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miR-122 – A key factor and therapeutic target in liver disease

  • Simonetta Bandiera
    Affiliations
    Inserm, U1110, Institut de Recherche sur les Maladies Virales et Hépatiques, Strasbourg, France

    Université de Strasbourg, Strasbourg, France
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  • Sébastien Pfeffer
    Affiliations
    Université de Strasbourg, Strasbourg, France

    Architecture et Réactivité de l’ARN – UPR 9002, Institut de Biologie Moléculaire et Cellulaire du CNRS, Strasbourg, France
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  • Thomas F. Baumert
    Correspondence
    Corresponding authors. Address: Inserm, U1110, Institut de Recherche sur les Maladies Virales et Hépatiques, Université de Strasbourg, 3 Rue Koeberlé, 67000 Strasbourg, France. Tel.: +33 3 68 85 37 03; fax: +33 3 68 85 37 24.
    Affiliations
    Inserm, U1110, Institut de Recherche sur les Maladies Virales et Hépatiques, Strasbourg, France

    Université de Strasbourg, Strasbourg, France

    Institut Hospitalo-Universitaire, Pôle Hépato-digestif, Hôpitaux Universitaires de Strasbourg, Strasbourg, France
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  • Mirjam B. Zeisel
    Correspondence
    Corresponding authors. Address: Inserm, U1110, Institut de Recherche sur les Maladies Virales et Hépatiques, Université de Strasbourg, 3 Rue Koeberlé, 67000 Strasbourg, France. Tel.: +33 3 68 85 37 03; fax: +33 3 68 85 37 24.
    Affiliations
    Inserm, U1110, Institut de Recherche sur les Maladies Virales et Hépatiques, Strasbourg, France

    Université de Strasbourg, Strasbourg, France
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Open AccessPublished:October 09, 2014DOI:https://doi.org/10.1016/j.jhep.2014.10.004

      Summary

      Being the largest internal organ of the human body with the unique ability of self-regeneration, the liver is involved in a wide variety of vital functions that require highly orchestrated and controlled biochemical processes. Increasing evidence suggests that microRNAs (miRNAs) are essential for the regulation of liver development, regeneration and metabolic functions. Hence, alterations in intrahepatic miRNA networks have been associated with liver disease including hepatitis, steatosis, cirrhosis and hepatocellular carcinoma (HCC). miR-122 is the most frequent miRNA in the adult liver, and a central player in liver biology and disease. Furthermore, miR-122 has been shown to be an essential host factor for hepatitis C virus (HCV) infection and an antiviral target, complementary to the standard of care using direct-acting antivirals or interferon-based treatment. This review summarizes our current understanding of the key role of miR-122 in liver physiology and disease, highlighting its role in HCC and viral hepatitis. We also discuss the perspectives of miRNA-based therapeutic approaches for viral hepatitis and liver disease.

      Abbreviations:

      miRNA (microRNA), HCC (hepatocellular carcinoma), HCV (hepatitis C virus), RNAi (RNA interference), Ago (Argonaute), RISC (RNA-induced silencing complex), mRNA (messenger RNA), 3′ UTR (3′ untranslated region), 5′ UTR (5′ untranslated region), DAA (direct-acting antiviral), IFN (interferon), NAFLD (non-alcoholic fatty-liver disease), LEFT (liver-enriched transcription factor), HNF (hepatocyte nuclear factor), CUTL1 (cut-like homeobox 1), APK (AMP-activated protein kinase), PPAR (peroxisome proliferator-activated receptor), KO (knock-out), KLF6 (Krüppel-like factor 6), Ccl2 ((C-C) motif ligand 2), AKT3 (v-akt murine thymoma viral oncogene homolog 3), ADAM10 (disintegrin and metalloproteinase domain-containing protein 10), IGF1R (insulin-like growth factor-1 receptor), SRF (serum response factor), Wnt1 (wingless-type MMTV integration site family, member 1), PFV-1 (primate foamy virus type 1), BACH1 (BTB and CNC homology 1), HMOX1 (heme oxygenase 1), KSHV (Kaposi’s sarcoma-associated herpesvirus), HSV-1 (herpes simplex virus-1), HCMV (human cytomegalovirus), HBsAg (hepatitis B surface antigen), IFITM1 (interferon induced transmembrane protein 1), HBV (hepatitis B virus), HBx (hepatitis B virus X protein), Akt (v-akt murine thymoma viral oncogene homolog 1), IRES (internal ribosome entry site), SVR (sustained virological response), rcDNA (relaxed circular partially double-stranded genome), cccDNA (covalently closed circular DNA), Gld2 (germline development 2), NDRG3 (N-myc downstream regulated gene 3), PTTG1 (pituitary tumor-transforming gene 1-binding factor)

