Profound Shut-Down of Antigen Specific T-Cell Effector Functions by Dasatinib.

Dasatinib (Sprycel®, Bristol-Myers Squibb) is a dual BCR-ABL/SRC kinase inhibitor. In 2006, it was approved for the treatment of imatinib-refractory/-resistant CML and Ph+ ALL. An increased infection rate has been described in patients undergoing dasatinib treatment; we postulated that this might be due to inhibition of the SRC kinases LCK and FYN, which play an important role in T cell signalling following antigen recognition. We evaluated dasatinib’s immunobiological effects on purified human CD3+ T cells from healthy blood donors using the promiscuous tyrosine kinase inhibitor staurosporine as a comparator; we also used antigen-specific T cell clones to dissect early T cell signaling events. All assays were performed at clinically relevant doses of dasatinib (1–100nM). In CFSE dilution assays, a dose-dependent inhibition of T cell proliferation was detected with dasatinib (IC50=11nM) and staurosporine (IC50=5nM). These effects of dasatinib, but not staurosporine, were reversible; thus, T cells that were pre-incubated with dasatinib for 24h and then washed, proliferated as well as untreated T cells in the absence of dasatinib. Furthermore, we observed significant inhibition of OKT3-induced up-regulation of the activation marker CD69 (dasatinib IC50=11nM, staurosporine IC50=5nM) and reduced IL-2 production (IC50=2nM, measured by ELISA, for both dasatinib and staurosporine) in purified T cells treated with dasatinib and staurosporine. CD4+ T cells were more sensitive than CD8+ T cells to the inhibitory effects of dasatinib on activation and proliferation (activation IC50=10nM for CD4+ and 15nM for CD8+ T cells; proliferation IC50=10nM for CD4+ and 13nM for CD8+ T cells) while no differences in sensitivity were observed for staurosporine (IC50=4–5 for activation and proliferation in both CD4+ and CD8+ T cells). Since CD8+ T cell-mediated immunity is essential for long-term control of persistent DNA viruses, we evaluated dasatinib’s impact on antigen-specific alpha/beta CD8+ T cell populations specific for CMV and EBV. Profound inhibition of proliferation, cytokine secretion and degranulation was observed in all cases; these observations potentially explain the increased frequency of viral infections in dasatinib-treated patients beyond the induction of myelosuppression. Of note, gamma/delta T cell functions were also inhibited by dasatinib. Dasatinib resulted in significant upregulation of TCR/CD8 at the T cell surface of EBV specific T cell clones indicating that the drug blocks TCR downregulation and recycling at the T cell surface and therefore exerts its effects by blockade of proximal signalling pathways. Overall, these findings provide a rationale to explore the potential of dasatinib as an immunosuppressant in the settings of transplantation and T cell driven autoimmune diseases.