Antigen recognizing construct capable of binding to a preferentially expressed antigen in melanoma (PRAME) peptide and a t cell receptor (TCR) having antigenic specificity for prame for use in a method of treating a disease in a subject as well as corresponding t-cells, nucleic acid sequences, vectors, pharmaceutical compositions and kits

Abstract

The present invention inter alia relates to (i) An antigen recognizing construct capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide, (ii) a chimeric human CD8 co-receptor comprising a polypeptide having an extracellular polypeptide region, a transmembrane polypeptide region, and a cytoplasmic polypeptide region, wherein said polypeptide comprises at least one CD8α polypeptide region, wherein said least one CD8α polypeptide region comprises an extracellular CD8α IG-like domain region, further wherein said polypeptide comprises at least one CD8β polypeptide region, wherein said least one CD8β polypeptide region comprises an extracellular CD8β IG-like domain region, and / or (iii) a chimeric human CD95 receptor; comprising a polypeptide, wherein said polypeptide comprises at least one CD95 polypeptide region comprising a CD95 extracellular ligand binding domain; further wherein said polypeptide comprises at least one non-CD95 co- stimulatory cytoplasmic polypeptide domain, region or motif; for use in a method of treating a disease in a subject, wherein the method comprises administering a therapeutically effective amount of the antigen recognizing construct, of the chimeric human CD8 co-receptor, and of the chimeric human CD95 receptor.The invention also relates to a corresponding method of treating a disease, as well as a to T- cells expressing (i), (ii) and (iii), to a pharmaceutical composition, and to a kit.

Description

New PCT ApplicationApplicant: T-Knife GmbHSWK Ref.: TKN18565PCTDate: 5 February 2026ANTIGEN RECOGNIZING CONSTRUCT CAPABLE OF BINDING TO A PREFERENTIALLY EXPRESSED ANTIGEN IN MELANOMA (PRAME) PEPTIDE AND A T CELL RECEPTOR (TCR) HAVING ANTIGENIC SPECIFICITY FOR PRAME FOR USE IN A METHOD OF TREATING A DISEASE IN A SUBJECT AS WELL AS CORRESPONDING T-CELLS, NUCLEIC ACID SEQUENCES, VECTORS, PHARMACEUTICAL COMPOSITIONS AND KITSREFERENCE TO SEQUENCE LISTING

[0001] This application contains a sequence listing in a computer readable form. The entire contents of the XML file is incorporated herein by reference.CROSS-REFERENCES

[0002] The present application claims the benefit of priority of the European Patent Application No. 25156079.3, filed 5 February 2025, the content of which is hereby incorporated by reference in its entirety for all purposes.FIELD OF THE INVENTION

[0003] The present invention relates to an antigen recognizing construct (e.g. a T-cell receptor) capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide for use in a method of treating a disease in a subject, a nucleic acid sequence encoding said antigen recognizing construct (e.g. said T cell receptor), a respective vector comprising the nucleic acid sequence, and a host cell comprising the antigen recognizing construct (e.g. the T cell receptor). The present invention further relates to a use of the antigen recognizing construct or the T cell receptor, the nucleic acid sequence, the vector, or the host cell, for the manufacture of a medicament for treating a disease. The present invention also relates to a method of treating a disease, comprising the step of administering a therapeutically effective amount of the antigen recognizing construct (e.g. the T cell receptor), the nucleic acid sequence, the vector, or the host cell. Additionally, the present invention also relates to a pharmaceutical composition comprisingthe antigen recognizing construct or the T cell receptor, the nucleic acid sequence, the vector, or the host cell of the invention and also to a respective kit for use in medicine.BACKGROUND OF THE INVENTION

[0004] Cancer testis antigens (CTAs) have been considered promising targets for adoptive T cell therapy thanks to their restricted expression in somatic normal tissues, reexpression in many cancer types (Fratta E. et al., Mol. Oncol. 2011;5: 164-182). CTA reexpression in tumors has been linked with worse clinical outcome in various cancers (Yao J. etal.; Cancer Immunol. Res. 2014; 2:371-379). Among all candidate CTAs, MAGE-A3, NY-ESO-1, and PRAME have shown great potential as prognostic biomarkers and immunotherapeutic targets.

[0005] PRAME is a tumor-associated antigen expressed in a wide variety of tumors. Specifically, PRAME is re-expressed in a wide range of cancers, including melanoma, ovarian cancer, endometrial cancer, esophageal cancer, renal cell cancer, non-small cell lung cancer (NSCLC), neuroblastoma, breast cancer, multiple myeloma, acute leukemia, chronic myeloid leukemia, multiple sarcoma subtypes, and primary and metastatic uveal melanoma (Al-Khadairi G. and Decock J.; Cancers (Basel). 2019 Jul; 11(7): 984). In normal tissues, it is expressed at high level only in testis.

[0006] The encoded protein acts as a repressor of retinoic acid receptor, and likely confers a growth advantage to cancer cells via this function. PRAME overexpression in triple negative breast cancer has also been found to promote cancer cell motility through induction of the epithelial-to-mesenchymal transition (Al-Khadairi et al., Journal of Translational Medicine 2019; 17: 9).

[0007] PRAME may be expressed in any of a variety of human cancers such as, for example, pancreatic cancers (e.g., pancreatic carcinoma), colorectal cancers, lung cancers (e.g., lung adenocarcinoma), endometrial cancers, ovarian cancers (e.g., epithelial ovarian cancer), prostate cancers, melanomas, thyroid cancers, and breast cancers, as well as some instances of myeloid leukemias such as AML.

[0008] Adoptive T-cell therapy (ACT) with T-cells expressing native or transgenic a -T-cell receptors (TCRs) is a promising treatment for cancer, as TCRs cover a wide range of potential target antigens [Chandran and Klebanoff, 2019], Native TCR specificities have successfully been exploited for ACT with tumor infiltrating lymphocytes (TILs) for melanoma [Dafni et al., 2019] and other tumors [Chandran and Klebanoff, 2019], or with virus-specific T-cells (VSTs) for viral-associated malignancies [Leung and Heslop, 2019], Transgenic TCR-based ACT allows the genetic redirection of T-cell specificity in a highly specific and reproducible manner, and has produced promising results in melanoma andseveral solid tumors [Robbins et al., 2015], multiple myeloma (MM) [Rapoport et al., 2015], viral-associated malignancies [Doran et al., 2019] and acute myeloid leukemia (AML) [Chapuis et al., 2019], Another promising option in ACT is the treatment with chimeric antigen receptor (CAR)-T-cells, which has produced remarkable clinical responses with certain subsets of B cell leukemia or lymphoma [Sterner and Sterner, 2019], Promising results have also been reported with multiple myeloma.

[0009] T-cell antigen recognition and subsequent T-cell activation is known to depend on the interaction between the T-cell receptor (TCR) and peptide-major histocompatibility complex (pMHC) molecules [Davis and Bjdrkman, 1988], In particular, the CD8 co-receptor plays a major role in CD8 T-cell activation, and the CD4 co-receptor stabilizes the interaction between the TCR on CD4 T-cells and the MHC class II molecule on antigen-presenting cells (APCs). Recently, it has been reported that in adoptive cell therapy experiments, the efficacy of high avidity CD4 T-cells in providing protective tumor immunity was similar to the therapeutic efficacy seen with CD8 T-cells. Specifically, it has been described that a co-transfer of Class I TCR- and CD8 co-receptor coding genes generated high avidity CD4 T-cells [Xue et al., 2013],

[0010] Furthermore, despite the progress made during recent years in developing specific ACT s targeting tumor cell specific antigen genes, a number of challenges of ACT s such as tumor heterogeneity, antigen escape, T-cell trafficking and an immunosuppressive tumor microenvironment remain to be addressed. For example, solid tumors can effectively evade the immune response, including the promising T cell therapies, through the expression of various inhibitory molecules that can hinder the function of T cells. For example, while, above, receptors of the immunoglobulin superfamily (IgSF) and tumor necrosis factor receptor superfamily (TNFRSF) have been mentioned that are known to have co-stimulatory function, there are also family members which are known to be bound by the inhibitory molecules - e.g. in the immunosuppressive tumor microenvironment -and to transmit the inhibitory effect to the T-cell. Within the IgSF, for example, PD1 and T cell immunoreceptor with Ig and ITIM domains (TIGIT) and TIM-3 have been described to transmit inhibitory signals coming from solid tumors that may inhibit activation, and / or promote exhaustion of T-cells.

[0011] In order to address the issue of the inhibitory effects to the T-cell described above, chimeric switch receptors (i.e. chimeric receptors comprising the extracellular domain of an inhibitory receptor and the cytoplasmic domain of an activating receptor) have been created to reverse the outcomes of their original signaling pathways in order to confer immune cells with the ability to overcome the immunosuppressive tumor microenvironment and to allow them to have greater in vivo persistence.

[0012] For example, PD1 switch receptors have been described, wherein the extracellular domain of PD1 has been fused to the cytoplasmic domain of ICOS or CD28, respectively [WO 2013019615A2], In particular, in this international patent application, the described PD1 switch receptors with ICOS or CD28 co-stimulatory domains, respectively, could be co-expressed with a specific chimeric antigen receptor (CAR) in T-cells. However, in view of the diversity of immunosuppressive tumor microenvironments, and in light of the huge number of involved actors which may both positively and negatively regulate the suppressive activity of T-cells in adoptive cell therapy, it still remains a challenging task to provide effective T-cells exhibiting sufficient cytotoxicity and / or cytokine secretion, in particular in the immunosuppressive tumor microenvironment.

[0013] There is thus a need for novel therapies targeting PRAME peptides.

[0014] Accordingly, it is an object of the invention to provide such therapies.SUMMARY OF THE INVENTION

[0015] This object is inter alia accomplished by the antigen recognizing constructs, and / or the chimeric CD8 co-receptors, and / or the chimeric CD95 receptors as herein described for use in a method of treating a disease, as well as by the nucleic acids, vectors, host cells, methods, compositions, and kits having the features of the respective independent claims.

[0016] In a first aspect, the invention relates to(i) An antigen recognizing construct capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide,(ii) a chimeric human CD8 co-receptor comprising a polypeptide having an extracellular polypeptide region, a transmembrane polypeptide region, and a cytoplasmic polypeptide region, wherein said polypeptide comprises at least one CD8a polypeptide region, wherein said at least one CD8a polypeptide region comprises an extracellular CD8a IG-like domain region, further wherein said polypeptide comprises at least one CD8p polypeptide region, wherein said at least one CD8p polypeptide region comprises an extracellular CD8p IG-like domain region, and / or(iii) a chimeric human CD95 receptor; comprising a polypeptide, wherein said polypeptide comprises at least one CD95 polypeptide region comprising a CD95 extracellular ligand binding domain; further wherein said polypeptide comprises at least one non-CD95 costimulatory cytoplasmic polypeptide domain, region or motif;for use in a method of treating a disease in a subject, wherein the method comprises administering a therapeutically effective amount of the antigen recognizing construct, of the chimeric human CD8 co-receptor, and of the chimeric human CD95 receptor.

[0017] The inventors have been found that the introduction of the specific combination of each of the specific antigen recognizing construct, of the specific chimeric human CD8 Co-receptor, and of the specific chimeric human CD95 receptor as herein provided into a host cell that is suitable for immunotherapy demonstrate improved cytotoxic activity and advantageous effector molecule secretion, thus representing an improved therapy for targeting PRAME expressing indications.

[0018] In a second aspect, the invention provides a method of treating a disease in a subject, comprising the step of administering a therapeutically effective amount of (i) An antigen recognizing construct capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide,(ii) a chimeric human CD8 co-receptor comprising a polypeptide having an extracellular polypeptide region, a transmembrane polypeptide region, and a cytoplasmic polypeptide region, wherein said polypeptide comprises at least one CD8a polypeptide region, wherein said least one CD8a polypeptide region comprises an extracellular CD8a IG-like domain region, further wherein said polypeptide comprises at least one CD8p polypeptide region, wherein said least one CD8p polypeptide region comprises an extracellular CD8p IG-like domain region, and(iii) a chimeric human CD95 receptor; comprising a polypeptide, wherein said polypeptide comprises at least one CD95 polypeptide region comprising a CD95 extracellular ligand binding domain; further wherein said polypeptide comprises at least one non-CD95 costimulatory cytoplasmic polypeptide domain, region or motif.

[0019] The specific co-administration (to a host cell) of each of the three constructs as specifically selected in accordance with the second aspect of the invention shows superior results in comparison with prior art PRAME administrations.

[0020] According to a third aspect, the invention provides a combination comprising (i) an antigen recognizing construct capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide,(ii) a chimeric human CD8 co-receptor comprising a polypeptide having an extracellular polypeptide region, a transmembrane polypeptide region, and a cytoplasmic polypeptide region, wherein said polypeptide comprises at least one CD8a polypeptide region, wherein said least one CD8a polypeptide region comprises an extracellular CD8a IG-like domain region, further wherein said polypeptide comprises at least one CD8p polypeptide region,wherein said least one CD8p polypeptide region comprises an extracellular CD8p IG-like domain region, and(iii) a chimeric human CD95 receptor; comprising a polypeptide, wherein said polypeptide comprises at least one CD95 polypeptide region comprising a CD95 extracellular ligand binding domain; further wherein said polypeptide comprises at least one non-CD95 costimulatory cytoplasmic polypeptide domain, region or motif.

[0021] In a fourth aspect, the invention provides a nucleic acid encoding(i) an antigen recognizing construct capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide,(ii) a chimeric human CD8 co-receptor comprising a polypeptide having an extracellular polypeptide region, a transmembrane polypeptide region, and a cytoplasmic polypeptide region, wherein said polypeptide comprises at least one CD8a polypeptide region, wherein said least one CD8a polypeptide region comprises an extracellular CD8a IG-like domain region, further wherein said polypeptide comprises at least one CD8p polypeptide region, wherein said least one CD8p polypeptide region comprises an extracellular CD8p IG-like domain region, and(iii) a chimeric human CD95 receptor; comprising a polypeptide, wherein said polypeptide comprises at least one CD95 polypeptide region comprising a CD95 extracellular ligand binding domain; further wherein said polypeptide comprises at least one non-CD95 costimulatory cytoplasmic polypeptide domain, region or motif.

[0022] In a fifth aspect, the invention provides a vector comprising the nucleic acid according to the fourth aspect.

[0023] In a sixth aspect, the invention provides a host cell (e.g. an isolated T-cell), comprising(i) an antigen recognizing construct capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide,(ii) a chimeric human CD8 co-receptor comprising a polypeptide having an extracellular polypeptide region, a transmembrane polypeptide region, and a cytoplasmic polypeptide region, wherein said polypeptide comprises at least one CD8a polypeptide region, wherein said least one CD8a polypeptide region comprises an extracellular CD8a IG-like domain region, further wherein said polypeptide comprises at least one CD8p polypeptide region, wherein said least one CD8p polypeptide region comprises an extracellular CD8p IG-like domain region, and(iii) a chimeric human CD95 receptor; comprising a polypeptide, wherein said polypeptide comprises at least one CD95 polypeptide region comprising a CD95 extracellular ligandbinding domain; further wherein said polypeptide comprises at least one non-CD95 costimulatory cytoplasmic polypeptide domain, region or motif.

[0024] Alternatively, in accordance with the sixth aspect, the host cell (e.g. the isolated T-cell) may also comprise a nucleic acid encoding for the polypeptides (i), (ii), and (iii), or a vector comprising said nucleic acid.

[0025] According to a seventh aspect, the invention provides a use of(i) an antigen recognizing construct capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide,(ii) a chimeric human CD8 co-receptor comprising a polypeptide having an extracellular polypeptide region, a transmembrane polypeptide region, and a cytoplasmic polypeptide region, wherein said polypeptide comprises at least one CD8a polypeptide region, wherein said least one CD8a polypeptide region comprises an extracellular CD8a IG-like domain region, further wherein said polypeptide comprises at least one CD8p polypeptide region, wherein said least one CD8p polypeptide region comprises an extracellular CD8p IG-like domain region, and(iii) a chimeric human CD95 receptor; comprising a polypeptide, wherein said polypeptide comprises at least one CD95 polypeptide region comprising a CD95 extracellular ligand binding domain; further wherein said polypeptide comprises at least one non-CD95 costimulatory cytoplasmic polypeptide domain, region or motif;for the manufacture of a medicament.

[0026] In accordance with said seventh aspect, there is alternatively provided the use of a combination comprising each of the polypeptides (i), (ii), and (iii), or the use of a nucleic acid encoding each of the polypeptides (i), (ii), and (iii), or the use of a vector comprising said nucleic acid, or the use of a cell comprising the nucleic acid, the vector, and / or the polypeptides (i), (ii), and (iii), for the manufacture of a medicament.

[0027] According to an eighth aspect, there is provided an isolated T-cell which expresses:(i) an antigen recognizing construct capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide,(ii) a chimeric human CD8 co-receptor comprising a polypeptide having an extracellular polypeptide region, a transmembrane polypeptide region, and a cytoplasmic polypeptide region, wherein said polypeptide comprises at least one CD8a polypeptide region, wherein said least one CD8a polypeptide region comprises an extracellular CD8a IG-like domain region, further wherein said polypeptide comprises at least one CD8p polypeptide region, wherein said least one CD8p polypeptide region comprises an extracellular CD8p IG-like domain region, and(iii) a chimeric human CD95 receptor; comprising a polypeptide, wherein said polypeptide comprises at least one CD95 polypeptide region comprising a CD95 extracellular ligand binding domain; further wherein said polypeptide comprises at least one non-CD95 costimulatory cytoplasmic polypeptide domain, region or motif.

[0028] According to a ninth aspect, there is provided a pharmacological composition comprising:(i) an antigen recognizing construct capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide,(ii) a chimeric human CD8 co-receptor comprising a polypeptide having an extracellular polypeptide region, a transmembrane polypeptide region, and a cytoplasmic polypeptide region, wherein said polypeptide comprises at least one CD8a polypeptide region, wherein said least one CD8a polypeptide region comprises an extracellular CD8a IG-like domain region, further wherein said polypeptide comprises at least one CD8p polypeptide region, wherein said least one CD8p polypeptide region comprises an extracellular CD8p IG-like domain region, and(iii) a chimeric human CD95 receptor; comprising a polypeptide, wherein said polypeptide comprises at least one CD95 polypeptide region comprising a CD95 extracellular ligand binding domain; further wherein said polypeptide comprises at least one non-CD95 costimulatory cytoplasmic polypeptide domain, region or motif; orb) the combination comprising each of the polypeptides as defined by i), ii), and iii); or c) the nucleic acid encoding for each of the polypeptides as defined by i), ii), and iii); or d) the vector comprising the nucleic acid, ore) the cell (e.g. host cell, or e.g. (isolated) T-cell) comprising and / or expressing each of the polypeptides as defined by i), ii) and iii).

[0029] According to a tenth aspect, there is provided a method of preparing a T-cell for immunotherapy, comprisingisolating T-cells from a human subject,introducing the vector comprising the nucleic acid encoding for the polypeptides as defined by i), ii) and iii) into the T-cell, or introducing the nucleic acid encoding for the polypeptides as defined by i), ii) and iii) into the T-cell ; andexpanding the T-cells.

[0030] According to an eleventh aspect, there is also provided a kit for use in medicine.BRIEF DESCRIPTION OF THE DRAWINGS

[0032] The invention will be better understood with reference to the detailed description when considered in conjunction with the non-limiting examples and the drawings, in which:

[0033] Fig. 1 shows a graphical representation of the results from an in-vitro T-cell killing assay of T-cells transduced with a specifically selected combination of a specific T cell receptor capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide, of a chimeric CD8 co-receptor polypeptide, and of a chimeric human CD95 receptor according to the invention in comparison with transduction of T -cells with other, previously known combinations of published T-cell receptors together with previously published Switch receptors or previously published Co-receptors.

[0034] Fig. 2 shows a graphical representation of results from a cytokine secretion assay of T-cells transduced with a specifically selected combination of a specific T cell receptor capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide, of a chimeric CD8 co-receptor polypeptide, and of a chimeric human CD95 receptor according to the invention in comparison with transduction of T-cells with other, previously known combinations of published T-cell receptors together with previously published Switch receptors or previously published Co-receptors, wherein the amount of secreted IFN-y is shown in pg / mL.

