Kaempferol 3‐O‐gentiobioside, an ALK5 inhibitor, affects the proliferation, migration, and invasion of tumor cells via blockade of the TGF‐β/ALK5/Smad signaling pathway

Overactivation of TGF‐β/ALK5/Smad signaling pathway has been observed in the advanced stage of various human malignancies. As a key component of TGF‐β/ALK5/Smad signaling pathway transduction, TGF‐β type I receptor (also known as ALK5) has emerged as a promising therapeutic target for cancer treatment. In this study, to discover a novel ALK5 inhibitor, a commercial natural products library was screened using docking‐based virtual screening, followed by luciferase reporter assay. A flavonoid glycoside kaempferol 3‐O‐gentiobioside (KPF 3‐O‐G) was identified as a potent ALK5 inhibitor through directly bound to the ATP‐site of ALK5, resulting in the inhibitory effects on phosphorylation and translocation of Smad2 and expression of Smad4. Additionally, we found that KPF 3‐O‐G reduced cell proliferation and inhibited TGF‐β‐induced cell migration and invasion. Moreover, western blotting and immunofluorescent analysis showed that KPF 3‐O‐G significantly reversed the TGF‐β‐induced EMT biomarkers, including upregulation of E‐cadherin and downregulation of N‐cadherin, vimentin, and snail. In vivo study showed that KPF 3‐O‐G administration reduced tumor growth in human ovarian cancer xenograft mouse model, without obvious toxic effect. This study provided novel insight into the anticancer effects of KPF‐3‐O‐G and indicated that KPF‐3‐O‐G might be developed as potential therapeutics for cancer treatment after further validation.