Several studies have also shown that HK may be

Several studies have also shown that HK2 may be an important downstream ARCA Cy5 EGFP mRNA mg of the PI3K/AKT/mTOR signaling pathway and may contribute to the development of cancer [15]. In our previous study, we found AKT2 expression was positively correlated with HK2 expression in primary colon cancer specimens (Spearman's R = 0.711, p < .01). However, how HK2 promotes the development and progression of colon cancer and whether it interacts directly with AKT2 remains poorly understood. In the present study, we aimed to elucidate how AKT2 mediates colon cancer invasion, tumorigenesis, and metastasis both in vitro and in vivo, and whether these effects are related to HK2.
Material and methods
Results
Discussion Although the role of AKT2 and HK2 in the development and progression of human cancer has been established [[24], [25], [26]], their specific mechanisms in the development of colon cancer remains unknown. In the present study, we found that AKT2 phosphorylates the T473 site of HK2, activates its enzymatic activity, increases intracellular lactate production, resists serum starvation-induced apoptosis, and promotes cell invasion, xenograft tumor growth, and metastasis. The overexpression of the AKT2 isoform is an early event and plays a predominant role in colon carcinogenesis. This capability of AKT2 has been attributed to the fact that AKT2, in addition to regulating cell migration and invasion, also inhibits apoptosis; however, the precise mechanism of AKT2-driven development of colon cancer is poorly understood. Recently, HK2 was been reported to be highly expressed in malignant tumors. HK2 may promote colon cancer progression via different mechanisms, of which one of the most important mechanisms may be aerobic glycolysis [28,29]. Although the correlation between HK2 and tumorigenesis has been confirmed by mouse models [30], the specific mechanism by which HK2 mediates the development of colon cancer remains unclear. Currently, some studies suggest that there may be interactions between AKT2 and HK2. It has been reported that miR-29b negatively regulates AKT2/AKT3 expression, resulting in the down-regulation of HK2/PKM2, which leads to a decrease in the Warburg effect and a slowing of ovarian cancer progression [31]. Moreover, Zhuo et al. demonstrated that the elevation of HK2 in osteosarcoma induced by activated PI3K/Akt signaling exerts anti-apoptotic and proliferative effects by regulating the Warburg effect [32]. In addition, double-stranded RNA-activated protein kinase (PKR)-like ER kinase (PERK) silencing results in decreased glioma cell viability and ATP/lactate production upon low glucose stress, which is mediated by partially blocked AKT activation and subsequent inhibition of HK2 mitochondria translocation [33]. However, how AKT2 and HK2 function together in colon cancer remains poorly understood to date. In the present study, an interaction between AKT2 and HK2 but not AKT1 or AKT3, in colon cancer cells was identified by immunoprecipitation assays. Since AKT is known to be closely related to insulin signaling and HK2 is a key rate-limiting enzyme in glycolysis, is a major subtype in insulin-sensitive tissue and is upregulated in tumors, one of the consequences of this interaction may be to alter the aerobic glycolysis metabolism of tumor cells. In addition, HK2 up-regulation results in increased glycolysis rates. In this study, we reported that the overexpression of AKT2 can simultaneously increase the expression level and phosphorylation level of HK2, enhanced the activity of cellular hexokinase, and promoted aerobic glycolysis in HCT-116 and HT-29 cells, while producing large amounts of lactic acid. A knockout of the hk2 gene following the overexpression of AKT2 can significantly reduce the hexokinase activity of the cells and the amount of lactic acid produced. Rescuing the stable overexpression of HK2 in OV AKT2Δhk2 cells significantly increased the hexokinase activity and amount of lactic acid, but the mutant HK2T473A did not. These results indicate that AKT2 contributes to increased levels of aerobic glycolysis through HK2 in colon cancer cells, can more efficiently obtain the energy required by these cells, and obtain a large number of intermediate products necessary for cellular proliferation, further promoting tumor development. This is similar to a previous report that c-Src can mediate HK2 to increase hexokinase activity and lactic acid production in tumor cells [34].