Natural compounds targeting E. Coli WRBA as antibiofilm agents: Virtual screening, design and synthesis of analogs, biological evaluation

Resistance to antibiotics has become one of the main concerns of modern medicine. This phenomenon is worsened by biofilm formation, which frequently leads to treatment failures.1 Hence, anti-biofilm agents represent useful tools in the treatment of microbial infections, increasing the effectiveness of antibiotics and preventing resistance mechanisms. Different studies have demonstrated that the protein WrbA plays a key role in the formation of biofilm in E. coli. Nonetheless, its specific function is still poorly understood, and only a few inhibitors are reported in the literature.2 For this reason, our research group has worked towards the identification of new WrbA inhibitors as antibiofilm agents. Through a target-based virtual database screening, we identified natural products endowed with a high affinity towards WrbA; the most promising compounds were purchased for biophysical and biological evaluation. Hence, we selected a natural scaffold as a starting point to create a new library of organic molecules and perform a virtual screening. Finally, five compounds were selected to be synthesized and subjected to biophysical evaluation. Both the natural and synthetic compounds were tested on recombinant E. coli WrbA by MicroScale Thermophoresis (MST), which allowed the determination of their dissociation constants (Kd). Among the natural compounds, one of them displayed a Kd in the nanomolar range. This value was lower than those measured for both caffeic and zosteric acid, which are known WrbA inhibitors.3 On the other hand, the synthesized compounds bound WrbA with Kd values in the low micromolar range. Based on our encouraging results, the inhibitors were assayed on E. coli and S. aureus to verify their antibiofilm effect. The outcomes of our study will be presented. References 1. Antimicrobial resistance https://www.who.int/news-room/fact-sheets/detail/antimicrobial-resistance (accessed 2022- 05-10). 2. Rossi, F.; Cattò, C.; Mugnai, G.; Villa, F.; Forlani, F.; Pinto, A.; Nuzzo, D. Effects of the Quinone Oxidoreductase WrbA on Escherichia Coli Biofilm Formation and Oxidative Stress. Antioxidants 2021, 10 (6), 919. 3. Cattò, C.; Dell’Orto, S.; Villa, F.; Villa, S.; Gelain, A.; Vitali, A.; Marzano, V.; Baroni, S.; Forlani, F.; Cappitelli, F. Unravelling the Structural and Molecular Basis Responsible for the Anti-Biofilm Activity of Zosteric Acid. PLoS ONE 2015, 10 (7), E0131519.