCR can bind to a peptide-MHC complex and is specific for a peptide or a limited range of peptides presented by a particular MHC complex. An intact TCR comprises a TCR variable region, a TCR constant region, a TCR transmembrane region, and a TCR cytoplasmic tail. Molecules of the present disclosure comprise a TCR variable region, or an antigen-binding portion thereof. Molecules of the present disclosure may also comprise a TCR constant region or a portion thereof, optionally a transmembrane region, and optionally a cytoplasmic tail. Thus, molecules of the present disclosure may be an intact TCR or a portion of an intact TCR.

[0112] The TCR, or its antigen-binding portion, may exist as a single polypeptide chain. For example, a single chain comprising a TCR-a variable region and a TCR-b variable region. The TCR-a chain and the TCR-b chain can be connected via a peptide linker. The TCR may be a single-chain TCR (e.g., as disclosed in Li et al., TCR extracellular domain genetically linked to CD28, 2B4 / 41BB and DAP10 / CD3ζ-engineered NK cells mediates antitumor effects, Cancer Immunology, Immunotherapy https: / / doi.org / 10.1007 / s00262-022-03275-5). Further examples of TCR formats are provided in Oh et al. (Nature Scientific Reports | (2019) 9:17291 | https: / / doi.org / 10.1038 / s41598-019-53756-4).

[0113] Alternatively, a TCR, or antigen-binding portion thereof, may comprise two or more polypeptide chains, e.g., one comprising a TCR-a variable region and one comprising a TCR-b variable region. The TCR-a and TCR-b chains may be linked by one or more disulfide bonds, non-covalent bonds (such as van der Waals forces and / or hydrogen bonds), and / or one or more covalent bonds (such as peptide bonds).

[0114] The TCR, or antigen-binding portion thereof, may be part of one or more longer polypeptides. For example, the TCR, or antigen-binding portion thereof, may be part of one or more fusion proteins that include other domains or proteins. Examples include the CD3 molecule or a portion thereof. In another example, the TCR, or antigen-binding portion thereof, may be part of a chimeric antigen receptor (CAR), thus conferring antigen specificity to the receptor. The TCR, or antigen-binding portion thereof, may be added to or replace the single-chain variable fragment (scFv) of the CAR.

[0115] The TCR, or antigen-binding portion thereof, may be part of a bispecific or bifunctional molecule. For example, the TCR, or antigen-binding portion thereof, may be part of a complex or fusion protein with at least one other antigen-binding protein, such as a TCR, or an antibody portion, such as an scFv. The second antigen-binding portion may have specificity for immune cells, for example, it may be specific for a T cell marker and therefore be able to recruit T cells to MAGE-A4-expressing cells.

[0116] The TCR, or antigen-binding portion thereof, may be part of a soluble molecule or composition. The TCR may be a soluble TCR.

[0117] The TCR may comprise a TCRa variable region and a TCRb variable region. The TCR may comprise an alpha chain constant region and / or a beta chain constant region. In some embodiments, the TCR comprises a TCRa variable region, a TCRb variable region, and an alpha chain constant region. In some embodiments, the TCR comprises a TCRa variable region, a TCRb variable region, and a beta chain constant region. In some embodiments, the TCR comprises a TCRa variable region, a TCRb variable region, an alpha chain constant region, and a beta chain constant region. Thus, the TCR may comprise the four domains, three domains, or two domains of a complete TCR. The TCR may be a single-chain TCR comprising four, three, or two domains. In some examples, the alpha and / or beta constant regions are human or derived from human sequences.

[0118] A TCR may be chimeric in that the variable and constant region or regions are not naturally associated with each other. For example, the variable region may comprise sequences derived from one species, and the constant region or regions may comprise sequences derived from a different species. In some instances, the constant domain may be murine or may be derived from a murine constant domain. The murine constant domain or domains may be connected to a human variable region. The alpha and / or beta constant regions may be derived from human sequences in which only some residues have been exchanged for the corresponding residues in the murine sequence (referred to as "minimal murinization").

[0119] A TCR may contain intervening sequences between domains not found in naturally occurring TCRs, such as linkers between domains, targeting sites for antibodies, hinge sequences, transmembrane sequences, etc. For example, a TCR may contain an epitope for an antibody to allow identification or elimination of cells expressing the TCR. Such features can be located anywhere within the TCR as long as they do not affect function. The TCR may be an engineered TCR.

[0120] The TCR, or antigen-binding portion thereof, may contain stabilizing mutations. For example, the transmembrane regions of the alpha and beta chains may contain mutations that favor correct TCR chain pairing. Examples of suitable mutations are disclosed in WO2016071343 (incorporated herein by reference). The TCR, or antigen-binding portion thereof, may contain mutations that enhance surface expression and functional avidity. Examples of suitable mutations are disclosed in Haga-Friedman et al., J Immunol 2012; 188:5538-5546 (incorporated herein by reference).

[0121] An "antigen-binding portion" of a TCR refers to one or more fragments of an intact TCR that retain the ability to bind to the same peptide-MHC complex. A particular peptide-MHC complex can be referred to as the "antigen" for a particular TCR. In the context of the present invention, an antigen-binding portion retains the ability to bind to a peptide derived from MAGE-A4 when presented by the appropriate MHC class I molecule. For example, an antigen-binding portion of a TCR may bind to a peptide derived from HLA-A4. * When presented by .0201, it can bind to a peptide of sequence KVLEHVVRV (SEQ ID NO: 122). The antigen-binding portion retains CDR3 of each chain. The antigen-binding portion may also retain CDR1 and / or CDR2 of each chain. The antigen-binding portion of the TCR may retain CDR3a and / or CDR3b. The antigen-binding portion of the TCR may retain CDR3a, CDR1b, CDR2b, and CDR3b. The antigen-binding portion of the TCR may retain CDR1a, CDR2a, CDR3a, and CDR3b. The antigen-binding portion of the TCR may retain CDR1a, CDR2a, CDR3a, CDR1b, CDR2b, and CDR3b.

[0122] As disclosed herein, the TCR, or antigen-binding portion thereof, may comprise SEQ ID NO: 4. The TCR, or antigen-binding portion thereof, may comprise a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 4. The TCR, or antigen-binding portion thereof, may comprise a sequence that is at least 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 4.

[0123] In some embodiments, the TCR, or antigen-binding portion thereof, comprises SEQ ID NO: 11, 138, or 139. The TCR, or antigen-binding portion thereof, may comprise a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 11, 138, and 139. The TCR, or antigen-binding portion thereof, may comprise a sequence that is at least 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 11, 138, and 139. In a specific embodiment, the TCR, or antigen-binding portion thereof, comprises SEQ ID NO: 11.

[0124] EDLNKVFPPEVAVFEPSEAEISHTQKATLVCLATGFFPDHVELSWWVNGKEVHSGVSTDPQPLKEQPALNDSRYCLSSRLRVSATFWQNPRNHFRCQVQFYGLSENDEWTQDRAKPVTQIVSAEAWGRADCGFTSVSYQQGVLSATILYEILLGKATLYAVLVSALVLMAMVKRKDF (SEQ ID NO: 138—an example of a beta chain constant region sequence).

[0125] EDLKNVFPPEVAVFEPSEAEISHTQKATLVCLATGFYPDHVELSWWVNGKEVHSGVSTDPQPLKEQPALNDSRYCLSSRLRVSATFWQNPRNHFRCQVQFYGLSENDEWTQDRAKPVTQIVSAEAWGRADCGFTSESYQQGVLSATILYEILLGKATLYAVLVSALVLMAMVKRKDSRG (SEQ ID NO: 139—an example of a beta chain constant region sequence).

[0126] In some embodiments, the TCR, or antigen-binding portion thereof, comprises SEQ ID NO: 4 and any one of SEQ ID NOs: 11, 138, and 139. The TCR, or antigen-binding portion thereof, may comprise a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 4 and a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 11, 138, and 139. The TCR, or antigen-binding portion thereof, may comprise a sequence that is at least 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 4 and a sequence that is at least 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 11, 138, and 139. The TCR, or antigen-binding portion thereof, may comprise a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99% or 100% identical to SEQ ID NO:4 and a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99% or 100% identical to SEQ ID NO:11.

[0127] Constructs encoding the TCR, or antigen-binding portion thereof, may encode a signal peptide or peptides, however, in some instances the signal peptide or peptides may be cleaved or removed during expression of the TCR or antigen-binding portion thereof.

[0128] The TCRa chain may comprise a signal peptide set forth in the sequence SEQ ID NO: 2. The TCRb chain may comprise a signal peptide set forth in the sequence SEQ ID NO: 9.

[0129] The TCRa chain may comprise a TCRa variable region set forth in any one of SEQ ID NOs: 82 to 116, 142, and 143, a signal peptide set forth in the sequence of SEQ ID NO: 2, and a constant region set forth in SEQ ID NO: 4. The TCRb chain may comprise a TCRb variable region set forth in any one of SEQ ID NOs: 117 to 121 and 144, a signal peptide set forth in the sequence of SEQ ID NO: 9, and a constant region set forth in any one of SEQ ID NOs: 11, 138, and 139.

[0130] As used herein, the term "conservative substitution" refers to the replacement of one amino acid with another amino acid that does not significantly adversely change functional activity. Conservative substitutions may be those described by Dayhoff, "The Atlas of Protein Sequence and Structure. Vol. 5," Natl. Biomedical Research, etc. For example, a conservative substitution may be the exchange of a residue from one of the following groups for another residue within the same group: Group 1: alanine, proline, glycine, asparagine, serine, threonine; Group 2: cysteine, serine, tyrosine, threonine; Group 3: valine, isoleucine, leucine, methionine, alanine, phenylalanine; Group 4: lysine, arginine, histidine; Group 5: phenylalanine, tyrosine, tryptophan, histidine; and Group 6: aspartic acid, glutamic acid.

[0131] In another example, conservative amino acid substitutions can be selected from T to A, A to I, T to V, A to M, T to I, A to V, T to G, and / or T to S.

