Abstract 131: Clonal expansion in bile duct associated with chronic inflammation
Authors: Maeda Hirona , Kakiuchi Nobuyuki , Ito Takashi , Ogawa Eri , Shiokawa Masahiro , Uza Norimitsu , Tanaka Hiroko , Nannya Yasuhito , Makishima Hideki , Yasuda Hiroaki , Kodama Yuzo , Miyano Satoru , Ogawa Seishi 
Published: June 2023

Abstract
Introduction: Chronic inflammation has been implicated in cancer development in many tissues and has recently been associated with promoted clonal expansion in non-cancer tissues. Primary sclerosing cholangitis (PSC) is a chronic progressive disease of unknown etiology affecting the bile ducts and has been associated with a substantially increased risk of cholangiocarcinoma. Clonal dynamics in the normal bile duct and PSC epithelium remains poorly understood because of the limited accessibility to normal/inflamed bile ducts. In this study, we established a method to detect somatic mutations accumulated in the bile duct epithelium and investigated a mutation rate in normal bile duct cells and clonal expansion in PSC epithelium.
Methods: We established single cell-derived organoids from bile, which were subjected to whole genome sequencing to measure the age-related mutation rate in normal epithelial cell in the bile ducts. We next collected multiple samples from perihilar bile ducts in patients with PSC who underwent total hepatectomy for liver transplantation, for which we performed whole exome sequencing to investigate clonal expansion in PSC-involved bile ducts.
Results: In total, 14 single cell-derived organoid lines were established from 2 patients with PSC and 3 control individuals. Based on the whole genome sequencing of those organoids, the number of mutations found in non-PSC bile duct epithelium increases with age at an annual mutation rate of 38.2 mutations per genome per year, which did not substantially differ from the rate in the PSC epithelium, suggesting that the inflammatory process in PSC hardly affected the mutations in PSC epithelium. To further investigate clonal expansion in the PSC epithelium, a total of 126 bulk organoid samples from perihilar bile ducts in 3 PSC patients were analyzed. Somatic mutations were detected in all samples (21 [4-101]/exome/sample on average), some of which showed high variant allele frequencies. In 2 PSC cases, a small number of somatic mutations (n=12[case 1], 11 and 8[case 2]) were shared between samples that were 13 mm distant apart from each other, suggesting a clonal expansion in PSC epithelium. Expanded clones harbored mutations in known cancer-driver genes, such as ARID2 and PIK3CA, suggesting their role in the expansion in this case.
Conclusions: Here, we newly established the method to detect somatic mutations accumulated in the normal/inflamed bile duct epithelium using an organoid-based analysis of clonal somatic mutations. Our results revealed clonal expansion existing in PSC epithelium and open up further potential for approaching carcinogenesis in the bile duct.
Citation Format: Hirona Maeda, Nobuyuki Kakiuchi, Takashi Ito, Eri Ogawa, Masahiro Shiokawa, Norimitsu Uza, Hiroko Tanaka, Yasuhito Nannya, Hideki Makishima, Hiroaki Yasuda, Yuzo Kodama, Satoru Miyano, Seishi Ogawa. Clonal expansion in bile duct associated with chronic inflammation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 131.