Whole blood flow cytometric measurement of NFATc1 and IL-2 expression to analyze cyclosporine A-mediated effects in T cells.

The calcineurin inhibitor Cyclosporine A (CsA) is one of the crucial immunosuppressive drugs given after organ transplantation The small therapeutic window of CsA generates the dilemma that efficient and toxic chug doses differ only slightly Moreover, these threshold concentrations differ considerably between individuals; therefore, functional assays are urgently needed We explored whether the transcription factor NFATc1, a direct as well as indirect target of CsA, can be used as a potential biomarker to determine the individual immunosuppressive activity of CsA. First, in isolated human T cells we showed that flow cytometry is practicable to measure NFATc1, the most abundant NFATc isoform in activated T cells. Second, for whole blood we developed a flow cytometric assay to determine in parallel the inducible transcription factor NFATc1 and the cytokine IL-2 in stimulated T cells. We found that added CsA inhibits both the expression of NFATc1 and IL-2 in T cells of stimulated whole blood samples with IC50 values of 200 and 150 nM, respectively The intra- and inter-assay variability was low, and clinical practicability was good. Further experiments have to demonstrate whether the parallel cytometric measurement of NFATc1 and IL-2 in whole blood is a good predictor of individual CsA efficacy and toxicity in CsA-treated patients (C) 2010 International Society for Advancement of Cytometry