In utero estrogenic exposures and transgenerational risk for breast cancer Background: Research in humans and animals suggests that what an expecting mother eats or is exposed to during pregnancy, can affect her offspring's susceptibility to disease in adulthood. Using a rat model, we have previously shown that a maternal diet high in fat during pregnancy is associated with increase in pregnancy estrogen levels and breast cancer risk in daughters. More recently, it was shown that what an expecting mother is exposed to during pregnancy affects risk of certain diseases not only in her offspring but subsequent generations as well. These effects are thought to be inheritable through epigenetic mechanisms. Objective/ hypothesis: We propose to study whether exposing pregnant rats to a high fat diet or an estrogen- supplemented diet will affect the risk of mammary cancer in the offspring and two subsequent generations. We hypothesize that in utero estrogenic exposures will increase the risk of mammary cancer of the female offspring (F1) and will be transmitted to the subsequent generations (F2/F3) through epigenetic mechanisms. A better understanding of how the in utero environment leads to modulation of breast cancer risk in the offspring and subsequent generations could have great implications for breast cancer prevention. Specific Aims: (1) Determine whether exposure of pregnant rats to estrogens or a high-fat diet is associated with increased mammary cancer risk in the offspring (F1) and subsequent generations(F2/F3). (2) Determine whether these in utero exposures are associated with changes in mammary gland developmental biology in the offspring (F1) and subsequent generations (F2/F3). (3) Determine whether the effects of these in utero exposures are transmitted to subsequent generations through epigenetic mechanisms. Study Design: Pregnant Sprague-Dawley rats will receive a high fat diet or an estradiol supplemented diet during gestation and mammary cancer development will be monitored in the offspring (F1). To address if the phenotype seen in the offspring is passed to subsequent generations, we will mate the female and male offspring (F1) and examine mammary cancer risk in the next generation (F2). Similarly, the F2 generation will be mated and mammary cancer examined in the offspring (F3). We will examine whether the mammary cancer phenotype in the F1, F2 and F3 generations is associated with changes in epigenetic and gene expression patterns in the mammary gland, employing genome-wide methylation and microarrays studies. We will also examine the mammary gland morphology to assess changes in mammary gland development.