Cell-free DNA genotyping using digital PCR for early detection of pancreatic neoplasm.

TPS464 Background: Pancreatic ductal adenocarcinoma (PDA) is still a dismal disease, and there is an urgent need to establish novel tool for early diagnosis of the tumor. There are two main types of pathologically and genetically distinct precursors for PDA — pancreatic intraepithelial neoplasia (PanIN) and intraductal papillary mucinous neoplasia (IPMN). Non-invasive markers for these precursor lesions have the potential to predict subsequent invasive tumor. Methods: Circulating cell-free DNA (cfDNA) released from tumor cells into the blood has been intensively studied as a novel way to monitor the genetic changes. To detect the cfDNA representing for the initiation and progression of PDA could be of the candidate for them. The role of cfDNA genotyping targeting the major driver mutations in these precursors, such as KRAS and GNAS, are currently under investigation in Japanese patients who have pancreatic tumors (UMIN000012810). The major technical challenge is to specifically detect the small fraction of tumor-derived DNA in patient plasma and urine. Since sequencing of target mutant alleles in cfDNA has a limitation to detect very low frequency variants, we sought to establish protocols for super-sensitive and absolute quantification of the “key drivers” for pancreatic tumor using a droplet digital PCR platform (Bio-Rad; QX200). The primary endpoint of this multi-center prospective analysis is to evaluate whether such an approach can appropriately monitor the risk of IPMN progression and detect localized early-stage PDA. Thirty cases of PDA and 90 cases of IPMN have been enrolled thus far. Detailed protocol for the study and improved technical points to quantify low-frequency variants will be discussed.