2-Sulfonylpyrimidines as Privileged Warheads for the Development of S. aureus Sortase A Inhibitors

Abstract

Staphylococcus aureus is one of the most frequent causes of nosocomial and community-acquired infections, with emerging multiresistant isolates causing a significant burden to public health systems. We identified 2-sulfonylpyrimidines as a new class of potent inhibitors against S. aureus sortase A acting by covalent modification of the active site cysteine 184. Series of derivatives were synthesized to derive structure-activity relationship (SAR) with the most potent compounds displaying low micromolar KI values. Studies on the inhibition selectivity of homologous cysteine proteases showed that 2-sulfonylpyrimidines reacted efficiently with protonated cysteine residues as found in sortase A, though surprisingly, no reaction occurred with the more nucleophilic cysteine residue from imidazolinium-thiolate dyads of cathepsin-like proteases. By means of enzymatic and chemical kinetics as well as quantum chemical calculations, it could be rationalized that the SNAr reaction between protonated cysteine residues and 2-sulfonylpyrimidines proceeds in a concerted fashion, and the mechanism involves a ternary transition state with a conjugated base. Molecular docking and enzyme inhibition at variable pH values allowed us to hypothesize that in sortase A this base is represented by the catalytic histidine 120, which could be substantiated by QM model calculation with 4-methylimidazole as histidine analog.

Dual Strategy to Design New Agents Targeting Schistosoma mansoni: Advancing Phenotypic and SmCB1 Inhibitors for Improved Efficacy

“…We initiated our search for lead structures with antischistosomal activity by screening an in-house library of 76 compounds. The library consisted of several cysteine protease-targeting inhibitors decorated with different covalent warheads, such as fluorovinylsulfon­(at)­es, nitriles, aldehydes, 4-oxoenoates, nitroalkenes, and acrylamides. − The compounds were tested in phenotypic assays on NTS and S. mansoni adults .…”

Investigation of the Compatibility between Warheads and Peptidomimetic Sequences of Protease Inhibitors—A Comprehensive Reactivity and Selectivity Study

“…Thus, to determine whether a reaction can be expected to take place, it is important to consider the whole reaction path, including the activation barriers, which determine the kinetics. Previous calculations have revealed that MeSH is often insufficiently nucleophilic to allow a reaction to occur at room temperature [ 57 ]. A base, such as triethylamine, serves as interim storage for the thiol proton before it is transferred to the warhead.…”

Emerging and Re-emerging Warheads for Targeted Covalent Inhibitors: An Update

“…Based on the DOCKovalent method, 367 a virtual library of more than 88,000 make-on-demand compounds was docked, out of which initially seven compounds (e.g., 193, Figure 56A) were synthesized. Further elaboration supported by docking led to the first covalent eIF4E inhibitor with cellular activity (194,Figure 56A), highlighting the utility of their docking approach. Even though hydrolytic and GSH stability was, as described previously in other cases, rather limited (t 1/2 = 30 min for hydrolytic stability and t 1/2 = 2.7 min for GSH stability of 194 at pH 7.5), the sufficiently fast reaction on target (k inact = 2.0 × 10 −2 s −1 , k inact /K I = 5.5 × 10 3 M −1 s −1 ) may partly compensate for this instability.…”

“…Notably, SrtA has been shown to feature a “reversely protonated” catalytic cysteine residue (>99.9% protonated at pH 7.5), − which seems to make it less amenable to classical protease-targeted warheads and Michael acceptors. While a variety of warheads, including disulfides, , benzisothiazolinones, thiadiazolidine-3,5-diones, and 1,3,4-thiadiazoles, have also been reported to target this enzyme, Barthels et al followed up on the above sulfonyl­pyrimidine series using a more sophisticated approach relying on covalent adduct detection by MS, kinetic experiments, enzyme inhibition at different pH values, and quantum mechanics (QM) calculations. Significant improvements in reversible binding affinity and warhead placement were achieved, but enzyme inhibition (described by the rate constant k second corresponding to k inact / K I ) also strongly correlated with the electron-withdrawing nature of substituents (and thus intrinsic reactivity) at the pyrimidine ring.…”