PROLIFERATION-DEPENDENT AND PROLIFERATION-INDEPENDENT CYTOTOXICITY BY ANTITUMOR DIARYLSULFONYLUREAS - INDICATION OF MULTIPLE MECHANISMS OF DRUG-ACTION

The mechanism(s) by which antitumor diarylsulfonylureas (DSU) cause cytotoxicity has been examined in GC3/Cl human colon adenocarcinoma cells and a subline selected for resistance to N-(5-indanylsulfonyl)-N'-(4-chlorophenyl)urea (ISCU). Resistance was stable in the absence of selection pressure. This mutant (designated LYC5) was 5.5-fold resistant to ISCU compared to parental GC3/C1 cells in serum containing medium when cells were exposed for 7 days. In contrast, LYC5 cells were not resistant to a 4-hr exposure to ISCU. These data indicated two possible mechanisms of action, dependent on concentration and time of exposure to ISCU. Proliferation-dependent and -independent mechanisms of cytotoxicity were identified in wild-type and resistant clones. In serum-free medium containing growth factors, the Ic50 for parental cells was 0.51 muM and for LYC5 7.0 muM (13.6-fold resistance), whereas without growth factors both lines were 8- to 9-fold resistant relative to conditions of cellular proliferation. Accumulation of ISCU was similar in quiescent and proliferating cells, and was reduced only slightly in resistant LYC5 cells. Analysis of DNA by agarose gel electrophoresis showed that in GC3/C1 cells nucleosomal ladders were formed only when proliferating cells were exposed to ISCU. No nucleosomal ladders were detected in quiescent cells during exposure to toxic concentrations of drug (IC90), or after removal of ISCU and addition of serum to stimulate growth. These data indicate several mechanisms by which diarylsulfonylurea antitumor agents may cause cell death. In serum-free medium at very high concentration (IC50 approximately 370 muM) for short periods of exposure (4 hr), cytotoxicity was proliferation independent, and GC3/Cl and LYC5 cells were equally sensitive. This mechanism may relate to the uncoupling activity of ISCU. However, at pharmacological relevant concentrations, the primary mechanism was proliferation dependent and led to formation of nucleosomal DNA ladders (IC50 approximately 0.5 muM). A possible additional mechanism occurred at higher concentration (IC50 approximately 7 muM) in quiescent cells, and was not associated with DNA degradation.