PO-269 Development of a fluorescence in situ hybridization (FISH) method for detection of intra-tumour bacteria involved in pancreatic cancer chemoresistance

Introduction Pancreatic cancer (PC) is projected to become the second leading cause of cancer-related deaths in 2020. Recent studies demonstrate that bacteria are a component of pancreatic tumour microenvironment, and they may also play a critical role in mediating resistance to chemotherapy.1 Remarkably, a prospective study showed that the presence of two oral pathogens, Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans, was associated with an elevated risk of developing PC.2 Material and methods This study aimed at developing a novel fluorescence in situ hybridization (FISH) based method to specifically detect oral bacteria in formalin-fixed paraffin-embedded (FFPE) sections from PC patients. Multi-wavelength confocal spinning disk microscopy was used to discriminate with high spatial resolution and sensitivity the tissue high autofluorescence signal from even low number of emitters/bacteria. Bacterial FISH probes were primers for the 16 s rRNA molecule. Results and discussions FISH allowed the in situ localisation and the study of spatial organisation of bacterial cells as they occur in PC. FISH results were always definitive and well-detectable both in in 4 and 20 µm FFPE sections. The intensity of the fluorescence was a direct measure for the activity of the bacterial cells, whereas inactive cells were recognised by their low intensity fluorescence. Thus, our method gives a optimal sensitivity of fluorescence and low background by subtracting autofluorescence components. Our FISH preliminary data confirmed the presence of bacteria in the pancreatic tumour microenvironment and support further studies aimed at elucidating how oral bacteria are involved in PC aetiology. It is well-known that the chemoresistance (intrinsic and acquired) has been a major problem for PC treatment3 and a recent study showed the role of bacteria in resistance to gemcitabine.1 To this end, the establishment of our reliable FISH protocol allows us to further investigate the potential role of oral bacteria in the failure of standard treatments for PC. Conclusion Our new robust methodology will contribute to obtain meaningful data and to unravel the complex interaction between oral bacteria and pancreatic cancer chemoresistance in clinically well-annotated tissues, ultimately identifying specific bacteria as potential biomarkers for early detection/treatment. References Geller, et al. Science2017;357:1156–1160. Farrell JJ, et al. Gut2012; 61:582–588. Coppola, et al. Drug Resist. Updates 2017;31:43–51.