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Cell Mol Gastroenterol Hepatol. 2019;7(4):803-817. doi: 10.1016/j.jcmgh.2019.01.008. Epub 2019 Feb 11.

MicroRNA-375 Suppresses the Growth and Invasion of Fibrolamellar Carcinoma.

Author information

1
Curriculum in Genetics and Molecular Biology, School of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina; Department of Biomedical Sciences, College of Veterinary Medicine, Cornell University, Ithaca, New York.
2
Department of Medicine, School of Medicine, Duke University, Durham, North Carolina.
3
Department of Biomedical Sciences, College of Veterinary Medicine, Cornell University, Ithaca, New York.
4
Department of Pharmacology, School of Medicine, University of Washington, Seattle, Washington.
5
Cancer Biology and Genetics Program, Memorial Sloan-Kettering Cancer Center, New York, New York.
6
Department of Cell Biology and Physiology, School of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina.
7
Department of Pathology, School of Medicine, Duke University, Durham, North Carolina.
8
Department of Surgery, University of Washington, Seattle, Washington.
9
Department of Medicine, School of Medicine, Duke University, Durham, North Carolina. Electronic address: annamae.diehl@duke.edu.
10
Department of Biomedical Sciences, College of Veterinary Medicine, Cornell University, Ithaca, New York. Electronic address: pr46@cornell.edu.

Abstract

BACKGROUND & AIMS:

Fibrolamellar carcinoma (FLC) is a rare liver cancer that primarily affects adolescents and young adults. It is characterized by a heterozygous approximately 400-kb deletion on chromosome 19 that results in a unique fusion between DnaJ heat shock protein family member B1 (DNAJB1) and the alpha catalytic subunit of protein kinase A (PRKACA). The role of microRNAs (miRNAs) in FLC remains unclear. We identified dysregulated miRNAs in FLC and investigated whether dysregulation of 1 key miRNA contributes to FLC pathogenesis.

METHODS:

We analyzed small RNA sequencing (smRNA-seq) data from The Cancer Genome Atlas to identify dysregulated miRNAs in primary FLC tumors and validated the findings in 3 independent FLC cohorts. smRNA-seq also was performed on a FLC patient-derived xenograft model as well as purified cell populations of the liver to determine whether key miRNA changes were tumor cell-intrinsic. We then used clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (Cas9) technology and transposon-mediated gene transfer in mice to determine if the presence of DNAJB1-PRKACA is sufficient to suppress miR-375 expression. Finally, we established a new FLC cell line and performed colony formation and scratch wound assays to determine the functional consequences of miR-375 overexpression.

RESULTS:

We identified miR-375 as the most dysregulated miRNA in primary FLC tumors (27-fold down-regulation; P = .009). miR-375 expression also was decreased significantly in a FLC patient-derived xenograft model compared to 4 different cell populations of the liver. Introduction of DNAJB1-PRKACA by clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 engineering and transposon-mediated somatic gene transfer in mice was sufficient to induce significant loss of miR-375 expression (P < .05). Overexpression of miR-375 in FLC cells inhibited Hippo signaling pathway proteins, including yes-associated protein 1 and connective tissue growth factor, and suppressed cell proliferation and migration (P < .05).

CONCLUSIONS:

We identified miR-375 as the most down-regulated miRNA in FLC tumors and showed that overexpression of miR-375 mitigated tumor cell growth and invasive potential. These findings open a potentially new molecular therapeutic approach. Further studies are necessary to determine how DNAJB1-PRKACA suppresses miR-375 expression and whether miR-375 has additional important targets in this tumor. Transcript profiling: GEO accession numbers: GSE114974 and GSE125602.

KEYWORDS:

Cancer Genomics; Fibrolamellar Carcinoma; Pediatric Cancer; miRNA

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