A genomic case study of mixed fibrolamellar hepatocellular carcinoma

Griffith, Obi L.; Griffith, Malachi; Krysiak, Kilannin; Magrini, Vincent; Ramu, Avinash; Skidmore, Zachary L.; Kunisaki, Jason; Austin, Rachel N.; McGrath, Sean; Zhang, Jin; Demeter, Ryan; Graves, Tina; Eldred, James M.; Walker, Jason; Larson, David E.; Maher, Christopher A.; Lin, Yiing; Chapman, William C.; Mahadevan, Anand; Miksad, Rebecca A.; Nasser, Imad; Hanto, Douglas W.; Mardis, Elaine R.

doi:10.1093/annonc/mdw135

Cited by 20 publications

(12 citation statements)

References 18 publications

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“…All the large and small Dnajb1–Prkaca -elicited lesions showed the cytological characteristics of FL-HCC described above. Only one lesion, in fact, contained premalignant clear cells intermingled with oncocytic hepatocytes with prominent nucleoli and eosinophilic cytoplasm (Supplementary Figure 3 C ), in accordance with the observation that FL-HCC may occasionally appear as a mixed-FL-HCC, which has also been reported positive for the fusion 25 . Genomic DNA analysis of laser capture microdissected small lesions confirmed the presence of the oncogenic Dnajb1–Prkaca aberration (Supplementary Figure 3 D ).…”

Section: Resultssupporting

confidence: 88%

CRISPR/Cas9 Engineering of Adult Mouse Liver Demonstrates That the Dnajb1–Prkaca Gene Fusion Is Sufficient to Induce Tumors Resembling Fibrolamellar Hepatocellular Carcinoma

et al. 2017

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Background & Aims Fibrolamellar hepatocellular carcinoma (FL-HCC) is a primary liver cancer that predominantly affects young adults with no underlying liver disease. A somatic, 400 Kb deletion on chromosome 19 that fuses part of the DnaJ heat shock protein family (Hsp40) member B1 gene (DNAJB1) to the protein kinase cAMP-activated catalytic subunit alpha gene (PRKACA) has been repeatedly identified in patients with FL-HCC. However, the DNAJB1–PRKACA gene fusion has not been shown to induce liver tumorigenesis. We used the CRISPR/Cas9 technique to delete in mice the syntenic region on chromosome 8 to create a Dnajb1–Prkaca fusion and monitored the mice for liver tumor development. Methods We delivered CRISPR/Cas9 vectors designed to juxtapose exon 1 of Dnajb1 with exon 2 of Prkaca to create the Dnajb1–Prkaca gene fusion associated with FL-HCC, or control Cas9 vector, via hydrodynamic tail vein injection to livers of 8 week-old female FVB/N mice. These mice did not have any other engineered genetic alterations and were not exposed to liver toxins or carcinogens. Liver tissues were collected 14 months after delivery; genomic DNA was analyzed by PCR to detect the Dnajb1–Prkaca fusion, and tissues were characterized by histology, immunohistochemistry, RNA sequencing, and whole-exome sequencing. Results Livers from 12 of the 15 mice given the vectors to induce the Dnajb1–Prkaca gene fusion, but none of the 11 mice given the control vector, developed neoplasms. The tumors contained the Dnajb1–Prkaca gene fusion and had histologic and cytologic features of human FL-HCCs: large polygonal cells with granular, eosinophilic, and mitochondria-rich cytoplasm, prominent nucleoli, and markers of hepatocytes and cholangiocytes. In comparing expression levels of genes between the mouse tumor and non-tumor liver cells, we identified changes similar to those detected in human FL-HCC, which included genes that affect cell cycle and mitosis regulation. Genomic analysis of mouse neoplasms induced by the Dnajb1–Prkaca fusion revealed a lack of mutations in genes commonly associated with liver cancers, as observed in human FL-HCC. Conclusions Using CRISPR/Cas9 technology, we found generation of the Dnajb1–Prkaca fusion gene in wild-type mice to be sufficient to initiate formation of tumors that have many features of human FL-HCC. Strategies to block DNAJB1–PRKACA might be developed as therapeutics for this form of liver cancer.

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Section: Resultssupporting

confidence: 88%

CRISPR/Cas9 Engineering of Adult Mouse Liver Demonstrates That the Dnajb1–Prkaca Gene Fusion Is Sufficient to Induce Tumors Resembling Fibrolamellar Hepatocellular Carcinoma

et al. 2017

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“…liver cancer is usually occult in onset and no effective screening measure is able to screen for the disease validly and reliably; therefore, two-thirds of cases are not diagnosed until an advanced stage (10,11). furthermore, liver cancer is often resistant to chemotherapeutic drugs because it usually originates from pre-existing liver diseases (12,13). these factors largely contribute to the high mortality rate of liver cancer.…”

