Abstract Tamoxifen, an antagonist at estrogen receptor alpha (ERα) in breast tissue, and the prototypical selective estrogen receptor modulator (SERM), is the standard of care for many patients with ER-positive breast cancer. However, continued tamoxifen treatment increases the risk of endometrial cancer and half of ER-positive breast cancer patients do not respond or relapse on treatment with tamoxifen. Tamoxifen-resistant tumors are often ER-positive and endocrine-independent and therefore resistant to aromatase inhibitors. Recent clinical trials have shown the efficacy of estrogen in patients who have undergone exhaustive tamoxifen therapy. Therefore, we propose the development of small molecules that mimic the actions of estradiol in ER-positive breast cancer without the uterotrophic actions of estradiol and tamoxifen: Selective Estrogen Mimics (SEMs). Evidence suggest that a significant subset of ER-positive breast cancer, over-expressing PKCα, and tamoxifen-resistant cancers will respond to SEMs. In order to design and optimize SEMs, tamoxifen-resistant cell lines, MCF-7:5C, T47D: PKCα and T47D: Tam1 were studied. The activation of classical ERα signaling by the estrogen mimics was profiled in MCF-7 cells and cell viability in 2D culture was examined. In an effort to increase throughput and mimic the in vivoenvironment more closely, we assessed the SEMs in a new 3D spheroid model. It was found that in MCF-7:5C cells in 2D and T47D spheroids that SEM induced cell death was mediated by activation of ERα. SEMs caused decreased spheroid viability and growth in all three models of tamoxifen resistance. Extranuclear localization of ER after SEM treatment previously reported in matrigel assays and in vivo xenografts was also observed in spheroids. This emphasizes the capability of the 3D spheroids to replicate in vivo environments, making them a high-throughput translational assay of choice for drug discovery. Three promising SEMs were evaluated in xenograft models of TAM-resistant, PKCα overexpressing breast cancer. While E2 caused regression of these tumors, a significant increase in uterine weight as predicted was observed. More importantly, SEM treated mice had negligible increase in uterine weight underlining the enhanced safety of these molecules. These data suggest that development of SEMs that retain the beneficial properties of estrogen while limiting the side effects is a feasible strategy for the treatment of tamoxifen resistant breast cancer. Citation Format: Hitisha Patel, Rui Xiong, Lauren Gutgesell, Jiong Zhao, Mary Ellen Molloy, Debra Tonetti, Gregory R.J Thatcher. Selective estrogen mimics (SEMs) for the treatment of tamoxifen resistant breast cancer. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr C98.