P-268 Using 16S rRNA gene sequencing to investigate the composition of Fusobacterium-associated microbiota and its impact on outcome in rectal cancer

The field of microbiome research has improved expeditiously over the years until becoming a topic of public interest for its association with human health and diseases. Many studies suggest a complex link between the gut microbiome, immunity, and molecular alterations in colorectal tumorigenesis. Fusobacterium is increasingly recognized as having an important role in human CRC carcinogenesis. We have previously shown that persistence of Fusobacterium after neoadjuvant chemoradiotherapy (nCRT) is significantly associated with relapse in locally advanced rectal cancer (LARC). However, the microbiota composition associated with the presence of Fusobacterium is still poorly elucidated. Twenty-nine non-consecutive patients diagnosed with LARC at Vall d’Hebron University Hospital between 2007 and 2018 and with available fresh frozen surgically resected tumor tissues were included in this study. Fourteen patients received nCRT. Microbial community analysis was performed by sequencing the V3-V4 region of the 16S rRNA gene using Illumina MiSeq platform. The output obtained from the sequencing was analysed by diversity (alfa diversity with Shannon index and Beta diversity with Bray-Curtis) and taxonomic (Linear discriminant analysis (LDA)) analyses. Samples were stratified into 3 groups according to the Fusobacterium relative abundance: negative (10%, n=6). The presence and abundance of intratumoral Fusobacterium was associated with a distinct tumour microbiota composition (Shannon index, Fuso neg vs Fuso low, p=0.013; Bray-Curtis, p=0.001). Genera already described in colorectal cancer and typically found in the oral cavity such as Gemella, Solobacterium, Peptostreptococcus and Parvimonas were found enriched in Fusobacterium-positive compared to negative tumors by LDA. No significant differences in microbial diversity between treated (n=14) and untreated (n=15) tumors were observed (Shannon index p=0.43, Bray-Curtis p=0.23). Taxonomic composition of microbial communities showed that several healthy-associated species were enriched in the treated group thus, suggesting a re-establishment of a eubiotic-like condition possibly as a consequence of treatment-induced tumor shrinkage. After stratifying the treated tumors according to outcome, patients who relapsed (n=6) exhibited a significantly different microbiota composition (Bray-Curtis p= 0.007) compared to those who did not relapse (n=8). Interestingly, taxonomic analysis showed an enrichment of Fusobacterium and associated taxa in tumors from patients who relapsed as compared to those who did not. These findings point to a distinct Fusobacterium-associated microbiota potentially impacting treatment response and outcome in LARC. This knowledge may have significant impacts for the development of novel diagnostic and therapeutic strategies targeting the gut microbiome and its connection with rectal cancer.