RESUMO
Tumors undergo dynamic immunoediting as part of a process that balances immunologic sensing of emerging neoantigens and evasion from immune responses. Tumor-infiltrating lymphocytes (TIL) comprise heterogeneous subsets of peripheral T cells characterized by diverse functional differentiation states and dependence on T-cell receptor (TCR) specificity gained through recombination events during their development. We hypothesized that within the tumor microenvironment (TME), an antigenic milieu and immunologic interface, tumor-infiltrating peripheral T cells could reexpress key elements of the TCR recombination machinery, namely, Rag1 and Rag2 recombinases and Tdt polymerase, as a potential mechanism involved in the revision of TCR specificity. Using two syngeneic invasive breast cancer transplantable models, 4T1 and TS/A, we observed that Rag1, Rag2, and Dntt in situ mRNA expression characterized rare tumor-infiltrating T cells. In situ expression of the transcripts was increased in coisogenic Mlh1-deficient tumors, characterized by genomic overinstability, and was also modulated by PD-1 immune-checkpoint blockade. Through immunolocalization and mRNA hybridization analyses, we detected the presence of rare TDT+RAG1/2+ cells populating primary tumors and draining lymph nodes in human invasive breast cancer. Analysis of harmonized single-cell RNA-sequencing data sets of human cancers identified a very small fraction of tumor-associated T cells, characterized by the expression of recombination/revision machinery transcripts, which on pseudotemporal ordering corresponded to differentiated effector T cells. We offer thought-provoking evidence of a TIL microniche marked by rare transcripts involved in TCR shaping.
Assuntos
Neoplasias da Mama/imunologia , Linfócitos T CD8-Positivos/imunologia , Linfócitos do Interstício Tumoral/imunologia , Recombinação Genética/imunologia , Especificidade do Receptor de Antígeno de Linfócitos T/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Mama/imunologia , Mama/patologia , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Linfócitos T CD8-Positivos/metabolismo , Dano ao DNA/imunologia , DNA Nucleotidilexotransferase/genética , DNA Nucleotidilexotransferase/metabolismo , Proteínas de Ligação a DNA/metabolismo , Conjuntos de Dados como Assunto , Modelos Animais de Doenças , Feminino , Proteínas de Homeodomínio/metabolismo , Humanos , Linfócitos do Interstício Tumoral/metabolismo , Camundongos , Camundongos Knockout , Pessoa de Meia-Idade , Proteína 1 Homóloga a MutL/genética , Proteína 1 Homóloga a MutL/metabolismo , Proteínas Nucleares/metabolismo , RNA-Seq , Receptores de Antígenos de Linfócitos T , Análise de Célula Única , Microambiente Tumoral/genética , Microambiente Tumoral/imunologiaRESUMO
Chimeric antigen receptor (CAR) T-cell immunotherapy is rapidly emerging as a promising novel treatment for malignancies. To broaden the success of CAR T-cell treatment for chronic myeloid leukaemia (CML), we attempted to construct a CD26 CAR T-cell product to target tyrosine kinase inhibitor-insensitive leukaemia stem cells (LSCs), which have been a challenge to cure for several decades and can be discriminated from healthy stem cells by the robust biomarker CD26. Of additional interest is that CD26 has also been reported to be a multi-purpose therapeutic target for other malignancies. Here, we constructed CD26 CAR T cells utilizing lentiviral transduction methods and verified them by flow cytometry analysis and RNA-seq. We found that the initial expansion of CD26 CAR-transduced T cells was delayed due to transient fratricide, but subsequent expansion was accelerated. CD26 CAR T cells exhibited cytotoxicity against the CD26+ T-cell lymphoma cell line Karpas 299, CD26-overexpressing K562 cells and primary CML LSCs, activated multiple effector functions in co-culture assays, and limited tumour progression in a mouse model; but there was some off-tumour cytotoxicity towards activated lymphocytes. In conclusion, these results establish the feasibility of using CD26 as an antigen for CAR T cells targeting CD26+ tumour cells.
