Twenty-four novel 14N-substituted tetrandrine (Tet) were synthesized and investigated for their effects on anti-proliferative activities against human cancer cell lines HepG2 and A549 by the MTT method. A structure-activity relationship (SAR) study disclosed that the introduction of heterocyclic group on 14N atom might be beneficial for both activity and safety. Compound 17 exhibited the strongest cytotoxic effect against HepG2 cells with an IC50 value of 2.09 μM and satisfactory SI value of 11.5, which was 5.3- and 6.4-fold than the activity of parental tetrandrine and adriamycin, respectively. Further in vitro experiments (flow cytometry assay, transwell assay, and scratching assay) were designed to validate the significant therapeutic effect of representative compound 17 on HCC. Network pharmacology was used to examine the mechanism of 17 in HCC treatment and the AKT signaling pathway was identified as the key pathway. Molecular docking, cellular thermal shift assay (CETSA) and ADP-Glo™ kinase enzyme inhibition assay experiment further demonstrated that 17 could specifically bind to AKT and inhibit its phosphorylation, then activated the proapoptotic protein BAX, inhibited the anti-apoptosis protein Bcl-2, with activating the caspase 3, so that to induce apoptosis. Moreover, it indicating a potent in vivo killing effect against liver cancers orthotopically transplanted HCC in AKT1-dependent manner with safety profile. Taken together, these results highlighted that compound 17 induced apoptosis, and provided powerful information for developing tetrandrine derivatives into a novel class of anti-cancer agents in liver cancer therapy in the future.

Twenty-four novel 14N-substituted tetrandrine (Tet) were synthesized and investigated for their effects on anti-proliferative activities against human cancer cell lines HepG2 and A549 by the MTT method. Compound 17 exhibited the strongest cytotoxic effect against HepG2 cells with an IC 50 value of 2.09 μM and satisfactory SI value of 11.5, which was 5.3- and 6.4-fold than the activity of parental tetrandrine and adriamycin, respectively. Further in vitro experiments (flow cytometry assay, transwell assay, and scratching assay) were designed to validate the significant therapeutic effect of representative compound 17 on HCC. Network pharmacology was used to examine the mechanism of 17 in HCC treatment and the AKT signaling pathway was identified as the key pathway. Molecular docking, cellular thermal shift assay (CETSA) and ADP-Glo™ kinase enzyme inhibition assay experiment further demonstrated that 17 could specifically bind to AKT and inhibit its phosphorylation, then activated the proapoptotic protein BAX, inhibited the anti-apoptosis protein Bcl-2, with activating the caspase 3, so that to induce apoptosis. Moreover, it indicating a potent in vivo killing effect against liver cancers orthotopically transplanted HCC in AKT1-dependent manner with safety profile. Taken together, these results highlighted that compound 17 induced apoptosis.