Discovering coactivator proteomimetic inhibitors

Nuclear receptor (NR) super-family of signaling proteins is responsible for the integration of physiological signals to maintain the dynamic homeostasis of an organism by regulating growth, development, and differentiation. This signaling network is composed of a series of highly conserved and mutually shared protein-protein interactions that permit the formation and disassembly of macromolecular protein transcriptional regulatory complexes composed of NR, coactivators, corepressors and transcription factors. Specific direct competitive inhibitors of these protein-protein interactions would aid in our understanding of the specific role of each of these interactions in the regulatory complex formation. We chose to target the coactivator interaction with the NR which has been identified to be a critical step in the activation complex formation following ligand binding. Coactivators bind receptors in a ligand dependent manner via the highly conserved alpha-helix LXXLL NR box motif. By mimicking this protein interface we have rationally designed chemical probes that are specific to different NR which inhibit coactivator binding despite the shared binding of the LXXLL NR box.; Here we present the design, synthesis, evaluation and selection of a series of inhibitors that are specific to human Estrogen Receptor Alpha (hERalpha), human Estrogen Receptor Beta (hERbeta) and human Thyroid Hormone Receptor Beta (hTRbeta). Additionally we have identified a set of inhibitors that are able to discriminate between three estrogens, 17beta-estradiol, genistein (Gen), and an estrogen analogue diethylstilbeserol (DES), with both ERalpha and ERbeta. These inhibitors have identified sub-site specific differences between several NR and their liganded states that have not been identified through traditional biochemical methods. Our 2.3 A resolution crystal structures of these inhibitors bound to ERalpha are providing insights behind the selectivity between receptors at this interface which are generating a new framework for drug design at the protein-protein interactions. We hope to utilize these probes to understand the role of this interaction in NR gene regulation and to investigate the therapeutic potential of targeting this interface in NR related disorders such as breast cancer. We also believe these results will play a seminal role in the identification of novel approaches to therapeutic intervention by characterizing protein interfaces in a variety of signaling networks for target validation and specific inhibitor design.