Characterization of the reactions of platinum antitumor agents with biologic and nonbiologic sulfur-containing nucleophiles

Abstract

Substitution reactions with biologic nucleophiles appear to govern the antitumor and toxic properties of platinum complexes. In this paper we have characterized the reactions of several platinum antitumor agents with sulfur-containing amino acids, peptides, proteins, and nonbiologic nucleophiles. The rate constants for the reactions of trans-diamminedichloroplatinum(II) (trans-DDP), cis-diamminedichloroplatinum(II) (DDP), diammine (1,1-cyclobutanedicarboxylato)platinum(II) (CBDCA) and cis-diisopropylamine-cis-dichloro-trans-dihydroxy platinum(IV) (CHIP) with cysteine (Cys), methionine (Met), and glutathione (GSH) were determined at 37 degrees. A reactivity ratio of 1:1.5:22:6500 was determined for the reaction of GSH with CHIP, CBDCA, DDP, and trans-DDP respectively. The rate constant for the binding of DDP to DNA, 7.4 X 10(-5) sec-1, decreased to 5.9 X 10(-5) sec-1 and 1.7 X 10(-5) sec-1 in the presence of 0.5 and 5 mM GSH respectively. The products formed in the reaction of GSH with trans-DDP, DDP, and CBDCA were also examined. Under conditions of high platinum concentration (2-3 mM), CBDCA and DDP form large molecular weight species with GSH as indicated by 1H-NMR and ultrafiltration experiments. The complex [Pt(GSH)2 X 3H2O]n was isolated from the reaction of 3 mM DDP with 6 mM GSH. The product formed in the reaction of 3 mM trans-DDP with 6 mM GSH was not macromolecular in nature, and 1H-NMR spectra revealed that platinum was bound to the Cys sulfhydryl group. Rate constants were determined for the reactions of these platinum complexes with diethyldithiocarbamate (DDTC) and thiosulfate, two agents known to reduce platinum-mediated nephrotoxicity. DDTC, but not thiosulfate, was shown to rapidly chelate platinum from [Pt(GSH)2 X 3H2O]n. The effects of DDP, CBDCA, and CHIP on the sulfhydryl-dependent rat renal proximal tubule membrane enzymes alkaline phosphatase (AP), gamma-glutamyltranspeptidase (GGTP), leucine aminopeptidase (LAP), and the Na+/K+- and Mg2+-adenosine-5'-triphosphatases (ATPases) were also investigated in vitro. The ability of platinum complexes to inhibit these enzymes parallels their reactivity with other nucleophiles. DDTC and thiourea were shown to restore activity to platinum-inhibited enzymes. Chloride ion was found to reduce platinum-mediated enzyme inhibition in an unpredictable manner, the greatest effect being observed with LAP and GGTP and the least with the ATPases. None of these renal enzymes was directly inhibited by DDP in vivo.