To determine the significance of inter and intra-individual variances of the gut microbiome in a homogenous cohort of cancer patients, and to define mucosal ecological stability with advancing cancer stage.. Microbiome metabonome interactions in colon cancer

Cancerous colonic mucosa maintains niche specific bacterial networks, however, these have yet to be fully defined and their key metabolic functions remain unclear. We aimed to determine the significance of inter and intra-individual variances of the gut microbiome in a homogenous cohort of cancer patients, and to define mucosal ecological stability with advancing cancer stage. In order to meet this aim a prospective, observational study in a cohort of patients undergoing elective surgery for colon cancer (n=20), without mechanical bowel preparation was performed at a single centre. Matched samples from tumour centre and healthy mucosa (5 and 10cm away from tumour) were acquired by a pathologist and stored at -800C. DNA was extracted from samples, and the V1-V3 region of the 16S rRNA gene sequences were sequenced on a 454 Genome Sequencer FLX Titanium platform (Roche Diagnostics and Beckman Coulter Genomics). Data was analysed by Mothur to determine OTU abundance per sample and further analysed in R. Metabonomic analysis was performed by 400MHz 1H magic angle spinning nuclear magnetic resonance (MAS-NMR) spectroscopy, and multivariate analysis was performed in Matlab. The median age was 76 (55-85), M/F 10:8, and a mean BMI of 26.6 (21-39). Partial least squares discriminant analysis (PLS-DA) of OTU data was unable to discriminate between biopsy distances. Inter-individual variation had a more significant impact on microbiome expression than the distance of the normal associated mucosal biopsy from the tumour site. Despite this, microbiome ecology is UICC (TNM) stage specific and was strongly associated with histological features of poor prognosis. Fusobacteria (p<0.007) and e- Proteobacteria (p<0.01) were enriched on tumour when compared to tissue at 5 and 10cm, and fusobaceria and ß-Proteobacteria levels increased with advancing cancer stage (p<0.014 and 0.002 respectively). Metabonomic analysis demonstrated that taurine, isoglutamine, choline, lactate, phenylalanine, tyrosine demonstrated increased concentrations in tumour tissue and lipids and triglycerides decreased in concentration. Network analysis revealed that passenger microbiota modify the CRC mucosal metabonome, but that pathobionts associated with poor prognostic features were not statistically important. The cancer mucosal microbiome is individualized, and evolves with cancer stage to meet the demands of cancer metabolism. Passenger microbiota in the CRC microbiome may play a role in the maintenance of mucosal metabolic homeostasis.