Murine L1210 leukemia cells resistant to the antineoplastic agent L-Phenylalinine Mustard have a 1.5-2.0 fold elevation in their cellular GSH and GSSG content. Cellular uptake of L-(14C(U)) cystine and its incorporation into GSH of the resistant tumor is correspondingly evelated. Synthesis of gamma-glutamyclysteine, GSH and GSSG is elevated 1.5-2.0 fold in cell-free preparations of the resistant tumor. This increased synthesis of GSH is attributed to increased cellular content (1.6-fold) of gamma- glutamylycsteine synthetase. GSH synthetase activity is equivalent in both drug sensitive and resistant cells. Investigation into the hydrolysis of selected peptides by cell-free preparations of both sensitive and resistant tumors suggest that Aminopeptidase M participates in the formation of L-cysteine from L-Cys-Gly. This is supported by the observation that these preparations readily degrade L-Leu-p-nitroanilide and L-Ala-L- Ala, known substrates for Aminopeptidase M, but not dipeptidase. The failure of the tumors to degrade Gly-D-Ala, a dipeptidase substrate, and the marked inhibition of L-Ala-Gly, L-Cys-Gly and L-Ala-L-Ala-L-Ala hydrolysis be bestatin further support a role for Aminopeptidase M in the generation of L-cysteine from L- Cys-Gly. These results suggest that the drug-resistant tumor cell has developed an efficient mechanism for maintenance of elevated GSH which involves both gamma GT initiated catobolism of GSH to cysteine and its reutilization by gamma-glutamyl- cysteine synthetase.
