Comparison of the Pharmacological Efficacies of Immunosuppressive Drugs Evaluated by the ATP Production and Mitochondrial Activity in Human Lymphocytes.

The lymphocyte immunosuppressant sensitivity test (LIST) using patient peripheral lymphocytes can predict the therapeutic efficacy of immunosuppressive drugs used in renal transplantation. We have evaluated the pharmacological efficacy of drugs by using the LIST with the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay, which measures the cellular mitochondrial activity. The LIST with the MTT assay requires a relatively large amount of blood. As such, we developed a new assay for examining drug sensitivity with a CellTiter-Glo assay, which measures the amount of cellular ATP to help increase the assay's sensitivity and reduce the amount of blood needed. Renal transplant recipients generally receive either cyclosporine or tacrolimus, in addition to mycophenolate mofetil and methylprednisolone, as an immunosuppressive therapy to prevent acute rejection. We evaluated the pharmacological efficacy of these immunosuppressive agents with both the MTT and CellTiter-Glo assays using the peripheral blood mononuclear cells of 21 healthy volunteers. Furthermore, we also examined the relationship between these immunosuppressive agents' pharmacological efficacy and the results of the MTT and CellTiter-Glo assays. The IC50 values for cyclosporine, tacrolimus, mycophenolic acid, and methylprednisolone were significantly correlated between the MTT and CellTiter-Glo assays. The amount of blood cells required for LIST with the CellTiter-Glo assay was able to be reduced to 25% of the amount required for the previously established LIST with the MTT assay procedure. We concluded from these observations that the LIST with the CellTiter-Glo assay should be used instead of the MTT assay for carrying out individualized immunosuppressive therapy in renal transplantation patients.

Peripheral Lymphocyte Response to Mycophenolic Acid In Vitro and Incidence of Cytomegalovirus Infection in Renal Transplantation.

The lymphocyte immunosuppressant sensitivity test (LIST) with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay procedure has been used to predict the pharmacological efficacy of immunosuppressive agents to prevent acute rejection episodes for renal transplant recipients. In this study, mycophenolic acid (MPA) pharmacological efficacies were evaluated by LIST at both prior to and just after renal transplantation. We compared the efficacies to the clinical outcome of these recipients. MPA's pharmacological efficacy was evaluated by LIST not only before the operation but also at 2, 4, and 6 weeks after transplantation in 16 renal transplant recipients. These recipients were divided into high- and low-sensitivity groups according to peripheral blood mononuclear cell (PBMC) sensitivity to MPA in vitro. The MPA sensitivities were compared to cytomegalovirus (CMV) infection and acute rejection episodes in these recipients under MPA immunosuppressive therapy. The rate of CMV infection episodes in the low-MPA pharmacological efficacy group categorized at 2 weeks after renal transplantation was 5/6 (83.3%), which was significantly higher than the rate of 1/10 (10.0%) (p < 0.01) in the high-MPA sensitivity group. However, the MPA pharmacological efficacy evaluated both before and after transplantation had no relationship with the incidence of rejection episodes. These findings suggest that the MPA pharmacological efficacy evaluated by LIST at 2 weeks after operation is a useful biomarker for predicting the following occurrence of CMV infection episodes in renal transplant recipients.