Identification of a Novel Sulfonamide Non-Nucleoside Reverse Transcriptase Inhibitor by a Phenotypic HIV-1 Full Replication Assay

<div><p>Classical target-based, high-throughput screening has been useful for the identification of inhibitors for known molecular mechanisms involved in the HIV life cycle. In this study, the development of a cell-based assay that uses a phenotypic drug discovery approach based on automated high-content screening is described. Using this screening approach, the antiviral activity of 26,500 small molecules from a relevant chemical scaffold library was evaluated. Among the selected hits, one sulfonamide compound showed strong anti-HIV activity against wild-type and clinically relevant multidrug resistant HIV strains. The biochemical inhibition, point resistance mutations and the activity of structural analogs allowed us to understand the mode of action and propose a binding model for this compound with HIV-1 reverse transcriptase.</p></div

Discovery of Novel <i>N</i>-Phenylphenoxyacetamide Derivatives as EthR Inhibitors and Ethionamide Boosters by Combining High-Throughput Screening and Synthesis

In this paper, we describe the screening of a 14640-compound library using a novel whole mycobacteria phenotypic assay to discover inhibitors of EthR, a transcriptional repressor implicated in the innate resistance of Mycobacterium tuberculosis to the second-line antituberculosis drug ethionamide. From this screening a new chemical family of EthR inhibitors bearing an <i>N</i>-phenylphenoxyacetamide motif was identified. The X-ray structure of the most potent compound crystallized with EthR inspired the synthesis of a 960-member focused library. These compounds were tested in vitro using a rapid thermal shift assay on EthR to accelerate the optimization. The best compounds were synthesized on a larger scale and confirmed as potent ethionamide boosters on M. tuberculosis-infected macrophages. Finally, the cocrystallization of the best optimized analogue with EthR revealed an unexpected reorientation of the ligand in the binding pocket

Identification of IPK1 as a potent antiretroviral hit compound.

<p>A. Chemical structure of the <b>IPK1</b> compound. B. Dose-response curve of <b>IPK1</b> and comparison to reference anti-HIV drugs. The IC₅₀ value was characterized in CEMx 174-LTR-GFP CG8 cells infected by HIV-1_LAI. C. <b>IPK1</b> activity defined by IC₅₀ characterization in HeLa-LTR-GFP cells upon HIV-1_LAI infection. D. <b>IPK1</b> activity against the multidrug resistant virus HIV-1_RTMDR/MT-2 in CEMx 174-LTR-GFP CG8.</p

HIV infection assay development.

<p>A. Measurement of GFP reporter activity, using Victor 3 and Opera reader, in CEMx 174-LTR-GFP CG8 cells upon HIV-1_LAI infection. B. Infection of CEMx 174-LTR-GFP CG8 and HeLa-LTR-GFP cells: Opera-acquired images showed infected (Panel b and d), unlike uninfected (Panel a and c) cells, carried the GFP reporter gene. Syncytia are indicated by red circles. C. Assay validation upon antiretroviral treatment: reference drugs were tested to evaluate the assay performance using an Opera reader. CEMx 174-LTR-GFP CG8 cells infected by HIV-1_LAI were treated with AZT (Panel b), nevirapine (Panel c) and saquinavir (Panel d). Images show nucleus detection in red and infected cells in green (Panel a: DMSO).</p

Table of calculated docking energy and IC₅₀ characterization of the IPK1 compound and analogs.

<p>Table of calculated docking energy and IC₅₀ characterization of the IPK1 compound and analogs.</p

IC₅₀ characterization of IPK1 and TMC125.

<p>Antiviral activity of <b>IPK1</b> and TMC125 compounds against defined reverse transcriptase resistant mutations using the Monogram Bioscience experimental assay.</p

Anti-reverse transcriptase activity of the IPK1 compound and nevirapine.

<p>The IC₅₀ value was determined <i>in vitro</i> using purified enzyme.</p

Triazolopyridines as Selective JAK1 Inhibitors: From Hit Identification to GLPG0634

Janus kinases (JAK1, JAK2, JAK3, and TYK2) are involved in the signaling of multiple cytokines important in cellular function. Blockade of the JAK-STAT pathway with a small molecule has been shown to provide therapeutic immunomodulation. Having identified JAK1 as a possible new target for arthritis at Galapagos, the compound library was screened against JAK1, resulting in the identification of a triazolopyridine-based series of inhibitors represented by <b>3</b>. Optimization within this chemical series led to identification of GLPG0634 (<b>65</b>, filgotinib), a selective JAK1 inhibitor currently in phase 2B development for RA and phase 2A development for Crohn’s disease (CD)

Discovery of <i>N</i>‑(3-Carbamoyl-5,5,7,7-tetramethyl-5,7-dihydro‑4<i>H</i>‑thieno[2,3‑<i>c</i>]pyran-2-yl)‑l<i>H</i>‑pyrazole-5-carboxamide (GLPG1837), a Novel Potentiator Which Can Open Class III Mutant Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) Channels to a High Extent

Cystic fibrosis (CF) is caused by mutations in the gene for the cystic fibrosis transmembrane conductance regulator (CFTR). With the discovery of Ivacaftor and Orkambi, it has been shown that CFTR function can be partially restored by administering one or more small molecules. These molecules aim at either enhancing the amount of CFTR on the cell surface (correctors) or at improving the gating function of the CFTR channel (potentiators). Here we describe the discovery of a novel potentiator GLPG1837, which shows enhanced efficacy on CFTR mutants harboring class III mutations compared to Ivacaftor, the first marketed potentiator. The optimization of potency, efficacy, and pharmacokinetic profile will be described

Ethionamide Boosters. 2. Combining Bioisosteric Replacement and Structure-Based Drug Design To Solve Pharmacokinetic Issues in a Series of Potent 1,2,4-Oxadiazole EthR Inhibitors

Mycobacterial transcriptional repressor EthR controls the expression of EthA, the bacterial monooxygenase activating ethionamide, and is thus largely responsible for the low sensitivity of the human pathogen <i>Mycobacterium tuberculosis</i> to this antibiotic. We recently reported structure–activity relationships of a series of 1,2,4-oxadiazole EthR inhibitors leading to the discovery of potent ethionamide boosters. Despite high metabolic stability, pharmacokinetic evaluation revealed poor mice exposure; therefore, a second phase of optimization was required. Herein a structure–property relationship study is reported according to the replacement of the two aromatic heterocycles: 2-thienyl and 1,2,4-oxadiazolyl moieties. This work was done using a combination of structure-based drug design and in vitro/ex vivo evaluations of ethionamide boosters on the targeted protein EthR and on the human pathogen <i>Mycobacterium tuberculosis</i>. Thanks to this process, we identified compound <b>42</b> (BDM41906), which displays improved efficacy in addition to high exposure to mice after oral administration