[0132] In further examples, conservative amino acid substitutions may include substitutions of one amino acid with another amino acid of the same class, for example, (1) non-polar: Ala, Val, Leu, Ile, Pro, Met, Phe, Trp; (2) uncharged polar: Gly, Ser, Thr, Cys, Tyr, Asn, Gln; (3) acidic: Asp, Glu; and (4) basic: Lys, Arg, His. Other conservative amino acid substitutions can also be made, such as: (1) aromatic: Phe, Tyr, His; (2) proton donor: Asn, Gln, Lys, Arg, His, Trp; and (3) proton acceptor: Glu, Asp, Thr, Ser, Tyr, Asn, Gln.

[0133] In another example, conservative substitutions can be made according to the following: Ala to Ser, Gly, or Cys Arg to Lys, Gln, or His Asn to Gln, His, Glu, or Asp Asp to Glu, Asn, or Gln Cys to Ser, Met, or Thr Gln to Asn, Lys, Glu, Asp, or Arg Glu to Asp, Asn, or Gln Gly to Pro, Ala, or Ser His to Asn, Gln, or Lys Ile to Leu, Val, Met, or Ala Leu to Ile, Val, Met, or Ala Lys to Arg, Gln, or His Met to Leu, Ile, Val, Ala, or Phe Phe to Met, Leu, Tyr, Trp, or His Ser to Thr, Cys, or Ala Thr to Ser, Val, or Ala Trp to Tyr or Phe Tyr to Trp, Phe, or His Val to Ile, Leu, Met, Ala, or Thr.

[0134] A "non-conservative" substitution is one that is not conservative as defined herein. A "naturally occurring" amino acid is one that forms part of proteins or peptides in organisms where it occurs in nature. A "standard" amino acid is one of the 20 amino acids that form proteins in humans.

[0135] The TCR, or antigen-binding portion thereof, may contain glycosylation sites and may be glycosylated.

[0136] The TCRs of the present disclosure, or antigen-binding portions thereof, may be isolated. The term "isolated" refers to a molecule that, depending on its origin or source, (1) is not associated with naturally associated components that accompany it in its natural state; (2) is substantially free from other molecules from the same species; (3) is expressed by cells from a different species; or (4) is not naturally occurring. Thus, a molecule that is chemically synthesized or expressed in a cell line different from the cell from which it naturally originated would be "isolated" from its naturally associated components. Molecules can also be rendered substantially free of naturally associated components by isolation using purification techniques well known in the art. The purity or homogeneity of a molecule can be assayed by several means well known in the art. For example, the purity of a polypeptide sample can be assayed using polyacrylamide gel electrophoresis and staining of the gel to visualize the polypeptide using techniques well known in the art. For certain purposes, higher resolution can be achieved by HPLC or other means well known in the art for purification.

[0137] The present invention encompasses modifications to the amino acid sequence of a TCR molecule or antigen-binding portion thereof as defined herein. For example, the present invention includes TCR molecules and their corresponding antigen-binding portions comprising functionally equivalent variable regions and CDRs that do not significantly affect these properties, as well as variants with enhanced or decreased activity and / or affinity. For example, the amino acid sequence can be mutated to obtain a TCR with the desired binding affinity for the peptide set forth in SEQ ID NO: 122 presented by the relevant MHC molecule. Amino acid sequence insertions are contemplated, including amino- and / or carboxyl-terminal fusions ranging in length from a single residue to polypeptides containing 100 or more residues, as well as intrasequence insertions of single or multiple amino acid residues. Examples of terminal insertions include TCR molecules with an N-terminal methionyl residue or TCR molecules fused to an epitope tag. Other insertional variants of TCR molecules include fusion to the N- or C-terminus of the TCR of an enzyme or polypeptide that increases the half-life of the TCR in the circulation.

[0138] The TCR, or antigen-binding portion thereof, can be linked to a payload or agent. Thus, provided herein is an immunoconjugate comprising a TCR disclosed herein, or antigen-binding portion thereof, linked to an agent.

[0139] The TCR can be linked to a therapeutic agent. For example, the TCR, or an antigen-binding portion thereof, can be linked to a substance that induces a change in cancer cells, such as a cytotoxic agent or a radioactive agent. The TCR, or an antigen-binding portion thereof, can be linked to a detection agent or label, such as a radioactive agent or a fluorescent agent.

[0140] Examples of suitable drugs include cytotoxins, radioisotopes, chemotherapeutic agents, immunomodulators, antiangiogenic agents, antiproliferative agents, proapoptotic agents, and cytostatic and cytolytic enzymes (e.g., RNAses). Additional drugs include nucleic acids, such as genes, that encode immunomodulators, antiangiogenic agents, antiproliferative agents, or proapoptotic agents. These drug descriptors are not mutually exclusive, and thus, one or more of the above terms can be used to describe a drug. Representative cytotoxins include antibiotics, inhibitors of tubulin polymerization, alkylating agents that bind to and destroy DNA, and drugs that disrupt protein synthesis or function of essential cellular proteins, such as protein kinases, phosphatases, topoisomerases, enzymes, and cyclins.

[0141] The linkage may be any suitable linkage for linking an agent or payload to a TCR or antigen-binding moiety. For example, the linkage may include a covalent or non-covalent bond. The linkage may be via a bifunctional linker. The linkage may be via two moieties capable of forming a non-covalent interaction, such as streptavidin and biotin.

[0142] In another aspect, there is provided at least one nucleic acid encoding a TCR disclosed herein, or an antigen-binding portion thereof.

[0143] A single nucleic acid may encode both the TCRa chain and the TCRb chain. For example, a TCRa chain comprising a TCRa variable region, or an antigen-binding portion thereof, and a TCRb chain comprising a TCRb variable region, or an antigen-binding portion thereof. The TCRa chain and the TCRb chain may be encoded as a single polypeptide, and thus the nucleic acid may encode a single-chain TCR. Alternatively, a single nucleic acid may encode a TCRa chain and another TCRb chain, for example, under the control of separate promoters. In another example, the TCRa chain and the TCRb chain may be encoded by the same nucleic acid and under the control of the same promoter, but separated by a region capable of inducing ribosome skipping or a self-cleaving peptide sequence. Examples of such features include P2A, an internal ribosome entry site (IRES), or an intein.

[0144] Alternatively, the first nucleic acid strand may encode the TCRa chain and the second nucleic acid may encode the TCRb chain.

[0145] Thus, a nucleic acid molecule of the present disclosure may encode (a) a TCRa variable region amino acid sequence; (b) a TCRb variable region amino acid sequence; or (c) the amino acid sequences of both the TCRa variable region and the TCRb variable region of a TCR or antigen-binding portion disclosed herein. A nucleic acid molecule as defined herein may be isolated.

[0146] In another aspect, there is provided a vector comprising a nucleic acid disclosed herein. The vector may be an expression vector or may be a vector designed for delivery to a subject, such as a human patient.

[0147] In another aspect, there is provided a cell comprising at least one nucleic acid disclosed herein, or a vector disclosed herein.

[0148] The cell may be a recombinant cell. The cell may be a cell suitable for commercial production of recombinant polypeptides and proteins. In some cases, the cell is of mammalian, bacterial, yeast, or plant origin. For example, the cell may be a CHO cell, an NS0 cell, an HEK cell, an E. coli cell, a Pichia pastoris cell, or other suitable cell.

[0149] In another aspect, there is provided a method of producing a TCR or antigen-binding portion thereof disclosed herein, the method comprising culturing a cell disclosed herein under conditions that result in expression and / or production of the TCR or antigen-binding portion thereof, and isolating the TCR or antigen-binding portion thereof from the cell or culture.

[0150] In another aspect, there is provided a cell comprising a TCR disclosed herein, or an antigen-binding portion thereof, at least one nucleic acid disclosed herein, or a vector disclosed herein.

[0151] The cell may be a mammalian or human cell. The cell may be an immune cell. The cell may be an immune cell suitable for use in immunotherapy. The immune cell may be a T cell, such as an alpha-beta T cell or a gamma-delta T cell. The alpha-beta T cell may be a CD4+ T cell or a CD8+ T cell. The alpha-beta T cell may be a naive cell, an effector cell, or a memory cell. The cell may be an invariant NK T cell. The cell may be an NK cell.

[0152] The immune cells may be primary immune cells, such as immune cells derived from PBMCs or tumor samples. The immune cells may be cell lines, such as T cell lines or NK cell lines. The immune cells may be differentiated in vitro, for example, from stem cells. The stem cells may be induced pluripotent stem cells (iPSCs), hematopoietic stem cells, or mesenchymal stem cells (MSCs). In certain embodiments, the cells are human NK cells differentiated from iPSCs.

[0153] The cells may be in vitro and may be T cells or NK cells. The cells may be an in vitro population of T cells or an in vitro population of NK cells suitable for adoptive transfer therapy. The cells may be an in vitro population of human NK cells.

[0154] The TCRs disclosed herein, or antigen-binding portions thereof, can be provided in lyophilized form for reconstitution prior to administration. For example, the lyophilized protein can be reconstituted in sterile water and mixed with saline before administration to an individual. The nucleic acids disclosed herein or the vectors disclosed herein can be provided in lyophilized form. The cells disclosed herein can be provided in a stable form for storage, e.g., the cells can be frozen in the presence of a cryoprotectant. In other embodiments, all of these products can be provided in a form suitable for administration.

[0155] Thus, in another aspect, there is provided a pharmaceutical composition comprising a TCR or antigen-binding portion thereof disclosed herein, at least one nucleic acid disclosed herein, a vector disclosed herein, or a cell disclosed herein.

[0156] Pharmaceutical compositions may contain pharmaceutically acceptable excipients, carriers, stabilizers or diluents. Pharmaceutically acceptable excipients may be compounds or combinations of compounds that do not induce secondary reactions and that allow, for example, easier administration, increased longevity and / or effectiveness in the body, or increased solubility in solution, to be incorporated into the pharmaceutical composition. These pharmaceutically acceptable vehicles are well known and are adapted by those skilled in the art according to the mode of administration.

