Epstein-Barr Virus-Mediated Metabolic Reprogramming In Cancer Cells And Its Molecular Basis

Metabolism is the set of life-sustaining chemical transformations within the cells of living organisms, which allows organisms to grow and reproduce, maintains their structures, and responds to their environments. Metabolism is usually divided into two categories. Catabolism breaks down organic matter, for example to harvest energy in cellular respiration. Anabolism uses energy to construct components of cells such as proteins and nucleic acids. Study on cellular metabolism not only help us to understand the physiological process, but also deepen our knowledge about complex disease.Carcinogenesis is a long-term, multifactorial and multistep process. During this special period, cancer cells evolve and gain abnormal metabolic pattern to sustain the uncontrollable proliferation, metastasis and other malignant phenotypes of tumor cells. At the very beginning of last century, one of the most important abnormal metabolic change in cancer was observed by a Germany biochemist Otto Warburg, which is called Warburg effect. The Warburg effect is the observation that most cancer cells predominantly produce energy by a high rate of glycolysis followed by lactic acid fermentation in the cytosol, rather than by a comparatively low rate of glycolysis followed by oxidation of pyruvate in mitochondria as in most normal cells. The latter process is aerobic (uses oxygen). Malignant, rapidly growing tumor cells typically have glycolytic rates up to200times higher than those of their normal tissues of origin; this occurs even if oxygen is plentiful. From then, with the great development of cancer metabolism and improvement metabolomics platform, scientists could much more systematic research the metabolic changes during carcinogenesis, and greatly enlarge this field.More and more evidences showed that multiple metabolic pathways were changed in cancer cells, including glycolysis, oxidative phosphorylation, amino acid metabolism, fatty acid metabolism and nucleic acid metabolism. Thus, scientists called this phenomenon as cancer cell metabolic reprogramming. Now it’s widely accepted that cancer cell metabolic reprogramming is the consequence of deregulation of intercellular signaling network, and also it’s the biochemical basis of malignant phenotypes. In2011, metabolic reprogramming has been recognized as the seventh hallmark of cancer by Professor Robet A. Weinberg from Whitehead Institute.Epstein-Barr Virus (EBV) is a hematological malignancies and epithelial malignancies-related oncogenic DNA virus, which has been demonstrated at the1960s. To sustain the EBV latent infection in host cells, multiple intercellular signaling pathways were disrupted by EBV encoded products, and one of the most important EBV encoded oncogene is latent membrane protein1(LMP1). It has been well demonstrated that LMP1could regulate the proliferation, differentiation, transformation and apoptosis via activating NF-kappaB、 PI3-K/Akt and MAPK signaling pathways. Our previous study showed that the lactate concentration in nasopharyngeal carcinoma (NPC) patients’serum is significantly higher than health population’s. And also, PI3K/Akt signaling pathway, which could be activated by LMP1, is the hub of cancer metabolic reprogramming. All of these clues implanted that EBV may induce metabolic reprogramming via its encoded products.To study the EBV or LMP1-mediated cancer metabolic changes, we performed a high-throughput metabolomics analysis in EBV-infected and LMP1-expressing in normal nasopharyngeal epithelial cells and NPC cells, and got a whole EBV or LMP1-mediated metabolic changes by using. We found that EBV and LMP1could influence glucose metabolism, oxidative phosphorylation and fatty acid metabolism in NPC cell lines. Furthermore, we found that glycolysis is the most significantly changed metabolic pathways by EBV and LMP1. Biochemical analysis confirmed that EBV and LMP1can significantly induce glycolysis in NPC cells.To clarify the molecular basis of LMP1-elevated glycolysis, we screened the expression of several glucose metabolism-related genes in LMP1-expressing and their parental cells. We found that glucose transporter1(glutl), hexokinase2(hk2) and lactate dehydrogenase A (Idha) are crucial for LMP1-mediated abnormal glucose metabolism. Next, we demonstrated that LMP1-elevated HK2not only promote glycolysis and proliferation in cancer cells, but also help cancer cells antagonizing apoptosis. Knock-down of HK2by shRNA or LMP1targeting DNAzyme strikingly induced apoptosis in LMP1-expressing NPC cells. Finally, by using PI3K/Akt small specific inhibitor, we conclused that PI3K/Akt-HK2signaling axis is crucial for LMP1-elevated glycolysis.To further investigate the clinical significance of LMP1-mediated overexpression of HK2, we detect the expression of LMP1and HK2in clinical specimens. We confirmed that HK2was overexpressed along with LMP1and its overexpression was associated with a worse clinical outcome of radiation therapy. Then, radiosensitivity assay was performed to evaluate radiosensitivity of NPC cells after stable knockdown of HK2. Our data suggested that knockdown of HK2could sensitize LMP1-overexpressing NPC cells to irradiation and thus improve the treatment outcome. These findings implicate HK2may sever as a potential target of radiation therapy of NPC.In summary, this study illustrates the interaction between metabolic changes and EBV infection. The ability of LMP1to induce glycolytic enzymes contributes to the significance of EBV in cellular energy metabolism, especially in glucose metabolism in NPC cells. HK2was suggested to promote malignant proliferation, inhibit apoptosis and especially causes resistance of NPC cells to radiation therapy, leading to the poor overall survival of NPC patients. With these discoveries, we could more fully understand infection-mediated carcinogenesis and provide a novel target to improve the therapeutic regimen of NPC.