RESUMO
Sepsis induces immune alterations, which last for months after the resolution of illness. The effect of this immunological reprogramming on the risk of developing cancer is unclear. Here we use a national claims database to show that sepsis survivors had a lower cumulative incidence of cancers than matched nonsevere infection survivors. We identify a chemokine network released from sepsis-trained resident macrophages that triggers tissue residency of T cells via CCR2 and CXCR6 stimulations as the immune mechanism responsible for this decreased risk of de novo tumor development after sepsis cure. While nonseptic inflammation does not provoke this network, laminarin injection could therapeutically reproduce the protective sepsis effect. This chemokine network and CXCR6 tissue-resident T cell accumulation were detected in humans with sepsis and were associated with prolonged survival in humans with cancer. These findings identify a therapeutically relevant antitumor consequence of sepsis-induced trained immunity.
Assuntos
Macrófagos , Neoplasias , Sepse , Humanos , Sepse/imunologia , Macrófagos/imunologia , Feminino , Neoplasias/imunologia , Neoplasias/terapia , Masculino , Receptores CXCR6/metabolismo , Animais , Linfócitos T/imunologia , Receptores CCR2/metabolismo , Pessoa de Meia-Idade , Camundongos , Idoso , Quimiocinas/metabolismo , AdultoRESUMO
The emerging tumor-on-chip (ToC) approaches allow to address biomedical questions out of reach with classical cell culture techniques: in biomimetic 3D hydrogels they partially reconstitute ex vivo the complexity of the tumor microenvironment and the cellular dynamics involving multiple cell types (cancer cells, immune cells, fibroblasts, etc.). However, a clear bottleneck is the extraction and interpretation of the rich biological information contained, sometime hidden, in the cell co-culture videos. In this work, we develop and apply novel video analysis algorithms to automatically measure the cytotoxic effects on human cancer cells (lung and breast) induced either by doxorubicin chemotherapy drug or by autologous tumor-infiltrating cytotoxic T lymphocytes (CTL). A live fluorescent dye (red) is used to selectively pre-stain the cancer cells before co-cultures and a live fluorescent reporter for caspase activity (green) is used to monitor apoptotic cell death. The here described open-source computational method, named STAMP (spatiotemporal apoptosis mapper), extracts the temporal kinetics and the spatial maps of cancer death, by localizing and tracking cancer cells in the red channel, and by counting the red to green transition signals, over 2-3 days. The robustness and versatility of the method is demonstrated by its application to different cell models and co-culture combinations. Noteworthy, this approach reveals the strong contribution of primary cancer-associated fibroblasts (CAFs) to breast cancer chemo-resistance, proving to be a powerful strategy to investigate intercellular cross-talks and drug resistance mechanisms. Moreover, we defined a new parameter, the 'potential of death induction', which is computed in time and in space to quantify the impact of dying cells on neighbor cells. We found that, contrary to natural death, cancer death induced by chemotherapy or by CTL is transmissible, in that it promotes the death of nearby cancer cells, suggesting the release of diffusible factors which amplify the initial cytotoxic stimulus.
Assuntos
Apoptose/fisiologia , Técnicas de Cocultura/métodos , Linfócitos T Citotóxicos , Microambiente Tumoral/fisiologia , Linhagem Celular Tumoral , Biologia Computacional , Fibroblastos/citologia , Fibroblastos/fisiologia , Humanos , Cinética , Técnicas Analíticas Microfluídicas , Microscopia de Vídeo , Linfócitos T Citotóxicos/citologia , Linfócitos T Citotóxicos/fisiologiaRESUMO
Hypoxia upregulates the core pluripotency factors NANOG, SOX2, and OCT4, associated with tumor aggressiveness and resistance to conventional anticancer treatments. We have previously reported that hypoxia-induced NANOG contributed in vitro to tumor cell resistance to autologous-specific CTL and in vivo to the in situ recruitment of immune-suppressive cells. In this study, we investigated the mechanisms underlying NANOG-mediated tumor cell resistance to specific lysis under hypoxia. We demonstrated the tumor-promoting effect of hypoxia on tumor initiation into immunodeficient mice using human non-small lung carcinoma cells. We next showed a link between NANOG and autophagy activation under hypoxia because inhibition of NANOG decreased autophagy in tumor cells. Chromatin immunoprecipitation and luciferase reporter assays revealed a direct binding of NANOG to a transcriptionally active site in a BNIP3L enhancer sequence. These data establish a new link between the pluripotency factor NANOG and autophagy involved in resistance to CTL under hypoxia.
