RESUMO
The costimulatory effects of CD27 on T lymphocyte effector function and memory formation has been confined to evaluations in mouse models, in vitro human cell culture systems, and clinical observations. Here, we tested whether CD27 costimulation actively enhances human T-cell function, expansion, and survival in vitro and in vivo. Human T cells transduced to express an antigen-specific chimeric antigen receptor (CAR-T) containing an intracellular CD3 zeta (CD3ζ) chain signaling module with the CD27 costimulatory motif in tandem exerted increased antigen-stimulated effector functions in vitro, including cytokine secretion and cytotoxicity, compared with CAR-T with CD3ζ alone. After antigen stimulation in vitro, CD27-bearing CAR-T cells also proliferated, up-regulated Bcl-X(L) protein expression, resisted apoptosis, and underwent increased numerical expansion. The greatest impact of CD27 was noted in vivo, where transferred CAR-T cells with CD27 demonstrated heightened persistence after infusion, facilitating improved regression of human cancer in a xenogeneic allograft model. This tumor regression was similar to that achieved with CD28- or 4-1BB-costimulated CARs, and heightened persistence was similar to 4-1BB but greater than CD28. Thus, CD27 costimulation enhances expansion, effector function, and survival of human CAR-T cells in vitro and augments human T-cell persistence and antitumor activity in vivo.
Assuntos
Neoplasias da Mama/terapia , Antígenos CD28/imunologia , Imunoterapia Adotiva , Mesotelioma/terapia , Neoplasias Ovarianas/terapia , Receptores de Antígenos de Linfócitos T/imunologia , Linfócitos T/imunologia , Animais , Western Blotting , Neoplasias da Mama/genética , Neoplasias da Mama/imunologia , Antígenos CD28/genética , Proliferação de Células , Citocinas/metabolismo , Feminino , Citometria de Fluxo , Imunofluorescência , Humanos , Ativação Linfocitária , Mesotelioma/genética , Mesotelioma/imunologia , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/imunologia , Receptores de Antígenos de Linfócitos T/genética , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Transforming growth factor-beta (TGF-beta) has an essential role in the generation of inducible regulatory T (iTreg) and T helper 17 (Th17) cells. However, little is known about the TGF-beta-triggered pathways that drive the early differentiation of these cell populations. Here, we report that CD4(+) T cells lacking the molecular adaptor tumor necrosis factor (TNF) receptor-associated factor 6 (TRAF6) exhibit a specific increase in Th17 differentiation in vivo and in vitro. We show that TRAF6 deficiency renders T cells more sensitive to TGF-beta-induced Smad2/3 activation and proliferation arrest. Consistent with this, in TRAF6-deficient T cells, TGF-beta more effectively down-regulates interleukin-2 (IL-2), a known inhibitor of Th17 differentiation. Remarkably, TRAF6-deficient cells generate normal numbers of Foxp3-expressing cells in iTreg differentiation conditions where exogenous IL-2 is supplied. These findings show an unexpected role for the adaptor molecule TRAF6 in Smad-mediated TGF-beta signaling and Th17 differentiation. Importantly, the data also suggest that a main function of TGF-beta in early Th17 differentiation may be the inhibition of autocrine and paracrine IL-2-mediated suppression of Th17 cell generation.
Assuntos
Diferenciação Celular/imunologia , Interleucina-2/imunologia , Transdução de Sinais/imunologia , Fator 6 Associado a Receptor de TNF/imunologia , Fator de Crescimento Transformador beta/imunologia , Animais , Comunicação Autócrina/genética , Comunicação Autócrina/imunologia , Diferenciação Celular/genética , Proliferação de Células , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/imunologia , Fatores de Transcrição Forkhead/metabolismo , Tolerância Imunológica/genética , Tolerância Imunológica/imunologia , Interleucina-2/genética , Interleucina-2/metabolismo , Camundongos , Camundongos Mutantes , Comunicação Parácrina/genética , Comunicação Parácrina/imunologia , Transdução de Sinais/genética , Proteína Smad2/genética , Proteína Smad2/imunologia , Proteína Smad2/metabolismo , Proteína Smad3/genética , Proteína Smad3/imunologia , Proteína Smad3/metabolismo , Linfócitos T Auxiliares-Indutores , Fator 6 Associado a Receptor de TNF/genética , Fator 6 Associado a Receptor de TNF/metabolismo , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/metabolismoRESUMO
CD8 T cells, which have a crucial role in immunity to infection and cancer, are maintained in constant numbers, but on antigen stimulation undergo a developmental program characterized by distinct phases encompassing the expansion and then contraction of antigen-specific effector (T(E)) populations, followed by the persistence of long-lived memory (T(M)) cells. Although this predictable pattern of CD8 T-cell responses is well established, the underlying cellular mechanisms regulating the transition to T(M) cells remain undefined. Here we show that tumour necrosis factor (TNF) receptor-associated factor 6 (TRAF6), an adaptor protein in the TNF-receptor and interleukin-1R/Toll-like receptor superfamily, regulates CD8 T(M)-cell development after infection by modulating fatty acid metabolism. We show that mice with a T-cell-specific deletion of TRAF6 mount robust CD8 T(E)-cell responses, but have a profound defect in their ability to generate T(M) cells that is characterized by the disappearance of antigen-specific cells in the weeks after primary immunization. Microarray analyses revealed that TRAF6-deficient CD8 T cells exhibit altered expression of genes that regulate fatty acid metabolism. Consistent with this, activated CD8 T cells lacking TRAF6 display defective AMP-activated kinase activation and mitochondrial fatty acid oxidation (FAO) in response to growth factor withdrawal. Administration of the anti-diabetic drug metformin restored FAO and CD8 T(M)-cell generation in the absence of TRAF6. This treatment also increased CD8 T(M) cells in wild-type mice, and consequently was able to considerably improve the efficacy of an experimental anti-cancer vaccine.
