Enzyme−Antibiotic Liquid Crystalline Nanoparticles for Enhanced Antibiofilm Activity, Bacterial Clearance, and Accelerated Healing of Infected Wounds

Biofilm-associated wound infections remain difficult to treat because bacteria are protected within extracellular polymeric matrices that restrict antibiotic penetration, promote antimicrobial tolerance, and delay tissue repair. Here, we developed liquid crystalline nanoparticles (LCNPs) for topical coformulation of gentamicin (GEN) and recombinant human DNase I (rhDNase), combining aminoglycoside therapy with an eDNA-targeting enzyme. GEN + rhDNase-LCNPs were successfully formulated as nanoscale particles with low polydispersity, high encapsulation efficiency, and formulation-dependent in vitro release behavior. LCNP formulation preserved GEN antibacterial activity against planktonic Escherichia coli Xen14 and reduced the MBIC against established biofilms by approximately 4-fold. In a minimum biofilm eradication concentration (MBEC) model, GEN + rhDNase-LCNPs achieved the greatest reduction in biofilm-associated viable bacteria, producing an approximately 6-log reduction compared with untreated biofilm controls and supporting the benefit of combining antibiotic delivery with rhDNase. Notably, GEN + rhDNase-LCNPs markedly suppressed bacterial burden and reduced recoverable bacteria at day 9 to near the limit of detection in a bioluminescent E. coli Xen14 biofilm-associated excisional wound infection mouse model. This antibacterial effect translated into accelerated macroscopic wound closure and improved histological repair, including an approximately 80% reduction in wound gape compared with infected controls. Masson's trichrome staining further confirmed enhanced tissue remodeling, with the collagen-positive wound area increasing to approximately 88% following GEN + rhDNase-LCNP treatment, compared with approximately 21% in infected controls. In conclusion, LCNP-mediated codelivery of GEN and rhDNase improved bacterial clearance, re-epithelialization, and collagen deposition, supporting this platform as a promising topical antibiofilm strategy for infected wounds.