      Keywords

      miRNAs and disease biology

      Given their involvement in regulating cell homeostasis and functions, miRNA expression is tightly controlled in a temporally restrained and tissue-specific manner [
      • Bartel D.P.
      MicroRNAs: genomics, biogenesis, mechanism, and function.
      ,
      • Landgraf P.
      • Rusu M.
      • Sheridan R.
      • Sewer A.
      • Iovino N.
      • Aravin A.
      • et al.
      A mammalian microRNA expression atlas based on small RNA library sequencing.
      ]. This suggests that miRNAs may be involved in determining and maintaining tissue identity. These specific expression patterns are controlled by both transcriptional and post-transcriptional regulatory systems that may target different steps of miRNA biogenesis and turnover (for a detailed discussion, see [
      • Krol J.
      • Loedige I.
      • Filipowicz W.
      The widespread regulation of microRNA biogenesis, function and decay.
      ]). It is thus not surprising that dysregulations of miRNA networks have been associated with various diseases. Indeed, several pieces of evidence have demonstrated that altered regulation of miRNA expression might contribute to disease processes, including genetic and infectious diseases as well as cancer. While some diseases have been linked to the altered functions of enzymes regulating miRNA biogenesis, others appear to involve altered modulation of miRNA expression or genetic alterations of genes, encoding miRNAs or their targets, including deletions and single-nucleotide polymorphisms that may ultimately lead to a gain or loss of miRNA-target interaction (reviewed in [
      • Kloosterman W.P.
      • Plasterk R.H.A.
      The diverse functions of microRNAs in animal development and disease.
      ,
      • Bandiera S.
      • Hatem E.
      • Lyonnet S.
      • Henrion-Caude A.
      MicroRNAs in diseases: from candidate to modifier genes.
      ,
      • Hrdlickova B.
      • de Almeida R.C.
      • Borek Z.
      • Withoff S.
      Genetic variation in the non-coding genome: involvement of micro-RNAs and long non-coding RNAs in disease.
      ]). Therefore, miRNAs represent potentially interesting druggable targets. Indeed, a miR-122 inhibitor (miravirsen) and a miR-34 mimic (MRX34) were the first miRNA-based molecules to enter the clinic [
      • Janssen H.L.A.
      • Reesink H.W.
      • Lawitz E.J.
      • Zeuzem S.
      • Rodriguez-Torres M.
      • Patel K.
      • et al.
      Treatment of HCV infection by targeting microRNA.
      ,
      • Agostini M.
      • Knight R.A.
      MiR-34: from bench to bedside.
      ]. First, clinical trials have provided the proof-of-concept of the potential of miravirsen as a novel therapeutic strategy against chronic hepatitis C virus (HCV) infection, complementary to the standard of care using direct-acting antivirals (DAAs) or interferon (IFN)-based treatment [
      • Janssen H.L.A.
      • Reesink H.W.
      • Lawitz E.J.
      • Zeuzem S.
      • Rodriguez-Torres M.
      • Patel K.
      • et al.
      Treatment of HCV infection by targeting microRNA.
      ]. MRX34 is currently in a phase 1 clinical trial in patients with unresectable primary liver cancer, and advanced or metastatic cancer with liver involvement (ClinicalTrials.gov identifier: NCT01829971A) [
      • Agostini M.
      • Knight R.A.
      MiR-34: from bench to bedside.
      ]. Furthermore, given the association of differential miRNA expression patterns with diseases, both tissue and circulating miRNA expression profiles can also be used as biomarkers for diagnostic, prognostic and therapeutic purposes.
      The liver is the largest internal organ of the human body with the unique ability of self-regeneration. It is involved in a wide variety of vital functions that require highly orchestrated and controlled biochemical processes. Increasing evidence suggests that miRNAs are essential for the regulation of liver development, regeneration and metabolic functions [
      • Chen Y.
      • Verfaillie C.M.
      MicroRNAs: the fine modulators of liver development and function.
      ]. Hence, alterations in intrahepatic miRNA networks have been associated with all aspects of liver disease, including hepatitis, steatosis, cirrhosis and HCC (reviewed in [
      • Szabo G.
      • Bala S.
      MicroRNAs in liver disease.
      ]). miR-122 is the most frequent miRNA in the adult liver [
      • Lagos-Quintana M.
      • Rauhut R.
      • Yalcin A.
      • Meyer J.
      • Lendeckel W.
      • Tuschl T.
      Identification of tissue-specific microRNAs from mouse.
      ,
      • Girard M.
      • Jacquemin E.
      • Munnich A.
      • Lyonnet S.
      • Henrion-Caude A.
      MiR-122, a paradigm for the role of microRNAs in the liver.
      ,
      • Hou W.
      • Tian Q.
      • Zheng J.
      • Bonkovsky H.L.
      MicroRNA-196 represses Bach1 protein and hepatitis C virus gene expression in human hepatoma cells expressing hepatitis C viral proteins.
      ]. Interestingly, miR-122 can be detected in the circulation and serum miR-122 has been shown to serve as a biomarker of liver injury in chronic hepatitis B or C, non-alcoholic fatty-liver disease (NAFLD) and drug-induced liver disease [
      • Cermelli S.
      • Ruggieri A.
      • Marrero J.A.
      • Ioannou G.N.
      • Beretta L.
      Circulating microRNAs in patients with chronic hepatitis C and non-alcoholic fatty liver disease.
      ,
      • Starkey Lewis P.J.
      • Dear J.
      • Platt V.
      • Simpson K.J.
      • Craig D.G.N.
      • Antoine D.J.
      • et al.
      Circulating microRNAs as potential markers of human drug-induced liver injury.
      ,
      • Waidmann O.
      • Bihrer V.
      • Pleli T.
      • Farnik H.
      • Berger A.
      • Zeuzem S.
      • et al.
      Serum microRNA-122 levels in different groups of patients with chronic hepatitis B virus infection.
      ,
      • Van der Meer A.J.
      • Farid W.R.R.
      • Sonneveld M.J.
      • de Ruiter P.E.
      • Boonstra A.
      • van Vuuren A.J.
      • et al.
      Sensitive detection of hepatocellular injury in chronic hepatitis C patients with circulating hepatocyte-derived microRNA-122.
      ,
      • Köberle V.
      • Waidmann O.
      • Kronenberger B.
      • Andrei A.
      • Susser S.
      • Füller C.
      • et al.
      Serum microRNA-122 kinetics in patients with chronic hepatitis C virus infection during antiviral therapy.
      ,
      • Miyaaki H.
      • Ichikawa T.
      • Kamo Y.
      • Taura N.
      • Honda T.
      • Shibata H.
      • et al.
      Significance of serum and hepatic microRNA-122 levels in patients with non-alcoholic fatty liver disease.
      ,
      • Yamada H.
      • Suzuki K.
      • Ichino N.
      • Ando Y.
      • Sawada A.
      • Osakabe K.
      • et al.
      Associations between circulating microRNAs (miR-21, miR-34a, miR-122, and miR-451) and non-alcoholic fatty liver.
      ]. Here, we review the key involvement of miR-122 in liver physiology and disease, highlighting its roles in HCC and viral hepatitis. We also discuss the perspectives of miRNA-based therapeutic approaches for viral hepatitis and liver disease.

