Basic Study
Copyright ©The Author(s) 2016. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Feb 14, 2016; 22(6): 2071-2080
Published online Feb 14, 2016. doi: 10.3748/wjg.v22.i6.2071
Oncogenic potential of IDH1R132C mutant in cholangiocarcinoma development in mice
Ning Ding, Li Che, Xiao-Lei Li, Yan Liu, Li-Jie Jiang, Biao Fan, Jun-Yan Tao, Xin Chen, Jia-Fu Ji
Ning Ding, Li Che, Biao Fan, Jia-Fu Ji, Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education), Department of Gastrointestinal Surgery, Peking University Cancer Hospital and Institute, Beijing 100142, China
Ning Ding, Li Che, Xiao-Lei Li, Yan Liu, Li-Jie Jiang, Jun-Yan Tao, Xin Chen, Department of Bioengineering and Therapeutic Sciences, Liver Center, University of California, San Francisco, CA 94143, United States
Xiao-Lei Li, Yan Liu, Department of Hepatobiliary Surgery, Xijing Hospital, The Fourth Military Medical University, Xi’an 710032, Shanxi Province, China
Jun-Yan Tao, Xin Chen, School of Pharmacy, Hubei University of Chinese Medicine, Wuhan 430065, Hubei Province, China
Author contributions: Ding N, Chen X and Ji JF designed the research; Ding N, Che L, Li XL, Liu Y, Jiang LJ, Fan B and Tao JY performed the research and analyzed the data; Ding N and Chen X wrote the paper.
Supported by Grants from National Institutes of Health, No. R01CA136606 (in part, to Chen X); UCSF Liver Center, No. P30DK026743; and China Scholarship Council, contract, No. 201206010086 (to Ding N) and No. 201306590021 (to Li XL).
Institutional animal care and use committee statement: All animal studies were approved by UCSF Institutional Animal Care and Use Committee (IACUC protocol number: AN108577).
Conflict-of-interest statement: The authors declare no conflict of interests.
Data sharing statement: No additional data are available.
Open-Access: This article is an open-access article which was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/
Correspondence to: Jia-Fu Ji, MD, Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education), Department of Gastrointestinal Surgery, Peking University Cancer Hospital and Institute, No. 52 Fucheng Road, Haidian District, Beijing 100142, China. jiafuj@gmail.com
Telephone: +86-10-88196048 Fax: +86-10-88122437
Received: January 15, 2015
Peer-review started: January 16, 2015
First decision: March 26, 2015
Revised: August 4, 2015
Accepted: September 2, 2015
Article in press: September 2, 2015
Published online: February 14, 2016
Processing time: 373 Days and 2.9 Hours
Abstract

AIM: To investigate whether IDH1R132C mutant in combination with loss of p53 and activated Notch signaling promotes intrahepatic cholangiocarcinoma (ICC) development.

METHODS: We applied hydrodynamic injection and sleeping beauty mediated somatic integration to induce loss of p53 (via shP53), activation of Notch [via intracellular domain of Notch1 (NICD)] and/or overexpression of IDH1R132C mutant together with the sleeping beauty transposase into the mouse liver. Specifically, we co-expressed shP53 and NICD (shP53/NICD, n = 4), shP53 and IDH1R132C (shP53/IDH1R132C, n = 3), NICD and IDH1R132C (NICD/IDH1R132C, n = 4), as well as NICD, shP53 and IDH1R132C (NICD/shP53/IDH1R132C, n = 9) in mice. Mice were monitored for liver tumor development and euthanized at various time points. Liver histology was analyzed by hematoxylin and eosin staining. Molecular features of NICD/shP53/IDH1R132C ICC tumor cells were characterized by Myc tag, Flag tag, Ki-67, p-Erk and p-AKT immunohistochemical staining. Desmoplastic reaction in tumor tissues was studied by Picro-Sirius red staining.

RESULTS: We found that co-expression of shP53/NICD, shP53/IDH1R132C or NICD/IDH1R132C did not lead to liver tumor formation. In striking contrast, co-expression of NICD/shP53/IDH1R132C resulted in ICC development in mice (P < 0.01). The tumors could be identified as early as 12 wk post hydrodynamic injection. Tumors rapidly progressed, and by 18 wk post hydrodynamic injection, multiple cystic lesions could be identified on the liver surface. NICD/shP53/IDH1R132C liver tumors shared multiple histological features of human ICCs, including hyperplasia of irregular glands. Importantly, all tumor cells were positive for the biliary epithelial cell marker cytokeratin 19. Extensive collagen fibers could be visualized in tumor tissues using Sirus red staining, duplicating the desmoplastic reaction observed in human ICC. Tumors were highly proliferative and expressed ectopically injected genes. Together these studies supported that NICD/shP53/IDH1R132C liver tumors were indeed ICCs. Finally, no p-AKT or p-ERK positive staining was observed, suggesting that NICD/shP53/IDH1R132C driven ICC development was independent of AKT/mTOR and Ras/MAPK signaling cascades.

CONCLUSION: We have generated a simple, non-germline murine ICC model with activated Notch, loss of p53 and IDH1R132C mutant. The study supported the oncogenic potential of IDH1R132C.

Keywords: IDH1 mutant; Notch pathway; Intrahepatic cholangiocarcinoma; Mouse liver cancer; p53

Core tip: We established a novel murine intrahepatic cholangiocarcinoma (ICC) model via hydrodynamic transfection of activated form of Notch1 (NICD), shP53 and IDH1R132C into the mouse liver. This study is the first to demonstrate that IDH1R132C mutant can cooperate with other oncogenes or tumor suppressor genes to promote ICC development in vivo. In addition, it provides the ICC research community with an innovative and convenient approach to generate IDH1R132C mutant ICC in mice. Finally, ICC induced by NICD/shP53/IDH1R132C provides a useful tool to study IDH mutant in ICC pathogenesis and a novel preclinical murine model for testing drugs against the deadly malignancy.