Evaluation of the effect of biochanin b and Sorafenib on cell survival, growth, migration, colony formation and apoptosis in hepatocellular carcinoma

Abstract Background and Purpose: Formononetin is a phytoestrogen isolated from several medicinal plants, and it is extensively investigated for its anti-cancer effects in several cells. However, the impact of formononetin on hepatocellular carcinoma (HCC) has been unclear. In this study, the influence of formononetin on cell growth, metabolism, and signaling molecules is examined in the HepG2 cells, singly and in combination with sorafenib. This study investigates the individual and combined effects of formononetin and sorafenib on growth, metabolism, and signaling in HepG2 cells, focusing on miR-21 regulation and theirdownstream effects for the first time. This study underscored the dependent effects of the two drugs on the expression of miR-21. The drugs, either alone or together, indirectly affect the expression of other genes via miR-21 as well as directly. this study aiming to explore its molecular mechanisms and potential therapeutic values. Materials and Methods: Materials and Methods: HepG2 cells were treated with formononetin, sorafenib, or a combination of both. cell growth, colony formation, migration, apoptosis, gene expression, and protein contents were investigated. Formononetin (1-640 µM) and sorafenib (0.01-64µM) or itscombination (0.01µM sorafenib+1µM formononetin) were tested. Results: After 48 h, treatments reduced the cell viability and colony formation and induced apoptosis. miR-21 expression was significantly lowered by combination therapy which demonstrated the most potent inhibitory effect on EGFR, VEGF-A, MMP-9 expression, increasing BAD expression with the highest levels in the combination group, then sorafenib and followed by formononetin. Isolated compounds or their combination caused a reduction in survivin expression levels, with the least effect for formononetin and the most notable effect for combination treatment. Formononetin produced a more considerable reduction in the expression of EGFR and VEGF-A thansorafenib, but their combination caused the most prominent reductions. Conclusion Formononetin is a promising compound as an adjuvant therapy for HCC, especially when used in combination with sorafenib. The combination therapy not only enhances the inhibition of key oncogenic pathways involved, such as EGFR, VEGF-A, and MMP-9, but also induces pro-apoptotic machinery by downregulating miR-21 and upregulating BAD expression. All these data suggest that formononetin, in combination with sorafenib, may present a more effective therapeutic approach againstHCC. More in-depth preclinical and clinical studies are needed to support the findings and establish its clinical applicability.