While hepatocellular carcinoma (HCC) is the fifth most commonly diagnosed cancer worldwide, the signaling pathways involved are not fully understood and treatment of advanced disease still represents an area of high unmet medical need. We previously reported that IQ-motif-containing GTPase-activating-like proteins IQGAP1 and IQGAP2 play opposing roles in hepatic carcinogenesis. IQGAP2 was identified as a novel tumor suppressor linked to the Wnt/β-catenin signaling pathway in HCC. In this study, mouse embryonic fibroblasts (MEFs) isolated from Iqgap2−/- mice displayed higher proliferation and migration rates compared to wild-type MEFs as evident by MTT proliferation assay and wound healing assay. In contrast to HepG2, the SK-Hep1 and SNU387 HCC cell lines are characterized by low expression of IQGAP2 and relatively high levels of IQGAP1. Migration assay using blind well chambers with polycarbonate filters revealed 2-fold increased motility for both SK-Hep1 and SNU387 compared to HepG2 (p < 0.05). HepG2 cells stably expressing IQGAP2-specific shRNA showed an increase in their motility. Conversely, overexpression of IQGAP2 in HEK293 resulted in up to 50% inhibition of their migration rate. qRT-PCR of naïve SK-Hep1 and SNU387 cells and HepG2 cells overexpressing IQGAP2 shRNA confirmed upregulation of genes encoding MET, MMP1, MMP2, MTA2 and TWIST1, reinforcing a role of IQGAP2 in cell migration. Akt kinase, one of the key regulators of cell migration, co-immunoprecipitated with IQGAP2 in both mouse liver lysates and HEK293 cells transiently expressing human IQGAP2. IQGAP2, but not IQGAP1, also co-immunoprecipitated with GSK3β kinase in mouse liver lysates. Furthermore, to address whether IQGAP2 affects Akt function in liver, insulin stimulation experiments were performed with wild-type, Iqgap2−/- and Iqgap1−/−/Iqgap2−/- mice. It was shown that upon stimulation with a single intraperitoneal injection of 5 U/kg insulin, levels of the phosphorylated (at Ser9) form of GSK3β were elevated in both Iqgap2−/- and Iqgap1−/−/Iqgap2−/- livers compared to wild-type, suggesting that in IQGAP2-deficiency, GSK3β kinase activity is inhibited by, perhaps, Akt activation. Additionally, the strength of the GSK3β - β-catenin interaction was found to be dependent on the presence of both IQGAP1 and IQGAP2, with Iqgap2−/- livers showing weaker interaction and Iqgap1−/−/Iqgap2−/- livers the weakest compared to wild-type. Using IQGAP2 deletional mutants, it was determined that IQ domains may be primarily responsible for IQGAP2 interactions with both Akt and GSK3β. While it remains unclear whether IQGAP2 binds Akt and GSK3β directly or as part of a larger protein complex, these findings suggest that tumor suppressive function of IQGAP2 in the liver may be realized through inhibition of cell proliferation and migration.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3989. doi:1538-7445.AM2012-3989