Established cell lines are widely used as in vitro models in toxicology studies. The choice of an appropriate cell line is critical when performing studies to elucidate drug-induced toxicity in humans. The porcine renal proximal tubular cell line LLC-PK1 is routinely used to study the nephrotoxic effects of drugs in humans. However, there are significant interspecies differences in drug pharmacokinetics and pharmacodynamics. The objective of this study was to determine whether the human renal proximal tubular cell line HK-2 is an acceptable model to use when performing in vitro toxicity studies to predict effects in humans. We examined 2 nephrotoxic agents, ifosfamide (IFO) and acyclovir, that exhibit different clinical nephrotoxic patterns. HK-2 cells metabolized IFO to its nephrotoxic metabolite, chloroacetaldehyde (CAA). Acyclovir induced a concentration-dependent decrease in HK-2 cell viability, suggesting that acyclovir may induce direct insult to renal proximal tubular cells. The results support clinical pathology data in humans and suggest that HK-2 cells are a suitable model to use in in vitro toxicity studies to determine drug-induced nephrotoxicity in humans.