Beta-lyase-dependent attenuation of cisplatin-mediated toxicity by selenocysteine Se-conjugates in renal tubular cell lines

Cisplatin [ cis -diamminedichloroplatinum(II)] is a widely used antitumor drug with dose-limiting nephrotoxic side effects due to selective toxicity to the proximal tubule. In the present study, the chemoprotective potential of three selenocysteine Se -conjugates, Se -methyl-l-selenocysteine, Se -(2-methoxyphenyl)-l-selenocysteine, and Se -(2-chlorobenzyl)-l-selenocysteine, belonging to three structural classes, against the nephrotoxic effects of cisplatin was investigated. Selenocysteine Se -conjugates have previously been proposed as kidney-selective prodrugs of pharmacologically active selenols because of their active uptake and bioactivation by cysteine conjugate β-lyases in the kidney. To elucidate whether chemoprotection is β-lyase-dependent wild-type LLC-PK1 cells, possessing a very low β-lyase activity, and LLC-PK1 cells stably transfected with full-length cDNA coding for rat kidney cysteine conjugate β-lyase/glutamine transaminase K (R1J) were used. The results indicate that all three selenocysteine Se -conjugates were able to attenuate the cisplatin-induced loss of viability in R1J cells but not in the parental LLC-PK1 cells, as determined by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and neutral red uptake. In addition, cisplatin-induced reactive oxygen species (ROS) production was determined using 2′,7′-dichlorodihydrofluorescein diacetate. The selenocysteine Se -conjugates were able to decrease ROS levels after cisplatin exposure in both cell types. However, this ROS-protective effect was more profound in R1J cells. Se -Methyl-l-selenocysteine provided the strongest protection. The protective activity against cisplatin-induced cytotoxicity and ROS generation was blocked by aminooxyacetic acid, a selective inhibitor of pyridoxal 5′-phosphate-dependent cysteine conjugate β-lyases, further supporting the role of β-lyase in the observed chemoprotection. The precise molecular mechanism by which selenols, generated by β-lyase, provide protection against cisplatin-induced cytotoxicity, however, remains to be established.