Inhibiting androgen receptor nuclear entry in castration-resistant prostate cancer.

Clinical resistance to the second-generation antiandrogen enzalutamide in castration-resistant prostate cancer (CRPC), despite persistent androgen receptor (AR) activity in tumors, highlights an unmet medical need for next-generation antagonists. We have identified and characterized tetra-aryl cyclobutanes (CBs) as a new class of competitive AR antagonists that exhibit a unique mechanism of action. These CBs are structurally distinct from current antiandrogens (hydroxyflutamide, bicalutamide, and enzalutamide) and inhibit AR-mediated gene expression, cell proliferation, and tumor growth in several models of CRPC. Conformational profiling revealed that CBs stabilize an AR conformation resembling an unliganded receptor. Using a variety of techniques, it was determined that the AR-CB complex was not recruited to AR-regulated promoters and, like apo AR, remains sequestered in the cytoplasm, bound to heat shock proteins. Thus, we have identified third-generation AR antagonists whose unique mechanism of action suggests that they may have therapeutic potential in CRPC.

π-π interaction energies as determinants of the photodimerization of mono-, di-, and triazastilbenes.

We describe the quantitative [2 + 2] photocycloaddition of crystalline trans-2,4-dichloro-6-styrylpyrimidine to produce the corresponding htt r-ctt cyclobutane dimer, and we present (1)H NMR analysis of the photolysis of this and six other mono-, di-, and triazastilbenes in solid and solution states. Density functional (M06-2X) and correlated ab initio (MP2) calculations were used to obtain interaction energies between two monomers of each azastilbene. These energies mirror the relative polarization of the stilbene moieties and can be quantitatively correlated with the rate of reaction and selective formation of the htt r-ctt dimers. In the solid state, poor correlation is observed between interaction energy and reactivity/selectivity. This lack of correlation is explained through X-ray analysis of the azastilbene monomers and is shown to be in accordance with the principles of Schmidt's topochemical postulate. Conversely, in solution there is a strong positive correlation (R(2) = 0.96) between interaction energies and formation of the htt r-ctt dimer. These results are the first to show this correlation and to demonstrate the utility of calculated interaction energies as a tool for the prediction of stereo- and regioselectivity in solution-state stilbene-type photocycloadditions.

Resveratrol modulates the inflammatory response via an estrogen receptor-signal integration network.

Resveratrol has beneficial effects on aging, inflammation and metabolism, which are thought to result from activation of the lysine deacetylase, sirtuin 1 (SIRT1), the cAMP pathway, or AMP-activated protein kinase. In this study, we report that resveratrol acts as a pathway-selective estrogen receptor-α (ERα) ligand to modulate the inflammatory response but not cell proliferation. A crystal structure of the ERα ligand-binding domain (LBD) as a complex with resveratrol revealed a unique perturbation of the coactivator-binding surface, consistent with an altered coregulator recruitment profile. Gene expression analyses revealed significant overlap of TNFα genes modulated by resveratrol and estradiol. Furthermore, the ability of resveratrol to suppress interleukin-6 transcription was shown to require ERα and several ERα coregulators, suggesting that ERα functions as a primary conduit for resveratrol activity.DOI: http://dx.doi.org/10.7554/eLife.02057.001.

Coupling of receptor conformation and ligand orientation determine graded activity.

Small molecules stabilize specific protein conformations from a larger ensemble, enabling molecular switches that control diverse cellular functions. We show here that the converse also holds true: the conformational state of the estrogen receptor can direct distinct orientations of the bound ligand. 'Gain-of-allostery' mutations that mimic the effects of ligand in driving protein conformation allowed crystallization of the partial agonist ligand WAY-169916 with both the canonical active and inactive conformations of the estrogen receptor. The intermediate transcriptional activity induced by WAY-169916 is associated with the ligand binding differently to the active and inactive conformations of the receptor. Analyses of a series of chemical derivatives demonstrated that altering the ensemble of ligand binding orientations changes signaling output. The coupling of different ligand binding orientations to distinct active and inactive protein conformations defines a new mechanism for titrating allosteric signaling activity.

Alternative inhibition of androgen receptor signaling: peptidomimetic pyrimidines as direct androgen receptor/coactivator disruptors.

Compounds that directly disrupt the androgen receptor/steroid receptor coactivator interaction could function as novel inhibitors of androgen signaling that would remain effective in the treatment of prostate cancer that is resistant to conventional endocrine therapies. A structure-based peptidomimetic approach was used to design and synthesize such compounds, based on a pyrimidine-core system. Using fluorescence resonance energy transfer and reporter gene assays, we identified members of this library that disrupt the androgen receptor/steroid receptor coactivator interaction selectively, without affecting the estrogen receptor/steroid receptor coactivator interaction. Unlike the activity of traditional androgen receptor antagonists, such as flutamide and bicalutamide, inhibition by these coactivator binding inhibitors is insurmountable by increased concentrations of androgen agonists and maintains effectiveness even on a mutant androgen receptor that is resistant to traditional antagonists. These findings support the feasibility of targeting the coactivator binding groove of the androgen receptor as an alternative approach to treatment-resistant prostate cancer therapy.

Blocking estrogen signaling after the hormone: pyrimidine-core inhibitors of estrogen receptor-coactivator binding.

As an alternative approach to blocking estrogen action, we have developed small molecules that directly disrupt the key estrogen receptor (ER)/coactivator interaction necessary for gene activation. The more direct, protein-protein nature of this disruption might be effective even in hormone-refractory breast cancer. We have synthesized a pyrimidine-core library of moderate size, members of which act as alpha-helix mimics to block the ERalpha/coactivator interaction. Structure-activity relationships have been explored with various C-, N-, O-, and S-substituents on the pyrimidine core. Time-resolved fluorescence resonance energy transfer and cell-based reporter gene assays show that the most active members inhibit the ERalpha/steroid receptor coactivator interaction with K i's in the low micromolar range. Through these studies, we have obtained a refined pharmacophore model for activity in this pyrimidine series. Furthermore, the favorable activities of several of these compounds support the feasibility that this coactivator binding inhibition mechanism for blocking estrogen action might provide a potential alternative approach to endocrine therapy.

Mechanistic analysis of the photocycloaddition of silyl-tethered alkenes.

The photochemistry of substituted cinnamyloxy silanes has been examined in both cyclohexane and acetonitrile solvents. Alkene isomerization occurs in addition to cycloaddition. Fluorescence quantum yields and excited singlet state lifetimes have been determined for each compound. We have used the information in order to better understand the regio- and stereoselectivity of photocycloaddition between silyl-tethered cinnamyl groups. This study allows us to conclude that the 2 + 2 photocycloaddition between alkenes is not a Woodward-Hoffmann orbital symmetry controlled event. The most consistent explanation for the excellent regio- and stereoselectivity is that the photocycloaddition is conformationally controlled.