Common tumor-specific T cell receptor

Abstract

The present invention relates to a common patient - general tumor - specific T - cell receptor (TCR), a nucleic acid encoding the TCR, a T cell comprising the TCR and / or the encoding nucleic acid, and these agents for use in cancer treatment.

Description

Technical Field

[0001] This application claims the priority of European Patent Application (EP) No. 22176133.1 filed on May 30, 2022 and European Patent Application (EP) No. 23152250.9 filed on January 18, 2023, both of which are incorporated herein by reference.

[0002] The present invention relates to common patient-spanning tumor-specific T cell receptors (TCRs), nucleic acids encoding TCRs, T cells comprising TCRs and / or encoding nucleic acids, and these agents for use in cancer treatment.

Background Art

[0003] Since it has been shown that the immune system has the ability to combat and reject tumors, significant efforts have been made in the development of cancer treatment or cancer prevention vaccines. Such efforts have faced difficult challenges in antigen discovery because tumor antigens suitable for vaccination need to meet three basic requirements. These need to be immunogenic in order to elicit an effective therapeutic response. These need to be tumor-specific in order to enable safe treatment, especially in a preventive scenario. However, cross-reactivity with pathogen-related antigens is within the scope of the present invention. Finally, it is necessary to identify shared antigens expressed in the tumors of many patients.

[0004] In the past decade, two high-throughput platforms have been developed and widely used in tumor antigen discovery. However, both only meet the conditions in a limited range. It is clear that tumor-specific mutant antigens can be identified by whole exome sequencing approaches. However, with the exception of certain recurrent driver mutations, most of the identified mutant-reactive antigens are patient-specific. On the other hand, mass spectrometry-based approaches can detect HLA-presented peptides shared by tumors from different patients. However, demonstrating the immunogenicity and tumor specificity of these peptides remains a difficult task.

[0005] Adoptive cell therapy (ACT) using T cells genetically engineered to express tumor-reactive chimeric antigen receptors (CAR-T cells) or T cell receptors (TCRs) is a promising treatment strategy for cancer patients. In contrast to hematological malignancies (where CAR-T cells against antigens on the surface of specific lineage-specific cells have been approved due to manageable side effects and high efficacy), in solid tumors, the application of CAR-T cells is (currently) not achievable due to the lack of cell surface target antigens that are expressed limitedly in tumors. Tumor-specific transgenic TCR (tsTCRtg-T cells) genetically modified (from a patient) to express a peptide recognized from a tumor-associated antigen or tumor-specific antigen (TAA or TSA) presented by an HLA molecule (pMHC) is an attractive alternative. TAAs (such as cancer / germline antigens, differentiation antigens, overexpressed antigens, etc.) and viral (v) TSAs (in tumors with a viral etiology) are widely shared among tumors, resulting in the presentation of common pMHCs in HLA-matched patients. However, most (non-viral) TSAs (neoantigens) are specific to individual cancers. The resulting diverse pMHCs have to be regarded as private antigens of individual subjects. However, in a few cases, it has been shown that TSAs caused by point mutations or chromosomal translocations that affect common driver genes of malignancy and are shared among tumors are immunogenic (e.g., RAS mutations, TP53 mutations, BRAF mutations, PIK3CA mutations, and translocations involving ALK, ROS, NTRK, RET, etc.). Also, antigen categories that are not yet clearly defined, such as tumor-specific cryptic ("dark matter (unknown)") transcripts or abnormally spliced transcripts, are also shared among tumors and may be recognized by T cells.

Prior Art Documents

Patent Documents

[0006] [Patent Document 1] International Publication (WO) No. 2017 / 025564 (A1) [Summary of the Invention] [Problems to be Solved by the Invention]

[0007] Previously, the inventors developed a method for identifying tumor-specific T cell receptors by comparing CDR3 sequences obtained from tumor-infiltrating lymphocytes (TILs) with T cells in adjacent tissues (WO2017 / 025564 A1). Therefore, it is possible to distinguish tumor specificity by the increased abundance of T cell clones in the tumor and non-tumor of a certain patient. However, most tumor-specific antigens are caused by mutations limited to individual patients. Privately owned neoantigens can be targeted by individualized tsTCRtg-T cell therapy, but shared TAAs or TSAs are ideal targets for off-the-shelf tsTCRtg-T cell therapy in patients with the expression of matching HLA alleles. Individualized treatment is time-consuming and costly, and the regulations of the FDA and EMA are also strict (ATMP, advanced medicinal products; gene therapy medicinal products). Furthermore, the diseases of many patients progress faster than individualized therapeutic agents can be manufactured. Therefore, it would be very beneficial to develop a method for identifying carriers of such common tumor-specific TCRs by scanning that the antigen recognition domains (CDR3α and β) of the TIL-repertoire are identical or very similar, whereby off-the-shelf therapeutic receptors are provided, and at the same time, the opportunity to identify shared tumor-specific antigens for further therapeutic options is expanded. [Means for Solving the Problems]

[0008] Based on the above state of the art, an object of the present invention is to provide a common tumor-specific TCR sequence. This object is achieved by the subject matter of the independent claims of the present specification, with further advantageous embodiments described in the dependent claims, examples, figures and general description of the present specification.

[0009] Summary of the Invention Rather than starting with antigen candidates that require critical validation in specificity and functionality tests, as an alternative approach, there is a method of analyzing the T cell repertoire of tumors in various cancer patients and exploring specific effects derived from shared tumor antigens. When T cells infiltrate a tumor and cause an interaction via a specific receptor with the tumor antigen, subsequent activation, proliferation, and enrichment of this clone within the tumor occur. Therefore, as determined quantitatively by the ratio of the frequency of TCR clonotypes (clone types) between the tumor and the non-tumor tissue adjacent to it, the preferred localization of this specific TCR clonotype serves as a predictor of tumor specificity. This technique is described in WO2017 / 025564 A1. When such specific tumor-specific TCR clonotypes, or structurally closely related TCR clonotypes called TCR clusters, are detected in the tumors of other patients, it indicates the presence of shared tumor antigens in these patients. This is particularly beneficial when TCR clusters are detected in HLA-matched patients and the nature of the HLA alleles presenting the shared antigen epitopes becomes clear. As a final step, for example, by single cell technology, if the clustered TCRs are fully elucidated, α / β-TCRs with specificity for the shared antigen will be obtained.

[0010] Such HLA-restricted α / β-TCRs with specificity for shared tumor antigens are the starting point for important applications.

[0011] As a novel tool without precedent, these can be used as specific probes in antigen discovery and can lead to a targeted search for shared tumor antigens.

[0012] As "off the shelf" TCRs in vector form, these can be used for introduction into the autologous T cells of cancer patients for immunotherapeutic intervention. The subjects are HLA-matched patients who are carriers of the clustered TCRs or carriers of known shared tumor antigens.

[0013] New genetic engineering technologies (CRISPR / Cas9, TALEN, zinc finger nucleases) are making it increasingly possible to produce allogeneic cell therapy products from a greater number of, and more accessible, healthy donors than from most patients, and to use a single product for the treatment of multiple patients. This is possible because (autologous and allogeneic) tsTCRtg-T cells can be genetically engineered to reduce immunogenicity (e.g., by knockout of endogenous HLA in the allogeneic setting), to be less prone to exhaustion / dysfunction (e.g., by knockout of checkpoint receptors), and to be less likely to cause graft-versus-host disease (GvHD) or unpredictable cross-reactivity by knockout of the endogenous TCR.

[0014] In addition to transducing conventional autologous or allogeneic CD4+ and CD8+ T cells with an α / β-tsTCR, it is also an option to transduce additional types of adaptive or innate immune cells, such as γδ-T cells, NKT cells, NK cells, that have these receptors: the above genetic engineering technologies enable co-introduction of an NK cell that has a tsTCR and the CD3 signaling domain required for cell activation upon engagement of the TCR with pMHC.

[0015] Therefore, it is highly advantageous to find T cell receptors and / or shared tumor antigens that are common to multiple individuals.

[0016] Therefore, there is a need to identify shared tumor-specific antigens and / or shared tumor-specific T cell receptors. If the HLA of patients is known to match, this enables off-the-shelf therapies for cancer.

[0017] A first aspect of the invention relates to an isolated TCR characterized by a specific CDR3 sequence.

[0018] The second aspect of the present invention relates to a nucleic acid sequence encoding a TCR according to the first aspect.

[0019] The third aspect of the present invention relates to an isolated autologous T cell comprising a TCR according to the first aspect and / or a nucleic acid sequence according to the second aspect.

[0020] The fourth aspect of the present invention relates to a TCR according to the first aspect, a nucleic acid sequence according to the second aspect, or an isolated autologous T cell according to the third aspect for use in the treatment of cancer.

[0021] In another embodiment, the present invention relates to a pharmaceutical composition comprising at least one of the TCR, nucleic acid sequence, or isolated autologous T cell of the present invention, and at least one pharmaceutically acceptable carrier, diluent, or excipient.

Mode for Carrying Out the Invention

[0022] Terms and Definitions For the purpose of interpreting this specification, the following definitions apply, and terms used in the singular form shall, where appropriate, include the plural form and vice versa. In the event of a conflict between the definitions set out below and the documents incorporated herein by reference, the definitions set out here shall prevail.

[0023] As used herein, the terms "comprising", "having", "containing", "including", and other similar forms, and their grammatically equivalent terms, are equivalent in meaning and are not intended to mean that one or more items following any one of these words comprehensively list, or are limited to, only the one or more listed items, but are intended to be open-ended. For example, an item "comprising" components A, B, and C can consist of (i.e., contain only) components A, B, and C, or can contain not only components A, B, and C but also one or more other components. Thus, "comprising" and its similar forms, and their grammatically equivalent terms, are intended and understood to include the disclosure of embodiments of "consisting essentially of" or "consisting of".

[0024] When a range of values is provided, unless the context clearly indicates otherwise, each intervening value between the upper and lower limits of that range, to one tenth of the unit of the lower limit, and any other stated value or intervening value within the stated range is understood to be included within the disclosure subject to any specifically excluded limitations of the stated range. When either or both of the limiting values of the stated range are included, the range excluding either or both of those included limiting values is also included in the disclosure.

[0025] As used herein, the reference to "about" a value or parameter includes (and describes) variations that are directed to that value or parameter itself. For example, a description that refers to "about X" includes the description of "X".

[0026] As used herein, including in the appended claims, the singular forms "a", "or", and "the" include plural referents unless the context clearly indicates otherwise.

[0027] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art (e.g., cell culture, molecular genetics, nucleic acid chemistry, hybridization techniques, and biochemistry). Standard techniques are used for molecular, genetic, and biochemical techniques (see generally: Sambrook et al., Molecular Cloning: A Laboratory Manual, 4th ed. (2012) Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y. and Ausubel et al., Short Protocols in Molecular Biology (2002) 5th ed., John Wiley & Sons, Inc.) and chemical techniques.

[0028] Sequence Sequences that are similar or identical (e.g., at least about 70% sequence identity) to the sequences disclosed herein are also part of the invention. In some embodiments, the sequence identity at the amino acid level can be about 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more than 99%. At the nucleic acid level, the sequence identity can be about 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more than 99%. Alternatively, substantial identity exists when a nucleic acid segment hybridizes to the complementary strand of its strand under selective hybridization conditions (e.g., under very high stringency hybridization conditions). The nucleic acid can be present in whole cells, in cell lysates, or in a partially purified or substantially pure form.

[0029] In the context of this specification, the terms "sequence identity" and "percentage of sequence identity" refer to a quantitative parameter representing the result of a sequence comparison determined by comparing two aligned sequences position by position. Methods for aligning sequences for comparison are well known in the art. Alignment of sequences for comparison can be performed by, but is not limited to, the local homology algorithm of Smith and Waterman, Adv. Appl. Math. 2:482 (1981), the global alignment algorithm of Needleman and Wunsch, J. Mol. Biol. 48:443 (1970), the similarity search method of Pearson and Lipman, Proc. Nat. Acad. Sci. 85:2444 (1988), or computerized implementations of these algorithms, including CLUSTAL, GAP, BESTFIT, BLAST, FASTA, and TFASTA. Software for performing BLAST analysis is publicly available, such as at the National Center for Biotechnology-Information (http: / / blast.ncbi.nlm.nih.gov / ).

