Mechanistic study of potentiation of chemotherapy by a haloenol lactone derivative in vitro.

PURPOSE: The objective of this study was to understand the biochemical mechanisms by which a haloenol lactone (HEL) derivative potentiates cisplatin-induced cytotoxicity in vitro. HEL was originally designed and synthesized as a site-directed inactivator of glutathione S-transferase pi isozyme (GST-pi). Over-expression of GST-pi has been found to be associated with chemotherapy resistance. METHODS: A concentration-dependent GST inhibition was assessed after UOK130 cells were exposed to HEL at concentrations of 10 and 20 muM. Potentiated cytotoxicity was evaluated by treatment of UOK130 cells with a selection of alkylating agents in the presence or absence of HEL. Intracellular glutathione (GSH) was determined after exposure to HEL. Protective effect of GSH was examined by co-treatment with GSH ester in UOK130 cells exposed with a combination of cisplatin and HEL. Multiple resistance-associated protein (MRP) 1-3 activity was assayed by determining the rate of (3)H-LTC(4) and (3)H-E(2)17betaG through the MRPs into recombinant membrane vesicles. RESULTS: Exposure of HEL at 10 and 20 muM caused 28 and 41% of inhibition of cellular GST activity. Cytotoxicity of cisplatin, chlorambucil, and melphalan was enhanced 1.8-2.7-fold by HEL at 10 muM. No significant protection effect by GSH ester exposure was observed on cisplatin toxicity co-treated with HEL. HEL was found to inhibit MRP1, MRP2, and MRP3 with IC(50) of 1.30, 28.2, and 3.66 muM, respectively. CONCLUSION: Haloenol lactone showed inhibitory effect on GST-pi and MRP1-3 (selective inhibition of MRP1 and MRP3), and it was also found to deplete intracellular GSH.