Decreased cytotoxicity and increased antimitotic activity of a proline analogue of chlorambucil as a prodrug susceptible to the action of fibroblast's prolidase

We synthesized an proline analogue of chlorambucil (CH-pro) as a prodrug su sceptible to the action of ubiquitously distributed, cytosolic imidopeptida se - prolidase [E.C.3.4.13.9]. A conjugation of chlorambucil (CH) with prol ine through an imido-bond resulted in the formation of a good substrate for prolidase. We have compared several aspects of biological actions of CH an d its prodrug in cultured normal human skin fibroblasts. The prodrug was fo und to be more effectively transported into the cells than the free drug. M oreover, in opposition to CH, CH-pro had no inhibitory effect on fibroblast 's prolidase activity against the endogenous substrate, glycyl-L-proline. L ower cytotoxicity and a higher antimitotic activity of the prodrug, compare d to the free drug, was observed. CH and CH-pro at concentrations of 25 muM led to a 30% and 10%. decrease in cell viability in confluent human skin f ibroblasts. IC50 values of CH and CH-pro for DNA synthesis was found to be 30 muM and 7 muM, suggesting higher antimitotic potency of the pro-drug com pared to the free drug. CH-pro also evoked lower ability to inhibit collage n biosynthesis in cultured fibroblasts than the free drug. IC50 values of C H and CH-pro for collagen biosynthesis were found at about 15 muM and 30 mu M, respectively. Targeting of prolidase as a prodrug-converting enzyme may serve as a novel strategy in pharmacotherapy of various diseases, leading t o the increase in therapeutic efficacy and reduction in untoward side effec ts of antineoplastic agents.