      miR-122 and liver physiology

      miR-122 has a liver-enriched expression and is one of the most abundant miRNAs in the liver, accounting for about 70% and 52% of the whole hepatic miRNome in adult mouse and human, respectively [
      • Lagos-Quintana M.
      • Rauhut R.
      • Yalcin A.
      • Meyer J.
      • Lendeckel W.
      • Tuschl T.
      Identification of tissue-specific microRNAs from mouse.
      ,
      • Girard M.
      • Jacquemin E.
      • Munnich A.
      • Lyonnet S.
      • Henrion-Caude A.
      MiR-122, a paradigm for the role of microRNAs in the liver.
      ,
      • Hou W.
      • Tian Q.
      • Zheng J.
      • Bonkovsky H.L.
      MicroRNA-196 represses Bach1 protein and hepatitis C virus gene expression in human hepatoma cells expressing hepatitis C viral proteins.
      ]. Consequently, miR-122 plays a central role in liver development, differentiation, homeostasis and functions (Fig. 1). miR-122 expression is driven by liver-enriched transcription factors (LETFs), including hepatocyte nuclear factor (HNF) 6 and 4a [
      • Xu H.
      • He J.-H.
      • Xiao Z.-D.
      • Zhang Q.-Q.
      • Chen Y.-Q.
      • Zhou H.
      • et al.
      Liver-enriched transcription factors regulate microRNA-122 that targets CUTL1 during liver development.
      ,
      • Laudadio I.
      • Manfroid I.
      • Achouri Y.
      • Schmidt D.
      • Wilson M.D.
      • Cordi S.
      • et al.
      A feedback loop between the liver-enriched transcription factor network and miR-122 controls hepatocyte differentiation.
      ,
      • Deng X.-G.
      • Qiu R.-L.
      • Wu Y.-H.
      • Li Z.-X.
      • Xie P.
      • Zhang J.
      • et al.
      Overexpression of miR-122 promotes the hepatic differentiation and maturation of mouse ESCs through a miR-122/FoxA1/HNF4a-positive feedback loop.
      ] that also fine-tune miR-122 dosage during liver development in vivo [
      • Xu H.
      • He J.-H.
      • Xiao Z.-D.
      • Zhang Q.-Q.
      • Chen Y.-Q.
      • Zhou H.
      • et al.
      Liver-enriched transcription factors regulate microRNA-122 that targets CUTL1 during liver development.
      ,
      • Laudadio I.
      • Manfroid I.
      • Achouri Y.
      • Schmidt D.
      • Wilson M.D.
      • Cordi S.
      • et al.
      A feedback loop between the liver-enriched transcription factor network and miR-122 controls hepatocyte differentiation.
      ,
      • Deng X.-G.
      • Qiu R.-L.
      • Wu Y.-H.
      • Li Z.-X.
      • Xie P.
      • Zhang J.
      • et al.
      Overexpression of miR-122 promotes the hepatic differentiation and maturation of mouse ESCs through a miR-122/FoxA1/HNF4a-positive feedback loop.
      ]. Particularly in liver development, the concertized expression of miR-122 and LETFs was suggested to regulate the proper balance between cell proliferation and differentiation in both the hepatocyte and cholangiocyte lineages [
      • Xu H.
      • He J.-H.
      • Xiao Z.-D.
      • Zhang Q.-Q.
      • Chen Y.-Q.
      • Zhou H.
      • et al.
      Liver-enriched transcription factors regulate microRNA-122 that targets CUTL1 during liver development.
      ,
      • Laudadio I.
      • Manfroid I.
      • Achouri Y.
      • Schmidt D.
      • Wilson M.D.
      • Cordi S.
      • et al.
      A feedback loop between the liver-enriched transcription factor network and miR-122 controls hepatocyte differentiation.
      ]. This temporal-regulation of miR-122 expression is particularly important as miR-122 promotes hepatobiliary segregation along with the acquisition and maintenance of a hepato-specific phenotype [
      • Xu H.
      • He J.-H.
      • Xiao Z.-D.
      • Zhang Q.-Q.
      • Chen Y.-Q.
      • Zhou H.
      • et al.
      Liver-enriched transcription factors regulate microRNA-122 that targets CUTL1 during liver development.
      ,
      • Laudadio I.
      • Manfroid I.
      • Achouri Y.
      • Schmidt D.
      • Wilson M.D.
      • Cordi S.
      • et al.
      A feedback loop between the liver-enriched transcription factor network and miR-122 controls hepatocyte differentiation.
      ,
      • Coulouarn C.
      • Factor V.M.
      • Andersen J.B.
      • Durkin M.E.
      • Thorgeirsson S.S.
      Loss of miR-122 expression in liver cancer correlates with suppression of the hepatic phenotype and gain of metastatic properties.
      ] (Fig. 1). Indeed, during mouse liver development, miR-122 was shown to gradually repress the transcription factor cut-like homeobox 1 (CUTL1), thus allowing terminal liver differentiation [
      • Xu H.
      • He J.-H.
      • Xiao Z.-D.
      • Zhang Q.-Q.
      • Chen Y.-Q.
      • Zhou H.
      • et al.
      Liver-enriched transcription factors regulate microRNA-122 that targets CUTL1 during liver development.
      ] (Fig. 1). This important role of miR-122 in liver development and differentiation was further demonstrated by studies reporting that antisense-mediated inhibition of miR-122 delayed liver development in zebrafish [
      • Laudadio I.
      • Manfroid I.
      • Achouri Y.
      • Schmidt D.
      • Wilson M.D.
      • Cordi S.
      • et al.
      A feedback loop between the liver-enriched transcription factor network and miR-122 controls hepatocyte differentiation.
      ] and switched on the expression of genes that were normally repressed in the adult mouse liver [
      • Krützfeldt J.
      • Rajewsky N.
      • Braich R.
      • Rajeev K.G.
      • Tuschl T.
      • Manoharan M.
      • et al.
      Silencing of microRNAs in vivo with “antagomirs”.
      ]. This is also corroborated by the fact that the repression of miR-122 in primary HCC with poor prognosis was associated with suppression of the hepatic phenotype [
      • Coulouarn C.
      • Factor V.M.
      • Andersen J.B.
      • Durkin M.E.
      • Thorgeirsson S.S.
      Loss of miR-122 expression in liver cancer correlates with suppression of the hepatic phenotype and gain of metastatic properties.
      ].
      Figure thumbnail gr1
      Fig. 1miR-122 is a key regulator of liver physiology and disease biology. The scheme illustrates the different roles of miR-122 in liver development and metabolism (red boxes) as well as in viral hepatitis and liver disease. Activation (+) or inhibition (−) is indicated dependent on the effect of miR-122 on a specific process. While host miR-122 targets are depicted outside of boxes, miR-122 targets of viral origin are indicated within grey boxes.
      miR-122 also plays a crucial role in the regulation of cholesterol and fatty acid metabolism in the adult liver (Fig. 1). In vivo antisense studies, coupled with microarray analysis, have been instrumental to uncover the role of miR-122 in lipid metabolism [
      • Krützfeldt J.
      • Rajewsky N.
      • Braich R.
      • Rajeev K.G.
      • Tuschl T.
      • Manoharan M.
      • et al.
      Silencing of microRNAs in vivo with “antagomirs”.
      ,
      • Esau C.
      • Davis S.
      • Murray S.F.
      • Yu X.X.
      • Pandey S.K.
      • Pear M.
      • et al.
      MiR-122 regulation of lipid metabolism revealed by in vivo antisense targeting.
      ,
      • Elmén J.
      • Lindow M.
      • Schütz S.
      • Lawrence M.
      • Petri A.
      • Obad S.
      • et al.
      LNA-mediated microRNA silencing in non-human primates.
      ]. Indeed, antisense-mediated inhibition of hepatic miR-122 markedly lowered plasma cholesterol levels in both mice and non-human primates [
      • Krützfeldt J.
      • Rajewsky N.
      • Braich R.
      • Rajeev K.G.
      • Tuschl T.
      • Manoharan M.
      • et al.
      Silencing of microRNAs in vivo with “antagomirs”.
      ,
      • Esau C.
      • Davis S.
      • Murray S.F.
      • Yu X.X.
      • Pandey S.K.
      • Pear M.
      • et al.
      MiR-122 regulation of lipid metabolism revealed by in vivo antisense targeting.
      ,
      • Elmén J.
      • Lindow M.
      • Schütz S.
      • Lawrence M.
      • Petri A.
      • Obad S.
      • et al.
      LNA-mediated microRNA silencing in non-human primates.
      ]. Transcriptomic analyses in mice further revealed that transient miR-122 sequestration downregulated the expression of genes involved in fatty acid metabolism as well as cholesterol biosynthesis, including the rate-limiting enzyme 3-hydroxy-3-methylglutaryl-CoA-reductase [
      • Krützfeldt J.
      • Rajewsky N.
      • Braich R.
      • Rajeev K.G.
      • Tuschl T.
      • Manoharan M.
      • et al.
      Silencing of microRNAs in vivo with “antagomirs”.
      ,
      • Esau C.
      • Davis S.
      • Murray S.F.
      • Yu X.X.
      • Pandey S.K.
      • Pear M.
      • et al.
      MiR-122 regulation of lipid metabolism revealed by in vivo antisense targeting.
      ]. Although the molecular mechanisms underlying regulation of lipid homeostasis by miR-122 are still unclear, both AMP-activated protein kinase (APK) and circadian metabolic regulators of the peroxisome proliferator-activated receptor (PPAR) family were suggested to be putative effectors of miR-122-mediated metabolic control [
      • Esau C.
      • Davis S.
      • Murray S.F.
      • Yu X.X.
      • Pandey S.K.
      • Pear M.
      • et al.
      MiR-122 regulation of lipid metabolism revealed by in vivo antisense targeting.
      ,
      • Gatfield D.
      • Le Martelot G.
      • Vejnar C.E.
      • Gerlach D.
      • Schaad O.
      • Fleury-Olela F.
      • et al.
      Integration of microRNA miR-122 in hepatic circadian gene expression.
      ] (Fig. 1). Interestingly, transcription of the miR-122 locus itself occurs in a circadian manner, suggesting the existence of a link between miR-122, circadian gene expression and hepatic lipid metabolism [
      • Gatfield D.
      • Le Martelot G.
      • Vejnar C.E.
      • Gerlach D.
      • Schaad O.
      • Fleury-Olela F.
      • et al.
      Integration of microRNA miR-122 in hepatic circadian gene expression.
      ].

