RESUMO
Adoptive cell therapy using engineered T cell receptors (TCRs) is a promising approach for targeting cancer antigens, but tumor-reactive TCRs are often weakly responsive to their target ligands, peptide-major histocompatibility complexes (pMHCs). Affinity-matured TCRs can enhance the efficacy of TCR-T cell therapy but can also cross-react with off-target antigens, resulting in organ immunopathology. We developed an alternative strategy to isolate TCR mutants that exhibited high activation signals coupled with low-affinity pMHC binding through the acquisition of catch bonds. Engineered analogs of a tumor antigen MAGE-A3-specific TCR maintained physiological affinities while exhibiting enhanced target killing potency and undetectable cross-reactivity, compared with a high-affinity clinically tested TCR that exhibited lethal cross-reactivity with a cardiac antigen. Catch bond engineering is a biophysically based strategy to tune high-sensitivity TCRs for T cell therapy with reduced potential for adverse cross-reactivity.
Assuntos
Imunoterapia Adotiva , Receptores de Antígenos de Linfócitos T , Linfócitos T , Antígenos de Neoplasias , Reações Cruzadas , Complexo Principal de Histocompatibilidade , Miocárdio/imunologia , Peptídeos , Linfócitos T/metabolismoRESUMO
The ideal cancer target antigen (Ag) is expressed at high copy numbers on neoplastic cells, absent on normal tissues, and contributes to the survival of cancer cells. Despite significant investments in the identification of cell surface Ags, there is a paucity of targets that meet such ideal cancer target criteria. Recent clinical trials in patients with cancer treated with immune checkpoint inhibitors (ICIs) indicate that cluster of differentiation (CD)8+ T cells, by means of their T cell receptors (TCRs) recognizing intracellular targets presented as peptides in the context of human leukocyte antigen (peptide-human leukocyte antigen complex; pHLA) molecules on tumor cells, can mediate deep and long-lasting antitumor responses in patients with solid tumors. Therefore, pHLA-target Ags may represent the long sought-after, ideal targets for solid tumor targeting by high-potency oncology compounds.
Assuntos
Antígenos de Neoplasias/metabolismo , Antineoplásicos Imunológicos/farmacologia , Descoberta de Drogas/métodos , Neoplasias/tratamento farmacológico , Receptores de Antígenos de Linfócitos T/metabolismo , Animais , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/imunologia , Antineoplásicos Imunológicos/uso terapêutico , Linhagem Celular Tumoral , Simulação por Computador , Reações Cruzadas/imunologia , Mapeamento de Epitopos/métodos , Epitopos de Linfócito T/genética , Epitopos de Linfócito T/imunologia , Antígenos HLA/genética , Antígenos HLA/imunologia , Antígenos HLA/metabolismo , Humanos , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/uso terapêutico , Ligantes , Neoplasias/imunologia , Neoplasias/patologia , Biblioteca de Peptídeos , Receptores de Antígenos de Linfócitos T/imunologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Linfócitos T/metabolismoRESUMO
T cell receptor cross-reactivity allows a fixed T cell repertoire to respond to a much larger universe of potential antigens. Recent work has emphasized the importance of peptide structural and chemical homology, as opposed to sequence similarity, in T cell receptor cross-reactivity. Surprisingly, though, T cell receptors can also cross-react between ligands with little physiochemical commonalities. Studying the clinically relevant receptor DMF5, we demonstrate that cross-recognition of such divergent antigens can occur through mechanisms that involve heretofore unanticipated rearrangements in the peptide and presenting MHC protein, including binding-induced peptide register shifts and extensions from MHC peptide binding grooves. Moreover, cross-reactivity can proceed even when such dramatic rearrangements do not translate into structural or chemical molecular mimicry. Beyond demonstrating new principles of T cell receptor cross-reactivity, our results have implications for efforts to predict and control T cell specificity and cross-reactivity and highlight challenges associated with predicting T cell reactivities.