Assuntos
Dipeptidil Peptidase 4/imunologia , Imunoterapia Adotiva , Receptores de Antígenos de Linfócitos T/imunologia , Receptores de Antígenos Quiméricos/imunologia , Especificidade do Receptor de Antígeno de Linfócitos T/imunologia , Linfócitos T/imunologia , Linfócitos T/metabolismo , Animais , Biomarcadores , Linhagem Celular Tumoral , Citotoxicidade Imunológica , Modelos Animais de Doenças , Perfilação da Expressão Gênica , Humanos , Imunofenotipagem , Imunoterapia Adotiva/efeitos adversos , Imunoterapia Adotiva/métodos , Camundongos , Neoplasias/imunologia , Neoplasias/terapia , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos Quiméricos/genética , Especificidade do Receptor de Antígeno de Linfócitos T/genética , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
MR1 is an MHC class I-like molecule with unique structural and biological features that make it an important member among the molecules involved in antigen presentation to T cells. Distinctive features include ubiquitous expression of the MR1 gene and its monomorphism. Another relevant property is that the MR1 protein appears at very low levels on the plasma membrane and its surface expression is regulated by antigen binding. Finally, the nature of presented antigens differs from those that bind other presenting molecules and includes small metabolites of microbial and self-origin, small drugs and tumor-associated antigens. This opinion paper describes in detail some of those features and discusses recent literature in the field.
Assuntos
Apresentação de Antígeno , Antígenos de Histocompatibilidade Classe I/metabolismo , Antígenos de Histocompatibilidade Menor/metabolismo , Especificidade do Receptor de Antígeno de Linfócitos T , Linfócitos T/metabolismo , Apresentação de Antígeno/genética , Apresentação de Antígeno/imunologia , Antígenos de Bactérias/imunologia , Antígenos de Bactérias/metabolismo , Antígenos de Histocompatibilidade Classe I/química , Antígenos de Histocompatibilidade Classe I/genética , Antígenos de Histocompatibilidade Classe I/imunologia , Humanos , Ligantes , Ativação Linfocitária/genética , Ativação Linfocitária/imunologia , Antígenos de Histocompatibilidade Menor/química , Antígenos de Histocompatibilidade Menor/genética , Antígenos de Histocompatibilidade Menor/imunologia , Ligação Proteica , Estrutura Terciária de Proteína , Especificidade do Receptor de Antígeno de Linfócitos T/genética , Especificidade do Receptor de Antígeno de Linfócitos T/imunologia , Linfócitos T/imunologiaRESUMO
High-throughput T cell receptor (TCR) sequencing allows the characterization of an individual's TCR repertoire and directly queries their immune state. However, it remains a non-trivial task to couple these sequenced TCRs to their antigenic targets. In this paper, we present a novel strategy to annotate full TCR sequence repertoires with their epitope specificities. The strategy is based on a machine learning algorithm to learn the TCR patterns common to the recognition of a specific epitope. These results are then combined with a statistical analysis to evaluate the occurrence of specific epitope-reactive TCR sequences per epitope in repertoire data. In this manner, we can directly study the capacity of full TCR repertoires to target specific epitopes of the relevant vaccines or pathogens. We demonstrate the usability of this approach on three independent datasets related to vaccine monitoring and infectious disease diagnostics by independently identifying the epitopes that are targeted by the TCR repertoire. The developed method is freely available as a web tool for academic use at tcrex.biodatamining.be.
Assuntos
Epitopos de Linfócito T/imunologia , Modelos Biológicos , Receptores de Antígenos de Linfócitos T/genética , Especificidade do Receptor de Antígeno de Linfócitos T/genética , Especificidade do Receptor de Antígeno de Linfócitos T/imunologia , Linfócitos T/imunologia , Linfócitos T/metabolismo , Algoritmos , Sequência de Aminoácidos , Evolução Clonal/genética , Bases de Dados Genéticas , Epitopos de Linfócito T/química , Humanos , Receptores de Antígenos de Linfócitos T/metabolismo , Reprodutibilidade dos Testes , Software , NavegadorRESUMO
High-throughput 3' single-cell RNA-sequencing (scRNA-seq) allows cost-effective, detailed characterization of individual immune cells from tissues. Current techniques, however, are limited in their ability to elucidate essential immune cell features, including variable sequences of T cell antigen receptors (TCRs) that confer antigen specificity. Here, we present a strategy that enables simultaneous analysis of TCR sequences and corresponding full transcriptomes from 3'-barcoded scRNA-seq samples. This approach is compatible with common 3' scRNA-seq methods, and adaptable to processed samples post hoc. We applied the technique to identify transcriptional signatures associated with T cells sharing common TCRs from immunized mice and from patients with food allergy. We observed preferential phenotypes among subsets of expanded clonotypes, including type 2 helper CD4+ T cell (TH2) states associated with food allergy. These results demonstrate the utility of our method when studying diseases in which clonotype-driven responses are critical to understanding the underlying biology.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Hipersensibilidade a Amendoim/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Células Th2/imunologia , Albuminas 2S de Plantas/imunologia , Animais , Antígenos de Plantas/imunologia , Células Cultivadas , Regiões Determinantes de Complementaridade/genética , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Imunização , Imunoglobulina E/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Proteínas E7 de Papillomavirus/imunologia , Análise de Célula Única , Especificidade do Receptor de Antígeno de Linfócitos T/genética , TranscriptomaRESUMO
