The prognosis for women with breast tumors that are estrogen receptor (ER)-negative is very poor because these tumors are aggressive and do not respond to standard treatments. This situation emphasizes the importance of developing new agents and protocols for preventing breast cancer. We address this gap of knowledge by examining the effect of Nexrutine, an herbal extract from the Phellodendron amurense plant, on breast cancer incidence and latency with the anticipation of developing this compound as a chemoprevention agent. In support of this study, we found that Nexrutine reduced excessive branching of mammary gland terminal end buds from HER-2/neu transgenic mice and decreased PGE2 production in the mammary glands. These findings suggest that Nexrutine may reduce risk of mammary tumor incidence by altering the early stages of development. Nexrutine also decreased survival of breast cancer cells by inducing cell death, which was associated with autophagy. The role of autophagy in cancer is unclear. This study offers an opportunity to investigate whether autophagy is involved in the prevention of breast cancer. Based upon this preliminary information, we hypothesize that the use of Nexrutine could provide a novel approach to reduce incidence and delay the progression of spontaneous mammary tumors. Additionally, we propose that the protective effects of Nexrutine are associated with induction of autophagy and cell death. We will test this hypothesis by completing the following: AIMS: 1. To determine whether Nexrutine will reduce incidence and multiplicity, and increase tumor latency of spontaneous mammary tumors in HER-2/neu transgenic mice. 2. To investigate mechanisms of Nexrutine-induced autophagic death in tumor cells and tumors from HER-2/neu transgenic mice by defining the Nexrutine-induced autophagic response and determining the role of ERK and Akt/mTOR, and the contribution of reactive oxygen species (ROS) in mediating increase in autophagy in response to Nexrutine treatment. The positive outcome of this preclinical study along with our mechanistic design will provide a strong rationale for establishing a clinical trial to determine the efficacy of Nexrutine as a prevention strategy against breast cancer. The importance of this study resides in the observation that Nexrutine may reduce early biomarkers of breast cancer risk. Completion of the proposed study has high potential of identifying additional biomarkers and signaling pathways to design effective prevention protocols. PUBLIC HEALTH RELEVANCE: Nexrutine is a botanical extract from the Phellodendron amurense tree that is native to Northern China and is one of the top 50 herbs used in traditional Chinese medicine for its anti-inflammatory properities. We have very compelling preliminary data showing that Nexrutine decreases growth of mammary gland terminal end buds and the biomarker, prostaglandin E2 in the HER-2/neu mouse model. Based upon our data showing that Nexrutine has bioavailability, we propose to evaluate the potential of Nexrutine for prevention of precancerous lesions and mammary tumors in a clinically relevant transgenic mouse model. Additionally, our preliminary findings demonstrate that Nexrutine induces autophagy and cell death. Since the role of autophagy in cancer is unclear, we will evaluate whether autophagy is associated with decreased development and incidence of mammary tumors. We also plan to determine mechanisms by which Nexrutine is inducing autophagy by interrogating the role of ERK and Akt/mTOR signaling and reactive oxygen species (ROS) in mediating these effects.