ζ链的纳入驱动 CD19-CAR-T 细胞中的磷酸酪氨酸信号传导

Although chimeric antigen receptor (CAR) T cell therapy has revolutionised individualised cancer therapies for relapsed/refractory lymphomas, signalling mechanisms underlying CAR T activation remain incompletely understood, especially among the three generations of CAR T exploiting different signalling domains.

Here, using Jurkat T cell as a model, we investigate how costimulation influences tyrosine phosphorylation cascades using LC-MS/MS based phosphotyrosine (pY) proteomics and CD69 expression in the presence of small molecule inhibitors of key TCR signalling regulators.

We find that including the -chain in first ( -CAR), second (28 -CAR and BB -CAR), and third (28BB -CAR) generation CARs largely determines pY signalling, irrespective of costimulation.

Further, we show that the phosphatase activity of PTPN22 and SHP-1 are largely negligible for activation of CARs, but indiscriminate inhibition of phosphatases using Pervanadate (PV) selectively activates BB -CARs without antigen encounter.

Finally, we find that selective, partial inhibition of Itk using Soquelitinib reduces basal CD69 expression in Jurkat CAR T cells while maintaining their ability to activate in response to antigen.

Our data suggest that the -chain determines the pY signalling profile of CD19-CAR Jurkat T cells and that Itk may drive antigen-independent CD19-CAR activation.