The long-term goals of this project are to understand the regulation and role of steroid hormone receptors in cancer development and therapeutics, through our development of a new technology that allows for the rapid analysis of steroid hormone receptors using X-ray crystallography. Specifically, our approach has demonstrated that we can increase both the rate and numbers of hits with crystal structures by at least a thousand fold, and that this allows rapid analysis of the ligand-binding domain of these receptors bound to chemotherapy agents, and pathway selective compounds. The estrogen and androgen receptors (ER and AR) are implicated in the development, diagnosis, and treatment for breast and prostate cancer, respectively. Glucocorticoids have a broader role, as up-front therapeutics for the treatment of several malignancies (e.g., leukemia and hormone-refractory prostate cancer), and as adjuvants that reduce the side effects of other chemotherapy agents. The synthetic compounds that target these receptors have, however, significant problems, including acquired resistance and undesirable side effects. They also display tissue and pathway selective signaling that is poorly understood, at both the molecular and structural level. It is possible to develop tissue and pathway selective compounds that ameliorate some of these problems, but there is very little understanding of the structural basis for such selectivity. The lack of good structural models for tissue selectivity is due to the difficulty in producing crystal structures. The steroid receptor ligand-binding domain (LBD) has proven very difficult to crystallize, due to conformational heterogeneity and protein misfolding. Here we propose to further develop our new technology for molecular analyses of steroid receptors, which we strongly believe will revolutionize the use of X-ray crystallography in both basic research and drug discovery, especially regarding steroid receptors. Specifically, we have identified and generated a series of surface mutations that stabilize the estrogen receptor in the conformations seen with both agonist and antagonist ligands. This advance has allowed us to add compounds in parallel to the purified protein, and to obtain the first structure of an apo steroid receptor LBD. We propose to apply these techniques to apply this high-throughput technology to other steroid receptors implicated in cancer, and to use this approach to define the structural basis through which the glucocorticoid receptor (GR) inhibits the NF-?B oncogenic pathway. We believe that these studies will establish new and robust techniques that will revolutionize the use of X-ray crystallography in defining how small molecules control tissue- and pathway-selective signaling through steroid hormone receptors. This """"""""class analysis"""""""" approach to studying groups of structures is highly novel, and allows for the incorporation of statistical power into structural analysis. Importantly, this approach will also directly impact the drug discovery process, by rapidly providing structural information that will guide the development of new therapeutics.

National Institute of Health (NIH)
National Cancer Institute (NCI)
Exploratory/Developmental Grants Phase II (R33)
Project #
Application #
Study Section
Special Emphasis Panel (ZCA1-SRLB-Q (M1))
Program Officer
Knowlton, John R
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
Scripps Research Institute
La Jolla
United States
Zip Code
Zhao, Yuechao; Laws, Mary J; Guillen, Valeria Sanabria et al. (2017) Structurally Novel Antiestrogens Elicit Differential Responses from Constitutively Active Mutant Estrogen Receptors in Breast Cancer Cells and Tumors. Cancer Res 77:5602-5613
Min, Jian; Guillen, Valeria Sanabria; Sharma, Abhishek et al. (2017) Adamantyl Antiestrogens with Novel Side Chains Reveal a Spectrum of Activities in Suppressing Estrogen Receptor Mediated Activities in Breast Cancer Cells. J Med Chem 60:6321-6336
Srinivasan, Sathish; Nwachukwu, Jerome C; Bruno, Nelson E et al. (2017) Full antagonism of the estrogen receptor without a prototypical ligand side chain. Nat Chem Biol 13:111-118
Nwachukwu, Jerome C; Srinivasan, Sathish; Zheng, Yangfan et al. (2016) Predictive features of ligand-specific signaling through the estrogen receptor. Mol Syst Biol 12:864
Kojetin, Douglas J; Matta-Camacho, Edna; Hughes, Travis S et al. (2015) Structural mechanism for signal transduction in RXR nuclear receptor heterodimers. Nat Commun 6:8013
Nwachukwu, Jerome C; Srinivasan, Sathish; Bruno, Nelson E et al. (2014) Resveratrol modulates the inflammatory response via an estrogen receptor-signal integration network. Elife 3:e02057
Nwachukwu, Jerome C; Southern, Mark R; Kiefer, James R et al. (2013) Improved crystallographic structures using extensive combinatorial refinement. Structure 21:1923-30
Srinivasan, Sathish; Nwachukwu, Jerome C; Parent, Alex A et al. (2013) Ligand-binding dynamics rewire cellular signaling via estrogen receptor-*. Nat Chem Biol 9:326-32
Bruning, John B; Parent, Alexander A; Gil, German et al. (2010) Coupling of receptor conformation and ligand orientation determine graded activity. Nat Chem Biol 6:837-43