[0157] The TCRs and antigen-binding moieties disclosed herein can be used to recognize MAGE-A4-expressing cells. Some types of cancer express MAGE-A4, and therefore, the TCRs and moieties can be used to recognize cancerous cells. In a non-limiting example, the TCRs and / or moieties can be expressed by immune cells, thereby conferring the immune cells the ability to recognize cancerous cells. In another example, the TCRs or moieties can be molecules that are independent of the cell, thus directing a payload (such as a cytotoxic agent) to tumor cells. In yet another example, the TCRs can be part of a bifunctional molecule, allowing tumor cell recognition by a molecule that also enables another function, such as T cell recruitment. The TCRs and moieties need not be administered directly but can be expressed in the subject. Thus, the nucleic acids disclosed herein and the vectors disclosed herein can also be administered to patients.

[0158] Thus, in another aspect, there is provided a TCR or antigen-binding portion thereof disclosed herein, at least one nucleic acid disclosed herein, a vector disclosed herein, a cell disclosed herein, or a pharmaceutical composition disclosed herein for use as a medicament.

[0159] In certain embodiments, there is provided a human immune cell, such as a T cell or an NK cell, comprising a TCR, or an antigen-binding portion thereof, for use as a medicament.

[0160] Provided are methods for enhancing an immune response in a subject, comprising administering to a subject in need thereof an effective amount of a TCR or antigen-binding portion disclosed herein, at least one nucleic acid disclosed herein, a vector disclosed herein, a cell disclosed herein, or a pharmaceutical composition disclosed herein. The administered or expressed TCR can enhance the immune response of cells expressing it, and / or the cells expressing the TCR can enhance the response of cells in the same microenvironment, for example, by release of cytokines.

[0161] Provided is the use of a TCR or antigen-binding portion thereof disclosed herein, at least one nucleic acid disclosed herein, a vector disclosed herein, a cell disclosed herein, or a pharmaceutical composition disclosed herein in the manufacture of a medicament.

[0162] The medicaments disclosed herein can be used in methods of treatment of the human or animal body, including prophylactic or preventative treatment (e.g., treatment prior to the onset of a condition in an individual, to reduce the risk of the condition occurring in the individual; delay its onset; or reduce its severity after onset).

[0163] The terms "prophylactic," "preventative," or "preventing," etc., refer to reducing the probability of developing a disorder or condition in a subject who does not have the disorder or condition but is at risk of developing or susceptible to it. Prevention, etc., does not mean that a subject does not acquire a particular disease or disorder in the first place. Prevention may require the administration of multiple doses. Prevention may include preventing the recurrence of disease in a subject in which all disease symptoms have been eliminated, or preventing recurrence in relapsing-remitting disease.

[0164] The method of treatment may comprise administering a medicament to an individual in need thereof.

[0165] The medicament may be for use in a mammalian subject. The medicament may be for use in a mouse, rat, rabbit, dog, cat, horse, or pig. Preferably, the subject is a human. The subject may be HLA-A * 0201 may have been determined to be positive.

[0166] It is known in the art that TCRs specific for peptides presented by a particular HLA subtype can also recognize target peptides presented in other HLA subtypes, e.g., related or similar subtypes. Thus, a subject can be determined to be positive for an HLA subtype to which a TCR of the present disclosure, or an antigen-binding portion thereof, can bind.

[0167] Administration is typically in a "therapeutically effective amount," which is sufficient to demonstrate benefit to the patient. Such benefit may be at least an improvement in at least one symptom. The actual amount administered, as well as the rate and duration of administration, will depend on the nature and severity of what is being treated, the particular mammal being treated, the clinical condition of the individual patient, the cause of the disorder, the site of delivery of the composition, the method of administration, the schedule of administration, and other factors known to physicians. Treatment prescriptions, such as decisions regarding dosage, are within the responsibility of medical professionals and may depend on the severity of the symptoms and / or the progression of the disease being treated. A therapeutically effective amount or appropriate dose of a drug can be determined by comparing in vitro activity and in vivo activity in animal models. Methods for extrapolating effective dosages in mice and other test animals to humans are known. The precise dose will depend on several factors, including whether the drug is for prevention or treatment, the size and location of the area to be treated, the precise nature of the drug, and the nature of any detectable label or other molecule attached to the antibody.

[0168] Administration may be by any suitable method, for example, by injection, intravenous administration, etc. Administration may be in a single dose or may be repeated periodically.

[0169] The medicaments of the present disclosure can be administered as monotherapy or in combination with other agents. Administration of the combination therapy can be simultaneous or sequential.

[0170] In another aspect, there is provided a TCR or antigen-binding portion disclosed herein, at least one nucleic acid disclosed herein, a vector disclosed herein, a cell disclosed herein, or a pharmaceutical composition disclosed herein for use in a method for treating, preventing, or ameliorating cancer.

[0171] For example, provided are methods for treating, preventing, or ameliorating cancer in a subject, the methods comprising administering to a subject in need thereof an effective amount of a TCR or antigen-binding portion disclosed herein, at least one nucleic acid disclosed herein, a vector disclosed herein, a cell disclosed herein, or a pharmaceutical composition disclosed herein.

[0172] Provided is the use of a TCR or antigen-binding portion disclosed herein, at least one nucleic acid disclosed herein, a vector disclosed herein, a cell disclosed herein, or a pharmaceutical composition disclosed herein in the manufacture of a medicament for the treatment of cancer.

[0173] In certain embodiments, provided are human immune cells, such as T cells or NK cells, comprising a TCR, or antigen-binding portion thereof, disclosed herein, for use in a method for treating, preventing, or ameliorating cancer.

[0174] The cancer may be a solid cancer. The cancer may comprise cells that express MAGE-A4. Some cancerous cells may present the peptide set forth in SEQ ID NO: 122. The subject may have precancerous cells that express MAGE-A4.

[0175] In one embodiment, the cancer is MAGE-A4 positive and is selected from the group consisting of lung cancer, head and neck cancer, liver cancer, skin cancer, renal cell carcinoma, brain tumors, gastric cancer, colorectal cancer, hepatocellular carcinoma, pancreatic cancer, prostate cancer, leukemia, breast cancer, Merkel cell carcinoma, bladder cancer, uterine cancer, gallbladder and bile duct cancer, osteosarcoma, liposarcoma, neuroblastoma, myeloma, melanoma, metastatic melanoma, synovial sarcoma, esophageal cancer, esophageal squamous cell carcinoma, ovarian cancer, ovarian epithelial cancer, pre- prostate cancer, breast cancer, malignant astrocytoma, glioblastoma multiforme, anaplastic astrocytoma, fallopian tube cancer, primary peritoneal cancer, advanced solid tumor, soft tissue sarcoma, sarcoma, myelodysplastic syndrome, acute myeloid leukemia, Hodgkin lymphoma, non-Hodgkin lymphoma, Hodgkin's disease, multiple myeloma, metastatic solid tumor, stomach cancer, gastric cancer, rhabdomyosarcoma, myxoid round cell liposarcoma, or recurrent non-small cell lung cancer.

[0176] In examples, the cancer is MAGE-A4 positive and is lung cancer, liver cancer, head and neck cancer, skin cancer, gastric cancer, colorectal cancer, hepatocellular carcinoma, pancreatic cancer, breast cancer, melanoma, ovarian cancer, bladder cancer, uterine cancer, gallbladder and bile duct cancer, and esophageal cancer.

[0177] In some examples, provided herein are methods of treating, preventing, or ameliorating lung cancer, esophageal cancer, head and neck cancer, ovarian cancer, and melanoma.

[0178] The lung cancer may be non-small cell lung cancer (NSCLC), such as non-small cell lung adenocarcinoma or squamous cell non-small cell lung cancer (SNSCLC), or small cell lung cancer (SCLC). The skin cancer may be melanoma. The head and neck cancer may be head and neck squamous cell carcinoma (HNSCC). The liver cancer may be hepatocellular carcinoma (HCC). The esophageal cancer may be gastroesophageal junction cancer.

[0179] In examples, the cancer is MAGE-A4 positive and is ovarian cancer, melanoma, non-small cell lung cancer, hepatocellular carcinoma, colorectal cancer, esophageal squamous cell carcinoma, esophageal adenocarcinoma, stomach cancer, bladder cancer, head and neck cancer, gastric cancer, synovial sarcoma, or myxoid round cell liposarcoma.

[0180] In preferred embodiments, the cancer is NSCLC, ovarian cancer, gastroesophageal cancer, head and neck synovial sarcoma, or urothelial cancer.

[0181] Sequence comparisons can be performed with the aid of readily available sequence comparison programs. These publicly and commercially available computer programs can calculate the sequence identity between two or more sequences.

[0182] Those skilled in the art will understand how to calculate the percentage identity between two nucleic acid or amino acid sequences. To calculate the percentage identity, an alignment of the two sequences must first be prepared, and then the sequence identity value must be calculated. The percentage identity between two sequences may vary depending on (i) the method used to align the sequences, such as the Needleman-Wunsch algorithm (e.g., as applied by Needle (EMBOSS) or Stretcher (EMBOSS)), the Smith-Waterman algorithm (e.g., as applied by Water (EMBOSS)), or the LALIGN application (e.g., as applied by Matcher (EMBOSS)); and (ii) the parameters used by the alignment method, such as partial versus global alignment, the matrix used, and the parameters applied for gaps. In certain embodiments, the sequence identity disclosed herein can be calculated based on a global alignment of relevant features. For example, a comparison of the complete length of a TCR variable region within a polypeptide with the complete reference sequence for a TCR variable region described herein. In some embodiments, antigen-binding portions of the described TCR sequences are disclosed, and in certain instances, comparison may be over the full length of the antigen-binding portion to respective fragments of the same length of the sequences described herein.

[0183] Once aligned, there are many different ways to calculate the identity percentage between two sequences.For example, the number of identities can be divided by (i) the length of the shortest sequence; (ii) the length of alignment; (iii) the average length of sequences; (iv) the number of non-gap positions; or (iv) the number of equivalent positions excluding overhangs.It is also understood that identity percentage is strongly length-dependent.Therefore, the shorter the sequence pair, the higher the sequence identity that can be expected to occur by chance.

[0184] The percentage identity between two nucleic acid sequences is then calculated as (N / T) *From such an alignment, the number of positions can be calculated as 100 (where N is the number of positions where the sequences share identical residues and T is the total number of positions compared, including gaps but not overhangs).

