Studies on the Reactions of Biapenem with VIM Metallo ß-Lactamases and the Serine ß-Lactamase KPC-2.

Carbapenems are important antibacterials and are both substrates and inhibitors of some ß-lactamases. We report studies on the reaction of the unusual carbapenem biapenem, with the subclass B1 metallo-ß-lactamases VIM-1 and VIM-2 and the class A serine-ß-lactamase KPC-2. X-ray diffraction studies with VIM-2 crystals treated with biapenem reveal the opening of the ß-lactam ring to form a mixture of the (2S)-imine and enamine complexed at the active site. NMR studies on the reactions of biapenem with VIM-1, VIM-2, and KPC-2 reveal the formation of hydrolysed enamine and (2R)- and (2S)-imine products. The combined results support the proposal that SBL/MBL-mediated carbapenem hydrolysis results in a mixture of tautomerizing enamine and (2R)- and (2S)-imine products, with the thermodynamically favoured (2S)-imine being the major observed species over a relatively long-time scale. The results suggest that prolonging the lifetimes of ß-lactamase carbapenem complexes by optimising tautomerisation of the nascently formed enamine to the (2R)-imine and likely more stable (2S)-imine tautomer is of interest in developing improved carbapenems.

Faropenem reacts with serine and metallo-ß-lactamases to give multiple products.

Penems have demonstrated potential as antibacterials and ß-lactamase inhibitors; however, their clinical use has been limited, especially in comparison with the structurally related carbapenems. Faropenem is an orally active antibiotic with a C-2 tetrahydrofuran (THF) ring, which is resistant to hydrolysis by some ß-lactamases. We report studies on the reactions of faropenem with carbapenem-hydrolysing ß-lactamases, focusing on the class A serine ß-lactamase KPC-2 and the metallo ß-lactamases (MBLs) VIM-2 (a subclass B1 MBL) and L1 (a B3 MBL). Kinetic studies show that faropenem is a substrate for all three ß-lactamases, though it is less efficiently hydrolysed by KPC-2. Crystallographic analyses on faropenem-derived complexes reveal opening of the ß-lactam ring with formation of an imine with KPC-2, VIM-2, and L1. In the cases of the KPC-2 and VIM-2 structures, the THF ring is opened to give an alkene, but with L1 the THF ring remains intact. Solution state studies, employing NMR, were performed on L1, KPC-2, VIM-2, VIM-1, NDM-1, OXA-23, OXA-10, and OXA-48. The solution results reveal, in all cases, formation of imine products in which the THF ring is opened; formation of a THF ring-closed imine product was only observed with VIM-1 and VIM-2. An enamine product with a closed THF ring was also observed in all cases, at varying levels. Combined with previous reports, the results exemplify the potential for different outcomes in the reactions of penems with MBLs and SBLs and imply further structure-activity relationship studies are worthwhile to optimise the interactions of penems with ß-lactamases. They also exemplify how crystal structures of ß-lactamase substrate/inhibitor complexes do not always reflect reaction outcomes in solution.

Broad Spectrum ß-Lactamase Inhibition by a Thioether Substituted Bicyclic Boronate.

ß-Lactamases comprise the most widely used mode of resistance to ß-lactam antibiotics. Cyclic boronates have shown promise as a new class of ß-lactamase inhibitor, with pioneering potential to potently inhibit both metallo- and serine-ß-lactamases. We report studies concerning a bicyclic boronate ester with a thioether rather than the more typical ß-lactam antibiotic "C-6/C-7" acylamino type side chain, which is present in the penicillin/cephalosporin antibiotics. The thioether bicyclic boronate ester was tested for activity against representative serine- and metallo-ß-lactamases. The results support the broad inhibition potential of bicyclic boronate based inhibitors with different side chains, including against metallo-ß-lactamases from B1, B2, and B3 subclasses. Combined with previous crystallographic studies, analysis of a crystal structure of the thioether inhibitor with the clinically relevant VIM-2 metallo-ß-lactamase implies that further SAR work will expand the already broad scope of ß-lactamase inhibition by bicyclic boronates.