Topoisomerase I levels in the NCI-60 cancer cell line panel determined by validated ELISA and microarray analysis and correlation with indenoisoquinoline sensitivity

Abstract

Topoisomerase I (Top1) is a proven target for cancer therapeutics, and the level of Top1 in tumors has been used as a biomarker for chemotherapeutic efficacy. In this study, we report the development and validation of a two-site enzyme chemiluminescent immunoassay for Top1, which we used to measure Top1 levels in the NCI-60 cancer cell line panel. Top1 levels ranged from 0.9 to 9.8 ng/mL/μg protein extract in these cell lines. Levels varied both within and between cancer types but were generally highest in colon cancer and leukemia cell lines and lowest in central nervous system and renal cancer cell lines. Top1 mRNA levels in the NCI-60 cell lines were also measured by microarray; mRNA values generally showed a good correlation with protein levels (Pearson correlation = 0.8). When these expression levels were compared with the activity of the indenoisoquinoline class of Top1 inhibitors across the NCI-60 cell panel, low levels of Top1 were associated with increased resistance to these drugs. The results of our studies indicate that our Top1 assay can be used to quantify Top1 levels in untreated cells as well as cells treated with Top1 inhibitors and that the assay has the potential to be adapted for use in predicting clinical response to Top1-active antineoplastic agents.

Sensitivity and Acquired Resistance of BRCA1;p53-Deficient Mouse Mammary Tumors to the Topoisomerase I Inhibitor Topotecan

“…Abcg2 −/− ;Brca1 −/− ;p53 −/− tumors can still develop full topotecan resistance, however, and a substantial decrease in the level of Top1 can explain resistance in several of these tumors. Remarkably, this profound decrease in Top1 protein was not accompanied by a corresponding decrease in Top1 mRNA in most tumors, in sharp contrast with the observations on camptothecin-resistant tumor cell lines (12,32,38). If downregulation of Top1 would also be posttranscriptional in human tumors, its detection by gene expression profiling would be impossible.…”

Trapping Poly(ADP-Ribose) Polymerase

“…Because the cytotoxicity of Top inhibitors appears to be positively correlated with the expression levels and activities of Top1 and Top2 (Burgess et al, 2008;O'Malley et al, 2009;Pfister et al, 2009), assays are being developed to measure the expression levels and activities of Top1 and Top2 (Pfister et al, 2009(Pfister et al, , 2012. However, as noted in a review by Pommier (2013), "there is no simple linear correlation between topoisomerase levels and drug response."…”

DNA Topoisomerase I Gene Copy Number and mRNA Expression Assessed as Predictive Biomarkers for Adjuvant Irinotecan in Stage II/III Colon Cancer

“…The active form of irinotecan stabilizes the Top1cc by interfacial inhibition, and as a result the rapid moving DNA and RNA polymerases catch up and collide with these stalled complexes whereby irreversible Top1-DNA cross-links are formed (20,21). Elevated Top1 protein levels will in theory result in increased cytotoxic activity of irinotecan, and based on in vitro cell line studies, Top1 protein has been suggested as a putative predictive biomarker (22)(23)(24)(25)(26). Attempts to clinically validate these findings in the setting of advanced colorectal cancer have been carried out in two well-designed retrospective biomarker studies (27,28).…”