N-Acetylneuraminate Pyruvate Lyase Promotes Cell Adaptation to Glucose Deprivation by Regulating Intracellular ATP Levels

N-acetylneuraminate pyruvate lyase (NPL) is a key enzyme in sialic acid catabolism that links sialylation to cellular metabolism, but its role in cancer cell metabolic adaptation is poorly defined. In particular, it remains unclear whether NPL contributes to ATP maintenance in hepatocellular carcinoma (HCC) under nutrient stress through its role in sialic acid catabolism. HCC is a highly lethal malignancy characterized by extensive metabolic reprogramming. Here, we investigated whether NPL links sialic acid catabolism to ATP maintenance in HCC cells under glucose deprivation. Glucose deprivation induced NPL expression in Huh7 and PLC/PRF/5 cells, which is consistent with an adaptive response to energetic stress. Stable NPL knockdown reduced intracellular ATP levels. Consistently, cell growth was significantly reduced, as assessed by Cell Counting Kit-8 (CCK-8) and colony formation assays. These effects were more pronounced in the absence of glucose. Exogenous pyruvate partially restored ATP levels and the growth inhibition caused by NPL knockdown, particularly in the absence of glucose. This rescue further suggests that NPL may support ATP maintenance under glucose deprivation partly through pyruvate metabolism. Together, these findings indicate that NPL contributes to maintaining intracellular ATP levels during glucose deprivation, thereby supporting HCC cell adaptation to metabolic stress. To extend these biological findings to potential therapeutic exploration, we performed virtual screening and molecular docking using a U.S. Food and Drug Administration (FDA)-approved drug library. Candidate compounds predicted to bind NPL were identified, providing a basis for further validation and optimization.