High-throughput cell-based assays for the preclinical development of DsbA inhibitors as antivirulence therapeutics

Abstract

21Antibiotics are failing fast, and the development pipeline is alarmingly dry. New drug research 22 and development is being urged by world health officials, with new antibacterials against 23 multidrug-resistant Gram-negative pathogens as the highest priority. Antivirulence drugs, 24 which are inhibitors of bacterial pathogenicity factors, are a class of promising antibacterials, 25 however, their development is often stifled by lack of standardised preclinical testing akin to 26 what guides antibiotic development. The lack of established target-specific microbiological 27 assays amenable to high-throughput, often means that cell-based testing of virulence inhibitors 28 is absent from the discovery (hit-to-lead) phase, only to be employed at later-stages of lead 29 optimization. Here, we address this by establishing a pipeline of bacterial cell-based assays 30 developed for the identification and early preclinical evaluation of DsbA inhibitors. Inhibitors 31 of DsbA block bacterial oxidative protein folding and were previously identified by biophysical 32 and biochemical assays. Here we use existing Escherichia coli DsbA inhibitors and 33 uropathogenic E. coli (UPEC) as a model pathogen, to demonstrate that a combination of a 34 cell-based AssT sulfotransferase assay and the UPEC motility assay, modified for a higher 35 throughput format, can provide a robust and target-specific platform for the evaluation of DsbA 36 inhibitors. Our pipeline could also be used in fragment and compound screening for the 37 identification of new DsbA inhibitor classes or hits with a broad spectrum of activity. In 38 conclusion, the establishment of accurate, high-throughput microbiological assays for 39 antivirulence drug identification and early preclinical development, is a significant first step 40 towards their translation into effective therapeutics. 41 42 3Importance: 43 The safety net of last resort antibiotics is quickly vanishing as bacteria become increasingly 44 resistant to most available drugs. If no action is taken, we will likely enter a post-antibiotic era, 45 where common infections and minor injuries are once again lethal. The paucity in new 46 antibiotic discovery of the past decades has compounded the problem of increasing antibiotic 47 resistance, to the point that it now constitutes a global health crisis that demands global action. 48There is currently an urgent need for new antibacterial drugs with new targets and modes of 49 action. To address this, research and development efforts into antivirulence drugs, such as 50 DsbA inhibitors, have been ramping up globally. However, the development of microbiological 51 assays as tools for effectively identifying and evaluating antivirulence drugs is lagging behind. 52Here, we present a high-throughput cell-based screening and evaluation pipeline, which could 53 significantly advance development of DsbA inhibitor as antivirulence therapeutics. 54 55 4Introduction: 56 In 2014 the World Health Organisation (WHO) released a statement declaring an...

DeepDDS: deep graph neural network with attention mechanism to predict synergistic drug combinations

“…Traditional and modern medicine has always taken advantage of the combined use of several active agents to treat different diseases. Compared with single-drug therapy, the combinatorial drugs instead of monotherapy can improve efficacy [1], reduce side effects [2] and overcome drug resistance [3,4].Drug combinations are increasingly used to treat a variety of complex diseases, such as hypertension [5], infectious diseases [6], and cancer [7,8].…”