Comet assay measures of DNA damage as biomarkers of irinotecan response in colorectal cancer in vitro and in vivo

The use of irinotecan to treat metastatic colorectal cancer (CRC) is limited by unpredictable response and variable toxicity; however, no reliable clinical biomarkers are available. Here, we report a study to ascertain whether irinotecan-induced DNA damage measures are suitable/superior biomarkers of irinotecan effect. CRC-cell lines (HCT-116 and HT-29) were treated in vitro with irinotecan and peripheral blood lymphocytes (PBL) were isolated from patients before and after receiving irinotecan-based chemotherapy. Levels of in vitro-, in vivo-, and ex vivo-induced DNA damage were measured using the Comet assay; correlations between damage levels with in vitro cell survival and follow-up clinical data were investigated. Irinotecan-induced DNA damage was detectable in both CRC cell-lines in vitro, with higher levels of immediate and residual damage noted for the more sensitive HT-29 cells. DNA damage was not detected in vivo, but was measurable in PBLs upon mitogenic stimulation prior to ex vivo SN-38 treatment. Results showed that, following corrections for experimental error, those patients whose PBLs demonstrated higher levels of DNA damage following 10 h of SN-38 exposure ex vivo had significantly longer times to progression than those with lower damage levels (median 291 vs. 173 days, P = 0.014). To conclude, higher levels of irinotecan-induced initial and residual damage correlated with greater cell kill in vitro and a better clinical response. Consequently, DNA damage measures may represent superior biomarkers of irinotecan effect compared to the more often-studied genetic assays for differential drug metabolism.