Adoptive immunotherapy based on chimeric antigen receptor-modified T (CAR-T) cells has been demonstrated as one of the most promising therapeutic strategies in the treatment of malignancies. However, CAR-T cell therapy has shown limited efficacy for the treatment of solid tumors. This is, in part, because of tumor heterogeneity and a hostile tumor microenvironment, which could suppress adoptively transferred T cell activity. In this study, we, respectively, engineered human- or murine-derived-armored glypican-3 (GPC3)-specific CAR-T cells capable of inducibly expressing IL-12 (GPC3-28Z-NFAT-IL-12) T cells. The results showed that GPC3-28Z-NFAT-IL-12 T cells could lyse GPC3+ tumor cells specifically and increase cytokine secretion compared with GPC3-28Z T cells in vitro. In vivo, GPC3-28Z-NFAT-IL-12 T cells augmented the antitumor effect when encountering GPC3+ large tumor burdens, which could be attributed to IL-12 increasing IFN-γ production, favoring T cells infiltration and persistence. Furthermore, in immunocompetent hosts, low doses of GPC3-m28Z-mNFAT-mIL-12 T cells exerted superior antitumor efficacy without prior conditioning in comparison with GPC3-m28Z T cells. Also, mIL-12 secretion decreased regulatory T cell infiltration in established tumors. In conclusion, these findings demonstrated that the inducible expression of IL-12 could boost CAR-T function with less potential side effects, both in immunodeficient and immunocompetent hosts. The inducibly expressed IL-12-armored GPC3-CAR-T cells could broaden the application of CAR-T-based immunotherapy to patients intolerant of lymphodepletion chemotherapy and might provide an alternative therapeutic strategy for patients with GPC3+ cancers.
Assuntos
Carcinoma Hepatocelular/terapia , Glipicanas/metabolismo , Imunoterapia Adotiva/métodos , Interleucina-12/metabolismo , Neoplasias Hepáticas/terapia , Linfócitos do Interstício Tumoral/fisiologia , Animais , Carcinoma Hepatocelular/imunologia , Glipicanas/genética , Glipicanas/imunologia , Células HEK293 , Humanos , Interleucina-12/genética , Interleucina-12/imunologia , Neoplasias Hepáticas/imunologia , Linfócitos do Interstício Tumoral/transplante , Camundongos , Camundongos Endogâmicos C57BL , Engenharia de Proteínas , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Especificidade do Receptor de Antígeno de Linfócitos T/genética , Microambiente Tumoral , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The experimental autoimmune encephalomyelitis (EAE) model is indispensable for autoimmunity research, but model-specific T cell dynamics are sparsely studied. We used next-generation immunosequencing across lymphoid organs, blood and spinal cord in response to immunization with myelin basic protein (MBP) to study T cell repertoires and migration patterns. Surprisingly, most spinal cord T cells were unique to the individual animal despite the existence of shared MBP-specific clones, suggesting a previously underestimated T cell diversity. Almost complete emigration of pathogenic clones from blood to spinal cord indicates that blood is not a suitable compartment to study EAE-mediating T cells.
Assuntos
Seleção Clonal Mediada por Antígeno/genética , Encefalomielite Autoimune Experimental/imunologia , Especificidade do Receptor de Antígeno de Linfócitos T/genética , Subpopulações de Linfócitos T/imunologia , Animais , Autoantígenos/imunologia , Células Sanguíneas/imunologia , Células Sanguíneas/patologia , Movimento Celular , Células Clonais/imunologia , Encefalomielite Autoimune Experimental/sangue , Encefalomielite Autoimune Experimental/patologia , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Tecido Linfoide/imunologia , Tecido Linfoide/patologia , Camundongos , Proteína Básica da Mielina/imunologia , Fragmentos de Peptídeos/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Organismos Livres de Patógenos Específicos , Medula Espinal/imunologia , Medula Espinal/patologiaRESUMO
Although the adaptive immune system can detect and eliminate malignant cells, patients with intact and fully functional immune systems develop head and neck cancer. How is this paradox explained? Manuscripts published in the English language from 1975 to 2018 were reviewed using search inputs related to tumor cell antigenicity and immunogenicity, immunodominance, cancer immunoediting and genomic alterations present within carcinomas. Early in tumor development, T cell responses to immunodominant antigens may lead to the elimination of cancer cells expressing these antigens and a tumor composed to tumor cells expressing only immunorecessive antigens. Conversely, other tumor cells may acquire genomic or epigenetic alterations that result in an antigen processing or presentation defect or other inability to be detected or killed by T cells. Such T cell insensitive tumor cells may also be selected for in a progressing tumor. Tumors harboring subpopulations of cells that cannot be eliminated by T cells may require non-T cell-based treatments, such as NK cell immunotherapies. Recognition of such tumor cell populations within a heterogeneous cancer may inform the selection of treatment for HNSCC in the future.
