Mechanistic insights into phthalylsulfacetamide-induced restoration of meropenem bactericidal activity in NDM-1-positive Escherichia coli

The emergence and global spread of New Delhi metallo-β-lactamase-1 (NDM-1) pose a severe threat to the efficacy of carbapenems, which are considered the last line of defense against infections caused by multidrug-resistant gram-negative bacteria. In this study, through virtual screening and enzyme activity inhibition assays, phthalylsulfacetamide (PSA) was identified as a compound exhibiting inhibitory activity against NDM-1. PSA dose-dependently inhibited NDM-1, with a 50% inhibitory concentration (IC50) of 15.4 ± 0.3 µM and an inhibition rate (IR%) of 89.2%. In vitro antibacterial activity assays, confirmed that PSA effectively restored the antibacterial activity of meropenem (MEM) against NDM-1-positive Escherichia. coli (E. coli), reducing the minimum inhibitory concentration (MIC) of MEM by 32-fold and demonstrating rapid bactericidal effects within 3 h. The combination of PSA and MEM had a significant synergistic effect with a fractional inhibitory concentration index (FICI) of 0.156. In vivo, the combination therapy significantly reduced the bacterial load, decreased inflammatory cytokine levels, alleviated histopathological damage and increased survival rates in a murine systemic infection model. Molecular dynamics simulation and site-directed mutagenesis revealed that PSA stably binds to the active hydrolysis center of NDM-1, interacts with Zn2+, and strongly interacts with the key amino acid residues Val73 and His122. Furthermore, the observed rapid bactericidal activity suggests that PSA’s mechanism of action may extend beyond NDM-1 inhibition. Molecular docking analysis indicates that PSA may potentially interact with LpxC, a key enzyme involved in lipopolysaccharide synthesis; however, this observation remains preliminary and requires further experimental validation. These results highlight PSA as a promising therapeutic agent against NDM-1-mediated carbapenem resistance.