Kinetic study of two novel enantiomeric tricyclic beta-lactams which efficiently inactivate class C beta-lactamases

A detailed kinetic study of the interaction between two ethylidene derivati ves of tricyclic carbapenems, Lek 156 and Lek 157, and representative beta -lactamases and D-alanyl-D-alanine peptidases (DD-peptidases) is presented. Both compounds are very efficient inactivators of the Enterobacter. cloaca e 908R beta -lactamase, which is usually resistant to inhibition. Prelimina ry experiments indicate that various extended-spectrum class C beta -lactam ases (ACT-1, CMY-1, and MIR-1) are also inactivated. With the E. cloacae 90 8R enzyme, complete inactivation occurs with a second-order rate constant, k(2)/K ', of 2 x 10(4) to 4 x 10(4) M-1 s(-1), and reactivation is very slo w, with a half-life of >1 h. Accordingly, Lek 157 significantly decreases t he MIC of ampicillin for E. cloacae P99, a constitutive class C beta -lacta mase overproducer. With the other serine beta -lactamases tested, the coval ent adducts exhibit a wide range of stabilities, with half-lives ranging fr om long (>4 h with the TEM-1 class A enzyme), to medium (10 to 20 min with the OXA-10 class D enzyme), to short (0.2 to 0.4 s with the NmcA class A be ta -lactamase). By contrast, both carbapenems behave as good substrates of the Bacillus cereus metallo-beta -lactamase (class B). The Streptomyces sp. strain R61 and K15 extracellular Do-peptidases exhibit low levels of sensi tivity to both compounds.