Evaluations of a human kidney (HK2 cells) model for drug-induced cytotoxicity and BK virus infection

Abstract

Renal transplant patients receive drugs such as tacrolimus (TAC), sirolimus (SRL), cyclosporin (CSA) for immunosuppression, and low-dose cidofovir (CID) for treatment of BK virus associated nephropathy (BKVN). In this series of studies, we evaluated the effects of clinically achievable concentrations of drug combination therapy on p-glycoprotein (P-GP) and organic anion (OA) transporters (OAT-1,-3 and MRP2) expression and function in human renal proximal tubule (HK2) cells. HK2 cells were further evaluated as a host for BK virus and a model for drug screening for the treatment of BKVN. HK2 cells were exposed to drug combinations over 5 days with determination of P-GP and OAT-1 expressions and functions in HK2 cells. P-GP function was assessed by rhodamine 123 (R123) uptake and OA transport activity was evaluated by para-aminohippurate transport (PAH) during acute and chronic exposures. Exposure to supraclinical concentrations of TAC, SRL and CSA resulted in significant reductions in cell viability whereas expression of P-GP, and OA transporters remained unchanged. A significant concentration-dependent toxicity was demonstrated by trypan blue exclusion but not by XTT assay, suggesting the importance of drug-uptake mechanism in designing experiments. R123 accumulation was significantly increased during TAC, SRL and CSA exposure. PAH transport remained unchanged in the presence of TAC, SRL, and CSA; however a trend towards decreased transport in the presence of CID was observed. HK2 cells were shown to express Caveolin-1, and were permeable to BK virus. Viral growth was gradual after 20 days post-infection, and stabilized after 55 days. A possible dormant stage of BK viral cycle was observed. The efficacy of CID was tested on the BK virus-infected HK2 cell model, and the quantitative PCR data showed that 24-hour CID exposure suppressed viral replications after 4 days incubation in a concentration-dependent manner. The key finding in these studies suggest that immunosuppressants used in renal patients may impair renal P-GP function. Exposure to drug combinations including CID may increase the risk of nephrotoxicity in patients receiving immunosuppressants. A BK virus infected HK2 cell model was developed that could be further utilized to study BK virus infection and new treatments for BKVN.