Assuntos
Autofagia , Hipóxia Celular , Elementos Facilitadores Genéticos , Proteínas de Membrana/genética , Proteínas Mitocondriais/genética , Proteína Homeobox Nanog/metabolismo , Regiões Promotoras Genéticas , Animais , Linhagem Celular Tumoral , Imunoprecipitação da Cromatina , Humanos , Proteínas de Membrana/antagonistas & inibidores , Proteínas de Membrana/metabolismo , Camundongos , Proteínas Mitocondriais/antagonistas & inibidores , Proteínas Mitocondriais/metabolismo , Interferência de RNA , Regulação para CimaRESUMO
We had previously demonstrated the role of CD103 integrin on lung tumor-infiltrating lymphocyte (TIL) clones in promoting specific TCR-mediated epithelial tumor cell cytotoxicity. However, the contribution of CD103 on intratumoral T cell distribution and functions and the prognosis significance of TIL subpopulations in non-small cell lung carcinoma (NSCLC) have thus far not been systematically addressed. In this study, we show that an enhanced CD103(+) TIL subset correlates with improved early stage NSCLC patient survival and increased intraepithelial lymphocyte infiltration. Moreover, our results indicate that CD8(+)CD103(+) TIL, freshly isolated from NSCLC specimens, display transcriptomic and phenotypic signatures characteristic of tissue-resident memory T cells and frequently express PD-1 and Tim-3 checkpoint receptors. This TIL subset also displays increased activation-induced cell death and mediates specific cytolytic activity toward autologous tumor cells upon blockade of the PD-1-PD-L1 interaction. These findings emphasize the role of CD8(+)CD103(+) tissue-resident memory T cells in promoting intratumoral CTL responses and support the rationale for using anti-PD-1 blocking Ab to reverse tumor-induced T cell exhaustion in NSCLC patients.
Assuntos
Memória Imunológica , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/mortalidade , Linfócitos do Interstício Tumoral/imunologia , Idoso , Idoso de 80 Anos ou mais , Antígenos CD/metabolismo , Antígenos CD8/metabolismo , Carcinoma Pulmonar de Células não Pequenas/imunologia , Carcinoma Pulmonar de Células não Pequenas/mortalidade , Carcinoma Pulmonar de Células não Pequenas/patologia , Citotoxicidade Imunológica , Feminino , Perfilação da Expressão Gênica , Receptor Celular 2 do Vírus da Hepatite A , Humanos , Imunofenotipagem , Cadeias alfa de Integrinas/metabolismo , Neoplasias Pulmonares/patologia , Ativação Linfocitária/imunologia , Linfócitos do Interstício Tumoral/metabolismo , Masculino , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Especificidade de Órgãos/imunologia , Prognóstico , Receptor de Morte Celular Programada 1/genética , Receptor de Morte Celular Programada 1/metabolismo , Fatores de RiscoRESUMO
Granzyme B (GzmB) plays a major role in CTLs and NK cell-mediated elimination of virus-infected cells and tumors. Human GzmB preferentially induces target cell apoptosis by cleaving the proapoptotic Bcl-2 family member Bid, which, together with Bax, induces mitochondrial outer membrane permeabilization. We previously showed that GzmB also induces a rapid accumulation of the tumor-suppressor protein p53 within target cells, which seems to be involved in GzmB-induced apoptosis. In this article, we show that GzmB-activated p53 accumulates on target cell mitochondria and interacts with Bcl-2. This interaction prevents Bcl-2 inhibitory effect on both Bax and GzmB-truncated Bid, and promotes GzmB-induced mitochondrial outer membrane permeabilization. Consequently, blocking p53-Bcl-2 interaction decreases GzmB-induced Bax activation, cytochrome c release from mitochondria, and subsequent effector caspases activation leading to a decreased sensitivity of target cells to both GzmB and CTL/NK-mediated cell death. Together, our results define p53 as a new important player in the GzmB apoptotic signaling pathway and in CTL/NK-induced apoptosis.
