With the emergence and rapid spreading of NDM-1 and existence of clinically relevant VIM-1 and IMP-1, discovery of pan inhibitors targeting metallo-beta-lactamases (MBLs) became critical in our battle against bacterial infection. Concurrent with our fragment and high-throughput screenings, we performed a knowledge-based search of known metallo-beta-lactamase inhibitors (MBLIs) to identify starting points for early engagement of medicinal chemistry. A class of compounds exemplified by 11, discovered earlier as B. fragilis metallo-beta-lactamase inhibitors, was selected for in silico virtual screening. From these efforts, compound 12 was identified with activity against NDM-1 only. Initial exploration on metal binding design followed by structureguided optimization led to the discovery of a series of compounds represented by 23 with a pan MBL inhibition profile. In in vivo studies, compound 23 in combination with imipenem (IPM) robustly lowered the bacterial burden in a murine infection model and became the lead for the invention of MBLI clinical candidates.

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Background Metallo-beta (β)-lactamases (MBLs) are in the class B group of β-lactamases due to zinc ions in the active site that are required for enzymatic activity. MK-3866 is a small molecule MBL inhibitor (MBLi) that restores antibacterial activity against resistant MBL-expressing gram-negative bacteria. Efflux is an important mechanism of antibiotic resistance in Pseudomonas aeruginosa (Pa). Imipenem (IMI) is not subject to efflux and neither is relebactam (REL), a β-lactamase inhibitor (BLi) of class A and C β-lactamases that is approved in combination with IMI/cilastatin. An efflux assay was devised to characterize MBLis for potentiation of IMI or cefepime (FEP) in isogenic strain pairs of efflux wild-type and multiply efflux deleted (MED) strains of Pa. Our objective was to determine if MK-3866 and related analogs are subject to efflux in Pa. Methods Bacterial isolates were engineered to demonstrate the ability of MBLis to be effluxed by introducing IMI metallo-β-lactamase-1 (IMP-1) (by electroporation) or Verona integron-encoded metallo-β-lactamases (VIM-1, VIM-2) (utilizing the cloning vector pFlp2) into isogenic MED and wild-type (WT) Pa isolates. Susceptibility testing was performed with IMI or FEP at a fixed concentration equal to the Pa Clinical and Laboratory Standards Institute susceptibility breakpoint for each and including a fixed 4 µg/mL of REL to inhibit class A or C enzymes, referred to as the SLICE assay. The concentration of MK-3866 and analogues required to restore susceptibility to either antibiotic in WT and MED strains was assessed. Results MICs to the combination of MK-3866/REL/IMI or MK-3866/REL/FEP vary by ≤4-fold between the WT and MED strain of each isogenic strain pair (Table 1). In contrast, large differential MIC values (efflux ratios) can be seen for the MK-3866 analogues A and B, with more moderate efflux ratios observed for other analogues. The exact efflux ratio depended on the MBL studied and the antibiotic partner. Conclusion MK-3866 showed a low potential for efflux whether IMP-1, VIM-1, or VIM-2 was expressed, in contrast to analogues of MK-3866 which exhibited differentials from nominal to extreme (≥ 128-fold) between efflux WT versus MED isolates. Disclosures Katherine Young, M.S., Merck & Co., Inc.: Stocks/Bonds Asra Mirza, MS, Merck & Co., Inc.: Stocks/Bonds Carl Balibar, PhD, Merck & Co., Inc.: Stocks/Bonds Shuzhi Dong, PhD, Merck & Co., Inc.: Stocks/Bonds Frank Bennett, PhD, Merck & Co., Inc.: Stocks/Bonds Jinlong Jiang, PhD, Merck & Co., Inc.: Stocks/Bonds Haiqun Tang, PhD, Merck & Co., Inc.: Stocks/Bonds Claire Tudge, PhD, Merck & Co., Inc.: Stocks/Bonds Jack Scott, PhD, Merck & Co., Inc.: Stocks/Bonds Dexi Yang, PhD, Merck & Co., Inc.: Stocks/Bonds Alexander Pasternak, PhD, Merck & Co., Inc.: Stocks/Bonds.

A renal outer medullary potassium channel (ROMK, Kir1.1) is a putative drug target for a novel class of diuretics with potential for treating hypertension and heart failure. Our first disclosed clinical ROMK compound, 2 (MK-7145), demonstrated robust diuresis, natriuresis, and blood pressure lowering in preclinical models, with reduced urinary potassium excretion compared to the standard of care diuretics. However, 2 projected to a short human half-life (∼5 h) that could necessitate more frequent than once a day dosing. In addition, a short half-life would confer a high peak-to-trough ratio which could evoke an excessive peak diuretic effect, a common liability associated with loop diuretics such as furosemide. This report describes the discovery of a new ROMK inhibitor 22e (MK-8153), with a longer projected human half-life (∼14 h), which should lead to a reduced peak-to-trough ratio, potentially extrapolating to more extended and better tolerated diuretic effects.