[0030] As an example of amino acid sequence comparison, there is the BLASTP algorithm using default settings: Expect threshold: 10; Word size: 3; Maximum matches within query range: 0; Matrix: BLOSUM62; Gap costs: Existence 11, Extension 1; Compositional adjustment: Conditional compositional score matrix adjustment. As an example of nucleic acid sequence comparison, there is the BLASTN algorithm using default settings: Expect threshold: 10; Word size: 28; Maximum matches in query range: 0; Match / mismatch scores: 1 / -2; Gap costs: Linear. Unless otherwise stated, the values of sequence identity provided herein refer to the values obtained using a series of BLAST programs with the default parameters specified above for protein and nucleic acid comparison, respectively (Altschul et al., J. Mol. Biol. 215:403-410 (1990)).

[0031] References to identical sequences without specifying a percentage include the meaning of 100% identical sequences (i.e., the same sequence).

[0032] General biochemistry: Peptides, amino acid sequences The term "polypeptide" in the context of this specification refers to a molecule consisting of 50 or more amino acids that form a linear chain in which the amino acids are connected by peptide bonds. The amino acid sequence of a polypeptide may represent the amino acid sequence of an entire protein (found physiologically) or a fragment thereof. The terms "polypeptide" and "protein" are used interchangeably herein and include proteins and their fragments. Polypeptides are disclosed herein as amino acid residue sequences.

[0033] The term "peptide" in the context of this specification relates to a molecule consisting of a maximum of 50 amino acids, particularly 8 to 30 amino acids, more particularly 8 to 15 amino acids, that form a linear chain in which the amino acids are connected by peptide bonds.

[0034] The sequence of amino acid residues is described from the amino terminus to the carboxyl terminus. Capital letters at the sequence positions refer to L-amino acids in one-letter code (Stryer, Biochemistry, 3rd ed., p. 21). Lowercase letters indicating the positions in the amino acid sequence mean the corresponding D- or (2R)-amino acids. The sequence is described from left to right from the amino terminus to the carboxyl terminus. According to standard nomenclature, the sequence of amino acid residues is represented in either three-letter or one-letter code as follows: alanine (Ala, A), arginine (Arg, R), asparagine (Asn, N), aspartic acid (Asp, D), cysteine (Cys, C), glutamine (Gln, Q), glutamic acid (Glu, E), glycine (Gly, G), histidine (His, H), isoleucine (Ile, I), leucine (Leu, L), lysine (Lys, K), methionine (Met, M), phenylalanine (Phe, F), proline (Pro, P), serine (Ser, S), threonine (Thr, T), tryptophan (Trp, W), tyrosine (Tyr, Y), and valine (Val, V).

[0035] General molecular biology: nucleic acid sequences, expression The term "gene" refers to a polynucleotide containing at least one open reading frame (ORF) that can encode a specific polypeptide or protein after transcription and translation. The polynucleotide sequence can be used to identify a larger fragment or the full-length coding sequence of the gene with which it is associated. Methods for isolating the sequence of a larger fragment are known to those skilled in the art.

[0036] The terms "gene expression" or "expression", or the term "gene product", may refer to either, or both, the process of production of nucleic acid (RNA) or of peptide or polypeptide - and their products - which are also referred to as transcription and translation respectively, or may refer to any of the intermediate processes that regulate the processing of genetic information to yield a polypeptide product. The term "gene expression" also applies to the transcription and processing of RNA gene products, such as regulatory RNAs or structural (e.g., ribosomal) RNAs. When the expressed polynucleotide is derived from genomic DNA, expression may include mRNA splicing in eukaryotic cells. Expression can be evaluated at the level of transcription and translation, i.e., both mRNA and / or protein product.

[0037] The term "nucleotide" in the context of this specification relates to the building blocks of nucleic acids or nucleic acid analogs, the oligomers of which are capable of forming selective hybrids with RNA oligomers or DNA oligomers based on base pairing. The term "nucleotide" in this context includes the classical ribonucleotide building blocks adenosine, guanosine, uridine (and ribosylthymine), cytidine, the classical deoxyribonucleotide deoxyadenosine, deoxyguanosine, thymidine, deoxyuridine, and deoxycytidine. Further included are nucleic acid analogs such as phosphorothioate, 2’O-methylphosphorothioate, peptide nucleic acid (PNA; N-(2-aminoethyl)-glycine units linked by peptide bonds with a nucleic acid base attached to the α-carbon of glycine), or locked nucleic acid (LNA; 2’O,4’C methylene-bridged RNA building units). When reference is made herein to a "hybridization sequence", such a hybridization sequence can be composed of any of the above nucleotides, or mixtures thereof.

[0038] T cell biology The term "CDR3" in the context of this specification refers to the hypervariable complementarity determining region 3. The size of CDR3 is particularly characterized by the total number of amino acids (AAs) from the conserved cysteine in the Vβ, or Vα or Vγ or Vδ segment to the position of the conserved phenylalanine in the Jβ or Jα, Jγ or Jδ segment and the respective nucleotides.

[0039] The term "TCR" or "TCR polypeptide" in the context of this specification means a T cell receptor. Depending on the context, the term TCR encompasses any of the following: 1) A heterodimeric transmembrane protein composed of one alpha chain and one beta chain that is expressed in its native configuration in T cells and associates with accessory proteins for signal transduction; 2) A soluble truncated derivative of 1), which is composed of the variable domains of one alpha chain and one beta chain in its native (antigen-binding) configuration and is expressed as a fusion construct with various fusion partners that provide various effector functions.

[0040] The minimum requirement for a TCR is that it contains the CDR3 region and includes (at least the truncated form of) an alpha chain and a beta chain that can specifically bind to an antigen.

[0041] (Cancer) immunotherapy The term "HLA" in the context of the present invention refers to the human leukocyte antigen as a specific subset of the major histocompatibility complex (MHC), which is a general term.

[0042] HLA supertypes are defined based on grouping together MHC alleles (alleles) that share similar binding specificities, i.e., peptides having the same or similar so-called anchor amino acid residues (e.g., positions 2 and 9 or 10 in 9-mer and 10-mer peptides). HLA supertypes are further described in Sidney et al. (BMC Immunology 2008, 9:1).

[0043] The term "essentially identical" in the context of this specification relates to nucleic acid sequences that are identical or have an identity of at least 95%, particularly at least 97%, more particularly at least 98%, more particularly at least 99%, and most particularly more than 99%.

[0044] The term "gene of the same HLA type" in the context of this specification relates to the HLA gene encoding the MHC molecule. As used herein, the same HLA type means that the HLA gene encodes the same variant of the MHC molecule. Since there are a wide variety of HLA genes in humans, in one embodiment of the method of the present invention, the HLA repertoire of the tested patient is determined, and patients sharing at least one gene of the same HLA type are selected for further analysis.

[0045] As used herein, the term "pharmaceutical composition" refers to a compound of the present invention or a pharmaceutically acceptable salt thereof, accompanied by at least one pharmaceutically acceptable carrier. In certain embodiments, the pharmaceutical composition according to the present invention is provided in a form suitable for topical administration, parenteral administration, or administration by infusion.

[0046] As used herein, the term "pharmaceutically acceptable carrier" includes, as known to those skilled in the art, any solvent, dispersion medium, coating, surfactant, antioxidant, preservative (e.g., antibacterial agent, antifungal agent), isotonic agent, absorption delaying agent, salt, preservative, drug, drug stabilizer, binder, excipient, disintegrant, lubricant, sweetening agent, flavoring agent, coloring agent, etc., and combinations thereof (see, for example, Remington: the Science and Practice of Pharmacy, ISBN0857110624).

[0047] As used herein, the term "treating" or "treatment" of any disease or disorder (e.g., cancer) refers, in one embodiment, to alleviating the disease or disorder (e.g., delaying, preventing, or reducing the onset of at least one of the disease or its clinical symptoms). In another embodiment, "treating" or "treatment" refers to alleviating or improving at least one physical parameter, including those that may not be distinguishable in a patient. In yet another embodiment, "treating" or "treatment" refers to modulating the disease or disorder, either physically (e.g., stabilization of distinguishable symptoms) or physiologically (e.g., stabilization of physical parameters), or both. Methods for evaluating the treatment and / or prevention of a disease are generally known in the art unless otherwise specifically described hereinbelow.

[0048] The term "mutation" of a gene or protein refers to a change in a nucleic acid sequence or an amino acid sequence. This change results in a difference in the activity of each protein. The difference in activity means that in the case of KRAS, EGFR, FGFR, or BRAF, the signal pathway in which the protein is involved is upregulated, and in the case of TP53, it is downregulated.

[0049] The term "KRAS" refers to the gene of GeneID3845 or the protein of UniProt-ID P01116.

[0050] The term "EGFR" refers to the gene of GeneID1956 or the protein of UniProt-ID P00533.

[0051] The term "FGFR1" refers to the gene of GeneID2260 or the protein of UniProt-ID P11362.

[0052] The term "BRAF" refers to the gene of GeneID673 or the protein of UniProt-ID P15056.

[0053] The term "TP53" refers to the gene of GeneID7157 or the protein of UniProt-ID P04637.

[0054] The term "KRAS G12 mutation" refers to the substitution of glycine at position 12 of the KRAS protein with a different amino acid.

[0055] The term "KRAS Q61 mutation" refers to the substitution of glutamine at position 61 of the KRAS protein with a different amino acid.

[0056] The term "mutation of a gene in the EGFR-Raf-Ras pathway" refers to the mutation of a single gene in this pathway. This gene can be selected from growth factor receptors, KRAS, BRAF, MEK, and ERK.

[0057] Detailed Description of the Invention TCR clusters are correlated with the presence of tumor driver mutations. One central aspect of this invention is that in the tumors of patients in whom TCRs are found in a large cluster of very similar TCRs, known tumor driver mutations mainly derived from the KRAS family occur significantly (where "similar" refers to the peptide sequences of preferably both chains (alpha chain and beta chain) of the TCRs that together construct a functional TCR). TCR clusters mostly include different patients sharing an HLA type. Furthermore, the inventors provide evidence that many of the clusters include patients with lung cancer and pancreatic cancer (see clusters a, e, j, l, m, p).

[0058] The discovery of common mutations (Table 17) does not mean that each mutation is the direct source of a tumor antigen, i.e., that a peptide that is part of the mutated protein is presented as a neoantigen by tumor cells via MHC molecules. Homologous and functional antigens may be products of oncogenic signaling pathway activation that result in the expression of gene transcripts that are under the control of and / or modified by the activity of each driver mutation. This has broad implications for the application of clustered TCRs in therapy and diagnosis: a. Verification of the direct association of a clustered TCR with oncogenic pathway activation involved in the de novo generation of (unknown) tumor-specific antigens enables the selection of the TCR as a tumor-specific TCR. This selection of TCR is antigen-agnostic and suggests a safe application of TCRs in adoptive cell therapy (ACT) approaches using TCR-transduced T cells (TCR-T cells). b. In newly diagnosed patients, if a clustered TCR is present in either the tumor tissue (e.g., by needle biopsy or preserved tumor material) or peripheral blood of patients sharing the relevant HLA type, that patient is identified as a prospective recipient * (of a method of administering a large number of T cells equipped with a verified clustered TCR). c. Identification of a panel of tumor mutations is a standard diagnostic tool in current oncological practice and is mainly performed by deep sequencing techniques. Identification of mutations in a patient's tumor associated with a clustered TCR is a strong indicator that TCR-T therapy with TCR-transduced T cells will be beneficial as long as the patient exhibits an HLA type associated with the cluster. This may also be true if the patient does not have a measurable frequency of each clustered TCR. d. There is strong evidence that TCR-T therapy may be beneficial for tumor patients who meet both criteria of b and c, even if each cancer type is still included in clusters a - o. *TCR-T cell therapy (abbreviation: TCR-T therapy): A cell therapy using autologous / allogeneic T cells having a disease-specific T cell receptor (TCR)

[0059] TCR-T therapy using clustered TCR The identification of clustered TCRs in one cancer type, or even in various patients with various cancer types, is a very promising basis for TCR-T therapy. Each T cell for therapeutic use can be produced by any of the following: a. Transduction and ex vivo expansion of autologous / allogeneic T cells using a recombinant TCR construct derived from peripheral blood, or b. Expansion of selected endogenous T cells expressing clustered TCR isolated from a patient's tumor-infiltrating lymphocytes or peripheral blood lymphocytes. Isolation of such existing autologous clustered T cells can be achieved by enrichment and FACS using one or more specific anti-TCR ligands (e.g., antibodies) or multimeric HLA / peptide complexes prior to the expansion step.