[0035] Fig. 3 shows a graphical representation of results from an effector molecule secretion assay of T-cells transduced with a specifically selected combination of a specific T cell receptor capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide, of a chimeric CD8 co-receptor polypeptide, and of a chimeric human CD95 receptor according to the invention in comparison with transduction of T -cells with other, previously known combinations of published T-cell receptors together with previously published Switch receptors or previously published Co-receptors, wherein the amount of secreted Granzyme A (GZMA) is shown in pg / mL.

[0036] Fig. 4 shows a graphical representation of results from an effector molecule secretion assay of T-cells transduced with a specifically selected combination of a specific T cell receptor capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide, of a chimeric CD8 co-receptor polypeptide, and of a chimeric human CD95 receptor according to the invention in comparison with transduction of T -cells with other, previously known combinations of published T-cell receptors together with previously published Switch receptors or previously published Co-receptors, wherein the amount of secreted Granzyme B (GZMB) is shown in pg / mL.

[0037] Fig. 5 shows a graphical representation of results from an effector molecule secretion of T-cells transduced with a specifically selected combination of a specific T cell receptor capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide, of a chimeric CD8 co-receptor polypeptide, and of a chimeric human CD 95 receptor according to the invention in comparison with transduction of T -cells with other, previously known combinations of published T-cell receptors together with previously published Switch receptors or previously published Co-receptors, wherein the amount of secreted perforin is shown in pg / mL.

[0038] Fig. 6 shows a graphical representation of results from a soluble secreted inhibitory molecule detection assay after a co-culture of cancer cells with T-cells transduced with a specifically selected combination of a specific T cell receptor capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide, of a chimeric CD8 co-receptor polypeptide, and of a chimeric human CD 95 receptor according to the invention in comparison with transduction of T -cells with other, previously known combinations of published T-cell receptors together with previously published Switch receptors or previously published Co-receptors, wherein the FasL level is shown in pg / mL.DETAILED DESCRIPTION OF THE INVENTION

[0039] As explained above, in a first aspect, the invention is directed to(i) An antigen recognizing construct capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide,(ii) a chimeric human CD8 co-receptor comprising a polypeptide having an extracellular polypeptide region, a transmembrane polypeptide region, and a cytoplasmic polypeptide region, wherein said polypeptide comprises at least one CD8a polypeptide region, wherein said least one CD8a polypeptide region comprises an extracellular CD8a IG-like domain region, further wherein said polypeptide comprises at least one CD8p polypeptide region, wherein said least one CD8p polypeptide region comprises an extracellular CD8p IG-like domain region, and / or(iii) a chimeric human CD95 receptor; comprising a polypeptide, wherein said polypeptide comprises at least one CD95 polypeptide region comprising a CD95 extracellular ligand binding domain; further wherein said polypeptide comprises at least one non-CD95 costimulatory cytoplasmic polypeptide domain, region or motif;for use in a method of treating a disease in a subject, wherein the method comprises administering a therapeutically effective amount of the antigen recognizing construct, of the chimeric human CD8 co-receptor, and of the chimeric human CD95 receptor.

[0040] The antigen recognizing construct capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide

[0041] According to an embodiment, the antigen recognizing construct for the use according to the first aspect of the invention may comprise a complementary determining region 3 (CDR3) of the a-chain having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 3, 9, 15, 21, 27, 33, 39, 45, 51, 57, 63, 69, and 75, and / or wherein the antigen recognizing construct comprises a complementary determining region 3 (CDR3) of the p-chain having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 6, 12, 18, 24, 30, 36, 42, 48, 54, 60, 66, 72 and 78.

[0042] It is herewith envisaged that the antigen recognizing construct may comprise a complementary determining region 3 (CDR3) of the a-chain which may have one or more conservative amino acid substitutions relative to the amino acid sequence as set forth in the group consisting of SEQ ID NOs: 3, 9, 15, 21, 27, 33, 39, 45, 51, 57, 63, 69, and 75.

[0043] It is further contemplated that the antigen recognizing construct may comprise a complementary determining region 3 (CDR3) of the p-chain which may have one or moreconservative amino acid substitutions relative to the amino acid sequence as set forth in the group consisting of SEQ ID NOs: 6, 12, 18, 24, 30, 36, 42, 48, 54, 60, 66, 72 and 78.

[0044] The antigen recognizing construct according to the first aspect of the invention may be an antibody, or fragment thereof, or a T cell receptor (TCR), or fragment thereof.

[0045] Thus, it is within the scope of the first aspect of the invention that the antigen recognizing construct is an antibody, or a fragment thereof. The term “antibody” in its various grammatical forms is used herein to refer to immunoglobulin molecules and immunologically active portions of immunoglobulin molecules, i.e. , molecules that contain an antibody combining site or a paratope. Such molecules are also referred to as "antigen binding fragments" of immunoglobulin molecules. The invention further provides an antibody, or antigen binding portion thereof, which specifically binds to the antigens described herein. The antibody can be any type of immunoglobulin that is known in the art. For instance, the antibody can be of any isotype, e.g., IgA, IgD, IgE, IgG, IgM, etc. The antibody can be monoclonal or polyclonal. The antibody can be a naturally-occurring antibody, e.g., an antibody isolated and / or purified from a mammal, e.g., mouse, rabbit, goat, horse, chicken, hamster, human, etc. Alternatively, the antibody can be a genetically-engineered antibody, e.g., a humanized antibody or a chimeric antibody. The antibody can be in monomeric or polymeric form.

[0046] A "T cell" or "T lymphocyte" is an immune system cell that matures in the thymus and produces T cell receptors (TCR). T cells can be naive ("TN"; not exposed to antigen; increased expression of CD62L, CCR7, CD28, CD3, CD127, and CD45RA, and decreased or no expression of CD45RO as compared to TCM), memory T cells (TM) (antigen experienced and long-lived), including stem cell memory T cells, and effector cells (antigen-experienced, cytotoxic). TM can be further divided into subsets of central memory T cells (TEM, express CD62L, CCR7, CD28, CD95, CD45RO, and CD127) and effector memory T cells (TEM express CD45RO, decreased expression of CD62L, CCR7, CD28, and CD45RA). Effector T cells (TE) refers to antigen-experienced CD8+ cytotoxic T lymphocytes that express CD45RA, have decreased expression of CD62L, CCR7, and CD28 as compared to TCM, and are positive for granzyme and perforin. CD4+ T-cells influence the activity of other immune cells by releasing cytokines. CD4+ T cells can activate and suppress an adaptive immune response, and which of those two functions is induced will depend on the presence of other cells and signals. T cells can be collected using known techniques, and the various subpopulations or combinations thereof can be enriched or depleted by known techniques, such as by affinity binding to antibodies, flow cytometry, or immunomagnetic selection. Other exemplary T cells include regulatory T cells, such as CD4+ CD25+ (Foxp3+) regulatory T cells and Treg17 cells, as well as Tr1, Th3, and Qa-1 restricted T cells.

[0047] "T cell receptor" (TCR) refers to an immunoglobulin superfamily member having a variable binding domain, a constant domain, a transmembrane region, and a short cytoplasmic tail; see, e. g., Janeway el al., Immunobiology: The Immune System in Health and Disease, 3rd Ed., Current Biology Publications, p. 433, 1997) capable of specifically binding to an antigen peptide presented on an MHC molecule. A TCR can be found on the surface of a cell or in soluble form and generally is comprised of a heterodimer having a and p chains (also known as TCR a and TCR p, respectively), or y and 8 chains (also known as TCRy and TCR8, respectively). A polynucleotide encoding a binding protein of this disclosure, e.g., a TCR, can be codon optimized to enhance expression in a particular host cell, such, for example, as a cell of the immune system, a hematopoietic stem cell, a T cell, a primary T cell, a T cell line, a NK cell, or a natural killer T cell (Scholten el al., Clin. Immunol. 119:135, 2006).

[0048] The terms "complementarity determining region" and "CDR," are synonymous with "hypervariable region" or "HVR," and are known in the art to refer to sequences of amino acids within immunoglobulin or TCR variable regions, which confer antigen specificity and / or binding affinity and are separated from one another in primary amino acid sequence by framework regions. In general, there are three CDRs in each TCR a-chain variable region (aCDR1, aCDR2, aCDR3) and three CDRs in each TCR p-chain variable region (PCDR1, PCDR2, PCDR3). In TCRs, CDR3 is thought to be the main CDR responsible for recognizing processed antigen. In general, CDR1 and CDR2 interact mainly or exclusively with the MHC.

[0049] The term “capable of binding” means that said PRAME peptide is specifically bound by said TCR. The term “specific(ally) binding” generally indicates that a TCR binds via its antigen binding site more readily to its intended antigenic target than to a random, unrelated non-target antigen. Particularly the term “specifically binds” may indicate that the binding affinity of the TCR will be at least about 5-fold, or at least 10-fold, or at least 25-fold, or at least 50-fold, or at least 100-fold or more, greater for its antigenic target than its binding specificity for a non-target antigen.

[0050] It is understood that the expression “PRAME peptide” relates to a human PRAME peptide / protein having an amino acid sequence according to UniProtKB database entry P78395 ■ PRAME_HUMAN, as set forth e.g. in SEQ ID No.: 79. The antigen recognizing construct according to the first aspect of the invention, which binds the PRAME peptide, may be presented by an MHC-I molecule. For example, the MHC-I molecule may be an HLA-A molecule. The HLA-A molecule may be, for example, HLA-A2. In embodiments, the HLA-A molecule may be HLA-A*02:01, HLA-A*02:02, HLA-A*02:03, HLA-A*02:04, HLA-A*02:05, HLA-A*02:06, HLA-A*02:07, HLA-A*68:02, or HLA-A*6901. According to a specific example, the HLA-A molecule may be HLA-A*02:01.

[0051] According to an embodiment the antigen recognizing construct may bind the amino acid sequence SLLQHLIGL (SEQ ID NO: 80) within the PRAME peptide. This epitope (“epitope SLL425-433 ) is known to be highly abundant on PRAM E-expressing tumor cells.

[0052] There are two classes of MHC molecules, MHC class I and MHC class II. Complexes of peptide and MHC class I are recognized by CD8-positive T cells bearing the appropriate T-cell receptor (TCR), whereas complexes of peptide and MHC class II molecules are recognized by CD4-positive T cells bearing the appropriate TCR. Since both types of responses, CD8 and CD4 T cell dependent responses, contribute jointly and synergistically to the anti-tumor effect, the identification and characterization of tumor-associated and tumor-specific antigens and corresponding T cell receptors is important in the development of cancer immunotherapies, such as vaccines and cell therapies.

[0053] A TCR as described herein is envisaged to bind to its antigenic target (for example a PRAME peptide presented on HLA-A*02:01 encoded molecules by antigen presenting cells), resulting in a high functional avidity of the TCR expressing cell. The term “functional avidity” refers to the capability of TCR expressing cells (in particular T cells expressing native TCRs as described herein) to respond in vitro to a given concentration of a ligand, and is thought to correlate with the in vivo effector capacity of TCR expressing cells. By definition, TCR expressing cells with high functional avidity respond in in-vitro tests to very low antigen concentrations, while such cells of lower functional avidity require higher amounts of antigen before they mount an immune response similar to that of the high-avidity TCR expressing cells. The functional avidity can be therefore considered as a quantitative determinant of the activation threshold of a TCR expressing cell. It is determined by exposing such cells in vitro to different amounts of cognate antigen. TCR expressing cells with high functional avidity respond to low antigen concentrations. Hence, the TCR of the present invention is a high-avidity TCR causing a half-maximal relative IFN-gamma secretion (EC50 value) of less than 10'85M, as measured by an IFN-gamma immunoassay (Figure 1).

[0054] Thus, in accordance with an embodiment, the antigen recognizing construct may be a TCR, and T-cells expressing said T-cell receptor may have a functional avidity characterized by an EC50 of less than about 1 x 10'85 MThe functional avidity may be assessed by evaluating secretion of interferon-gamma by a population of T cells expressing the T cell receptor. In illustrated examples, the functional avidity may be characterized by an EC50 of less than about 1 x 10'86M or of less than about 1 x 10'87M or of less than about 1 x 10'88M, or of less than about 1 x 10'89M, or of less than about 1x 10'9M. The EC50 value may be determined by the procedure as carried out in Example 1 as described herein.

[0055] According to an embodiment, the antigen recognizing construct, e.g. the T cell receptor, may comprise a complementary determining region 1 (CDR1) of the a-chain having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 7, 13, 19, 25, 31, 37, 43, 49, 55, 61, 67, and 73, and / or the antigen recognizing construct, e.g. the T cell receptor may comprise a complementary determining region 1 (CDR1) of the p-chain having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 10, 16, 22, 28, 34, 40, 46, 52, 58, 64, 70 and 76.

[0056] It is herewith envisaged that the antigen recognizing construct may comprise a complementary determining region 1 (CDR1) of the a-chain which may have one or more conservative amino acid substitutions relative to the amino acid sequence as set forth in the group consisting of SEQ ID NOs: 1, 7, 13, 19, 25, 31, 37, 43, 49, 55, 61, 67, and 73.

[0057] It is further contemplated that the antigen recognizing construct may comprise a complementary determining region 1 (CDR1) of the p-chain which may have one or more conservative amino acid substitutions relative to the amino acid sequence as set forth in the group consisting of SEQ ID NOs: 4, 10, 16, 22, 28, 34, 40, 46, 52, 58, 64, 70 and 76.

[0058] According to an embodiment, the antigen recognizing construct, e.g. the T cell receptor, may comprise a complementary determining region 2 (CDR2) of the a-chain having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 2, 8, 14, 20, 26, 32, 36, 44, 50, 56, 62, 68, and 74, and / or the T cell receptor may comprise a complementary determining region 2 (CDR2) of the p-chain having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 5, 11, 17, 23, 29, 35, 41, 47, 53, 59, 65, 71 and 77.

[0059] It is herewith envisaged that the antigen recognizing construct may comprise a complementary determining region 2 (CDR2) of the a-chain which may have one or more conservative amino acid substitutions relative to the amino acid sequence as set forth in the group consisting of SEQ ID Nos 2, 8, 14, 20, 26, 32, 36, 44, 50, 56, 62, 68, and 74.

[0060] It is further contemplates that the antigen recognizing construct may comprise a complementary determining region 2 (CDR2) of the p-chain which may have one or more conservative amino acid substitutions relative to the amino acid sequence as set forth in the group consisting of SEQ ID NOs: 5, 11, 17, 23, 29, 35, 41, 47, 53, 59, 65, 71 and 77.

[0061] It is also comprised in the first aspect of the invention that the antigen recognizing construct, e.g. the T cell receptor, may comprise a complementary determining region 3 (CDR3) of the a-chain having at least 95%, or at least 96%, or at least 97%, or at least 98%, or at least 99%, or 100% sequence identity to an amino acidsequence selected from the group consisting of SEQ ID NOs: 3, 9, 15, 21, 27, 33, 39, 45, 51, 57, 63, 69, and 75, and / or the antigen recognizing construct, e.g. the T cell receptor, may comprise a complementary determining region 3 (CDR3) of the p-chain having at least 95%, or at least 96%, or at least 97%, or at least 98%, or at least 99%, or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 6, 12, 18, 24, 30, 36, 42, 48, 54, 60, 66, 72 and 78. The antigen recognizing construct, e.g. the T cell receptor, may comprise a complementary determining region 1 (CDR1) of the a-chain having at least 95%, or at least 96%, or at least 97%, or at least 98%, or at least 99%, or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 7, 13, 19, 25, 31, 37, 43, 49, 55, 61, 67, and 73, and / or the antigen recognizing construct, e.g. the T cell receptor, may comprise a complementary determining region 1 (CDR1) of the p-chain having at least 95%, or at least 96%, or at least 97%, or at least 98%, or at least 99%, or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 10, 16, 22, 28, 34, 40, 46, 52, 58, 64, 70 and 76. The antigen recognizing construct, e.g. the T cell receptor, may comprise a complementary determining region 2 (CDR2) of the a-chain having at least 95%, or at least 96%, or at least 97%, or at least 98%, or at least 99%, or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 2, 8, 14, 20, 26, 32, 36, 44, 50, 56, 62, 68, and 74, and / or the antigen recognizing construct, e.g. the T cell receptor, may comprise a complementary determining region 2 (CDR2) of the p-chain having at least 95%, or at least 96%, or at least 97%, or at least 98%, or at least 99%, or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 5, 11, 17, 23, 28, 35, 41, 47, 53, 59, 65, 71 and 77.

[0062] The antigen recognizing construct, e.g. the T cell receptor, according to the first aspect of the invention may comprise(a) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 1,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 2, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 3; or(b) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 7,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 8, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 9; or(c) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 13,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 14, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 15; or(d) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 19,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 20, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 21 ; or(e) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 25,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 26, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 27, or(f) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 31,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 32, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 33, or(g) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 37,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 38, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 39, or(h) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 43,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 44, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 45, or(i) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 49,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 50, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 51 , or(j) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 55,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 56, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 57, or(k) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 61,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 62, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 63, or(l) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 67,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 68, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 69, or(m) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 73,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 74, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 75.

[0063] The antigen recognizing construct, e.g, the T cell receptor, according to the first aspect of the invention may comprise(a) a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 4,a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 5, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 6; or (b) a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 10,a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 11 , anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 12; or(c) a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 16,a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 17, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 18; or(d) a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 22,a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 23, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 24; or(e) a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 28,a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 29, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 30, or(f) a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 34,a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 35, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 36, or(g) a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 40,a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 41 , anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 42, or(h) a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 46,a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 47, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 48, or(i) a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 52,a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 53, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 54, or(j) a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 58,a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 59, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 60, or(k) a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 64,a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 65, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 66, or(l) a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 70,a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 71 , anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 72, or(m) a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 76,a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 77, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 78.

[0064] The antigen recognizing construct, e.g. the T cell receptor, for use according to the first aspect of the invention may comprise(a) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 1,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 2, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 3; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 4, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 5, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 6; or (b) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 7,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 8, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 9; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 10, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 11 , anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 12; or(c) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 13,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 14, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 15; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 16, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 17, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 18; or(d) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 19,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 20, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 21 ; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 22, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 23, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 24; or(e) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO:an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 26, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 27; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 28, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 39, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 30, or(f) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 31,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 32, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 33; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 34, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 35, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 36, or(g) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 37,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 38, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 39; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 40, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 41 , anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 42, or(h) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 43,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 44, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 45; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 46, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 47, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 48, or(i) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 49,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 50,an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 51 ; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 52, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 53, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 54, or(j) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 55,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 56, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 57; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 58, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 59, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 60, or(k) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 61,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 62, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 63; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 64, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 65, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 66, or(l) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 67,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 68, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 69; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 70, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 71 , anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 72, or(m) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 73,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 74, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 75;a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 76, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 77, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 78.

[0065] In embodiments according to the first aspect, the antigen recognizing construct for the use according to the first aspect of the invention may be a T cell receptor (TCR) having antigenic specificity for Preferentially Expressed Antigen in Melanoma (PRAME), wherein the TCR comprises:(a) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 1,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 2, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 3; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 4, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 5, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 6; or (b) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 7,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 8, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 9; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 10, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 11 , anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 12; or(c) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 13,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 14, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 15; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 16, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 17, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 18; or(d) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 19,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 20, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 21 ; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 22, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 23, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 24; or(e) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 25,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 26, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 27; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 28, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 39, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 30, or(f) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 31,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 32, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 33; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 34, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 35, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 36, or(g) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 37,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 38, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 39; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 40, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 41 , anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 42, or(h) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 43,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 44,an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 45; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 46, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 47, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 48, or(i) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 49,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 50, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 51 ; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 52, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 53, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 54, or(j) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 55,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 56, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 57; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 58, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 59, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 60, or(k) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 61,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 62, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 63; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 64, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 65, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 66, or(l) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 67,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 68, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 69;a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 70, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 71 , anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 72, or(m) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 73,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 74, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 75; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 76, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 77, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 78.

