Longitudinal Profiling of Circulating Tumour DNA for Tracking Tumour Dynamics in Pancreatic Cancer

Background: The utility of circulating tumour DNA (ctDNA) for longitudinal tumour monitoring in pancreatic ductal adenocarcinoma (PDAC) has not been explored beyond mutations in the KRAS proto-oncogene. Here, we aimed to characterise and track patient-specific somatic ctDNA variants, to assess longitudinal changes in disease burden and explore the landscape of actionable alterations. Methods: We followed 3 patients with resectable disease and 4 patients with unresectable disease, including 4 patients with >= 3 serial follow-up samples, of whom 2 were rare long survivors (>5 years). We performed whole exome sequencing of tumour gDNA and plasma ctDNA (n =20) collected over a similar to 2-year period from diagnosis through treatment to death or final follow-up. Plasma from 3 chronic pancreatitis cases was used as a comparison for analysis of ctDNA mutations. Results: We detected > 55% concordance between somatic mutations in tumour tissues and matched serial plasma. Mutations in ctDNA were detected within known PDAC driver genes (KRAS, TP53, SMAD4, CDKN2A), in addition to patient-specific variants within alternative cancer drivers (KRAS, HRAS, MTOR, ERBB2, EGFR, PBRM1), with a trend towards higher overall mutation loads in advanced disease. ctDNA alterations with potential for therapeutic actionability were identified in all 7 patients, including DNA damage response (DDR) variants co-occurring with hypermutation signatures predictive of response to platinum chemotherapy. Longitudinal tracking in 4 patients with follow-up >2 years demonstrated that ctDNA mutant allele fractions and clonal trends were consistent with CA19-9 measurements and/or clinically reported disease burden. The estimated prevalence of'stem clones'was highest in an unresectable patient where changes in ctDNA dynamics preceded CA19-9 levels. Longitudinal evolutionary trajectories revealed ongoing subclonal evolution following chemotherapy. Conclusion: These results provide proof-of-concept for the use of exome sequencing of serial plasma to characterise patient-specific ctDNA profiles, and demonstrate the sensitivity of ctDNA in monitoring disease burden in PDAC even in unresectable cases without matched tumour genotyping. They reveal the value of tracking clonal evolution in serial ctDNA to monitor treatment response, establishing the potential of applied precision medicine to guide stratified care by identifying and evaluating actionable opportunities for intervention aimed at optimising patient outcomes for an otherwise intractable disease.