[0060] In any case, prior to TCR-T therapy, patients can be stratified by screening blood or even tumor samples using DNA sequencing techniques and the methods developed by the inventors to identify TCR sequences from blood or tissue samples. When a patient has a TCR that is identical or very similar to a known clustered TCR and has TCRs of each HLA type, it can be expected to benefit from the corresponding TCR-T therapy.

[0061] The first aspect of the present invention relates to an isolated TCR polypeptide, wherein the TCR polypeptide comprises a CDR3 alpha sequence and a CDR3 beta sequence, and the CDR3 alpha sequence and the CDR3 beta sequence are identical to the sequences shown below, or have one or two amino acid substitutions per CDR3 sequence, where a. For group a, the CDR3 alpha sequence is selected from the group of sequences including SEQ ID NO: 007 - 009 or SEQ ID NO: 436 - 439, and the CDR3 beta sequence is selected from the group of sequences including SEQ ID NO: 001 - 006 or SEQ ID NO: 432 - 435, or b. For group b, the CDR3 alpha sequence is selected from the group of sequences including SEQ ID NO: 031 - 032, and the CDR3 beta sequence is selected from the group of sequences including SEQ ID NO: 027 - 030, or c. For group c, the CDR3 alpha sequence is selected from the group of sequences including SEQ ID NO: 052 - 054, and the CDR3 beta sequence is selected from the group of sequences including SEQ ID NO: 048 - 051, or d. For group d, the CDR3 alpha sequence is selected from the sequence of SEQ ID NO: 073, and the CDR3 beta sequence is selected from the group of sequences including SEQ ID NO: 070 - 072, or e. For group e, the CDR3 alpha sequence is selected from the sequence of SEQ ID NO: 100 or SEQ ID NO: 455 - 459, and the CDR3 beta sequence is selected from the group of sequences including SEQ ID NO: 086 - 099 or SEQ ID NO: 450 - 454, or f. For group f, the CDR3 alpha sequence is selected from the group of sequences including SEQ ID NO: 131 - 132 or SEQ ID NO: 472 - 473, and the CDR3 beta sequence is selected from the group of sequences including SEQ ID NO: 124 - 130 or SEQ ID NO: 470 - 471, or g. For group g, the CDR3 alpha sequence is selected from the sequence of SEQ ID NO: 156, and the CDR3 beta sequence is selected from the group of sequences including SEQ ID NO: 150 - 155, or h. For group h, the CDR3 alpha sequence is selected from the group of sequences including SEQ ID NO: 175 - 176, and the CDR3 beta sequence is selected from the group of sequences including SEQ ID NO: 172 - 174, or i. For group i, the CDR3 alpha sequence is selected from the group of sequences including SEQ ID NO: 193 - 194, and the CDR3 beta sequence is selected from the group of sequences including SEQ ID NO: 190 - 192, or j. For the j group, the CDR3 alpha sequence is selected from the group of sequences including SEQ ID NO: 212-213 or SEQ ID NO: 513, and the CDR3 beta sequence is selected from the group of sequences including SEQ ID NO: 208-211 or SEQ ID NO: 478, or k. For the k group, the CDR3 alpha sequence is selected from the group of sequences including SEQ ID NO: 232-235, and the CDR3 beta sequence is selected from the group of sequences including SEQ ID NO: 228-231, or l. For the l group, the CDR3 alpha sequence is selected from the group of sequences including SEQ ID NO: 261-269 or SEQ ID NO: 484-486, and the CDR3 beta sequence is selected from the group of sequences including SEQ ID NO: 252-260 or SEQ ID NO: 481-483, or m. For the m group, the CDR3 alpha sequence is selected from the group of sequences including SEQ ID NO: 302-307 or SEQ ID NO: 496-498, and the CDR3 beta sequence is selected from the group of sequences including SEQ ID NO: 296-301 or SEQ ID NO: 493-495, or n. For the n group, the CDR3 alpha sequence is selected from the group of sequences including SEQ ID NO: 334-339, and the CDR3 beta sequence is selected from the group of sequences including SEQ ID NO: 328-333, or o. For the o group, the CDR3 alpha sequence is selected from the group of sequences including SEQ ID NO: 363-365, and the CDR3 beta sequence is selected from the group of sequences including SEQ ID NO: 360-362, or p. For the p group, the CDR3 alpha sequence is selected from the group of sequences including SEQ ID NO: 391-401 or SEQ ID NO: 507-508, and the CDR3 beta sequence is selected from the group of sequences including SEQ ID NO: 380-390 or SEQ ID NO: 505-506, In particular, the CRD3 alpha sequence and the CDR3 beta sequence are identified within the same row of Tables 1-16.

[0062] In a particular embodiment, the substitution is selected according to the following substitution rules, The substitution rules are: - Glycine (G) and alanine (A) are interchangeable; valine (V), leucine (L), and isoleucine (I) are interchangeable, and A and V are interchangeable; - Tryptophan (W) and phenylalanine (F) are interchangeable, and tyrosine (Y) and F are interchangeable; - Serine (S) and threonine (T) are interchangeable; - Aspartic acid (D) and glutamic acid (E) are interchangeable; - Asparagine (N) and glutamine (Q) are interchangeable, N and S are interchangeable, N and D are interchangeable, and E and Q are interchangeable; - Methionine (M) and Q are interchangeable; - Cysteine (C) and A, S are interchangeable; - Proline (P) and G, A are interchangeable; - Arginine (R) and lysine (K) are interchangeable.

[0063] The group of CDR3 sequences can be called a cluster.

[0064] In certain embodiments, the CDR3 sequence is selected from the group of a, b, c, d, e, f, g, h, i, j, k.

[0065] In certain embodiments, the TCR polypeptide further comprises a variable (V) alpha sequence, a joining-constant (JC) alpha sequence, a V beta sequence, and a JC beta sequence, or a sequence having at least 80%, 85%, 90%, 92%, 94%, 96%, 98%, or 99% sequence identity to said sequences, wherein the complete TCR sequence retains its biological activity, wherein, a. For group a, the V alpha sequence is SEQ ID NO: 025, the JC alpha sequence is SEQ ID NO: 026, the V beta sequence is SEQ ID NO: 018, and the JC beta sequence is SEQ ID NO: 019, or b. For group b, the V alpha sequence is SEQ ID NO: 046, the JC alpha sequence is SEQ ID NO: 047, the V beta sequence is SEQ ID NO: 040, and the JC beta sequence is SEQ ID NO: 041, or c. For group c, the V alpha sequence is SEQ ID NO: 068, the JC alpha sequence is SEQ ID NO: 069, the V beta sequence is SEQ ID NO: 061, and the JC beta sequence is SEQ ID NO: 062, or d. For group d, the V alpha sequence is SEQ ID NO: 084, the JC alpha sequence is SEQ ID NO: 085, the V beta sequence is SEQ ID NO: 079, and the JC beta sequence is SEQ ID NO: 080, or e. For group e, the V alpha sequence is SEQ ID NO: 122, the JC alpha sequence is SEQ ID NO: 123, the V beta sequence is SEQ ID NO: 117, and the JC beta sequence is SEQ ID NO: 118, or f. For group f, the V alpha sequence is SEQ ID NO: 148, the JC alpha sequence is SEQ ID NO: 149, the V beta sequence is SEQ ID NO: 142, and the JC beta sequence is SEQ ID NO: 143, or g. For group g, the V alpha sequence is SEQ ID NO: 170, the JC alpha sequence is SEQ ID NO: 171, the V beta sequence is SEQ ID NO: 165, and the JC beta sequence is SEQ ID NO: 166, or h. For group h, the V alpha sequence is SEQ ID NO: 188, the JC alpha sequence is SEQ ID NO: 189, the V beta sequence is SEQ ID NO: 182, and the JC beta sequence is SEQ ID NO: 183, or i. For group i, the V alpha sequence is SEQ ID NO: 206, the JC alpha sequence is SEQ ID NO: 207, the V beta sequence is SEQ ID NO: 200, and the JC beta sequence is SEQ ID NO: 201, or j. For group j, the V alpha sequence is SEQ ID NO: 226, the JC alpha sequence is SEQ ID NO: 227, the V beta sequence is SEQ ID NO: 220, and the JC beta sequence is SEQ ID NO: 221, or k. For the k group, the V alpha sequence is SEQ ID NO: 250, the JC alpha sequence is SEQ ID NO: 251, the V beta sequence is SEQ ID NO: 242, and the JC beta sequence is SEQ ID NO: 243, or l. For the l group, the V alpha sequence is SEQ ID NO: 294, the JC alpha sequence is SEQ ID NO: 295, the V beta sequence is SEQ ID NO: 281, and the JC beta sequence is SEQ ID NO: 282, or m. For the m group, the V alpha sequence is SEQ ID NO: 326, the JC alpha sequence is SEQ ID NO: 327, the V beta sequence is SEQ ID NO: 316, and the JC beta sequence is SEQ ID NO: 317, or n. For the n group, the V alpha sequence is SEQ ID NO: 358, the JC alpha sequence is SEQ ID NO: 359, the V beta sequence is SEQ ID NO: 348, and the JC beta sequence is SEQ ID NO: 349, or o. For the o group, the V alpha sequence is SEQ ID NO: 378, the JC alpha sequence is SEQ ID NO: 379, the V beta sequence is SEQ ID NO: 371, and the JC beta sequence is SEQ ID NO: 372, or p. For the p group, the V alpha sequence is SEQ ID NO: 430, the JC alpha sequence is SEQ ID NO: 431, the V beta sequence is SEQ ID NO: 415, and the JC beta sequence is SEQ ID NO: 416.

[0066] The biological activity of the TCR is determined through the activation of T cells containing the TCR by tumor cells or APCs. If the TCR can still recognize an APC presenting a specific HLA-peptide complex, the TCR maintains its biological activity even if it has a deviant sequence.

[0067] The second aspect of the present invention relates to a nucleic acid sequence encoding the TCR polypeptide according to the first aspect.

[0068] The third aspect of the present invention relates to an isolated autologous T cell containing the TCR polypeptide according to the first aspect. An alternative aspect of the third aspect of the present invention relates to an isolated autologous T cell containing the nucleic acid sequence according to the second aspect.

[0069] In certain embodiments, the isolated autologous T cells are recombinant T cells that recombinantly express the TCR polypeptide.

[0070] A fourth aspect of the invention relates to a TCR polypeptide according to the first aspect for use in the treatment of cancer. An alternative aspect of the fourth aspect of the invention relates to a nucleic acid sequence according to the second aspect for use in the treatment of cancer. An alternative aspect of the fourth aspect of the invention relates to an isolated autologous T cell according to the third aspect for use in the treatment of cancer.

[0071] In certain embodiments of the fourth aspect, the agent of the fourth aspect is administered to a patient characterized by the following HLA types: a. for group a, HLA-A * 02:01; or b. for group b, HLA-B * 08:01 and / or HLA-C * 07:01; or c. for group c, HLA-A * 02:01; or d. for group d, HLA-A * 02:01; or e. for group e, HLA-B * 15:01; or f. for group f, HLA-A * 02:01; or g. for group g, HLA-B * 08:01; or h. for group h, HLA-B * 07:02; or i. for group i, HLA-A * 01:01 and / or HLA-B * 08:01 and / or HLA-C * 07:01; or j. for group j, HLA-A * 02:01; or k. for group k, HLA-A * 02:01.

