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It has been suggested that mutagen sensitivity is a constitutional factor which may be useful in identification of patients with an increased risk for the development of tumors. In this study, the chromosome sensitivity to bleomycin was measured according to Hsu in patients with hereditary non-polyposis colorectal cancer (HNPCC), sporadic colorectal cancer and in control persons with no tumor history in family. In vitro lymphocytes were exposed to bleomycin according to Hsu and chromosomal damage was quantified by scoring breaks of 100 cells. A significant difference (P < 0.01) in the mean number of breaks per cell (b/c) was found between HNPCC patients (0.59 ± 0.14; n = 12; mean age 55.4 yrs) and control individuals (0.35 ± 0.13: n = 12; mean age 55.8 yrs). In contrast, patients with sporadic colorectal cancer showed a mean b/c value of 0.43 ± 0.14 (n = 14; mean age 63.4 yrs) which was not significantly higher than that in control individuals for this group (0.42 ± 0.15; n = 14; mean age 63.1 yrs). Selenium protected lymphocytes of HNPCC patients against bleomycin activity in vitro.
Using the alkaline comet assay, we showed that bleomycin at 0.1-5 μg/ml induced DNA strand breaks and/or alkali-labile sites, measurable as the comet tail moment, in human colonic mucosa cells. This DNA damage was completely repaired during a 120-minute post-treatment incubation of the cells. Post-treatment of the bleomycin-damaged DNA with 3-methyladenine-DNA glycosylase II (AlkA), an enzyme recognizing alkylated bases, gave rise to a significant increase in the extent of DNA damage, indicating that the drug could induce alkylative bases in DNA. We did not observe any change in the comet tail moment in the presence of catalase. Vitamin E ((+)-α-tocopherol) decreased DNA damage induced by bleomycin. The results obtained suggest that hydrogen peroxide might not be involved in the formation of DNA lesions induced by bleomycin in the colonic mucosa cells, and that vitamin E may exert protective effects on these cells.
Bleomycin (BLM) is a natural antibiotic that is effective in treatment of selected cancers. Although the exact therapeutic mechanism of bleomycin is not known, its target is thought to be a nucleic acid. Besides cleaving DNA, in vitro, Fe-bleomycin cleaves the anticodon of yeast tRNAPhe specifically. Using CD and fluorescence spectroscopy we have found that apo-bleomycin binds to synthetic RNA analogs of the anticodon of yeast tRNAPhe with an affinity similar to that previously reported for DNA. In order to understand BLM's selectivity, the role magnesium ions play in RNA recognition should be explained. Many RNA substrates for Fe-BLM, including yeast tRNAPhe, are not cleaved by the drug when the Mg2+ concentration exceeds 1 mM. Competition experiments with anticodon analogs provide insight into the role of magnesium ions in RNA recognition by BLM. These simple modified RNAs may be useful as model systems for investigating BLM/RNA recognition and development of highly selective drugs toward RNA targets.
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