      miR-122 and pathogenesis of liver disease and hepatocellular carcinoma

      In line with its essential role in maintaining liver homeostasis and differentiation, reduced expression of miR-122 has been associated with liver disease. The generation of both germline knock-out (KO) mice and liver-specific KO mice has been pivotal to revealing a key involvement of miR-122 in liver disease [
      • Hsu S.-H.
      • Wang B.
      • Kota J.
      • Yu J.
      • Costinean S.
      • Kutay H.
      • et al.
      Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.
      ,
      • Tsai W.-C.
      • Hsu S.-D.
      • Hsu C.-S.
      • Lai T.-C.
      • Chen S.-J.
      • Shen R.
      • et al.
      MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.
      ,
      • Zeisel M.B.
      • Pfeffer S.
      • Baumert T.F.
      MiR-122 acts as a tumor suppressor in hepatocarcinogenesis in vivo.
      ]. Indeed, in contrast to transient miR-122 sequestration, genetic deletion of miR-122 was shown not only to severely impact on lipid metabolism, but also to drive microsteatosis and inflammation, which progressed to steatohepatitis and fibrosis as mice aged [
      • Hsu S.-H.
      • Wang B.
      • Kota J.
      • Yu J.
      • Costinean S.
      • Kutay H.
      • et al.
      Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.
      ,
      • Tsai W.-C.
      • Hsu S.-D.
      • Hsu C.-S.
      • Lai T.-C.
      • Chen S.-J.
      • Shen R.
      • et al.
      MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.
      ]. Consistently, miR-122 expression was also lowered in a carbon tetrachloride-induced mouse model of liver fibrosis [
      • Li J.
      • Ghazwani M.
      • Zhang Y.
      • Lu J.
      • Li J.
      • Fan J.
      • et al.
      MiR-122 regulates collagen production via targeting hepatic stellate cells and suppressing P4HA1 expression.
      ]. Of note, the restoration of miR-122 levels in miR-122 KO mice reversed liver inflammation, at least in part, by repressing two miR-122 targets, namely the chemokine Ccl2, which was shown to recruit CD11bhiGr1+ inflammatory cells intrahepatically [
      • Hsu S.-H.
      • Wang B.
      • Kota J.
      • Yu J.
      • Costinean S.
      • Kutay H.
      • et al.
      Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.
      ] and the pro-fibrogenic Krüppel-like factor 6 (KLF6), whose expression was enhanced in the miR-122 KO mouse liver [
      • Tsai W.-C.
      • Hsu S.-D.
      • Hsu C.-S.
      • Lai T.-C.
      • Chen S.-J.
      • Shen R.
      • et al.
      MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.
      ]. This piece of data clearly highlights the anti-inflammatory and anti-fibrotic properties of miR-122 in the liver (Fig. 2). Although this knowledge has been acquired using mouse models, it is important to note that reduced miR-122 expression has been associated with human non-alcoholic steatohepatitis [
      • Cheung O.
      • Puri P.
      • Eicken C.
      • Contos M.J.
      • Mirshahi F.
      • Maher J.W.
      • et al.
      Nonalcoholic steatohepatitis is associated with altered hepatic MicroRNA expression.
      ] extending the relevance of these findings to human liver disease. Furthermore, decreased miR-122 levels have been associated with poor prognosis and metastasis of liver cancer, and several targets of miR-122 have been implicated in tumourigenesis [
      • Hsu S.-H.
      • Wang B.
      • Kota J.
      • Yu J.
      • Costinean S.
      • Kutay H.
      • et al.
      Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.
      ,
      • Coulouarn C.
      • Corlu A.
      • Glaise D.
      • Guénon I.
      • Thorgeirsson S.S.
      • Clément B.
      Hepatocyte-stellate cell cross-talk in the liver engenders a permissive inflammatory microenvironment that drives progression in hepatocellular carcinoma.
      ,
      • Bai S.
      • Nasser M.W.
      • Wang B.
      • Hsu S.-H.
      • Datta J.
      • Kutay H.
      • et al.
      MicroRNA-122 inhibits tumorigenic properties of hepatocellular carcinoma cells and sensitizes these cells to sorafenib.
      ,
      • Fornari F.
      • Gramantieri L.
      • Giovannini C.
      • Veronese A.
      • Ferracin M.
      • Sabbioni S.
      • et al.
      MiR-122/cyclin G1 interaction modulates p53 activity and affects doxorubicin sensitivity of human hepatocarcinoma cells.
      ,
      • Zeng C.
      • Wang R.
      • Li D.
      • Lin X.-J.
      • Wei Q.-K.
      • Yuan Y.
      • et al.
      A novel GSK-3 beta-C/EBP alpha-miR-122-insulin-like growth factor 1 receptor regulatory circuitry in human hepatocellular carcinoma.
      ,
      • Nassirpour R.
      • Mehta P.P.
      • Yin M.-J.
      MiR-122 regulates tumorigenesis in hepatocellular carcinoma by targeting AKT3.
      ,
      • Xu J.
      • Zhu X.
      • Wu L.
      • Yang R.
      • Yang Z.
      • Wang Q.
      • et al.
      MicroRNA-122 suppresses cell proliferation and induces cell apoptosis in hepatocellular carcinoma by directly targeting Wnt/β-catenin pathway.
      ,
      • Nakao K.
      • Miyaaki H.
      • Ichikawa T.
      Antitumor function of microRNA-122 against hepatocellular carcinoma.
      ] (Fig. 1). Indeed, a number of validated miR-122 targets including cyclin G1, ADAM10, IGF1R, SRF, and Wnt1, were shown to be involved in hepatocarcinogenesis, epithelial-mesenchymal transition, and angiogenesis [
      • Nakao K.
      • Miyaaki H.
      • Ichikawa T.
      Antitumor function of microRNA-122 against hepatocellular carcinoma.
      ] (Fig. 1). Altogether, these data suggested that miR-122 acts as a tumour suppressor in the liver.
      Figure thumbnail gr2
      Fig. 2Therapeutic effects of miR-122-modulating agents in liver disease. Current state-of-the-art approaches in modulating miRNAs in vivo comprise restoration of miRNA expression, using synthetic miRNA mimics or viral vectors driving miRNA expression, as well as inhibition of miRNA expression via chemically modified anti-miR oligonucleotides
      [
      • van Rooij E.
      • Kauppinen S.
      Development of microRNA therapeutics is coming of age.
      ]
      . While antisense-mediated inhibition of miR-122 (anti-miR-122) has been demonstrated of clinical interest to treat chronic HCV infection and to represent a potential therapeutic strategy against hypercholesterolemia, restoration of miR-122 (miR-122 mimics), was suggested as a therapeutic approach against liver fibrosis and HCC development.
      The proof-of-concept that miR-122 has an anti-tumour function in the liver was again provided using miR-122 KO mice [
      • Hsu S.-H.
      • Wang B.
      • Kota J.
      • Yu J.
      • Costinean S.
      • Kutay H.
      • et al.
      Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.
      ,
      • Tsai W.-C.
      • Hsu S.-D.
      • Hsu C.-S.
      • Lai T.-C.
      • Chen S.-J.
      • Shen R.
      • et al.
      MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.
      ]. These mice spontaneously develop liver tumours and demonstrate abnormal expression of genes involved in cell growth and cell death, epithelial-mesenchymal transition and cancer [
      • Hsu S.-H.
      • Wang B.
      • Kota J.
      • Yu J.
      • Costinean S.
      • Kutay H.
      • et al.
      Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.
      ,
      • Tsai W.-C.
      • Hsu S.-D.
      • Hsu C.-S.
      • Lai T.-C.
      • Chen S.-J.
      • Shen R.
      • et al.
      MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.