Assuntos
Suscetibilidade a Doenças , Neoplasias de Cabeça e Pescoço/etiologia , Neoplasias de Cabeça e Pescoço/metabolismo , Sistema Imunitário/imunologia , Sistema Imunitário/metabolismo , Animais , Antígenos de Neoplasias/imunologia , Suscetibilidade a Doenças/imunologia , Regulação Neoplásica da Expressão Gênica , Neoplasias de Cabeça e Pescoço/patologia , Neoplasias de Cabeça e Pescoço/terapia , Humanos , Imunoterapia , Transdução de Sinais , Especificidade do Receptor de Antígeno de Linfócitos T/genética , Linfócitos T/imunologia , Linfócitos T/metabolismo , Evasão Tumoral/genética , Microambiente TumoralRESUMO
The antigenic specificity of T cells occurs via generation and rearrangement of different gene segments producing a functional T cell receptor (TCR). High-throughput sequencing (HTS) allows in-depth assessment of TCR repertoire patterns. There are limited data concerning whether TCR repertoires are altered in inflammatory bowel disease. We hypothesized that pediatric ulcerative colitis (UC) patients possess unique TCR repertoires, resulting from clonotypical expansions in the gut. Paired blood and rectal samples were collected from nine newly diagnosed treatment-naive pediatric UC patients and four healthy controls. DNA was isolated to determine the TCR-ß repertoire by HTS. Significant clonal expansion was demonstrated in UC patients, with inverse correlation between clinical disease severity and repertoire diversity in the gut. Using different repertoire variables in rectal biopsies, a clear segregation was observed between patients with severe UC, those with mild-moderate disease and healthy controls. Moreover, the overlap between autologous blood-rectal samples in UC patients was significantly higher compared with overlap among controls. Finally, we identified several clonotypes that were shared in either all or the majority of UC patients in the colon. Clonal expansion of TCR-ß-expressing T cells among UC patients correlates with disease severity and highlights their involvement in mediating intestinal inflammation.
Assuntos
Células Clonais/fisiologia , Colite Ulcerativa/imunologia , Colo/imunologia , Genes Codificadores da Cadeia beta de Receptores de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T alfa-beta/metabolismo , Especificidade do Receptor de Antígeno de Linfócitos T/genética , Linfócitos T/fisiologia , Adolescente , Proliferação de Células , Criança , Seleção Clonal Mediada por Antígeno , Colite Ulcerativa/genética , DNA/análise , Progressão da Doença , Humanos , Ativação Linfocitária , Receptores de Antígenos de Linfócitos T alfa-beta/genéticaRESUMO
CD20 is an effective immunotherapy target for CD20+ B-cell malignant cells. Monoclonal antibody, especially rituximab, has been a conventional strategy in the treatment of B-cell malignancies such as non-Hodgkin's lymphoma. However, treatment with monoclonal antibodies has not been enough to overcome the refractory/relapse problems. Chimeric antigen receptor engineered T (CAR-T) cells have exhibited excellent therapeutic effect on lymphocytic leukemia in recent years. In this study, a CD20-specific CAR was constructed and the cytotoxic efficacy of CD20 CAR-T cells on B-cell malignant cells was evaluated by CD107a degranulation, pro-inflammation cytokine production, and true lytic ability in vitro and in vivo. It was found that CD20 CAR-T cells possessed stronger cytotoxic ability against CD20 highly expressed cells. Furthermore, when histone deacetylase inhibitor was used to enhance the expression of CD20 antigen on the surface of B-cell malignant cells via inducing acetylation of H3K9 on CD20 promoter site, it revealed that the cytotoxicity of CD20 CAR-T cells against histone deacetylase inhibitor-treated B-cell malignant cells was significantly enhanced.
