Profound Inhibition of Antigen-Specific T-Cell Effector Functions by Dasatinib

Abstract

Purpose: The dual BCR-ABL/SRC kinase inhibitor dasatinib entered the clinic for the treatment of chronic myeloid leukemia and Ph + acute lymphoblastic leukemia. Because SRC kinases are known to play an important role in physiologicT-cell activation, we analyzed the immunobiological effects of dasatinib on T-cell function. The effect of dasatinib on multiple T-cell effector functions was examined at clinically relevant doses (1-100 nmol/L); the promiscuous tyrosine kinase inhibitor staurosporine was used as a comparator. Experimental Design: Purified human CD3 + cells and virus-specific CD8 + Tcells from healthy blood donors were studied directly ex vivo ; antigen-specific effects were confirmed in defined T-cell clones. Functional outcomes included cytokine production (interleukin-2, IFNg, and tumor necrosis factor a), degranulation (CD107a/b mobilization), activation (CD69 up-regulation), proliferation (carboxyfluorescein diacetate succinimidyl ester dilution), apoptosis/necrosis induction, and signal transduction. Results: Both dasatinib and staurosporine inhibited T-cell activation, proliferation, cytokine production, and degranulation in a dose-dependent manner. Mechanistically, this was mediated by the blockade of early signal transduction events and was not due to loss of T-cell viability. Overall, CD4 + T cells seemed to be more sensitive to these effects than CD8 + Tcells, and naBve Tcells more sensitive than memoryT-cell subsets. The inhibitory effects of dasatinib were so profound that all T-cell effector functions were shut down at therapeutically relevant concentrations. Conclusion: These findings indicate that caution is warranted with use of this drug in the clinical setting and provide a rationale to explore the potential of dasatinib as an immunosuppressant in the fields of transplantation and T-cell^driven autoimmune diseases.

Clonal expansion of T/NK-cells during tyrosine kinase inhibitor dasatinib therapy

“…[9][10][11][12] To resolve the discrepancy between the in vitro and our in vivo data, we hypothesize that in a proportion of dasatinib treated patients, off-target kinase inhibition results in hyperreactivity of terminal memory CD8 þ T cells or NK cells on antigen challenge (for example, CMV) owing to activation through an alternative, non-TCR-dependent pathway (for example, IL-2R -mediated). Lower proportion of regulatory T cells in these patients may contribute to the enhanced immune reactivity.…”

“…[9][10][11][12] The putative in vivo immunomodulatory effects of dasatinib have thus far not been explored.…”

Expansion of highly differentiated CD8+ T-cells or NK-cells in patients treated with dasatinib is associated with cytomegalovirus reactivation

“…7,10 All functional assays were conducted in RPMI 1640 medium supplemented with 10% fetal calf serum, 2 mM L-glutamine, 100 U/ml penicillin and 100 mg/ml streptomycin (R10 medium). DMSO as a solvent control for dasatinib was included in all functional assays performed ex vivo.…”

“…4,5 The broader target profile of dasatinib not only implies its less susceptibility to the development of resistance, but also the potential for more potent modulation of immune cell subsets. Indeed, immunosuppressive effects on T-cells and NK-cells have been described in vitro for dasatinib [6][7][8][9] and other tyrosine kinase inhibitors. [10][11][12] This is in line with observations in single center clinical case series.…”

“…Dasatinib was used in vitro at clinically relevant doses of 10 and 50 nM, thereby mimicking peak and steady state concentrations. 7,18 In addition, polychromatic flow cytometric analyses were conducted to determine the phenotypic characteristics of T-cell subsets and NK-cells in patients on continuous dasatinib treatment with and without LGL expansions. Plasma samples were assayed for soluble immune mediators in parallel, and cytomegalovirus (CMV) load measurements were conducted as indicated on clinical grounds.…”