Project Summary Among men, prostate cancer is the most prevalent form of cancer and the third most common cause of cancer- related death. While patients with localized disease have a positive prognosis, those with metastatic disease have a five-year survival rate of 30%. The main therapeutic target in prostate cancer is the androgen receptor (AR). Most patients treated with anti-androgens will develop resistance and continue disease progression to metastatic castration resistant prostate cancer (CRPC). Second-generation anti-androgens, including enzalutamide and abiraterone, extend life expectancy, but eventually result in resistance to the treatment. These findings highlight the need for new treatment approaches necessary to overcome or limit the development of resistance. As many as 20% of aggressive prostate cancers have mutations resulting in upregulation of Wnt/β-catenin signaling. β-catenin is a transcription factor that is activated in response to Wnt pathway signaling. To cause changes in gene transcription β-catenin interacts with the DNA binding protein called TCF. Importantly, recent analysis identified the Wnt/β-catenin pathway as the foremost differentially modulated pathway among enzalutamide-resistant individuals. To target the Wnt/β-catenin pathway, we used structure-based design of oligomeric macrocycles to identify new molecules that potently inhibit Wnt/β-catenin signaling and the growth of prostate cancer cells. Our goal is to evaluate the ability of macrocycle 13, the lead inhibitor identified in our screen, to suppress β-catenin signaling in castration resistant prostate cancer using in vivo models of prostate cancer. We will also determine the impact of β-catenin inhibition on immune cells.