Design, synthesis, and optimization of Honokiol-Piperazine derivatives as lung-enriched agents against MRSA pneumonia infection.

Methicillin-resistant Staphylococcus aureus (MRSA) causes severe bacterial pneumonia; current therapies (e.g., vancomycin) suffer from poor pulmonary bioavailability and dose-limiting safety concerns, demanding precision lung-enriched agents. However, small-molecule lung-enriched antibacterials are relatively rare. Honokiol (HNK), a natural phenolic compound with broad pharmacological activities but moderate antibacterial potency and limited tissue selectivity, was selected as the scaffold. We integrated a piperazine moiety into HNK and synthesized a series of HNK-piperazine derivatives. Among them, 13f showed potent activity primarily against Gram-positive bacteria, including MRSA (MIC: 2-4 μM; 16-32-fold more potent than HNK) together with preferential lung exposure. 13f exhibited rapid bactericidal activity and showed minimal MIC shift under sub-MIC selection over 31 serial passages (MICₙ/MIC₀ = 1-2), whereas ciprofloxacin displayed a 256-fold increase over the same period; it also attenuated staphyloxanthin production and reduced biofilm biomass. Mechanistic assays suggested time- and concentration-dependent membrane-associated effects, as reflected by permeability/depolarization readouts, nucleic acid/protein leakage, SEM imaging, and complementary MD analyses, with more pronounced effects at higher exposures. Pharmacokinetic and biodistribution analyses revealed a 13-fold longer half-life and a 147.92-fold higher lung AUC for 13f than for HNK. In a murine MRSA pneumonia model, 13f substantially reduced pulmonary bacterial burden and showed in vivo activity in the same overall range as vancomycin under the tested dosing regimen. No obvious histopathological alterations were observed under the tested short-term conditions. Overall, this work demonstrated a small-molecule approach to combine lung enrichment with potent anti-MRSA activity, supporting the further preclinical evaluation of 13f for MRSA pneumonia.