Inhibition of gamma-glutamyl transpeptidase or cysteine S-conjugate beta-lyase activity blocks the nephrotoxicity of cisplatin in mice

Abstract

Cisplatin is nephrotoxic. The mechanism underlying this organ-specific toxicity is unknown. We hypothesize that cisplatin is metabolized via a gamma-glutamyl transpeptidase (GGT) and cysteine S-conjugate beta-lyase-dependent pathway that has been shown to activate several haloalkenes to nephrotoxins. To test this hypothesis, we inhibited GGT and cysteine S-conjugate beta-lyase in C57BL/6 mice and analyzed the effect of the inhibitors on the nephrotoxicity of cisplatin. GGT was inhibited by pretreating the mice with acivicin. Cysteine S-conjugate beta-lyase was inhibited by aminooxyacetic acid (AOAA). Male C57BL/6 mice were treated with 15 mg/kg cisplatin (i.p.) and sacrificed on day 5. Half the mice treated with cisplatin alone died before sacrifice. The cisplatin-treated mice sacrificed at 5 days had significantly elevated levels of blood urea nitrogen (BUN). Histologic analysis revealed severe damage to the renal proximal tubules. Pretreatment with acivicin or AOAA protected the mice from the nephrotoxicity of cisplatin. None of the pretreated animals died before sacrifice. BUN levels and quantitative histologic analysis of the kidneys confirmed the protective effect of acivicin and AOAA. Platinum levels in the kidneys were not altered by acivicin or AOAA, indicating that neither affected the uptake of cisplatin into the kidney. Likewise, cisplatin-induced weight loss was not altered by acivicin or AOAA, suggesting that weight loss and nephrotoxicity are via distinct mechanisms. These data support the hypothesis that the nephrotoxicity of cisplatin is due to the metabolism of a platinum-glutathione conjugate by GGT and cysteine S-conjugate beta-lyase to a potent nephrotoxin.

Fig. 1.

Structure of cisplatin and nephrotoxic haloalkenes and proposed metabolism of cisplatin to a nephrotoxin. A, the structure of cisplatin and two nephrotoxic haloalkenes: trichloroethene and hexachloro-1,3-butadiene. B, the proposed pathway for the metabolism of cisplatin to a nephrotoxin. The pathway is based on the metabolic pathway through which the haloalkenes are activated to nephrotoxins. The first step is the conjugation to glutathione, followed by the cleavage by GGT to a cysteinyl-glycine conjugate, the cleavage by aminopeptidase to a cysteine S-conjugate, and the β-elimination by cysteine S-conjugate β-lyase to an unstable thiol that yields a highly reactive electrophile.

Fig. 2.

BUN values 5 Days after treatment. Mice were pretreated with saline, acivicin, or AOAA followed by an injection of either saline or 15 mg/kg cisplatin. Five days after treatment blood was collected and analyzed for BUN. Data are shown for mice pretreated with saline (solid bar), acivicin (stippled bar), and AOAA (striped bar) and treated with saline (three left bars) or cisplatin (three right bars). The bars indicate the mean of each group ± S.D. a, the mean differed significantly from all other groups (p < 0.05).

Fig. 3.

Effect of pretreatment with acivicin or AOAA on cisplatin-induced proximal tubule damage. Hematoxylin and eosin stained sections of kidneys from mice 5 days after treatment. Shown are representative sections from the saline treatment group (A), the cisplatin treatment group (B), the acivicin cisplatin treatment group (C), and the AOAA cisplatin treatment group (D). Arrows indicate protein casts.