[0072] In certain embodiments of the fourth aspect, the cancer is a solid tumor. In certain embodiments of the fourth aspect, the cancer is selected from lung cancer, pancreatic cancer, colon cancer, and breast cancer. In certain embodiments of the fourth aspect, the cancer is selected from lung cancer and pancreatic cancer.

[0073] In certain embodiments of the fourth aspect, the cancer is selected from the group consisting of bladder urothelial carcinoma, breast invasive carcinoma, cervical squamous cell carcinoma and endocervical adenocarcinoma, cholangiocarcinoma, colon adenocarcinoma, lymphoid neoplasm diffuse large B-cell lymphoma, esophageal cancer, glioblastoma multiforme, head and neck squamous cell carcinoma, kidney chromophobe, renal papillary cell carcinoma of the kidney, acute myeloid leukemia, low-grade glioma of the brain, lung adenocarcinoma, lung squamous cell carcinoma, mesothelioma, ovarian serous cystadenocarcinoma, pancreatic adenocarcinoma, rectal adenocarcinoma, sarcoma, cutaneous melanoma, gastric adenocarcinoma, testicular germ cell tumor, thyroid cancer, endometrial carcinoma of the uterine body, carcinosarcoma of the uterus.

[0074] In certain embodiments of the fourth aspect, for group a, the cancer is characterized by a mutation in a gene selected from the group consisting of KRAS, EGFR, and / or TP53, and in particular here the mutation in KRAS is a KRAS G12 mutation.

[0075] In certain embodiments of the fourth aspect, for group b, the cancer is characterized by a mutation in a gene selected from the group consisting of KRAS, FGFR, and / or TP53, and in particular here the mutation in KRAS is a KRAS Q61 mutation.

[0076] In certain embodiments of the fourth aspect, for group c, the cancer is characterized by a mutation in a gene selected from the group consisting of KRAS and / or EGFR, and in particular here the mutation in KRAS is a KRAS G12 or KRAS Q61 mutation.

[0077] In certain embodiments of the fourth aspect, for group d, the cancer is characterized by a mutation in a gene selected from the group consisting of KRAS and / or TP53, and in particular here the mutation in KRAS is a KRAS G12 mutation.

[0078] In a particular embodiment of the fourth aspect, the cancer is characterized by a mutation in a gene selected from the group consisting of KRAS, EGFR and / or BRAF for group e, and in particular here the mutation in KRAS is a KRAS G12 or KRAS Q61 mutation.

[0079] In a particular embodiment of the fourth aspect, the cancer is characterized by a mutation in a gene selected from the group consisting of KRAS, EGFR and / or BRAF for group f, and in particular here the mutation in KRAS is a KRAS G12 or KRAS Q61 mutation.

[0080] In a particular embodiment of the fourth aspect, the cancer is characterized by a mutation in the gene TP53 for group g.

[0081] In a particular embodiment of the fourth aspect, the cancer is characterized by a mutation in a gene selected from the group consisting of KRAS, EGFR, BRAF and / or TP53 for group h, and in particular here the mutation in KRAS is a KRAS G12 or KRAS Q61 mutation.

[0082] In a particular embodiment of the fourth aspect, the cancer is characterized by a mutation in a gene selected from the group consisting of EGFR, FGFR and / or TP53 for group i.

[0083] In a particular embodiment of the fourth aspect, the cancer is characterized by a mutation in a gene selected from the group consisting of KRAS, EGFR and / or TP53, and in particular here the mutation in KRAS is a KRAS G12 mutation.

[0084] In a particular embodiment of the fourth aspect, the cancer is characterized by a mutation in a gene selected from the group consisting of KRAS, EGFR, BRAF and / or TP53 for group k, and in particular here the mutation in KRAS is a KRAS G12 or KRAS Q61 mutation.

[0085] In certain embodiments of the fourth aspect, the cancer is characterized by mutations in genes selected from the group consisting of KRAS, EGFR, BRAF, and / or TP53 for group n, and in particular here the mutations in KRAS are KRAS G12 or KRAS Q61 mutations.

[0086] In certain embodiments of the fourth aspect, the cancer is characterized by mutations in genes selected from the group consisting of KRAS, EGFR, BRAF, and / or TP53 for group o, and in particular here the mutations in KRAS are KRAS G12 or KRAS Q61 mutations.

[0087] In certain embodiments, the cancer is characterized by mutations in genes selected from the group of KRAS for group a, and in particular here the mutations in KRAS are KRAS G12 mutations.

[0088] In certain embodiments, the cancer is characterized by mutations in genes selected from the group of TP53 for group b.

[0089] In certain embodiments, the cancer is characterized by mutations in genes selected from the group of KRAS for group d, and in particular here the mutations in KRAS are KRAS G12 mutations.

[0090] In certain embodiments, the cancer is characterized by mutations in genes selected from the group consisting of KRAS and / or EGFR for group e, and in particular here the mutations in KRAS are KRAS G12 mutations.

[0091] In certain embodiments, the cancer is characterized by mutations in genes selected from the group consisting of KRAS and / or EGFR for group f, and in particular here the mutations in KRAS are KRAS G12 or KRAS Q61 mutations.

[0092] In certain embodiments, the cancer is characterized by mutations in genes selected from the group of TP53 for group i.

[0093] In certain embodiments, the cancer is characterized by a mutation in a gene selected from the group consisting of KRAS and / or EGFR, particularly where the KRAS mutation is a KRAS G12 mutation.

[0094] In certain embodiments, the cancer is characterized by a mutation in a gene selected from the group consisting of EGFR and / or TP53 for group k.

[0095] In certain embodiments, the cancer is characterized by a mutation in a gene selected from the group consisting of KRAS, EGFR and / or TP53 for group n, particularly where the KRAS mutation is a KRAS G12 mutation.

[0096] In certain embodiments, the cancer is characterized by a mutation in a gene selected from the group consisting of KRAS, EGFR and / or TP53 for group o, particularly where the KRAS mutation is a KRAS G12 mutation.

[0097] A further aspect of the invention relates to - a TCR polypeptide, - an isolated nucleic acid sequence encoding the TCR polypeptide, or - an isolated autologous T cell comprising the TCR polypeptide and / or the nucleic acid sequence; for use in the treatment of cancer, selected from The TCR polypeptide comprises a CDR3 alpha sequence and a CDR3 beta sequence, the CDR3 alpha sequence and the CDR3 beta sequence are identical to the sequences shown below or have one amino acid substitution per CDR3 sequence, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 007 - 009 or SEQ ID NOs: 436 - 439, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 001 - 006 or SEQ ID NOs: 432 - 435. The TCR polypeptide further comprises a variable (V) alpha sequence, a joining constant (JC) alpha sequence, a V beta sequence, and a JC beta sequence, or a sequence having at least 80%, at least 85%, at least 90%, at least 92%, at least 94%, at least 96%, at least 98%, or at least 99% sequence identity to said sequences, wherein the V alpha sequence is SEQ ID NO: 025, the JC alpha sequence is SEQ ID NO: 026, the V beta sequence is SEQ ID NO: 018, and the JC beta sequence is SEQ ID NO: 019. The agent is HLA-A * administered to a patient characterized by an HLA type of 02:01, and the cancer is characterized by a mutation in a gene of the EGFR-Raf-Ras pathway.

[0098] In certain embodiments of this further aspect, the mutation in the gene of the EGFR-Raf-Ras pathway is a mutation in KRAS and / or EGFR.

[0099] In certain embodiments of this further aspect, the mutation in the gene of the EGFR-Raf-Ras pathway is a KRAS G12 mutation.

[0100] Medical treatment, dosage form, and salt Similarly, within the scope of the present invention, there is a method for treating cancer in a patient in need thereof, comprising administering to the patient a TCR according to the third aspect, a nucleic acid sequence according to the fourth aspect, or an isolated autologous T cell according to the fifth aspect.

[0101] Similarly, within the scope of the present invention, there is provided a formulation for preventing or treating cancer, comprising a TCR according to the third aspect, a nucleic acid sequence according to the fourth aspect, or an isolated autologous T cell according to the fifth aspect.

[0102] Pharmaceutical composition and administration Another aspect of the present invention relates to a pharmaceutical composition comprising a TCR according to the third aspect, a nucleic acid sequence according to the fourth aspect, or an isolated autologous T cell according to the fifth aspect.

[0103] In certain embodiments of the invention, the compounds of the invention are typically formulated into pharmaceutical dosage forms in order to provide dosage amounts that are easily controllable for drugs and to provide a product that is clear and easy to manage for patients.

[0104] The pharmaceutical composition can be formulated for parenteral administration, for example, for intravenous (i.v.) injection.

[0105] Manufacturing methods and treatment methods according to the present invention The present invention further includes, as additional aspects, the use of the TCR according to the third aspect, the nucleic acid sequence according to the fourth aspect, or the isolated autologous T cell according to the fifth aspect, as specifically identified above, for use in the manufacture of a medicament for the treatment or prevention of cancer.

[0106] Similarly, the present invention includes a method for treating a patient diagnosed with a disease associated with cancer. The method involves administering to the patient a TCR according to the third aspect, a nucleic acid sequence according to the fourth aspect, or an isolated autologous T cell according to the fifth aspect.

[0107] This specification further includes the following items:

[0108] Item 1. An isolated TCR polypeptide, wherein the TCR polypeptide comprises a CDR3 alpha sequence and a CDR3 beta sequence, and the CDR3 alpha sequence and the CDR3 beta sequence are identical to the sequences shown below, or have one or two amino acid substitutions per CDR3 sequence, wherein for group a, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 007 - 009 or SEQ ID NOs: 436 - 439, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 001 - 006 or SEQ ID NOs: 432 - 435, The isolated TCR polypeptide.

[0109] Item 2. The TCR polypeptide further includes a variable (V) alpha sequence, a joining-constant (JC) alpha sequence, a V beta sequence, and a JC beta sequence, or a sequence having a sequence identity of ≧80%, ≧85%, ≧90%, ≧92%, ≧94%, ≧96%, ≧98% or ≧99% with respect to these sequences. For group a, the V alpha sequence is SEQ ID NO: 025, the JC alpha sequence is SEQ ID NO: 026, the V beta sequence is SEQ ID NO: 018, and the JC beta sequence is SEQ ID NO: 019. The isolated TCR polypeptide according to Item 1.

[0110] Item 3. An isolated TCR polypeptide, wherein the TCR polypeptide includes a CDR3 alpha sequence and a CDR3 beta sequence, and the CDR3 alpha sequence and the CDR3 beta sequence are identical to the sequences shown below, or have one or two amino acid substitutions per CDR3 sequence. For group b, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 031 to 032, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 027 to 030. The isolated TCR polypeptide.

[0111] Item 4. The TCR polypeptide further includes a variable (V) alpha sequence, a joining-constant (JC) alpha sequence, a V beta sequence, and a JC beta sequence, or a sequence having a sequence identity of ≧80%, ≧85%, ≧90%, ≧92%, ≧94%, ≧96%, ≧98% or ≧99% with respect to these sequences. For group b, the V alpha sequence is SEQ ID NO: 046, the JC alpha sequence is SEQ ID NO: 047, the V beta sequence is SEQ ID NO: 040, and the JC beta sequence is SEQ ID NO: 041. The isolated TCR polypeptide according to Item 3.

[0112] Item 5. An isolated TCR polypeptide, wherein the TCR polypeptide includes a CDR3 alpha sequence and a CDR3 beta sequence, and the CDR3 alpha sequence and the CDR3 beta sequence are identical to the sequences shown below, or have one or two amino acid substitutions per CDR3 sequence. For group c, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 052 to 054, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 048 to 051, said isolated TCR polypeptide.

[0113] Item 6. The TCR polypeptide further comprises a variable (V) alpha sequence, a joining constant (JC) alpha sequence, a V beta sequence and a JC beta sequence, or a sequence having a sequence identity of ≧80%, ≧85%, ≧90%, ≧92%, ≧94%, ≧96%, ≧98%, or ≧99% with respect to said these sequences. For group c, the V alpha sequence is SEQ ID NO: 068, the JC alpha sequence is SEQ ID NO: 069, the V beta sequence is SEQ ID NO: 061, and the JC beta sequence is SEQ ID NO: 062. The isolated TCR polypeptide according to Item 5.

