We have found that certain tetrasubstituted pyrazoles are high-affinity ligands for the estrogen receptor (ER) (Fink et al. Chem. Biol. 1999, 6, 205-219) and that one pyrazole is considerably more potent as an agonist on the ERalpha than on the ERbeta subtype (Sun et al. Endocrinology 1999, 140, 800-804). To investigate what substituent pattern provides optimal ER binding affinity and the greatest enhancement of potency as an ERalpha-selective agonist, we prepared a number of tetrasubstituted pyrazole analogues with defined variations at certain substituent positions. Analysis of their binding affinity pattern shows that a C(4)-propyl substituent is optimal and that a p-hydroxyl group on the N(1)-phenyl group also enhances affinity and selectivity for ERalpha. The best compound in this series, a propylpyrazole triol (PPT, compound 4g), binds to ERalpha with high affinity (ca. 50% that of estradiol), and it has a 410-fold binding affinity preference for ERalpha. It also activates gene transcription only through ERalpha. Thus, this compound represents the first ERalpha-specific agonist. We investigated the molecular basis for the exceptional ERalpha binding affinity and potency selectivity of pyrazole 4g by a further study of structure-affinity relationships in this series and by molecular modeling. These investigations suggest that the pyrazole triols prefer to bind to ERalpha with their C(3)-phenol in the estradiol A-ring binding pocket and that binding selectivity results from differences in the interaction of the pyrazole core and C(4)-propyl group with portions of the receptor where ERalpha has a smaller residue than ERbeta. These ER subtype-specific interactions and the ER subtype-selective ligands that can be derived from them should prove useful in defining those biological activities in estrogen target cells that can be selectively activated through ERalpha.

A high-throughput screen of the NIH molecular libraries sample collection and subsequent optimization of a lead dipeptide-like series of severe acute respiratory syndrome (SARS) main protease (3CLpro) inhibitors led to the identification of probe compound ML188 (16-(R), (R)-N-(4-(tert-butyl)phenyl)-N-(2-(tert-butylamino)-2-oxo-1-(pyridin-3-yl)ethyl)furan-2-carboxamide, Pubchem CID: 46897844). Unlike the majority of reported coronavirus 3CLpro inhibitors that act via covalent modification of the enzyme, 16-(R) is a non-covalent SARS-CoV 3CLpro inhibitor with moderate MW and good enzyme and antiviral inhibitory activity. A multi-component Ugi reaction was utilized to rapidly explore structure activity relationships within S1′, S1, and S2 enzyme binding pockets. The X-ray structure of SARS-CoV 3CLpro bound with 16-(R) was instrumental in guiding subsequent rounds of chemistry optimization. 16-(R) provides an excellent starting point for the further design and refinement of 3CLpro inhibitors that act by a non-covalent mechanism of action.

Modulators of metabotropic glutamate receptor subtype 5 (mGluR5) may provide novel treatments for multiple central nervous system (CNS) disorders, including anxiety and schizophrenia. Although compounds have been developed to better understand the physiological roles of mGluR5 and potential usefulness for the treatment of these disorders, there are limitations in the tools available, including poor selectivity, low potency, and limited solubility. To address these issues, we developed an innovative assay that allows simultaneous screening for mGluR5 agonists, antagonists, and potentiators. We identified multiple scaffolds that possess diverse modes of activity at mGluR5, including both positive and negative allosteric modulators (PAMs and NAMs, respectively). 3-Fluoro-5-(3-(pyridine-2-yl)-1,2,4-oxadiazol-5-yl)benzonitrile (VU0285683) was developed as a novel selective mGluR5 NAM with high affinity for the 2-methyl-6-(phenylethynyl)-pyridine (MPEP) binding site. VU0285683 had anxiolytic-like activity in two rodent models for anxiety but did not potentiate phencyclidine-induced hyperlocomotor activity. (4-Hydroxypiperidin-1-yl)(4-phenylethynyl)phenyl)methanone (VU0092273) was identified as a novel mGluR5 PAM that also binds to the MPEP site. VU0092273 was chemically optimized to an orally active analog, N-cyclobutyl-6-((3-fluorophenyl)ethynyl)nicotinamide hydrochloride (VU0360172), which is selective for mGluR5. This novel mGluR5 PAM produced a dose-dependent reversal of amphetamine-induced hyperlocomotion, a rodent model predictive of antipsychotic activity. Discovery of structurally and functionally diverse allosteric modulators of mGluR5 that demonstrate in vivo efficacy in rodent models of anxiety and antipsychotic activity provide further support for the tremendous diversity of chemical scaffolds and modes of efficacy of mGluR5 ligands. In addition, these studies provide strong support for the hypothesis that multiple structurally distinct mGluR5 modulators have robust activity in animal models that predict efficacy in the treatment of CNS disorders.