Development of a Novel Liquid Biopsy for Colorectal Cancer in Symptomatic Patients with Elevated Faecal Haemoglobin

Colorectal cancer (CRC) is the fourth most common cancer in the UK and remains a significant cause of cancer-related death. However, diagnosis can be challenging as symptoms are a poor predictor of underlying CRC or any significant bowel disease. Faecal immunochemical testing, which identifies blood in the stool, has become the cornerstone of referral pathways for suspected CRC. The data in this thesis show that faecal immunochemical test (FIT) requesting has increased significantly since its introduction, without significant change in the distribution of faecal haemoglobin (f-Hb) results. This has led to an increase in the number of patients with the highest level of f-Hb, >400 μg Hb/g faeces, who have the greatest risk of significant bowel disease (SBD) and are prioritised for colonoscopy. However, the yield of pathology in this group has decreased significantly. This thesis aimed to develop a liquid biopsy blood test for the identification of CRC prior to colonoscopy, potentially allowing the prioritisation of waiting lists. The prospective study was designed to be translational to NHS workflows and incorporated validation of new laboratory procedures at each stage. Cell-free DNA was extracted from a development cohort consisting of individuals with CRC and controls with high f-Hb but benign or absent bowel disease. Overall, using a combination of next-generation sequencing of oncogenic hotspots and methylation-specific quantitative PCR of two genes, approximately one in five CRC cases could be identified prior to colonoscopy. At present, these analyses of cell-free DNA are at the limit of capability of currently available technology in the NHS setting. Further investment in technology and staffing will be required to bring liquid biopsy into mainstream use. Finally, total proteome analysis was performed in a subset of samples to identify differentially expressed proteins in CRC. Several promising targets have been identified for confirmation by immunoassay.