ku-0063794 and Liver-Neoplasms

It is challenging to overcome the low response rate of everolimus in the treatment of patients with hepatocellular carcinoma (HCC). To overcome this challenge, we combined everolimus with Ku0063794, the inhibitor of mTORC1 and mTORC2, to achieve higher anticancer effects. However, the precise mechanism for the synergistic effects is not clearly understood yet. To achieve this aim, the miRNAs were selected that showed the most significant variation in expression according to the mono- and combination therapy of everolimus and Ku0063794. Subsequently, the roles of specific miRNAs were determined in the processes of the treatment modalities. Compared to individual monotherapies, the combination therapy significantly reduced viability, increased apoptosis, and reduced autophagy in HepG2 cells. The combination therapy led to significantly lower expression of miR-4790-3p and higher expression of zinc finger protein225 (ZNF225)-the predicted target of miR-4790-3p. The functional study of miR-4790-3p and ZNF225 revealed that regarding autophagy, miR-4790-3p promoted it, while ZNF225 inhibited it. In addition, regarding apoptosis, miR-4790-3p inhibited it, while ZNF225 promoted it. It was also found that HCC tissues were characterized by higher expression of miR-4790-3p and lower expression of ZNF225; HCC tissues were also characterized by higher autophagic flux. We, thus, conclude that the potentiated anticancer effect of the everolimus and Ku0063794 combination therapy is strongly associated with reduced autophagy resulting from diminished expression of miR-4790-3p, as well as higher expression of ZNF225.

Topics: Antineoplastic Agents; Apoptosis; Autophagy; Carcinoma, Hepatocellular; Cell Line, Tumor; Cell Survival; Drug Synergism; Early Growth Response Protein 1; Enzyme Inhibitors; Everolimus; Gene Expression Regulation, Neoplastic; Hep G2 Cells; Humans; Liver Neoplasms; Mechanistic Target of Rapamycin Complex 1; Mechanistic Target of Rapamycin Complex 2; MicroRNAs; Morpholines; Pyrimidines

Everolimus only inhibits mammalian target of rapamycin complex 1 (mTORC1), whereas Ku0063794 inhibits both mTORC1 and mTORC2. Although they have similar anticancer effects, their combination has a synergistic effect against hepatocellular carcinoma (HCC) cells. We aimed to determine the mechanism underlying the synergistic effects of everolimus and Ku0063794 associated with autophagy in HCC cells.. We compared the effects of everolimus and Ku0063794, individually or in combination, on both the in vitro and in vivo models of HCCs.. HepG2 cells treated with both agents had significantly lower rates of cell proliferation and higher apoptosis than the individual monotherapies (p < 0.05). Autophagic studies consistently indicated that, unlike the monotherapies, the combination therapy significantly reduced autophagy (p < 0.05). Autophagic blockage directly promoted the pro-apoptotic effects of combination therapy, suggesting autophagy as the survival mechanism of HCC cells. Unlike the monotherapies, combination therapy showed the potential to inhibit sirtuin 1 (SIRT1), the positive regulator of autophagy. SIRT1 overexpression abrogated the autophagy-inhibiting and pro-apoptotic effects of combination therapy. In a nude mouse xenograft model, the shrinkage of tumors was more prominent in mice treated with combination therapy than in mice treated with the respective monotherapies (p < 0.05). The immunohistochemical and immunofluorescence stains of the tumor obtained from the xenograft model showed that combination therapy had the potential of reducing autophagy and promoting apoptosis.. The combination of everolimus and Ku0063794 potentiates anticancer effects on HCCs through a decrease in autophagy, which is prompted by SIRT1 downregulation.

Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Autophagy; Carcinoma, Hepatocellular; Cell Proliferation; Drug Synergism; Everolimus; Gene Expression Regulation, Neoplastic; Hep G2 Cells; Humans; Liver Neoplasms; Mice; Morpholines; Pyrimidines; Sirtuin 1; Up-Regulation; Xenograft Model Antitumor Assays