[0185] The sequence alignment may be a pairwise sequence alignment. Suitable services include Needle (EMBOSS), Stretcher (EMBOSS), Water (EMBOSS), Matcher (EMBOSS), LALIGN, or GeneWise. In one example, the identity between two amino acid sequences can be calculated using the Needle (EMBOSS) service set to default parameters, such as matrix (BLOSUM62), gap open (10), gap extension (0.5), end gap penalty (false), end gap open (10), and end gap extension (0.5). In another example, the identity between two amino acid sequences can be calculated using the Matcher (EMBOSS) service set to default parameters, such as matrix (BLOSUM62), gap open (14), gap extension (4), and alternative match (1). In one example, identity between two nucleic acid sequences can be calculated using the Needle (EMBOSS) service set to default parameters, such as matrix (DNAfull), gap open (10), gap extension (0.5), end gap penalty (false), end gap open (10), and end gap extension (0.5). In another example, identity between two nucleic acid sequences can be calculated using the Matcher (EMBOSS) service set to default parameters, such as matrix (DNAfull), gap open (16), gap extension (4), and alternative match (1).

[0186] All of the features described in this specification (including the accompanying claims, abstract and drawings), and / or all of the steps of any method or process so disclosed, may be combined with any of the above embodiments in any combination, except combinations in which at least some of such features and / or steps are mutually exclusive.

[0187] For a better understanding of the present invention, and to show how to put into practice embodiments thereof, reference will now be made to examples which are not intended to limit the invention in any way. [Example]

[0188] Example 1 Parent TCR To generate a new MAGE-A4 TCR, a parental TCR was selected for analysis. The parental TCR was HLA-A * 0201 and is disclosed in WO2021 / 236548 A1 (incorporated herein by reference). The sequence of the parent TCR is set forth in the table below. This TCR is referred to herein as "wild-type" in the appropriate places.

[0189] [Table 5]

[0190] Example 2 Structural biology analysis of the parental TCR Structural biology analysis was performed on the parental TCR when bound to peptide-HLA.

[0191] Crystal diffraction data were collected. The data were scaled using STARANISO at a resolution of 2.39 Å. The structure was then solved by molecular replacement using a structural model from the Protein Data Bank (PDB). Molecular replacement searches for single-copy pHLA and then single-copy TCR were successful, allowing the binding mode and protein-protein interface to be clearly defined.

[0192] The crystal structure showed that the majority of interactions are made through the TCR beta chain, and in particular, most interactions are made by CDR3b.

[0193] In particular, the underlined residues in the sequences below represent residues in CDR3b that interact with the HLA molecule, and the bolded residues represent those that interact with the peptide.

[0194] [ka]

[0195] Of the contributions made by the alpha chain, CDR3a is responsible for most of the interaction.

[0196] In particular, the underlined residues in the sequences below represent residues that interact with the HLA molecule, and the bolded residues represent those that interact with the peptide.

[0197] [ka]

[0198] Example 3 Library generation - in silico modeling We used an alanine scanning screening method to map important amino acid residues in the TCR-pMHC binding interface.

[0199] A first TCR library containing mutations of the parental TCR (also known as the WT TCR) was then prepared using artificial intelligence. This involved the use of a machine learning platform trained to find patterns in the TCR repertoire and apply these patterns to specific PCR reactions. Part of the in silico modeling was expert-guided.

[0200] The first set of modeling was performed based on sequence-only and alanine-scan datasets. Four 3D structural models were generated from the TCR docked into the peptide-HLA complex. Variants were then produced by both expert-guided mutagenesis and in silico generation. The effects of mutations on the 3D models were then evaluated in silico.

[0201] Variants were tested against the modeled structure to search for predicted increases in affinity.

[0202] Changes in ΔG, total buried surface area, pHLA solvent accessible surface area (SASA) ratio, number of hydrogen bonds interacting with peptide-HLA, and number of van der Waals interactions with peptide-HLA were evaluated. Candidate TCRs were then selected for further testing.

[0203] Example 4 Binding analysis of TCR variants A subset of TCRs resulting from in silico modeling (Example 3) was entered into binding analyses, as described below.

[0204] Molecular cloning for protein expression The Novagen pET21(+) expression plasmid was used for bacterial expression using E. coli (BL21 DE3) as the host organism. Due to the nature of the pHLA and TCR domains, they are expressed as insoluble inclusion bodies (IBs). The total protein expressed may represent more than 50% of the total cellular protein. IB preparation, solubilization and refolding of TCR and pHLA are well-known and established methods (e.g., Willcoj et al., Protein Science (1999), 8:2418-2423, https: / / doi.org / 10.1110 / ps.8.11.2418; Boulter et al., Protein Engineering vol. 16 no. 9, pp. 707-711, 2003, https: / / doi.org / 10.1093 / protein / gzg087; or Garboczi et al., PNAS, Vol. 89, pp. 3429-3433, April 1992, https: / / doi.org / 10.1073 / pnas.89.8.3429), which often result in highly pure, homogeneous protein preparations (free of N-linked glycosylation).

[0205] TCR construct design The alpha and beta chains were cloned into the bacterial expression vector pET21(+). The alpha and beta chains were cloned into the vector using NdeI and NotI restriction sites, which utilize the ATG start codon in the NdeI site to initiate transcription, ensuring that no extra amino acids were expressed at the N-terminus of the protein chains. The TCR sequence was truncated to remove the transmembrane region and modified by the incorporation of a non-native alpha-beta disulfide bond between the constant domains (Boulter et al., 2003) and the substitution of a free cysteine ​​to produce a soluble and stabilized protein.

[0206] The TCR alpha constant domain (UNIPROT P01848) was modified at position 47; at position 85, a threonine was replaced with a cysteine, terminating the domain (similar to SEQ ID NO: 67). IQNPDPAVYQLRDSKSSDKSVCLFTDFDSQTNVSQSKDSDVYITDKCVLDMRSMDFKSNSAVAWSNKSDFACANAFNNSIIPEDT (SEQ ID NO: 67).

[0207] The TCR beta constant domain (UNIPROT A0A5B9) was modified at position 9; at position 56, a lysine was replaced with glutamic acid; at position 74, a serine was replaced with cysteine; at position 88, a cysteine ​​was replaced with alanine; and at position 129, an asparagine was replaced with aspartic acid, terminating the domain (similar to SEQ ID NO: 68). DLKNVFPPEVAVFEPSEAEISHTQKATLVCLATGFYPDHVELSWWVNGKEVHSGVCTDPQPLKEQPALNDSRYALSSRLRVSATFWQDPRNHFRCQVQFYGLSENDEWTQDRAKPVTQIVSAEAWGRAD (SEQ ID NO: 68).

[0208] Human leukocyte antigen (HLA) and beta 2 microglobulin (B2M) construct design The constructs used to produce the pHLA protein complex (biotinylated monomer) were generated in the pET21a vector using an NdeI restriction site start codon to initiate transcription. The HLA A201 construct (pET21|HLA_A2_25-300_Avi) encodes HLA A201 amino acids 25-300 (alpha regions 1, 2, and 3), terminating in the transmembrane domain and leading directly into the biotinylation sequence (AviTag: GLNDIFEAQKIEWHE (SEQ ID NO: 69)). The beta2 microglobulin construct (pET21|B2M_21-119) corresponds to amino acids 21-119 of the naturally secreted protein.

[0209] MGSHSMRYFFTSVSRPGRGEPRFIAVGYVDDTQFVRFDSDAASQRMEPRAPWIEQEGPEYWDGETRKVKAHSQTHRVDLGTLRGYYNQSEAGSHTVQRMYGCDVGSDWRFLRGYHQYAYDGKDYIALKEDLRSWTAADMAAQTTKH KWEAAHVAEQLRAYLEGTCVEWLRRYLENGKETLQRTDAPKTHMTHAVSDHEATLRCWALSFYPAEITLTWQRDGEDQTQDTELVETRPAGDGTFQKWAAVVVPSGQEQRYTCHVQHEGLPKPLTLRWEPGLNDIFEAQKIEWHE (pET21|HLA_A2_25-300_Avi - SEQ ID NO: 70).

[0210] MIQRTPKIQVYSRHPAENGKSNFLNCYVSGFHPSDIEVDLLKNGERIEKVEHSDLSFSKDWSFYLLYYTEFTPTEKDEYACRVNHVTLSQPKIVKWDRDM (pET21|B2M_21-119 - SEQ ID NO: 123).

[0211] MAGE-A4 286-294 and mimetic peptides Engineered TCRs (modified from WT as discussed above) and affinity-enhanced TCR variants were used in surface plasmon resonance (SPR) experiments to estimate binding affinities to the cognate MAGE-A4(286-294) peptide-HLA complexes. A set of 15 pHLA complexes with sequence-related mimetic peptides was also evaluated for potential cross-reactivity (below). KVLEHVVRV (MAGE-A4 286-294 (Melanoma-associated antigen 4, Uniprot P43358 MAGA4_HUMAN - SEQ ID NO: 122) FLLETVVRV (RABGAP1L_408-416 (RAB GTPase-activating protein 1-like, Uniprot: Q5R372 · RBG1L_HUMAN - SEQ ID NO: 124) ILDEHVQRV (AXIN1_470-478 (Axin-1, Uniprot: O15169 · AXIN1_HUMAN) - SEQ ID NO: 125) KILEDVVGV (TPX2_465-473 (targeting protein of Xklp2, Uniprot: Q9ULW0 · TPX2_HUMAN - SEQ ID NO: 126) KLLDLQVRV (SYNE3_563-571 (Nesprin-3, Uniprot: Q6ZMZ3 · SYNE3_HUMAN - SEQ ID NO: 127) KVLEILHRV (HERC4_595-603 (putative E3 ubiquitin protein ligase HERC4, Uniprot: Q5GLZ8 · HERC4_HUMAN - SEQ ID NO: 128) KVLERVNAV (PSME2_145-153 (proteasome activator complex subunit 2, Uniprot: Q9UL46 · PSME2_HUMAN - SEQ ID NO: 129) KVLETLVTV (HEATR5A_1925-1933 (HEAT repeat-containing protein 5A, Uniprot: Q86XA9 · HTR5A_HUMAN - SEQ ID NO: 130) KVLGIVVGV (CNOT1_1965-1973 (CCR4-NOT transcription complex subunit 1, Uniprot: A5YKK6 · CNOT1_HUMAN - SEQ ID NO: 131) KVLNKVITV (KIAA1033_85-93 (WASC4_HUMAN WASH complex subunit 4, Uniprot: Q2M389 · WASC4_HUMAN - SEQ ID NO: 132) KVTEMLVNV (PPP3CB_369-377 (Serine / Threonine Protein Phosphatase 2B Catalytic Subunit Beta Isoform, Uniprot: P16298 · PP2BB_HUMAN - SEQ ID NO: 133) KVLETTVEI (VPS35_371-379 (vacuolar protein sorting-associated protein 35, Uniprot: Q96QK1 · VPS35_HUMAN - SEQ ID NO: 134) KVLEMETTV (RNF40_271-279 (E3 ubiquitin protein ligase BRE1B, Uniprot: O75150 · BRE1B_HUMAN - SEQ ID NO: 135) KVMEHVYEK (WDR48_595-603 (WD repeat-containing protein 48, Uniprot: Q8TAF3 · WDR48_HUMAN - SEQ ID NO: 136) KVQEQVHKV (NSD1_1300-1308 (histone-lysine N-methyltransferase, H3 lysine-36 specific, Uniprot: Q96L73 · ​​NSD1_HUMAN - SEQ ID NO: 137)