[0114] Item 7. An isolated TCR polypeptide, the TCR polypeptide comprising a CDR3 alpha sequence and a CDR3 beta sequence, the CDR3 alpha sequence and the CDR3 beta sequence being identical to the sequences shown below, or having one or two amino acid substitutions per CDR3 sequence, For group d, the CDR3 alpha sequence is selected from the sequence of SEQ ID NO: 073, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 070 to 072, said isolated TCR polypeptide.

[0115] Item 8. The TCR polypeptide further comprises a variable (V) alpha sequence, a joining constant (JC) alpha sequence, a V beta sequence and a JC beta sequence, or a sequence having a sequence identity of ≧80%, ≧85%, ≧90%, ≧92%, ≧94%, ≧96%, ≧98% or ≧99% with respect to said these sequences. For group d, the V alpha sequence is SEQ ID NO: 084, the JC alpha sequence is SEQ ID NO: 085, the V beta sequence is SEQ ID NO: 079, and the JC beta sequence is SEQ ID NO: 080. The isolated TCR polypeptide according to Item 7.

[0116] Item 9. An isolated TCR polypeptide, wherein the TCR polypeptide comprises a CDR3 alpha sequence and a CDR3 beta sequence, and the CDR3 alpha sequence and the CDR3 beta sequence are identical to the sequences shown below, or have one or two amino acid substitutions per CDR3 sequence, For group e, the CDR3 alpha sequence is selected from the sequences of SEQ ID NO: 100 or SEQ ID NOs: 455-459, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 086-099 or SEQ ID NOs: 450-454, said isolated TCR polypeptide.

[0117] Item 10. The TCR polypeptide further comprises a variable (V) alpha sequence, a joining constant (JC) alpha sequence, a V beta sequence and a JC beta sequence, or a sequence having a sequence identity of ≧80%, ≧85%, ≧90%, ≧92%, ≧94%, ≧96%, ≧98% or ≧99% to said sequences thereof. For group e, the V alpha sequence is SEQ ID NO: 122, the JC alpha sequence is SEQ ID NO: 0123, the V beta sequence is SEQ ID NO: 117, and the JC beta sequence is SEQ ID NO: 118. The isolated TCR polypeptide according to Item 9.

[0118] Item 11. An isolated TCR polypeptide, wherein the TCR polypeptide comprises a CDR3 alpha sequence and a CDR3 beta sequence, and the CDR3 alpha sequence and the CDR3 beta sequence are identical to the sequences shown below, or have one or two amino acid substitutions per CDR3 sequence, For group f, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 131-132 or SEQ ID NOs: 472-473, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 124-130 or SEQ ID NOs: 470-471. Said isolated TCR polypeptide.

[0119] Item 12. The TCR polypeptide further includes a variable (V) alpha sequence, a joining constant (JC) alpha sequence, a V beta sequence, and a JC beta sequence, or a sequence having a sequence identity of ≧80%, ≧85%, ≧90%, ≧92%, ≧94%, ≧96%, ≧98% or ≧99% with respect to these sequences. For group f, the V alpha sequence is SEQ ID NO: 148, the JC alpha sequence is SEQ ID NO: 149, the V beta sequence is SEQ ID NO: 142, and the JC beta sequence is SEQ ID NO: 143. The isolated TCR polypeptide according to Item 11.

[0120] Item 13. An isolated TCR polypeptide, wherein the TCR polypeptide includes a CDR3 alpha sequence and a CDR3 beta sequence, and the CDR3 alpha sequence and the CDR3 beta sequence are identical to the sequences shown below, or have one or two amino acid substitutions per CDR3 sequence. For group g, the CDR3 alpha sequence is selected from the sequences of SEQ ID NO: 156, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 150 to 155. The isolated TCR polypeptide.

[0121] Item 14. The TCR polypeptide further includes a variable (V) alpha sequence, a joining constant (JC) alpha sequence, a V beta sequence, and a JC beta sequence, or a sequence having a sequence identity of ≧80%, ≧85%, ≧90%, ≧92%, ≧94%, ≧96%, ≧98% or ≧99% with respect to these sequences. For group g, the V alpha sequence is SEQ ID NO: 170, the JC alpha sequence is SEQ ID NO: 171, the V beta sequence is SEQ ID NO: 165, and the JC beta sequence is SEQ ID NO: 166. The isolated TCR polypeptide according to Item 13.

[0122] Item 15. An isolated TCR polypeptide, wherein the TCR polypeptide includes a CDR3 alpha sequence and a CDR3 beta sequence, and the CDR3 alpha sequence and the CDR3 beta sequence are identical to the sequences shown below, or have one or two amino acid substitutions per CDR3 sequence. For the h group, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 175 to 176, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 172 to 174, said isolated TCR polypeptide.

[0123] Item 16. The TCR polypeptide further comprises a variable (V) alpha sequence, a joining constant (JC) alpha sequence, a V beta sequence and a JC beta sequence, or a sequence having a sequence identity of ≧80%, ≧85%, ≧90%, ≧92%, ≧94%, ≧96%, ≧98% or ≧99% with respect to said sequences. For the h group, the V alpha sequence is SEQ ID NO: 188, the JC alpha sequence is SEQ ID NO: 189, the V beta sequence is SEQ ID NO: 182, and the JC beta sequence is SEQ ID NO: 183, the isolated TCR polypeptide according to Item 15.

[0124] Item 17. An isolated TCR polypeptide, wherein the TCR polypeptide comprises a CDR3 alpha sequence and a CDR3 beta sequence, and the CDR3 alpha sequence and the CDR3 beta sequence are identical to the sequences shown below, or have one or two amino acid substitutions per CDR3 sequence, For the i group, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 193 to 194, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 190 to 192, said isolated TCR polypeptide.

[0125] Item 18. The TCR polypeptide further comprises a variable (V) alpha sequence, a joining constant (JC) alpha sequence, a V beta sequence and a JC beta sequence, or a sequence having a sequence identity of ≧80%, ≧85%, ≧90%, ≧92%, ≧94%, ≧96%, ≧98% or ≧99% with respect to said sequences. For the i group, the V alpha sequence is SEQ ID NO: 206, the JC alpha sequence is SEQ ID NO: 207, the V beta sequence is SEQ ID NO: 200, and the JC beta sequence is SEQ ID NO: 201, the isolated TCR polypeptide according to Item 17.

[0126] Item 19. An isolated TCR polypeptide, wherein the TCR polypeptide comprises a CDR3 alpha sequence and a CDR3 beta sequence, and the CDR3 alpha sequence and the CDR3 beta sequence are identical to the sequences shown below or have one or two amino acid substitutions per CDR3 sequence, For group j, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 212-213 or SEQ ID NO: 513, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 208-211 or SEQ ID NO: 478, said isolated TCR polypeptide.

[0127] Item 20. The TCR polypeptide further comprises a variable (V) alpha sequence, a joining constant (JC) alpha sequence, a V beta sequence, and a JC beta sequence, or a sequence having a sequence identity of ≥ 80%, ≥ 85%, ≥ 90%, ≥ 92%, ≥ 94%, ≥ 96%, ≥ 98% or ≥ 99% to said sequences thereof. For group j, the V alpha sequence is SEQ ID NO: 226, the JC alpha sequence is SEQ ID NO: 227, the V beta sequence is SEQ ID NO: 220, and the JC beta sequence is SEQ ID NO: 221, the isolated TCR polypeptide according to Item 19.

[0128] Item 21. An isolated TCR polypeptide, wherein the TCR polypeptide comprises a CDR3 alpha sequence and a CDR3 beta sequence, and the CDR3 alpha sequence and the CDR3 beta sequence are identical to the sequences shown below or have one or two amino acid substitutions per CDR3 sequence, For group k, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 232-235, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 228-231, said isolated TCR polypeptide.

[0129] Item 22. The TCR polypeptide further comprises a variable (V) alpha sequence, a joining constant (JC) alpha sequence, a V beta sequence, and a JC beta sequence, or a sequence having a sequence identity of ≧80%, ≧85%, ≧90%, ≧92%, ≧94%, ≧96%, ≧98%, or ≧99% with respect to these sequences. For group k, the V alpha sequence is SEQ ID NO: 250, the JC alpha sequence is SEQ ID NO: 251, the V beta sequence is SEQ ID NO: 242, and the JC beta sequence is SEQ ID NO: 243. The isolated TCR polypeptide according to Item 21.

[0130] Item 23. An isolated TCR polypeptide, wherein the TCR polypeptide comprises a CDR3 alpha sequence and a CDR3 beta sequence, and the CDR3 alpha sequence and the CDR3 beta sequence are identical to the sequences shown below, or have one or two amino acid substitutions per CDR3 sequence. For group l, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 261 to 269 or SEQ ID NOs: 484 to 486, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 252 to 260 or SEQ ID NOs: 481 to 483. The isolated TCR polypeptide.

[0131] Item 24. The TCR polypeptide further comprises a variable (V) alpha sequence, a joining constant (JC) alpha sequence, a V beta sequence, and a JC beta sequence, or a sequence having a sequence identity of ≧80%, ≧85%, ≧90%, ≧92%, ≧94%, ≧96%, ≧98%, or ≧99% with respect to these sequences. For group l, the V alpha sequence is SEQ ID NO: 294, the JC alpha sequence is SEQ ID NO: 295, the V beta sequence is SEQ ID NO: 281, and the JC beta sequence is SEQ ID NO: 282. The isolated TCR polypeptide according to Item 23.

[0132] Item 25. An isolated TCR polypeptide, wherein the TCR polypeptide comprises a CDR3 alpha sequence and a CDR3 beta sequence, and the CDR3 alpha sequence and the CDR3 beta sequence are identical to the sequences shown below, or have one or two amino acid substitutions per CDR3 sequence, For group m, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 302 - 307 or SEQ ID NOs: 496 - 498, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 296 - 301 or SEQ ID NOs: 493 - 495, The isolated TCR polypeptide.

[0133] Item 26. The TCR polypeptide further comprises a variable (V) alpha sequence, a joining constant (JC) alpha sequence, a V beta sequence, and a JC beta sequence, or a sequence having at least 80%, at least 85%, at least 90%, at least 92%, at least 94%, at least 96%, at least 98% or at least 99% sequence identity to said sequences thereof. For group m, the V alpha sequence is SEQ ID NO: 326, the JC alpha sequence is SEQ ID NO: 327, the V beta sequence is SEQ ID NO: 316, and the JC beta sequence is SEQ ID NO: 317. The isolated TCR polypeptide according to Item 25.

[0134] Item 27. An isolated TCR polypeptide, wherein the TCR polypeptide comprises a CDR3 alpha sequence and a CDR3 beta sequence, and the CDR3 alpha sequence and the CDR3 beta sequence are identical to the sequences shown below, or have one or two amino acid substitutions per CDR3 sequence, For group n, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 334 - 339, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 328 - 333. The isolated TCR polypeptide.

[0135] Item 28. The TCR polypeptide further comprises a variable (V) alpha sequence, a joining constant (JC) alpha sequence, a V beta sequence, and a JC beta sequence, or a sequence having a sequence identity of ≧80%, ≧85%, ≧90%, ≧92%, ≧94%, ≧96%, ≧98% or ≧99% with respect to these sequences. For group n, the V alpha sequence is SEQ ID NO: 358, the JC alpha sequence is SEQ ID NO: 359, the V beta sequence is SEQ ID NO: 348, and the JC beta sequence is SEQ ID NO: 349. The isolated TCR polypeptide according to Item 27.

[0136] Item 29. An isolated TCR polypeptide, wherein the TCR polypeptide comprises a CDR3 alpha sequence and a CDR3 beta sequence, and the CDR3 alpha sequence and the CDR3 beta sequence are identical to the sequences shown below, or have one or two amino acid substitutions per CDR3 sequence. For group o, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 363 to 365, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 360 to 362. The isolated TCR polypeptide.

