6-Methoxyflavone inhibits glycolytic energy metabolism in HeLa cells

Enhanced glycolytic levels in cancer cells are a common characteristic of many cancer types. Modulation of glycolytic metabolism is crucial for enhancing the efficacy of cancer therapy. The specific role of 6-methoxyflavone in regulating glycolytic metabolism in cancer cells remains unclear. This study aimed to elucidate the impact of 6-methoxyflavone on glycolytic metabolism in cervical cancer cells and its clinical relevance.

The tandem mass tag (TMT) proteomic analysis was used to identify significantly enriched biological processes and pathways in HeLa cells after treatment with 6-methoxyflavone. Additionally, the differential expression of glycolysis-related proteins was validated using parallel reaction monitoring (PRM) proteomics. Untargeted and targeted metabolomics analyses were used to identify differentially expressed glycolysis-related metabolites. Furthermore, alternative splicing, new transcripts, and domain analyses were used to detect the effects of 6-methoxyflavone on the structures of glycolysis-related genes and proteins. Subcellular localization, molecular docking, and non-covalent interaction analyses were used to detect the subcellular localization, affinity of 6-methoxyflavone for glycolysis-related proteins, and sites of non-covalent interactions. Clinical characteristics and immunological correlation analyses were used to elucidate the relationships between glycolysis-related genes and clinicopathological characteristics, survival, prognosis, and immune-related indicators of patients with cervical cancer. Finally, glycolysis stress tests and enzyme activity assays were used to verify the effect of 6-methoxyflavone on glycolysis in HeLa cells.

TMT and PRM proteomics, as well as untargeted and targeted metabolomics results, showed that 6-methoxyflavone downregulated the expression levels of glycolysis-related proteins and metabolites in HeLa cells, and that the structures and functions of glycolysis-related genes and proteins in the cytoplasm underwent changes. 6-Methoxyflavone had a good affinity for nine glycolysis-related proteins, all of which had non-covalent interaction sites. Clinical characteristics and immune correlation analyses showed relationships between 6-methoxyflavone and five clinical characteristics, survival prognosis, and four immune-related indicators in patients with cervical cancer. After treatment with 6-methoxyflavone, the basal glycolytic level, maximum glycolytic capacity, and glycolytic reserve of HeLa cells were downregulated. Additionally, 6-methoxyflavone inhibited the activity of pyruvate kinase.

6-Methoxyflavone inhibited energy metabolism in HeLa cells through the glycolysis pathway. 6-Methoxyflavone may be related to five clinical characteristics, prognosis, tumor microenvironment, immune cells, immune checkpoints, and immunotherapy efficacy in patients with cervical cancer.