[0212] Soluble biotinylated class I HLA-A * 02 Protein expression, refolding, and purification of protein complexes HLA and B2M expression, IB isolation, protein refolding in the presence of the peptide of interest, biotinylation and purification were performed using established methods known to those skilled in the art (Garboczi et al. (1992) or O'Callaghan et al., Analytical Biochemistry, Volume 266, Issue 1, 1 January 1999, pp. 9-1, https: / / doi.org / 10.1006 / abio.1998.2930).

[0213] Briefly, HLA and B2M proteins were expressed and IBs were isolated as described herein for TCR. Solubilized and reduced HLA, B2M, and peptide (at a molar ratio of 1:2:4) were refolded by rapid dilution in refolding buffer (0.4 M L-arginine-HCl, 100 mM Tris pH 8.0, 2 mM EDTA, 6.6 mM cysteamine hydrochloride, and 3.7 mM cysteamine dihydrochloride). The refolded material was dialyzed into 10 mM Tris-HCl pH 8.1, purified by anion exchange, biotinylated in vitro, and purified by size-exclusion chromatography.

[0214] Protein expression, refolding, and purification of soluble TCRs DNA sequences encoding the variable and constant regions of the TCR alpha and TCR beta chains were cloned into the pET21(+) expression plasmid between the NdeI and NotI restriction sites using standard methods. The TCR alpha and TCR beta plasmids were transformed separately into the E. coli strain Rosetta 2(DE3)pLysS. Individual colonies were used to inoculate LB (+ampicillin; 100 μg / ml) medium and grown to an OD of 0.5. 600 The culture was grown at 37°C until it reached a concentration of 0.01%. The culture was then used to inoculate autoinduction medium (AIM) - Super Broth (ForMedium) supplemented with 100 μg / ml ampicillin - for 5 hours at 37°C and an additional 16 hours at 30°C. Cells were harvested by centrifugation. The cell pellet was lysed using BugBuster® Protein Extraction Reagent (Merck Millipore) according to the manufacturer's instructions, and inclusion bodies were collected by centrifugation. The inclusion bodies were washed three times with Triton wash buffer (50 mM Tris pH 8.1, 100 mM NaCl, 1% Triton-X100, and 10 mM EDTA), once in resuspension buffer (50 mM Tris pH 8.1, 100 mM NaCl, and 10 mM EDTA), and finally resuspended in the same buffer. Inclusion body protein concentrations were determined by dissolving in solubilization buffer (6 M guanidine hydrochloride, 50 mM Tris pH 8.1, 100 mM NaCl, and 10 mM EDTA) and measuring OD 280 The inclusion body purity was assessed by dissolving approximately 2 μg of protein in 8 M urea and SDS-PAGE under reducing conditions. Purity was estimated by densitometry (GelDoc, Bio-Rad). The inclusion bodies were stored at +4°C or -80°C for short-term or long-term storage, respectively.

[0215] For TCR refolding, 45 mg of alpha and 30 mg of beta chain inclusion bodies were mixed and solubilized in 10 ml of denaturing buffer (6 M guanidine hydrochloride, 50 mM Tris pH 8.1, 100 mM NaCl, 10 mM EDTA, and 20 mM DTT) and incubated at 30° C. for 10 minutes. Refolding was achieved by rapid dilution into 1 L of vigorously stirred refolding buffer (100 mM Tris pH 8.1, 400 mM L-arginine-HCl, 4 M urea, 2 mM EDTA, 6.5 mM cysteamine hydrochloride, and 1.9 mM cystamine dihydrochloride) and incubated at 4° C. for 3 hours. The protein solution was dialyzed against 20 L of water at 4° C. for 16 hours. The protein solution was further dialyzed overnight into 20 L of 10 mM Tris pH 8.1, and this process was repeated. The protein solution was filtered through a 0.45 μm cellulose membrane.

[0216] The TCR was purified by anion exchange followed by size exclusion chromatography. The protein solution was passed over a 5 ml Q Sepharose (Cytiva) column and eluted with a continuous gradient of 0 to 500 mM NaCl in 10 mM Tris pH 8.1 over 25 column volumes using an FPLC instrument (Cytiva). Protein fractionation was assessed by SDS-PAGE, and TCR heterodimer fractions were pooled and concentrated. The concentrated TCR protein was further purified by size exclusion chromatography using a Superdex 200 Increase column in PBS (10 mM phosphate buffer, 2.7 mM KCl, and 137 mM NaCl). The concentration of the pooled peak TCR fractions was determined by OD 280 was estimated by measuring the value and calculated taking into account the attenuation coefficient.

[0217] Join analysis Affinity analysis for soluble TCR binding to pHLA complexes was performed by surface plasmon resonance (SPR) using a BIAcore T200 instrument. All measurements were performed at 25°C.

[0218] SPR method Biotinylated peptide-HLA monomers were immobilized on a sensor chip (Sensor Chip SA) preloaded with streptavidin. Equilibrium binding constants were determined by flowing serial dilutions of TCR at 10 μl / min for 60 seconds over flow cells loaded with 50–600 response units of peptide-HLA complexes. Specific responses were determined by subtracting the nonspecific response of a flow cell immobilized with streptavidin alone from the response of the flow cell loaded with the relevant peptide-HLA.

[0219] K was calculated by nonlinear curve fitting using Prism software and a binding isotherm equivalent to the Langmuir adsorption isotherm for gas adsorption onto surfaces. d value was determined. Y=Bmax * X / (K d +X) K d is the equilibrium dissociation constant, in units equal to X (molar concentration). It is the concentration required to achieve half-maximal binding at equilibrium. Bmax is the maximum specific binding, in units equal to Y (RU). Y is the equilibrium binding in RU at the injected TCR concentration X.

[0220] For the highest TCR affinity interactions, there is sufficient curvature in the injection and dissociation phases of the sensorgram to allow for the kinetic parameter k on and k off can be fitted, which was implemented in the BIAevaluation software. d k off / k on Dissociation half-life: t = 0.693 / k off was calculated.

[0221] result The experiments described above demonstrate that certain variant CDR3b

[0222] [ka]

[0223] was identified.

[0224] This variant was tested by surface plasmon resonance (SPR). The CDR3b WQ mutation was found to result in an 8.3-fold increase in Kd (to 19 μM) compared to the parental TCR. This is shown in Figure 1(D). Alternative mutations were tested as described in Example 6.

[0225] Another variant identified is

[0226] [ka]

[0227] The NQx motif is found in several CDR3a sequences disclosed herein (e.g., SEQ ID NOs: 35-44—experimentally tested as described in Example 6). This motif was predicted to change the potential energy for binding from −1765.34 kcal / mol to −2018.19 kcal / mol.

[0228] The M94N, D95Q, S96 and W99 mutations were tested in combination (see Example 6 - especially KVL-01-073 to KVL-01-77) and found to be functional.

[0229] Example 5 Library generation In silico prediction of favorable TCR framework and CDR mutations A set of TCR single-point mutations targeting the framework and CDRs of the TR and TRB chains was predicted based on custom-built TCR and pHLA homology models and supervised structural evaluation of an optimized in silico trimeric TCR-pHLA docking complex. Initial TCR and pHLA models were constructed using LYRA (Klausen, MS et al., Nucleic Acids Res 43, W349–355, doi:10.1093 / nar / gkv535 (2015)) and DockTope (Menegatti Rigo, M. et al., Scientific Reports 5, 18413, doi:10.1038 / srep18413 (2015)), respectively. Analysis included evaluation of TCR scaffold charge and folding liability, as well as refinement of suboptimal TCR-pHLA interaction regions, which were streamlined by in silico mutagenesis evaluation based on options generated by rotamer library scanning. TCR reconstituted SKW3 cells, which gave rise to secondary TCR combination mutant versions that were validated in the same manner, so only candidate mutations with wet-lab interaction profiles (e.g., alanine scanning data) were advanced for subsequent wet-lab confirmation.

[0230] In particular, alanine scanning data confirmed the importance of W99 in the beta chain. Replacement of residue 7 of KVLEHVVRV (SEQ ID NO: 122) with alanine reduced recognition of pHLA as measured by ELISPOT. This suggests that the TCR beta chain CDR3-W99 This was hypothesized to be due to the stabilizing effect of the linchpin.

[0231] Design and generation of CDR1 and CDR2 libraries A second TCR library was prepared containing mutations in CDR1 and CDR2 of the parental TCR (also known as the WT TCR).

[0232] To generate variants, different approaches were used: i) a focused, unbiased approach, and ii) a focused, information-based approach based on interrogation of structural models and known structures.

[0233] i) involved strong randomization of CDR1a, CDR2a, and CDR1b, and ii) involved focusing on positions close to critical residues likely to be involved in peptide contacts, HLA contacts, or CDR stabilization.

