The Novel β-Lactamase Inhibitor, ETX-2514, in Combination with Sulbactam Effectively Inhibits Acinetobacter baumannii
Authors: Barnes Melissa D , Bethel Christopher R , Rutter Joseph D , Akker Focco Van Den , Papp-Wallace Krisztina M , Bonomo Robert A 
Published: October 2017

Abstract

Background
Multidrug resistant (MDR) Acinetobacter sp. were deemed a “serious” health threat by the Centers for Disease Control and Prevention with a daunting 63% of infections being nearly untreatable. Underlying this challenging pathogen are the presence of a chromosomal class C β-lactamase, Acinetobacter-derived cephalosporinase (ADC), as well as the abundant prevalence of class D OXA β-lactamases that hydrolyze carbapenems in conjunction with a lack of potent β-lactamase inhibitors. Based on the ability of ETX2514, a rationally designed novel diazabicyclooctane inhibitor (Figure 1A), to inhibit class D β-lactamases as well as PBPs, we hypothesized that highly resistant clinical isolates of A. baumannii will demonstrate susceptibility to the sulbactam-ETX2514 combination and that A. baumannii β-lactamases, ADC-7 and OXA-58 will be readily inactivated by ETX2514.

Methods
Susceptibility testing according to Clinical and Laboratory Standards Institute was performed for sulbactam ±4 mg/L ETX2514 using 72 A. baumannii strains. More than half of the isolates are MDR, have ≥ eight resistant determinants, contain an ADC β-lactamase, and have OXA-23 and OXA-58-like β-lactamases in the carbapenem resistant isolates. ADC-7 and OXA-58 β-lactamases were purified and characterized with ETX2514 by steady-state inhibition kinetics and Q-TOF mass spectrometry.

Results
The A. baumannii strains demonstrated an MIC90 of 32 mg/L for sulbactam and 2 mg/L for the sulbactam-ETX-2514 combination. The addition of ETX2514 lowered the MIC50 from 8 to 1 mg/L (Figure 1B). ETX-2514 effectively inhibited purified OXA-58 (k2/K = 2.5 ± 0.3 × 105 M−1 s−1 and Ki apP = 0.39 ± 0.01 µM) and ADC-7 (k2/K =1.0 ± 0.1 × 106 M−1 s−1 and Ki app = 0.11 ± 0.04 µM). The two β-lactamases displayed similar dissociation constants (Kd =1 ± 0.1 nM), but ADC-7 possessed a faster dissociation rate (koff =8 ± 1 × 10−4 s−1 for ADC-7 and 1.6 ± 0.3 × 10−4 s−1for OXA-58). Using mass spectrometry, ADC-7 and OXA-58 were found to stably bind the ETX-2514 compound for up to 24 hours (Figure 1C).

Conclusion
ETX2514 is a new β-lactamase inhibitor that is strikingly effective at restoring susceptibility to highly drug-resistant A. baumannii isolates when combined with sulbactam via inhibition of the ADC-7 and OXA-58 β-lactamases.

Disclosures
F. Van Den Akker, Entasis: Grant Investigator, Research grant Wockhardt: Grant Investigator, Research grant; K. M. Papp-Wallace, Entasis: Grant Investigator, Research grant Allecra: Grant Investigator, Research grant Merck: Grant Investigator, Research grant Roche: Grant Investigator, Research grant Allergan: Grant Investigator, Research grant; R. A. Bonomo, Entasis: Grant Investigator, Research grant Allecra: Grant Investigator, Research grant Wockhardt: Grant Investigator, Research grant Merck: Grant Investigator, Research grant Roche: Grant Investigator, Research grant GSK: Grant Investigator, Research grant Allergan: Grant Investigator, Research grant Shionogi: Grant Investigator, Research grant