Assuntos
Apoptose/imunologia , Proteína Agonista de Morte Celular de Domínio Interatuante com BH3/metabolismo , Granzimas/metabolismo , Linfócitos T Citotóxicos/imunologia , Proteína Supressora de Tumor p53/metabolismo , Proteína Agonista de Morte Celular de Domínio Interatuante com BH3/genética , Benzotiazóis/farmacologia , Caspase 3/metabolismo , Linhagem Celular Tumoral , Citocromos c/metabolismo , Ativação Enzimática , Granzimas/antagonistas & inibidores , Granzimas/farmacologia , Humanos , Células Matadoras Naturais/imunologia , Células MCF-7 , Mitocôndrias/imunologia , Membranas Mitocondriais/metabolismo , Interferência de RNA , RNA Interferente Pequeno , Tolueno/análogos & derivados , Tolueno/farmacologia , Proteína Supressora de Tumor p53/genética , Proteína X Associada a bcl-2/metabolismoRESUMO
The interaction of integrin αE(CD103)ß7, often expressed on tumor-infiltrating T lymphocytes, with its cognate ligand, the epithelial cell marker E-cadherin on tumor cells, plays a major role in antitumor CTL responses. CD103 is induced on CD8 T cells upon TCR engagement and exposure to TGF-ß1, abundant within the tumor microenvironment. However, the transcriptional mechanisms underlying the cooperative role of these two signaling pathways in inducing CD103 expression in CD8 T lymphocytes remain unknown. Using a human CTL system model based on a CD8(+)/CD103(-) T cell clone specific of a lung tumor-associated Ag, we demonstrated that the transcription factors Smad2/3 and NFAT-1 are two critical regulators of this process. We also identified promoter and enhancer elements of the human ITGAE gene, encoding CD103, involved in its induction by these transcriptional regulators. Overall, our results explain how TGF-ß1 can participate in CD103 expression on locally TCR-engaged Ag-specific CD8 T cells, thus contributing to antitumor CTL responses and cancer cell destruction.
Assuntos
Antígenos CD/imunologia , Antígenos de Neoplasias/imunologia , Linfócitos T CD8-Positivos/imunologia , Cadeias alfa de Integrinas/imunologia , Neoplasias Pulmonares/imunologia , Fatores de Transcrição NFATC/imunologia , Proteína Smad2/imunologia , Proteína Smad3/imunologia , Antígenos CD/biossíntese , Antígenos CD/genética , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Linfócitos T CD8-Positivos/patologia , Células HEK293 , Humanos , Cadeias alfa de Integrinas/biossíntese , Cadeias alfa de Integrinas/genética , Células Jurkat , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Fatores de Transcrição NFATC/genética , Fatores de Transcrição NFATC/metabolismo , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T/imunologia , Receptores de Antígenos de Linfócitos T/metabolismo , Elementos de Resposta/genética , Elementos de Resposta/imunologia , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Proteína Smad2/genética , Proteína Smad2/metabolismo , Proteína Smad3/genética , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta1/genética , Fator de Crescimento Transformador beta1/imunologia , Fator de Crescimento Transformador beta1/metabolismoRESUMO
The lytic function of CTL relies on the polarized release of cytotoxic granules (CG) at the immune synapse (IS) with target cells. CTL also contain CCL5 in cytoplasmic storage vesicles (CCL5V) distinct from CG, the role of which, in regulating T cell effector functions, is not understood. Using human CD8(+) T cells specific to a lung tumor-associated Ag, we show in this article that CTL release both secretory compartments into the immune synapse with autologous tumor cells. Moreover, we demonstrate that disorganization of the T cell microtubule cytoskeleton and defects in hMunc13-4 or Rab27a abrogate CG exocytosis and synaptic secretion of the chemokine. Mechanistically, synaptic release of CCL5 cytoplasmic storage vesicles likely occurs upon their coalescence with the Rab27a-hMunc13-4 compartment and results in autocrine, CCR5-dependent induction of CXCR4 cell surface expression, thereby promoting T cell migration in response to CXCL12. We propose that CCL5 polarized delivery represents a mechanism by which CTL control immune synapse duration.
Assuntos
Antígenos de Neoplasias/imunologia , Quimiocina CCL5/imunologia , Quimiocina CXCL12/imunologia , Citotoxicidade Imunológica , Receptores CXCR4/imunologia , Linfócitos T Citotóxicos/imunologia , Antígenos de Neoplasias/genética , Linhagem Celular Tumoral , Quimiocina CCL5/genética , Quimiocina CCL5/metabolismo , Quimiocina CXCL12/genética , Quimiotaxia , Grânulos Citoplasmáticos/química , Grânulos Citoplasmáticos/imunologia , Exocitose/imunologia , Regulação da Expressão Gênica , Humanos , Sinapses Imunológicas , Microtúbulos/imunologia , Proteínas do Tecido Nervoso/deficiência , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/imunologia , Receptores CXCR4/genética , Transdução de Sinais , Linfócitos T Citotóxicos/patologia , Proteínas rab de Ligação ao GTP/deficiência , Proteínas rab de Ligação ao GTP/genética , Proteínas rab de Ligação ao GTP/imunologia , Proteínas rab27 de Ligação ao GTPRESUMO
There is a compelling need for approaches to predict the efficacy of immunotherapy drugs. Tumor-on-chip technology exploits microfluidics to generate 3D cell co-cultures embedded in hydrogels that recapitulate simplified tumor ecosystems. Here, we present the development and validation of lung tumor-on-chip platforms to quickly and precisely measure ex vivo the effects of immune checkpoint inhibitors on T cell-mediated cancer cell death by exploiting the power of live imaging and advanced image analysis algorithms. The integration of autologous immunosuppressive FAP+ cancer-associated fibroblasts impaired the response to anti-PD-1, indicating that tumors-on-chips are capable of recapitulating stroma-dependent mechanisms of immunotherapy resistance. For a small cohort of non-small cell lung cancer patients, we generated personalized tumors-on-chips with their autologous primary cells isolated from fresh tumor samples, and we measured the responses to anti-PD-1 treatment. These results support the power of tumor-on-chip technology in immuno-oncology research and open a path to future clinical validations.
