Breast cancer represents a global problem, accounting for an astonishing one quarter of all cancer diagnoses in women. In just the US alone, it is estimated that more than 268,000 women will be diagnosed with an invasive form of breast cancer this year. Therefore, there is a substantially unmet medical need to identify novel mechanisms of breast cancer etiology and therapeutic approaches that will significantly improve the lives of these patients. Moreover, triple negative breast carcinoma ranks among the most aggressive forms known, with severely limited treatment options while accounting for 12% of total tumors diagnosed. This new application proposes to investigate the role of mitostatin to establish feasibility and provide proof of concept for our central hypothesis that mitostatin drives mitophagy to suppress breast cancer proliferation and migration. This working hypothesis is based on several key observations. Mitostatin localizes to mitochondria and is frequently found to be mutated or is significantly reduced or absent. We have discovered that mitostatin expression is significantly reduced across numerous subtypes of breast cancer. Strikingly, we found that patients with a high tumor expression of mitostatin was significantly correlated to have improved relapse free survival in total breast cancer as well as in triple negative circumstances. We have further discovered that mitostatin is required for stimulus- dependent mitophagy and is able to concurrently suppress proliferation and migration from 2D and 3D matrices. Therefore, these preliminary data indicate that mitostatin is of high clinical and biological relevance and is of potential importance to help stop the progression of breast cancer. The precise mechanism(s) mitostatin uses to exert these oncosuppressive functions remain unknown; and further, whether mitostatin-dependent mitophagy is required to suppress breast cancer proliferation and migration. Therefore, we plan to examine the following to decipher the in vitro and in vivo role of mitostatin in breast cancer: Aim #1: Decipher the mechanism of mitostatin-evoked mitophagy and the connection to suppressed breast cancer growth and migration Aim #2: Utilize novel mouse models of mammary tumorigenesis to investigate the in vivo role of mitostatin-driven mitophagy and suppression of growth. These innovative studies will greatly augment our understanding of the mechanisms governing mitostatin to evoke mitophagy as a new way to combat breast cancer proliferation and migration. The short-term impact will be to provide proof-of-concept and feasibility that targeting mitochondria vis-à-vis mitostatin is a valid approach. The long-term impact will be to develop novel therapeutic strategies, based on mitostatin evoked mitophagy, to effectively stop breast cancer and improve the lives of patients.