Abstract: Enolase 1 (ENO1), a key enzyme in the glycolytic pathway, catalyzes the conversion of 2-phosphoglycerate (2-PG) to phosphoenolpyruvate (PEP). As a multifunctional protein, ENO1 is expressed on the cell surface and in the cytoplasm. Its role in glycolysis extends beyond metabolic regulation to promoting tumor cell proliferation and inhibiting apoptosis. The expression of ENO1 is not regulated by growth inhibitors, thus facilitating immune escape in tumor cells. Overexpression of ENO1 in various cancer types makes it a potential target for tumor therapy, and also an important biomarker for cancer prognosis and diagnosis. This review summarizes the latest research advances in the relationship between ENO1 and cancer, explores its potential as a cancer biomarker, and discusses the potential applications of small-molecule drugs targeting ENO1 in tumor treatment.