PROJECT SUMMARY β-lactams including carbapenems are the most widely used antibiotic class. Carbapenem-resistant A. baumannii (CRAB) represents the most extreme resistant human pathogen among the Acinetobacter species, where acquisition and evolution of β-lactamases have essentially eliminated all therapeutic options. CRAB infections are classified by the US Centers for Disease Control and Prevention as an “Urgent threat” and by the World Health Organization as a “Priority 1 critical threat”. To provide a new therapeutic option against this prominent pathogen, Venatorx has identified a novel chemical series of highly selective cyclic boronates that inhibit penicillin-binding proteins (PBPs) like β-lactams but that are impervious to the action of all β-lactamases. This research program aims to deliver a cyclic boronate PBP inhibitor (boro-PBPi) for the treatment of CRAB infections that exploits porin mediated uptake rather than siderophore conjugation, in order to avoid both inactivation by β-lactamases and reduced activity through loss of siderophore uptake as has been observed with cefiderocol (Fetroja®). Significant strides in microbiological activity have been achieved with the lead boro-PBPi VNRX-14791 exhibiting an MIC90 of 16 µg/mL relative to 128/64 µg/mL for ampicillin-sulbactam and 64/4 µg/mL for sulbactam-durlobactam against 26 CRAB clinical isolates. Prototype boro-PBPi VNRX-6884 was bactericidal in vitro and achieved proof of concept efficacy in the murine thigh infection model in carbapenem-producing A. baumannii while possessing favorable selectivity, ADME and PK properties. Optimization efforts proposed herein target a 4-fold improvement in antibacterial activity to reach an MIC90 ≤4 µg/mL, driven by rational structure-guided design for AbPBP3 affinity as well as using specific physicochemical properties to improve cellular accumulation in gram-negative bacteria. As contingency, we will also explore optimization of inhibitors that target both AbPBP2 and AbPBP3 using a recently identified starting point (VNRX-14841) to enhance efficacy in vivo through dual inhibition AbPBP2 and AbPBP3 that function in the elongasome and divisome, respectively. Such an optimized boro-PBPi will be a first new class antibiotic addressing resistance to β-lactams for the treatment of CRAB infections and a long-term therapeutic solution to resistance development in A. baumannii.