Vactosertib reverses ABCG2-mediated multidrug resistance through dual inhibition of drug efflux and transporter expression

ATP-binding cassette (ABC) transporter ABCG2 is a key mediator of multidrug resistance (MDR) in Cancer. Its overexpression limits the intracellular retention and cytotoxicity of chemotherapeutic agents. Identifying clinically relevant modulators capable of suppressing ABCG2 activity remains an important therapeutic objective. In this study, we examined whether selective ALK4/5 inhibitor vactosertib can reverse ABCG2-associated MDR. At non-cytotoxic concentrations, vactosertib significantly reduced the IC₅₀ values of multiple ABCG2 substrates in both wild-type and R482-variant mutant ABCG2-overexpressing cell models, indicating that it restored drug sensitivity. Similar re-sensitizing effects of mitoxantrone were also observed in colony formation and multicellular tumor spheroid assays. Accumulation and efflux assays showed that vactosertib increased intracellular mitoxantrone accumulation by suppressing ABCG2-mediated efflux, which meet the result of docking simulations and ATPase assays. In addition to functional inhibition, vactosertib reduced the expression levels in both ABCG2 mRNA and protein without altering membrane localization, suggesting transcriptional downregulation. As a result, these findings indicate that vactosertib reverses ABCG2-mediated MDR through dual effects on inhibiting ABCG2 function and downregulating and the expression of ABCG2, supporting its potency as a chemo sensitizing agent in ABCG2-mediated MDR Cancer models.