Abstract 2909: Rapid autopsy of metastatic cholangiocarcinoma demonstrates diverse patterns of clonal heterogeneity
Authors: Krook Melanie A. , Bonneville Russell , Chen Hui-Zi , Reeser Julie W. , Wing Michele R. , Miya Jharna , Smith Amy M. , Paruchuri Anoosha , Dao Thuy , Martin Dorrelyn M. , Samorodnitsky Eric , Baker Kaitlin R. , Timmers Cynthia , Dittmar Kristin , Cole Sharon , Yu Lianbo , Freud Aharon G. , Allenby Patricia , Roychowdhury Sameek 
Published: August 2019

Abstract
Background: Cholangiocarcinoma is an aggressive and deadly bile duct cancer. Due to its poor prognosis and limited treatments beyond first-line therapy, in-depth characterization of cholangiocarcinoma is needed to facilitate the development of novel therapies. Tumor heterogeneity has emerged as a contributor to treatment resistance, but it is largely unknown how it contributes to cholangiocarcinoma. Characterization of tumor heterogeneity in cholangiocarcinoma has largely been limited to biopsies and surgical resections and may not accurately reflect the complex and heterogeneous nature of the disease. A partnership with patients and their families for body donation is a powerful strategy to characterize the landscape of tumor heterogeneity in cholangiocarcinoma.
Study Design: We performed whole exome sequencing (WES) of a pre-treatment tumor biopsy and multiple tumor samples collected through rapid research autopsy from 9 patients with metastatic cholangiocarcinoma. This will allow us to determine spatial heterogeneity, delineate cancer phylogeny, and identify spatial origin of treatment-resistant clones as well as drivers of metastasis and recurrence.
Results and Conclusions: We completed research autopsies of 9 patients with metastatic cholangiocarcinoma including 5 with FGFR alterations, 1 with an IDH1 mutation, 1 with a BRCA mutation, 1 with a BAP1 splice site mutation, and 1 with hypermutation by an unknown mechanism. We observed varying patterns of clonal evolution in this cohort including persistent, sensitive, and emergent clones. One patient with metastatic cholangiocarcinoma with a novel FGFR2-CLIP1fusion received the FGFR inhibitor, INCB054828 and had a 5.5-month response but unfortunately progressed and succumbed to his disease. Multi-site WES and analysis of tumor heterogeneity through subclonal inference identified four genetically distinct clonal cancer cell populations, two of which were only observed in post-treatment samples (emergent clones). Additionally, WES revealed an FGFR2N549H mutation in a single tumor sample hypothesized to confer resistance to INCB054828. This hypothesis was corroborated in vitro, in which cells expressing the FGFR2-CLIP1 fusion protein were sensitive to INCB054828 (IC50of 10.16 nM), whereas cells expressing FGFR2-CLIP1 N549H were resistant (IC50of 1257.57 nM). Furthermore,the FGFR2 N549H mutation displayed cross-resistance to other selective FGFR inhibitors, but remained sensitive to the non-selective inhibitor, ponatinib.
Cancer Relevance: These studies will augment our understanding of tumor heterogeneity in cholangiocarcinoma. The broader impact of this research includes opportunities to develop new therapies for cholangiocarcinoma, centralized storage of cholangiocarcinoma tumor samples, and demonstration of how research autopsy can be applied to study tumor heterogeneity in other cancer types.
Citation Format: Melanie A. Krook, Russell Bonneville, Hui-Zi Chen, Julie W. Reeser, Michele R. Wing, Jharna Miya, Amy M. Smith, Anoosha Paruchuri, Thuy Dao, Dorrelyn M. Martin, Eric Samorodnitsky, Kaitlin R. Baker, Cynthia Timmers, Kristin Dittmar, Sharon Cole, Lianbo Yu, Aharon G. Freud, Patricia Allenby, Sameek Roychowdhury. Rapid autopsy of metastatic cholangiocarcinoma demonstrates diverse patterns of clonal heterogeneity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2909.