? ? Investigations of the genetic etiology of cancer have mainly focused on sequence variants that alter protein function (protein quality) such as truncating mutations, missense substitutions, and splice junction mutations. However, recent studies suggest that, for complex diseases, milder changes in protein quantity may be more important than radical changes in the protein quality. Increasing evidence has shown that regulatory polymorphisms that affect gene expression abundance can modify disease risk. The insulin-like growth factor (IGF) system plays an important role in regulating cellular proliferation and apoptosis. Deregulation of the IGF system leads to cancer cell growth and survival. Converging data from genetic epidemiology and clinical studies provide strong evidence that high circulating levels of IGF1 protein and low circulating levels of IGFBP3 protein in serum significantly predict increased risks of breast cancer. Although IGF1 and IGFBP3 circulating levels are associated with increased risk of breast cancer at the population level, individual risk cannot be easily assigned by directly measuring the protein concentration in serum, because the normal IGF1 and IGFBP3 protein ranges are wide and circulating levels are influenced by several personal factors including diet, exercise, and body weight. Limited data also suggest that both IGF1 and IGFBP3 exhibit allele-specific gene expression (ASGE), which is usually controlled by regulatory polymorphisms. Therefore, the elucidation of regulatory polymorphisms responsible for increasing the risk of breast cancer and development of simple DNA-based assays may allow us to accurately identify healthy individuals who are at high risk of breast cancer in the future. Hypothesis: Regulatory polymorphisms in genes in the IGF signaling pathway, which control allele-specific gene expression (ASGE), affect risk of developing breast cancer. Specifically, we will investigate the following hypotheses: 1). the inter-individual variation in IGF1 and IGFBP3 protein levels are due to ASGE in these genes; 2). Specific haplotype(s) in IGF1 gene are correlated with over-expression of IGF1 and specific haplotype(s) in IGFBP3 correlated with reduced expression of IGFBP3. This resubmission is focused on ASGE in the IGF and IGFBP3 genes. In this application, we will identify ASGE in cell lines for these two genes, by recently developed techniques, in Aim 1, determine the haplotypes, correlated to ASGE, through re- sequencing in Aim 2, and then test if the ASGE in mRNA level results in alterations in protein level in Aim 3. We will identify haplotypes responsible for ASGE and inter-individual variation in protein levels of IGF1 and IGFBP3. This approach will lead to a new diagnostic test in which ASGE can be assayed with a DNA-based blood test. In future work, we will then determine if there is an association with breast cancer risk, as well as test for ASGE in additional genes. If there is an association with breast cancer, this test could be a valuable predictive marker of an individual's risk for breast cancer. The knowledge could then be used to assist clinicians and their patients to more accurately assess breast cancer risk and help in designing effective preventative strategies. ? ? ? ?
