Proliferation of CD8⁺ T-Cells Specific for CMV and EBV in Response to TCR Mediated Recognition of Cognate Antigen Is Inhibited by Imatinib in a Dose-Dependent Manner.

We and others have shown that the tyrosine kinase inhibitor imatinib (STI571, Gleevec®) inhibits T-cell proliferation and activation at concentrations achieved in vivo. At 10μM, imatinib inhibited T-cell receptor (TCR)-mediated proliferation of purified peripheral blood T-cells almost completely. Up-regulation of the activation markers CD25 and CD69 at 24h in response to TCR cross-linking was suppressed by imatinib at a mean IC₅₀ of 5.4μM and 7.3μM, respectively and IL-2 production was also severely impaired. However, these assays may not fully reflect the response to clinical relevant antigens. Therefore, we chose to investigate the antigen-triggered proliferation of memory CD8⁺ T-cells specific for immunodominant CMV and EBV HLA-A2 peptide epitopes. We used HLA-peptide tetramers to identify healthy blood donors with detectable CMV- or EBV-specific CD8⁺ T-cell populations. Purified T-cells from these donors were then stimulated with the CMV peptide pp65_495–503 or the EBV peptide BMFLI_259–267. Antigen-induced proliferation was measured by dilution of the vital dye CFSE over a period of 4 or 8 days. The magnitude of the virusspecific CD8⁺ T-cell population ranged from 0.5 % to 7.1% of CD8⁺ T-cells for CMV and from 0.05% to 0.35% of CD8⁺ T-cells for EBV. Antigen-specific CD8⁺ T-cells from all 10 donors studied proliferated in response to the CMV peptide. In 8 from 10 donors, imatinib reduced CMV peptide induced proliferation. With increasing imatinib concentrations (range: 5 – 10μM), we observed dose dependent reduction of both the number of cells undergoing cell division and the average number of divisions completed per cell. Comparable inhibition of specific T-cell proliferation in response to the EBV-derived peptide was observed in two donors. Immunoblots demonstrated that imatinib substantially reduced tyrosine phosphorylation of ZAP70 and LAT in response to TCR-mediated activation in Jurkat T-cells. Sequence comparisons of all 90 tyrosine kinase genes in the human genome for homology in the ATP binding pocket identified Lck, which is required for ZAP70 activation, as a likely target for imatinib. Our results indicate that imatinib may interfere with clinically important T-cell effector functions. As concentrations sufficient for half-maximal inhibition of TCR signalling are achieved in vivo, imatinib could increase the risk of opportunistic infections and impact on GVH and GVL reactions post-transplantation especially when used in conjuction with other immunosuppressive agents. Therefore, close monitoring of patients on imatinib for CMV reactivation or EBV-induced lymphoproliferative diseases, especially in stem cell transplant recipients, appears warranted.