A New Promising Class of pyrazolobenzothiazine inhibitors of S. aureus NorA Efflux Pump

The increasing resistance to antibacterials commonly employed in the clinic, together with the spread of multidrug resistant bacterial strains, suggests that the development of new therapeutic approaches should be of primary importance (1). Among the mechanisms whereby the bacteria develop resistance to antimicrobial agents, the overexpression of efflux pumps provide a self-defense mechanism by which antibacterials are actively removed from the cell. For antibacterials, this results in sub-lethal drug concentrations at the active site that in turn may predispose the organism to the development of high-level target-based resistance. Therefore, efflux pumps are viable antibacterial targets and identification and development of potent efflux pump inhibitors (EPIs) is a promising and valid strategy, which can restore the susceptibility of resistant strains to antibacterial substrates of efflux pumps. Among the multidrug resistant bacteria, methicillin-resistant Staphylococcus aureus (MRSA) accounted for 44% (n = 171200) of healthcare-associated infections and 22% (n = 5400) of attributable extra deaths in 2008 in EU. The increased expression of one or more MDR efflux pump genes was identified in 151 out of 309 S. aureus clinical strains (49%). Among those overexpressing a single gene, norA was the most common (43%), followed by its strict homologue norB (23.2%) and mepA (9.9%)(2). It has been recently reported that celecoxib, a COX-2 specific inhibitor, increases the sensitivity of some S. aureus strains to different antibacterials (3). Taking this evidence into account, we decided to bring to light as possible EPIs a focused in-house library of more than 150 compounds, bearing the 1,4-dihydropyrazolo[4,3-c]-benzothiazine 5,5-dioxide nucleus, previously designed and synthesized as celecoxib analogues. We then used our previously reported NorA in silico model for ethidium bromide (EtBr) efflux inhibition (4) to select a focused subset of our library to be tested. In the present work, EPI activity of celecoxib was confirmed against the NorA efflux pump and a new chemotype (3g) with potent NorA EPI activity, a limited intrinsic antibacterial activity, and a reduced cytotoxicity was identified (5). (1) Piddock, L.J. Lancet Infect. Dis. 2012, 12, 249-253; (2) Patel, D.; Kosmidis, C.; Seo, S.M.; Kaatz, G.W. Antimicrob. Agents Chemother. 2010, 54, 5070-5073; (3) Kalle, A.M.; Rizvi, A. Antimicrob. Agents Chemother. 2011, 55, 439-442; (4) Brincat, J. P.; Carosati, E.; Sabatini, S.; Manfroni, G.; Fravolini, A.; Raygada, J. L.; Patel, D.; Kaatz, G. W.; Cruciani, G. J. Med. Chem. 2011, 54, 354−365; (5) Sabatini, S.; Gosetto, F.; Serritella, S.; Manfroni, G.; Tabarrini, O.; Iraci, N.; Brincat, J.P.; Carosati, E.; Villarini, M.; Kaatz, G.W.; Cecchetti V. J. Med. Chem. 2012, 55, 3568-3572.