title: Oncogenic potential of IDH1R132C mutant in cholangiocarcinoma development in mice.
abstract: OBJECTIVE
  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.
  
  
  CONCLUSIONS
  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.
authors.affiliation: 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.
authors.forename: Ning
authors.initials: N
authors.lastname: Ding
authors.affiliation: 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.
authors.forename: Li
authors.initials: L
authors.lastname: Che
authors.affiliation: 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.
authors.forename: Xiao-Lei
authors.initials: XL
authors.lastname: Li
authors.affiliation: 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.
authors.forename: Yan
authors.initials: Y
authors.lastname: Liu
authors.affiliation: 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.
authors.forename: Li-Jie
authors.initials: LJ
authors.lastname: Jiang
authors.affiliation: 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.
authors.forename: Biao
authors.initials: B
authors.lastname: Fan
authors.affiliation: 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.
authors.forename: Jun-Yan
authors.initials: JY
authors.lastname: Tao
authors.affiliation: 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.
authors.forename: Xin
authors.initials: X
authors.lastname: Chen
authors.affiliation: 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.
authors.forename: Jia-Fu
authors.initials: JF
authors.lastname: Ji
mesh_terms: D000818:Animals; D001650:Bile Duct Neoplasms / Q000201:enzymology / Q000235:genetics* / Q000473:pathology; D002471:Cell Transformation, Neoplastic / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D018281:Cholangiocarcinoma / Q000201:enzymology / Q000235:genetics* / Q000473:pathology; D015971:Gene Expression Regulation, Enzymologic; D015972:Gene Expression Regulation, Neoplastic; D020022:Genetic Predisposition to Disease; D006801:Humans; D007521:Isocitrate Dehydrogenase / Q000235:genetics* / Q000378:metabolism; D008099:Liver / Q000201:enzymology* / Q000473:pathology; D051379:Mice; D009154:Mutation; D009857:Oncogenes; D010641:Phenotype; D051880:Receptors, Notch / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D013997:Time Factors; D014162:Transfection; D016159:Tumor Suppressor Protein p53 / Q000172:deficiency / Q000235:genetics
chemical_list: D051880:Receptors, Notch; D016159:Tumor Suppressor Protein p53; D007521:Isocitrate Dehydrogenase; C543588:IDH1 protein, human
keywords: IDH1 mutant; Intrahepatic cholangiocarcinoma; Mouse liver cancer; Notch pathway; p53