[0137] Item 30. The TCR polypeptide further comprises a variable (V) alpha sequence, a joining constant (JC) alpha sequence, a V beta sequence, and a JC beta sequence, or a sequence having a sequence identity of ≧80%, ≧85%, ≧90%, ≧92%, ≧94%, ≧96%, ≧98% or ≧99% with respect to these sequences. For group o, the V alpha sequence is SEQ ID NO: 378, the JC alpha sequence is SEQ ID NO: 379, the V beta sequence is SEQ ID NO: 371, and the JC beta sequence is SEQ ID NO: 372. The isolated TCR polypeptide according to Item 29.

[0138] Item 31. An isolated TCR polypeptide, wherein the TCR polypeptide comprises a CDR3 alpha sequence and a CDR3 beta sequence, and the CDR3 alpha sequence and the CDR3 beta sequence are identical to the sequences shown below, or have one or two amino acid substitutions per CDR3 sequence. For the p group, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 391 to 401 or SEQ ID NOs: 507 to 508, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 380 to 390 or SEQ ID NOs: 505 to 506. The isolated TCR polypeptide.

[0139] Item 32. The TCR polypeptide further includes a variable (V) alpha sequence, a joining constant (JC) alpha sequence, a V beta sequence, and a JC beta sequence, or a sequence having at least 80%, at least 85%, at least 90%, at least 92%, at least 94%, at least 96%, at least 98%, or at least 99% sequence identity to said sequences. For the p group, the V alpha sequence is SEQ ID NO: 430, the JC alpha sequence is SEQ ID NO: 431, the V beta sequence is SEQ ID NO: 415, and the JC beta sequence is SEQ ID NO: 416. The isolated TCR polypeptide according to item 29.

[0140] Item 33. The TCR polypeptide includes a CDR3 alpha sequence and a CDR3 beta sequence, and the CDR3 alpha sequence and the CDR3 beta sequence are identical to the shown sequences or have one amino acid substitution per CDR3 sequence. The isolated TCR polypeptide according to any one of items 1 to 32.

[0141] Item 34. The TCR polypeptide includes a CDR3 alpha sequence and a CDR3 beta sequence, and the CDR3 alpha sequence and the CDR3 beta sequence are identical to the shown sequences without any amino acid substitution. The isolated TCR polypeptide according to any one of items 1 to 32.

[0142] Item 35. The CDR3 alpha sequence and the CDR3 beta sequence are identified within the same row of Tables 1 to 16. The isolated TCR polypeptide according to any one of items 1 to 34.

[0143] Item 36. The substitution is selected according to the substitution rules shown below. The substitution rules are: - Glycine (G) and alanine (A) are interchangeable; valine (V), leucine (L), and isoleucine (I) are interchangeable, and A and V are interchangeable; - Tryptophan (W) and phenylalanine (F) are interchangeable, and tyrosine (Y) and F are interchangeable; - Serine (S) and threonine (T) are interchangeable; - Aspartic acid (D) and glutamic acid (E) are interchangeable; - Asparagine (N) and glutamine (Q) are interchangeable, N and S are interchangeable, N and D are interchangeable, and E and Q are interchangeable; - Methionine (M) and Q are interchangeable; - Cysteine (C) is interchangeable with A and S; - Proline (P) is interchangeable with G and A; - Arginine (R) and lysine (K) are interchangeable. An isolated TCR polypeptide according to any one of items 1 to 35.

[0144] Item 37. A library of TCR polypeptides, comprising at least two TCR polypeptides from different clusters a to p as described in any one of items 1 to 36.

[0145] Item 38. An isolated nucleic acid sequence encoding a TCR polypeptide according to any one of items 1 to 36.

[0146] Item 39. A library of isolated nucleic acid sequences encoding TCR polypeptides, comprising at least two isolated nucleic acid sequences encoding TCR polypeptides from different clusters a to p as described in any one of items 1 to 36.

[0147] Item 40. An isolated autologous T cell comprising the TCR polypeptide according to any one of Items 1 to 36 and / or the nucleic acid sequence according to Item 38.

[0148] Item 41. The isolated autologous T cell according to Item 40, wherein the isolated autologous T cell is a recombinant T cell.

[0149] Item 42. A library of isolated autologous T cells comprising the TCR polypeptide, the library comprising at least two isolated autologous T cells each comprising a TCR polypeptide from different clusters a to p as described in any one of Items 1 to 36.

[0150] Item 43. A medicament for use in the treatment of cancer, selected from the TCR polypeptide according to any one of Items 1 to 36, the nucleic acid sequence according to Item 38, or the isolated autologous T cell according to Item 40 or 41.

[0151] Item 44. The medicament for use according to Item 43, wherein the medicament is administered to a patient characterized by the following HLA types: a. For group a, HLA-A * 02:01; or b. For group b, HLA-B * 08:01 and / or HLA-C * 07:01; or c. For group c, HLA-A * 02:01; or d. For group d, HLA-A * 02:01; or e. For group e, HLA-B * 15:01; or f. For group f, HLA-A * 02:01; or g. For group g, HLA-B * 08:01; or h. For group h, HLA-B * 07:02; or i. For group i, HLA-A *01:01 and / or HLA-B * 08:01 and / or HLA-C * 07:01; or j. For group j, HLA-A * 02:01; or k. For group k, HLA-A * 02:01.

[0152] Item 45. The cancer is a solid tumor, and the drug for the use according to any one of Items 43 or 44.

[0153] Item 46. The cancer is selected from lung cancer, pancreatic cancer, colon cancer, and breast cancer, and the drug for the use according to any one of Items 43 or 44.

[0154] Item 47. The cancer is selected from the group consisting of bladder urothelial carcinoma, breast invasive carcinoma, cervical squamous cell carcinoma and cervical endometrial adenocarcinoma, cholangiocarcinoma, colon adenocarcinoma, lymphoid neoplasm diffuse large B-cell lymphoma, esophageal cancer, glioblastoma multiforme, head and neck squamous cell carcinoma, chromophobe renal cell carcinoma, renal papillary cell carcinoma, acute myeloid leukemia, low-grade glioma of the brain, lung adenocarcinoma, lung squamous cell carcinoma, mesothelioma, ovarian serous cystadenocarcinoma, pancreatic adenocarcinoma, rectal adenocarcinoma, sarcoma, cutaneous melanoma, gastric adenocarcinoma, testicular germ cell tumor, thyroid cancer, endometrial carcinoma of the uterine body, uterine carcinosarcoma, and the drug for the use according to any one of Items 43 or 44.

[0155] Item 48. The cancer is the following group: a. For group a, KRAS, EGFR and / or TP53, especially where the mutation of KRAS is the KRAS G12 mutation; or b. For group b, KRAS, FGFR and / or TP53, especially where the mutation of KRAS is the KRAS Q61 mutation; or c. For group c, KRAS and / or EGFR, especially where the mutation of KRAS is the KRAS G12 mutation or the KRAS Q61 mutation; or d. For group d, KRAS and / or TP53, especially where the mutation of KRAS is the KRAS G12 mutation; or e. For group e, it is KRAS, EGFR and / or BRAF, especially where the mutation of KRAS is a KRAS G12 or KRAS Q61 mutation; or f. For group f, it is KRAS, EGFR and / or BRAF, especially where the mutation of KRAS is a KRAS G12 or KRAS Q61 mutation; or g. For group g, it is TP53; or h. For group h, it is KRAS, EGFR, BRAF and / or TP53, especially where the mutation of KRAS is a KRAS G12 or KRAS Q61 mutation; or i. For group i, it is EGFR, FGFR and / or TP53; or j. It is KRAS, EGFR and / or TP53, especially where the mutation of KRAS is a KRAS G12 mutation; or k. For group k, it is KRAS, EGFR, BRAF and / or TP53, especially where the mutation of KRAS is a KRAS G12 or KRAS Q61 mutation; or n. For group n, it is KRAS, EGFR, BRAF and / or TP53, especially where the mutation of KRAS is a KRAS G12 or KRAS Q61 mutation; or o. For group o, it is KRAS, EGFR, BRAF and / or TP53, especially where the mutation of KRAS is a KRAS G12 or KRAS Q61 mutation A medicament for use according to any one of items 43 to 47, characterized by a mutation of a gene selected from

[0156] Item 49. The cancer is the following groups: a. For group a, it is KRAS, especially where the mutation of KRAS is a KRAS G12 mutation; or b. For group b, it is TP53; or d. For group d, it is KRAS, especially where the mutation of KRAS is a KRAS G12 mutation; or e. The e group is KRAS and / or EGFR, particularly where the mutation of KRAS is a KRAS G12 mutation; or f. The f group is KRAS and / or EGFR, particularly where the mutation of KRAS is a KRAS G12 or KRAS Q61 mutation; or i. The i group is TP53; or j. KRAS and / or EGFR, particularly where the mutation of KRAS is a KRAS G12 mutation; or k. The k group is EGFR and / or TP53; or n. The n group is KRAS, EGFR and / or TP53, particularly where the mutation of KRAS is a KRAS G12 mutation; or o. The o group is KRAS, EGFR and / or TP53, particularly where the mutation of KRAS is a KRAS G12 mutation A medicament for use according to any one of items 7 to 9, characterized by a mutation of a gene selected from

[0157] The present invention is further illustrated by the following examples and figures, from which further embodiments and advantages can be derived. These examples are for illustrative purposes only and do not limit the scope of the invention.

Brief Description of Drawings

[0158]

Figure 1

Figure 2

Figure 3

Figure 4

Figure 5

Example

[0159] A.1: Preparation of non-small cell lung cancer (NSCLC) tumor and normal lung tissue specimens and TCRSafe analysis Each tumor specimen is dissected free of surrounding normal tissue and necrotic areas. Cubes of approximately 1 g from tumor and normal lung tissue are cut into small pieces with each dimension approximately 2 - 3 mm. Sliced tumor (and non-tumor) biopsies are subjected to a commercially available mechanical / enzymatic tissue dissociation system (GentleMACS, Miltenyi Biotec, Bergisch Gladbach, Germany) using the Tumor Dissociation Kit (Miltenyi Biotech) according to the manufacturer's instructions. After GentleMACS dissociation, the cell suspension is passed through a 70 μm cell strainer. Aliquots of tumor cells and lung cells are taken and cryopreserved in 10% DMSO (Sigma-Aldrich) and 90% FCS (Life Technologies) for later use. The remaining cell suspension is subjected to density gradient centrifugation using a 40% / 80% step gradient of Percoll® (GE Healthcare Europe GmbH) in PBS / RPMI 1640. T lymphocytes are harvested from the interphase and washed in complete medium (RPMI 1640, Lonza). Tumor-infiltrating T lymphocytes (TIL) and lymphocytes from normal lung tissue are then placed in a 24-well tissue culture plate containing 2 mL of recovery medium (RM) at 0.5x10 6Add at a concentration of cells / ml. RM is RPMI 1640 supplemented with 25 mM HEPES pH 7.2 and L-glutamine (Lonza), 100 IU / mL penicillin, 100 mg / mL streptomycin and 50 mM β-mercaptoethanol (ThermoFisher Scientific, Waltham, Massachusetts, USA), and 10% autologous human serum. Place the plates in a 5% CO 2 , humidified incubator at 37 °C and culture overnight. The next day, collect and pool cells from the TIL and normal lung cultures and isolate the following subpopulations by FACS: · CD4+ T cells · CD8+ T cells · PD1+ cells (from TIL only) · PD1-negative cells (from TIL only)

[0160] Extract genomic DNA from the subpopulations and subject it to TCRsafe analysis (as disclosed in WO2014 / 096394A1). The resulting T cell clonotype frequencies are compared between the identified subpopulations and tumor-specific clonotypes as detailed in WO2017 / 025564A1.

[0161] All subsequent steps in these examples refer to CD8+ T cells isolated from tumor and non-tumor tissues as described above.

[0162] Patient IDs that are numbers following "P" relate to non-small cell lung cancer patients. Patient IDs that are numbers following "PANC" relate to pancreatic cancer patients.

