Hepatocellular carcinoma (HCC) is the primary cause of cancer-related mortality in the world. Cytochrome P450 2E1 (CYP2E1) plays a key role in metabolizing xenobiotics and toxic substances. The attempts to target CYP2E1 with inhibitors have yielded limited success in cancer treatment. Loss and gain-of-function experiments were performed to investigate the effects of CYP2E1 on HepG2 and Hepa 1-6 cells. Migration, invasion, intracellular reactive oxygen species (ROS) accumulation, and mitochondrial membrane potential were assessed in HepG2 cells. In vivo histopathological alterations were studied in DEN-administered rat models. Western blotting was performed to evaluate the mechanistic activation of the Wnt/β-catenin signaling pathway. In our pursuit, to elucidate the impact of CYP2E1 in carcinogenesis we revealed that in vitro, overexpression of CYP2E1 enhances the migration, invasion, increases the accumulation of intracellular ROS, and reduces the loss of mitochondrial membrane potential of HepG2 cells. In contrast, siRNA-mediated silencing of CYP2E1 produced the opposite effects. Histopathological analysis of liver sections showed pathological features such as clear cell foci, bile duct proliferation, inflammatory cell infiltration and nodule formation. In vitro and in vivo, mechanistic analysis demonstrated that CYP2E1 overexpression significantly activated the Wnt/β-catenin signaling pathway in HCC cells, as evidenced by increased protein levels of β-catenin, LEF, Cyclin D1 and survivin in immunoblot study. In vivo increased β catenin, LEF and GPC3 enhance the carcinogenicity as revealed in immunohistochemistry results. Altogether, the above findings indicates that CYP2E1 has a pivotal role in the pathogenesis and progression of HCC identifying it as a potential target for liver cancer treatment.