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Establishment and characterization of a human intrahepatic cholangiocarcinoma cell line derived from an Italian patient.

ABSTRACT: Biliary tract carcinoma is a rare malignancy with multiple causes, which underlie the different genetic and molecular profiles. Cancer cell lines are affordable models, reflecting the characteristics of the tumor of origin. They represent useful tools to identify molecular targets for treatment. Here, we established and characterized from biological, molecular, and genetic point of view, an Italian intrahepatic cholangiocarcinoma cell line (ICC), the MT-CHC01. MT-CHC01 cells were isolated from a tumor-derived xenograft. Immunophenotypical characterization was evaluated both at early and after stabilization passages. In vitro biological, genetic, and molecular features were also investigated. In vivo tumorigenicity was assessed in NOD/SCID mice. MT-CHC01cells retain epithelial cell markers, EPCAM, CK7, and CK19, and some stemness and pluripotency markers, i.e., SOX2, Nanog, CD49f/integrin-?6, CD24, PDX1, FOXA2, and CD133. They grow as a monolayer, with a population double time of about 40 h; they show a low migration and invasion potential. In low attachment conditions, they are able to form spheres and to growth in anchorage-independent manner. After subcutaneous injection, they retain in vivo tumorigenicity; the expression of biliary markers as CA19-9 and CEA were maintained from primary tumor. The karyotype is highly complex, with a hypotriploid to hypertriploid modal number (3n+/-) (52 to 77 chromosomes); low level of HER2 gene amplification, TP53 deletion, gain of AURKA were identified; K-RAS G12D mutation were maintained from primary tumor to MT-CHC01 cells. We established the first ICC cell line derived from an Italian patient. It will help to study either the biology of this tumor or to test drugs both in vitro and in vivo.

SUBMITTER: Cavalloni G

PROVIDER: S-EPMC4844644 | biostudies-literature | 2016 Mar

REPOSITORIES: biostudies-literature

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Establishment and characterization of a human intrahepatic cholangiocarcinoma cell line derived from an Italian patient.

Cavalloni Giuliana G Peraldo-Neia Caterina C Varamo Chiara C Casorzo Laura L Dell'Aglio Carmine C Bernabei Paola P Chiorino Giovanna G Aglietta Massimo M Leone Francesco F

Tumour biology : the journal of the International Society for Oncodevelopmental Biology and Medicine 20151020 3

Biliary tract carcinoma is a rare malignancy with multiple causes, which underlie the different genetic and molecular profiles. Cancer cell lines are affordable models, reflecting the characteristics of the tumor of origin. They represent useful tools to identify molecular targets for treatment. Here, we established and characterized from biological, molecular, and genetic point of view, an Italian intrahepatic cholangiocarcinoma cell line (ICC), the MT-CHC01. MT-CHC01 cells were isolated from a ...[more]

PMID: 26486326

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Project description:Chemotherapy resistance is a relevant clinical issue in the tumor treatment, in particular in biliary tract carcinoma (BTC), for which there are no effective therapies, neither in the first nor in the second line. The development of chemoresistant cell lines as experimental models to investigate the mechanisms of resistance and identify alternative druggable pathways is mandatory. In BTC, in which genetics and biological behavior depend on the etiology, ethnicity and anatomical site of origin, the creation of models that better recapitulate these characteristics is even more crucial. Here we have established and characterized an intrahepatic cholangiocarcinoma (iCCA) cell line derived from an Italian patient, called 82.3. Cells were isolated from a patient-derived xenograft and, after establishment, immunophenotypic, biological, genetic, molecular characteristics and tumorigenicity in vivo in NOD/SCID mice were investigated. 82.3 cells exhibited epithelial morphology and cell markers (EPCAM, CK7 and 19); they also expressed different cancer stem markers (CD44, CD133, CD49b, CD24, Stro1, PAX6, FOXA2, OCT3 / 4) and under anchorage-independent and serum-free conditions were capable of originating cholangiospheres. The population doubling time was approximately 53 hours. In vitro they demonstrated a poor ability to migrate, but in vivo, 82.3 cells retained their tumorigenicity, with a long latency period (16 weeks). Genetic identity using DNA fingerprinting analysis revealed 16 different loci and the cell line was characterized by a complex hyperdiploid karyotype. Furthermore, 82.3 cells showed cross-resistance to gemcitabine, 5-fluorouracil, carboplatin and oxaliplatin; in fact their genetic profile showed that in a panel of genes (n = 168), specific for drug resistance and related to the epithelial-mesenchymal transition, 60% of them were deregulated in 82.3 cells compared to a control iCCA cell line sensitive to chemotherapeutics. In conclusion, we have created new iCCA cell line of Caucasian origin that could be exploited as a preclinical model to study drug resistance mechanisms and to identify alternative therapies to improve the prognosis of this tumor type.

Project description:BackgroundExtrahepatic cholangiocarcinoma sarcoma is extremely rare in clinical practice. These cells consist of both epithelial and mesenchymal cells. Patient-derived cell lines that maintain tumor characteristics are valuable tools for studying the molecular mechanisms associated with carcinosarcoma. However, cholangiocarcinoma sarcoma cell lines are not available in cell banks.AimTo establish and characterize a new extrahepatic cholangiocarcinoma sarcoma cell line, namely CBC2T-2.MethodsWe conducted a short tandem repeat (STR) test to confirm the identity of the CBC2T-2 cell line. Furthermore, we assessed the migratory and invasive properties of the cells and performed clonogenicity assay to evaluate the ability of individual cells to form colonies. The tumorigenic potential of CBC2T-2 cells was tested in vivo using non-obese diabetic/severe combined immunodeficient (NOD/SCID) mice. The cells were injected subcutaneously and tumor formation was observed. In addition, immunohistochemical analysis was carried out to examine the expression of epithelial marker CK19 and mesenchymal marker vimentin in both CBC2T-2 cells and xenografts. The CBC2T-2 cell line was used to screen the potential therapeutic effects of various clinical agents in patients with cholangiocarcinoma sarcoma. Lastly, whole-exome sequencing was performed to identify genetic alterations and screen for somatic mutations in the CBC2T-2 cell line.ResultsThe STR test showed that there was no cross-contamination and the results were identical to those of the original tissue. The cells showed round or oval-shaped epithelioid cells and mesenchymal cells with spindle-shaped or elongated morphology. The cells exhibited a high proliferation ratio with a doubling time of 47.11 h. This cell line has migratory, invasive, and clonogenic abilities. The chromosomes in the CBC2T-2 cells were polyploidy, with numbers ranging from 69 to 79. The subcutaneous tumorigenic assay confirmed the in vivo tumorigenic ability of CBC2T-2 cells in NOD/SCID mice. CBC2T-2 cells and xenografts were positive for both the epithelial marker, CK19, and the mesenchymal marker, vimentin. These results suggest that CBC2T-2 cells may have both epithelial and mesenchymal characteristics. The cells were also used to screen clinical agents in patients with cholangiocarcinoma sarcoma, and a combination of paclitaxel and gemcitabine was found to be the most effective treatment option.ConclusionWe established the first human cholangiocarcinoma sarcoma cell line, CBC2T-2, with stable biogenetic traits. This cell line, as a research model, has a high clinical value and would facilitate the understanding of the pathogenesis of cholangiocarcinoma sarcoma.

Dataset Information

Establishment and characterization of a human intrahepatic cholangiocarcinoma cell line derived from an Italian patient.

Publications

Establishment and characterization of a human intrahepatic cholangiocarcinoma cell line derived from an Italian patient.

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