Benzofuran compound from Sophora tonkinensis Gagnep suppresses nasopharyngeal carcinoma growth via PI3K/AKT/mTOR pathway: evidence from molecular dynamics simulation and in vitro experiments

Introduction Shandougen (SDG), the dried root and rhizome of Sophora tonkinensis Gagnep. , has been historically documented in Ben Cao Qiu Zhen for its properties of “clearing heat, detoxifying, reducing swelling, and relieving throat disorders.” It is highly valued as a primary herb for relieving throat swelling and abscesses. SDG-based formulas, such as “Bi Yan Ling Tablets” and the “Bi’ai Formula,” have shown definite efficacy against heat-toxin accumulation type nasopharyngeal carcinoma (NPC). Despite this, the specific active monomer in SDG inhibiting NPC remains unclear, warranting further investigation. Objective This study aimed to identify the bioactive monomers underlying SDG’s anti-NPC efficacy and elucidate their molecular mechanisms. Methods Guided by HPLC fingerprinting, targeted isolation and purification of components from the active fraction were performed. 13 compounds were screened for activity, with benzofuran compound 2-(2′,4′-dihydroxyphenyl)-5,6-methylenedioxybenzofuran (ABF) showing the most potent in vitro effects. Molecular dynamics (MD) simulations were employed to evaluate the binding affinity and stability of ABF with the target protein. The anti-proliferative mechanisms of ABF were investigated in cells, and its anti-tumor capacity was evaluated using a nude mouse xenograft model. Results ABF exhibited stronger anti-proliferative activity against CNE1 and CNE2 cells compared to SDG extract, with a 10-fold lower 48-h IC 50 value. ABF inhibits the invasion and metastasis of NPC cells, which is associated with S-phase cell cycle arrest and the downregulation of cyclin E1 and CDK2 proteins. Apoptosis induction by ABF was associated with increased caspase family proteins and lowered the Bcl-2/Bax ratio. PCR and Western blot analyses confirmed that ABF is involved in the downregulation of the PI3K/AKT/mTOR pathway, inhibiting NPC cell growth. Additionally, in a nude mouse xenograft model, ABF treatment at high doses achieved tumor inhibition rates of 46.59% and 45.71%, respectively,and the expression of the tumor proliferation marker KI67 was significantly reduced. Conclusion ABF demonstrates promising anti-NPC potential, providing scientific evidence for the traditional anti-cancer use of SDG. This molecule may serve as a natural lead compound for NPC growth inhibitors.