Assuntos
Inibidores de Checkpoint Imunológico , Neoplasias Pulmonares , Medicina de Precisão , Receptor de Morte Celular Programada 1 , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/imunologia , Medicina de Precisão/métodos , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptor de Morte Celular Programada 1/metabolismo , Receptor de Morte Celular Programada 1/imunologia , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/uso terapêutico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/patologia , Carcinoma Pulmonar de Células não Pequenas/imunologia , Dispositivos Lab-On-A-Chip , Imunoterapia/métodos , Microambiente Tumoral/efeitos dos fármacos , Microambiente Tumoral/imunologia , Linhagem Celular TumoralRESUMO
Various T cell adhesion molecules and their cognate receptors on target cells promote T cell receptor (TCR)-mediated cell killing. In this report, we demonstrate that the interaction of epithelial cell marker E-cadherin with integrin alpha(E)(CD103)beta(7), often expressed by tumor-infiltrating lymphocytes (TILs), plays a major role in effective tumor cell lysis. Indeed, we found that although tumor-specific CD103(+) TIL-derived cytotoxic T lymphocyte (CTL) clones are able to kill E-cadherin(+)/intercellular adhesion molecule 1(-) autologous tumor cells, CD103(-) peripheral blood lymphocyte (PBL)-derived counterparts are inefficient. This cell killing is abrogated after treatment of the TIL clones with a blocking anti-CD103 monoclonal antibody or after targeting E-cadherin in the tumor using ribonucleic acid interference. Confocal microscopy analysis also demonstrated that alpha(E)beta(7) is recruited at the immunological synapse and that its interaction with E-cadherin is required for cytolytic granule polarization and subsequent exocytosis. Moreover, we report that the CD103(-) profile, frequently observed in PBL-derived CTL clones and associated with poor cytotoxicity against the cognate tumor, is up-regulated upon TCR engagement and transforming growth factor beta1 treatment, resulting in strong potentiation of antitumor lytic function. Thus, CD8(+)/CD103(+) tumor-reactive T lymphocytes infiltrating epithelial tumors most likely play a major role in antitumor cytotoxic response through alpha(E)beta(7)-E-cadherin interactions.
Assuntos
Caderinas/fisiologia , Polaridade Celular/imunologia , Grânulos Citoplasmáticos/imunologia , Exocitose/imunologia , Integrinas/fisiologia , Neoplasias Pulmonares/imunologia , Linfócitos T Citotóxicos/imunologia , Caderinas/metabolismo , Carcinoma de Células Grandes/imunologia , Carcinoma de Células Grandes/prevenção & controle , Linhagem Celular Tumoral , Grânulos Citoplasmáticos/metabolismo , Humanos , Integrinas/metabolismo , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/prevenção & controle , Linfócitos T Citotóxicos/metabolismoRESUMO
The CD5 coreceptor is expressed on all T cells and on the B1a B cell subset. It is associated with TCR and BCR, and modulates intracellular signals initiated by both Ag receptor complexes. Human CD5 contributes to regulation of the antitumor immune response and susceptibility of specific CTL to activation-induced cell death (AICD) triggered by the tumor. In this study, we compared the T cell response to the B16F10 melanoma engrafted into CD5-deficient and wild-type C57BL/6 mice. Compared with wild-type mice, CD5 knockout animals displayed delayed tumor growth, associated with tumor infiltration by T cell populations exhibiting a more activated phenotype and enhanced antitumor effector functions. However, control of tumor progression in CD5(-/-) mice was transient due to increased AICD of CD8(+) tumor-infiltrating T lymphocytes. Remarkably, in vivo protection of T cells from TCR-mediated apoptosis by an adenovirus engineered to produce soluble Fas resulted in a dramatic reduction in tumor growth. Our data suggest that recruitment of tumor-specific T cells in the tumor microenvironment occurs at early stages of cancer development and that tumor-mediated AICD of tumor-infiltrating T lymphocytes is most likely involved in tumor escape from the immune system.