[0066] According to an embodiment, the T cell receptor for the use according to the first aspect may comprise:an a-chain variable domain having at least 70%, or at least 80%, or at least 90%, or at least 95%, or at least 96%, or at least 97%, or at least 98%, or at least 99%, or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 81, 83, 85, 87, 89, 91, 93, 95, 97, 99, 101, 103, and 105 and / ora p-chain variable domain having at least 70%, or at least 80%, or at least 90%, or at least 95%, or at least 96%, or at least 97%, or at least 98%, or at least 99%, or 100% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 82, 84, 86, 88, 90, 92, 94, 96, 98, 100, 102, 104 and 106.

[0067] In accordance with a preferred embodiment of the first aspect, the T-cell receptor for the use in accordance with the first aspect comprises: an a-chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 111, and a p-chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 112.

[0068] According to an embodiment, the T cell receptor for the use according to the first aspect may comprise:(a) an a-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 81; anda p-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 82;or(b) (a) an a-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 83; anda p-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 84;or(c) an a-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 85; anda p-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 86;or(d) an a-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 87; anda p-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 88;or(e) an a-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 89; anda p-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 90;or(f) an a-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 91; anda p-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 92;or(g) an a-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 93; anda p-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 94;or(h) an a-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 95; anda p-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 96;or(i) an a-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 97; anda p-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 98;or(j) an a-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 99; anda p-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 100;or(k) an a-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 101; anda p-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 102;or(l) an a-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 103; anda p-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 104;or(m) an a-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 105; anda p-chain variable domain having at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID No. 106.

[0069] In accordance with a preferred embodiment of the first aspect, the T-cell receptor for the use in accordance with the first aspect comprises: an a-chain variable region comprising or consisting of a polypeptide having at least 90% sequence identity, or at least 95% identity, or at least 96% identity, or at least 97% identity, or at least 98% identity, or at least 99% identityto an amino acid sequence as set forth in SEQ ID NO: 111, and a p-chain variable region comprising or consisting of a polypeptide having at least 90% sequence identity , or at least 95% identity, or at least 96% identity, or at least 97% identity, or at least 98% identity, or at least 99% identity, to an amino acid sequence as set forth in SEQ ID NO: 112.

[0070] According to an embodiment of the first aspect, the T-cell receptor for the use in accordance with the first aspect comprises a TCR a-chain comprising or consisting of the amino acid sequence of SEQ ID NO: 107, and a TCR p-chain comprising or consistingof the amino acid sequence of SEQ ID NO: 108. It is envisaged herein that the T-cell receptor for the use in accordance with the first aspect may comprise a TOR a-chain having at least 70%, or at least 75 %, or at least 80%, or at least 85%, or at least 90% sequence identity to an amino acid sequence as set forth in SEQ ID NO: 107, and a TOR p-chain having at least 70%, or at least 75 %, or at least 80%, or at least 85%, or at least 90% sequence identity to an amino acid sequence as set forth in the amino acid sequence of SEQ ID NO: 108.

[0071] Included in the scope of the invention are functional variants of the inventive antigen recognizing constructs or TCRs for the use according to the first aspect described herein. The term “functional variant,” as used herein, refers to an antigen recognizing construct or TCR having substantial or significant sequence identity or similarity to a parent antigen recognizing construct or TCR, which functional variant retains the biological activity of the antigen recognizing construct or TCR of which it is a variant. Functional variants encompass, for example, those variants of the antigen recognizing construct or TCR described herein (the parent antigen recognizing construct or TCR) that retain the ability to specifically bind to a PRAME peptide for which the parent antigen recognizing construct or TCR has antigenic specificity to a similar extent, the same extent, or to a higher extent, as the parent antigen recognizing construct or TCR. In reference to the parent antigen recognizing construct or TCR, the functional variant can, for instance, be at least about 30%, 50%, 75%, 80%, 90%, 95%, 96%, 97%, 98%, 99% or more identical in amino acid sequence to the parent antigen recognizing construct or TCR.

[0072] The functional variant can, for example, comprise the amino acid sequence of the parent antigen recognizing construct or TCR with at least one conservative amino acid substitution. Conservative amino acid substitutions are known in the art, and include amino acid substitutions in which one amino acid having certain physical and / or chemical properties is exchanged for another amino acid that has the same, or very close, chemical or physical properties. For instance, the conservative amino acid substitution can be an acidic amino acid substituted for another acidic amino acid (e.g., Asp or Glu), an amino acid with a nonpolar side chain substituted for another amino acid with a nonpolar side chain (e.g., Ala, Gly, Vai, He, Leu, Met, Phe, Pro, Trp, Vai, etc.), a basic amino acid substituted for another basic amino acid (Lys, Arg, etc.), an amino acid with a polar side chain substituted for another amino acid with a polar side chain (Asn, Cys, Gin, Ser, Thr, Tyr, etc.), etc.

[0073] The term "sequence identity" or "identity" as used in the present invention means the percentage of pair-wise identical residues, following homology alignment of asequence of a polypeptide and or nucleic acid of the present invention with a sequence in question, with respect to the number of residues in the longer of these two sequences.

[0074] The percentage of sequence homology or sequence identity can, for example, be determined herein using the program BLASTP, version blastp 2.2.5 (November 16, 2002; cf. Altschul, S. F. et al. (1997) Nucl. Acids Res. 25, 3389-3402). The percentage of homology is based on the alignment of the entire polypeptide sequences (matrix: BLOSLIM 62; gap costs: 11.1; cutoff value set to 10-3) including the respective sequences. It is calculated as the percentage of numbers of "positives" (homologous amino acids) indicated as result in the BLASTP program output divided by the total number of amino acids selected by the program for the alignment.

[0075] There is also provided herein a T cell receptor (TCR) having antigenic specificity for Preferentially Expressed Antigen in Melanoma (PRAME), wherein the TCR comprises an a-chain variable region comprising or consisting of the amino acid sequence of SEQ ID No 111 , and wherein the TCR comprises a p-chain variable region comprising or consisting of the amino acid sequence of SEQ ID No. 112. According to an embodiment, the TCR may comprise an a-chain variable region having at least 90%, or at leat 95%, or at least 96%, or at least 97%, or at least 98%, or at least 99%, or 100% identity with the amino acid sequence of SEQ ID No 111, and the TCR may comprise a p-chain variable region having at least 90%, or at leat 95%, or at least 96%, or at least 97%, or at least 98%, or at least 99%, or 100% identity with the amino acid sequence of SEQ ID No. 112.

[0076] The chimeric human CD8 co-receptor

[0077] Turning now to the chimeric human CD8 co-receptor for the use according to the first aspect of the invention, it is contemplated herein that the chimeric CD8 co-receptor may be a single chain polypeptide.

[0078] According to an embodiment, the cytoplasmic polypeptide region of the chimeric human CD8 co-receptor may further comprise at least two TNF receptor associated factor 6 (TRAF 6) binding motifs, and / or said cytoplasmic polypeptide region may comprise at least two CD40 TNF receptor associated factor 1 / 2 / 3 (TRAF 1 / 2 / 3) binding motifs.

[0079] As demonstrated in the present application, this specifically designed cytoplasmic costimulatory domain / region of a chimeric CD8 coreceptor as herein described enables the chimeric human CD8 coreceptor for the use according to the first aspect of the present invention to increase (e.g. synergistically) the costimulatory activity, such that it is particularly suitable in the combination together with the antigen recognizing construct and together with the chimeric human CD95 receptor for the use according to the first aspect.

[0080] According to an embodiment, at least one, or at least two, of the at least two TRAF 6 binding motifs may comprise a CD40 TRAF 6 binding motif.

[0081] According to an embodiment, the at least one cytoplasmic polypeptide region may comprise the at least two TNF receptor associated factor 6 (TRAF 6) binding motifs (such as e.g. two CD40 TRAF6 binding motifs), and may further comprise at least one TNF receptor associated factor 1, 2, 3 (TRAF 1 / 2 / 3) binding motif.

[0082] For example, the at least one TRAF 1 / 2 / 3 binding motif may comprise a CD40 TRAF 1 / 2 / 3 binding motif.

[0083] According to an embodiment, the at least one cytoplasmic polypeptide region may comprise the at least two TNF receptor associated factor 6 (TRAF 6) binding motifs (such as e.g. two CD40 TRAF6 binding motifs) and the least two CD40 TRAF 1 / 2 / 3 binding motifs.

[0084] According to an embodiment, the at least one cytoplasmic polypeptide region may comprise the at least two CD40 TNF receptor associated factor 1 / 2 / 3 (TRAF 1 / 2 / 3) binding motifs and at least one TNF receptor associated factor 6 (TRAF 6) binding motifs (such as at least one CD40 TRAF6 binding motif).

[0085] TRAF 6 binding motifs and TRAF 123 binding motifs have been described in the art (e.g. Park HH, Front. Immunol. 9:1999, 2018). There are also numerous suitable methods known in the art, such as e.g. yeast two hybrid assays or surface plasmon resonance assay) for determining binding of TRAF6 and / or TRAF1 , TRAF2 and TRAF3 to a specific binding motif. Thus, “TRAF6 binding” motif as referred to herein may relate to any polypeptide region which is functional in binding TRAF6. “TRAF 1 / 2 / 3” binding motifs as referred to herein may relate to any polypeptide region which is functional in binding TRAF1, and / or TRAF2, and / or TRAF3.

[0086] “CD40 TRAF6 binding motif” as referred to herein relates to the specific polypeptide sequence of human wildtype CD40 which is necessary and sufficient for binding to TRAF6.

[0087] According to an embodiment, the CD40 TRAF6 binding motif may comprise or may consists of an amino acid sequence having at least 70%, or at least 71%, or at least 72%, or at least 73%, or at least 74%, or at least 75%, or at least 76%, or at least 77%, or at least 78%, or at least 79%, or at least 80%, or at least 81%, or at least 82%, or at least 83%, or at least 84%, or at least 85%, or at least 86%, or at least 87%, or at least 88%, or at least 89% or at least 90%, or at least 91%, or at least 92%, or at least 93%, or at least 94%, or at least 95%, or at least 96% or at least 97%, or at least 98%, or at least 99%, or 100% sequence identity with the amino acid sequence as set forth in Seq ID No:130. For example, the CD40 TRAF6 binding motif may have one or more conservative amino acid substitutions relative to the functional polypeptide domain, region or motif of the human CD40 (Seq ID No. 130). For example, the “TRAF6” binding motif of CD40 as included in the chimeric CD8 co-receptor as herein provided may comprise a polypeptide region of CD40 including amino acid position 229 of SEQ ID No. 141 (relating to the complete polypeptide sequence of human wildtype CD40), and further wherein, at said position, the TRAF6 binding motif may be mutated, optionally wherein said mutation may consist of an exchange of an proline (P) to an alanine (A). In some embodiments, for example, the “TRAF6” binding motif of wildtype CD40 as included in the chimeric CD8 coreceptor as herein provided may comprise a polypeptide region of CD40 including amino acid position 237 of SEQ ID No 141 (wildtype CD40), and further wherein, at said position, the TRAF6 binding motif may be mutated, optionally wherein said mutation may consist of an exchange of an asparagine (N) to an aspartic acid (D). In particular, all amino acid substitutions that maintain the functional activity of the “TRAF6” motif of wildtype CD40 are envisaged.

[0088] For example, the at least one CD40 TRAF6 binding motif may have at least 70%, or at least 71 %, or at least 72%, or at least 73%, or at least 74%, or at least 75%, or at least 76%, or at least 77%, or at least 78%, or at least 79%, or at least 80%, or at least 81 %, or at least 82%, or at least 83%, or at least 84%, or at least 85%, or at least 86%, or at least 87%, or at least 88%, or at least 89% or at least 90%, or at least 91%, or at least 92%, or at least 93%, or at least 94%, or at least 95%, or at least 96% or at least 97%, or at least 98%, or at least 99%, or 100% sequence identity with the functional polypeptide domain or a functional polypeptide motif of a wildtype human CD40 TRAF6 binding motif (e.g. being comprised in or consisting of a polypeptide having an amino acid sequence as set forth in Seq ID No. 130).

[0089] “CD40 TRAF1 / 2 / 3 binding motif” as referred to herein relates to the specific polypeptide sequence of human wildtype CD40 which is necessary and sufficient for binding to TRAF 1 , TRAF 2 and / or TRAF3.

[0090] According to an embodiment, the CD40 TRAF1 / 2 / 3 binding motif may comprise or may consists of an amino acid sequence having at least 70%, or at least 71%, or at least 72%, or at least 73%, or at least 74%, or at least 75%, or at least 76%, or at least 77%, or at least 78%, or at least 79%, or at least 80%, or at least 81%, or at least 82%, or at least 83%, or at least 84%, or at least 85%, or at least 86%, or at least 87%, or at least 88%, or at least 89% or at least 90%, or at least 91%, or at least 92%, or at least 93%, or at least 94%, or at least 95%, or at least 96% or at least 97%, or at least 98%, or at least 99%, or 100%sequence identity with the amino acid sequence as set forth in Seq ID No:131 . For example, the CD40 TRAF 1 / 2 / 3 binding motif may have one ore moreconservative amino acid substitutions relative to the functional polypeptide domain, region or motif of the human CD40 (Seq ID No. 131).

[0091] For example, the at least one CD40 TRAF1 / 2 / 3 binding motif may have at least 70%, or at least 71%, or at least 72%, or at least 73%, or at least 74%, or at least 75%, or at least 76%, or at least 77%, or at least 78%, or at least 79%, or at least 80%, or at least 81%, or at least 82%, or at least 83%, or at least 84%, or at least 85%, or at least 86%, or at least 87%, or at least 88%, or at least 89% or at least 90%, or at least 91 %, or at least 92%, or at least 93%, or at least 94%, or at least 95%, or at least 96% or at least 97%, or at least 98%, or at least 99%, or 100% sequence identity with the functional polypeptide domain or a functional polypeptide motif of a wildtype human CD40TRAF1 / 2 / 3 binding motif (e.g. being comprised in or consisting of a polypeptide having an amino acid sequence as set forth in Seq ID No. 131).

[0092] According to an embodiment, the cytoplasmic polypeptide region of the chimeric CD8 co-receptor may comprise a complete cytoplasmic domain of CD40.

[0093] According to an embodiment, the complete cytoplasmic domain of CD40 may comprise or may consists of an amino acid sequence having at least 70%, or at least 75%, or at least 80%, or at least 85%, or at least 90%, or at least 95%, or at least 97%, or 100% sequence identity with the amino acid sequence as set forth in Seq ID No: 18. For example, the complete cytoplasmic domain of CD40 may have one ore more conservative amino acid substitutions relative to the functional polypeptide domain of the human wildtype CD40 complete cytoplasmic domain (Seq ID No. 18).

[0094] For example, the complete cytoplasmic domain of CD40 may have at least 70%, or at least 71 %, or at least 72%, or at least 73%, or at least 74%, or at least 75%, or at least 76%, or at least 77%, or at least 78%, or at least 79%, or at least 80%, or at least 81 %, or at least 82%, or at least 83%, or at least 84%, or at least 85%, or at least 86%, or at least 87%, or at least 88%, or at least 89% or at least 90%, or at least 91%, or at least 92%, or at least 93%, or at least 94%, or at least 95%, or at least 96% or at least 97%, or at least 98%, or at least 99%, or 100% sequence identity with the functional polypeptide domain of a wildtype human CD40 cytoplasmic domain (e.g. Seq ID No. 129).

[0095] It is unserstood herein that a complete cytoplasmic domain of CD40 comprises both the at least one CD40 TRAF1 / 2 / 3 binding motif, and the at least one CD40 TRAF6 binding motif as referred to herein.

[0096] According to an embodiment, the cytoplasmic polypeptide region may comprise a complete cytoplasmic domain of CD40, a CD40 TRAF6 motif, and a CD40 TRAF 1 / 2 / 3 motif.

[0097] According to a preferred embodiment, the cytoplasmic polypeptide region of the chimeric CD8 co-receptor may comprise - in an N-terminal to C-terminal order - a CD40 TRAF 1 / 2 / 3 binding motif, a CD40 TRAF 6 binding motif, and a further CD40 TRAF 1 / 2 / 3 binding motif.

[0098] It is contemplated that the cytoplasmic polypeptide region may comprise at least one linker between the at least two TRAF6 binding motifs, and / or between the at least two CD40 TRAF1 / 2 / 3 binding motifs, and / or between at least one TRAF1 / 2 / 3 binding motif and at least one TRAF6 motif, and / or between the complete CD40 cytoplasmic domain and at least one TRAF6 domain, and / or between the complete CD40 cytoplasmic domain and at least one TRAF1 / 2 / 3 domain.

[0099] For example, the linker may comprise or consist of an amino acid sequence as set forth by SEQ ID No. 149 (GGGGS), or of an amino acid sequence as set fprth by SEQ ID No. 148 (SGGGS).

[0100] According to an embodiment, the polypeptide of the chimeric CD8 co-receptor may comprise at least one CD8a (-derived) polypeptide region having at least 60% sequence identity with a functional polypeptide domain or a functional polypeptide motif of a human wildtype CD8a co-receptor (e.g. Seq ID No. 113), wherein said at least one CD8a (-derived) polypeptide region comprises said CD8a (-derived) IG-like domain region. According to an embodiment, said polypeptide of the chimeric CD8 co-receptor may comprise at least one CD8p (-derived) polypeptide region having at least 60% sequence identity with a functional polypeptide domain or a functional polypeptide motif of a human wildtype CD8p co-receptor (e.g. Seq ID No. 114), wherein said at least one CD8p (-derived) polypeptide region comprises an CD8p (-derived) IG-like domain region.

[0101] It is understood that the expression of human wildtype CD8a co-receptor relates to a protein having an amino acid sequence according to UniProtKB database entry No. P01732 ■ CD8A_HUMAN, as set forth e.g. in SEQ ID No. 113. It is further understood that the expression of human wildtype CD8p co-receptor relates to a protein having an amino acid sequence according to UniProtKB database entry No. P10966 ■ CD8B_HUMAN, as set forth e.g. in SEQ ID No. 114.

[0102] The term "sequence identity" or "identity" as used in the present invention means the percentage of pair-wise identical residues, following homology alignment of a sequence of a polypeptide and or nucleic acid of the present invention with a sequence in question, with respect to the number of residues in the longer of these two sequences.

[0103] The percentage of sequence homology or sequence identity can, for example, be determined herein using the program BLASTP, version blastp 2.2.5 (November 16, 2002; cf. Altschul, S. F. et al. (1997) Nucl. Acids Res. 25, 3389-3402). The percentage of homology is based on the alignment of the entire polypeptide sequences (matrix: BLOSLIM 62; gap costs: 11.1; cutoff value set to 10-3) including the respective sequences. It is calculated as the percentage of numbers of "positives" (homologous amino acids) indicated as result in the BLASTP program output divided by the total number of amino acids selected by the program for the alignment.

[0104] It is noted in this context that it has been found that the chimeric human CD8 Co-receptor polypeptide comprising both an CD8a IG-like domain region together with an CD8 IG-like domain region in combination with the specific costimulatory cytoplasmic domain as herein described is able to enhance T-cell activation, proliferation, cytokine production, and cytotoxicity, thereby ultimately improving the therapeutic efficacy of TCR-T-cell therapy in the specific combination with the antigen recognizing construct capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide, and with the chimeric human CD 95 receptor. The provision of a chimeric CD8 co-receptor polypeptide comprising both an CD8a IG-like domain region together with an CD8p IG-like domain region in combination with the costimulatory domain as described above allows for advantages in terms of viral vector packaging efficiency and knock-in (KI) efficiency, e.g. if non-viral methods such as electroporation and plasmids are used, by utilizing a smaller chimeric CD8 co-receptor polypeptide. Co-engineering of cells in combination with the antigen recognizing construct capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide, and with the chimeric human CD 95 receptor for the use as herewith disclosed may consequently be more effective.

[0105] According to an embodiment, the at least one CD8a polypeptide region has at least 70%, or at least 75%, or at least 80%, or at least 85%, or at least 90%, or at least 95%, or at least 97% sequence identity with the functional polypeptide domain or a functional polypeptide motif of a wildtype human CD8a co-receptor (e.g. Seq ID No. 113).