Assuntos
Oligopeptídeos/química , Receptores de Antígenos de Linfócitos T/química , Antígenos/química , Autoimunidade , Reações Cruzadas , Cristalografia por Raios X , Epitopos/química , Humanos , Cinética , Ligantes , Mimetismo Molecular , Ligação Proteica , Domínios Proteicos , Retroviridae , Ressonância de Plasmônio de Superfície , Linfócitos T/químicaRESUMO
TCR-signaling strength generally correlates with peptide-MHC binding affinity; however, exceptions exist. We find high-affinity, yet non-stimulatory, interactions occur with high frequency in the human T cell repertoire. Here, we studied human TCRs that are refractory to activation by pMHC ligands despite robust binding. Analysis of 3D affinity, 2D dwell time, and crystal structures of stimulatory versus non-stimulatory TCR-pMHC interactions failed to account for their different signaling outcomes. Using yeast pMHC display, we identified peptide agonists of a formerly non-responsive TCR. Single-molecule force measurements demonstrated the emergence of catch bonds in the activating TCR-pMHC interactions, correlating with exclusion of CD45 from the TCR-APC contact site. Molecular dynamics simulations of TCR-pMHC disengagement distinguished agonist from non-agonist ligands based on the acquisition of catch bonds within the TCR-pMHC interface. The isolation of catch bonds as a parameter mediating the coupling of TCR binding and signaling has important implications for TCR and antigen engineering for immunotherapy.
Assuntos
Antígenos de Histocompatibilidade Classe I/fisiologia , Ativação Linfocitária/fisiologia , Adulto , Feminino , Humanos , Cinética , Ligantes , Complexo Principal de Histocompatibilidade/fisiologia , Masculino , Pessoa de Meia-Idade , Simulação de Dinâmica Molecular , Oligopeptídeos , Peptídeos , Ligação Proteica/fisiologia , Receptores de Antígenos de Linfócitos T/metabolismo , Receptores de Antígenos de Linfócitos T/fisiologia , Transdução de Sinais , Imagem Individual de Molécula , Linfócitos T/fisiologiaRESUMO
Cancer immunotherapy based on genetically redirecting T cells has been used successfully to treat B cell malignancies1-3. In this strategy, the T cell genome is modified by integration of viral vectors or transposons encoding chimaeric antigen receptors (CARs) that direct tumour cell killing. However, this approach is often limited by the extent of expansion and persistence of CAR T cells4,5. Here we report mechanistic insights from studies of a patient with chronic lymphocytic leukaemia treated with CAR T cells targeting the CD19 protein. Following infusion of CAR T cells, anti-tumour activity was evident in the peripheral blood, lymph nodes and bone marrow; this activity was accompanied by complete remission. Unexpectedly, at the peak of the response, 94% of CAR T cells originated from a single clone in which lentiviral vector-mediated insertion of the CAR transgene disrupted the methylcytosine dioxygenase TET2 gene. Further analysis revealed a hypomorphic mutation in this patient's second TET2 allele. TET2-disrupted CAR T cells exhibited an epigenetic profile consistent with altered T cell differentiation and, at the peak of expansion, displayed a central memory phenotype. Experimental knockdown of TET2 recapitulated the potency-enhancing effect of TET2 dysfunction in this patient's CAR T cells. These findings suggest that the progeny of a single CAR T cell induced leukaemia remission and that TET2 modification may be useful for improving immunotherapies.
