Integrating m5C methylation sequencing and transcriptome profiling to investigate key genes and molecular mechanisms of ginsenoside Rg1 intervention in head and neck cancer

Head and neck cancer (HNC), characterized by aggressive progression and poor prognosis, remains a critical clinical challenge. In this study, we pioneered the integration of m5C methylome and transcriptome profiling to decipher the therapeutic mechanism of ginsenoside Rg1 in HNC. Using FaDu cells treated with 200 µg/mL Rg1, we systematically investigated the antitumor mechanisms of ginsenoside Rg1 in HNC through an integrative multi-omics approach combining m5C methylation profiling and transcriptome sequencing. Additionally, immune-related analyses and molecular docking were conducted, and cellular heterogeneity was explored at the single-cell level. Three key genes were identified, THBS1, COL12A1 and ADAMTS1, exhibiting hypomethylation-high expression patterns. Gene set enrichment analysis (GSEA) revealed their robust involvement in important biological processes such as DNA replication, ribosome biogenesis, and lysosomal function. Immune infiltration analysis demonstrated Rg1-mediated promotion of naïve B cells and uncovered ICOSLG and PDCD1LG2 as novel immunotherapeutic biomarkers. Furthermore, molecular docking verified stable binding between ginsenoside Rg1 and each of the three key genes. Single-cell analysis showed that tissue stem cells and endothelial cells were highly expressed at the single-cell level. And Rg1 may promote the differentiation of tissue stem cells toward hematopoietic lineages, leading to an increased pool of naïve B cells. Collectively, our findings show that ginsenoside Rg1 reduces HNC cell viability and suppresses FaDu cell migration and invasion, while being associated with coordinated changes in m5C enrichment and gene expression. THBS1, COL12A1, and ADAMTS1 emerged as key candidate genes, providing a basis for further mechanistic and preclinical investigation.