[0106] For example, the at least one CD8a polypeptide region may have at least 70%, or at least 71 %, or at least 72%, or at least 73%, or at least 74%, or at least 75%, orat least 76%, or at least 77%, or at least 78%, or at least 79%, or at least 80%, or at least 81 %, or at least 82%, or at least 83%, or at least 84%, or at least 85%, or at least 86%, or at least 87%, or at least 88%, or at least 89% or at least 90%, or at least 91%, or at least 92%, or at least 93%, or at least 94%, or at least 95%, or at least 96% or at least 97%, or at least 98%, or at least 99%, or 100% sequence identity with the functional polypeptide domain or a functional polypeptide motif of a wildtype human CD8a co-receptor (e.g. Seq ID No. 113).

[0107] According to an embodiment, the at least one CD8p polypeptide region has at least 70%, or at least 75%, or at least 80%, or at least 85%, or at least 90%, or at least 95%, or at least 97% sequence identity with the functional polypeptide domain or a functional polypeptide motif of a wildtype human CD8p co-receptor (Seq ID No. 114).

[0108] For example, the at least one CD8p polypeptide region may have at least 70%, or at least 71 %, or at least 72%, or at least 73%, or at least 74%, or at least 75%, or at least 76%, or at least 77%, or at least 78%, or at least 79%, or at least 80%, or at least 81 %, or at least 82%, or at least 83%, or at least 84%, or at least 85%, or at least 86%, or at least 87%, or at least 88%, or at least 89% or at least 90%, or at least 91%, or at least 92%, or at least 93%, or at least 94%, or at least 95%, or at least 96% or at least 97%, or at least 98%, or at least 99%, or 100% sequence identity with the functional polypeptide domain or a functional polypeptide motif of a wildtype human CD8p co-receptor (e.g. Seq ID No. 114).

[0109] The at least one CD8a polypeptide region having at least 60% sequence identity with CD8a IG-like domain region of a human wildtype CD8a co-receptor may have generally a sufficient portion of the human wildtype CD8a IG-like domain region to be able to bind to MHC. For example, said at least one CD8a polypeptide region having at least 60% sequence identity with CD8a IG-like domain region of a human wildtype CD8a Co-receptor may comprise the complete or a considerable part of the human wildtype CD8a IG-like domain region, as set forth e.g. in SEQ ID NO.: 116, but in an embodiment may comprise at least 10, 20, 30, 40, 50, 60, 70, 80, 80, 100, or 110 amino acids of the human wildtype CD8a IG-like domain region. Thus, the expression “Ig-like domain” as used herein may - in principle - refer to a polypeptide region that is homologous to the V and / or C domains in immunoglobuline proteins.

[0110] The at least one CD8p polypeptide region having at least 60% sequence identity with CD8p IG-like domain region of a human wildtype CD8p co-receptor may have generally a sufficient portion of the human wildtype CD8p IG-like domain region to be ableto bind to MHC. For example, said at least one CD8p polypeptide region having at least 60% sequence identity with CD8p IG-like domain region of a human wildtype CD8p coreceptor may comprise the complete or a considerable part of the human wildtype CD8p IG-like domain region, as set forth e.g. in SEQ ID NO.: 117, but in an embodiment may comprise at least 10, 20, 30, 40, 50, 60, 70, 80, 80, 100, or 110 amino acids of the human wildtype CD8p IG-like domain region. Thus, the expression “Ig-like domain” as used herein may - in principle - refer to a polypeptide region that is homologous to the V and / or C domains in immunoglobuline proteins.

[0111] According to an embodiment, the chimeric human CD8 co-receptor polypeptide may be a single-chain polypeptide.

[0112] According to an embodiment, the chimeric CD8 co-receptor comprises the CD8a IG-like domain region, the CD8p IG-like domain region, a stalk region, a transmembrane domain region; and an intracellular / cytoplasmic domain region, wherein the intracellular domain region comprises a palmitoylation motif region and a LCK binding site region.

[0113] The expressions “domain region”, “binding site region”, “motif region” as used herein are understood to relate to e.g. a region of the chimeric receptor which is necessary and / or sufficient for a biological function of the chimeric receptor, or to a region of the chimeric receptor (e.g. of the chimeric CD8 co-receptor) which is defined e.g. by a localization with respect to a cell, or to a structurally defined unit of the chimeric receptor polypeptide. Furthermore, the expression “cytoplasmic polypeptide domain” and “cytoplasmic polypeptide motif’ as used herein may be understood as relating to a region of the chimeric receptor which is defined by its localization in the cytoplasm of a cell, and which is necessary and / or sufficient for a biological function of the chimeric receptor.

[0114] According to an embodiment, the at least one CD8 polypeptide region further comprises at least one polypeptide region selected from the group consisting of a stalk domain region, a transmembrane domain region, and a LCK binding site motif region.

[0115] According to another embodiment, the at least one CD8p polypeptide region further comprises at least one polypeptide region selected from the group consisting of a stalk domain region, a transmembrane domain region, a cytoplasmic region, and a palmitoylation motif region.

[0116] Thus, it is contemplated that the chimeric CD8 co-receptor polypeptide may comprise, in addition to comprising the CD8a IG-like domain region and the CD8p IG-like domain region, further domain regions / motif regions / binding site regions from one or both of a wildtype human CD8a co-receptor and / or a wildtype human CD8p co-receptor in every conceivable combination to establish a functional chimeric CD8 co-receptor polypeptide, together with the specific cytoplasmic costimulatory domain as herein described.

[0117] For example, it has been described that wildtype human CD8a co-receptor comprises a stalk domain region, a transmembrane domain region, and a LCK binding site motif region. Furthermore, it has been reported that the human wildtype CD8p- co-receptor comprises a stalk domain region, a transmembrane domain region, and a palmitoylation motif region (e.g. Wong et al.; 2003).

[0118] For example, the expression “wildtype human CD8a co-receptor stalk domain region” as referred to herein may relate to a polypeptide having an amino acid sequence as set forth in SEQ ID NO: 118. For example, the expression “wildtype human CD8a co-receptor stalk domain region” as referred to herein may relate to a polypeptide comprising amino acid sequence 136-182 of UniProtKB database entry No. P01732 ■ CD8A_HUMAN, as set forth e.g. in SEQ ID No. 113. Thus, the expression “stalk region” as used herein may refer to a protein portion that preserves the ability to propagate signal transduction of a functional CD8 co-receptor analogous to a wildtype human CD8a co-receptor stalk domain region. For example, the expression “wildtype human CD8a co-receptor transmembrane domain region” as referred to herein may relate to a polypeptide having an amino acid sequence as set forth in SEQ ID NO: 119. For example, the expression “wildtype human CD8a co-receptor transmembrane domain region” as referred to herein may relate to a polypeptide comprising amino acid sequence 183-203 of UniProtKB database entry No. P01732 ■ CD8A_HUMAN, as set forth e.g. in SEQ ID No. 113. For example, the expression “wildtype human CD8a co-receptor LCK binding site motif region” as referred to herein may relate to a polypeptide having an amino acid sequence as set forth in SEQ ID NO: 120. For example, the expression “wildtype human CD8a co-receptor LCK binding site motif domain region” as referred to herein may relate to a polypeptide comprising amino acid sequence CKCP at amino acid positions 215-218 of UniProtKB database entry No. P01732 ■ CD8A_HUMAN, as set forth e.g. in SEQ ID No. 113. Thus, the expression “LCK binding site” as used herein may refer to a protein portion that preserves the ability to recruite the Src family kinase Lek of a functional CD8 Co-receptor.

[0119] For example, the expression “wildtype human CD8p co-receptor stalk domain region” as referred to herein may relate to a polypeptide having an amino acid sequenceas set forth in SEQ ID NO: 121. For example, the expression “wildtype human CD8p coreceptor stalk domain region” as referred to herein may relate to a polypeptide comprising amino acid sequence 133-170 of UniProtKB database entry No. P10966 ■ CD8B_HUMAN, as set forth e.g. in SEQ ID No. 114. Thus, the expression “stalk region” as used herein may refer to a protein portion that preserves the ability to propagate signal transduction of a functional CD8 co-receptor analogous to a wildtype human CD8p co-receptor stalk domain region. For example, the expression “wildtype human CD8p co-receptor transmembrane domain region” as referred to herein may relate to a polypeptide having an amino acid sequence as set forth in SEQ ID NO: 122. For example, the expression “wildtype human CD8p co-receptor transmembrane domain region” as referred to herein may relate toa polypeptide comprising amino acid sequence 171-191 of UniProtKB database entry No. P10966 ■ CD8B_HUMAN, as set forth e.g. in SEQ ID No. 114. For example, expression “the wildtype human CD8p co-receptor palmitoylation motif region” as referred to herein may relate to a polypeptide having an amino acid sequence as set forth in SEQ ID NO: 123. For example, the expression “wildtype human CD8p co-receptor palmitoylation motif region” as referred to herein may relate to the two conserved amino acids Cysteine at amino acid positions 194 and 195 with respect to the amino acid sequence of UniProtKB database entry No. P10966 ■ CD8B_HUMAN, as set forth e.g. in SEQ ID No. 114. Thus, the expression “palmitoylation motif region” as used herein may refer to a protein portion that preserves the ability to propagate signal transduction of a functional CD8 co-receptor analogous to a wildtype human CD8p palmitoylation motif region region.

[0120] According to an embodiment, the chimeric CD8 co-receptor may further comprise at least one CD4 (-derived) polypeptide region having at least 60% sequence identity with a functional polypeptide domain or a functional polypeptide motif of a human CD4 co-receptor.

[0121] It is understood that the expression of human wildtype CD4 co-receptor relates to a protein having an amino acid sequence according to UniProtKB database entry No. P01730 ■ CD4_HUMAN, as set forth e.g. in SEQ ID No. 115.

[0122] According to an embodiment, the at least one CD4polypeptide region has at least 70%, or at least 75%, or at least 80%, or at least 85%, or at least 90%, or at least 95%, or at least 97% sequence identity with the functional polypeptide domain or a functional polypeptide motif of a wildtype human CD4 co-receptor (Seq ID No. 115).

[0123] For example, the at least one CD4 polypeptide region may have at least 70%, or at least 71%, or at least 72%, or at least 73%, or at least 74%, or at least 75%, or at least 76%, or at least 77%, or at least 78%, or at least 79%, or at least 80%, or at least 81 %, or at least 82%, or at least 83%, or at least 84%, or at least 85%, or at least 86%, or at least 87%, or at least 88%, or at least 89% or at least 90%, or at least 91%, or at least 92%, or at least 93%, or at least 94%, or at least 95%, or at least 96% or at least 97%, or at least 98%, or at least 99%, or 100% sequence identity with the functional polypeptide domain ora functional polypeptide motif of a wildtype human CD4 co-receptor (Seq ID No.115).

[0124] Thus, it is contemplated that the chimeric CD8 co-receptor polypeptide may comprise, in addition to comprising the CD8a IG-like domain region and the CD8p- IG-like domain region, at least one domain region / motif region / binding site region from a wildtype human CD4 co-receptor. Such at least one CD4 domain region / motif region / binding site region may be combined with further CD4 domain region / motif region / binding site region and / or further domain regions / motif regions / binding site regions from one or both of a wildtype human CD8a co-receptor and / or a wildtype human CD8p co-receptor in every conceivable combination to establish a functional chimeric CD8 co-receptor polypeptide. Without being bound to theory, it is herewith contemplated that inclusion of a CD4 co-receptor derived functional polypeptide domain / motif region / binding site region into the chimeric CD8 co-receptor may contribute to incorporating CD4 T-cells into TCR-T-cell therapy.

[0125] As described herein, the expressions “CD8a-derived polypeptide region”, “CD8p-derived polypeptide region” and “CD4 derived polypeptide region” may be used herein interchangeably with the expressions “CD8a polypeptide region”, “CD8p polypeptide region” and “CD4 polypeptide region”, respectively, to refer to a (functional) portion of a wildtype CD8a, a wildtype CD8p ora wildtype CD4 protein (or variants thereof), respectively. With respect to variants of wildtype CD8a, wildtype CD8p or wildtype CD4 protein portions, such a variant may have at least 60% sequence identity (e.g., at least 70%, 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or more) to the corresponding portion of the amino acid sequence of SEQ ID NO: 113, SEQ ID NO: 114 or SEQ ID NO 115, respectively. For example, variants of wildtype CD8a as herein described may comprise an extracellular domain that preserves the ability to bind peptide-MCH class I. Suitable methods (such as e.g. surface plasmon resonance assay) for determining said functional ability are known to the skilled person. For example, variants of wildtype CD8p as herein described may comprise an extracellular domain that preserves the ability to bind peptide-MCH class I. Suitable methods (such as e.g. surfaceplasmon resonance assay) for determining said functional ability are known to the skilled person. For example, variants of wildtype CD4 protein portions as described herein may comprise a CD4 stalk, a CD4 transmembrane and / or a CD4 cytoplasmic domain that preserve the ability to propagate signal transduction. Suitable methods for determining said functional ability are known to the skilled person (e.g. as described by [Capone M., et al., 2021]).

[0126] The wildtype human CD4 co-receptor polypeptide is known to comprise -among others - a stalk domain region, a transmembrane domain region, and a cytoplasmic region comprising a palmitoylation motif region and an LCK binding site region.

[0127] For example, the expression “wildtype human CD4 co-receptor stalk domain region” as referred to herein may relate to a polypeptide having an amino acid sequence as set forth in SEQ ID NO: 124. For example, the expression “wildtype human CD4 co-receptor stalk domain region” as referred to herein may relate to a polypeptide comprising amino acid sequence 375-396 of UniProtKB database entry No. P01730 ■ CD4_HUMAN, as set forth e.g. in SEQ ID No. 115. For example, the expression “wildtype human CD4 co-receptor transmembrane domain region” as referred to herein may relate to a polypeptide having an amino acid sequence as set forth in SEQ ID NO: 125. For example, the expression “wildtype human CD4 co-receptor transmembrane domain region” as referred to herein may relate to a polypeptide comprising amino acid sequence 397-418 of UniProtKB database entry No. P01730 ■ CD4_HUMAN, as set forth e.g. in SEQ ID No.115. For example, the expression “wildtype human CD4 co-receptor LCK binding site motif region” as referred to herein may relate to a polypeptide having an amino acid sequence as set forth in SEQ ID NO: 126. For example, the expression “wildtype human CD4 co-receptor LCK binding site motif domain region” as referred to herein may relate to a polypeptide comprising amino acid sequence 442-449 (KKTCQCPH) of UniProtKB database entry No. P01730 ■ CD4_HUMAN, as set forth e.g. in SEQ ID No. 115. For example, the expression “wildtype human CD4 co-receptor palmitoylation motif region” as referred to herein may relate to a polypeptide having an amino acid sequence as set forth in SEQ ID NO: 127. For example, the expression “wildtype human CD4 co-receptor palmitoylation motif region” as referred to herein may relate to a polypeptide comprising amino acid sequence 419-422 (CVRC) of UniProtKB database entry No. P01730 ■ CD4_HUMAN, as set forth e.g. in SEQ ID No. 115.

[0128] According to an embodiment, the at least one CD4 polypeptide region comprises at least one polypeptide region selected from the group consisting of a stalkdomain region, a transmembrane domain region, a cytoplasmic domain, a palmitoylation motif region, and a LCK binding site region.

[0129] In certain embodiments, the chimeric CD8 co-receptor polypeptide may comprise a CD4 polypeptide region comprising a wildtype human CD4 co-receptor cytoplasmic region including a palmitoylation motif region and an LCK binding site region. For example, the expression “wildtype human CD4 co-receptor cytoplasmic region” as referred to herein may relate to a polypeptide having an amino acid sequence as set forth in SEQ ID NO: 128. For example, the expression “wildtype human CD4 co-receptor cytoplasmic region” as referred to herein may relate to a polypeptide comprising amino acid sequence 419 - 458 of UniProtKB database entry No. P01730 ■ CD4_HUMAN, as set forth e.g. in SEQ ID No. 115.

[0130] According to a preferred embodiment, the chimeric CD8 co-receptor comprises, in an N-terminal to C-terminal order, at least the following polypeptide regions: a CD8a IG-like domain, a CD8p IG-like domain, a CD8p stalk region, a CD8a transmembrane region, a complete cytoplasmic domain from CD40, a first CD40 TRAF 1 / 2 / 3 binding motif, a CD40 TRAF 6 binding motif, and a second CD40 TRAF 1 / 2 / 3 binding motif.

[0131] According to an embodiment, the cytoplasmic polypeptide region of the chimeric CD8 co-receptor may comprise or consist of a polypeptide having an amino acid sequence having at least at least 70%, or at least 71%, or at least 72%, or at least 73%, or at least 74%, or at least 75%, or at least 76%, or at least 77%, or at least 78%, or at least 79%, or at least 80%, or at least 81%, or at least 82%, or at least 83%, or at least 84%, or at least 85%, or at least 86%, or at least 87%, or at least 88%, or at least 89% or at least 90%, or at least 91%, or at least 92%, or at least 93%, or at least 94%, or at least 95%, or at least 96% or at least 97%, or at least 98%, or at least 99%, or 100% sequence identity with the amino acid sequence as set forth in Seq ID No: 132.

[0132] According to a preferred embodiment, the chimeric human CD8 co-receptor, may comprise or consist of a polypeptide having an amino acid sequence having at least 70%, or at least 71 %, or at least 72%, or at least 73%, or at least 74%, or at least 75%, or at least 76%, or at least 77%, or at least 78%, or at least 79%, or at least 80%, or at least 81 %, or at least 82%, or at least 83%, or at least 84%, or at least 85%, or at least 86%, or at least 87%, or at least 88%, or at least 89% or at least 90%, or at least 91%, or at least 92%, or at least 93%, or at least 94%, or at least 95%, or at least 96% or at least 97%, orat least 98%, or at least 99%, or 100% sequence identity with the amino acid sequence as set forth in Seq ID No: 133.

[0133] The chimeric human CD95 receptor

[0134] Turning now to the chimeric human CD95 receptor for the use according to the first aspect of the invention, according to an embodiment, the polypeptide of the chimeric human CD95 receptor may comprise at least one CD95 polypeptide region comprising or consisting of a polypeptide domain, a polypeptide region or a polypeptide motif of a human CD95 receptor as set forth in SEQ ID No 134, or said polypeptide may comprise at least one CD95 polypeptide region having at least 60% sequence identity with a polypeptide domain, a polypeptide region or a polypeptide motif of a human CD95 receptor as set forth in SEQ ID No. 134, wherein said CD95 polypeptide region comprises the CD95 extracellular ligand binding domain.

[0135] According to an embodiment, the polypeptide of the chimeric human CD95 receptor may further comprise at least two non-CD95 cytoplasmic co-stimulatory polypeptide domains, regions or motifs, wherein a first non-CD95 co-stimulatory region may comprise a cytoplasmic costimulatory region of 0X40, and wherein a second non-CD95 co-stimulatory region may comprise a cytoplasmic costimulatory region of CD40.

[0136] In accordance with an embodiment, the extracellular ligand binding domain of the chimeric CD95 receptor may be functional in binding FAS-ligand (CD95L) or any other protein / polypeptide having the ability of binding to the wildtype CD95 receptor ligand binding domain. In the art, numerous suitable assays for testing FAS-ligand binding functionality have been described. Any suitable method may be selected.

[0137] In some embodiments, the polypeptide may be a single-chain polypeptide.

[0138] For example, the chimeric CD95 receptor may have one, two, or three cysteine-rich domains (CRDs) of a human CD95 receptor, said one, two, or three CRDs selected from the group consisting of SEQ ID NO: 135, SEQ ID NO: 136, and SEQ ID NO: 137, wherein said human CD95 polypeptide region comprises a CD95 extracellular ligand binding domain; further wherein said polypeptide comprises a first non-CD95 co-stimulatory region of CD40, and a second non-CD95 cytoplasmic costimulatory region of 0X40. A particular example for a CRD to be included in the chimeric CD95 receptor may be the CRD as set forth in SEQ ID NO: 137.