      ]. Importantly, tumour development in these mice could be prevented by restoration of miR-122 expression in vivo [
      • Hsu S.-H.
      • Wang B.
      • Kota J.
      • Yu J.
      • Costinean S.
      • Kutay H.
      • et al.
      Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.
      ,
      • Tsai W.-C.
      • Hsu S.-D.
      • Hsu C.-S.
      • Lai T.-C.
      • Chen S.-J.
      • Shen R.
      • et al.
      MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.
      ]. Moreover, by using a mouse model where tumours developed in the absence of inflammation, it has been demonstrated that miR-122 has an anti-tumour function that is independent of its role in preventing liver disease and inflammation [
      • Hsu S.-H.
      • Wang B.
      • Kota J.
      • Yu J.
      • Costinean S.
      • Kutay H.
      • et al.
      Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.
      ]. miR-122 may thus be used as a potential therapeutic tool against HCC. Indeed, given that the decrease of miR-122 can promote hepatocarcinogenesis, and that restoration of miR-122 in HCC cells can reverse the tumourigenic properties of these cells, preventing HCC development in vivo [
      • Hsu S.-H.
      • Wang B.
      • Kota J.
      • Yu J.
      • Costinean S.
      • Kutay H.
      • et al.
      Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.
      ,
      • Tsai W.-C.
      • Hsu S.-D.
      • Hsu C.-S.
      • Lai T.-C.
      • Chen S.-J.
      • Shen R.
      • et al.
      MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.
      ,
      • Coulouarn C.
      • Corlu A.
      • Glaise D.
      • Guénon I.
      • Thorgeirsson S.S.
      • Clément B.
      Hepatocyte-stellate cell cross-talk in the liver engenders a permissive inflammatory microenvironment that drives progression in hepatocellular carcinoma.
      ,
      • Bai S.
      • Nasser M.W.
      • Wang B.
      • Hsu S.-H.
      • Datta J.
      • Kutay H.
      • et al.
      MicroRNA-122 inhibits tumorigenic properties of hepatocellular carcinoma cells and sensitizes these cells to sorafenib.
      ,
      • Fornari F.
      • Gramantieri L.
      • Giovannini C.
      • Veronese A.
      • Ferracin M.
      • Sabbioni S.
      • et al.
      MiR-122/cyclin G1 interaction modulates p53 activity and affects doxorubicin sensitivity of human hepatocarcinoma cells.
      ,
      • Xu Y.
      • Xia F.
      • Ma L.
      • Shan J.
      • Shen J.
      • Yang Z.
      • et al.
      MicroRNA-122 sensitizes HCC cancer cells to adriamycin and vincristine through modulating expression of MDR and inducing cell cycle arrest.
      ,
      • Hsu S.-H.
      • Yu B.
      • Wang X.
      • Lu Y.
      • Schmidt C.R.
      • Lee R.J.
      • et al.
      Cationic lipid nanoparticles for therapeutic delivery of siRNA and miRNA to murine liver tumor.
      ], miR-122 mimics represent an interesting strategy to prevent and treat HCC (Fig. 2). Furthermore, it has also been shown that restoration of miR-122 also sensitizes HCC cells to chemotherapy, suggesting that combination of miR-122 and chemotherapeutic agents may have an additive or synergistic effect against liver cancer [
      • Bai S.
      • Nasser M.W.
      • Wang B.
      • Hsu S.-H.
      • Datta J.
      • Kutay H.
      • et al.
      MicroRNA-122 inhibits tumorigenic properties of hepatocellular carcinoma cells and sensitizes these cells to sorafenib.
      ,
      • Fornari F.
      • Gramantieri L.
      • Giovannini C.
      • Veronese A.
      • Ferracin M.
      • Sabbioni S.
      • et al.
      MiR-122/cyclin G1 interaction modulates p53 activity and affects doxorubicin sensitivity of human hepatocarcinoma cells.
      ,
      • Xu Y.
      • Xia F.
      • Ma L.
      • Shan J.
      • Shen J.
      • Yang Z.
      • et al.
      MicroRNA-122 sensitizes HCC cancer cells to adriamycin and vincristine through modulating expression of MDR and inducing cell cycle arrest.
      ]. It is worth noting that the first miRNA mimic reached phase 1 clinical studies, indicating the feasibility of modulating miRNA expression in human liver (ClinicalTrials.gov identifier: NCT01829971A) [
      • Agostini M.
      • Knight R.A.
      MiR-34: from bench to bedside.
      ]. Taken together, results from these studies broadened our understanding of HCC-development, enabled researchers to draw arresting conclusions regarding the association between loss of miR-122 and diverse aspects of liver disease as well as HCC, and highlighted important implications regarding the therapeutic potential of miR-122 [
      • Hsu S.-H.
      • Wang B.
      • Kota J.
      • Yu J.
      • Costinean S.
      • Kutay H.
      • et al.
      Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.
      ,
      • Tsai W.-C.
      • Hsu S.-D.
      • Hsu C.-S.
      • Lai T.-C.
      • Chen S.-J.
      • Shen R.
      • et al.
      MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.
      ,
      • Zeisel M.B.
      • Pfeffer S.
      • Baumert T.F.
      MiR-122 acts as a tumor suppressor in hepatocarcinogenesis in vivo.
      ].
      Despite the fact that proof-of-concept studies have elegantly demonstrated the tumour suppressor function of miR-122 [
      • Hsu S.-H.
      • Wang B.
      • Kota J.
      • Yu J.
      • Costinean S.
      • Kutay H.
      • et al.
      Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.
      ,
      • Tsai W.-C.
      • Hsu S.-D.
      • Hsu C.-S.
      • Lai T.-C.
      • Chen S.-J.
      • Shen R.
      • et al.
      MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.
      ], it is important to point out that HCC is not consistently associated with loss of miR-122. Indeed, HCC is a multifactorial and heterogeneous disease and miR-122 expression appears to be dependent on the aetiology of the liver cancer. Interestingly, reduced miR-122 expression has been associated with hepatitis B virus (HBV)-related HCC, while miR-122 levels appear normal or increased in HCV-related HCC [
      • Varnholt H.
      • Drebber U.
      • Schulze F.
      • Wedemeyer I.
      • Schirmacher P.
      • Dienes H.-P.
      • et al.
      MicroRNA gene expression profile of hepatitis C virus-associated hepatocellular carcinoma.
      ,
      • Spaniel C.
      • Honda M.
      • Selitsky S.R.
      • Yamane D.
      • Shimakami T.
      • Kaneko S.
      • et al.
      MicroRNA-122 abundance in hepatocellular carcinoma and non-tumor liver tissue from Japanese patients with persistent HCV versus HBV infection.
      ]. One can hypothesize that this is due to different roles of miR-122 in the life cycle of these two viruses (see below), and at least with respect to miR-122, each of the two viruses causes HCC in different ways (Fig. 1). These data underscore that HCC is not the result of the deregulated expression of a single gene, and rather several lines of evidence indicate that various signalling pathways are deregulated in HCC (reviewed in [
      • Szabo G.
      • Bala S.
      MicroRNAs in liver disease.
      ,
      • Negrini M.
      • Gramantieri L.
      • Sabbioni S.
      • Croce C.M.
      MicroRNA involvement in hepatocellular carcinoma.
      ,
      • Imbeaud S.
      • Ladeiro Y.
      • Zucman-Rossi J.
      Identification of novel oncogenes and tumor suppressors in hepatocellular carcinoma.
      ]). Further studies are required to better understand the molecular mechanisms underlying HCC and the role of miRNAs in this disease.

