An in-vitro model of the effects of pH on cisplatin-induced nephrotoxicity

Abstract

2077 Background: There is evidence that acidic conditions potentiate the cytotoxicity of Cisplatin (Cp) (F. Tanaka et al, Hearing Research: 177, 2003, 21–31). However, little is known about the effects of pH on Cp nephrotoxicity in humans. It was indicated that the proximal renal tubules in humans are more affected by Cp cytotoxicity than the distal renal tubules (G. Daugaard et al, Clin Pharmacol Ther.: 44(2), 1988,164–172). We hypothesize that pH variations in renal tubules are important modulators of Cp nephrotoxicity. Methods: LLC-PK1 (porcine proximal) and MDCK (canine distal) renal tubular epithelium cells were used as in-vitro models to investigate the effect of pH variations on Cp-induced nephrotoxicity. The IC50 of Cp at different pHs after one-hour exposure was determined for these two cell lines using the MTT assay. For LLC-PK1 cells, C-14 thymidine assay was used to examine the effect of Cp on DNA synthesis, and Platinum (Pt) uptake was measured by atomic absorption spectroscopy. Similar experiments are ongoing for MDCK cells. Results: The IC50 of Cp for LLC-PK1 cells at pH 6.0, was 33 μM Cp. The IC50s for MDCK cells was 75 μM (n=3, P< 0.005, r2=0.98). The table below shows the results of LLC-PK1 cells as a function of extracellular pH (pHe) (n=3, P<0.005). Standard errors are not listed, to save space. The highest inhibition in DNA synthesis (12%±3 SD, P<0.0001), and the highest uptake of platinum (Pt) (180ng/106 cells) were detected at pH 6.0 compared to the other pH values tested. Conclusions: These cells developed maximal Cp-induced cytotoxicity at pHe=6.0. This cytotoxicity in LLC-PK1 cells is correlated with intracellular Cp uptake, and with inhibition of DNA synthesis. If impairment of DNA synthesis is ultimately found to be the mechanism of clinical Cp-induced nephrotoxicity, then strategies can be developed to overcome this process, and improve the therapeutic index of Cp. No significant financial relationships to disclose.