[0139] It is understood that the expression of human wildtype CD95 receptor relates to a protein having an amino acid sequence according to UniProtKB database entry No. P25445 ■ TNR6_HUMAN, as set forth e.g. in SEQ ID No. 134.

[0140] It is noted in this context that it has been found here that the chimeric CD95 receptor comprising both a functional extracellular CD95 receptor ligand binding domain and a first non-CD95 co-stimulatory region comprising a cytoplasmic costimulatory regionof 0X40 in combination with a second non-CD95 co-stimulatory region comprising a cytoplasmic costimulatory region of CD40, is - in the specific combination together with the PRAME antigen recognizing construct as herein described, and together with the chimeric human CD8 Co-receptor as herein described- able to turn negative signals (e.g. present in a tumor microenvironment) into positive signals for T-cell activation, if all three constructs are used in a method of treating a disease in a subject, wherein the method comprises administering a therapeutically effective amount of the antigen recognizing construct, of the chimeric human CD8 Co-receptor, and of the chimeric human CD95 receptor.

[0141] Advantageously, replacing e.g. at least the cytoplasmic “death domain” of wildtype CD 95 (amino acids 230-314 of SEQ ID No. 134), which is characterized by its ability to recruite Fas-Associated Death Domain-Containing Protein (FADD) through homotypic interactions, thereby potentially initiating apoptosis, or replacing e.g. the complete cytoplasmic domain of CD95 by a at least one non-CD95-derived co-stimulatory cytoplasmic polypeptide domain as herein provided enables the T-cell comprising the chimeric CD95 receptor and the engineered T-cell receptor, in further presence of the chimeric CD8 Co-receptor, to bypass the inhibitory effects of FASL expressed by the tumor microenvironment, thus creating resistance to tumor-mediated immune suppression. Secondly, the chimeric CD95 receptors as herein described is thus believed to act as a molecular switch, redirecting the signaling pathways triggered by FAS engagement with FASL. Instead of inducing apoptosis and T cell death, the fusion of the CD95 receptor ligand binding domain to the co-stimulatory domain as herein described, comprising both an 0X40 polypeptide region and a CD40 polypeptide region, alters the intracellular signaling events, promoting T cell activation, persistence and enhanced anti-tumor responses.

[0142] According to an embodiment, the extracellular ligand binding domain of the chimeric CD95 receptors as herein described for the use according to the first aspect of the invention may be functional in binding FAS-ligand (CD95L) or any other protein / polypeptide having the ability of binding to the wildtype CD95 receptor ligand binding domain. Only for example, e.g. scFv and Fab have been described to have the capacity of binding to the ligand binding domain of wildtype CD95 receptor.

[0143] According to an embodiment, the at least one CD95 (-derived) polypeptide region may have at least 70%, or at least 75%, or at least 80%, or at least 85%, or at least 90%, or at least 95%, or at least 97% sequence identity with the functional polypeptide domain or a functional polypeptide motif of a wildtype human CD95 receptor (e.g. Seq ID No. 134). According to an embodiment, the at least one CD95 (-derived) polypeptideregion may have one or more conservative amino acid substitutions relative to the amino acid sequence of the wildtype CD95 receptor.

[0144] For example, the at least one CD95 (-derived) polypeptide region may have at least 70%, or at least 71%, or at least 72%, or at least 73%, or at least 74%, or at least 75%, or at least 76%, or at least 77%, or at least 78%, or at least 79%, or at least 80%, or at least 81 %, or at least 82%, or at least 83%, or at least 84%, or at least 85%, or at least 86%, or at least 87%, or at least 88%, or at least 89% or at least 90%, or at least 91 %, or at least 92%, or at least 93%, or at least 94%, or at least 95%, or at least 96% or at least 97%, or at least 98%, or at least 99%, or 100% sequence identity with the functional polypeptide domain or a functional polypeptide motif of a wildtype human CD95 receptor (e.g. Seq ID No. 134).

[0145] The human CD95 polypeptide region comprising a CD95 extracellular ligand binding domain may have generally a sufficient portion of the human wildtype CD95 extracellular ligand binding domain to be functional in binding FASL. For example, said at least one CD95-derived polypeptide region having at least 60% sequence identity with CD95-derived FASL binding domain of a human wildtype CD95 receptor may comprise the complete or a considerable part of the human wildtype CD95 FASL binding domain and / or all amino acids at respective amino acid positions of the wildtype CD95 receptor that are necessary and sufficient for FASL binding to the CD95 receptor.

[0146] The expressions “domain region”, “binding site region”, “motif region” as used herein are understood to relate to e.g. a region of the chimeric CD95 receptor polypeptide which is necessary and / or sufficient for a biological function of the chimeric receptor, or to a region of the chimeric CD95 receptor which is defined e.g. by a localization with respect to a cell, or to a structurally defined unit of the chimeric CD95 receptor polypeptide. Furthermore, the expression “cytoplasmic polypeptide domain” and “cytoplasmic polypeptide motif” as used herein may be understood as relating to a region of the chimeric CD95 receptor which is defined by its localization in the cytoplasm of a cell, and which is necessary and / or sufficient for a biological function of the chimeric receptor.

[0147] The chimeric CD95 receptor polypeptide may be a single-chain polypeptide.

[0148] According to an embodiment, as mentioned above, the CD95 (-derived) polypeptide region may comprise CRD 1, CRD 2 and CRD 3 of the wildtype CD95 receptor. It is herewith envisaged that CRD 1, 2 and 3 polypeptide regions as included in the chimeric CD95 receptor of the T-cell as herein provided may have one or more conservative amino acid substitutions relative to the CRD 1, 2, and 3 polypeptide regions of the wildtype CD95 receptor. In particular, all amino acid substitutions that maintain the functional activity of the wildtype CRD domains are envisaged. For example, (Gary C. Starling et al., 1998), which is incorporated by reference herein in its entirety, discloses amutagenesis study for identifying amino acid residues contributing to the Fas-FasL interaction. The skilled person is therefore aware of protein portions of CRD 1 , CRD 2 and / or CRD 3 of the wildtype CD95 receptor and potential amino acid substitutions that maintain functional activity. For example, the CD95 polypeptide region may comprise CRD 2 of the CD95 wildtype receptor as referred to herein may relate to a polypeptide having an amino acid sequence as set forth in SEQ ID NO: 136 (optionally having one or more conservative amino acid substitutions relative to the CRD 2 polypeptide regions of the wildtype CD 95 receptor).

[0149] According to an embodiment, the at least one CD95 (-derived) polypeptide region may further comprise an extracellular N-terminal pre ligand assembly domain (PLAD) region. The N-Terminal PLAD region of the CD95 wildtype receptor as referred to herein may relate to a polypeptide comprising amino acid sequence 17-82 of UniProtKB database entry No. P25445 ■ TNR6_HUMAN, as set forth e.g. in SEQ ID No. 134. For example, the N-terminal PLAD region as referred to herein may relate to a polypeptide having an amino acid sequence as set forth in SEQ ID NO: 138. It is herewith envisaged that the PLAD region as included in the chimeric CD95 receptor of the T-cell as herein provided may have one or more conservative amino acid substitutions relative to the PLAD region of the wildtype CD95 receptor. In particular, all amino acid substitutions that maintain the functional activity of the wildtype PLAD region are envisaged.

[0150] According to another embodiment, the at least one CD95-derived polypeptide region may further comprise a CD95 wildtype homotypic interaction domain. CD95 homotypic interaction domain as referred to herein may relate to a polypeptide comprising amino acid sequence 59 - 82 of UniProtKB database entry No. P25445 ■ TNR6_HUMAN, as set forth e.g. in SEQ ID No. 134. For example, the homotypic interaction domain region as referred to herein may relate to a polypeptide having an amino acid sequence as set forth in SEQ ID NO: 139. It is herewith envisaged that the homotypic interaction domain as included in the chimeric CD95 receptor of the T-cell as herein provided may have one or more conservative amino acid substitutions relative to the homotypic interaction domain of the wildtype CD95 receptor. In particular, all amino acid substitutions that maintain the functional activity of the wildtype homotypic interaction domain are envisaged.

[0151] According to another embodiment, the at least one CD95 (-derived) polypeptide region may comprise a CD95-derived transmembrane region. The transmembrane domain of the CD95 wildtype receptor as referred to herein may relate to a polypeptide comprising amino acid sequence 174-190 of UniProtKB database entry No. P25445 ■ TNR6_HUMAN, as set forth e.g. in SEQ ID No. 134. For example, the transmembrane domain of wildtype CD95 as referred to herein may relate to a polypeptide having an amino acid sequence as set forth in SEQ ID NO: 140. It is herewith envisagedthat the transmembrane domain as included in the chimeric CD95 receptor of the T-cell as herein provided may have one or more conservative amino acid substitutions relative to the transmembrane domain of the wildtype CD95 receptor. In particular, all amino acid substitutions that maintain the functional activity of the wildtype transmembrane domain are envisaged.

[0152] Thus, it is contemplated that the chimeric CD95 receptor polypeptide as herein provided may comprise, in addition to comprising the CD95 (-derived) ligand binding domain, and in addition to comprising the at least two cytoplasmic costimulatory domains of 0X40 and CD40, further domain regions / motif regions / binding site regions from a wildtype human CD95 receptor in every conceivable combination to establish a functional chimeric CD95 receptor polypeptide. “Functional” chimeric CD 95 receptor in this context relates to a chimeric CD95 receptor that is capable of redirecting the signaling pathways triggered by FAS engagement with FASL such that - instead of inducing apoptosis and T-cell death - binding of FASL promotes T-cell activation, persistence and enhanced anti-tumor responses of the T-cell as herein provided. For example, an optional test for functionality of a chimeric CD95 receptor may be an in-vitro T-cell killing assay as described e.g. by Kalbasi, A., Siurala, M., Su, L.L. et al. “Potentiating adoptive cell therapy using synthetic IL-9 receptors”. Nature 607, 360-365 (2022) using cells expressing FASL. Thus, the expression “every conceivable combination” of CD95 receptor regions as described above is meant to exclude a combination with wildtype human CD95 receptor domains / regions / motifs being inhibitory and / or promoting apoptosis and T-cell death. For example, the chimeric CD95 receptor polypeptide of the T-cell as herein provided may lack the “death domain” of wildtype CD95 (amino acids 230-314 of SEQ ID No. 1), or may merely comprise an altered “death domain” which no longer functions in promoting cell death and / or apoptosis of the T-cell, e.g. due to mutations which abolish any inhibitory and / or apoptotic and / or cell-death promoting functionality of the “death domain”.

[0153] According to an embodiment, the CD95 (-derived) polypeptide region may comprise a complete wildtype CD95 receptor extracellular domain. For example, the expression “wildtype CD95 receptor extracellular domain” as referred to herein may relate to a polypeptide comprising amino acid sequence 26-173 of UniProtKB database entry No. P25445 ■ TNR6_HUMAN, as set forth e.g. in SEQ ID No. 134. Specifically, the wildtype CD95 receptor extracellular domain is depicted herein by SEQ ID No. 150. It is herewith envisaged that the CD95 receptor extracellular domain as included in the chimeric CD95 receptor of the T-cell as herein provided may have one or more conservative amino acid substitutions relative to the extracellular domain of the wildtype CD95 receptor. In particular, all amino acid substitutions that maintain the functional activity of the wildtype CD95 extracellular domain are envisaged.

[0154] According to an embodiment, the CD95-derived polypeptide region may comprise a complete wildtype CD95 receptor extracellular domain, and an entire CD95-derived transmembrane domain.

[0155] According to some embodiments, the transmembrane domain of the chimeric CD95 receptor of the T-cell as herein described for the use according to the first aspect of the invention may be derived from other proteins which comprise a transmembrane domain. For example, the transmembrane domain may be derived from a co-stimulatory molecule. In principle, any transmembrane domain which is functional and allows surface detectable expression of the chimeric CD95 receptor is herewith envisaged.

[0156] The chimeric CD95 receptor of the T-cell as herein provided may further comprise at least one linker region. This may be e.g. a polypeptide linker region. Such linker(s) may be included e.g. between functional domains / regions / motifs of the chimeric CD95 receptor. It may be a linker region naturally occurring e.g. in wildtype CD95 receptor, or e.g. in co-stimulatory proteins, e.g. in costimulatory proteins from which the costimulatory domain of the receptor is derived. For example, polypeptide linker regions may be included between the transmembrane domain and the ligand binding domain, and / or between the transmembrane domain and a CRD domain of the chimeric CD95 receptor, and / or between individual CRDs (in embodiments comprising more than one CRD), and / or between the transmembrane domain and the intracellular co-stimulatory domain, and / or between individual co-stimulatory domains (in embodiments comprising more than one co-stimulatory domains).

[0157] Such linker region may comprise 1-100 amino acids, or e.g. 1-80 amino acids, or e.g. 1-50 amino acids, or e.g. 5-100 amino acids.

[0158] According to an embodiment, a linker region of the chimeric CD95 receptor of the T-cell as herein provided may comprise the amino acid sequence as set forth in SEQ ID No. 9 (GGGS)n or as set forth in Seq ID No. 149 (GGGGS)n, or SEQ ID No 148 (SGGGS)n, wherein n is between 0 and 20, or wherein n is between 0 and 10, or where n is between 0 and 5, or where n is between 3 and 5, or wherein n is 1.

[0159] However, in principle, each (polypeptide) linker known in the art is herewith envisaged as being potentially included in the chimeric CD95 switch receptor as described in context with the use according to the first aspect of the invention.

[0160] Turning now in more detail to the co-stimulatory domain, according to the use of the chimeric CD95 receptor in combination with the PRAME recognizing construct and the chimeric human CD8 Co-receptor for the use according to the first aspect, the chimeric CD95 receptor polypeptide may comprise at least two non-CD95 cytoplasmic co-stimulatory polypeptide domains, regions or motifs, wherein a first non-CD95 co-stimulatory region comprises a cytoplasmic costimulatory region of 0X40, and wherein asecond non-CD95 co-stimulatory region comprises a cytoplasmic costimulatory region of CD40. The authors have found for the first time that the specific combination of costimulatory regions / motifs as herein described may be fused to the at least one CD95-derived polypeptide region comprising a functional CD95 receptor extracellular ligand binding domain in order to generate a functional chimeric CD95 switch receptor capable of redirecting the signaling pathways triggered by FAS engagement with FASL such that -instead of inducing apoptosis and T-cell death - binding of FASL promotes T-cell activation, persistence and enhanced anti-tumor responses of the T-cell as herein provided, when the T-cell at the same time comprises / expresses an engineered T-cell receptor. Thus, for example, the expression “T-cell co-stimulatory domain or motif” as used herein may relate to a protein portion which is necessary and / or sufficient to preserve the ability to propagate a co-stimulatory signal in a co-stimulatory molecule / protein as herein described. For example, Hong Ye et al., 1999, which is incorporated herein by reference, discloses - by way of example - TRAF2 binding sites of diverse TNF-family members that may serve as T-cell co-stimulatory domain or motif in the sense of the present application. The skilled person is further aware of numerous further “T-cell co-stimulatory domain or motif’ that have been described in literature.

[0161] According to an embodiment, the chimeric CD95 receptor as herein provided may comprise e.g. the complete cytoplasmic region of CD40, and / or e.g the complete cytoplasmic region of 0X40. According to an embodiment, e.g. the first non-CD95 co-stimulatory region as herein defined may comprise the complete cytoplasmic costimulatory region of 0X40, and / or the second non-CD95 co-stimulatory region as herein defined may comprise the complete cytoplasmic costimulatory region of 0X40.

[0162] According to some embodiments,, the at least one cytoplasmic costimulatory polypeptide domain, region or motif of CD40, and and / or of 0X40 may have an amino acid sequence having at least 70%, or at least 71%, or at least 72%, or at least 73%, or at least 74%, or at least 75%, or at least 76%, or at least 77%, or at least 78%, or at least 79%, or at least 80%, or at least 81 %, or at least 82%, or at least 83%, or at least 84%, or at least 85%, or at least 86%, or at least 87%, or at least 88%, or at least 89% or at least 90%, or at least 91 %, or at least 92%, or at least 93%, or at least 94%, or at least 95%, or at least 96% or at least 97%, or at least 98%, or at least 99%, or 100% sequence identity with the respective functional polypeptide domain or a functional polypeptide motif of a wildtype human CD40 and / or 0X40, respectively.

[0163] According to an embodiment, the chimeric CD95 receptor polypeptide may be able to enhance cytotoxicity of the T-cell as herein provided.

[0164] According to an embodiment, the chimeric CD95 receptor polypeptide may be able to enhance activation, proliferation, and / or production of activating cytokines of / in the T-cell as herein provided.

[0165] According to an embodiment, the chimeric CD95 receptor may be capable of increasing resistance of T-cells as herein provided to CD95L expressing cancer cells.

[0166] As mentioned above, the chimeric CD95 receptor as herein provided may comprise at least one cytoplasmic polypeptide domain, region or motif of wildtype CD40. For example, the expression “wildtype human CD40 cytoplasmic domain” as referred to herein may relate to a polypeptide comprising amino acid sequence 216-277 of UniProtKB database entry No. P25942- TNR5_HUMAN, as set forth e.g. in SEQ ID No. 141. For example, the wildtype cytoplasmic domain of CD40 is set forth herein by SEQ ID No 129. It is herewith envisaged that the cytoplasmic polypeptide domain, region or motif of wildtype CD40 as included in the chimeric CD95 receptor as herein provided may have one or more conservative amino acid substitutions relative to the amino acid sequence as set forth in SEQ ID No. 129. In particular, all amino acid substitutions that maintain the functional activity of the cytoplasmic polypeptide domain, region or motif of wildtype CD40 are envisaged.

[0167] According to an embodiment, the second non-CD95 co-stimulatory region may comprise at least one TRAF binding motif of CD40. TRAF binding motifs of CD40 have been previously described in the art, as discussed above.

[0168] According to an embodiment, the second non-CD95 co-stimulatory region may comprise at least one TRAF6 binding motif of CD40.

[0169] According to an embodiment, the second non-CD95 co-stimulatory region may comprise or may consist of the polypeptide region of human CD40 as set forth in SEQ ID No. 130.

[0170] It is contemplated herein that the second non-CD95 co-stimulatory region may comprise or may consists of the polypeptide region of human CD40 as set forth in SEQ ID No. 130, or that the second non-CD 95 co-stimulatory region may comprise or consist of a polypeptide region of human CD40 having at least 85% or at least 86%, or at least 87%, or at least 88%, or at least 89% or at least 90%, or at least 91%, or at least 92%, or at least 93%, or at least 94%, or at least 95%, or at least 96% or at least 97%, or at least 98%, or at least 99%, or 100% identity to the amino acids as set forth in SEQ ID No. 130. For example, the second non-CD95 co-stimulatory region may comprise or may consist of a polypeptide region of human CD40 that may have one or more conservative amino acid substitutions relative to the amino acid sequence as set forth in SEQ ID No.130.

[0171] According to an embodiment, the second non-CD95 co-stimulatory region may comprise a polypeptide region of CD40 including amino acid position 237 of SEQ ID No 141 (wildtype CD40) , and further wherein, at said position, the first non-CD95 costimulatory region may be mutated, optionally wherein said mutation may consist of an exchange of an asparagine (N) to an aspartic acid (D). According to an embodiment, the “TRAF6” binding motif of CD40 as included in second non-CD95 co-stimulatory region may comprise a polypeptide region of CD40 including amino acid position 229 of SEQ ID No. 141 (wildtype CD40) , and further wherein, at said position, the TRAF6 binding motif may be mutated, optionally wherein said mutation may consist of an exchange of an proline (P) to an alanine (A).

[0172] It is envisaged herein that a CD40 TRAF binding domain (e.g. a CD40 TRAF 6 binding domain) that is used in context with the chimeric CD8 Co-receptor as herein provided may also be used in context with the second non-CD95 costimulatory region of the chimeric CD 95 receptor as herein provided, and vice versa.

[0173] According to an embodiment, the second non-CD95 co-stimulatory region may comprise or may consists of the polypeptide region having at least 85% or at least 86%, or at least 87%, or at least 88%, or at least 89% or at least 90%, or at least 91 %, or at least 92%, or at least 93%, or at least 94%, or at least 95%, or at least 96% or at least 97%, or at least 98%, or at least 99%, or 100% identity to the amino acids as set forth in SEQ ID No. 143.