      miRNAs and virus-host interactions

      Chronic viral hepatitis due to HBV or HCV infection is a major cause of chronic liver disease and HCC. HBV and HCV are both characterized by a tight species and tissue tropism, almost exclusively infecting human hepatocytes. This cell specificity may be explained by the fact that both viruses depend at each step of their respective life cycle on several host factors, which happen to be expressed in hepatocytes. Within the past years, numerous proteins have been uncovered to be required for either the HBV or the HCV life cycle, and increasing evidence indicates that non-protein-coding RNAs, such as miRNAs also plays important roles in these processes (reviewed in [
      • Zeisel M.B.
      • Lupberger J.
      • Fofana I.
      • Baumert T.F.
      Host-targeting agents for prevention and treatment of chronic hepatitis C – Perspectives and challenges.
      ,
      • Lohmann V.
      Hepatitis C virus RNA replication.
      ,
      • Lindenbach B.D.
      Virion assembly and release.
      ,
      • Grimm D.
      • Thimme R.
      • Blum H.E.
      HBV life cycle and novel drug targets.
      ,
      • Baumert T.F.
      • Meredith L.
      • Ni Y.
      • Felmlee D.J.
      • McKeating J.A.
      • Urban S.
      Entry of hepatitis B and C viruses – Recent progress and future impact.
      ,
      • Van der Ree M.H.
      • de Bruijne J.
      • Kootstra N.A.
      • Jansen P.L.
      • Reesink H.W.
      MicroRNAs: role and therapeutic targets in viral hepatitis.
      ]). Furthermore, in addition of using host miRNAs for their replicative cycle, HBV and HCV have also been reported to modulate the expression profile of the cellular miRNome to favour viral persistence, which may contribute to pathogenesis of liver disease (reviewed in [
      • Liu W.-H.
      • Yeh S.-H.
      • Chen P.-J.
      Role of microRNAs in hepatitis B virus replication and pathogenesis.
      ,
      • Shrivastava S.
      • Mukherjee A.
      • Ray R.B.
      Hepatitis C virus infection, microRNA and liver disease progression.
      ]). Accumulating evidence points to a role of human miRNAs in modulating viral infectivity, cell tropism and host immune responses [
      • Ghosh Z.
      • Mallick B.
      • Chakrabarti J.
      Cellular versus viral microRNAs in host-virus interaction.
      ,
      • Roberts A.P.E.
      • Lewis A.P.
      • Jopling C.L.
      The role of microRNAs in viral infection.
      ]. The outcome of this miRNA-virus interplay can have either a positive (proviral) or negative (antiviral) effect on the virus. In addition, there are different levels of interactions, which are not mutually exclusive, as described below.
      Cellular miRNAs have been demonstrated to directly target defined viral genomes or transcripts (Table 1). The best described example so far is the binding of miR-122 within the HCV genomic RNA that has a positive effect on viral translation, replication and infectious particle production (see below) [
      • Jopling C.L.
      • Yi M.
      • Lancaster A.M.
      • Lemon S.M.
      • Sarnow P.
      Modulation of hepatitis C virus RNA abundance by a liver-specific MicroRNA.
      ,
      • Villanueva R.A.
      • Jangra R.K.
      • Yi M.
      • Pyles R.
      • Bourne N.
      • Lemon S.M.
      MiR-122 does not modulate the elongation phase of hepatitis C virus RNA synthesis in isolated replicase complexes.
      ]. Actually, the positive outcome of miR-122 for HCV is more of an oddity than the rule, as most direct binding of miRNAs to viral RNAs is deleterious. Indeed, miR-199a also directly targets HCV RNA but this leads to an inhibition of HCV replication [
      • Murakami Y.
      • Aly H.H.
      • Tajima A.
      • Inoue I.
      • Shimotohno K.
      Regulation of the hepatitis C virus genome replication by miR-199a.
      ]. Likewise, HBV transcripts have also been reported to contain binding sites for cellular miRNAs, including miR-122, miR-199a, and miR-210 that all repress HBV mRNA expression [
      • Chen Y.
      • Shen A.
      • Rider P.J.
      • Yu Y.
      • Wu K.
      • Mu Y.
      • et al.
      A liver-specific microRNA binds to a highly conserved RNA sequence of hepatitis B virus and negatively regulates viral gene expression and replication.
      ,
      • Zhang G.
      • Li Y.
      • Zheng S.
      • Liu M.
      • Li X.
      • Tang H.
      Suppression of hepatitis B virus replication by microRNA-199a-3p and microRNA-210.
      ]. Noteworthy, to counteract inhibition by cellular miRNAs, RNA viruses appear to have evolved strategies to escape direct miRNA-mediated repression. Indeed, a recent comprehensive survey on the roles of miRNAs in different virus infections using Dicer KO HEK293 cells indicated that miRNAs had only a limited impact on the viruses tested, and hence that most of the viruses have evolved to be resistant to cellular miRNAs [
      • Bogerd H.P.
      • Skalsky R.L.
      • Kennedy E.M.
      • Furuse Y.
      • Whisnant A.W.
      • Flores O.
      • et al.
      Replication of many human viruses is refractory to inhibition by endogenous cellular microRNAs.
      ]. While the molecular mechanisms underlying viral evasion from miRNAs remain to be determined, first evidence indicated that HIV-1 was able to adopt extensive RNA secondary structures to avoid efficient inhibition by host miRNAs [
      • Whisnant A.W.
      • Bogerd H.P.
      • Flores O.
      • Ho P.
      • Powers J.G.
      • Sharova N.
      • et al.
      In-depth analysis of the interaction of HIV-1 with cellular microRNA biogenesis and effector mechanisms.
      ]. Taken together, these data suggest that the crosstalk between miRNAs and viral RNAs likely lead viruses to develop strategies to escape antiviral immunity and indicate that the dependence on a host miRNA as seen with miR-122 and HCV is rare [
      • Bogerd H.P.
      • Skalsky R.L.
      • Kennedy E.M.
      • Furuse Y.
      • Whisnant A.W.
      • Flores O.
      • et al.
      Replication of many human viruses is refractory to inhibition by endogenous cellular microRNAs.
      ].
      Table 1miRNA-mediated regulation of viral infection.
