DOWN-REGULATION OF C-MYC GENE-EXPRESSION WITH INDUCTION OF HIGH-MOLECULAR-WEIGHT DNA FRAGMENTS BY FLUORODEOXYURIDINE

5-Fluoro-2'-deoxyuridine (FdUrd), a potent inhibitor of thymidylate sy nthase, induces extensive bulk DNA damage at drug concentrations that produce significant in vitro growth inhibition of human ileocecal carc inoma (HCT-8) cells. Constant- and pulsed-field gel electrophoresis (C FGE and PFGE), to detect size distribution of DNA double-strand breaks and repair kinetics, in parallel with northern and western blot analy ses, to quantitate c-myc gene and protein expression, were utilized to analyze drug effects. At 24-hr post in vitro drug treatment, when max imum bulk DNA damage was detected, FdUrd produced a broad range of hig h molecular weight DNA fragments, clustering between 0.1 and 5.7 megab ases in size, and resulted in a decrease in the level of c-myc transcr ipts and protein with no significant effect on the level of v-myc and H-ras. These effects preceded the observed cellular growth inhibition. Addition of the reduced folate leucovorin potentiated the effects ind uced by FdUrd, indicating that thymidylate synthase inhibition is an i mportant initial step in drug effect followed by DNA fragmentation and suppression of c-myc expression. Changes in the integrity of the gene tic materials and regulatory genes occurred prior to the observed cell growth inhibition by FdUrd, suggesting that these molecular alteratio ns by FdUrd may be associated with subsequent FdUrd-induced cell growt h inhibition.