[0174] According to the present invention, the chimeric CD95 receptor as herein provided may further comprise at least one cytoplasmic polypeptide domain, region or motif of 0X40. For example, the first non-CD95 cytoplasmic co-stimulatory region of the chimeric CD95 receptor may comprise the complete cytoplasmic domain of wildtype human 0X40. In other embodiments, the cytoplasmic polypeptide region of 0X40 included in the chimeric CD95 receptor may comprise at least one functional, co-stimulatory motif / domain / region of the complete wildtype human 0X40 cytoplasmic domain. For example, the expression “wildtype human 0X40 cytoplasmic domain” as referred to herein may relate to a polypeptide comprising amino acid sequence 236-277 of UniProtKB database entry No. P43489- TNR4_HUMAN, as set forth e.g. in SEQ ID No. 142. It is herewith envisaged that the cytoplasmic polypeptide domain, region or motif of wildtype 0X40 as included in the chimeric CD95 receptor of the T-cell as herein provided may have one or more conservative amino acid substitutions relative to the amino acid sequence 236 - 277 as set forth in SEQ ID No. 142. In particular, all amino acid substitutions that maintain the functional activity of the cytoplasmic polypeptide domain, region or motif of wildtype 0X40 are envisaged.

[0175] Thus, it is envisaged that the first non-CD95 co-stimulatory region may comprise or may consist od a polypeptide having the amino acid sequence 236 - 277 as set forth in SEQ ID No. 142, or that the first non-CD95 co-stimulatory region may comprise or consist of a polypeptide region of human 0X40 having at least 85% or at least 86%, or at least 87%, or at least 88%, or at least 89% or at least 90%, or at least 91 %, or at least 92%, or at least 93%, or at least 94%, or at least 95%, or at least 96% or at least 97%, or at least 98%, or at least 99%, or 100% identity to the amino acids 236 - 277 as set forth in SEQ ID No. 142. For example, the first non-CD95 co-stimulatory region may comprise or may consist of a polypeptide region of human 0X40 that may have one or more conservative amino acid substitutions relative to the amino acid sequence 236 - 277 as set forth in SEQ ID No. 142.

[0176] According to an embodiment, the first non-CD95 co-stimulatory region may comprise at least one TRAF binding site of 0X40. TRAF binding domains of 0X40 have been previously described in the art.

[0177] According to an embodiment, the first non-CD95 co-stimulatory region may comprise at least one potential PI3K binding motif / potential ubiquitination site of 0X40. Potential PI3K binding motif / potential ubiquitination site of 0X40 have been previously described in the art (e.g. Croft, M. et al., Immunol Rev. 2009 May ; 229(1)).

[0178] According to an embodiment, the first non-CD95 co-stimulatory region may comprise a polypeptide region of 0X40 including amino acid positions 266 to 268 of SEQ ID No 142 as set forth herein (human wildtype 0X40), and further wherein at said position, the first non-CD95 co-stimulatory region is mutated, wherein said mutation consists of an exchange of the wildtype 0X40 amino acids EQA to TEP.

[0179] It is contemplated herein that the first non-CD95 co-stimulatory region may comprise or may consists of the polypeptide region of human 0X40 as set forth in SEQ ID No. 144, or that the first non-CD95 co-stimulatory region may comprise or consist of a polypeptide region of human 0X40 having at least 85% or at least 86%, or at least 87%, or at least 88%, or at least 89% or at least 90%, or at least 91%, or at least 92%, or at least 93%, or at least 94%, or at least 95%, or at least 96% or at least 97%, or at least 98%, or at least 99%, or 100% identity to the amino acids as set forth in SEQ ID No. 144. For example, the first non-CD95 co-stimulatory region may comprise or may consist of a polypeptide region of human 0X40 that may have one or more conservative amino acid substitutions relative to the amino acid sequence as set forth in SEQ ID No. 144.

[0180] According to some embodiments, the second costimulatory region (CD40) may be located closer to the C-terminal tail of the polypeptide than the first costimulatory region (0X40).

[0181] It is further envisaged that the cytoplasmic non-CD95 co-stimulatory region may comprise a linker between the first and the second non-CD95 co-stimulatory regions,optionally wherein said linker comprises or consists of a sequence as set forth in SEQ ID No.148.

[0182] According to an embodiment, the chimeric human CD 95 receptor may comprise or consists of an amino acid sequence having at least 70%, or at least 75%, or at least 80%, or at least 85%, or at least 90%, or at least 95%, or at least 97%, or 100% sequence identity with the amino acid sequence as set forth in Seq ID No: 145.

[0183] According to further embodiments of the use of the above-described polypeptides in a method of treating a disease in a subject, wherein the method comprises administering a therapeutically effective amount of the antigen recognizing construct, of the chimeric human CD8 co-receptor, and of the chimeric human CD95 receptor according to the first aspect, the disease may be a cancer.

[0184] For example, the disease may be a tumor, optionally an advanced-stage tumor, that expresses PRAME.

[0185] According to an embodiment, the disease may be selected from the group consisting of melanoma, bladder carcinoma, colon carcinoma, and breast adenocarcinoma, sarcoma, prostate cancer, uterine cancer, uveal cancer, uveal melanoma, squamous head and neck cancer, synovial carcinoma, Ewing’s sarcoma, breast cancer such as triple negative breast cancer, thyroid cancer, testicular cancer, renal cancer, pancreatic cancer, ovarian cancer, esophageal cancer, non-small-cell lung cancer (NSCLC), non-Hodgkin’s lymphoma, multiple myeloma, melanoma, hepatocellular carcinoma, head and neck cancer, gastric cancer, endometrial cancer, colorectal cancer, cholangiocarcinoma, breast cancer, bladder cancer, myeloid leukemia, acute lymphoblastic leukemia, small-cell lung cancer (SCLC), sarcoma and osteosarcoma.

[0186] For example, the breast cancer may be selected from the group consisting of ductal breast cancer, tubular breast cancer, medullary breast cancer, triple negative breast cancer and combinations thereof.

[0187] The gastric cancer may be gastric adenocarcinoma cancer.

[0188] In accordance with an embodiment, the sarcoma cancer is selected from the group consisting of chondrosarcoma cancer, osteosarcoma cancer, synovial sarcoma cancer and combinations thereof.

[0189] In accordance with the first aspect of the invention, there is further provided herein(i) A T-cell receptor capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide, wherein the T-cell receptor comprises: a TCR a-chain comprising or consisting of the amino acid sequence of SEQ ID NO: 107, or having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 107; and a TCR p-chaincomprising or consisting of the amino acid sequence of SEQ ID NO: 108, or having at least 90% sequence identity with the amino acid sequence of SEQ ID NO: 108;(ii) a chimeric human CD8 co-receptor consisting of a polypeptide having an amino acid sequence as set forth in SEQ ID No. 133, or having at least 90% sequence identity with the amino acid sequence as set forth in SEQ ID No. 133; and / or(iii) a chimeric human CD 95 receptor consisting of a polypeptide having an amino acid sequence as set forth in SEQ ID No. 145, or having at least 90% sequence identity with the amino acid sequence as set forth in SEQ ID No. 145;for use in a method of treating a cancer in a subject, wherein the method comprises administering a therapeutically effective amount of the antigen recognizing construct, of the chimeric human CD8 co-receptor, and of the chimeric human CD95 receptor.

[0190] In accordance with the first aspect of the invention, there is also provided herein(i) a T-cell receptor capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide, wherein the T-cell receptor comprises: a TCR a-chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 111 , or having at least 95% sequence identity with the amino acid sequence of SEQ ID NO: 111 and a TCR p-chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 112, or having at least 95% sequence identity with the amino acid sequence of SEQ ID NO: 112;(ii) a chimeric human CD8 co-receptor consisting of a polypeptide having an amino acid sequence as set forth in SEQ ID No. 133, or having at least 90% sequence identity with the amino acid sequence as set forth in SEQ ID No. 133; and / or(iii) a chimeric human CD 95 receptor consisting of a polypeptide having an amino acid sequence as set forth in SEQ ID No. 145, or having at least 90% sequence identity with the amino acid sequence as set forth in SEQ ID No. 145;for use in a method of treating a cancer in a subject, wherein the method comprises administering a therapeutically effective amount of the antigen recognizing construct, of the chimeric human CD8 co-receptor, and of the chimeric human CD95 receptor.

[0191] According to a further aspect of the invention, there is provided a method of treating a disease in a subject (e.g. in a patient), comprising the step of administering a therapeutically effective amount of an antigen (e.g. PRAME) recognizing construct or an (PRAME recognizing) T cell receptor as herein described, of a chimeric human CD8 co-receptor as herein described, and of a chimeric human CD95 receptor as herein described.

[0192] Another aspect of the invention relates to a combination (or composition) comprising an antigen (e.g. PRAME) recognizing construct or an (PRAME recognizing) Tcell receptor as herein described, a chimeric human CD8 co-receptor as herein described, and a chimeric human CD 95 receptor as herein described.According to an embodiment, the combination may be used in a method of treating a disease, optionally a tumor, in a subject (e.g. in a patient).

[0193] According to a further aspect, the invention also relates to a nucleic acid sequence encoding an antigen recognizing construct or a T cell receptor as herein defined, a chimeric human CD8 co-receptor according as herein defined, and the chimeric human CD95 receptor as defined herein.

[0194] It is envisaged that the nucleic acid sequence may further comprise at least one nucleic acid sequence encoding for a 2A peptide sequence, optionally wherein the nucleic acid sequence comprises a 2A peptide sequence between each of the sequences encoding for a TCRa chain, for a TCRp chain, for the chimeric human CD8 Co-receptor, and for the chimeric human CD 95 receptor.

[0195] A used herein, “nucleic acid” refers to single or double-stranded molecules, which may be DNA or RNA, including genomic DNA, mRNA, cRNA, miRNA, tRNA, although without limitation thereto. A nucleic acid comprises a nucleotide sequence which typically includes nucleotides that comprise an A, G, C, T or II base. However, nucleotide sequences may include other bases such as inosine, methylcytosine, hydroxymethycytosine, methylinosine, methyladenosine and / or thioitridine, although without limitation thereto.

[0196] In a further aspect, the inventon is directed to a vector comprising the nucleic acid sequence according to the invention.

[0197] According to an embodiment, the vector may be a viral vector or a non-viral vector.

[0198] For example, the viral vector may be selected from adenoviruses, poxviruses, alphaviruses, arenaviruses, flaviruses, rhabdoviruses, retroviruses, lentiviruses, herpesviruses, paramyxoviruses, picornaviruses, and combinations thereof.

[0199] The invention will be further explained in the following making reference to either, several or all of these aspects. If reference is only made to one of these aspects, it is understood by the person skilled in the art, that this reference nevertheless includes references to all other aspects of the invention, if applicable.

[0200] In a further aspect, the invention is directed to a host cell comprising i) the antigen recognizing construct or T cell receptor according as defined with respect to the first aspect of the invention, the chimeric human CD8 co-receptor as defined with respect to the first aspect of the invention, and the chimeric human CD 95 receptor as defined with respect to the first aspect of the invention, and / orii) the nucleic acid sequence as herein provided, and / oriii) the vector as herein provided.

[0201] According to an embodiment, the host cell may comprise a T cell, a hematopoietic stem cell, an NK cell, a primary T cell, or a natural killer T cell.

[0202] According to an embodiment, the host cell may be a T-cell.

[0203] According to a further aspect, there is also provided an isolated T-cell, wherein the T-cell expresses the antigen recognizing construct or the T cell receptor according to the first aspect, the chimeric human CD8 co-receptor according to the first aspect, and the chimeric human CD95 receptor according to the first aspect.

[0204] According to a further aspect, there is provided an isolated T-cell, wherein the T-cell expresses:(i) a T-cell receptor capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide, wherein the T-cell receptor comprises: a TCR a-chain comprising or consisting of the amino acid sequence of SEQ ID NO: 107, and a TOR p-chain comprising or consisting of the amino acid sequence of SEQ ID NO: 108,(ii) a chimeric human CD8 Co-receptor consisting of a polypeptide having an amino acid sequence as set forth in SEQ ID No. 133 ;and(iii) a chimeric human CD 95 receptor consisting of a polypeptide having an amino acid sequence as set forth in SEQ ID No. 145.

[0205] There is provided an isolated T-cell, wherein the T-cell expresses:(i) a T-cell receptor capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide, wherein the T-cell receptor comprises: a TCR a-chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 111 , or having at least 95% sequence identity with the amino acid sequence of SEQ ID NO: 111 and a TCR p-chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 112, or having at least 95% sequence identity with the amino acid sequence of SEQ ID NO: 112;(ii) a chimeric human CD8 co-receptor consisting of a polypeptide having an amino acid sequence as set forth in SEQ ID No. 133, or having at least 90% sequence identity with the amino acid sequence as set forth in SEQ ID No. 133; and / or(iii) a chimeric human CD 95 receptor consisting of a polypeptide having an amino acid sequence as set forth in SEQ ID No. 145, or having at least 90% sequence identity with the amino acid sequence as set forth in SEQ ID No. 145.

[0206] According to an embodiment, the cell may be an p T-cell, y8 T-cell, and / or a natural killer T-cell.

[0207] For example, the ap T-cell may be a CD4 T-cell, or the T-cell may be a CD8 T-cell, or the y5 T-cell may be a Vy9V82+ T-cell, or the y5 T-cell may comprise a V81 T-cell.

[0208] It is envisaged that the T-cell may be derived from an induced pluripotent stem cell (iPSCs).

[0209] According to an embodiment, the T-cells herein provided may have an enhanced cytotoxicity.

[0210] According to an embodiment, the T-cells herein provided may have an increased resistance to CD95L expressing cancer cells.

[0211] According to a further aspect, the invention also provides:(a) a combination (or composition) comprising an antigen (e.g. PRAME) recognizing construct or a (PRAME recognizing) T cell receptor as herein described, a chimeric human CD8 co-receptor as herein described, and a chimeric human CD95 receptor as herein described, or(b) a nucleic acid, wherein the nucleic acid encodes for an antigen (e.g. PRAME) recognizing construct or a (PRAME recognizing) T cell receptor as herein described, a chimeric human CD8 co-receptor as herein described, and a chimeric human CD95 receptor as herein described, or(c) a vector combrising the nucleic acid as herein provide, or(d) a host cell (e.g. T-cell), wherein the cell comprises:i) the antigen recognizing construct or T cell receptor according as defined with respect to the first aspect of the invention, the chimeric human CD8 co-receptor as defined with respect to the first aspect of the invention, and the chimeric human CD95 receptor as defined with respect to the first aspect of the invention, and / orii) the nucleic acid sequence as herein provided, and / oriii) the vector as herein provided,for use in medicine, e.g. for use in detection, diagnosis, prognosis, prevention and / or treatment of a disease.

[0212] According to another aspect, there is also provided herein a use of(a) an antigen (e.g. PRAME) recognizing construct or a (PRAME recognizing) T cell receptor as herein described, a chimeric human CD8 co-receptor as herein described, and a chimeric human CD95 receptor as herein described.(b) a combination (or composition) comprising an antigen (e.g. PRAME) recognizing construct ora (PRAME recognizing) T cell receptor as herein described, a chimeric human CD8 co-receptor as herein described, and a chimeric human CD95 receptor as herein described, or(c) a nucleic acid, wherein the nucleic acid encodes for an antigen (e.g. PRAME) recognizing construct or an (PRAME recognizing) T cell receptor as herein described, a chimeric human CD8 co-receptor as herein described, and a chimeric human CD95 receptor as herein described, or(d) a vector combrising the nucleic acid as herein provide, or(e) a host cell (e.g. T-cell), wherein the cell comprises:i) the antigen recognizing construct or T cell receptor according as defined with respect to the first aspect of the invention, the chimeric human CD8 co-receptor as defined with respect to the first aspect of the invention, and the chimeric human CD95 receptor as defined with respect to the first aspect of the invention, and / orii) the nucleic acid sequence as herein provided, and / oriii) the vector as herein providedfor the manufacture of a medicament for treating a disease.

[0213] According to an embodiment, the disease may be a cancer.

[0214] For example, the disease may be a tumor that expresses PRAME, optionally wherein the disease may be an advanced-stage tumor that expresses PRAME.

[0215] The terms “treat” and “prevent” as well as words stemming therefrom, like “treatment” or “prevention”, as used herein, do not necessarily imply 100% or complete treatment or prevention. Rather, there are varying degrees of treatment or prevention of which one of ordinary skill in the art recognizes as having a potential benefit or therapeutic effect. In this respect, the inventive methods can provide any amount of any level of treatment or prevention of cancer in a mammal. Furthermore, the treatment or prevention provided by the inventive method can include treatment or prevention of one or more conditions or symptoms of the cancer being treated or prevented. For example, treatment or prevention can include promoting the regression of a tumor. Also, for purposes herein, “prevention” can encompass delaying the onset of the cancer, or a symptom or condition thereof.

[0216] According to an embodiment, the disease may be selected from the group consisting of melanoma, bladder carcinoma, colon carcinoma, and breast adenocarcinoma, sarcoma, prostate cancer, uterine cancer, uveal cancer, uveal melanoma, squamous head and neck cancer, synovial carcinoma, Ewing’s sarcoma, breast cancer such as triple negative breast cancer, thyroid cancer, testicular cancer, renal cancer, pancreatic cancer, ovarian cancer, esophageal cancer, non-small-cell lung cancer (NSCLC), non-Hodgkin’s lymphoma, multiple myeloma, melanoma, hepatocellular carcinoma, head and neck cancer, gastric cancer, endometrial cancer, colorectal cancer, cholangiocarcinoma, breast cancer, bladder cancer, myeloid leukemia, acute lymphoblastic leukemia, small-cell lung cancer (SCLC), sarcoma and osteosarcoma.

[0217] According to an embodiment, the breast cancer may be selected from the group consisting of ductal breast cancer, tubular breast cancer, medullary breast cancer, triple negative breast cancer and combinations thereof.

[0218] According to an embodiment, the sarcoma cancer may be selected from the group consisting of chondrosarcoma cancer, osteosarcoma cancer, synovial sarcoma cancer and combinations thereof.

[0219] In an embodiment, the gastric cancer may be gastric adenocarcinoma cancer.

[0220] The present invention also provides a pharmaceutical composition comprising:(a) an antigen (e.g. PRAME) recognizing construct or an (PRAME recognizing) T cell receptor as herein described, a chimeric human CD8 co-receptor as herein described, and a chimeric human CD95 receptor as herein described;(b) a combination (or composition) comprising an antigen (e.g. PRAME) recognizing construct ora (PRAME recognizing) T cell receptor as herein described, a chimeric human CD8 co-receptor as herein described, and a chimeric human CD95 receptor as herein described, or(c) a nucleic acid, wherein the nucleic acid encodes for an antigen (e.g. PRAME) recognizing construct or an (PRAME recognizing) T cell receptor as herein described, a chimeric human CD8 Co-receptor as herein described, and a chimeric human CD95 receptor as herein described, or(d) a vector combrising the nucleic acid as herein provide, or(e) a host cell (e.g. T-cell), wherein the cell comprises:i) the antigen recognizing construct or T cell receptor according as defined with respect to the first aspect of the invention, the chimeric human CD8 co-receptor as defined with respect to the first aspect of the invention, and the chimeric human CD95 receptor as defined with respect to the first aspect of the invention, and / orii) the nucleic acid sequence as herein provided, and / oriii) the vector as herein provided.

[0221] According to an embodiment, the pharmaceutical composition may further comprise an adjuvant, excipient, buffer, diluent, carrier, stabilizer or combination thereof.

[0222] According to another aspect, the invention further provides a method of preparing a T-cell for immunotherapy, comprisingisolating T-cells from a human subject,introducing the vector according to any one of claims 43 to 45 into the T-cell, and expanding the T-cells

[0223] It is contemplated herein that the method may comprise transforming, transfecting or transducing the isolated T-cell with the vector.

