Exploring fluorination in MBL inhibitor design

Abstract

<p>Antimicrobial resistance (AMR) achieved through, among others, β-lactamase activity, remains an ongoing challenge for the treatment of infections caused by antibiotic-resistant pathogens. Metallo-β-lactamase (MBL)-expressing bacteria, resistant to most β-lactam antibiotics, are of growing clinical relevance with currently no approved treatment options available. The most common approaches to combating AMR include the modification of existing compounds or the development of new drugs and inhibitors. Alternative strategies are also being explored, including novel drug delivery systems. Recently, an approach based on exploiting the ability of MBLs to hydrolyze β-lactams has been investigated using cephalosporin prodrugs. The reported conjugates consisted of aromatic thiol-based MBL inhibitors linked to a cephalosporin scaffold that was designed to release the inhibitors upon enzymatic hydrolysis. <p>The goal of the current work was to extend the concept of thiol-releasing conjugates to aliphatic thiolbased MBL inhibitors, as aliphatic thiols have been described to lack the required leaving group properties. The work is based on the hypothesis that fluorination in the vicinity of the thiol moiety can be used as a tool in the design of aliphatic thiol-releasing conjugates. <p>Thus, the initial focus was on designing fluorinated captopril analogues, exhibiting inhibitory activity against relevant MBLs, and investigating the influence of fluorination on their inhibitory effect. A library of fluorinated thiols, of which several exhibited IC₅₀ values in the low micromolar region against NDM-1, was developed. Furthermore, the use of fluorinated inhibitors as probes for NMR-based binding studies was demonstrated on a representative compound. <p>Among the active thiols, a few were selected for attachment to the cephalosporin core, resulting in a series of conjugates. The hydrolysis of the conjugates was examined and the inhibitory activity against several MBLs was determined. The conjugates were shown to release the thiols, as determined by LCMS and NMR. In addition, IC₅₀ values of the conjugates were found to be in a similar range to the respective thiol inhibitors. <p>In summary, the obtained conjugates verified that fluorination facilitates the release of aliphatic thiolbased inhibitors and provide directions for future conjugate development. Moreover, fluorinated thiol inhibitors can assist in studies of inhibitor-enzyme interactions and conjugate hydrolysis utilizing ¹⁹F NMR methods.