Assuntos
Antígenos CD5/genética , Ativação Linfocitária/imunologia , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/patologia , Melanoma Experimental/prevenção & controle , Linfócitos T Citotóxicos/imunologia , Linfócitos T Citotóxicos/patologia , Regulação para Cima/imunologia , Animais , Antígenos CD5/metabolismo , Carcinoma Pulmonar de Lewis/genética , Carcinoma Pulmonar de Lewis/patologia , Carcinoma Pulmonar de Lewis/prevenção & controle , Morte Celular/genética , Morte Celular/imunologia , Linhagem Celular Tumoral , Citotoxicidade Imunológica/genética , Humanos , Tolerância Imunológica/genética , Ativação Linfocitária/genética , Melanoma Experimental/genética , Melanoma Experimental/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores de Antígenos de Linfócitos T/biossíntese , Receptores de Antígenos de Linfócitos T/genética , Linfócitos T Citotóxicos/metabolismo , Regulação para Cima/genéticaRESUMO
Decreased antigenicity of cancer cells is a major problem in tumor immunology. This is often acquired by an expression defect in the TAP. However, it has been reported that certain murine Ags appear on the target cell surface upon impairment of TAP expression. In this study, we identified a human CTL epitope belonging to this Ag category. This epitope is derived from preprocalcitonin (ppCT) signal peptide and is generated within the endoplasmic reticulum by signal peptidase and signal peptide peptidase. Lung cancer cells bearing this antigenic peptide displayed low levels of TAP, but restoration of their expression by IFN-γ treatment or TAP1 and TAP2 gene transfer abrogated ppCT Ag presentation. In contrast, TAP upregulation in the same tumor cells increased their recognition by proteasome/TAP-dependent peptide-specific CTLs. Thus, to our knowledge, ppCT(16-25) is the first human tumor epitope whose surface expression requires loss or downregulation of TAP. Lung tumors frequently display low levels of TAP molecules and might thus be ignored by the immune system. Our results suggest that emerging signal peptidase-generated peptides represent alternative T cell targets, which permit CTLs to destroy TAP-impaired tumors and thus overcome tumor escape from CD8(+) T cell immunity.
Assuntos
Transportadores de Cassetes de Ligação de ATP/biossíntese , Apresentação de Antígeno/imunologia , Antígenos de Neoplasias/imunologia , Linfócitos T Citotóxicos/imunologia , Evasão Tumoral/imunologia , Transportadores de Cassetes de Ligação de ATP/genética , Transportadores de Cassetes de Ligação de ATP/imunologia , Apresentação de Antígeno/genética , Western Blotting , Linhagem Celular Tumoral , Epitopos de Linfócito T/imunologia , Imunofluorescência , Perfilação da Expressão Gênica , Humanos , Neoplasias Pulmonares/imunologia , Interferência de RNA , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Evasão Tumoral/genéticaRESUMO
Hypoxia is a major feature of the solid tumor microenvironment and is known to be associated with tumor progression and poor clinical outcome. Recently, we reported that hypoxia protects human non-small cell lung tumor cells from specific lysis by stabilizing hypoxia-inducible factor-1α and inducing STAT3 phosphorylation. In this study, we show that NANOG, a transcription factor associated with stem cell self renewal, is a new mediator of hypoxia-induced resistance to specific lysis. Our data indicate that under hypoxic conditions, NANOG is induced at both transcriptional and translational levels. Knockdown of the NANOG gene in hypoxic tumor cells is able to significantly attenuate hypoxia-induced tumor resistance to CTL-dependent killing. Such knockdown correlates with an increase of target cell death and an inhibition of hypoxia-induced delay of DNA replication in these cells. Interestingly, NANOG depletion results in inhibition of STAT3 phosphorylation and nuclear translocation. To our knowledge, this study is the first to show that hypoxia-induced NANOG plays a critical role in tumor cell response to hypoxia and promotes tumor cell resistance to Ag-specific lysis.
