Effects of diethyldithiocarbamate and nine other nucleophiles on the intersubunit protein cross-linking and inactivation of purified human alpha 2-macroglobulin by cis-diamminedichloroplatinum(II).

The cross-linking and inactivation of the plasma protein alpha 2-macroglobulin by cis-diamminedichloroplatinum(II) (cisplatin; Gonias, S. L., and Pizzo, S. V. J. Biol. Chem., 256: 12478-12484, 1981) was used to study platinum(II)-protein binding in the presence of compounds of therapeutic or biochemical significance. Diethyldithiocarbamate, potassium cyanide (KCN), sodium thiocyanate, L-methionine, N-acetyl-L-cysteine, 2-aminoethanethiol, L-cysteine, L-lysine, L-histidine, and L-arginine demonstrated variable capacity to inhibit reaction of cisplatin with protein and to reverse bidentate platinum(II)-protein binding in the in vitro model system. alpha 2-Macroglobulin lost 90% of its activity and was completely cross-linked, as determined with polyacrylamide gel electrophoresis, after reaction with cisplatin (0.6 to 1.0 mM). When diethyldithiocarbamate (4 to 15 mM) was incubated with alpha 2-macroglobulin and cisplatin, protein inactivation and cross-linking were totally prevented. In experiments with alpha 2-macroglobulin-platinum(II) complex, purified by gel filtration chromatography, 1.0 mM diethyldithiocarbamate completely reactivated the protein and eliminated nearly all of the intersubunit cross-links. Only KCN was comparably effective as an inhibitor of the reaction of cisplatin with alpha 2-macroglobulin; however, KCN was significantly less reactive with preformed platinum(II)-protein bonds than was diethyldithiocarbamate. N-Acetyl-L-cysteine, 2-aminoethanethiol, and L-cysteine were moderately reactive with free cisplatin. This group of compounds also demonstrated a low level of reactivity with the purified alpha 2-macroglobulin-platinum(II) complex. L-Methionine inhibited reaction of cisplatin with the protein, but was ineffective at reversing the reaction in the concentration range studied. The remaining compounds had little or no effect on the reaction of cisplatin with alpha 2-macroglobulin. The ability of diethyldithiocarbamate to displace nucleophilic protein groups from highly stable bonds with platinum(II) may be critical in its function as a rescue agent, limiting cisplatin toxicity towards nontumor cells.