The Mechanism of Monolaurate Derived Glycolipids Inhibit Sindbis Virus Replication and Release
Alphaviruses transmitted by mosquitoes represent an expanding global health concern, yet no specific antiviral agents are currently available for clinical use. Trehalose‐derived glycolipids have emerged as promising host‐modulating compounds with antiviral potential, but their relevance to alphavirus infection has remained largely unexplored. Here, we investigated the antiviral properties and underlying mechanism of trehalose 6‐monolaurate (TML) against Sindbis virus (SINV), a representative Old World alphavirus. In cell‐based infection models, TML demonstrated strong antiviral efficacy with favorable selectivity, substantially outperforming the structurally related analog trehalose 6‐monooleate. Exposure to TML led to a concentration‐dependent reduction in intracellular viral RNA synthesis, viral protein accumulation, and release of infectious progeny across distinct cell types. Kinetic and mechanistic analyses indicated that TML exerts its antiviral activity predominantly after viral entry, while exerting limited influence on virion integrity, cellular attachment, or internalization. Global transcriptomic analysis revealed that TML counterbalances SINV‐driven host gene expression programs, particularly those involving immediate‐early transcriptional regulators and pathways linked to Wnt signaling, cellular metabolism, and biosynthetic processes. Network‐based interrogation further highlighted EGR1‐associated transcriptional hubs as central targets of TML‐mediated modulation. Together, these results establish TML as a host‐directed inhibitor acting at post‐entry stages of alphavirus replication and underscore its potential as a lead scaffold for the development of therapeutics against mosquito‐borne RNA viruses.