Assuntos
Hipóxia Celular/imunologia , Proteínas de Homeodomínio/biossíntese , Neoplasias/imunologia , Linfócitos T Citotóxicos/imunologia , Western Blotting , Hipóxia Celular/genética , Linhagem Celular Tumoral , Separação Celular , Citometria de Fluxo , Expressão Gênica , Regulação da Expressão Gênica/imunologia , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/imunologia , Humanos , Microscopia Confocal , Proteína Homeobox Nanog , Neoplasias/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Linfócitos T Citotóxicos/metabolismo , Microambiente Tumoral/genética , Microambiente Tumoral/imunologiaRESUMO
Tumor-infiltrating lymphocytes (TIL) often include a substantial subset of CD8+ tissue-resident memory T (TRM) cells enriched in tumor-specific T cells. These TRM cells play a major role in antitumor immune response. They are identified on the basis of their expression of the CD103 (αE(CD103)ß7) and/or CD49a (α1(CD49a)ß1) integrins, and the C-type lectin CD69, which are involved in tissue residency. TRM cells express several T-cell inhibitory receptors on their surface but they nevertheless react strongly to malignant cells, exerting a strong cytotoxic function, particularly in the context of blocking interactions of PD-1 with PD-L1 on target cells. These TRM cells form stable conjugates with autologous tumor cells and interact with dendritic cells and other T cells within the tumor microenvironment to orchestrate an optimal in situ T-cell response. There is growing evidence to indicate that TGF-ß is essential for the formation and maintenance of TRM cells in the tumor, through the induction of CD103 expression on activated CD8+ T cells, and for the regulation of TRM effector functions through bidirectional integrin signaling. CD8+ TRM cells were initially described as a prognostic marker for survival in patients with various types of cancer, including ovarian, lung and breast cancers and melanoma. More recently, these tumor-resident CD8+ T cells have been shown to be a potent predictive biomarker of the response of cancer patients to immunotherapies, including therapeutic cancer vaccines and immune checkpoint blockade. In this review, we will highlight the major characteristics of tumor TRM cell populations and the possibilities for their exploitation in the design of more effective immunotherapy strategies for cancer.
Assuntos
Linfócitos T CD8-Positivos , Neoplasias , Humanos , Integrina alfa1/metabolismo , Células T de Memória , Memória Imunológica , Neoplasias/metabolismo , Biomarcadores/metabolismo , Imunoterapia , Microambiente TumoralRESUMO
Chimeric antigen receptor (CAR) T cells have shown promising results in the treatment of B-cell malignancies. Despite the successes, challenges remain. One of them directly involves the CAR T-cell manufacturing process and especially the ex vivo activation phase. While this is required to allow infection and expansion, ex vivo activation dampens the antitumor potential of CAR T cells. Optimizing the nature of the T cells harboring the CAR is a strategy to address this obstacle and has the potential to improve CAR T-cell therapy, including for solid tumors. Here, we describe a protocol to create CAR T cells without ex vivo preactivation by inhibiting the transcription factor FOXO1 (CAR TAS cells). This approach made T cells directly permissive to lentiviral infection, allowing CAR expression, with enhanced antitumor functions. FOXO1 inhibition in primary T cells (TAS cells) correlated with acquisition of a stem cell memory phenotype, high levels of granzyme B, and increased production of TNFα. TAS cells displayed enhanced proliferative and cytotoxic capacities as well as improved migratory properties. In vivo experiments showed that CAR TAS cells were more efficient at controlling solid tumor growth than classical CAR T cells. The production of CAR TAS from patients' cells confirmed the feasibility of the protocol in clinic.
Assuntos
Imunoterapia Adotiva , Linfócitos T , Humanos , Linhagem Celular Tumoral , Imunoterapia Adotiva/métodos , Fenótipo , Proteína Forkhead Box O1/metabolismoRESUMO
Major advances in cancer treatment have emerged with the introduction of immunotherapies using blocking antibodies that target T-cell inhibitory receptors, such as programmed death-1 (PD-1) and cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4), known as immune checkpoints. However, most cancer patients do not respond to immune checkpoint blockade (ICB) therapies, suggesting the development of resistance mechanisms associated with either an insufficient number of preexisting tumor-specific T-cell precursors and/or inappropriate T-cell reactivation. To broaden clinical benefit, anti-PD-1/PD-1 ligand (PD-L1) neutralizing antibodies have been combined with therapeutic cancer vaccines based on non-mutant and/or mutant tumor antigens, to stimulate and expand tumor-specific T lymphocytes. Although these combination treatments achieve the expected goal in some patients, relapse linked to alterations in antigen presentation machinery (APM) of cancer cells often occurs leading to tumor escape from CD8 T-cell immunity. Remarkably, an alternative antigenic peptide repertoire, referred to as T-cell epitopes associated with impaired peptide processing (TEIPP), arises on these malignant cells with altered APM. TEIPP are derived from ubiquitous non-mutant self-proteins and represent a unique resource to target immune-edited tumors that have acquired resistance to cytotoxic T lymphocytes (CTLs) related to defects in transporter associated with antigen processing (TAP) and possibly also to ICB. The present review discusses tumor-associated antigens (TAAs) and mutant neoantigens and their use as targets in peptide- and RNA-based therapeutic cancer vaccines. Finally, this paper highlights TEIPP as a promising immunogenic non-mutant neoantigen candidates for active cancer immunotherapy and combination with TAA and mutant neoantigens. Combining these polyepitope cancer vaccines with ICB would broaden T-cell specificity and reinvigorate exhausted antitumor CTL, resulting in the eradication of all types of neoplastic cells, including immune-escaped subtypes.