Assuntos
Antígenos CD20/genética , Linfócitos B/imunologia , Linfócitos B/metabolismo , Citotoxicidade Imunológica , Expressão Gênica , Receptores de Antígenos Quiméricos/metabolismo , Linfócitos T/imunologia , Linfócitos T/metabolismo , Animais , Antineoplásicos/farmacologia , Linfócitos B/patologia , Biomarcadores , Linhagem Celular Tumoral , Terapia Combinada , Citotoxicidade Imunológica/efeitos dos fármacos , Modelos Animais de Doenças , Feminino , Genes Reporter , Inibidores de Histona Desacetilases/farmacologia , Humanos , Imunofenotipagem , Imunoterapia Adotiva/métodos , Linfoma de Células B/genética , Linfoma de Células B/imunologia , Linfoma de Células B/terapia , Camundongos , Especificidade do Receptor de Antígeno de Linfócitos T/genética , Especificidade do Receptor de Antígeno de Linfócitos T/imunologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Patients with high risk neuroblastoma have a poor prognosis and survivors are often left with debilitating long term sequelae from treatment. Even after integration of anti-GD2 monoclonal antibody therapy into standard, upftont protocols, 5-year overall survival rates are only about 50%. The success of anti-GD2 therapy has proven that immunotherapy can be effective in neuroblastoma. Adoptive transfer of chimeric antigen receptor (CAR) T cells has the potential to build on this success. In early phase clinical trials, CAR T cell therapy for neuroblastoma has proven safe and feasible, but significant barriers to efficacy remain. These include lack of T cell persistence and potency, difficulty in target identification, and an immunosuppressive tumor microenvironment. With recent advances in CAR T cell engineering, many of these issues are being addressed in the laboratory. In this review, we summarize the clinical trials that have been completed or are underway for CAR T cell therapy in neuroblastoma, discuss the conclusions and open questions derived from these trials, and consider potential strategies to improve CAR T cell therapy for patients with neuroblastoma.
Assuntos
Imunoterapia Adotiva , Neuroblastoma/imunologia , Neuroblastoma/terapia , Receptores de Antígenos de Linfócitos T/metabolismo , Receptores de Antígenos Quiméricos/metabolismo , Linfócitos T/imunologia , Linfócitos T/metabolismo , Fatores Etários , Animais , Antígenos de Neoplasias/imunologia , Ensaios Clínicos como Assunto , Engenharia Genética , Humanos , Imunoterapia Adotiva/efeitos adversos , Imunoterapia Adotiva/métodos , Neuroblastoma/patologia , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos Quiméricos/genética , Especificidade do Receptor de Antígeno de Linfócitos T/genética , Especificidade do Receptor de Antígeno de Linfócitos T/imunologia , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Resultado do TratamentoRESUMO
BACKGROUND: Merkel cell carcinoma (MCC) is an aggressive skin cancer that frequently responds to anti-PD-1 therapy. MCC is associated with sun exposure and, in 80% of cases, Merkel cell polyomavirus (MCPyV). MCPyV-specific T and B cell responses provide a unique opportunity to study cancer-specific immunity throughout PD-1 blockade therapy. METHODS: Immune responses were assessed in patients (n = 26) with advanced MCC receiving pembrolizumab. Peripheral blood mononuclear cells (PBMC) were collected at baseline and throughout treatment. MCPyV-oncoprotein antibodies were quantified and T cells were assessed for MCPyV-specificity via tetramer staining and/or cytokine secretion. Pre-treatment tumor biopsies were analyzed for T cell receptor clonality. RESULTS: MCPyV oncoprotein antibodies were detectable in 15 of 17 (88%) of virus-positive MCC (VP-MCC) patients. Antibodies decreased in 10 of 11 (91%) patients with responding tumors. Virus-specific T cells decreased over time in patients who had a complete response, and increased in patients who had persistent disease. Tumors that were MCPyV(+) had a strikingly more clonal (less diverse) intratumoral TCR repertoire than virus-negative tumors (p = 0.0001). CONCLUSIONS: Cancer-specific T and B cell responses generally track with disease burden during PD-1 blockade, in proportion to presence of antigen. Intratumoral TCR clonality was significantly greater in VP-MCC than VN-MCC tumors, suggesting expansion of a limited number of dominant clones in response to fewer immunogenic MCPyV antigens. In contrast, VN-MCC tumors had lower clonality, suggesting a diverse T cell response to numerous neoantigens. These findings reveal differences in tumor-specific immunity for VP-MCC and VN-MCC, both of which often respond to anti-PD-1 therapy.