Section: Microrna-493-5p Promotes Apoptosis and Suppresses Proliferatmentioning

confidence: 99%

MicroRNA-493-5p promotes apoptosis and suppresses proliferation and invasion in liver cancer cells by targeting VAMP2

et al. 2018

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The aim of the present study was to explore the role of miR‑493-5p in liver cancer tissues and cell lines, and its effect on cell behavioral characteristics. The expression of miR-493-5p was detected by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) in liver cancer tissues and cell lines (hepatic cell line HL-7702 and the liver cancer cell lines HCCC-9810, HuH-7 and HepG2). In addition, the mechanism by which miR-493-5p mediates its effects was analyzed via the transfection of miR-493-5p mimic and negative control miRNA into HepG2 cells. The viability, proliferation, apoptosis and invasion of the cells were analyzed using MTT assay, flow cytometry and Transwell chamber experiments. Furthermore, the effect of miR-493-5p on the expression of vesicle associated membrane protein 2 (VAMP2) was assayed using a dual-luciferase reporter system, and VAMP2 protein levels were determined by western blot analysis. In addition, following the cotransfection of HepG2 cells with pcDNA3.1‑VAMP2 plasmid and miR‑493-5p mimic, the role of miR-493-5p as a regulator of VAMP2 was evaluated using MTT assay, flow cytometry and Transwell chamber experiments. RT-qPCR analysis indicated that the expression of miR-493-5p in liver cancer tissues and cell lines was decreased significantly compared with that in adjacent normal liver tissues and normal liver cell lines, respectively. Compared with the control group, the cells transfected with miR-493-5p mimic (the miR-493-5p overexpression group) exhibited reduced cell viability, a reduced percentage of cells in the S phase and an increased percentage of apoptotic cells. In addition, fewer cells passed through the Transwell membrane in the miR-493-5p overexpression group compared with the control group. In the dual-luciferase reporter assay, luciferase activity in the miR‑493-5p overexpression group was attenuated compared with that in the control group. In addition, western blot analysis indicated that the VAMP2 protein levels in the miR‑493-5p overexpression group were lower than those in the control group. Furthermore, in cells overexpressing miR-493-5p and VAMP2 simultaneously, the biological behavior of the cells, including cell viability, cell cycle and cell invasiveness, was significantly rescued compared with that of the control group transfected with miR‑493-5p alone. In conclusion, miR-493-5p is indicated to be a tumor suppressor gene, and is downregulated in human liver cancer. miR-493-5p overexpression promotes cell apoptosis and inhibits the proliferation and migration of liver cancer cells by negatively regulating the expression of VAMP. These observations suggest the potential of treating liver cancer by the overexpression of microRNA-493-5p.

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Section: Genetic Abnormalities In Liver Cancermentioning

confidence: 99%

“…Recent studies have characterized at molecular level a variant of FL-HCC, known as mixed fibrolamellar hepatocellular carcinoma (Mix-FL-HCC) and characterized by the presence of both pure FL-HCC and conventional HCC component within the same tumors [ 53 ]. In these rare tumors, the DNAJB1-PRKACA fusion transcript is expressed at high levels [ 53 ]. Importantly, the analysis of RNA-sequencing data from The Cancer Genome Atlas (TCGA) for more than 9000 tumors across about 30 cancer types showed that the DNAJB1-PRKACA fusion is specific for FLHCCs [ 54 ].…”

Section: Genetic Abnormalities In Liver Cancermentioning

confidence: 99%

Liver Cancer: Molecular Characterization, Clonal Evolution and Cancer Stem Cells

Castelli

Pelosi

Testa

2017

Cancers

120

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Liver cancer is the second most common cause of cancer-related death. The major forms of primary liver cancer are hepatocellular carcinoma (HCC) and intrahepatic cholangiocarcinoma (iCCA). Both these tumors develop against a background of cirrhotic liver, non-alcoholic fatty liver disease, chronic liver damage and fibrosis. HCC is a heterogeneous disease which usually develops within liver cirrhosis related to various etiologies: hepatitis B virus (HBV) infection (frequent in Asia and Africa), hepatitis C virus (HCV), chronic alcohol abuse, or metabolic syndrome (frequent in Western countries). In cirrhosis, hepatocarcinogenesis is a multi-step process where pre-cancerous dysplastic macronodules transform progressively into HCC. The patterns of genomic alterations observed in these tumors were recently identified and were instrumental for the identification of potential targeted therapies that could improve patient care. Liver cancer stem cells are a small subset of undifferentiated liver tumor cells, responsible for cancer initiation, metastasis, relapse and chemoresistance, enriched and isolated according to immunophenotypic and functional properties: cell surface proteins (CD133, CD90, CD44, EpCAM, OV-6, CD13, CD24, DLK1, α2δ1, ICAM-1 and CD47); the functional markers corresponding to side population, high aldehyde dehydrogenase (ALDH) activity and autofluorescence. The identification and definition of liver cancer stem cells requires both immunophenotypic and functional properties.

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A genomic case study of mixed fibrolamellar hepatocellular carcinoma

Cited by 20 publications

References 18 publications

CRISPR/Cas9 Engineering of Adult Mouse Liver Demonstrates That the Dnajb1–Prkaca Gene Fusion Is Sufficient to Induce Tumors Resembling Fibrolamellar Hepatocellular Carcinoma

CRISPR/Cas9 Engineering of Adult Mouse Liver Demonstrates That the Dnajb1–Prkaca Gene Fusion Is Sufficient to Induce Tumors Resembling Fibrolamellar Hepatocellular Carcinoma

MicroRNA-493-5p promotes apoptosis and suppresses proliferation and invasion in liver cancer cells by targeting VAMP2

Liver Cancer: Molecular Characterization, Clonal Evolution and Cancer Stem Cells

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