Abstract Breast cancer accounts for one of the most common causes of cancer-related deaths in women. Despite the available treatment modalities to cure breast cancer, therapeutic resistance and recurrence remain major challenges, owing to the culminating evidence of the presence of cancer stem cell subpopulations in breast tumors. The purpose of the study is to identify chemopreventive and dietary compounds that effectively target cancer stem cells to inhibit breast cancer development and progression. The mammosphere culture system has been proven to enrich cancer stem-like cells, and cells from mammospheres are shown to be more tumorigenic in vivo. By culturing mammospheres from breast cancer cell lines, we have shown that 1,25 vitamin D3, a vitamin D3 analog, BXL0124, pterostilbene (trans-3,5-dimethoxy-4-hydroxystilbene), tocopherols and triterpenoids (CDDO-Im) decreased the mammosphere forming efficiency (MFE) in a dose-dependent manner. Vitamin D compounds (1,25D3 and BXL0124) decreased the MFE in SUM159, MCF10DCIS and MCF-7 breast cancer cell lines (dose range 1-100 nM). Pterostilbene, a bioactive component of blueberries, effectively decreased the MFE in the MCF-7 cell line. The size of the mammospheres was found to be smaller with increasing concentrations (10-100 μM). Tocopherols significantly decreased estrogen-stimulated MFE in MCF-7 cells (1-10 μM). CDDO-Im, a synthetic triterpenoid derived from natural oleanolic acid, has also been shown to decrease MFE and sphere size in MCF-7 cells (50-100 nM). The effect on the shape of the spheres was also observed, suggesting the differentiation-inducing properties of these compounds. Taken together, we demonstrate that naturally occurring compounds as well as synthetic analogs derived from natural and dietary compounds significantly inhibit sphere forming efficiency and cancer stemness in breast cancer cells. Our study suggests that chemopreventive compounds, 1,25D3, BXL0124, pterostilbene, tocopherols, and CDDO-Im, are agents with the potential to target cancer stem-like cells to prevent breast cancer. Citation Format: Naing Lin Shan, Jeffrey Yang, Min Ji Bak, Joseph Wahler, Nanjoo Suh. Targeting cancer stem cells with chemopreventive agents for breast cancer prevention [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 276.