For sotrastaurin-treated patients the absolute number of FoxP3+CD127low Tregs remained stable: median numbers were 23, 16 and 28 cells/μl pre-, 3 and 6 months after transplantation (Fig. 4b). In neoral-treated patients, the number of FoxP3+CD127low Tregs decreased significantly at month 3 but returned to levels pretransplantation at month 6 (median learn more 12 and 18 cells/μl 3 and 6 months after transplantation, P = 0·008 for neoral 3 months versus pretransplantation (Fig. 4b). Trough levels of sotrastaurin correlated with Treg numbers: the AUC of trough levels at 0–3 months

correlated with the absolute number of FoxP3+CD127lowCD4+CD25high T regulatory cells at 3 months (n = 10, Pearson’s r = 0·65, P = 0·04). The AUC of trough levels at 0–6 months also www.selleckchem.com/products/ulixertinib-bvd-523-vrt752271.html correlated with Treg numbers at 6 months (n = 10, Pearson’s r = 0·68, P = 0·03) (Fig. 5). The functional capacity of patients’ effector and regulator T cells was tested in MLR. Of two sotrastaurin- and two neoral-treated patients, the number of isolated CD4+CD25high T cells was insufficient to determine their inhibitory capacity at each timepoint. The proliferative response of effector CD25low cells in samples of three patients was <1000 cpm. Co-culture experiments with isolated CD4+CD25high Tregs in a 1 : 10 ratio were performed (n = 4 sotrastaurin and n = 6 neoral). We analysed whether

co-culture with Tregs and time after transplantation influenced the proliferation in sotrastaurin- versus neoral-treated patients. The inhibitory capacity of Tregs in sotrastaurin-treated patients

remained intact: the median percentages of inhibition by Tregs were 82% pretransplantation, 71% at 3 months and 67% at 6 months against donor cells (months 3 and 6, P > 0·05 due to small sample size) (Fig. 6). The protein kinase C inhibitor sotrastaurin is a novel, calcineurin-independent drug in autoimmune disease, oncology and clinical organ transplantation. Currently, the effect of sotrastaurin 2-hydroxyphytanoyl-CoA lyase on cell populations that control immune responses is unknown. We therefore investigated the number and function of regulatory T cells in samples of healthy volunteers and in renal allograft recipients during sotrastaurin treatment. First, we determined the IC50 of sotrastaurin in MLR to confirm previous reported findings: Evenou et al. have shown that sotrastaurin potently inhibited alloreactivity of mouse and human T cells [6]. In a two-way MLR performed with human T cells, the IC50 of sotrastaurin to inhibit [3H]-thymidine incorporation after 6 days was 37 nM. The studies by Matz et al. also revealed dose-dependent inhibition by sotrastaurin of carboxyfluorescein succinimidyl ester (CFSE)-labelled CD4+ T cells, after allogeneic stimulation [17]. In one-way MLR with irradiated stimulator cells, we demonstrated that sotrastaurin blocked alloreactivity dose-dependently (Fig. 1). In the high concentration of 250 ng/ml, the mean percentage of inhibition was 92%. The mean IC50 of sotrastaurin in our experiments was 89 nM.