Hydroxychloroquine alters the cytoskeleton to impair cell migration

Hydroxychloroquine (HCQ), a clinically relevant quinoline derivative, impairs both collective and individual cell migration by disrupting actin and vimentin cytoskeletal dynamics during wound healing. In scratch assays with HeLa cells, HCQ treatment significantly reduces wound closure rates and inhibits bursts of coordinated migration, as well as single-cell motility. Quantitative imaging reveals that HCQ diminishes actin filament density at the wound edge and induces reorganization of the vimentin network, resulting in smaller nuclei and compromised structural connectivity. Particle tracking micro-rheology demonstrates that HCQ softens the cytoplasm and decreases cellular mechanical heterogeneity. In vitro spectroscopic studies show that HCQ binds cooperatively to actin with micromolar affinity, perturbing its secondary structure, reducing filament polymerisation rates, and impairs interactions between actin and actin binding proteins (ABPs). HCQ also significantly suppresses lamellipodial protrusive activity, indicating a link between cytoskeletal remodelling and impaired cell migration. Collectively, these findings establish that HCQ disrupts essential mechanisms for directed migration by modulating the cytoskeleton and cell mechanics. This multifaceted impairment may underlie therapeutic potential of HCQ as an anticancer agent by restricting cellular invasive capacity and remodelling required for tumour progression.