Examining the Feasibility of Using Free Energy Perturbation (FEP+) in Predicting Protein Stability

The importance of engineering protein stability is well-known and has the potential to impact many fields ranging from pharmaceuticals to food sciences. Engineering proteins can be both a time-consuming and expensive experimental process. The use of computation is a potential solution to mitigating some of the time and expenses required to engineer a protein. This process has been previously hindered by inaccurate force fields or energy equations and slow computational processors; however, improved software and hardware have made this goal much more attainable. Here we find that Schrödinger’s new FEP+, although still imperfect, proves more successful in predicting protein stability than other simpler methods of investigation. This increased accuracy comes at a cost of computational time and resources when compared to simpler methods. This work adds to the initial testing of FEP+ by offering options for more accurately predicting protein stability in an efficient manner

Novel inhibitors of an emerging target in Mycobacterium tuberculosis; substituted thiazolidinones as inhibitors of dTDP-rhamnose synthesis

The emergence of multi-drug resistant tuberculosis, coupled with the increasing overlap of the AIDS and tuberculosis pandemics has brought tuberculosis to the forefront as a major worldwide health concern. In an attempt to find new inhibitors of the enzymes in the essential rhamnose biosynthetic pathway, a virtual library of 2,3,5 trisubstituted-4-thiazolidinones was created. These compounds were then docked into the active site cavity of 6'hydroxyl; dTDP-6-deoxy-D-xylo-4-hexulose 3,5-epimerase (RmlC) from Mycobacterium tuberculosis. The resulting docked conformations were consensus scored and the top 5% were slated for synthesis. Thus far, 94 compounds have been successfully synthesized and initially tested. Of those, 30 (32%) have > or =50% inhibitory activity (at 20 microM) in the coupled rhamnose synthetic assay with seven of the 30 also having modest activity against whole-cell M. tuberculosis

Comprehensive Mechanistic Analysis of Hits from High-Throughput and Docking Screens against beta-Lactamase.

High-throughput screening (HTS) is widely used in drug discovery. Especially for screens of unbiased libraries, false positives can dominate “hit lists”; their origins are much debated. Here we determine the mechanism of every active hit from a screen of 70,563 unbiased molecules against β-lactamase using quantitative HTS (qHTS). Of the 1274 initial inhibitors, 95% were detergent-sensitive and were classified as aggregators. Among the 70 remaining were 25 potent, covalent-acting β-lactams. Mass spectra, counter-screens, and crystallography identified 12 as promiscuous covalent inhibitors. The remaining 33 were either aggregators or irreproducible. No specific reversible inhibitors were found. We turned to molecular docking to prioritize molecules from the same library for testing at higher concentrations. Of 16 tested, 2 were modest inhibitors. Subsequent X-ray structures corresponded to the docking prediction. Analog synthesis improved affinity to 8 µM. These results suggest that it may be the physical behavior of organic molecules, not their reactivity, that accounts for most screening artifacts. Structure-based methods may prioritize weak-but-novel chemotypes in unbiased library screens

Discovery of an Oral Respiratory Syncytial Virus (RSV) Fusion Inhibitor (GS-5806) and Clinical Proof of Concept in a Human RSV Challenge Study

GS-5806 is a novel, orally bioavailable RSV fusion inhibitor discovered following a lead optimization campaign on a screening hit. The oral absorption properties were optimized by converting to the pyrazolo­[1,5-<i>a</i>]-pyrimidine heterocycle, while potency, metabolic, and physicochemical properties were optimized by introducing the <i>para</i>-chloro and aminopyrrolidine groups. A mean EC₅₀ = 0.43 nM was found toward a panel of 75 RSV A and B clinical isolates and dose-dependent antiviral efficacy in the cotton rat model of RSV infection. Oral bioavailability in preclinical species ranged from 46 to 100%, with evidence of efficient penetration into lung tissue. In healthy human volunteers experimentally infected with RSV, a potent antiviral effect was observed with a mean 4.2 log₁₀ reduction in peak viral load and a significant reduction in disease severity compared to placebo. In conclusion, a potent, once daily, oral RSV fusion inhibitor with the potential to treat RSV infection in infants and adults is reported

Discovery of 6‑(Fluoro-¹⁸<i>F</i>)‑3-(1<i>H</i>‑pyrrolo[2,3‑<i>c</i>]pyridin-1-yl)isoquinolin-5-amine ([¹⁸F]-MK-6240): A Positron Emission Tomography (PET) Imaging Agent for Quantification of Neurofibrillary Tangles (NFTs)

Neurofibrillary tangles (NFTs) made up of aggregated tau protein have been identified as the pathologic hallmark of several neurodegenerative diseases including Alzheimer’s disease. In vivo detection of NFTs using PET imaging represents a unique opportunity to develop a pharmacodynamic tool to accelerate the discovery of new disease modifying therapeutics targeting tau pathology. Herein, we present the discovery of 6-(fluoro-¹⁸<i>F</i>)-3-(1<i>H</i>-pyrrolo­[2,3-<i>c</i>]­pyridin-1-yl)­isoquinolin-5-amine, <b>6</b> ([¹⁸F]-MK-6240), as a novel PET tracer for detecting NFTs. <b>6</b> exhibits high specificity and selectivity for binding to NFTs, with suitable physicochemical properties and in vivo pharmacokinetics