Rapid screening of healthy donors suggests strategies for clinical purification of virus-specific T cells

Abstract

Transfer of virus-specific memory T cells is an emerging treatment for viral diseases in immunocompromised patients. One approach is to purify antigen-specific cells based on their up-regulation of activation markers after short-term in vitro stimulation with peptides. In order to rapidly screen the frequency of antigenspecific CD4 and CD8 T cells in potential donors we have modified a commercial kit fromMiltenyi so that we can simultaneouslymeasure induction of IFN-gamma, CD40L and CD137 on both cell types following peptide stimulation. In a prestudy using blood from healthy donors we found that the presence of IFN-gamma producing cells following stimulation with EBV or CMV peptides correlated with the presence of serum antibodies against the viruses. Simultaneous stimulationwith peptides derived frommultiple proteins lead to activation of similar number of cells as stimulation with separate peptides. Whereas it was relatively easy to identify memory cells reactive to CMV and EBV, fewer donors maintained cells that could be activated by peptides derived from JC and BK viruses. IFN-gamma producing cells had increased expression of CD137, and CD4 cells also CD40L. However, more cells up-regulated their expression of CD137 than IFN-gamma following peptide stimulation and background levels of CD137 expression was higher, demonstrating that care has to be taken when selecting activation marker for purification. Taken together, this pre-study to clinical purification of virus-specific memory T cells shows that rapid screening of potential donors is possible, and suggests strategies for activation and purification of virus-specific cells.