Over 30 compounds, including para-, meta-, and ortho-phenylenediboronic acids, ortho-substituted phenylboronic acids, benzenetriboronic acids, di- and triboronated thiophenes, and pyridine derivatives were investigated as potential β-lactamase inhibitors. The highest activity against KPC-type carbapenemases was found for ortho-phenylenediboronic acid 3a, which at the concentration of 8/4 mg/L reduced carbapenems' MICs up to 16/8-fold, respectively. Checkerboard assays revealed strong synergy between carbapenems and 3a with the fractional inhibitory concentrations indices of 0.1-0.32. The nitrocefin hydrolysis test and the whole cell assay with E. coli DH5α transformant carrying blaKPC-3 proved KPC enzyme being its molecular target. para-Phenylenediboronic acids efficiently potentiated carbapenems against KPC-producers and ceftazidime against AmpC-producers, whereas meta-phenylenediboronic acids enhanced only ceftazidime activity against the latter ones. Finally, the statistical analysis confirmed that ortho-phenylenediboronic acids act synergistically with carbapenems significantly stronger than other groups. Since the obtained phenylenediboronic compounds are not toxic to MRC-5 human fibroblasts at the tested concentrations, they can be considered promising scaffolds for the future development of novel KPC/AmpC inhibitors. The complexation of KPC-2 with the most representative isomeric phenylenediboronic acids 1a, 2a, and 3a was modeled by quantum mechanics/molecular mechanics calculations. Compound 3a reached the most effective configuration enabling covalent binding to the catalytic Ser70 residue.
Keywords: KPC/AmpC β-lactamase inhibitors; antibacterial activity; arylboronic acids; molecular docking; time-dependent QM/MM.