Assuntos
Carcinoma de Célula de Merkel/tratamento farmacológico , Carcinoma de Célula de Merkel/etiologia , Poliomavírus das Células de Merkel/imunologia , Infecções por Polyomavirus/complicações , Infecções por Polyomavirus/imunologia , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Infecções Tumorais por Vírus/complicações , Infecções Tumorais por Vírus/imunologia , Antineoplásicos Imunológicos/farmacologia , Antineoplásicos Imunológicos/uso terapêutico , Linfócitos B/efeitos dos fármacos , Linfócitos B/imunologia , Linfócitos B/metabolismo , Biomarcadores Tumorais , Carcinoma de Célula de Merkel/diagnóstico , Humanos , Imunomodulação/efeitos dos fármacos , Ativação Linfocitária/imunologia , Terapia de Alvo Molecular , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T/metabolismo , Especificidade do Receptor de Antígeno de Linfócitos T/genética , Especificidade do Receptor de Antígeno de Linfócitos T/imunologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Linfócitos T/metabolismo , Resultado do TratamentoRESUMO
IgE is the key mediator of allergic responses. Omalizumab, an IgE-specific monoclonal antibody that depletes IgE, is effective for treating severe allergic asthma. The need for frequent administration of the expensive drug, however, limits its applications. Taking advantage of T cell memory, adoptive T cell therapy (ACT) targeting IgE-producing cells has the potential to achieve long-term suppression of IgE and relief of symptoms for severe allergic diseases. The transmembrane form of IgE (mIgE), which is present on all IgE-producing cells, serves as an excellent molecular target for ACT that employs chimeric antigen receptors (CARs). Here, we designed and tested CARs that use the extracellular domain of high affinity IgE receptor, FcεRIα, for mIgE recognition. When expressed on Jurkat T cells, FcεRIα-based CARs mediated robust responses in terms of CD69 upregulation to U266 myeloma cells expressing low levels of mIgE. FcεRIα-based CARs specifically recognized cells expressing mIgE, but not cells with secreted IgE captured through Fcε receptors. CAR+ Jurkat cells did not respond to LAD2 mast cells with secreted IgE bound through FcεRI or Ramos cells with secreted IgE bound through FcεRII. Co-culture of CAR+ Jurkat cells and LAD2 mast cells with IgE bound did not trigger LAD2 cell degranulation. The activity of CAR using wild type FcεRIα for mIgE binding was inhibited by the presence secreted IgE, which likely blocked CAR-mIgE interaction. The activities of CARs using low affinity mutants of FcεRIα, however, tolerated secreted IgE at relatively high concentrations. Moreover, primary human CD8+ T cells expressing a low affinity mutant CAR responded to U266 cells with INFγ production and cytotoxicity despite the presence of secreted IgE. The potency, specificity, and robustness of our CAR design, combined with repaid advances in the safety of ACT, hold promise for novel and highly effective cell-based therapies against severe allergic diseases.
Assuntos
Imunoglobulina E/imunologia , Receptores de Antígenos Quiméricos/imunologia , Receptores de IgE/imunologia , Especificidade do Receptor de Antígeno de Linfócitos T/imunologia , Afinidade de Anticorpos/imunologia , Citotoxicidade Celular Dependente de Anticorpos/imunologia , Asma/imunologia , Asma/terapia , Linhagem Celular , Linhagem Celular Tumoral , Humanos , Imunoglobulina E/genética , Imunoglobulina E/metabolismo , Imunoterapia Adotiva/métodos , Células Jurkat , Mutação , Receptores de Antígenos Quiméricos/genética , Receptores de Antígenos Quiméricos/metabolismo , Receptores de IgE/genética , Receptores de IgE/metabolismo , Especificidade do Receptor de Antígeno de Linfócitos T/genética , Linfócitos T/imunologia , Linfócitos T/metabolismoRESUMO
Engineering T cells with chimeric antigen receptors (CARs) is an effective method for directing T cells to attack tumors, but may cause adverse side effects such as the potentially lethal cytokine release syndrome. Here the authors show that the T cell antigen coupler (TAC), a chimeric receptor that co-opts the endogenous TCR, induces more efficient anti-tumor responses and reduced toxicity when compared with past-generation CARs. TAC-engineered T cells induce robust and antigen-specific cytokine production and cytotoxicity in vitro, and strong anti-tumor activity in a variety of xenograft models including solid and liquid tumors. In a solid tumor model, TAC-T cells outperform CD28-based CAR-T cells with increased anti-tumor efficacy, reduced toxicity, and faster tumor infiltration. Intratumoral TAC-T cells are enriched for Ki-67+ CD8+ T cells, demonstrating local expansion. These results indicate that TAC-T cells may have a superior therapeutic index relative to CAR-T cells.
