Design and SAR Analysis of Phenylboronic Acid-Based Inhibitors for Sensitizing KPC-2-Producing <i>Klebsiella pneumoniae</i> to β-Lactam Antibiotics

The global threat posed by multidrug-resistant bacteria, particularly those producing KPC-2 carbapenemases, has compromised the effectiveness of carbapenems, the last-resort antibiotics. To address this, we utilized a bioisosteric replacement approach to synthesize phenylboronic acid (PBA) derivatives targeting KPC-2. The enzyme kinetics study revealed that electron-withdrawing substituents enhanced the residence time of lead compounds, enhancing KPC-2 inhibition. Lead molecule <b>3b</b>, in synergy with Meropenem, achieved a 3-log CFU/mL reduction in bacterial count within 3 h in time-kill kinetics and exhibited a postantibiotic effect of 71 ± 5 min. A nanomolar range inhibition constant (<i>K</i>_<i>i</i>), a second-order inactivation rate constant (<i>k</i>₂/<i>K</i>) of ∼10³ M^-1 s^-1, and a slow off-rate (<i>k</i>_off ∼ 0.0018 s^-1) were observed. Importantly, the PBA derivatives were nontoxic to human cells and significantly reduced lung bacterial load in a murine pneumonia model. Overall, PBA-based non-β-lactam β-lactamase inhibitors showed promise in restoring carbapenem efficacy against KPC-2-producers.