[0163] A.2: T cell receptor (TCR) α / β pairing using 10x Genomics' high-throughput single cell sequencing Start with a TIL single cell suspension and use the 10x Genomics Chromium Next GEM Single Cell V(D)J Reagent Kit in combination with the Chromium Single Cell V(D)J Enrichment Kit (human) to subject 5000 - 10000 T cells to high-throughput single cell RNA Seq analysis. The 10x Genomics® GemCodeTM Technology disperses thousands of individual cells into Gel Bead-in-Emulsion (GEM) droplets. The single cells captured in GEMs are lysed, and during GEM lysis, the barcoded primers, oligos, master mix, and lysed cell components attached to the beads are mixed to generate a full-length oligo-dT primed cDNA library by RT-PCR. Single-stranded cDNA synthesis using a template-switching mechanism is completed, which includes the barcoded sequences attached to the beads. All cDNA molecules within a single GEM are labeled with the same barcode. The GEMs are disassembled, and further library preparation is continued as a bulk reaction. After cDNA cleanup, the Chromium Single Cell V(D)J Enrichment Kit efficiently amplifies the TCR sequences to generate a sequencing library compatible with Illumina sequencing. In combination with whole cDNA amplification, Illumina sequencing reveals the paired α / β TCR sequences and the corresponding full transcriptome for each single T cell analyzed. Both kits, the 10x Genomics Chromium Next GEM Single Cell V(D)J Reagent Kit and the Chromium Single Cell V(D)J Enrichment Kit (human), are used according to the manufacturer's recommendations.

[0164] The same nucleotide sequences between the TCR described in A.1 above and the TCRs derived from single cell VDJ pairings are used to establish a complete annotation of the TCRs regarding the α and β chains, frequency, and tumor specificity.

[0165] A.3: Clustering of TCRs (TCRpolyClust) As described in the schematic diagram of the TCRpolyClust method, once the combination of A.1 and A.2 is established, subsequent TCR cluster analysis can identify patients with common TCRs and matching HLA types, enabling the screening of shared tumor antigens.

[0166] Correlation between known tumor driver mutations and common tumor-specific TCRs There is a set of well-known tumor mutations (such as KRAS, EGFR, etc.), which are routinely screened by the use of sequencing technologies and available primer tumor panels (such as QIAseq Targeted DNA Panel, AmpliSeq for Illumina Focus Panel). Under this set of tumor mutations, little success has been achieved in finding general (public) antigens of tumors so far, but there is strong evidence that the presence of common tumor-specific TCRs is highly correlated with the occurrence of dedicated frequent tumor mutations. Therefore, if patients belong to their respective HLA types and are carriers of their respective mutations, it is possible to sufficiently select patients for TCR-T therapy via common TCRs.

[0167] As shown in Fig. 5, the TCR-T cells transfected with the three TCRs of cluster a showed almost the same response pattern: spontaneous IFN-γ secretion (by TCR-T cells only) was low to moderate (separated by dashed lines), but in response to NSCLC cell lines NCI-H1792 and MZ-LC-16, all three TCR-T cells secreted large amounts of IFN-γ specifically (calculated as the number of IFN-γ spot-forming cells per TCR-T cell per test reaction). No IFN-γ response was detected in co-culture with tumor cell lines NCI-H661 and MOR / CPR. From NGS analysis of the tumor mutation profiles of the cell lines, it was revealed that the only mutation commonly found in cell lines NCI-H1792 and MZ-LC-16 was the oncogenic driver mutation KRAS p.G12C. This mutation was not seen in the two tumor lines not recognized by the TCR-T cells. Surprisingly, in cluster a of TCRs, the tumors of 7 out of 9 patients tested were positive for different KRAS p.G12 mutations (Table 17).

[0168] The frequencies of common mutations along most of the TCR clusters a - o are summarized in Table 17, and the respective frequencies of these mutations in all relevant cancers are shown in Tables 18 and 19. The numbers in Table 17 were obtained from mutation screening (oncopanel) of each patient's tumor.

[0169] Materials and Methods T lymphocytes isolated from the meninges of healthy donors were removed from the endogenous TCR by CRISPR / CAS9 gene knockout (KO). Then, three recombinant TCRs from TCR cluster a were introduced into these T cells by retroviral transduction. After in vitro proliferation and confirmation (by FACS) of the expression of the recombinant TCRs on the cell surface, the resulting TCR-T cells were tested for recognition of HLA-A02-positive NSCLC cell lines MOR / CPR, NCI-H1792, NCI-H661, MZ-LC-16. In patients of cluster a, HLA-A *To show the common expression at 02:01, the inventors focused on the recognition of HLA-A02 positive tumor cells. To test whether TCR-T cells secrete IFN-γ in response to co-culture with these cell lines, a 96-well format IFN-γ ELISpot assay was performed. The spontaneous IFN-γ release by TCR-T cells served as a background control. All reactions were performed in duplicate. The results (Figure 5) are shown as the average value of the number of IFN-γ spot-producing cells for each duplicate.

[0170] Synthesis, cloning, and ectopic expression of clustered TCRs in T cells isolated from autologous patients or healthy donors The paired clustered TCRs are codon-optimized, synthesized, and cloned into a retroviral (or comparable) expression vector as a 2-cistronic chimeric construct (βTCR-VDJ-mC_P2A-element_αTCR-VJ-mC; mC represents the murine constant domain) and transduced into autologous T cells or allogeneic T cells collected from the blood of each patient or healthy donor. The recipient T cells are pre-treated with CRISPR / Cas9 to knock out the endogenous TCR, preventing off-target immune responses mediated by mixed TCR dimers (endogenous × exogenous chains in autologous and allogeneic settings) or allo-responses by the endogenous TCR (in allogeneic settings). The chimeric (c)TCR recombinant T cells are expanded in vitro and applied to functional experiments such as the recognition of autologous tumor cells (if available), allogeneic tumor cell lines, and / or antigen screening as described below.

[0171] Targeted approach for screening shared tumor antigens Apply comparative sequencing of whole-exome (WES) and whole transcriptome (WTS) of genomic and total RNA of tumor tissue and corresponding normal tissue, including samples from all patients of each TCR cluster, to identify shared neoantigens (SNV, MNV, InDel, fusion gene products, structural variations), abnormally expressed canonical genes (cancer / germline antigens and overexpressed antigens), and abnormally expressed and translated non-canonical transcripts (dark matter transcripts or cryptic transcripts). Then, test all categories of candidates for recognition by cTCR-introduced recombinant T cells. The antigen format is either an expression plasmid encoding the full-length antigen cDNA or a tandem minigene (TMG) encoding only the peptide coding region where immunogenicity of the candidate antigen is expected. Both formats are tested by co-transfecting the plasmid encoding the antigen and the plasmid encoding HLA-cDNA into 293T cells or COS-7 cells and subjecting the transformants to a recognition test by T cells in an IFN-γ ELISpot assay. Alternatively, using publicly available prediction algorithms (IEDB, NetMHC), it is possible to predict that the antigen peptide candidate binds to the relevant HLA allele, synthesize the peptide, and pulse the antigen-presenting cells with a matching HLA. The latter is then subjected to an ELISpot assay to test for recognition by recombinant T cells.

[0172] Screening approach for tumor cDNA expression library Target identification of antigen candidates is not equally effective for all antigen categories. For example, screening for nonsynonymous somatic mutations in tumor cells using whole exome and whole transcriptome sequencing can be highly sensitive and reproducible, and can provide a quantitative list of potential neoantigens, but the identification of cryptic translatable transcripts (dark matter antigens) is less effective because specific traits for their reliable identification are generally lacking. This dilemma can be resolved by probing the complete transcriptome of tumor cells by a cDNA expression library screening approach. A cDNA expression library generated from total RNA is co-expressed with the appropriate HLA allele of antigen-presenting cells (293T cells or COS-7 cells) from either an HLA-matched tumor cell line or sorted autologous tumor cells shown prior to recognition by cTCR-transduced T cells. The transformants are then tested for recognition by cTCR-transduced T cells in an ELISpot assay. To obtain the opportunity to express even rare transcripts after transfection, the screening procedure requires a high-throughput approach to test highly fractionated cDNA libraries. For this purpose, the cDNA library is prepared to consist of 2000 pools of 100 cDNAs per well prepared in, for example, a 96-well plate format. Transfection and ELISpot assays using cTCR-transduced T cells as effector cells are performed in this 96-well format, and by progressively reducing and testing the pools from the 100 pools recognized (e.g., 10 cDNAs / pool and well, cDNA clones / pool and well), cDNA clones encoding the antigen are selected.

[0173] For the isolation of RNA and the preparation of cDNA libraries, since it is limited to only a few patients to obtain a sufficiently pure and highly sufficient number of viable autologous tumor cell populations, a pre-screening can be performed to identify tumor cell lines with a matching type. It is possible to either select the cell lines for the expression of shared HLA alleles or transduce them with the HLA of interest. The recognized cell lines are used as proof of the presence of common antigens and as a source for RNA extraction and cDNA library generation.

[0174] Table 17: For each cluster (a - o), the table outlines the total number of patients (A) within each cluster and the number of patients (B) with tumors analyzed for recurrent mutations. Six genes with recurrent mutations in tumors were detected, and columns C - H indicate how frequently the mutations in these genes were found. The mutated genes in columns C - G are involved in the same signaling pathway, and it can be predicted that the mutations have overlapping effects.

Chem.

[0175] Table 18: Summary of the most frequent known tumor mutations published in TCGA. The percentages indicate each cancer type in which the mutation was found. The abbreviations of the cancer types are explained in Table 19.

Chem.

[0176]

Chem.

[0177] Sequence The TCR sequence is constructed as follows (from the N - terminus to the C - terminus):

Chem.

[0178]

Table 1

[0179]

Chem.

[0180]

Table 2

[0181]

Chem.

[0182]

Table 3

[0183]

Chem.

[0184]

Table 4

[0185]

Chem.

[0186]

Table 5

[0187]

Chem.

[0188]

Table 6

[0189]

Chem.

[0190]

Table 7

[0191]

Chem.

[0192]

Table 8

[0193]

Chem.

[0194]

Table 9

[0195]

Chem.

[0196]

Table 10

[0197]

Chem.

[0198]

Table 11

[0199]

Chem.

[0200]

Table 12

[0201]

Chem.

[0202]

Table 13

[0203]

Chem.

[0204]

Table 14

[0205]

Chem.

[0206]

Table 15

[0207]

Chem.

[0208]

Table 16

[0209]

Chem.

