Drug Repurposing Identifies Natamycin and Avacopan as Urease‐Targeted Therapeutic Candidates Against Helicobacter pylori

The global rise of antimicrobial resistance necessitates innovative strategies to combat persistent infections, including those caused by Helicobacter pylori. This study investigates potential inhibitors of the H. pylori urease enzyme, a key virulence factor that enables survival in the highly acidic gastric environment. A virtual screening of 1773 FDA‐approved drugs was performed, followed by molecular dynamics simulations. Among the top candidates, Natamycin, Zavegepant, Midostaurin, Avacopan, and Metolazone displayed significant binding affinities towards urease. Further simulations confirmed the stability and favorable interaction profiles of these drug–enzyme complexes, with the Avacopan–urease complex exhibiting the highest stability based on RMSD and binding free energy analyses. Natamycin and Avacopan were subsequently selected for in vitro validation, where both significantly inhibited urease activity and demonstrated antibacterial effects against clinical isolates and the reference strain H. pylori B128. Avacopan exhibited inhibition ranging from 30.0% in strain CL2 to 99.7% in B128, whereas Natamycin showed consistently stronger inhibition across all clinical strains, ranging from 89.2% in CL4 to 99.9% in CL1, and exceeding 90% in CL1, CL2, CL3, and B128. Both inhibitors were effective against cag‐positive and cag‐negative clinical isolates, indicating broad‐spectrum urease inhibition. However, the bactericidal activities of these compounds were more strain‐specific with increased efficacies observed in cag− isolates. Therefore, further studies are warranted to fully explore the specific mechanisms and selectivity of their individual inhibitory activities. These findings highlight Avacopan and Natamycin as promising urease‐targeted drug repurposing candidates for future therapeutic development against H. pylori.