RESUMO
BACKGROUND: Cancer stem cells (CSC) define a population of rare malignant cells endowed with 'stemness' properties, such as self-renewing, multipotency and tumorigenicity. They are responsible for tumor initiation and progression, and could be associated with resistance to immunotherapies by negatively regulating antitumor immune response and acquiring molecular features enabling escape from CD8 T-cell immunity. However, the immunological hallmarks of human lung CSC and their potential interactions with resident memory T (TRM) cells within the tumor microenvironment have not been investigated. METHODS: We generated a non-small cell lung cancer model, including CSC line and clones, and autologous CD8+CD103+ TRM and CD8+CD103- non-TRM clones, to dissect out immune properties of CSC and their susceptibility to specific T-cell-mediated cytotoxic activity. RESULTS: Unlike their parental tumor cells, lung CSC are characterized by the initiation of an epithelial-to-mesenchymal transition program defined by upregulation of the SNAIL1 transcription factor and downregulation of phosphorylated-GSK-3ß and cell surface E-cadherin. Acquisition of a CSC profile results in partial resistance to TRM-cell-mediated cytotoxicity, which correlates with decreased surface expression of the CD103 ligand E-cadherin and human leukocyte antigen-A2-neoepitope complexes. On the other hand, CSC gained expression of intercellular adhesion molecule (ICAM)-1 and thereby sensitivity to leukocyte function-associated antigen (LFA)-1-dependent non-TRM-cell-mediated killing. Cytotoxicity is inhibited by anti-ICAM-1 and anti-major histocompatibility complex class I neutralizing antibodies further emphasizing the role of LFA-1/ICAM-1 interaction in T-cell receptor-dependent lytic function. CONCLUSION: Our data support the rational design of immunotherapeutic strategies targeting CSC to optimize their responsiveness to local CD8+CD103+ TRM cells for more efficient anticancer treatments.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Linfócitos T CD8-Positivos , Caderinas/metabolismo , Glicogênio Sintase Quinase 3 beta/metabolismo , Humanos , Memória Imunológica , Pulmão , Linfócitos do Interstício Tumoral , Células-Tronco Neoplásicas , Microambiente TumoralRESUMO
Besides tumor cells, the tumor microenvironment harbors a variety of host-derived cells, such as endothelial cells, fibroblasts, innate and adaptive immune cells. It is a complex and highly dynamic environment, providing very important cues to tumor development and progression. Tumor-associated endothelial cells play a key role in this process. On the one hand, they form tumor-associated (angiogenic) vessels through sprouting from locally preexisting vessels or recruitment of bone marrow-derived endothelial progenitor cells, to provide nutritional support to the growing tumor. On the other hand, they are the interface between circulating blood cells, tumor cells and the extracellular matrix, thereby playing a central role in controlling leukocyte recruitment, tumor cell behavior and metastasis formation. Hypoxia is a critical parameter modulating the tumor microenvironment and endothelial/tumor cell interactions. Under hypoxic stress, tumor cells produce factors that promote tumor angiogenesis, tumor cell motility and metastasis. Among these factors, VEGF, a main angiogenesis modulator, can also play a critical role in the control of immune tolerance. This review discusses some aspects of the role of endothelial cells within tumor microenvironment and emphasizes their interaction with tumor cells, the extracellular matrix and with immune killer cells. We will also address the role played by circulating endothelial progenitor cells and illustrate their features and mechanism of recruitment to the tumor microenvironment and their role in tumor angiogenesis.
Assuntos
Comunicação Celular , Células Endoteliais/imunologia , Matriz Extracelular/imunologia , Células Matadoras Naturais/imunologia , Neoplasias/imunologia , Microambiente Tumoral , Animais , Humanos , Neoplasias/irrigação sanguíneaRESUMO
Cellular interactions in the tumor stroma play a major role in cancer progression but can also induce tumor rejection. To explore the role of endothelial cells in these interactions, we used an in vitro three-dimensional collagen matrix model containing a cytotoxic T lymphocyte CTL clone (M4.48), autologous tumor cells (M4T), and an endothelial cell (M4E) line that are all derived from the same tumor. We demonstrate in this study that specific killing of the endothelial cells by the CTL clone required the autologous tumor cells and involved Ag cross-presentation. The formation of gap junctions between endothelial and tumor cells is required for antigenic peptide transfer to endothelial cells that are then recognized and eliminated by CTL. Our results indicate that gap junctions facilitate an effective CTL-mediated destruction of endothelial cells from the tumor microenvironment that may contribute to the control of tumor progression.