Claims

Claim 1 An isolated TCR polypeptide, wherein the TCR polypeptide comprises a CDR3 alpha sequence and a CDR3 beta sequence, and the CDR3 alpha sequence and the CDR3 beta sequence are identical to the sequences shown below, or have one or two amino acid substitutions per CDR3 sequence, where a. For group a, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 007-009 or SEQ ID NOs: 436-439, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 001-006 or SEQ ID NOs: 432-435, or b. For group b, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 031-032, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 027-030, or c. For group c, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 052-054, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 048-051, or d. For group d, the CDR3 alpha sequence is selected from the sequence of SEQ ID NO: 073, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 070-072, or e. For group e, the CDR3 alpha sequence is selected from the sequences of SEQ ID NO: 100 or SEQ ID NOs: 455-459, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 086-099 or SEQ ID NOs: 450-454, or f. For group f, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 131-132 or SEQ ID NOs: 472-473, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 124-130 or SEQ ID NOs: 470-471, or g. For group g, the CDR3 alpha sequence is selected from the sequence of SEQ ID NO: 156, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 150-155, or h. For group h, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 175-176, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 172-174, or i. For group i, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 193-194, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 190-192, or j. For group j, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 212-213 or SEQ ID NO: 513, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 208-211 or SEQ ID NO: 478, or k. For the k group, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 232 to 235, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 228 to 231, or l. For the l group, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 261 to 269 or SEQ ID NOs: 484 to 486, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 252 to 260 or SEQ ID NOs: 481 to 483, or m. For the m group, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 302 to 307 or SEQ ID NOs: 496 to 498, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 296 to 301 or SEQ ID NOs: 493 to 495, or n. For the n group, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 334 to 339, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 328 to 333, or o. For the o group, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 363 to 365, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 360 to 362, or p. For the p group, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 391 to 401 or SEQ ID NOs: 507 to 508, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 380 to 390 or SEQ ID NOs: 505 to 506, In particular, the CRD3 alpha sequence and the CDR3 beta sequence are identified within the same row of Tables 1 to 16, In particular, the substitution is selected according to the following substitution rules, The substitution rules are: - Glycine (G) and alanine (A) are interchangeable; valine (V), leucine (L), and isoleucine (I) are interchangeable, and A and V are interchangeable; - Tryptophan (W) and phenylalanine (F) are interchangeable, and tyrosine (Y) and F are interchangeable; - Serine (S) and threonine (T) are interchangeable; - Aspartic acid (D) and glutamic acid (E) are interchangeable; - Asparagine (N) and glutamine (Q) are interchangeable; N and S are interchangeable; N and D are interchangeable; E and Q are interchangeable; - Methionine (M) and Q are interchangeable; - Cysteine (C) and A and S are interchangeable; - Proline (P) and G and A are interchangeable; - Arginine (R) and lysine (K) are interchangeable, The isolated TCR polypeptide. **Claim 2** The isolated TCR polypeptide according to claim 1, wherein the CDR3 sequence is selected from the group consisting of a, b, c, d, e, f, g, h, i, j, k. **Claim 3** The TCR polypeptide further comprises a variable (V) alpha sequence, a joining constant (JC) alpha sequence, a V beta sequence and a JC beta sequence, or a sequence having a sequence identity of ≧80%, ≧85%, ≧90%, ≧92%, ≧94%, ≧96%, ≧98%, or ≧99% with respect to the above sequences, where a. For group a, the V alpha sequence is SEQ ID NO: 025, the JC alpha sequence is SEQ ID NO: 026, the V beta sequence is SEQ ID NO: 018, and the JC beta sequence is SEQ ID NO: 019, or b. For group b, the V alpha sequence is SEQ ID NO: 046, the JC alpha sequence is SEQ ID NO: 047, the V beta sequence is SEQ ID NO: 040, and the JC beta sequence is SEQ ID NO: 041, or c. For group c, the V alpha sequence is SEQ ID NO: 068, the JC alpha sequence is SEQ ID NO: 069, the V beta sequence is SEQ ID NO: 061, and the JC beta sequence is SEQ ID NO: 062, or d. For group d, the V alpha sequence is SEQ ID NO: 084, the JC alpha sequence is SEQ ID NO: 085, the V beta sequence is SEQ ID NO: 079, and the JC beta sequence is SEQ ID NO: 080, or e. For group e, the V alpha sequence is SEQ ID NO: 122, the JC alpha sequence is SEQ ID NO: 0123, the V beta sequence is SEQ ID NO: 117, and the JC beta sequence is SEQ ID NO: 118, or f. For group f, the V alpha sequence is SEQ ID NO: 148, the JC alpha sequence is SEQ ID NO: 149, the V beta sequence is SEQ ID NO: 142, and the JC beta sequence is SEQ ID NO: 143, or g. For group g, the V alpha sequence is SEQ ID NO: 170, the JC alpha sequence is SEQ ID NO: 171, the V beta sequence is SEQ ID NO: 165, and the JC beta sequence is SEQ ID NO: 166, or h. For group h, the V alpha sequence is SEQ ID NO: 188, the JC alpha sequence is SEQ ID NO: 189, the V beta sequence is SEQ ID NO: 182, and the JC beta sequence is SEQ ID NO: 183, or i. For group i, the V alpha sequence is SEQ ID NO: 206, the JC alpha sequence is SEQ ID NO: 207, the V beta sequence is SEQ ID NO: 200, and the JC beta sequence is SEQ ID NO: 201, or j. For group j, the V alpha sequence is SEQ ID NO: 226, the JC alpha sequence is SEQ ID NO: 227, the V beta sequence is SEQ ID NO: 220, and the JC beta sequence is SEQ ID NO: 221, or k. For group k, the V alpha sequence is SEQ ID NO: 250, the JC alpha sequence is SEQ ID NO: 251, the V beta sequence is SEQ ID NO: 242, and the JC beta sequence is SEQ ID NO: 243, or l. For group l, the V alpha sequence is SEQ ID NO: 294, the JC alpha sequence is SEQ ID NO: 295, the V beta sequence is SEQ ID NO: 281, and the JC beta sequence is SEQ ID NO: 282, or m. For group m, the V alpha sequence is SEQ ID NO: 326, the JC alpha sequence is SEQ ID NO: 327, the V beta sequence is SEQ ID NO: 316, and the JC beta sequence is SEQ ID NO: 317, or n. For group n, the V alpha sequence is SEQ ID NO: 358, the JC alpha sequence is SEQ ID NO: 359, the V beta sequence is SEQ ID NO: 348, and the JC beta sequence is SEQ ID NO: 349, or o. For group o, the V alpha sequence is SEQ ID NO: 378, the JC alpha sequence is SEQ ID NO: 379, the V beta sequence is SEQ ID NO: 371, and the JC beta sequence is SEQ ID NO: 372, or p. For group p, the V alpha sequence is SEQ ID NO: 430, the JC alpha sequence is SEQ ID NO: 431, the V beta sequence is SEQ ID NO: 415, and the JC beta sequence is SEQ ID NO: 416 An isolated TCR polypeptide according to claim 1 or 2.

4. An isolated nucleic acid sequence encoding a TCR polypeptide according to any one of claims 1 to 3.

5. An isolated autologous T cell comprising a TCR polypeptide according to any one of claims 1 to 3 and / or a nucleic acid sequence according to claim 4.

6. The isolated autologous T cell according to claim 5, wherein the isolated autologous T cell is a recombinant T cell.

7. An agent for use in the treatment of cancer, selected from the TCR polypeptide according to any one of claims 1 to 3, the nucleic acid sequence according to claim 4, or the isolated autologous T cells according to claim 5 or 6.

8. The agent is administered to a patient characterized by the following HLA types: a. For group a, HLA-A ＊ 02:01; or b. For group B, HLA-B ＊ 08:01 and / or HLA-C ＊ 07:01; or c. For group C, HLA-A ＊ 02:01; or d. For the d group, HLA-A ＊ 02:01; or e. For the e group, HLA-B ＊ 15:01; or For the f group, HLA-A ＊ 02:01; or g. For group g, HLA-B ＊ 08:01; or h. For the h group, HLA-B ＊ 07:02; or i. For group i, HLA-A ＊ 01:01 and / or HLA-B ＊ 08:01 and / or HLA-C ＊ 07:01; or For group j, HLA-A ＊ 02:01; or k. For the k group, HLA-A ＊ 02:01 The agent for use according to claim 7, which is administered to a patient.

9. The cancer is a solid tumor, in particular the cancer is selected from lung cancer, pancreatic cancer, colon cancer and breast cancer, the agent for use according to claim 7 or 8.

10. The cancer is selected from the group consisting of bladder urothelial carcinoma, breast invasive carcinoma, cervical squamous cell carcinoma and endocervical adenocarcinoma, cholangiocarcinoma, colon adenocarcinoma, lymphoid neoplasm diffuse large B-cell lymphoma, esophageal cancer, glioblastoma multiforme, head and neck squamous cell carcinoma, chromophobe renal cell carcinoma, papillary renal cell carcinoma of the kidney, acute myeloid leukemia, low-grade glioma of the brain, lung adenocarcinoma, lung squamous cell carcinoma, mesothelioma, ovarian serous cystadenocarcinoma, pancreatic adenocarcinoma, rectal adenocarcinoma, sarcoma, cutaneous melanoma, gastric adenocarcinoma, testicular germ cell tumor, thyroid cancer, endometrial carcinoma of the uterine body, carcinosarcoma of the uterus, the agent for use according to claim 7 or 8.

11. The cancer is from the following group: a. For group a, it is KRAS, EGFR and / or TP53, in particular where the mutation of KRAS is a KRAS G12 mutation; or b. For group b, it is KRAS, FGFR and / or TP53, in particular where the mutation of KRAS is a KRAS Q61 mutation; or c. For group c, it is KRAS and / or EGFR, in particular where the mutation of KRAS is a KRAS G12 or KRAS Q61 mutation; or d. For group d, it is KRAS and / or TP53, in particular where the mutation of KRAS is a KRAS G12 mutation; or e. For group e, it is KRAS, EGFR and / or BRAF, in particular where the mutation of KRAS is a KRAS G12 or KRAS Q61 mutation; or f. For group f, it is KRAS, EGFR and / or BRAF, in particular where the mutation of KRAS is a KRAS G12 or KRAS Q61 mutation; or g. For group g, it is TP53; or h. For group h, it is KRAS, EGFR, BRAF and / or TP53, in particular where the mutation of KRAS is a KRAS G12 or KRAS Q61 mutation; or i. For group i, it is EGFR, FGFR and / or TP53; or j. KRAS, EGFR and / or TP53, in particular where the mutation of KRAS is a KRAS G12 mutation; or k. for group k, KRAS, EGFR, BRAF and / or TP53, in particular where the mutation of KRAS is a KRAS G12 or KRAS Q61 mutation; or n. for group n, KRAS, EGFR, BRAF and / or TP53, in particular where the mutation of KRAS is a KRAS G12 or KRAS Q61 mutation; or o. for group o, KRAS, EGFR, BRAF and / or TP53, in particular where the mutation of KRAS is a KRAS G12 or KRAS Q61 mutation A medicament for use according to any one of claims 7 to 10, characterized by a mutation of a gene selected from the group consisting of:

12. The cancer is from the following groups: a. for group a, KRAS, in particular where the mutation of KRAS is a KRAS G12 mutation; or b. for group b, TP53; or d. for group d, KRAS, in particular where the mutation of KRAS is a KRAS G12 mutation; or e. for group e, KRAS and / or EGFR, in particular where the mutation of KRAS is a KRAS G12 mutation; or f. for group f, KRAS and / or EGFR, in particular where the mutation of KRAS is a KRAS G12 or KRAS Q61 mutation; or i. for group i, TP53; or j. KRAS and / or EGFR, in particular where the mutation of KRAS is a KRAS G12 mutation; or k. for group k, EGFR and / or TP53; or n. for group n, KRAS, EGFR and / or TP53, in particular where the mutation of KRAS is a KRAS G12 mutation; or o. for group o, KRAS, EGFR and / or TP53, in particular where the mutation of KRAS is a KRAS G12 mutation A medicament for use according to any one of claims 7 to 9, characterized by a mutation of a gene selected from the group consisting of:

13. - A TCR polypeptide, - An isolated nucleic acid sequence encoding the TCR polypeptide, or - An isolated autologous T cell comprising the TCR polypeptide and / or the nucleic acid sequence; A medicament for use in the treatment of cancer, selected from the group consisting of: The TCR polypeptide comprises a CDR3 alpha sequence and a CDR3 beta sequence, and the CDR3 alpha sequence and the CDR3 beta sequence are identical to the sequences shown below, or have one amino acid substitution per CDR3 sequence, the CDR3 alpha sequence is selected from the sequences of SEQ ID NOs: 007-009 or SEQ ID NOs: 436-439, and the CDR3 beta sequence is selected from the sequences of SEQ ID NOs: 001-006 or SEQ ID NOs: 432-435. The TCR polypeptide further comprises a variable (V) alpha sequence, a joining constant (JC) alpha sequence, a V beta sequence and a JC beta sequence, or a sequence having at least 80%, at least 85%, at least 90%, at least 92%, at least 94%, at least 96%, at least 98% or at least 99% sequence identity to said sequences, the V alpha sequence is SEQ ID NO: 025, the JC alpha sequence is SEQ ID NO: 026, the V beta sequence is SEQ ID NO: 018, and the JC beta sequence is SEQ ID NO:

019. The agent is administered to a patient characterized by the HLA-A ＊ type of HLA at 02:01, and Said cancer is characterized by a mutation in a gene of the EGFR-Raf-Ras pathway, said agent. Claims 14 The agent for use according to claim 13, wherein said mutation in a gene of the EGFR-Raf-Ras pathway is a mutation in KRAS and / or EGFR. Claims 15 The agent for use according to claim 13, wherein said mutation in a gene of the EGFR-Raf-Ras pathway is a KRAS G12 mutation.

Images