[0234] The quality of the CDR1 and CDR2 libraries was measured by sequencing to assess the quality of the inserts, to check the background frequency, and to check the ratio between the mixed libraries. The functionality of the libraries was tested by TCR cellular expression, activation with 10 μM MAGE-A4 peptide, and flow cytometry analysis, as described in Example 4 and in more detail below.

[0235] Generation of CDR1 and CDR2 libraries The designed TCR library was provided as subcloning-compatible dsDNA. CDR loop definition and library design were based on IMGT and predicted 3D models. The library was subcloned into the pMSCVneo vector (Clontech) carrying the target gene with the mouse TCR domain (Cohen, CJ et al., Cancer research 66, 8878-8886, doi:10.1158 / 0008-5472.CAN-06-1450 (2006)). The resulting plasmid DNA for cell transfection was prepared from the resulting E. coli material using standard methods.

[0236] Plasmid DNA from the library was transfected into GP2-293 packaging cells (ClonTech) along with the pAmpho (ClonTech) packaging vector using Lipofectamine 2000 transfection reagent (Invitrogen). Retroviral supernatant was used to transduce target cells (SKW-3, Cell Line Services, Eppelheim, Germany). Transduced SKW-3 cells were stained with H57-PE (BD Pharmingen) and anti-CD3-A647 (BD Biosciences) mAbs to identify TCR-positive populations. Double-positive events were isolated on a SONY SH800 flow cytometer.

[0237] Functional selection of cell-presented TCR CDR1 and CDR2 Briefly, T2 cells (ATCC) were pulsed with 10 nM KVL peptide (Genscript) and cocultured with SKW-3 cells carrying the library. After 16 hours of coculture, cells were stained with a-HLA-A2-FITC (BD Pharmingen), a-CD25-PE (Miltenyi Biotech), and a-CD69-APC (BD Biosciences) mAbs. HLA-A2-negative and CD25 and CD69-double-positive cells were single-cell sorted in growth medium containing 20% ​​FCS using a Sony SH800 or BD FACSAria II cytometer. Expanded single cells were sequenced to confirm clonal identity. Briefly, 40,000 cells were lysed using Cell Lysis Buffer (TaKaRa). RNA transcription library candidates were amplified using a One-Step RT-PCR kit (QIAGEN). The resulting PCR products were sequenced (GeneWiz) and clone identities were revealed using standard sequence analysis tools.

[0238] Example 6 Variant screening The TCR panel generated by the experiments in Examples 3 to 5 was then screened. The candidate TCRs tested and disclosed herein are listed in the table below.

[0239] [Table 6A]

[0240] [Table 6B]

[0241] Viral vectors were prepared for transduction of peripheral blood mononuclear cells (PBMCs) with candidate TCRs. All TCRs in the table above were sufficiently expressed in the cells. These cells were then tested in an IFNγ ELISPOT assay to measure the activity of each TCR against antigen-presenting cells pulsed with the target peptide. The cells were also tested against various cancer cell lines, including NCI-H1703 and C33A, both of which express the target peptide in the correct HLA context.

[0242] [Table 7A]

[0243] [Table 7B]

[0244] [Table 7C]

[0245] [Table 7D]

[0246] Materials and Methods for Example 6 Production of lentiviral supernatant HEK293T (ATCC) cells are seeded 5–6 hours prior to transfection in DMEM supplemented with 5% FBS and 50 μg / ml gentamicin in poly-L-lysine-precoated 6-well plates. When the cells are fully attached to the plastic, they are transfected by adding a mixture of X-tremeGene™ 9 (Merck), helper plasmids pALD-Rev-K, pALD-GagPol-K, and pALD-VSV-GK (Aldevron), and 1 μg of the plasmid of interest, or nuclease-free HO for a mock control. Cells are initially incubated at 37°C and 5% CO for 16–18 hours, after which the medium is replaced with DMEM supplemented with 5% FBS and 50 μg / ml gentamicin. After a further 48 hours of incubation at 37°C, 5% CO2, the supernatant is filtered through a 0.45 μm syringe filter and stored at +4°C until use.

[0247] Activation of PBMCs HLA-A2-negative PBMCs were thawed and plated at 1 × 10 in a 24-well plate pre-coated with α-CD3 and α-CD28 (both from Thermo Fisher Scientific). 6 PBMCs were seeded at a density of 1 ml / well in X-vivo15 (Lonza) supplemented with 10% CTS™ Immune Cell SR (Thermo Fisher) and 100 IU / ml IL-2 (Peprotech) (hereafter referred to as complete X-vivo15). After 48 h of incubation, activated PBMCs were collected, washed, and counted.

[0248] Transduction of Tc with viral supernatant Activated PBMCs were cultured at 1 × 10 in complete X-vivo 15 containing 200 IU / ml IL-2 and Lentiboost (Sirion Biotech). 6The cells were seeded into a 6-well plate at a concentration of 1000 / ml and mixed 1:1 with 1 ml of viral supernatant. The plate was then spun down at 900 x g for 1 hour at 32°C and incubated at 37°C, 5% CO2 for 20 hours.

[0249] Checking transduction efficiency and subculturing T cells Transduced T cells were collected, washed, and cultured at 0.33 x 10 in complete X-vivo 15 containing 50 UI / ml IL-2, 2 ng / ml IL-7, and 2 ng / ml IL-15. 6 Tc cultures are seeded at 1000 / ml and incubated at 37°C, 5% CO2. After 72 h, a small aliquot of cells is used to assess transduction efficiency by staining with chain-specific monoclonal antibodies conjugated to fluorescent dyes and acquired on a Cytoflex (Beckman Coulter) flow cytometer. Tc cultures are diluted approximately 1:4 with fresh complete X-vivo15 containing 50 UI / ml IL-2, 2 ng / ml IL-7, and 2 ng / ml IL-15 and incubated for 24 h before use in functional assays.

[0250] Peptide susceptibility and efficacy testing To assess TCR sensitivity, transduced Tc were incubated with T2 cells (DSMZ) pulsed with titrated amounts of the peptide of interest (Genscript) at an effector to target ratio (E:T) of 1:5. IFNγ detection by ELISpot was used as the readout. Briefly, on the day before the assay, 96-well filter plates with 0.45 μm pore size hydrophobic PVDF membranes (Merck) were activated with 35% ethanol and washed four times with sterile dH2O. An α-IFNγ coating antibody (ELISpot Flex: Human IFN-γ (ALP) Kit (Mabtech)) was then added and incubated overnight at +4°C. On the day of the assay, the coating antibody was washed off with PBS, and the plates were blocked in complete Xvivo15 at room temperature for 30 minutes. T2 cells were counted and cultured at 0.5 × 10 6The cells were resuspended at a concentration of 1000 / ml and incubated with 12 titrated concentrations of the target peptide for at least 2 h at 37°C, 5% CO2, with unpulsed T2 cells and T2 cells pulsed with an irrelevant peptide serving as controls. At the end of the incubation, the pulsed T2 cells were seeded in triplicate into conditioned filter plates.

[0251] To evaluate efficacy, TCR-transduced Tc are incubated with HLA-A2-expressing cancer cell lines and TCR target antigens at two E:T ratios: 1:5 and 1:10. IFNγ detection by ELISpot is used as the readout. Briefly, filter plates are prepared as described in the previous paragraph. NCI-H1703, C33A, and control cell lines (all from ATCC) are collected and incubated at 0.5 × 10 in complete Xvivo15. 6 The cells are resuspended at a concentration of 1000 / ml and plated twice in triplicate into conditioned plates.

[0252] Effector cells are counted and resuspended in complete Xvivo15 without cytokines. To rest the cells prior to functional assays, Tc are incubated alone for at least 3 hours before adding them to the plates containing target cells. The percentage of positive TCR in each condition is normalized to the condition showing the lowest expression by diluting the cells with mock-transduced Tc. The normalized Tc are then diluted to 0.1 x 10 6 The remaining effector cells are resuspended at 0.05 x 10 cells / ml and seeded into the plates containing the pulsed T2 cells and the cancer cell line targets (1:5 E:T condition only). 6 The cells were diluted to a concentration of 1000 / ml and added to the 1:10 E:T co-culture with the cancer cell lines. The plates were incubated overnight at 37°C and 5% CO. Plate detection and color development were performed using the ELISpot Flex: Human IFN-γ (ALP) Kit (Mabtech) according to the manufacturer's instructions.

[0253] Example 7 Further screening of variants Further TCRs are screened. The candidate TCRs tested are listed in Table 6 as KVL-01-145, KVL-01-149, KVL-01-151, and KVL-01-153.

[0254] The results of the screening are shown in Figure 4, which shows the results of an IFNγ ELISPOT assay. The target cell line in the assay was NCI-H1703 (MAGE-A4+ and HLA-A2+). Further details of the IFNγ ELISPOT assay method are provided in Example 6.

Claims

1. A T cell receptor (TCR), or an antigen-binding portion thereof, capable of specifically binding to the peptide set forth in SEQ ID NO: 122 presented on an MHC class I molecule, TCR-alpha (TCR-a) variable region, or a portion thereof, and TCR-beta (TCR-b) variable region, or a portion thereof Including, the TCR-a variable region, or a portion thereof, comprises a complementarity determining region 3 alpha (CDR3a) comprising SEQ ID NO:49 or SEQ ID NO:53, wherein the CDR3a does not comprise SEQ ID NO:7; and / or the TCR-b variable region, or a portion thereof, comprises a complementarity determining region 3 beta (CDR3b) comprising SEQ ID NO: 62 or SEQ ID NO: 64, wherein CDR3b does not comprise SEQ ID NO: 14; TCR, or its antigen-binding portion.

2. CDR3a comprises SEQ ID NO: 49 or SEQ ID NO: 53, but does not comprise SEQ ID NO: 7; The TCR-b variable region, or a portion thereof, comprises a CDR3b comprising any one of SEQ ID NOs: 14 and 57 to 66; 2. The TCR of claim 1, or an antigen-binding portion thereof.

3. CDR3a comprises SEQ ID NO: 55, CDR3b comprises any one of SEQ ID NOs: 14 and 57-61; A TCR, or an antigen-binding portion thereof, according to claim 2.

