Targeting envelope lipids with 2,6‑di‑O‑methyl‑3‑acetyl‑β‑cyclodextrin impairs infectivity of SARS‑CoV‑2 and Japanese encephalitis virus.

Until pathogen-specific treatments and vaccines are available, early containment and prevention of further spread are essential. Here, we evaluated the antiviral activity of cyclodextrin (CyD) derivatives against enveloped RNA viruses, including SARS-CoV-2 and Japanese encephalitis virus (JEV). Among the 19 derivatives tested, 2,6-di-O-methyl-3-acetyl-β-cyclodextrin (DMA-β-CyD) exhibited the most potent antiviral activity against SARS-CoV-2 Wuhan, Beta, and Delta variants and demonstrated antiviral activity against the Wuhan variant (IC50 = 0.28 mM, CC50 ≥ 100 mM, selectivity index (SI) ≥ 357). Pretreatment of viral particles with DMA-β-CyD significantly reduced infectivity (by 2.25-2.67 log₁₀ units after 180 min of treatment) without affecting spike protein-ACE2 binding. DMA-β-CyD attenuated the procoagulant activity mediated by virion-associated aminophospholipids via interaction with the viral envelope. This effect was suppressed by 1-adamantanecarboxylic acid, suggesting that the inclusion of the target molecule of DMA-β-CyD is involved. Furthermore, DMA-β-CyD disrupted SARS-CoV-2 particles and reduced entry efficiency. Although the JEV-associated procoagulant activity was weaker than the SARS-CoV-2-associated procoagulant activity at 37°C, DMA-β-CyD recognized virion-associated aminophospholipids exposed on the surface of JEV particles. Against JEV, DMA-β-CyD exhibited comparable antiviral activity (IC50 = 0.84 mM, CC50 ≥ 100 mM, SI ≥119). These findings demonstrate that DMA-β-CyD exerts antiviral effects against both SARS-CoV-2 and JEV by targeting envelope aminophospholipids, which supports lipid-targeting strategies as a promising basis for broad-spectrum antiviral development.IMPORTANCEThe SARS-CoV-2 pandemic has highlighted the urgent need for effective antiviral strategies that can interrupt viral transmission before pathogen-specific vaccines or therapeutic interventions become available. In this study, we demonstrate that DMA-β-CyD is the first compound shown to reduce the infectivity of SARS-CoV-2 and Japanese encephalitis virus (JEV) particles by interacting with aminophospholipids, such as phosphatidylserine (PS). By exploiting differences in lipid composition between viral envelopes and host cell membranes, DMA-β-CyD provides a novel, lipid-targeting antiviral mechanism and may serve as a valuable platform for addressing infections caused by emerging and drug-resistant viral pathogens. This study underscores the therapeutic potential of disrupting envelope lipids as a broad-spectrum antiviral strategy.