Fusobacteria is Associated with a Th1 and Th17 Immune Microenvironment in Colon Cancer Patients and Germ-Free Mice

Sporadic colorectal cancer (CRC) remains a global health burden with a rising incidence in younger patients (<50 years), enhancing the need to identify novel targets for diagnosis and treatment. Certain T cell subsets appear to be associated with CRC prognosis; Th1 subsets, marked by interferon gamma (IFNγ), with better prognoses, and Th17, marked by interleukin‐17 (IL‐17), with worse prognoses. Several studies have shown that Fusobacteria (Fuso), specifically Fusobacterium nucleatum (Fn), are prevalent in CRC tissues despite their predominant location in the oral microbiome. Herein, we tested the hypothesis that Fuso are associated with colonic mucosal Th17 responses. First, using tissues from surgically‐resected CRC patients (tumor and paired normal flanking tissue, n=19 tissues) that were positive for Fuso by 16S ribosomal RNA gene (16S) sequencing, we analyzed tissue‐associated Th1 and Th17 cell populations by flow cytometry. Fuso 16S reads positively correlated with both IL‐17 + and IFNγ + cells in these patient tissues (R 2 =0.3, p=0.03). We next tested whether subspecies (subsp.) of Fn isolated from CRC patients would differentially impact colonic mucosal immune responses in germ‐free (GF) mice. GF mice (n=4 GF controls; n=5/subsp.) were inoculated with Fn subsp. animalis, nucleatum, vincentii , or polymorphum for 2 weeks before tissue harvest. Colonization of GF mice with bacteria typically results in decreased ceca weights, however all subsp. resulted in increased ceca weights compared to GF mice (Kruskal‐Wallis 1way ANOVA, p=0.0148). Stools (n=2/subsp.) were positive for Fuso colonization by fluorescence in situ hybridization (FISH), except for mice colonized with subsp. polymorphum . Fuso‐specific fecal 16S rRNA qPCR and colon FISH are in progress. Mice (n=3/subsp.) were analyzed for expression of inflammatory cytokines associated with CRC and/or Fn. Compared to the GF control, subsp. animalis and vincentii induced IL‐17a expression by an average fold‐change (ΔΔCt) of 1.6 and 4.1, respectively; subsp. nucleatum and polymorphum did not differ from the GF mouse control. All subsp. induced an increase in IL‐6 expression that contributes to IL‐17 T cell polarization. Similar to observations in our patient cohort, all subspecies increased IFNγ expression. In summary, these findings suggest human CRC tissue Th1 and Th17 immune responses correlate with Fuso, specifically, Fn, tissue abundance. This dual response was mimicked by select Fn subsp. in GF mice, and certain CRC‐derived Fn subsp. may be more proinflammatory than others. Further investigations into the immune‐modulating and tumorigenic potential of CRC‐associated Fn subsp. are needed. Support or Funding Information Bloomberg~Kimmel Institute for Cancer Immunotherapy; Cancer Research Institute This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .