RESUMO
The chimeric antigen receptor (CAR) directs T cells to target and kill specific cancer cells. Despite the success of CAR T therapy in clinics, the intracellular signaling pathways that lead to CAR T cell activation remain unclear. Using CD19 CAR as a model, we report that, similar to the endogenous T cell receptor (TCR), antigen engagement triggers the formation of CAR microclusters that transduce downstream signaling. However, CAR microclusters do not coalesce into a stable central supramolecular activation cluster (cSMAC). Moreover, LAT, an essential scaffold protein for TCR signaling, is not required for microcluster formation, immunological synapse formation, nor actin remodeling following CAR activation. However, CAR T cells still require LAT for an optimal production of the cytokine IL-2. Together, these data show that CAR T cells can bypass LAT for a subset of downstream signaling outputs, thus revealing a rewired signaling pathway as compared to native T cells.
Assuntos
Sinapses Imunológicas/imunologia , Interleucina-2/imunologia , Receptores de Antígenos Quiméricos/imunologia , Transdução de Sinais/imunologia , Linfócitos T/imunologia , Células HEK293 , Humanos , Sinapses Imunológicas/genética , Interleucina-2/genética , Células Jurkat , Receptores de Antígenos Quiméricos/genética , Transdução de Sinais/genéticaRESUMO
BRCA1/2-mutated ovarian cancers (OCs) are defective in homologous recombination repair (HRR) of double-strand breaks (DSBs) and thereby sensitive to platinum and PARP inhibitors (PARPis). Multiple PARPis have recently received US Food and Drug Administration (FDA) approval for treatment of OCs, and resistance to PARPis is a major clinical problem. Utilizing primary and recurrent BRCA1/2-mutated carcinomas from OC patients, patient-derived lines, and an in vivo BRCA2-mutated mouse model, we identified a microRNA, miR-493-5p, that induced platinum/PARPi resistance exclusively in BRCA2-mutated carcinomas. However, in contrast to the most prevalent resistance mechanisms in BRCA mutant carcinomas, miR-493-5p did not restore HRR. Expression of miR-493-5p in BRCA2-mutated/depleted cells reduced levels of nucleases and other factors involved in maintaining genomic stability. This resulted in relatively stable replication forks, diminished single-strand annealing of DSBs, and increased R-loop formation. We conclude that impact of miR-493-5p on multiple pathways pertinent to genome stability cumulatively causes PARPi/platinum resistance in BRCA2 mutant carcinomas.