      Host miRNAs are also able to indirectly target a virus through the miRNA-mediated regulation of specific host factors (Table 1). This kind of interaction has for example been described in the context of the antiviral response to HCV infection. Indeed, recent studies indicated that miR-196 may play a role in counteracting HCV infection in vitro by both enhancing antioxidant and anti-inflammatory responses and direct targeting of the HCV genome [
      • Hou W.
      • Tian Q.
      • Zheng J.
      • Bonkovsky H.L.
      MicroRNA-196 represses Bach1 protein and hepatitis C virus gene expression in human hepatoma cells expressing hepatitis C viral proteins.
      ,
      • Pedersen I.M.
      • Cheng G.
      • Wieland S.
      • Volinia S.
      • Croce C.M.
      • Chisari F.V.
      • et al.
      Interferon modulation of cellular microRNAs as an antiviral mechanism.
      ], which merits further validation in vivo. Furthermore, HBV replication has been shown to be regulated by different miRNAs, which modulate the expression of transcription factors, having an impact on the virus life cycle [
      • Hu W.
      • Wang X.
      • Ding X.
      • Li Y.
      • Zhang X.
      • Xie P.
      • et al.
      MicroRNA-141 represses HBV replication by targeting PPARA.
      ,
      • Zhang X.
      • Zhang E.
      • Ma Z.
      • Pei R.
      • Jiang M.
      • Schlaak J.F.
      • et al.
      Modulation of hepatitis B virus replication and hepatocyte differentiation by MicroRNA-1.
      ]. Given the widely spread regulation of cellular proteins by cellular miRNAs, it is likely that the tuning of host cell gene expression by host miRNAs contributes to modulating viral life cycles.
      If host miRNAs modulate viral RNA expression, likewise viruses can impact on host miRNA expression, which in turn could target either host or viral RNAs (Table 1). Viral infection has been reported to modulate the expression of miRNAs that can promote viral replication and/or contribute to viral evasion as well as pathogenesis. For instance, HCV infection promotes the expression of miRNAs that suppress the innate immune response pathways, thereby leading to an increase of viral replication [
      • Ishida H.
      • Tatsumi T.
      • Hosui A.
      • Nawa T.
      • Kodama T.
      • Shimizu S.
      • et al.
      Alterations in microRNA expression profile in HCV-infected hepatoma cells: Involvement of miR-491 in regulation of HCV replication via the PI3 kinase/Akt pathway.
      ,
      • Bhanja Chowdhury J.
      • Shrivastava S.
      • Steele R.
      • Di Bisceglie A.M.
      • Ray R.
      • Ray R.B.
      Hepatitis C virus infection modulates expression of interferon stimulatory gene IFITM1 by upregulating miR-130A.
      ,
      • Chen Y.
      • Chen J.
      • Wang H.
      • Shi J.
      • Wu K.
      • Liu S.
      • et al.
      HCV-induced miR-21 contributes to evasion of host immune system by targeting MyD88 and IRAK1.
      ]. Furthermore, the HBV X protein (HBx) has been reported to modulate the expression of cellular miRNAs that likely contribute to the pathogenesis of liver disease [
      • Wang Y.
      • Lu Y.
      • Toh S.T.
      • Sung W.-K.
      • Tan P.
      • Chow P.
      • et al.
      Lethal-7 is down-regulated by the hepatitis B virus x protein and targets signal transducer and activator of transcription 3.
      ,
      • Dai X.
      • Zhang W.
      • Zhang H.
      • Sun S.
      • Yu H.
      • Guo Y.
      • et al.
      Modulation of HBV replication by microRNA-15b through targeting hepatocyte nuclear factor 1α.
      ]. Beside viral proteins, virus-encoded transcripts can also play a role in regulating miRNA abundance in host cells by degrading miRNAs or interfering with their biogenesis [
      • Cazalla D.
      • Yario T.
      • Steitz J.A.
      • Steitz J.
      Down-regulation of a host microRNA by a Herpesvirus saimiri noncoding RNA.
      ,
      • Marcinowski L.
      • Tanguy M.
      • Krmpotic A.
      • Rädle B.
      • Lisnić V.J.
      • Tuddenham L.
      • et al.
      Degradation of cellular mir-27 by a novel, highly abundant viral transcript is important for efficient virus replication in vivo.
      ,
      • Lu S.
      • Cullen B.R.
      Adenovirus VA1 noncoding RNA can inhibit small interfering RNA and MicroRNA biogenesis.
      ] (Table 1). Taken together, these data demonstrate that viruses have evolved several strategies to modulate cellular miRNAs. While this may allow the virus to escape antiviral immunity and establish persistent infection, virus-induced changes in the host miRNome may ultimately also contribute to cellular transformation and oncogenesis.
      Finally, viruses can also encode miRNAs, which can target either host or viral RNAs [
      • Roberts A.P.E.
      • Lewis A.P.
      • Jopling C.L.
      The role of microRNAs in viral infection.
      ,
      • Cullen B.R.
      Viruses and microRNAs: RISCy interactions with serious consequences.
      ,
      • Kincaid R.P.
      • Burke J.M.
      • Sullivan C.S.
      RNA virus microRNA that mimics a B-cell oncomiR.
      ] (Table 1). Virus-encoded miRNAs can either be specific to a virus or be analogues of host miRNAs, and they usually promote viral infection by prolonging the longevity of infected cells, inhibiting immune responses, and/or regulating host or viral genes to limit the lytic cycle (reviewed in [
      • Kincaid R.P.
      • Sullivan C.S.
      Virus-encoded microRNAs: an overview and a look to the future.
      ]). Interestingly, although a computational approach indicated that HBV putatively encodes a candidate pre-miRNA that might yield a mature miRNA with putative binding sites within the HBV mRNA [
      • Jin W.-B.
      • Wu F.-L.
      • Kong D.
      • Guo A.-G.
      HBV-encoded microRNA candidate and its target.
      ], to date there is no experimental evidence for any HBV- or HCV-encoded miRNA.

      miR-122 and HCV infection: Host-dependency factor and antiviral target

      HCV is a single-stranded RNA virus of positive polarity [

      Lindenbach BD, Thiel HJ, Rice CM. Flaviviridae: the viruses and their replication. Fields Virol., vol. 1. 5th edition, 2007; 1101–52.

