AFN‐1252 is a potent inhibitor of enoyl‐ACPreductase fromBurkholderia pseudomallei—Crystal structure, mode of action, and biological activity

Abstract

Melioidosis is a tropical bacterial infection caused by Burkholderia pseudomallei (B. pseudomallei; Bpm), a Gram-negative bacterium. Current therapeutic options are largely limited to trimethoprim-sulfamethoxazole and b-lactam drugs, and the treatment duration is about 4 months. Moreover, resistance has been reported to these drugs. Hence, there is a pressing need to develop new antibiotics for Melioidosis. Inhibition of enoyl-ACP reducatase (FabI), a key enzyme in the fatty acid biosynthesis pathway has shown significant promise for antibacterial drug development. FabI has been identified as the major enoyl-ACP reductase present in B. pseudomallei. In this study, we evaluated AFN-1252, a Staphylococcus aureus FabI inhibitor currently in clinical development, for its potential to bind to BpmFabI enzyme and inhibit B. pseudomallei bacterial growth. AFN-1252 stabilized BpmFabI and inhibited the enzyme activity with an IC 50 of 9.6 nM. It showed good antibacterial activity against B. pseudomallei R15 strain, isolated from a melioidosis patient (MIC of 2.35 mg/L). X-ray structure of BpmFabI with AFN-1252 was determined at a resolution of 2.3 Å . Complex of BpmFabI with AFN-1252 formed a symmetrical tetrameric structure with one molecule of AFN-1252 bound to each monomeric subunit. The kinetic and thermal melting studies supported the finding that AFN-1252 can bind to BpmFabI independent of cofactor. The structural and mechanistic insights from these studies might help the rational design and development of new FabI inhibitors.

Triclosan depletes the membrane potential in Pseudomonas aeruginosa biofilms inhibiting aminoglycoside induced adaptive resistance

“…aeruginosa biofilms. AFN-1252 forms a binary complex with the active site of FabI, an enoyl-acyl reductase responsible for the final elongation step in fatty acid synthesis and the molecular target of triclosan [ 50 ]. We reasoned that if triclosan inhibition of FabI was the mechanism responsible for increased activity of tobramycin, we should similarly observe synergy of AFN-1252 and tobramycin to kill P .…”

“…To determine if the inhibition of fatty acid biosynthesis and membrane biogenesis by triclosan is responsible for increasing P. aeruginosa susceptibility to tobramycin, we tested if the specific fatty acid synthesis inhibitor AFN-1252 synergized with tobramycin to kill P. aeruginosa biofilms. AFN-1252 forms a binary complex with the active site of FabI, an enoyl-acyl reductase responsible for the final elongation step in fatty acid synthesis and the molecular target of triclosan [50]. We reasoned that if triclosan inhibition of FabI was the mechanism responsible for increased activity of tobramycin, we should similarly observe synergy of AFN-1252 and tobramycin to kill P. aeruginosa biofilms.…”

A Review of Antibacterial Candidates with New Modes of Action

“…52−54 Crystal structures of S. aureus and Burkholderia pseudomallei FabI−AFN-1252 complexes showed that AFN-1252 (6) bound noncompetitively at the active site but independently of NADPH. 52,55 Afabicin (7) is currently being evaluated in a phase 2 trial (NCT03723551) using an iv/oral switch strategy for the treatment of S. aureus bone or joint infections 56 and has previously completed a phase 2 trial (NCT02426918) for staphylococcal ABSSSI infections. 57 Investigation of β-hairpin-shaped peptidomimetics stabilized by a D-proline−L-proline template based on the host-defense peptide protegrin I led to the identification of murepavadin (8) (POL7080, RG7929) which has potent activity against Pseudomonas spp.…”

“…A high throughput screening campaign at GSK against bacterial enzymes identified and then optimized active compounds against S. aureus FabI . These compounds had potent activity against staphylococci but did not possess broad spectrum activity, as many pathogens had salvage pathways or isoforms of FabI, such as FabK, that could undertake the biochemical function of the enzyme. , A new analog, AFN-1252 ( 6 ) (Debio 1452) and its prodrug afabicin ( 7 ) (Debio 1450, AFN-1720), were later discovered. − Crystal structures of S. aureus and Burkholderia pseudomallei FabI–AFN-1252 complexes showed that AFN-1252 ( 6 ) bound noncompetitively at the active site but independently of NADPH. , Afabicin ( 7 ) is currently being evaluated in a phase 2 trial (NCT03723551) using an iv/oral switch strategy for the treatment of S. aureus bone or joint infections and has previously completed a phase 2 trial (NCT02426918) for staphylococcal ABSSSI infections …”