[0224] Thus, the pharmaceutical composition as herein provided may also comprise T-cells (e.g. also comprising CD4 cells) expressing the antigen recognizing construct or the T cell receptor as herein defined e.g. in the first aspect of the invention, the chimeric human CD8 co-receptor as herein defined e.g. in the first aspect of the invention, and the chimeric human CD95 receptor as herein defined e.g. in the first aspect of the invention.

[0225] The invention further provides a method of treating a patient having a disease, comprising administering the pharmaceutical composition as herein described to the patient.

[0226] introducing in vivo the vector according to any one of claims 43 to 45 into a T-cell of the patient.

[0227] According to an embodiment, the vector may be DNA or mRNA.

[0228] According to an embodiment, the vector may be a non-replicating viral vector.

[0229] It is envisaged herein that in methods wherein the vector is mRNA, said mRNA may be introduced into the T-cell of the patient using nanoparticles.

[0230] In the methods and uses provided herein, the disease may be a cancer or an autoimmune disease.

[0231] According to an embodiment wherein the cancer may be selected from the group consisting of non-small cell lung cancer, small cell lung cancer, pancreatic cancer, ovarian cancer, melanoma, uveal melanoma, synovial sarcoma, breast cancer, triple negative breast cancer, endometrial cancer, liver cancer, kidney cancer, esophageal cancer, brain cancer, gastric cancer, Merkel cell carcinoma, leukemia, urinary bladder cancer, uterine cancer, colorectal cancer, gallbladder cancer, bile duct cancer, and prostate cancer.

[0232] For example, the cancer cells may express FASL.

[0233] It is further provided a method for increasing cytotoxicity of a T-cell in adoptive cell therapy, comprisingintroducing a vector as herein defined into a T-cell.

[0234] Dealing now with the kit of the invention, such a kit is a kit for use in medicine comprisingi) the antigen recognizing construct or the T cell receptor according as herein defined, the chimeric human CD8 co-receptor as herein defined, and the chimeric human CD95 receptor as herein defined, orii) the combination as herein provided, oriii) the nucleic acid sequence as herein provided, oriv) the vector as herein provided, orv) the cell as herein provided.

[0235] The invention will be further illustrated by the following non-limiting Experimental Examples.

[0236] Sequences, as used herein, are depicted in below Table 1.

[0237] Table 1. Sequences as used herein.&Experimental Examples

[0238] Example 1:

[0239] PBMCs from a healthy donor buffy coat were isolated by density gradient centrifugation with Lymphoprep. Purified polyclonal CD4+ and CD8+ T cells were obtained by positive selection with anti-CD4+ and anti-CD8+ microbeads. CD3+ T cells were activated using TransAct in presence of IL-7 / IL-15. Two days post activation, CD4 / 8+ T cells were electroporated with gRNA / Cas9 protein complex together with a template plasmid bearing transgene of interest. Electroporated CD4 / 8+ T cells were further expanded, and at Day 10 harvested and cryopreserved. T cell characterization was based on transgene expression levels using FACS and killing assay.

[0240] Cell killing assay in NCI-H1703 cells (60% No Inhibitory Ligands+20%PDL1 +20%FASL):

[0241] The in-vitro T-cell killing assay was performed according to the method described e.g. by Kalbasi, A., Siurala, M., Su, L.L. et al. “Potentiating adoptive cell therapy using synthetic IL-9 receptors”. Nature 607, 360-365 (2022). In particular, the human T-cell repetitive killing assay was conducted using IncuCyte Live Cell Analysis. Using 20% PDL1+, 20% FASL+ and 60% PDL1-FASL- NCI-H1703 cells, 1x104tumor cells were plated per well in 96-well plates. Transfected human T-cells (with no plasmid (Mock T cells), or transfected with a specific combination of the antigen recognizing construct capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide, the chimeric human CD8 co-receptor, and the chimeric human CD95 receptor according to the present invention (pl_2324); or transduced with published known PRAME-specific T-Cell Receptor in respectively published combinations with either a wildtype CD8 coreceptor (pl_2387 and pl_2344) or with a PD1-41BB Switch receptor (pl_2388)) were added in triplicates at 2 to 1 E:T ratio (for the experiments shown by Fig 1).

[0242] T-cell killing assay analysis

[0243] The Relative cell growth has been observed over time for each transduced T-cell fraction. The results are shown in Fig. 1. “Mock” relates to mock-transfected T-cell fraction, “pl_2324” relates to T-cells transfected with the specific combination of the antigen recognizing construct capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide, the chimeric human CD8 co-receptor, and the chimeric human CD95 receptor according to the present invention. In particular, “pl_2324” relates to a vector used for transfection of T-cells which comprises a nucleic acid encoding for the PRAME specific T-cell receptor comprising a TCR a-chain consisting of the amino acid sequence of SEQ ID NO: 107, and a TCR b-chain consisting of the amino acid sequence of SEQ ID NO: 108 (S109 / T01), further encoding for the chimeric human CD8 co-receptorconsisting of a polypeptide having an amino acid sequence as set forth in SEQ ID No. 133, and further encoding for the chimeric human CD95 receptor consisting of a polypeptide having an amino acid sequence as set forth in SEQ ID No. 145. “pl_2388” relates to a vector used for transfection of T-cells which comprises a nucleic acid encoding for a publicly known PRAM E-specific T-Cell Receptor from Medigene Immunotherapies GmbH and BioNtech SE as published in the international Patent application Publication No. WO 2021 / 099360 A1 (TCR 027-004), and further encoding for a publicy known PD1-41BB Switch receptor from Medigene Immunotherapies GmbH (WO2022 / 234116A1). “pl_2387” relates to a vector used for transfection of T-cells which comprises a nucleic acid encoding for a publicly known PRAM E-specific T-Cell Receptor from TSCAN THERAPEUTICS as published in the international Patent application Publication No. WO2023215183 and WO2024077134A1 (TCR366), and further encoding for a wildtype CD8 co-receptor consisting of a wildtype CD8a polypeptide having an amino acid sequence as set forth in SEQ ID No.113, and a wildtype CD8p polypeptide having an amino acid sequence as set forth in SEQ ID No. 114 respectively. “pl_2344” relates to a vector used for transfection of T-cells which comprises a nucleic acid encoding for a publicly known PRAM E-specific T-Cell Receptor from Immatics Biotechnologies as published in the international Patent application Publication No. WO2018172533A2 (R11P3D3_KE), and further encoding for a wildtype CD8 co-receptor consisting of a wildtype CD8a polypeptide having an amino acid sequence as set forth in SEQ ID No. 113, and a wildtype CD8p polypeptide having an amino acid sequence as set forth in SEQ ID No. 114 respectively.

[0244] The results are given herein in Fig. 1. As visible, human T-cells Cotransfected with the specific combination of the antigen recognizing construct capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide, the chimeric human CD8 co-receptor, and the chimeric human CD95 receptor according to the present invention show an increased cytotoxicity in the T-cell serial killing assay in comparison with the reference TCRs co-transfected with either CD8 wildtype receptor, or with a PD1-41 BB Switch receptor. (pl_2344 and pl_2387 or pl_2388, respectively).

[0245] Example 2:

[0246] Human T cells (mix of both CD4 and CD8 T cells) from a human donor were either non-transfected, or transfected with “Mock”, “pl_2387”, “pl_2344”, “pl_2388”, or pl_2324, respectively. “Mock” relates to mock-transfected T-cell fraction, “pl_2324” relates to T-cells transfected with the specific combination of the antigen recognizing construct capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide, the chimeric human CD8 co-receptor , and the chimeric human CD95 receptor according to the present invention. In particular, “pl_2324” relates to a vector used for transfection ofT-cells which comprises a nucleic acid encoding for the PRAME specific T-cell receptor comprising a TCR a-chain consisting of the amino acid sequence of SEQ ID NO: 107, and a TCR b-chain consisting of the amino acid sequence of SEQ ID NO: 108 (S109 / T01), further encoding for the chimeric human CD8 co-receptor consisting of a polypeptide having an amino acid sequence as set forth in SEQ ID No. 133, and further encoding for the chimeric human CD95 receptor consisting of a polypeptide having an amino acid sequence as set forth in SEQ ID No. 145. “pl_2388” relates to a vector used for transfection of T-cells which comprises a nucleic acid encoding for a publicly known PRAME-specific T-Cell Receptor from Medigene Immunotherapies GmbH and BioNtech SE as published in the international Patent application Publication No. WO 2021 / 099360 A1 (TCR 027-004), and further encoding fora publicy known PD1-41BB Switch receptor from Medigene Immunotherapies GmbH (WO2022 / 234116A1). “pl_2387” relates to a vector used for transfection of T-cells which comprises a nucleic acid encoding for a publicly known PRAME-specific T-Cell Receptor from TSCAN THERAPEUTICS as published in the international Patent application Publication No. WO2023215183 and WQ2024077134A1 (TCR366), and further encoding for a wildtype CD8 co-receptor consisting of a wildtype CD8a polypeptide having an amino acid sequence as set forth in SEQ ID No. 113, and a wildtype CD8p polypeptide having an amino acid sequence as set forth in SEQ ID No. 114 respectively. “pl_2344” relates to a vector used for transfection of T-cells which comprises a nucleic acid encoding for a publicly known PRAME-specific T-Cell Receptor from Immatics Biotechnologies as published in the international Patent application Publication No. WO2018172533A2 (R11P3D3_KE), and further encoding for a wildtype CD8 co-receptor consisting of a wildtype CD8a polypeptide having an amino acid sequence as set forth in SEQ ID No. 113, and a wildtype CD8p polypeptide having an amino acid sequence as set forth in SEQ ID No. 114 respectively. The transfected T cells were co-cultured with PRAME and HLA-A*02:01 expressing cell lines NCI-H1703 (60% of the cells express No Inhibitory Ligands, 20% express PDL1, and 20% express FASL). Cytokine / effector molecule secretion in the culture supernatants upon T cell activation was measured by LegendPlex assay. Specifically, the secretion of interferon-y (IFNy), of granzyme A (GZMA), of granzyme B (GZMB), and of perforin, and the level of soluble FAS ligand (FASL) has been measured. The results are shown in Fig. 2, Fig.3, Fig.4, Fig.5, and Fig.6, respectively.

[0247] As demonstrated by the results in Fig. 2, Fig.3, Fig.4, Fig.5, and Fig. 6, as visible, human T-cells co-transfected with the specific combination of the antigen recognizing construct capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide, the chimeric human CD8 co-receptor , and the chimerichuman CD95 receptor according to the present invention, show increased secretion of interferon-y (IFNy), of granzyme A (GZMA), of granzyme B (GZMB), of perforin upon T cell activation in comparison with the reference TCRs co-transduced with either wildtype CD8 ap co-receptor, or with a PD1-41 BB switch receptor. Furthermore, the level of the inhibitory soluble FasL is reduced. Thus, use of the specific combination of the antigen recognizing construct capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide, the chimeric human CD8 co-receptor, and the chimeric human CD95 receptor for T-cell transduction results in T-cells having superior advantageous characteristics for the use in immunotherapy in comparison with published combinations of PRAME TCRs transfected together with additional polypeptides.Materials and Methods

[0248] Animal immunizationsTo generate CD8+ T cell responses to PRAME epitopes, humanized TCR / HLA class I transgenic mice according to the ABabDII mouse strain described in Li etal. (2010, Nature Medicine 16, 1029-1034), were used for immunizations with either full length human PRAME in adenoviral vector i.p., or with a PRAME peptide “SLLQHLIGL” plus adjuvant s.c. For peptide immunizations, the mice were subcutaneously primed and boosted monthly with the PRAME peptide “SLLQHLIGL” emulsified in adjuvant. For adenoviral immunizations, the mice received intraperitoneal injections of an adenoviral vector that expresses the full length human PRAME protein under control of the CMV promoter.

[0249] TCR identification and characterizationExpanded TCR clonotypes were screened in a reporter-based screening assay. Most functional TCRs were selected and cloned into a retroviral vector and transduced into human peripheral blood lymphocytes for detailed functional characterization.

[0250] It will be readily apparent to a person skilled in the art that varying substitutions and modifications may be made to the invention disclosed herein without departing from the scope and spirit of the invention.

[0251] All patents and publications mentioned in the specification are indicative of the levels of those of ordinary skill in the art to which the invention pertains. All patents and publications are herein incorporated by reference to the same extent as if each individual publication was specifically and individually indicated to be incorporated by reference.

[0252] As used herein, the term "about" means ± 20% of the indicated range, value, or structure, unless otherwise indicated. It should be understood that the terms "a" and"an", as used herein, refer to "one or more" of the enumerated components. The use of the alternative (e.g., "or") should be understood to mean either one, both, or any combination thereof of the alternatives. As used herein, the terms "include", "have", and "comprise" are used synonymously, which terms and variants thereof are intended to be construed as non-limiting.

[0253] The inventions illustratively described herein may suitably be practiced in the absence of any element or elements, limitation or limitations, not specifically disclosed herein. Thus, for example, the terms "comprising", "including", "containing", etc. shall be read expansively and without limitation. Additionally, the terms and expressions employed herein have been used as terms of description and not of limitation, and there is no intention in the use of such terms and expressions of excluding any equivalents of the features shown and described or portions thereof, but it is recognized that various modifications are possible within the scope of the invention claimed. Thus, it should be understood that although the present invention has been specifically disclosed by preferred embodiments and optional features, modification and variation of the inventions embodied therein and herein disclosed may be resorted to by those skilled in the art, and that such modifications and variations are considered to be within the scope of this invention. The invention has been described broadly and generically herein. Each of the narrower species and subgeneric groupings falling within the generic disclosure also form part of the invention. This includes the generic description of the invention with a proviso or negative limitation removing any subject matter from the genus, regardless of whether or not the excised material is specifically recited herein. In addition, where features or aspects of the invention are described in terms of Markush groups, those skilled in the art will recognize that the invention is also thereby described in terms of any individual member or subgroup of members of the Markush group. Further embodiments of the invention will become apparent from the following claims.

Claims

Claims:What is claimed is:

1. (i) An antigen recognizing construct capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide,(ii) a chimeric human CD8 co-receptor comprising a polypeptide having an extracellular polypeptide region, a transmembrane polypeptide region, and a cytoplasmic polypeptide region, wherein said polypeptide comprises at least one CD8a polypeptide region, wherein said least one CD8a polypeptide region comprises an extracellular CD8a IG-like domain region, further wherein said polypeptide comprises at least one CD8p polypeptide region, wherein said least one CD8p polypeptide region comprises an extracellular CD8p IG-like domain region, and / or(iii) a chimeric human CD95 receptor; comprising a polypeptide, wherein said polypeptide comprises at least one CD95 polypeptide region comprising a CD95 extracellular ligand binding domain; further wherein said polypeptide comprises at least one non-CD95 costimulatory cytoplasmic polypeptide domain, region or motif;for use in a method of treating a disease in a subject, wherein the method comprises administering a therapeutically effective amount of the antigen recognizing construct, of the chimeric human CD8 Co-receptor, and of the chimeric human CD95 receptor.

2. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of claim 1 ,wherein the antigen recognizing construct comprises a complementary determining region 3 (CDR3) of the a-chain having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 3, 9, 15, 21, 27, 33, 39, 45, 51, 57, 63, 69, and 75, and / orwherein the antigen recognizing construct comprises a complementary determining region 3 (CDR3) of the p-chain having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 6, 12, 18, 24, 30, 36, 42, 48, 54, 60, 66, 72 and 78.

3. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of claim 1 or 2, wherein the antigen recognizingconstruct is an antibody, or fragment thereof, or a T cell receptor (TCR), or fragment thereof.

4. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of the preceding claims, wherein the antigen recognizing construct is a T cell receptor (TCR).

5. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of the preceding claims, wherein the antigen recognizing construct binds the peptide being presented by a MHC I molecule, optionally wherein the MHC I molecule is an HLA-A molecule.

6. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of the preceding claims, wherein the antigen recognizing construct binds the amino acid sequence SLLQHLIGL (SEQ ID NO: 80) within the PRAME peptide.

7. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of claims 4 to 6, wherein the T cell receptor comprises a complementary determining region 1 (CDR1) of the a-chain having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 7, 13, 19, 25, 31, 37, 43, 49, 55, 61, 67, and 73, and / or wherein the T cell receptor comprises a complementary determining region 1 (CDR1) of the p-chain having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 10, 16, 22, 28, 34, 40, 46, 52, 58, 64, 70 and 76.

8. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of claims 4 to 7, wherein the T cell receptor comprises a complementary determining region 2 (CDR2) of the a-chain having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 2, 8, 14, 20, 26, 32, 36, 44, 50, 56, 62, 68, and 74, and / or wherein the T cell receptor comprises a complementary determining region 2 (CDR2) of the p-chain having at least 90% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 5, 11, 17, 23, 29, 35, 41, 47, 53, 59, 65, 71 and 77.

9. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of claims 4 to 8, wherein the T cell receptor comprises a complementary determining region 3 (CDR3) of the a-chain having at least 95% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 3, 9, 15, 21, 27, 33, 39, 45, 51, 57, 63, 69, and 75, and / or wherein the T cell receptor comprises a complementary determining region 3 (CDR3) of the p-chain having at least 95% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 6, 12, 18, 24, 30, 36, 42, 48, 54, 60, 66, 72 and 78.

10. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of claims 4 to 9, wherein the T cell receptor comprises a complementary determining region 1 (CDR1) of the a-chain having at least 95% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 7, 13, 19, 25, 31, 37, 43, 49, 55, 61, 67, and 73, and / or wherein the T cell receptor comprises a complementary determining region 1 (CDR1) of the p-chain having at least 95% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 10, 16, 22, 28, 34, 40, 46, 52, 58, 64, 70 and 76.

11. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of claims 5 to 10, wherein the T cell receptor comprises a complementary determining region 2 (CDR2) of the a-chain having at least 95% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 2, 8, 14, 20, 26, 32, 36, 44, 50, 56, 62, 68, and 74, and / or wherein the T cell receptor comprises a complementary determining region 2 (CDR2) of the p-chain having at least 95% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 5, 11, 17, 23, 29, 35, 41, 47, 53, 59, 65, 71 and 77.

12. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of claims 5 to 11 , wherein the T cell receptor comprises:(a) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 1,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 2, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 3; or(b) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 7,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 8, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 9; or(c) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 13,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 14, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 15; or(d) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 19,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 20, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 21 ; or(e) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 25,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 26, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 27, or(f) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 31,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 32, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 33, or(g) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 37,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 38, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 39, or(h) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 43,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 44, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 45, or(i) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 49,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 50, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 51 , or(j) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 55,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 56, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 57, or(k) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 61,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 62, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 63, or(l) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 67,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 68, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 69, or(m) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 73,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 74, andan a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 75.

13. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD 95 receptor for the use of any one of claims 3 to 12, wherein the T cell receptor comprises:(a) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 1,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 2, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 3; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 4, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 5, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 6; or (b) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 7,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 8, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 9; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 10, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 11 , anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 12; or(c) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 13,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 14, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 15; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 16, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 17, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 18; or(d) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 19,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 20, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 21 ; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 22, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 23, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 24; or(e) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 25,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 26, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 27; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 28, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 39, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 30, or(f) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 31,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 32, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 33; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 34, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 35, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 36, or(g) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 37,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 38, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 39; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 40, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 41 , anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 42, or(h) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 43,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 44, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 45; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 46, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 47, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 48, or(i) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 49,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 50, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 51 ; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 52, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 53, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 54, or(j) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 55,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 56, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 57; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 58, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 59, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 60, or(k) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 61,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 62, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 63; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 64, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 65, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 66, or(l) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 67,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 68, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 69; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 70, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 71 , anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 72, or(m) an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 73,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 74, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 75; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 76, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 77, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 78; and / orwherein the T cell receptor comprises: an a-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 31 ,an a-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 32, an a-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 33; a p-chain CDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 34, a p-chain CDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 35, anda p-chain CDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 36.

14. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of claims 5 to 13, wherein the T cell receptor comprises: an a-chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 111, and a p-chain variable region comprising or consisting of the amino acid sequence of SEQ ID NO: 112.

15. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of claims 5 to 13, wherein the T cell receptor comprises: a TCR a-chain comprising or consisting of the amino acid sequence of SEQ ID NO: 107, and a TCR p-chain comprising or consisting of the amino acid sequence of SEQ ID NO: 108.

16. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of claims 1 to 15, wherein the cytoplasmic polypeptide region of the chimeric human CD8 co-receptor further comprisesat least two TNF receptor associated factor 6 (TRAF 6) binding motifs, and / or wherein said cytoplasmic polypeptide region comprises at least two CD40 TNF receptor associated factor 1 / 2 / 3 (TRAF 1 / 2 / 3) binding motifs.

17. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of claims 1 to 16, wherein the cytoplasmic polypeptide region of the chimeric human CD8 co-receptor comprises at least two CD40 TNF receptor associated factor 1 / 2 / 3 (TRAF 1 / 2 / 3) binding motifs.

18. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of claim 17, wherein the cytoplasmic polypeptide region of the chimeric human CD8 co-receptor further comprises at least one CD40 TRAF6 binding motif.

19. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of claims 16 to 18, wherein the at least one TRAF 6 binding motif comprises or consists of an amino acid sequence having at least 70%, or at least 75%, or at least 80%, or at least 85%, or at least 90%, or at least 95%, or at least 97%, or 100% sequence identity with the amino acid sequence as set forth in Seq ID No: 130, and / or wherein the the at least one TRAF 1 / 2 / 3 binding motif comprises or consists of an amino acid sequence having at least 70%, or at least 75%, or at least 80%, or at least 85%, or at least 90%, or at least 95%, or at least 97%, or 100% sequence identity with the amino acid sequence as set forth in Seq ID No: 131.

20. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of claims 1 to 19, wherein the cytoplasmic polypeptide region of the chimeric human CD8 co-receptor further comprises a complete cytoplasmic domain of CD4.

21. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of claims 1 to 20, wherein the cytoplasmic polypeptide region of the chimeric human CD8 co-receptor comprises or consists of an amino acid sequence having at least 70%, or at least 75%, or at least 80%, or at least 85%, or at least 90%, or at least 95%, or at least 97%, or 100% sequence identity with the amino acid sequence as set forth in Seq ID No: 132.

22. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of claims 1 to 21, wherein the transmembrane region of the chimeric human CD8 co-receptor comprises a CD8a transmembrane domain.

23. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of claims 1 to 22, wherein the chimeric human CD8 co-receptor comprises or consists of an amino acid sequence having at least 70%, or at least 75%, or at least 80%, or at least 85%, or at least 90%, or at least 95%, or at least 97%, or 100% sequence identity with the amino acid sequence as set forth in Seq ID No: 133.

24. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of claims 1 to 22, wherein the polypeptide of the chimeric human CD95 receptor comprises at least two non-CD95 cytoplasmic co-stimulatory polypeptide domains, regions or motifs, wherein a first non-CD95 co-stimulatory region comprises a cytoplasmic costimulatory region of 0X40, and wherein a second non-CD95 co-stimulatory region comprises a cytoplasmic costimulatory region of CD40.

25. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of claim 24, wherein the second non-CD95 co-stimulatory region comprises at least a TRAF 6 binding motif of CD40, and / or wherein the first non-CD95 co-stimulatory region comprises the complete cytoplasmic region of 0X40.

26. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of claim 25, wherein the at least one TRAF 6 binding motif of CD40 comprises or consists of an amino acid sequence having at least 70%, or at least 75%, or at least 80%, or at least 85%, or at least 90%, or at least 95%, or at least 97%, or 100% sequence identity with the amino acid sequence as set forth in Seq ID No: 130.

27. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of claims 1 to 25, wherein the polypeptide of the chimeric human CD95 receptor comprises a cytoplasmic polypeptide region comprising or consisting of an amino acid sequence having at least 70%, or at least75%, or at least 80%, or at least 85%, or at least 90%, or at least 95%, or at least 97%, or 100% sequence identity with the amino acid sequence as set forth in Seq ID No: 146.

28. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of claims 1 to 25, wherein the at least one CD95 polypeptide region comprises a CD95 transmembrane region.

29. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of claims 1 to 28, wherein said CD95 polypeptide region comprises a complete CD95 extracellular domain.

30. The antigen recognizing construct, the chimeric human CD8 Co-receptor, and / or the chimeric human CD95 receptor for the use of any one of claims 1 to 29, wherein the chimeric human CD95 receptor comprises or consists of an amino acid sequence having at least 70%, or at least 75%, or at least 80%, or at least 85%, or at least 90%, or at least 95%, or at least 97%, or 100% sequence identity with the amino acid sequence as set forth in Seq ID No: 145.

31. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of claims 1 to 30, wherein the disease is a cancer.

32. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of claims 1 to 31, wherein the disease is a tumor, optionally an advanced-stage tumor, that expresses PRAME.

33. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of any one of claims 1 to 32, wherein the disease is selected from the group consisting of melanoma, bladder carcinoma, colon carcinoma, and breast adenocarcinoma, sarcoma, prostate cancer, uterine cancer, uveal cancer, uveal melanoma, squamous head and neck cancer, synovial carcinoma, Ewing’s sarcoma, breast cancer such as triple negative breast cancer, thyroid cancer, testicular cancer, renal cancer, pancreatic cancer, ovarian cancer, esophageal cancer, non-small-cell lung cancer (NSCLC), non-Hodgkin’s lymphoma, multiple myeloma, melanoma, hepatocellular carcinoma, head and neck cancer, gastric cancer, endometrial cancer, colorectal cancer, cholangiocarcinoma, breast cancer, bladder cancer, myeloid leukemia, acute lymphoblastic leukemia, small-cell lung cancer (SCLC), sarcoma and osteosarcoma.

34. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of claim 33, wherein the breast cancer is selected from the group consisting of ductal breast cancer, tubular breast cancer, medullary breast cancer and combinations thereof.

35. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of claim 33, wherein the gastric cancer is gastric adenocarcinoma cancer.

36. The antigen recognizing construct, the chimeric human CD8 co-receptor, and / or the chimeric human CD95 receptor for the use of claim 33, wherein the sarcoma cancer is selected from the group consisting of chondrosarcoma cancer, osteosarcoma cancer and combinations thereof.

37. (i) An antigen recognizing construct comprising an alpha chain comprising a CDR1 having the amino acid sequence of any one of SEQ ID NOs: 1, 7, 13, 19, 25, 31, 37, 43, 49, 55, 61 , 67, or 73, a CDR2 having the amino acid sequence of any one of SEQ ID NOs: 2, 8, 14, 20, 26, 32, 36, 44, 50, 56, 62, 68, or 74, and a CDR3 having the amino acid sequence of any one of SEQ ID NOs: 3, 9, 15, 21, 27, 33, 39, 45, 51, 57, 63, 69, or 75; and a beta chain comprising a CDR1 having the amino acid sequence of any one of SEQ ID NOs: 4, 10, 16, 22, 28, 34, 40, 46, 52, 58, 64, 70 or 76, a CDR2 having the amino acid sequence of any one of SEQ ID NOs: 5, 11, 17, 23, 29, 35, 41 , 47, 53, 59, 65, 71 or 77, and a CDR3 having the amino acid sequence of any one of SEQ ID NOs: 6, 12, 18, 24, 30, 36, 42, 48, 54, 60, 66, 72 or 78;(ii) a chimeric human CD8 Co-receptor comprising a polypeptide having an extracellular polypeptide region, a transmembrane polypeptide region, and a cytoplasmic polypeptide region, wherein said polypeptide comprises at least one CD8a polypeptide region having the amino acid sequence of SEQ ID NO: 116, further wherein said polypeptide comprises at least one CD8p polypeptide region having the amino acid sequence of SEQ ID NO: 117, and / or(iii) a chimeric human CD95 receptor; comprising a polypeptide, wherein said polypeptide comprises at least one CD95 polypeptide region having the amino acid sequence of SEQ ID NO: 150; further wherein said polypeptide comprises at least one non-CD95 costimulatory cytoplasmic polypeptide domain, region or motif;for use in a method of treating a disease in a subject, wherein the method comprises administering a therapeutically effective amount of the antigen recognizing construct, of the chimeric human CD8 Co-receptor, and of the chimeric human CD95 receptor.

38. (i) A T-cell receptor capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide, wherein the T-cell receptor comprises: a TCR a-chain comprising or consisting of the amino acid sequence of SEQ ID NO: 107, and a TOR p-chain comprising or consisting of the amino acid sequence of SEQ ID NO: 108,(ii) a chimeric human CD8 co-receptor comprising or consisting of a polypeptide having an amino acid sequence as set forth in SEQ ID No. 133 and / or(iii) a chimeric human CD 95 receptor comprising or consisting of a polypeptide having an amino acid sequence as set forth in SEQ ID No. 145,for use in a method of treating a cancer in a subject, wherein the method comprises administering a therapeutically effective amount of the antigen recognizing construct, of the chimeric human CD8 co-receptor, and of the chimeric human CD95 receptor.

39. A method of treating a disease in a subject, comprising the step of administering a therapeutically effective amount of the antigen recognizing construct or the T cell receptor according to any one of claims 1 to 38, of the chimeric human CD8 co-receptor according to any one of claims 1 to 38, and of the chimeric human CD95 receptor according to any one of claims 1 to 38.

40. A combination comprising the antigen recognizing construct or the T cell receptor according to any one of claims 1 to 38, of the chimeric human CD8 co-receptor according to any one of claims 1 to 38, and of the chimeric human CD95 receptor according to any one of claims 1 to 38.

41. The combination of claim 40 for use in a method of treating a disease, optionally a tumor, in a subject.

42. A nucleic acid sequence encoding an antigen recognizing construct or a T cell receptor as defined in any one of claims 1 to 38, the chimeric human CD8 co-receptor according as defined in any one of claims 1 to 38, and the chimeric human CD95 receptor as defined in any one of claims claims 1 to 38.

43. The nucleic acid sequence according to claim 42, further comprising at least one nucleic acid sequence encoding for a 2A peptide sequence, optionally wherein the nucleic acid sequence comprises a 2A peptide sequence between each of the sequences encoding for a TCRa chain, for a TCRp chain, for the chimeric human CD8 co-receptor, and for the chimeric human CD95 receptor.

44. A vector comprising a nucleic acid sequence as defined in claim 42 or 43.

45. The vector of claim 44, wherein the vector is a viral vector or a non-viral vector.

46. The vector of claim 45, wherein the viral vector is selected from adenoviruses, poxviruses, alphaviruses, arenaviruses, flaviruses, rhabdoviruses, retroviruses, lentiviruses, herpesviruses, paramyxoviruses, picornaviruses, and combinations thereof.

47. A host cell comprisingi) the antigen recognizing construct or T cell receptor according as defined in any one of claims 1 to 38, the chimeric human CD8 co-receptor according as defined in any one of claims 1 to 38, and the chimeric human CD 95 receptor as defined in any one of claims claims 1 to 38, and / orii) comprising the nucleic acid sequence according to claim 42 or 43, and / oriii) comprising the vector according to any one of claims 44 to 46.

48. The host cell according to claim 47, wherein the host cell comprises a T cell, a hematopoietic stem cell, a NK cell, a primary T cell, or a natural killer T cell.

49. The host cell according to claim 47 or 48, wherein the host cell is a T cell.

50. An isolated T-cell, wherein the T-cell expresses the antigen recognizing construct or the T cell receptor according to any one of claims 1 to 38, the chimeric human CD8 co-receptor according to any one of claims 1 to 38, and the chimeric human CD95 receptor according to any one of claims 1 to 38.

51. An isolated T-cell, wherein the T-cell expresses:(i) a T-cell receptor capable of binding to a Preferentially Expressed Antigen in Melanoma (PRAME) peptide, wherein the T-cell receptor comprises: a TCR a-chain comprising orconsisting of the amino acid sequence of SEQ ID NO: 107, and a TOR p-chain comprising or consisting of the amino acid sequence of SEQ ID NO: 108,(ii) a chimeric human CD8 co-receptor comprising or consisting of a polypeptide having an amino acid sequence as set forth in SEQ ID No. 133; and(iii) a chimeric human CD95 receptor comprising or consisting of a polypeptide having an amino acid sequence as set forth in SEQ ID No. 145.

52. An isolated T-cell, wherein the vector of any one of claims 44 to 46 has been introduced in said T-cell.

53. The T-cell of any one of claims 50 to 52, wherein the cell is a p T-cell, y8 T-cell, and / or a natural killer T-cell.

54. The T-cell of according to claim 52, wherein the ap T-cell is a CD4 T-cell, or wherein the ap T-cell is a CD8 T-cell, or wherein the \| / 8 T-cell is a V / 9V82+ T-cell, or wherein the \| / 8 T-cell comprises a V81 T-cell.

55. The T-cell according to any one of claims 50 to 54, wherein the T-cell is derived from an induced pluripotent stem cell (iPSCs).

56. The T-cell of any one of the claims 50 to 55 , having an enhanced cytotoxicity.

57. The T-cell of any one of the foregoing claims, wherein the T-cell is having an increased resistance to CD95L expressing cancer cells.

58. The combination according to claim 40 or 41, or the nucleic acid sequence according to claim 42 or 43, or the vector according to claim any one of claims 44 to 46, or the cell according to any one of claims 47 to 57, for use in medicine.

59. The combination according to claim 40 or 41, or the nucleic acid sequence according to claim 42 or 43, or the vector according to claim 44 to 46, or the cell according to any one of claims 47 to 57, for use in detection, diagnosis, prognosis, prevention and / or treatment of a disease.

60. Use ofi) the antigen recognizing construct or the T cell receptor according to any one of claims 1 to 38, of the chimeric human CD8 co-receptor according to any one of claims 1 to 38, and of the chimeric human CD95 receptor according to any one of claims 1 to 38, or ii) the combination according to claim 40 or 41 , oriii) the nucleic acid sequence according to claim 42 or 43, oriv) of the vector according to any one of claims 44 to 46, orv) of the cell according to any one of claims 47 to 57,for the manufacture of a medicament for treating a disease.

61. The use ofi) the antigen recognizing construct or the T cell receptor, of the chimeric human CD8 coreceptor and of the chimeric human CD95 receptor, orii) the combination, oriii) the nucleic acid sequence, oriv) of the vector according, orv) of the cellaccording claim 60, wherein the disease is a cancer.

62. The use ofi) the antigen recognizing construct or the T cell receptor, of the chimeric human CD8 coreceptor and of the chimeric human CD95 receptor, orii) the combination, oriii) the nucleic acid sequence, oriv) of the vector, orv) of the cellaccording to claim 60 or 61, wherein the disease is a tumor that expresses PRAME, optionally wherein the disease is an advanced-stage tumor that expresses PRAME.

63. The use ofi) the antigen recognizing construct or the T cell receptor, of the chimeric human CD8 coreceptor and of the chimeric human CD95 receptor, orii) the combination, oriii) the nucleic acid sequence, oriv) the vector, orv) the cellaccording to any one of claims 60 - 62, wherein the disease is selected from the group consisting of melanoma, bladder carcinoma, colon carcinoma, and breast adenocarcinoma, sarcoma, prostate cancer, uterine cancer, uveal cancer, uveal melanoma, squamous head and neck cancer, synovial carcinoma, Ewing’s sarcoma, breast cancer such as triple negative breast cancer, thyroid cancer, testicular cancer, renal cancer, pancreatic cancer, ovarian cancer, esophageal cancer, non-small-cell lung cancer (NSCLC), non-Hodgkin’s lymphoma, multiple myeloma, melanoma, hepatocellular carcinoma, head and neck cancer, gastric cancer, endometrial cancer, colorectal cancer, cholangiocarcinoma, breast cancer, bladder cancer, myeloid leukemia, acute lymphoblastic leukemia, small-cell lung cancer (SCLC), sarcoma and osteosarcoma.

64. The use ofi) the antigen recognizing construct or the T cell receptor, of the chimeric human CD8 coreceptor and of the chimeric human CD95 receptor, orii) the combination, oriii) the nucleic acid sequence, oriv) the vector, orv) the cellaccording to claim 63, wherein the breast cancer is selected from the group consisting of ductal breast cancer, tubular breast cancer, medullary breast cancer, triple negative breast cancer and combinations thereof.

65. The use ofi) the antigen recognizing construct or the T cell receptor, of the chimeric human CD8 coreceptor and of the chimeric human CD95 receptor, orii) the combination, oriii) the nucleic acid sequence, oriv) the vector, orv) of the cellaccording to claim 63, wherein the gastric cancer is gastric adenocarcinoma cancer.

66. The use ofi) the antigen recognizing construct or the T cell receptor, of the chimeric human CD8 coreceptor and of the chimeric human CD95 receptor, orii) the combination, oriii) the nucleic acid sequence, oriv) the vector, orv) the cellaccording to claim 63, wherein the sarcoma cancer is selected from the group consisting of chondrosarcoma cancer, osteosarcoma cancer, synovial sarcoma and combinations thereof.

67. A pharmaceutical composition comprising:i) the antigen recognizing construct or the T cell receptor according to any one of claims 1 to 38, the chimeric human CD8 co-receptor according to any one of claims 1 to 38, and the chimeric human CD95 receptor according to any one of claims 1 to 38, orii) the combination according to claim 40 or 41 , oriii) the nucleic acid sequence according to claim 42 or 43, oriv) the vector according to any one of claims 44 to 46, orv) the cell according to any one of claims 47 to 57.

68. The pharmaceutical composition of claim 67, wherein the composition further comprises an adjuvant, excipient, buffer, diluent, carrier, stabilizer or combination thereof.

69. A method for preparing a T-cell for immunotherapy, comprisingisolating T-cells from a human subject,introducing the vector according to any one of claims 44 to 46, or the nucleic acid according to claim 42 or 43 into the T-cell, andexpanding the T-cells.

70. The method according to claim 69, comprising transforming, transfecting or transducing the isolated T-cells with the vector, or with the nucleic acid.

71. The pharmaceutical composition according to claim 69 or 70, comprising T-cells expressing the antigen recognizing construct or the T cell receptor according to any one of claims 1 to 38, the chimeric human CD8 co-receptor according to any one of claims 1 to 38, and the chimeric human CD95 receptor according to any one of claims 1 to 38.

72. The pharmaceutical composition according to claim 71, further comprising CD4 cells expressing the antigen recognizing construct or the T cell receptor according to any one of claims 1 to 38, the chimeric human CD8 co-receptor according to any one of claims 1 to 38, and the chimeric human CD95 receptor according to any one of claims 1 to 38.9973. A method for treating a patient having a disease, comprising administering to the patient the composition of any one of claims 67, 68, 71 or 72.

74. A method for treating a patient having a disease, comprising introducing in vivo the vector according to any one of claims 44 to 46 into a T-cell of the patient.

75. The method according to claim 74, wherein the vector is DNA or an mRNA.

76. The method according to claim 74, wherein the vector is a non-replicating viral vector.

77. The method according to claim 74 or 75, wherein the vector is mRNA, and wherein the mRNA is introduced into the T-cell of the patient using nanoparticles.

78. The method according to any one of claims 73 to 77, wherein the disease is a cancer or an autoimmune disease.

79. The method according to claim 78, wherein the cancer is selected from the group consisting of non-small cell lung cancer, small cell lung cancer, pancreatic cancer, ovarian cancer, melanoma, breast cancer, liver cancer, kidney cancer, esophageal cancer, brain cancer, gastric cancer, Merkel cell carcinoma, leukemia, urinary bladder cancer, uterine cancer, colorectal cancer, gallbladder cancer, bile duct cancer, and prostate cancer.

80. The method according to claim 78 or 79, wherein cancer cells express FasL.

81. A method for increasing cytotoxicity of a T-cell in adoptive cell therapy, comprising introducing a vector according to any one of claims 44 to 46 into a T-cell.

82. A kit for use in medicine comprising:i) the antigen recognizing construct or the T cell receptor according to any one of claims 1 to 38, the chimeric human CD8 co-receptor according to any one of claims 1 to 38, and the chimeric human CD95 receptor according to any one of claims 1 to 38, orii) the combination according to claim 40 or 41 , oriii) the nucleic acid sequence according to claim 42 or 43, oriv) the vector according to any one of claims 44 to 46, orv) the cell according to any one of claims 47 to 57.

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