ALTERED GENE-EXPRESSION IN HUMAN LEUKEMIA K562 CELLS SELECTED FOR RESISTANCE TO ETOPOSIDE

Sublines of K562 human leukemia cells were selected for resistance (30 - to 80-fold) to etoposide by continuous exposure to 0.5 mu M VP-16. T wo etoposide-resistant cell lines, K/VP.S and K/VP.5-1, showed a 5-fol d reduction in levels of topoisomerase II alpha protein compared with K562 cells. Northern analysis indicated a 2.5-fold reduction in topois omerase II alpha mRNA in etoposide-resistant cell lines, due in part t o a 1.2-fold decrease in topoisomerase II mRNA stability with no chang e in transcription rate. Immunoblotting assays of electrophoresed cell lysates from VP-16-treated cells revealed less drug-induced covalent topoisomerase II/DNA adducts in resistant than in sensitive cells, sug gesting a functional alteration in resistant cell topoisomerase II. Re cent reports of specific topoisomerase II DNA binding sites near the p romoter sites of growth response genes and alterations of gene express ion in cells treated with topoisomerase II inhibitory drugs led to exp eriments to determine if the apparent functional alterations of topois omerase II were accompanied by changes in the regulation of these gene s. Therefore, the expression of several growth response genes was comp ared by northern analysis in parental K562 and both VP-16-resistant ce ll lines. Basal levels of c-myc were comparable for all three cell lin es, but revels of c-jun and c-fos were elevated 2- to 4-fold in VP-16- resistant cell Lines. Increased levels of c-fos and c-jun were not a r esult of altered rates of transcription, as determined by nuclear run- off assays. Exposure of both sensitive and resistant cells to 200 mu M VP-16 for 5 hr resulted in no further changes in topoisomerase II mRN A levels but caused an additional 2- to 3-fold elevation in the level of c-jun mRNA, indicating that altered basal levels of this gene were not due to deregulation of this gene. Acquired VP-16 resistance in K/V P.5 and K/VP.5-1 cells was accompanied by reduced levels and altered a ctivities of DNA topoisomerase II as well as changes affecting the exp ression of genes important for growth and differentiation.