DNA repair capacity of lymphoblastoid cell lines from sisters discordant for breast cancer.

BACKGROUND: Interindividual differences in DNA repair capacity may influence cancer risk. We tested whether the nucleotide excision repair pathway was deficient in breast cancer case patients by analyzing sister pairs. METHODS: Cell lines derived from sisters discordant for breast cancer (137 families containing 158 case patients and 154 control sisters) were obtained from the Metropolitan New York Registry of Breast Cancer Families. Lymphoblastoid cells were treated with benzo[a]pyrene diolepoxide (BPDE) for 30 minutes and were either harvested immediately or were washed and cultured in complete medium for 4 hours to allow DNA repair. Immunofluorescence using a polyclonal anti-BPDE-DNA primary antibody was used to quantify BPDE-DNA adducts. Percent DNA repair capacity was calculated from the difference between staining immediately after treatment minus that after 4 hours of repair, divided by the initial damage and was categorized into quartiles based on control values. Odds ratios and 95% confidence intervals (CIs) were calculated using conditional logistic regression models adjusted for age at blood donation, body mass index, and smoking. Statistical tests were two-sided. RESULTS: Mean percent DNA repair capacity was lower in breast cancer case patients than in control subjects (difference = 8.6, 95% CI = 4.3 to 13.8, P = .001). Using the quartile with the highest percent DNA repair capacity as the referent group, adjusted odds ratios of breast cancer increased from 1.23 (95% CI = 0.57 to 2.65) to 2.38 (95% CI = 1.17 to 4.86) to 2.99 (95% CI = 1.45 to 6.17) (P(trend) = .002) as DNA repair capacity decreased. CONCLUSIONS: Deficient DNA repair capacity is associated with increased breast cancer risk.

Removal of benzo(a)pyrene diol epoxide (BPDE)-DNA adducts as a measure of DNA repair capacity in lymphoblastoid cell lines from sisters discordant for breast cancer.

The mutagen sensitivity assay is one of the approaches used to investigate individual DNA repair capacity. This method is based on the premise that after in vitro treatment with a test mutagen, DNA from subjects with defective repair will be more damaged than DNA from those with an efficient repair system. However, very little is known about unmeasured processes that occur between cell treatment and final assessment of DNA damage. To develop a more precise assay, we modified the traditional mutagen sensitivity assay to also include measurement of DNA damage after culturing cells in the absence of mutagen. First, we treated apparently normal and xeroderma pigmentosum lymphoblastoid cell lines with various doses of benzo(a)pyrene diol epoxide (BPDE) and harvested cells at different time points. A polyclonal antiserum against BPDE-DNA was used to quantitate levels of adducts by immunoslot-blot and immunohistochemistry. Selected conditions included treatment with 10 microM BPDE, a 4-hr culture in mutagen-free medium, and immunohistochemical measurement of BPDE-DNA adducts. The method was then applied in a pilot study to 50 lymphoblastoid lines from sisters discordant for breast cancer. There was no significant difference between cases and controls in the level of BPDE-DNA adducts in lymphoblasts harvested immediately after BPDE treatment. However, after a 4-hr culture in mutagen-free medium, the level of adducts was significantly higher (P = 0.006) among cases than in controls. There was a two-fold increase in mean adduct removal in lines from nonaffected as compared to affected sisters (44% and 22% decrease, respectively). DNA repair capacity was predictive of case status (P = 0.04) in logistic regression analysis. This method, which can be easily applied to large numbers of samples, should be useful in studies to investigate the role of DNA repair in cancer risk.