DNA Damage as Women Age: Implications for Breast Cancer

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DESCRIPTION (provided by applicant): The objectives of the proposed research are to delineate age-related changes in DNA structure in the epithelium, myoepithelium, and stroma of the healthy and neoplastic breast. Reciprocal interactions between these cellular fractions have been shown to be pivotal in tumor development in the breast and other hormone response tissues and may at least partially result from alterations in DNA structure produced by reactive oxygen species (e.g., free radicals) and other factors in the microenvironment. Our specific aims are: I. To isolate nuclear DNA of pure epithelium, myoepithelium, and fibroblast-enriched stroma from normal tissue from the non-cancerous breast (NN) and from histologically normal tissue from the cancerous breast (NC) of women aged 20-85 years. We will obtain the required quantities of DNA to perform the analyses outlined in aims II and III. II. To determine base lesion concentrations of 8-hydroxyadenine (8-OH-Ade), 8-hydroxyguanine (8-OH-Gua), 4,6- diamino-5-formamidopyrimide (Fapyadenine [FapyAde]), and 2,6-diamino-4-hydroxy-5-formamidopyrimide (Fapyguanine [FapyGua]) and the ratio of mutagenic 8-hydroxy (8-OH) to non-mutagenic Fapy lesions in DNA from each of the isolated cellular fractions in relation to a woman's age using gas chromatography-mass spectrometry (GC-MS). We hypothesize (a) that the mutagenic and other base lesion profiles will reflect a relatively high degree of base oxidation which will change as a function of a woman's age, and (b) that age-base lesion relationships will be different for each of the three cellular fractions. III. To determine Fourier transform-infrared (FT-IR) spectral profiles for the isolated DNA reflecting vibrations in base and phosphodiester-deoxyribose backbone structures, thus obtaining a broad understanding of alterations in functional group (e.g., NH2, C-O) and conformational structure as biomarkers for DNA changes in each cellular fraction in relation to a woman's age. We hypothesize (a) that the FT-IR spectral profiles will change as a function of a woman's age, and (b) that age-spectral profile relationships will be different for each of the three cellular fractions. IV. To integrate, using established statistical protocols, the age data with the GC-MS and FT-IR biomarker data for the three cellular fractions (epithelium, myoepithelium, and stroma) isolated from the two tissue types (histologically normal tissues from cancerous and non-cancerous breasts). We hypothesize that this integrated approach will lead to an initial, first level of understanding of age-related DNA changes in cellular fractions whose reciprocal interactions have been widely shown to be pivotal in breast cancer development.