Chemical Space Exploration around Thieno[3,2-d]pyrimidin-4(3H)-one Scaffold Led to a Novel Class of Highly Active Clostridium difficile Inhibitors

Abstract

<i>Clostridium difficile</i> infection (CDI) is the leading cause of healthcare-associated infection in the United States. Therefore, development of novel treatments for CDI is a high priority. Toward this goal, we began in vitro screening of a structurally diverse in-house library of 67 compounds against two pathogenic <i>C. difficile</i> strains (ATCC BAA 1870 and ATCC 43255), which yielded a hit compound, 2-methyl-8-nitroquinazolin-4(3<i>H</i>)-one (<b>2</b>) with moderate potency (MIC = 312/156 μM). Optimization of <b>2</b> gave lead compound <b>6a</b> (2-methyl-7-nitrothieno[3,2-<i>d</i>]pyrimidin-4(3<i>H</i>)-one) with improved potency (MIC = 19/38 μM), selectivity over normal gut microflora, CC₅₀s > 606 μM against mammalian cell lines, and acceptable stability in simulated gastric and intestinal fluid. Further optimization of <b>6a</b> at C2-, N3-, C4-, and C7-positions resulted in a library of >50 compounds with MICs ranging from 3 to 800 μM against clinical isolates of <i>C. difficile</i>. Compound <b>8f</b> (MIC = 3/6 μM) was identified as a promising lead for further optimization.