Targeting AMPK, mTOR and β-Catenin by Combined Metformin and Aspirin Therapy in HCC: An Appraisal in Egyptian HCC Patients

Background Hepatocellular carcinoma (HCC) is an expanding health problem with a great impact on morbidity and mortality, both in Egypt and worldwide. Recently, metformin and aspirin showed a potential anticancer effect on HCC, although the mechanism of this effect is not fully elucidated. Objective The current work aimed to investigate the possibility of targeting AMP-activated protein kinase (AMPK), mammalian target of rapamycin (mTOR), and beta-catenin proteins through combined metformin/aspirin treatment in the HepG2 cell line, and to explore such molecular targets in Egyptian HCC patients. Materials and Methods HepG2 cells were exposed to increasing concentrations of metformin, aspirin and combined treatment, and an MTT assay was performed to determine half maximal inhibitory concentration (IC50). Caspase-3 activity, cell cycle analysis, and protein expression of AMPK, phosphorylated AMPK (pAMPK) and mTOR proteins were assessed. Furthermore, the expression and localization of beta-catenin protein was assessed by immunocytochemistry, and protein expression of pAMPK, mTOR and beta-catenin was assessed in Egyptian HCC and cirrhotic tissue specimens. Results Metformin/aspirin combined treatment had a synergistic effect on cell cycle arrest at the G2/M phase and apoptosis induction in a caspase-dependent manner via downregulation of pAMPK and mTOR protein expression. Additionally, metformin/aspirin combined treatment enhanced cell-cell membrane localization of beta-catenin expression in HepG2 cells, which might inhibit the metastatic potential of HepG2 cells. In Egyptian HCC specimens, pAMPK, mTOR and beta-catenin proteins showed a significant increased expression compared with cirrhotic controls. Conclusions Targeting AMPK, mTOR and beta-catenin by combined metformin/aspirin treatment could be a promising therapeutic strategy for Egyptian HCC patients, and possibly other HCC patients.