Inhibition of metallo-beta-lactamases by a series of mercaptoacetic acid thiol ester derivatives.

A series of mercaptoacetic acid thiol esters have been identified as metallo-beta-lactamase inhibitors. Electrospray mass spectrometry (ESMS) has shown that irreversible inhibition of the Bacillus cereus II metallo-beta-lactamase by SB214751, SB214752, and SB213079 was concomitant with a 90-Da increase in mass of the enzyme. Tryptic digestion of the B. cereus II inhibited with SB214751 illustrated that the peptide fragment, containing the only cysteine of the enzyme, had undergone a mass increment of 90 Da. It was further demonstrated that B. cereus II hydrolyzed this type of compound across the thiol ester bond to yield mercaptoacetic acid. Mercaptoacetic acid is the only molecular fragment common to SB214751, SB214752, and SB213079, and free mercaptoacetic acid does not bind covalently to B. cereus II. Therefore, it is concluded that these compounds inhibit B. cereus II by the mechanism-based delivery of mercaptoacetic acid, forming a disulfide linkage with the active sites cysteine (predicted mass shift = +90 Da) under the aerobic conditions of the assay. The different thiol esters examined had a broad range of potencies against the metallo-beta-lactamases tested. For example SB214751, SB214752, and SB213079 all had 50% inhibitory concentrations of < 10 and > 1,000 microM for the Stenotrophomonas maltophilia L-1 and Bacteroides fragilis CfiA enzymes, respectively. SB216968 was particularly active against the Aeromonas hydrophila CphA metallo-beta-lactamase and was found to be an uncompetitive inhibitor of this enzyme (Ki = 3.9 microM), whereas it exhibited irreversible inhibition of the L-1 enzyme. These observations with this series of compounds have revealed subtle differences between the active sites of different metallo-beta-lactamases. Finally, a novel application for isothermal titration calorimetry for assessing the zinc chelating activity of candidate inhibitors is also presented.

Inhibition of metallo-beta-lactamases by a series of thiol ester derivatives of mercaptophenylacetic acid.

A series of mercaptophenylacetic acid thiol esters bearing a phenyl substituent adjacent to the carboxylic acid function has been shown to be inhibitors of metallo-beta-lactamases. The inhibition of the Bacteroides fragilis CfiA and Bacillus cereus II metallo-beta-lactamases was Zn2- dependent, greater inhibition being observed at 1 microM ZnSO4 than at 100 microM ZnSO4. Despite this Zn2+ dependency, isothermal titration calorimetry studies illustrated that representative compounds had no detectable affinity for Zn2+ (K > 1 mM). This indicates that their mode of inhibition was not by chelation of the active site Zn2+. Greatest potency was observed against the Stenotrophomonas maltophilia L1 metallo-beta-lactamase with I50 values of between < 1.95 microM and 6 microM and SB-217843 exhibited a similar level of inhibition of this enzyme at 1 and 100 microM Zn2+ (I50 values 5 and 6 microM, respectively). Inhibition of B. cereus II metallo-beta-lactamase by SB-218018 and SB-217782 was competitive with Ki values of 185 microM and 1500 microM, respectively. Therefore, these compounds are specific inhibitors of metallo-beta-lactamases and provide further probes of the active sites of these enzymes.

Aryloxyalkyloxy- and aralkyloxy-4-hydroxy-3-nitrocoumarins which inhibit histamine release in the rat and also antagonize the effects of a slow reacting substance of anaphylaxis.

The syntheses and structure--activity relationships of a number of 4-hydroxy-3-nitrocoumarins, which are both antagonists of a slow reacting substance of anaphylaxis and potent inhibitors of antigen-induced histamine release in the rat, are described. Most active among these are 7-[3-(4-acetyl-3-hydroxy-2-n-propylphenoxy(-2-hydroxypropoxy] derivatives having hydrogen or lower alkyl substituents at the C-8 position of the coumarin ring, 168, 171, 173, and 174.