POLYMERASE-CHAIN REACTION AS A TOOL FOR INVESTIGATIONS ON SEQUENCE-SELECTIVITY OF DNA DRUGS INTERACTIONS

Sequence-selectivity of DNA-binding drugs was recently reported in a n umber of studies employing footprinting and gel retardation approaches . In this paper we performed polymerase-chain reaction (PCR) experimen ts to study the in vitro effects of distamycin, daunomycin, chromomyci n and mithramycin. As model systems we employed the human estrogen rec eptor (ER) gene and the Harvey-ras (Ha-ras) oncogene, in order to obta in PCR products significantly differing for the A + T/G + C frequency ratio. Distamycin, daunomycin, chromomycin and mithramycin are indeed known to differentially bind to different DNA regions depending upon t he DNA sequences recognized. The main conclusion of our experiments is that distamycin, daunomycin, chromomycin and mithramycin inhibit poly merase-chain reaction in a sequence-dependent manner. Distamycin inhib its indeed PCR mediated amplification of AT-rich regions of the human estrogen receptor gene, displaying no inhibitory effects on PCR-mediat ed amplification of GC-rich sequences of Ha-ras oncogene. By contrast daunomycin, chromomycin and mithramycin were found to inhibit PCR-medi ated amplification of the Ha-ras GC-rich oncogene sequences. We propos e that polymerase-chain reaction technique could be applied to study t he in vivo interactions of DNA-binding drugs to specific genes in inta ct cells.