Shuhao Shi

Inhibition of influenza virus replication via small molecules that induce the formation of higher-order nucleoprotein oligomers

Influenza nucleoprotein (NP) plays multiple roles in the virus life cycle, including an essential function in viral replication as an integral component of the ribonucleoprotein complex, associating with viral RNA and polymerase within the viral core. The multifunctional nature of NP makes it an attractive target for antiviral intervention, and inhibitors targeting this protein have recently been reported. In a parallel effort, we discovered a structurally similar series of influenza replication inhibitors and show that they interfere with NP-dependent processes via formation of higher-order NP oligomers. Support for this unique mechanism is provided by site-directed mutagenesis studies, biophysical characterization of the oligomeric ligand:NP complex, and an X-ray cocrystal structure of an NP dimer of trimers (or hexamer) comprising three NP_A:NP_B dimeric subunits. Each NP_A:NP_B dimeric subunit contains two ligands that bridge two composite, protein-spanning binding sites in an antiparallel orientation to form a stable quaternary complex. Optimization of the initial screening hit produced an analog that protects mice from influenza-induced weight loss and mortality by reducing viral titers to undetectable levels throughout the course of treatment.

Acyl guanidine inhibitors of β-secretase (BACE-1): optimization of a micromolar hit to a nanomolar lead via iterative solid- and solution-phase library synthesis.

This report describes the discovery and optimization of a BACE-1 inhibitor series containing an unusual acyl guanidine chemotype that was originally synthesized as part of a 6041-membered solid-phase library. The synthesis of multiple follow-up solid- and solution-phase libraries facilitated the optimization of the original micromolar hit into a single-digit nanomolar BACE-1 inhibitor in both radioligand binding and cell-based functional assay formats. The X-ray structure of representative inhibitors bound to BACE-1 revealed a number of key ligand:protein interactions, including a hydrogen bond between the side chain amide of flap residue Gln73 and the acyl guanidine carbonyl group, and a cation-π interaction between Arg235 and the isothiazole 4-methoxyphenyl substituent. Following subcutaneous administration in rats, an acyl guanidine inhibitor with single-digit nanomolar activity in cells afforded good plasma exposures and a dose-dependent reduction in plasma Aβ levels, but poor brain exposure was observed (likely due to Pgp-mediated efflux), and significant reductions in brain Aβ levels were not obtained.

Discovery and optimization of (R)-prolinol-derived agonists of the Growth Hormone Secretagogue receptor (GHSR).

The discovery and optimization of a novel series of prolinol-derived GHSR agonists is described. This series emerged from a 11,520-member solid-phase library targeting the GPCR protein superfamily, and the rapid optimization of low micromolar hits into single-digit nanomolar leads can be attributed to the solid-phase synthesis of matrix libraries, which revealed multiple non-additive structure-activity relationships. In addition, the separation of potent diastereomers highlighted the influence of the alpha-methyl stereochemistry of the phenoxyacetamide sidechain on GHSR activity.

Solid-phase synthesis of a library based on biphenyl-containing trypsin-like serine protease inhibitors

The solid-phase synthesis of a library based on an unusual biphenyl-containing trypsin-like serine protease inhibitor is described. Key to this effort was the synthesis of a highly functionalized aryl boronic acid reagent which required the development of a novel and efficient method to convert a triflate to a pinacolboronate in large scale.