Combining sorafenib with spermine and sphingosine synergistically enhances anticancer efficacy by modulating metabolic pathways and gut microbiome in hepatocellular carcinoma

Hepatocellular carcinoma (HCC), which makes up about 90% of liver cancer, is the third leading cause of cancer-related death. Recent studies suggest that metabolites derived from the gut microbiome may offer new therapeutic opportunities for HCC. In this study, we explored whether microbial metabolites could enhance the effectiveness of sorafenib, a first-line multi-kinase inhibitor used in advanced HCC. Through a screen of a microbiome metabolite library, we identified spermine and sphingosine as potential candidates that boosted anticancer effects of sorafenib in HepG2, Huh7, and SK-Hep-1 cells. These metabolites worked synergistically with sorafenib to suppress tumor growth in cultured HCC cells, patients-derived HCC organoids, and a xenograft mouse model. Mechanistically, spermine triggered cell cycle arrest at the S phase, while sphingosine and sorafenib induced G1 arrest, contributing to an increased sub-G1 population and apoptosis when combined. Notably, sorafenib treatment led to the downregulation of SMOX (a key catabolic enzyme for spermine), as well as SPHK1 and CERS1 (critical enzymes involved in sphingosine metabolism), whose high expression levels are associated with poorer survival outcomes in liver cancer patients according to TCGA data. A 16S rRNA sequencing analysis revealed that combination of sorafenib with spermine or sphingosine alters the gut microbiome, increasing the relative abundance of Faecalibaculum, inversely correlated with tumor sizes in a xenograft mouse model. Therefore, we propose that combining sorafenib with spermine or sphingosine could enhance its anti-HCC effects by promoting apoptosis and reducing the expression of metabolic enzymes. Moreover, Faecalibaculum may serve as a potential microbiome-based prognostic marker for HCC.