Quantitative determination of decitabine incorporation into DNA and its effect on mutation rates in human cancer cells.
Bottom Line: The mode of action of decitabine strictly depends on the incorporation of the drug into DNA.Finally, we also used next-generation sequencing to comprehensively analyze the effects of decitabine incorporation on the DNA sequence level.These results indicate that standard rates of decitabine incorporation are not genotoxic in myeloid leukemia cells.
Affiliation: Division of Epigenetics, DKFZ-ZMBH Alliance, German Cancer Research Center, 69120 Heidelberg, Germany.Show MeSH
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Mentions: In following experiments, we further expanded the scope of our analysis by investigating decitabine incorporation in a panel of six leukemia cell lines (ML-2, K562, KG-1, KG-1a, U937 and HL-60) and a colon cancer cell line (HCT116). When these cell lines were treated with the same dose (100 nM, 24 h) of [3H]-decitabine, we did not observe any detectable drug-induced cytotoxicity (Supplementary Table S3). However, incorporation rates ranged from 0.1 pmol/μg DNA to 0.3 pmol/μg DNA (Figure 4A, upper panel). Total intracellular decitabine levels appeared similar to the DNA-incorporated levels, with HL-60 and U937 cells showing the highest uptake (Figure 4A, lower panel). Drug-dependent differences in the cell cycle profiles were minor and not significant (P > 0.5, two-way ANOVA) for any of the seven cell lines (Figure 4B), which indicates that our results are a consequence of differential drug uptake and not influenced by differential drug toxicity. Interestingly, non-cancerous cell lines showed significantly (P = 0.007, t-test) lower levels of incorporated decitabine (Figure 4C). Similarly, total levels of intracellular decitabine were significantly (P = 0.018, t-test) lower in non-cancerous cells (Figure 4D). These findings are in agreement with the reduced cellular responses to low-dose decitabine in normal bone marrow cells (12).
Affiliation: Division of Epigenetics, DKFZ-ZMBH Alliance, German Cancer Research Center, 69120 Heidelberg, Germany.