4. the TCR-a variable region, or portion thereof, comprises a CDR3a comprising any one of SEQ ID NOs: 7, 26-55, 140, and 141, or optionally any one of SEQ ID NOs: 7 and 26-55; CDR3b comprises SEQ ID NO: 62 or SEQ ID NO: 64, and CDR3b does not comprise SEQ ID NO: 14; 2. The TCR of claim 1, or an antigen-binding portion thereof.

5. CDR3a comprises any one of SEQ ID NOs: 7, 26-48, 140, and 141, or optionally SEQ ID NOs: 7 and 26-48; CDR3b comprises SEQ ID NO: 66; A TCR, or an antigen-binding portion thereof, according to claim 4.

6. A TCR, or an antigen-binding portion thereof, according to any one of claims 1 to 5, wherein the TCR-a variable region, or a portion thereof, comprises a CDR1a comprising any one of SEQ ID NOs: 5 and 15 to 17.

7. A TCR, or an antigen-binding portion thereof, according to any one of claims 1 to 6, wherein the TCR-a variable region, or a portion thereof, comprises CDR2a comprising any one of SEQ ID NOs: 6 and 18 to 25.

8. A TCR, or an antigen-binding portion thereof, according to any one of claims 1 to 7, wherein the TCR-b variable region, or a portion thereof, comprises CDR1b comprising sequence number 12.

9. A TCR, or an antigen-binding portion thereof, according to any one of claims 1 to 8, wherein the TCR-b variable region, or a portion thereof, comprises CDR2b comprising sequence number 13 or sequence number 56.

10. CDR3a comprises SEQ ID NO: 49 or SEQ ID NO: 53, but does not comprise SEQ ID NO: 7; the TCR-a variable region, or a portion thereof, comprises a CDR1a comprising any one of SEQ ID NOs: 5, 15 to 17 and a CDR2a comprising any one of SEQ ID NOs: 6 and 18 to 25; the TCR-b variable region, or a portion thereof, comprises a CDR1b comprising SEQ ID NO: 12, a CDR2b comprising SEQ ID NO: 13 or SEQ ID NO: 56, and a CDR3b comprising any one of SEQ ID NOs: 14 and 57 to 61; 2. The TCR of claim 1, or an antigen-binding portion thereof.

11. the TCR-a variable region, or a portion thereof, comprises a CDR1a comprising any one of SEQ ID NOs: 5 and 15-17, a CDR2a comprising any one of SEQ ID NOs: 6 and 18-25, and a CDR3a comprising any one of SEQ ID NOs: 7, 26-48, 140, and 141, or optionally any one of SEQ ID NOs: 7 and 26-48; the TCR-b variable region, or a portion thereof, comprises CDR1b comprising SEQ ID NO: 12 and CDR2b comprising SEQ ID NO: 13 or SEQ ID NO: 56; CDR3b comprises SEQ ID NO: 62 or SEQ ID NO: 64, and CDR3b does not comprise SEQ ID NO: 14; 2. The TCR of claim 1, or an antigen-binding portion thereof.

12. A TCR, or an antigen-binding portion thereof, capable of specifically binding to a peptide set forth in SEQ ID NO: 122 presented on an MHC class I molecule, TCR-a variable region, or a portion thereof, and TCR-b variable region, or a portion thereof Including, a TCR-a variable region, or a portion thereof, comprising CDR3a, and a TCR-b variable region, or a portion thereof, comprising CDR3b; wherein the CDR3a comprises any one of the CDR3a sequences listed in Table 1; wherein the CDR3b comprises any one of the CDR3b sequences listed in Table 2; A TCR, or antigen-binding portion thereof, in which the CDR3a and CDR3b are associated with the same TCR ID listed in Table 1 and Table 2.

13. A TCR, or an antigen-binding portion thereof, capable of specifically binding to a peptide set forth in SEQ ID NO: 122 presented on an MHC class I molecule, TCR-a variable region, or a portion thereof, and TCR-b variable region, or a portion thereof Including, the TCR-a variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a, and the TCR-b variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b; CDR1a comprises any one of the CDR1a sequences listed in Table 1; CDR2a comprises any one of the CDR2a sequences listed in Table 1; wherein the CDR3a comprises any one of the CDR3a sequences listed in Table 1; CDR1b comprises any one of the CDR1b sequences listed in Table 2; CDR2b comprises any one of the CDR2b sequences listed in Table 2; wherein the CDR3b comprises any one of the CDR3b sequences listed in Table 2; A TCR, or antigen-binding portion thereof, in which CDR1a, CDR2a, CDR3a, CDR1b, CDR2b, and CDR3b all associate with the same TCR ID (i.e., are derived from the same clone) listed in Tables 1 and 2.

14. A TCR, or an antigen-binding portion thereof, capable of specifically binding to a peptide set forth in SEQ ID NO: 122 presented on an MHC class I molecule, TCR-a variable region, or a portion thereof, and TCR-b variable region, or a portion thereof Including, (1) the TCRa variable region, or a portion thereof, comprises a CDR3a comprising or in accordance with SEQ ID NO:7 or SEQ ID NO:26, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or in accordance with SEQ ID NO:14 or SEQ ID NO:57; or (2) the TCRa variable region, or a portion thereof, comprises a CDR3a comprising or in accordance with SEQ ID NO: 7 or SEQ ID NO: 26, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or in accordance with SEQ ID NO: 14 or SEQ ID NO: 57; or (3) the TCRa variable region, or a portion thereof, comprises a CDR3a comprising or in accordance with SEQ ID NO: 7 or SEQ ID NO: 26, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or in accordance with SEQ ID NO: 14 or SEQ ID NO: 57; or (4) the TCRa variable region, or a portion thereof, comprises a CDR3a comprising or in accordance with SEQ ID NO: 7 or SEQ ID NO: 26, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or in accordance with SEQ ID NO: 14 or SEQ ID NO: 57; or (5) the TCRa variable region, or a portion thereof, comprises a CDR3a comprising or in accordance with SEQ ID NO: 27 or SEQ ID NO: 28, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or in accordance with SEQ ID NO: 14 or SEQ ID NO: 57; or (6) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 29 or SEQ ID NO: 30, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (7) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 31 or SEQ ID NO: 32, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (8) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 33 or SEQ ID NO: 34, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (9) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 29 or SEQ ID NO: 30, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (10) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 31 or SEQ ID NO: 32, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (11) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 29 or SEQ ID NO: 30, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (12) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 31 or SEQ ID NO: 32, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (13) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 7 or SEQ ID NO: 26, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 58 or SEQ ID NO: 59; or (14) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 7 or SEQ ID NO: 26, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 58 or SEQ ID NO: 59; or (15) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 7 or SEQ ID NO: 26, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 58 or SEQ ID NO: 59; or (16) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 7 or SEQ ID NO: 26, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 58 or SEQ ID NO: 59; or (17) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 27 or SEQ ID NO: 28, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 58 or SEQ ID NO: 59; or (18) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 7 or SEQ ID NO: 26, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 58 or SEQ ID NO: 59; or (19) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 31 or SEQ ID NO: 32, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 58 or SEQ ID NO: 59; or (20) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 35 or SEQ ID NO: 36, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (21) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 37 or SEQ ID NO: 38, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (22) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 39 or SEQ ID NO: 40, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (23) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 41 or SEQ ID NO: 42, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (24) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 35 or SEQ ID NO: 36, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 58 or SEQ ID NO: 59; or (25) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 37 or SEQ ID NO: 38, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 58 or SEQ ID NO: 59; or (26) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 39 or SEQ ID NO: 40, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 58 or SEQ ID NO: 59; or (27) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 41 or SEQ ID NO: 42, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 58 or SEQ ID NO: 59; or (28) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 43 or SEQ ID NO: 44, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 58 or SEQ ID NO: 59; or (29) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 29 or SEQ ID NO: 30, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (30) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 29 or SEQ ID NO: 30, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (31) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 31 or SEQ ID NO: 32, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (32) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 31 or SEQ ID NO: 32, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (33) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 31 or SEQ ID NO: 32, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (34) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 7 or SEQ ID NO: 26, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (35) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 31 or SEQ ID NO: 32, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (36) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 31 or SEQ ID NO: 32, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (37) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 31 or SEQ ID NO: 32, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (38) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 31 or SEQ ID NO: 32, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (39) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 31 or SEQ ID NO: 32, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (40) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 29 or SEQ ID NO: 30, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (41) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 29 or SEQ ID NO: 30, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (42) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 29 or SEQ ID NO: 30, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (43) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 45 or SEQ ID NO: 46, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (44) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 47 or SEQ ID NO: 48, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (45) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 45 or SEQ ID NO: 46, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 58 or SEQ ID NO: 59; or (46) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 45 or SEQ ID NO: 46, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (47) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 47 or SEQ ID NO: 48, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (48) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 31 or SEQ ID NO: 32, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (49) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 31 or SEQ ID NO: 32, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; or (50) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 31 or SEQ ID NO: 32, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 60 or SEQ ID NO: 61; or (51) The TCRa variable region, or a portion thereof, comprises a CDR3a comprising or according to SEQ ID NO: 140 or SEQ ID NO: 141, and the TCRb variable region, or a portion thereof, comprises a CDR3b comprising or according to SEQ ID NO: 14 or SEQ ID NO: 57; TCR, or its antigen-binding portion.

