Elevation of topoisomerase I messenger RNA, protein, and catalytic activity in human tumors: demonstration of tumor-type specificity and implications for cancer chemotherapy

Abstract

Topoisomerase I has been identified as an intracellular target of camptothecin, a plant alkaloid with anticancer activity. Various lines of evidence suggest that the sensitivity of cells to this drug is directly related to the topoisomerase I content. In humans, the levels of topoisomerase I have been shown to be elevated in colorectal tumors, compared to normal colon mucosa. The aim of our study was to determine whether (a) topoisomerase I levels are elevated in other solid tumors, (b) the elevated enzyme is catalytically active in these tumors, and (c) the increase in topoisomerase I levels in colorectal tumors is a result of increased transcription or translation. Topoisomerase I levels were quantitated in crude extracts from colorectal, prostate, and kidney tumors and their matched normal counterparts by Western blotting and by direct determination of catalytic activity, and mRNA levels were determined by Northern blotting. By Western blotting, colorectal tumors showed 5-35-fold increases in topoisomerase I levels, compared to their normal colon mucosa. In the case of prostate tumors, the increase was 2-10-fold, compared with benign hyperplastic prostate tissue from the same patients. However, no difference was observed in topoisomerase I levels in kidney tumors, compared to their normal counterparts. The catalytic activity of topoisomerase I was determined by a quantitative 32P-transfer assay in crude homogenates, without isolating nuclei. Colorectal and prostate tumors exhibited 11-40- and 4-26-fold increases, respectively, in catalytic activity. However, kidney tumors did not show any alteration in catalytic activity, compared to their normal matched samples. Thus, for all three tumor types there was a good correlation between enzyme levels and catalytic activity. Finally, colorectal tumors were analyzed for steady state mRNA levels. A 2-33-fold increase in mRNA levels was found in colorectal tumors, compared to normal colon mucosa. These results suggest that alterations in topoisomerase I expression in humans are tumor type specific and that the increase in topoisomerase I levels results from either increased transcription of the topoisomerase I gene or increased mRNA stability.