OBJECTIVES/GOALS: The molecular basis of increased risk of triple negative breast cancer in non-Hispanic Black women represents a critical knowledge gap that this research is designed to address; successful completion of this work could lead to better prevention, earlier stage diagnoses, and possible discovery of novel therapeutic strategies for this population. METHODS/STUDY POPULATION: We have recently generated a living tissue cohort of 11 non-Hispanic Black and 25 non-Hispanic White women who underwent breast surgery at Mayo Clinic. Gene expression profiling of normal breast tissue from this cohort has identified a pattern of gene expression differences that have been associated with the development of basal breast cancer and are also reflective of Hedgehog (Hh) signaling. We will identify protein-based biomarkers for Hedgehog signaling within normal breast tissue using immunohistochemistry methods. We will culture primary human mammary epithelial cells and further separate luminal and myoepithelial cells using flow cytometry to then decipher Hedgehog signaling. RESULTS/ANTICIPATED RESULTS: We anticipate identifying and localizing protein-based biomarkers for Hedgehog signaling within myoepithelial cells of non-Hispanic Black women. Using our findings, we aim to create a biomarker risk model for triple negative breast cancer and validate this model within a separate and larger cohort of women to predict breast cancer risk. DISCUSSION/SIGNIFICANCE: In addition to immediate benefits from improved risk prediction, the proposed work has the potential to provide new insight into the driving forces underlying basal breast carcinogenesis and the distinct biological differences that distinguish non-Hispanic Black women from non-Hispanic White women.