In a prospective cohort of pancreatic cancer patients, we show how longitudinal monitoring using liquid biopsy samples through exoDNA and ctDNA provides both predictive and prognostic information relevant to therapeutic stratification.

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Background: Prognostic and therapeutic stratification of pancreatic ductal adenocarcinoma (PDAC) patients remains elusive due to a lack of effective biomarkers, and a predilection towards metastatic disease. Peripheral blood-based liquid biopsies for tumor markers has emerged as a potential minimally invasive strategy for tumor monitoring. We have implemented a liquid biopsy assay utilizing DNA derived from vesicles known as exosomes (exoDNA) and circulating tumor DNA (ctDNA) in the metastatic setting to determine the potential utility of these liquid biopsy compartments in tumor management. Methods: A total of 318 plasma samples from 123 metastatic pancreatic patients were prospectively collected. ExoDNA and ctDNA were then extracted from matched plasma samples. Digital PCR, was used to identify codon 12/13 KRAS gene mutations. We assessed clinical endpoints in relation to progression free survival (PFS) and overall survival (OS) using univariate and multivariate analyses. In a metastatic PDAC patient, six serial liquid biopsies and seven tissue biopsies taken throughout disease progression underwent whole genome sequencing for detection of copy number events. Results: Detection rates of KRAS mutations in exoDNA and ctDNA at baseline treatment naïve staus were 61.0% and 52.9%, respectively. On multivariate COX regression analysis, exoDNA KRAS mutant allelic fraction (MAF) ≥ 5% was a significant predictor of poorer PFS (HR 2.28, 95% CI 1.18-4.40, P=0.014) and OS (HR 3.46, 95% CI 1.40-8.50, P=0.007) in metastatic patients. Among 34 patients, liquid biopsy tumor monitoring was performed across 127 serial blood draws during a median followup time of 11.1 months. The presence of an exoDNA MAF peak ≥ 1% during tumor monitoring was significantly correlated to radiological progression (p=0.0003). Specifically, detection of an exoDNA MAF peak ≥ 1% preceded radiological progression by a median of 50 days compared to a median of 0 days for CA19-9 (p=0.03). CtDNA did not emerge as a significant predictor of survival outcomes in our cohort. In a patient with multiple longitudinal liquid biopsies, comprehensive genomic profiling of exoDNA further demonstrated our ability to capture additional mutational events as they emerged during therapy and correlated to progression including amplifications in ERBB2 and MYC and deletions in CDKN2A and SMAD4. Conclusions: Liquid biopsies in PDAC provide direct evidence of those patients likely to experience poorer outcomes allowing for more effective therapeutic stratification. Liquid biopsies also demonstrate utility in characterization of putative emerging driver events during disease progression. Citation Format: Vincent Bernard, Dong U. Kim, F. Anthony San Lucas, Jonathan Castillo, Kelvin Allenson, Feven C. Mulu, Bret M. Stephens, Jonathan Huang, Eugene Koay, Cullen M. Taniguchi, Milind Javle, Robert A. Wolff, Matthew H. Katz, Gauri R. Varadhachary, Hector A. Alvarez, Anirban Maitra. Circulating nucleic acids as biomarkers of prognosis and chemorefractory status in metastatic pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2599.