Abstract PR04: Clonal composition and clonal selection during PanIN progression

Abstract

Abstract Pancreatic cancer is the fourth most common cause of cancer death in the United States. It remains a highly lethal malignancy despite surgical and chemotherapeutic advances. In spite of progress made delineating the mechanisms underlying pancreatic cancer, there has been little improvement in survival rates over the past fifty years. Copy number analysis of human invasive pancreatic ductal adenocarcinoma (PDAC) revealed massively rearranged cellular karyotypes as well as significant clonal complexity. Given the genetic complexity of already invasive PDAC, we believe it will be important to understand earlier events in pancreatic cancer, i.e. PanIN formation and progression. We hypothesized that stage-specific bottlenecks may lead to clonal selection during pancreatic intraepithelial neoplasia (PanIN) formation and progression. In order to test this hypothesis, we examined evolving clonal complexity during the progression of pancreatic intraepithelial neoplasia in the Mist1Tg/wt; LSL-KrasG12D (KCiMist1) mouse model of pancreatic cancer. Following KrasG12D activation in adult acinar cells, these mice develop acinar to ductal metaplasia (ADM) and preinvasive PanIN 1 - PanIN 3 lesions in a manner that faithfully recapitulates the human disease. Our strategy involves crossing MistTg/wt; LSL-KrasG12D to Brainbow2.1Tg/Tg “Confetti” mice, in which individual cells undergo stochastic recombination of the Confetti reporter to activate expression of either nuclear GFP, membrane-associated CFP, cytoplasmic RFP or cytoplasmic YFP. Following tamoxifen induction, mice were given cerulein injections to increase PanIN density, and sacrificed at four and nine weeks post-tamoxifen. Following additional immunofluorescent labeling with EpCAM to demarcate individual cells, confocal imaging was performed. Lesions were characterized by grade (ADM, early PanIN and late PanIN), and the number of clones, defined as adjacent cells sharing an identical fluorescent protein signature, was compared at 4 and 9 weeks. For AMDs, early PanIN and late PanIN, we observe both monoclonal and polyclonal lesions. Our analysis of evolving clonal complexity over time is ongoing, but preliminary data suggests a time-dependent decrease in clonal complexity. The average number of clones seen in individual ADM and late PanIN lesions decreased in the mice sacrificed at nine week post tamoxifen induction compared to those sacrificed at four weeks post tamoxifen induction (1.5 +/- 0.52 clones per lesion vs. 2.29 +/- 1.14 clones per lesion for ADM and 2.5 +/- 1.29 vs. 3.75 +/- 1.89 clones per lesion for late PanIN). In contrast, there was little difference in clonal complexity noted for early PanINs at 4 vs. 9 weeks (2.17 +/- 1.19 clones per lesion in the four week cohort vs. 2.42 +/- 1.24 clones per lesion in the nine week time point). These preliminary data suggest an overall trend towards decreased clonal complexity of PanIN lesions over time, suggesting significant selective pressure at even the earliest stages of pancreatic cancer. This abstract is also presented as Poster A62. Citation Format: Kelly J. Lafaro, Audrey Hendley, Jennifer Bailey, Steven Leach. Clonal composition and clonal selection during PanIN progression. [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Innovations in Research and Treatment; May 18-21, 2014; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2015;75(13 Suppl):Abstract nr PR04.