15. A TCR, or an antigen-binding portion thereof, capable of specifically binding to a peptide set forth in SEQ ID NO: 122 presented on an MHC class I molecule, TCR-a variable region, or a portion thereof, and TCR-b variable region, or a portion thereof Including, (1) the TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 18, and 7 / 26, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (2) the TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 19, 7 / 26, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (3) the TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 20, 7 / 26, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (4) the TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 21, 7 / 26, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (5) the TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 6, 27 / 28, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (6) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 15, 6, 29 / 30, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (7) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 6, 31 / 32, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (8) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 15, 6, 33 / 34, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (9) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 6, 29 / 30, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (10) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 15, 6, 31 / 32, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (11) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 15, 6, 29 / 30, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (12) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 6, 31 / 32, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (13) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 18, 7 / 26, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 58 / 59, respectively; or (14) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 19, 7 / 26, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 58 / 59, respectively; or (15) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 20, 7 / 26, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 58 / 59, respectively; or (16) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 21, 7 / 26, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 58 / 59, respectively; or (17) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 6, 27 / 28, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 58 / 59, respectively; or (18) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 6, 7 / 26, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 58 / 59, respectively; or (19) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 6, 31 / 32, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 58 / 59, respectively; or (20) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 6, 35 / 36, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (21) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 6, 37 / 38, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (22) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 6, 39 / 40, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (23) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 6, 41 / 42, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (24) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 6, 35 / 36, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 58 / 59, respectively; or (25) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 6, 37 / 38, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 58 / 59, respectively; or (26) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 6, 39 / 40, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 58 / 59, respectively; or (27) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 6, 41 / 42, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 58 / 59, respectively; or (28) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 6, 43 / 44, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 58 / 59, respectively; or (29) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 15, 22, 29 / 30, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (30) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 15, 23, 29 / 30, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (31) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 15, 22, 31 / 32, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (32) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 15, 23, 31 / 32, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (33) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 15, 6, 31 / 32, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (34) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 15, 6, 7 / 26, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (35) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 16, 22, 31 / 32, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (36) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 17, 23, 31 / 32, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (37) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 17, 6, 31 / 32, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (38) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 23, 31 / 32, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (39) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 22, 31 / 32, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (40) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 6, 29 / 30, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (41) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 23, 29 / 30, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (42) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 22, 29 / 30, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (43) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 6, 45 / 46, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (44) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 19, 47 / 48, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (45) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 19, 45 / 46, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 58 / 59, respectively; or (46) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 19, 45 / 46, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (47) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 19, 47 / 48, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (48) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 19, 31 / 32, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (49) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 19, 31 / 32, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 56, 14 / 57, respectively; or (50) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 6, 31 / 32, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 60 / 61, respectively; or (51) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 6, 140 / 141, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; or (52) The TCRa variable region, or a portion thereof, comprises CDR1a, CDR2a, and CDR3a comprising or according to SEQ ID NOs: 5, 22, 140 / 141, respectively, and the TCRb variable region, or a portion thereof, comprises CDR1b, CDR2b, and CDR3b comprising or according to SEQ ID NOs: 12, 13, 14 / 57, respectively; TCR, or its antigen-binding portion.

16. the TCR-a variable region comprises a sequence at least 70%, 80%, 90%, 95% or 100% identical to SEQ ID NO: 71, a sequence at least 70%, 80%, 90%, 95% or 100% identical to SEQ ID NO: 72, a sequence at least 70%, 80%, 90%, 95% or 100% identical to SEQ ID NO: 73, and a sequence at least 70%, 80%, 90%, 95% or 100% identical to SEQ ID NO: 74; and / or the TCR-b variable region comprises a sequence at least 70%, 80%, 90%, 95%, or 100% identical to SEQ ID NO: 75, a sequence at least 70%, 80%, 90%, 95%, or 100% identical to any one of SEQ ID NOs: 76-79, a sequence at least 70%, 80%, 90%, 95%, or 100% identical to SEQ ID NO: 80, and a sequence at least 70%, 80%, 90%, 95%, or 100% identical to SEQ ID NO: 81; A TCR, or an antigen-binding portion thereof, according to any one of claims 1 to 15.

17. the TCR-a variable region comprises a sequence, or a portion thereof, that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 3, 82-116, 142, and 143; and / or the TCR-b variable region comprises a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 10, 117-121, and 144, or a portion thereof; optionally, the TCR-a variable region comprises a sequence, or a portion thereof, that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 3 and 82-116; and / or the TCR-b variable region comprises a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 10, 117-121, or a portion thereof; A TCR, or an antigen-binding portion thereof, according to any one of claims 1 to 16.

18. A TCR, or an antigen-binding portion thereof, capable of specifically binding to a peptide set forth in SEQ ID NO: 122 presented on an MHC class I molecule, a TCR-a variable region comprising a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 3, 82-116, 142, and 143, or a portion thereof; and A TCR-b variable region comprising a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 10, 117-121, and 144, or a portion thereof. Including, the TCR does not comprise both SEQ ID NO: 3 and SEQ ID NO: 10, and optionally the TCR-a variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 3 and 82-116, or a portion thereof; the TCR-b variable region comprises a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 10 and 117-121, or a portion thereof; TCR, or its antigen-binding portion.

19. A TCR, or an antigen-binding portion thereof, capable of specifically binding to a peptide set forth in SEQ ID NO: 122 presented on an MHC class I molecule, a TCR-a variable region comprising a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 3, 82-116, 142, and 143, or a portion thereof; and A TCR-b variable region comprising a sequence that is at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 10, 117-121, and 144, or a portion thereof. Including, The TCR-a variable region sequence and the TCR-b variable region sequence are associated with the same TCR ID listed in Table 3 and Table 4. TCR, or its antigen-binding portion.

20. A TCR, or an antigen-binding portion thereof, capable of specifically binding to a peptide set forth in SEQ ID NO: 122 presented on an MHC class I molecule, TCR-a variable region, or a portion thereof, and TCR-b variable region, or a portion thereof Including, (1) the TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 82 and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (2) the TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 83, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (3) the TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 84 and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (4) the TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 85 and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (5) the TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 86, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (6) the TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 87, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (7) the TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 88 and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 117; or (8) the TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 89 and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 117; or (9) the TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 90, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 117; or (10) the TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 91, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 117; or (11) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 92, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 117; or (12) the TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 93, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (13) the TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 82, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 118; or (14) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 83, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 118; or (15) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 84, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 118; or (16) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 85, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 118; or (17) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 86, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 118; or (18) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO:3, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO:118; or (19) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 93, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 118; or (20) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 94, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (21) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 95, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (22) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 96, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (23) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 97, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (24) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 94, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 118; or (25) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 95, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 118; or (26) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 96, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 118; or (27) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 97, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 118; or (28) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 98, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 118; or (29) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 99, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (30) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 100, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (31) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 101, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (32) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 102, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (33) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 103, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (34) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 104, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (35) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 105, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (36) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 106, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (37) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 107, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 119; or (38) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 108, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (39) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 109, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (40) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 110, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (41) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 111, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (42) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 112, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (43) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 113, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (44) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 114, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (45) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 115, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 118; or (46) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 115, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (47) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 114, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (48) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 116, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (49) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 116, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 120; or (50) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 93, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 121; or (51) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 142, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (52) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 143, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 10; or (53) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 142, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 119; or (54) The TCRa variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 98, and the TCRb variable region comprises a sequence at least 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO:

144. TCR, or its antigen-binding portion.

21. (i) the CDR3a sequence is constant and the CDR3b sequence is constant, the CDR3a sequence being defined in Table 3 and the CDR3b sequence being defined in Table 4; and / or (ii) the CDR1a, CDR2a, and CDR3a sequences are unchanged, and the CDR1b, CDR2b, and CDR3b sequences are unchanged, and the CDR sequences are as defined in Table 3 and Table 4; and / or (iii) carry the relevant mutations listed in Table 6; 21. A TCR, or an antigen-binding portion thereof, according to any one of claims 18 to 20.

22. MHC class I molecules are HLA-A * 0201, or an antigen-binding portion thereof, according to any one of claims 1 to 21.

23. K less than or equal to 200 μM, 175 μM, 143 μM, 100 μM, 75 μM, 50 μM, 25 μM, 20 μM, 15 μM, 10 μM, 5 μM, or 2 μM d 23. A TCR, or an antigen-binding portion thereof, according to any one of claims 1 to 22, having:

24. 24. A TCR, or an antigen-binding portion thereof, according to any one of claims 1 to 23, comprising a TCRa constant region and / or a TCRb constant region.

25. the TCRa constant region sequence is 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to SEQ ID NO:4; and / or the TCRb constant region sequence is 70%, 80%, 85%, 90%, 95%, 97%, 98%, 99%, or 100% identical to any one of SEQ ID NOs: 11, 138, or 139; 25. A TCR, or an antigen-binding portion thereof, according to claim 24.

26. 26. A TCR, or antigen-binding portion thereof, according to any one of claims 1 to 25, linked to an agent.

27. 27. The TCR, or antigen-binding portion thereof, of claim 26, wherein the agent is a therapeutic agent.

28. (a) TCRa variable region amino acid sequence; (b) a TCRb variable region amino acid sequence; or (c) both the TCRa variable region and the TCRb variable region amino acid sequence of the TCR or antigen-binding portion of any one of claims 1 to 25 A nucleic acid molecule encoding

29. 29. A vector comprising the nucleic acid of claim 28.

30. 30. A cell comprising a TCR, or an antigen-binding portion thereof, described in any one of claims 1 to 27, a nucleic acid described in claim 28, or a vector described in claim 29.

31. 31. The cell of claim 30, which is an immune cell.

32. The cell of claim 31 , wherein the immune cell is a T cell or an NK cell.

33. A pharmaceutical composition comprising a TCR, or an antigen-binding portion thereof, described in any one of claims 1 to 27, a nucleic acid described in claim 28, a vector described in claim 29, or a cell described in any one of claims 30 to 32.

34. 34. A TCR, or an antigen-binding portion thereof, according to any one of claims 1 to 27, a nucleic acid according to claim 28, a vector according to claim 29, a cell according to any one of claims 30 to 32, or a pharmaceutical composition according to claim 33, for use as a pharmaceutical.

35. 34. A TCR, or antigen-binding portion thereof, according to any one of claims 1 to 27, a nucleic acid according to claim 28, a vector according to claim 29, a cell according to any one of claims 30 to 32, or a pharmaceutical composition according to claim 33, for use in a method for treating, preventing, or ameliorating cancer.

36. The TCR, antigen-binding portion thereof, nucleic acid, vector, cell, or pharmaceutical composition of claim 35, wherein the cancer comprises cells that express MAGE-A4.

37. The TCR, its antigen-binding portion, nucleic acid, vector, cell, or pharmaceutical composition of claim 35 or 36, wherein the cancer is non-small cell lung cancer (NSCLC), ovarian cancer, gastroesophageal cancer, head and neck synovial sarcoma, or urothelial cancer.

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