Hepatocarcinogenesis is a multi-step process involving a number of different genetic and epigenetic alterations that ultimately lead to malignant transformation of the hepatocyte. It has been recently demonstrated that concerted upregulation of both the insulin receptor (IR) and insulin-like growth factor-1 receptor (IGF-1R) are early events in hepatocarcinogenesis induced in the rat by N-Nitrosomorpholine. The small molecule IGF-1R inhibitor picropodophyllin (PPP) has been found to potently inhibit IGF-1R autophosphorylation in different cancer cell lines and xenografts with selective effect against a panel of receptor tyrosine kinases including IR. However, the precise molecular mechanism of PPP is not yet fully elucidated. The goal of the present work was to study the antitumour effects of PPP, and the underlying molecular mechanism using the HepG2 cell line, which expresses a high basal level of IGF-1R. PPP inhibited proliferation and colony formation of HepG2 cells. At low concentrations and as early as after 2 hr. of treatment, PPP increased the percentage of cells in the G2/M phase of the cell cycle. At higher concentrations (0.5 µM, 1 µM), PPP induced apoptosis measured by Terminal deoxynucleotidyl transferase mediated dUTP Nick End Labelling (TUNEL) assay. Furthermore, PPP induced downregulation of the total protein levels of IGF-1R at the 16 and 24 hr. without affecting the levels of AKT or its active form serine 473 (pAKT.S473). PPP induced the accumulation of p27 in a dose-dependent manner with no effect on the levels of Cyclin-dependent kinase 1 (CDK1) or CDK2. Taken together, the present results highlight some molecular mechanism for the multiple antitumour effects of PPP. The data provide also the rationale for further preclinical evaluation of PPP for therapeutic implications in hepatocellular carcinoma in vivo.
Key words: HepG2 cells, apoptosis, cell cycle, PPP, IGF-1R, proliferation
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