Assuntos
Receptores de Antígenos/imunologia , Receptores de Antígenos Quiméricos/imunologia , Proteínas Recombinantes/imunologia , Especificidade do Receptor de Antígeno de Linfócitos T/imunologia , Linfócitos T/imunologia , Transferência Adotiva , Animais , Antígenos CD28/imunologia , Linhagem Celular Tumoral , Citocinas/sangue , Citocinas/metabolismo , Citotoxicidade Imunológica , Feminino , Engenharia Genética , Células HEK293 , Humanos , Imunoterapia Adotiva/métodos , Lentivirus/genética , Ativação Linfocitária , Masculino , Camundongos , Camundongos Endogâmicos NOD , Engenharia de Proteínas , Receptor ErbB-2/imunologia , Receptores de Antígenos/genética , Receptores de Antígenos Quiméricos/genética , Anticorpos de Domínio Único , Especificidade do Receptor de Antígeno de Linfócitos T/genética , Linfócitos T Citotóxicos/imunologia , Visão Ocular , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The generation of allergen-specific TCR transgenic animals allows for the characterization of allergen-specific T-cell responses in vivo and in vitro and is a powerful tool to study adaptive immunity to allergens. Here we describe an approach starting from the isolation of antigen-specific T-cell hybridomas and using PCR, flow cytometric, and co-culture methods to obtain antigen-specific MHC class II-restricted CD4+ TCR transgenic mice on the Rag2-/- background.
Assuntos
Alérgenos/imunologia , Receptores de Antígenos de Linfócitos T/metabolismo , Especificidade do Receptor de Antígeno de Linfócitos T/imunologia , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Animais , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Comunicação Celular/imunologia , Linhagem Celular , Clonagem Molecular , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Expressão Gênica , Ordem dos Genes , Vetores Genéticos/genética , Linfonodos/inervação , Linfonodos/metabolismo , Camundongos , Camundongos Transgênicos , Receptores de Antígenos de Linfócitos T/genética , Especificidade do Receptor de Antígeno de Linfócitos T/genéticaRESUMO
The unique capabilities of gamma-delta (γδ) T cells to recognize cells under stressed conditions, particularly infected or transformed cells, and killing them or regulating the immune response against them, paved the way to the development of promising therapeutic strategies for cancer and infectious diseases. From a mechanistic standpoint, numerous studies have unveiled a remarkable flexibility of γδ T cells in employing their T cell receptor and/or NK cell receptors for target cell recognition, even if the relevant ligands often remain uncertain. Here, we review the accumulated knowledge on the diverse mechanisms of target cell recognition by γδ T cells, focusing on human γδ T cells, to provide an integrated perspective of their therapeutic potential in cancer and infectious diseases.
Assuntos
Receptores de Antígenos de Linfócitos T gama-delta/metabolismo , Especificidade do Receptor de Antígeno de Linfócitos T/genética , Especificidade do Receptor de Antígeno de Linfócitos T/imunologia , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Animais , Antígenos de Neoplasias/imunologia , Antígenos de Neoplasias/metabolismo , Comunicação Celular , Doenças Transmissíveis/etiologia , Doenças Transmissíveis/metabolismo , Epitopos de Linfócito T/imunologia , Humanos , Ligantes , Neoplasias/genética , Neoplasias/imunologia , Neoplasias/metabolismo , Ligação Proteica/imunologiaRESUMO
During infections and cancer, the composition of the T-cell receptor (TCR) repertoire of antigen-specific CD8+ T cells changes over time. TCR avidity is thought to be a major driver of this process, thereby interacting with several additional regulators of T-cell responses to form a composite immune response architecture. Infections with latent viruses, such as cytomegalovirus (CMV), can lead to large T-cell responses characterized by an oligoclonal TCR repertoire. Here, we review the current status of experimental studies and theoretical models of TCR repertoire evolution during CMV infection. We will particularly discuss the degree to which this process may be determined through structural TCR avidity. As engineered TCR-redirected T cells have moved into the spotlight for providing more effective immunotherapies, it is essential to understand how the key features of a given TCR influence T-cell expansion and maintenance in settings of infection or malignancy. Deeper insights into these mechanisms will improve our basic understanding of T-cell immunology and help to identify optimal TCRs for immunotherapy.
