Reprograming of Glucose Metabolism by Zerumbone Suppresses Hepatocarcinogenesis

Hepatocellular carcinoma (HCC) is the most prevalent and highly aggressive liver malignancy with limited therapeutic options. Here, the therapeutic potential of zerumbone, a sesquiterpene derived from the ginger plant Zingiber zerumbet, against HCC was explored. Zerumbone inhibited proliferation and clonogenic survival of HCC cells in a dose-dependent manner by arresting cell at the G2/M phase, and inducing apoptosis. To elucidate the underlying molecular mechanisms, a phosphokinase array was performed that showed significant inhibition of the PI3K/AKT/mTOR and STAT3 signaling pathways in zerumbone treated HCC cells. Gene expression profiling using microarray and analysis of microarray data using Gene Set Enrichment Analysis (GSEA) and Ingenuity Pathway Analysis (IPA) revealed that zerumbone treatment resulted in significant deregulation of genes regulating apoptotic, cell cycle and metabolism. Indeed, tracing glucose metabolic pathways by growing HCC cells with 13C-glucose and measuring extracellular and intracellular metabolites by 2D-NMR showed a reduction in glucose consumption and reduced lactate production, suggesting glycolytic inhibition. Additionally, zerumbone impeded shunting of glucose-6-phosphate through the pentose-phosphate-pathway, thereby forcing tumor cells to undergo cell cycle arrest and apoptosis. Importantly, zerumbone treatment suppressed subcutaneous and orthotopic growth and lung metastasis of HCC xenografts in immune-compromised mice. In conclusion, these findings reveal a novel and potentially effective therapeutic strategy for HCC using a natural product that targets cancer cell metabolism. MHCC-LM3 cells were treated with Zerumbone or DMSO as a control. Total RNA from the MHCC-LM3 cells was isolated using TRIzol (Invitrogen) and purified using RNeasy Mini columns (QIAGEN), and the integrity and quantity of the RNA were assessed using an Agilent Bioanalyzer and Nanodrop RNA 6000, respectively. Total RNA was labeled using the Affymetrix Whole Transcript Sense Target Labeling kit and hybridized to the Affymetrix human Exon 2.0 ST array following the manufacturer’s protocol at the Microarray Shared Resource Facility at the Ohio State University Comprehensive Cancer Center.