Adenosine triphosphate-dependent transport of leukotriene c₄ by membrane vesicles prepared from cisplatin-resistant human epidermoid carcinoma tumor cells

Background: Cisplatin accumulation is decreased in many cisplatin-resistant cells. An active efflux pump for cisplatin exists in cisplatin-resistant human epidermoid carcinoma cells (called KB cells). A previous study has suggested that the adenosine triphosphate (ATP)-dependent glutathione S-con-jugate export pump (GS-X pump), which exports the bis-(glutathionato)-platinum (II) (GS-platinum) complex, could contribute to cellular resistance to cisplatin. Purpose: In this study, we examined whether the active efflux pump for cisplatin in the cisplatin-resistant KB cells is the GS-X pump and tested its activity by using an endogenous substrate, [³H]leukotriene C₄ ([³H]LTC₄). Methods: Membrane vesicles were prepared from KB-3-1 (clone from parental KB cells) cells and from cisplatin-resistant KCP-4 (a mutant clone derived from KB-3-1 cells) cells. Using a filtration technique, we measured the uptake and transport of [³H]LTC₄, a substrate for the GS-X pump, into membrane vesicles at 37°C. Results: The uptake of [³H]LTC₄ in the membrane vesicles from both the KB-3-1 and KCP-4 cells was ATP-de-pendent. In contrast, the ATP-de-pendent transport of [³H]LTC₄ was observed only in KCP-4 membrane vesicles but not in KB-3-1 membrane vesicles. The ATP-dependent transport was vanadate sensitive and was inhibited by GS-platinum complex but only marginally by cisplatin and glutathione and not by vincristine or ver-apamil. The nucleotide triphosphates, guanosine triphosphate, cytidine triphosphate, uridine triphosphate, and deoxythymidine triphosphate could be substituted for ATP but were less efficient. A nonhydrolyzable ATP analogue, adenosine 5'-(β, γ-methylene) triphosphate, was not effective. Conclusions: The transport of LTC₄ in membrane vesicles prepared from KCP-4 cells was facilitated by an ATP-dependent pump that appeared very similar to the GS-X pump. Implications: Our study suggests that the GS-X pump is involved in the decreased accumulation of cisplatin in KCP-4 cells. [J Natl Cancer Inst 86:1781-1784, 1994]