Assuntos
5-Metilcitosina/metabolismo , Antígenos CD19/imunologia , Dioxigenases/genética , Imunoterapia/métodos , Leucemia Linfocítica Crônica de Células B/imunologia , Leucemia Linfocítica Crônica de Células B/terapia , Linfócitos T/imunologia , Linfócitos T/transplante , Transferência Adotiva , Idoso , Alelos , Diferenciação Celular , Ensaios Clínicos como Assunto , Células Clonais/citologia , Células Clonais/imunologia , Dioxigenases/metabolismo , Epigênese Genética , Células HEK293 , Humanos , Leucemia Linfocítica Crônica de Células B/genética , Leucemia Linfocítica Crônica de Células B/patologia , Masculino , Mutação , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Linfócitos T/citologia , Linfócitos T/metabolismo , TransgenesRESUMO
The immune system can mount T cell responses against tumors; however, the antigen specificities of tumor-infiltrating lymphocytes (TILs) are not well understood. We used yeast-display libraries of peptide-human leukocyte antigen (pHLA) to screen for antigens of "orphan" T cell receptors (TCRs) expressed on TILs from human colorectal adenocarcinoma. Four TIL-derived TCRs exhibited strong selection for peptides presented in a highly diverse pHLA-A∗02:01 library. Three of the TIL TCRs were specific for non-mutated self-antigens, two of which were present in separate patient tumors, and shared specificity for a non-mutated self-antigen derived from U2AF2. These results show that the exposed recognition surface of MHC-bound peptides accessible to the TCR contains sufficient structural information to enable the reconstruction of sequences of peptide targets for pathogenic TCRs of unknown specificity. This finding underscores the surprising specificity of TCRs for their cognate antigens and enables the facile indentification of tumor antigens through unbiased screening.
Assuntos
Adenocarcinoma/imunologia , Antígenos de Neoplasias/imunologia , Neoplasias Colorretais/imunologia , Linfócitos do Interstício Tumoral/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , Idoso , Animais , Antígenos de Neoplasias/química , Linhagem Celular Tumoral , Células Cultivadas , Células HEK293 , Antígenos HLA-A/química , Antígenos HLA-A/imunologia , Humanos , Masculino , Pessoa de Meia-Idade , Biblioteca de Peptídeos , Células Sf9 , SpodopteraRESUMO
T cells engineered to express a tumor-specific αß T cell receptor (TCR) mediate anti-tumor immunity. However, mispairing of the therapeutic αß chains with endogenous αß chains reduces therapeutic TCR surface expression and generates self-reactive TCRs. We report a general strategy to prevent TCR mispairing: swapping constant domains between the α and ß chains of a therapeutic TCR. When paired, domain-swapped (ds)TCRs assemble with CD3, express on the cell surface, and mediate antigen-specific T cell responses. By contrast, dsTCR chains mispaired with endogenous chains cannot properly assemble with CD3 or signal, preventing autoimmunity. We validate this approach in cell-based assays and in a mouse model of TCR gene transfer-induced graft-versus-host disease. We also validate a related approach whereby replacement of αß TCR domains with corresponding γδ TCR domains yields a functional TCR that does not mispair. This work enables the design of safer TCR gene therapies for cancer immunotherapy.
Assuntos
Genes Codificadores dos Receptores de Linfócitos T , Terapia Genética/efeitos adversos , Terapia Genética/métodos , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Animais , Modelos Animais de Doenças , Doença Enxerto-Hospedeiro , Camundongos , Domínios Proteicos , Recombinação GenéticaRESUMO
The T cell antigen receptor (TCR)-peptide-major histocompatibility complex (MHC) interface is composed of conserved and diverse regions, yet the relative contribution of each in shaping recognition by T cells remains unclear. Here we isolated cross-reactive peptides with limited homology, which allowed us to compare the structural properties of nine peptides for a single TCR-MHC pair. The TCR's cross-reactivity was rooted in highly similar recognition of an apical 'hot-spot' position in the peptide with tolerance of sequence variation at ancillary positions. Furthermore, we found a striking structural convergence onto a germline-mediated interaction between the TCR CDR1α region and the MHC α2 helix in twelve TCR-peptide-MHC complexes. Our studies suggest that TCR-MHC germline-mediated constraints, together with a focus on a small peptide hot spot, might place limits on peptide antigen cross-reactivity.