<i>CATENIBACTERIUM. MITSUOKAI</i> GENERATED QUINOLINIC ACID PROMOTES HEPATOCELLULAR CARCINOGENESIS

Background: We identified that Catenibacterium is enriched in feces and liver tissues of hepatocellular carcinoma (HCC). We aimed to examine the role and mechanism of Catenibacterium mitsuokai (the only known species in genus Catenibacterium) in hepato-carcinogenesis.
Methods: The role of C. mitsuokai in HCC formation was evaluated in mice with orthotopic injection of mouse HCC cell lines (Hepa 1-6 and RIL175). The attachment ability of C. mitsuokai on tumor cells was evaluated by cell attachment assay and transmission electron microscopy (TEM). The functional bacteria surface protein and the hepatocyte receptor were identified by biotinylation-based far-western assay. The metabolites produced by C. mitsuokai in feces, portal vein serum and liver tumor tissue samples were identified by liquid chromatography-mass spectrometry. C. mitsuokai, its conditional medium, or candidate metabolites were co-cultured with human HCC cell lines (Hep3B, Huh7, PLC5) and normal hepatocyte cell line MIHA. C. mitsuokai associated the host gene expression was identified by PCR array.
Results: Oral gavage of C. mitsuokai significantly promoted HCC formation with increased tumor volume (P<0.05) and tumor weight (P<0.05) in two orthotopic mouse models. The gut barrier was also impaired by C. mitsuokai as indicated by increased intestinal permeability and down-regulated tight junction proteins Occludin and Claudin-3 and adherent protein E-cadherin. We confirmed that C. mitsuokai could colonize in mouse colon and liver tumors by fluorescence in situ hybridization (FISH). Mechanistically, C. mitsuokai had greater favorite to adhere to HCC cells compared to normal hepatocytes, of which, C. mitsuokai through its surface protein Gtrl1/RagA attached to the receptor of HCC cells. Moreover, both C. mitsuokai and its conditional medium significantly promoted the proliferation of HCC cells. Through integrated metabolomic analysis, quinolinic acid derived from C. mitsuokai was consistently increased in the feces, portal vein serum and HCC tissues of C. mitsuokai-treated mice. Quinolinic acid significantly promoted the proliferation of HCC cells. Quinolinic acid directly bund to and activated the tyrosine kinase receptor Tie2 on HCC cells. The phosphorylated Tie2 further stimulated the oncogenic PI3K/Akt signaling pathway, thereby promoting HCC development.
Conclusion: We demonstrated that C. mitsuokai is a novel pathogenic bacterium capable of promoting hepato-carcinogenesis. The enriched C. mitsuokai colonizes in the HCC tissue through its surface protein Gtrl1/RagA. C. mitsuokai derived quinolinic acid directly bund to Tie2 of hepatocytes to activate the PI3K/Akt oncogenic pathway therefore promoting hepatocarcinogenesis.