Combining Cell Envelope Stress Reporter Assays in a Screening Approach to Identify BAM Complex Inhibitors.

The development of new antibiotics is particularly problematic in Gram-negative bacteria due to the presence of the outer membrane (OM), which serves as a permeability barrier. Recently, the ß-barrel assembly machine (BAM), located in the OM and responsible for ß-barrel type OM protein (OMP) assembly, has been validated as a novel target for antibiotics. Here, we identified potential BAM complex inhibitors using a screening approach that reports on cell envelope σE and Rcs stress in Escherichia coli. Screening a library consisting of 316 953 compounds yielded five compounds that induced σE and Rcs stress responses, while not inducing the intracellular heat-shock response. Two of the five compounds (compounds 2 and 14) showed the characteristics of known BAM complex inhibitors: synergy with OMP biogenesis mutants, decrease in the abundance of various OMPs, and loss of OM integrity. Importantly, compound 2 also inhibited BAM-dependent OMP folding in an in vitro refolding assay using purified BAM complex reconstituted in proteoliposomes.

Discovery of GLPG2451, a Novel Once Daily Potentiator for the Treatment of Cystic Fibrosis.

Cystic fibrosis (CF) is a life-threatening recessive genetic disease caused by mutations in the gene encoding for the cystic fibrosis transmembrane conductance regulator (CFTR). With the discovery of Ivacaftor and Lumacaftor, it has been shown that administration of one or more small molecules can partially restore the CFTR function. Correctors are small molecules that enhance the amount of CFTR on the cell surface, while potentiators improve the gating function of the CFTR channel. Herein, we describe the discovery and optimization of a novel potentiator series. Scaffold hopping, focusing on retaining the different intramolecular contacts, was crucial in the whole discovery process to identify a novel series devoid of genotoxic liabilities. From this series, the clinical candidate GLPG2451 was selected based on its pharmacokinetic properties, allowing QD dosing and based on its low CYP induction potential.