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• W1996885664 abstract "Purpose/Objective: Predicting the radiosensitivity of normal and tumor cells has great potential importance for radiation oncology. The surviving fraction after 2Gy (SF2) has been shown in many instances to predict normal tissue toxicity and tumor control probability. SF2 measurements, however, are time consuming, expensive, and very lab-dependent. We hypothesized that an analysis of proteins that bind to DNA double-strand breaks could predict radiosensitivity, and would be more practical than SF2 determination. Materials/Methods: DNA end binding complexes (DNA-EBCs) were identified by mixing nuclear protein extracts with a radiolabeled oligonucleotide, in the presence of a vast excess of supercoiled DNA (without ends, to remove non-end-specific binding proteins like histones), followed by electrophoresis and autoradiography. DNA-EBCs from 21 primary fibroblast and 15 tumor cell lines with a variety of radiosensitivities (SF2 range 0.01–0.40 and 0.30–0.80 respectively) were then correlated with SF2. To identify protein components of the DNA-EBCs, antibodies to DNA repair proteins were added to the DNA-EBC reaction. Super-shifting of DNA-EBCs demonstrated the presence of the specific protein. The nuclear protein levels of each band A component were determined in triplicate by western analysis. Results: Ten DNA-EBCs were identified. DNA-EBC analysis identified a rapidly migrating ATM-containing band (called: band-A) whose density correlated with SF2. The density of other bands, or total DNA-EBC binding, correlated more poorly with SF2 (r2<0.45). There were no differences in DNA-EBC migration pattern noted. Band-A density correlated well with SF2 (0.02 < SF2 < 0.41) in 21 primary fibroblast lines (r2=0.77). The DNA-EBC pattern of peripheral blood lymphocytes was identical to that of fibroblasts, and the band-A density was identical in matched pairs of lymphocytes/fibroblasts from an individual. Band-A density also correlated with SF2 (0.35 < SF2 < 0.80) in 15 human tumor cell lines (r2=0.91). When normal and tumor cell results are combined (Fig.), there is an excellent correlation between band-A density and Sf2 (r2=0.85). Mixing studies demonstrated that band-A density predicted SF2 of tumor cells even when contaminated with normal cells, or (in the case of xenografts) rodent cells. DNA-EBC analysis also predicted radiosensitization by two unrelated radiosensitizers: the COX-2 inhibitor SC-236, and the histone deacetylase inhibitor sodium butyrate. Supershift analysis revealed that band-A contains the following proteins: ATM, Ku70, DNA Ligase III, Rpa32, Rpa14, DNA ligase IV, XRCC4, WRN, BLM, RAD51 and p53. In a subset of primary cells that were relatively radiosensitive because of BRCA1 mutation, there was no correlation between the levels of any of these proteins and band-A density or SF2. Conclusions: Band-A density predicts SF2 of primary and tumor cells, and can predict radiosensitizing effects of biologic agents. The band-A density of lymphocytes is identical to that of fibroblasts from the same individual, demonstrating that band-A analysis of lymphocytes would be a good predictor of normal tissue SF2. DNA-EBC analysis may be a clinically useful test to predict toxicity and tumor control in individual patients." @default.
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• W1996885664 date "2003-10-01" @default.
• W1996885664 modified "2023-09-27" @default.
• W1996885664 title "Radiosensitivity can be predicted by DNA-end binding complex analysis" @default.
• W1996885664 doi "https://doi.org/10.1016/s0360-3016(03)01010-1" @default.
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