Assuntos
Comunicação Celular/imunologia , Apresentação Cruzada/imunologia , Células Endoteliais/imunologia , Células Endoteliais/patologia , Junções Comunicantes/imunologia , Melanoma/imunologia , Melanoma/patologia , Linfócitos T Citotóxicos/imunologia , Antígenos de Neoplasias/metabolismo , Biomarcadores/metabolismo , Biomarcadores Tumorais/metabolismo , Linhagem Celular Tumoral , Células Clonais , Técnicas de Cocultura , Citosol/imunologia , Citosol/metabolismo , Citotoxicidade Imunológica/imunologia , Células Endoteliais/metabolismo , Junções Comunicantes/metabolismo , Junções Comunicantes/patologia , Humanos , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/metabolismo , Linfócitos do Interstício Tumoral/patologia , Melanoma/terapia , Linfócitos T Citotóxicos/metabolismo , Linfócitos T Citotóxicos/patologiaRESUMO
Hypoxia is an essential component of tumor microenvironment. In this study, we investigated the influence of hypoxia (1% PO(2)) on CTL-mediated tumor cell lysis. We demonstrate that exposure of target tumor cells to hypoxia has an inhibitory effect on the CTL clone (Heu171)-induced autologous target cell lysis. Such inhibition correlates with hypoxia-inducible factor-1alpha (HIF-1alpha) induction but is not associated with an alteration of CTL reactivity as revealed by granzyme B polarization or morphological change. Western blot analysis indicates that although hypoxia had no effect on p53 accumulation, it induced the phosphorylation of STAT3 in tumor cells by a mechanism at least in part involving vascular endothelial growth factor secretion. We additionally show that a simultaneous nuclear translocation of HIF-1alpha and phospho-STAT3 was observed. Interestingly, gene silencing of STAT3 by small interfering RNA resulted in HIF-1alpha inhibition and a significant restoration of target cell susceptibility to CTL-induced killing under hypoxic conditions by a mechanism involving at least in part down-regulation of AKT phosphorylation. Moreover, knockdown of HIF-1alpha resulted in the restoration of target cell lysis under hypoxic conditions. This was further supported by DNA microarray analysis where STAT3 inhibition resulted in a partly reversal of the hypoxia-induced gene expression profile. The present study demonstrates that the concomitant hypoxic induction of phospho-STAT3 and HIF-1alpha are functionally linked to the alteration of non-small cell lung carcinoma target susceptibility to CTL-mediated killing. Considering the eminent functions of STAT3 and HIF-1alpha in the tumor microenvironment, their targeting may represent novel strategies for immunotherapeutic intervention.
Assuntos
Citotoxicidade Imunológica , Subunidade alfa do Fator 1 Induzível por Hipóxia/biossíntese , Hipóxia/imunologia , Neoplasias Pulmonares/imunologia , Fator de Transcrição STAT3/biossíntese , Linfócitos T Citotóxicos/imunologia , Linhagem Celular Tumoral , Células Clonais , Regulação Neoplásica da Expressão Gênica/imunologia , Humanos , Hipóxia/metabolismo , Hipóxia/patologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/fisiologia , Imunidade Inata , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Fator de Transcrição STAT3/antagonistas & inibidores , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/fisiologia , Linfócitos T Citotóxicos/metabolismo , Linfócitos T Citotóxicos/patologiaRESUMO
We identified an antigen recognized on a human non-small-cell lung carcinoma by a cytotoxic T lymphocyte clone derived from autologous tumor-infiltrating lymphocytes. The antigenic peptide is presented by HLA-A2 and is encoded by the CALCA gene, which codes for calcitonin and for the alpha-calcitonin gene-related peptide. The peptide is derived from the carboxy-terminal region of the preprocalcitonin signal peptide and is processed independently of proteasomes and the transporter associated with antigen processing. Processing occurs within the endoplasmic reticulum of all tumoral and normal cells tested, including dendritic cells, and it involves signal peptidase and the aspartic protease, signal peptide peptidase. The CALCA gene is overexpressed in medullary thyroid carcinomas and in several lung carcinomas compared with normal tissues, leading to recognition by the T cell clone. This new epitope is, therefore, a promising candidate for cancer immunotherapy.