Abstract 3529: Cisplatin-induced downregulation of OCTN2 affects carnitine (vitamin BT) wasting

Abstract

Abstract Carnitine is an essential cofactor for mitochondrial fatty acid oxidation. It is normally maintained at a steady level in the blood, and is retained in the body by a rescue mechanism that involves renal tubular reabsorption of filtered carnitine by OCTN2, a luminal organic cation transporter. The physiological significance of OCTN2 has been confirmed by the identification of mutations that cause a potentially lethal autosomal recessive disease known as primary systemic carnitine deficiency. Heterozygosity for OCTN2 mutations is associated with an intermediate carnitine-deficiency phenotype, suggesting that even partial loss of transporter function may be detrimental. The nephrotoxic agent cisplatin was previously reported to cause urinary loss of carnitine in cancer patients, although the mechanism by which this occurs remains unknown. We hypothesized that cisplatin may influence the function of OCTN2 and/or catalytic enzymes involved in carnitine homeostasis. In wildtype mice, we found that the urinary excretion of carnitine and acetylcarnitine at baseline over a 24-hour period was about ∼3 fold higher than in mice lacking the organic cation transporters Oct1 and Oct2 [Oct1/2(−/−) mice] (P<0.001), indicating that carnitine may undergo carrier-mediated basolateral uptake in renal tubular cells. Transport of carnitine (50 nM) was confirmed in 293 Flp-In cells overexpressing OCT2 (P=0.0034), and carnitine (1 mM) itself caused approximately 40% inhibition of OCT2 function as assessed by changes in transport of [14C]tetraethylammonium, a known OCT2 substrate. In wildtype mice, a single dose of cisplatin (10 mg/kg, i.p) caused a time-dependent increase in urinary carnitine excretion on days 2 and 3 (2-3 fold), and a 7-14 fold increase in acetylcarnitine excretion on days 1-3 post-treatment. Cisplatin-induced changes in urinary carnitine and acetylcarnitine were not observed in Oct1/2(−/−) mice, suggesting that renal tubular transport of cisplatin by Oct1 and Oct2 is a prerequisite for treatment-related disturbances in carnitine excretion. One possible explanation for this phenomenon involves inhibition of OCTN2 activity by cisplatin. However, cisplatin and a mixture of cisplatin-glutathione conjugates (100 µM; 30 min) did not substantially affect carnitine transport in HEK293 cells stably transfected with OCTN2. Next, we considered the possibility that cisplatin affects the expression of Octn2 and/or carnitine-palmitoyltransferases (Cpt1/2) involved in the formation of acetylcarnitine. In wildtype mice, mRNA expression analysis in kidney biopsies following treatment with cisplatin indicated a dramatic downregulation of Octn2 as well as upregulation of Cpt1b in a time-dependent manner, which changes were absent in the Oct1/2(−/−) mice. This study suggests that carnitine wasting associated with cisplatin chemotherapy is at least partially dependent on drug-induced downregulation of OCTN2 Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3529.