Assuntos
Infecções por Citomegalovirus/genética , Infecções por Citomegalovirus/imunologia , Citomegalovirus/imunologia , Receptores de Antígenos de Linfócitos T/genética , Especificidade do Receptor de Antígeno de Linfócitos T/imunologia , Linfócitos T/imunologia , Linfócitos T/metabolismo , Animais , Infecções por Citomegalovirus/metabolismo , Epitopos de Linfócito T/imunologia , Variação Genética , Humanos , Imunoterapia , Ativação Linfocitária/imunologia , Receptores de Antígenos de Linfócitos T/metabolismo , Especificidade do Receptor de Antígeno de Linfócitos T/genética , Vacinas/imunologiaRESUMO
The adoptive transfer of neoantigen-reactive tumor-infiltrating lymphocytes (TILs) can result in tumor regression in patients with metastatic cancer. To improve the efficacy of adoptive T cell therapy targeting these tumor-specific mutations, we have proposed a new therapeutic strategy, which involves the genetic modification of autologous T cells with neoantigen-specific T cell receptors (TCRs) and the transfer of these modified T cells back to cancer patients. However, the current techniques to isolate neoantigen-specific TCRs are labor intensive, time consuming, and technically challenging, not suitable for clinical applications. To facilitate this process, a new approach was developed, which included the co-culture of TILs with tandem minigene (TMG)-transfected or peptide-pulsed autologous antigen-presenting cells (APCs) and the single-cell RNA sequencing (RNA-seq) analysis of T cells to identify paired TCR sequences associated with cells expressing high levels of interferon-γ (IFN-γ) and interleukin-2 (IL-2). Following this new approach, multiple TCRs were identified, synthesized, cloned into a retroviral vector, and then transduced into donor T cells. These transduced T cells were shown to specifically recognize the neoantigens presented by autologous APCs. In conclusion, this approach provides an efficient procedure to isolate neoantigen-specific TCRs for clinical applications, as well as for basic and translational research.
Assuntos
Antígenos de Neoplasias/imunologia , Sequenciamento de Nucleotídeos em Larga Escala , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T/metabolismo , Análise de Célula Única , Linfócitos T/imunologia , Linfócitos T/metabolismo , Linhagem Celular Tumoral , Humanos , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/metabolismo , Mutação , Proteínas Proto-Oncogênicas p21(ras)/genética , Proteínas Proto-Oncogênicas p21(ras)/imunologia , Especificidade do Receptor de Antígeno de Linfócitos T/genética , Especificidade do Receptor de Antígeno de Linfócitos T/imunologiaRESUMO
Progress in our understanding of basic immunology along with the advent of bioengineering technologies have made possible the production of human T-cells expressing Chimeric Antigen Receptors (CAR T-cells). These CAR T-cells are designed to target specific antigens presented by cancer cells. Once CARs are bound to these antigens, CAR T-cells get activated and can initiate potent anti-tumor effects. We will here overview the bioengineering advances which made possible the clinical application of CAR T-cell therapy. We will review the data to date regarding anti-CD19 CAR T-cell therapy for acute lymphoblastic leukemia, non-Hodgkin lymphomas, and chronic lymphocytic leukemia. Besides CD19, CAR T-cells directed against the B-cell maturation antigen have also shown encouraging results to treat patients with refractory multiple myeloma. The more limited body of clinical research in the field of solid tumors will also be reviewed. Moreover, we will elaborate on the main toxicities of limitations of CAR T-cell therapy, namely cytokine release syndrome and neurotoxicity. While enjoying an undeniable hype, CAR T-cell therapy bears significant limitations. We will conclude by exposing the possible approaches to make CAR T-cells safer and more efficient beyond the CD19 target.
Assuntos
Imunoterapia Adotiva/métodos , Neoplasias/terapia , Receptores de Antígenos de Linfócitos T , Receptores de Antígenos Quiméricos , Linfócitos T/transplante , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/imunologia , Antígenos de Neoplasias/metabolismo , Humanos , Imunoterapia Adotiva/efeitos adversos , Neoplasias/epidemiologia , Neoplasias/imunologia , Neoplasias/patologia , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T/imunologia , Receptores de Antígenos de Linfócitos T/metabolismo , Receptores de Antígenos Quiméricos/genética , Receptores de Antígenos Quiméricos/imunologia , Receptores de Antígenos Quiméricos/metabolismo , Especificidade do Receptor de Antígeno de Linfócitos T/genética , Linfócitos T/imunologia , Linfócitos T/metabolismo , Resultado do TratamentoRESUMO
Measles is an acute viral disease associated both with immune suppression and development of life-long immunity. Clearance of measles virus (MeV) involves rapid elimination of infectious virus during the rash followed by slow elimination of viral RNA. To characterize cellular immune responses during recovery, we analyzed the appearance, specificity and function of MeV-specific T cells for 6 months after respiratory infection of rhesus macaques with wild type MeV. IFN-γ and IL-17-producing cells specific for the hemagglutinin and nucleocapsid proteins appeared in circulation in multiple waves approximately 2-3, 8 and 18-24 weeks after infection. IFN-γ-secreting cells were most abundant early and IL-17-secreting cells late. Both CD4+ and CD8+ T cells were sources of IFN-γ and IL-17, and IL-17-producing cells expressed RORγt. Therefore, the cellular immune response evolves during MeV clearance to produce functionally distinct subsets of MeV-specific CD4+ and CD8+ T cells at different times after infection.