Downregulation of intracellular nm23-H1 prevents cisplatin-induced DNA damage in oesophageal cancer cells: possible association with Na+, K+-ATPase

Previously, we showed that expression of nm23-H1 is associated inversely wi th sensitivity to cisplatin in human oesophageal squamous cell carcinoma (O SCC). The present study was undertaken to investigate the association of nm 23-H1 expression with cisplatin-induced DNA damage in OSCC using antisense nm23-H1 transfectants. YES-2/AS-12, an antisense nm23-H1-transfected OSCC c ell line, showed significantly reduced expression of intracellular nm23-H1 protein compared with that in parental YES-2 cells and YES-2/Neo transfecta nts. Surface expression of nm23-H1 protein was not observed in any of the t hree cell lines. PCR analysis for DNA damage demonstrated that YES-2/AS-12 cells were more resistant to nuclear and mitochondrial DNA damage by cispla tin than were YES-2/Neo cells. In addition, mitochondrial membrane potentia ls and DNA fragmentation assays confirmed that YES-2/AS-12 was more resista nt than YES-2/Neo to apoptosis induced by cisplatin. In contrast, YES-2/AS- 12 was more sensitive to ouabain, a selective inhibitor of Na+, K+-ATPase, than YES-2 and YES-2/Neo. Pre-treatment with ouabain resulted in no differe nces in cisplatin sensitivity between the three cell lines examined. Intrac ellular platinum level in YES-2/AS-12, was significantly lower than that in YES-2 and YES-2/Neo following incubation with cisplatin, whereas ouabain p re-treatment resulted in no differences in intracellular platinum accumulat ions between the three cell lines. Our data support the conclusion that red uced expression of intracellular nm23-H1 in OSCC cells is associated with c isplatin resistance via the prevention of both nuclear and mitochondrial DN A damage and suggest that it may be related to Na+, K+-ATPase activity, whi ch is responsible for intracellular cisplatin accumulation. (C) 2000 Cancer Research Campaign.