      ]. The role for miR-122 in HCV infection was first demonstrated by sequestration of endogenous miR-122, which led to a substantial reduction in HCV RNA abundance [
      • Jopling C.L.
      • Yi M.
      • Lancaster A.M.
      • Lemon S.M.
      • Sarnow P.
      Modulation of hepatitis C virus RNA abundance by a liver-specific MicroRNA.
      ]. Unlike most miRNAs that repress their targets through binding the 3′ UTR of mRNAs, miR-122 directly pairs with two adjacent sites in the 5′ UTR of the viral RNA, thus enhancing viral replication [
      • Pedersen I.M.
      • Cheng G.
      • Wieland S.
      • Volinia S.
      • Croce C.M.
      • Chisari F.V.
      • et al.
      Interferon modulation of cellular microRNAs as an antiviral mechanism.
      ,
      • Jopling C.L.
      • Schütz S.
      • Sarnow P.
      Position-dependent function for a tandem microRNA miR-122-binding site located in the hepatitis C virus RNA genome.
      ,
      • Jangra R.K.
      • Yi M.
      • Lemon S.M.
      Regulation of hepatitis C virus translation and infectious virus production by the microRNA miR-122.
      ,
      • Machlin E.S.
      • Sarnow P.
      • Sagan S.M.
      Masking the 5′ terminal nucleotides of the hepatitis C virus genome by an unconventional microRNA-target RNA complex.
      ,
      • Shimakami T.
      • Yamane D.
      • Welsch C.
      • Hensley L.
      • Jangra R.K.
      • Lemon S.M.
      Base pairing between hepatitis C virus RNA and microRNA 122 3′ of its seed sequence is essential for genome stabilization and production of infectious virus.
      ] (Fig. 1). These target sites are located upstream of the HCV internal ribosome entry site (IRES), and are conserved across HCV genotypes. Recent studies indicated that miR-122 positively acts on the HCV life cycle by enhancing viral translation and genome stabilization. Indeed, it has been shown that miR-122 binding to the 5′ UTR of the HCV genome enhances the association of ribosomes with the viral RNA [
      • Jangra R.K.
      • Yi M.
      • Lemon S.M.
      Regulation of hepatitis C virus translation and infectious virus production by the microRNA miR-122.
      ,
      • Henke J.I.
      • Goergen D.
      • Zheng J.
      • Song Y.
      • Schüttler C.G.
      • Fehr C.
      • et al.
      MicroRNA-122 stimulates translation of hepatitis C virus RNA.
      ,
      • Goergen D.
      • Niepmann M.
      Stimulation of Hepatitis C Virus RNA translation by microRNA-122 occurs under different conditions in vivo and in vitro.
      ]. Furthermore, the association of miR-122 and the HCV genome together with Ago2 within the RISC complex also stabilizes viral RNA by protecting it from degradation by exonucleases [
      • Shimakami T.
      • Yamane D.
      • Welsch C.
      • Hensley L.
      • Jangra R.K.
      • Lemon S.M.
      Base pairing between hepatitis C virus RNA and microRNA 122 3′ of its seed sequence is essential for genome stabilization and production of infectious virus.
      ,
      • Wilson J.A.
      • Zhang C.
      • Huys A.
      • Richardson C.D.
      Human Ago2 is required for efficient microRNA 122 regulation of hepatitis C virus RNA accumulation and translation.
      ,
      • Conrad K.D.
      • Giering F.
      • Erfurth C.
      • Neumann A.
      • Fehr C.
      • Meister G.
      • et al.
      MicroRNA-122 dependent binding of Ago2 protein to hepatitis C virus RNA is associated with enhanced RNA stability and translation stimulation.
      ,
      • Li Y.
      • Masaki T.
      • Yamane D.
      • McGivern D.R.
      • Lemon S.M.
      Competing and noncompeting activities of miR-122 and the 5′ exonuclease Xrn1 in regulation of hepatitis C virus replication.
      ]. The importance of miR-122 in HCV infection is also underscored by a number of studies, which indicated the involvement of this miRNA in allowing HCV replication in non-HCV permissive cell lines. Indeed, hepatoma cell lines as well as non-liver derived HEK-293T or HeLa cells, which do not express significant amounts of miR-122 and are unable to sustain HCV replication, were rendered permissive to HCV replication upon ectopic miR-122 expression [
      • Narbus C.M.
      • Israelow B.
      • Sourisseau M.
      • Michta M.L.
      • Hopcraft S.E.
      • Zeiner G.M.
      • et al.
      HepG2 cells expressing microRNA miR-122 support the entire hepatitis C virus life cycle.
      ,
      • Kambara H.
      • Fukuhara T.
      • Shiokawa M.
      • Ono C.
      • Ohara Y.
      • Kamitani W.
      • et al.
      Establishment of a novel permissive cell line for the propagation of hepatitis C virus by expression of microRNA miR122.
      ,
      • Chang J.
      • Guo J.-T.
      • Jiang D.
      • Guo H.
      • Taylor J.M.
      • Block T.M.
      Liver-specific microRNA miR-122 enhances the replication of hepatitis C virus in nonhepatic cells.
      ,
      • Costa D.D.
      • Turek M.
      • Felmlee D.J.
      • Girardi E.
      • Pfeffer S.
      • Long G.
      • et al.
      Reconstitution of the entire hepatitis C virus life cycle in nonhepatic cells.
      ,
      • Fukuhara T.
      • Kambara H.
      • Shiokawa M.
      • Ono C.
      • Katoh H.
      • Morita E.
      • et al.
      Expression of microRNA miR-122 facilitates an efficient replication in nonhepatic cells upon infection with hepatitis C virus.
      ,
      • Hueging K.
      • Doepke M.
      • Vieyres G.
      • Bankwitz D.
      • Frentzen A.
      • Doerrbecker J.
      • et al.
      Apolipoprotein E codetermines tissue tropism of hepatitis C virus and is crucial for viral cell-to-cell transmission by contributing to a postenvelopment step of assembly.
      ]. Interestingly, in addition to the direct effect, mediated by miR-122 targeting of the HCV RNA, an indirect effect has been reported that involves the downregulation of HMOX1, the latter having been shown to inhibit HCV replication [
      • Shan Y.
      • Zheng J.
      • Lambrecht R.W.
      • Bonkovsky H.L.
      Reciprocal effects of micro-RNA-122 on expression of heme oxygenase-1 and hepatitis C virus genes in human hepatocytes.
      ] (Fig. 1). miR-122 was also discovered to prompt alcohol-induced HCV RNA replication [
      • Hou W.
      • Bukong T.N.
      • Kodys K.
      • Szabo G.
      Alcohol facilitates HCV RNA replication via up-regulation of miR-122 expression and inhibition of cyclin G1 in human hepatoma cells.
      ,
      • Bukong T.N.
      • Hou W.
      • Kodys K.
      • Szabo G.
      Ethanol facilitates hepatitis C virus replication via up-regulation of GW182 and heat shock protein 90 in human hepatoma cells.
      ]. In particular, acute alcohol exposure in HCC cell lines was shown to enhance HCV replication by upregulating miR-122 expression while downregulating the miR-122 target cyclin G1 [
      • Hou W.
      • Bukong T.N.
      • Kodys K.
      • Szabo G.
      Alcohol facilitates HCV RNA replication via up-regulation of miR-122 expression and inhibition of cyclin G1 in human hepatoma cells.
      ]. Taken together, these data indicate that miR-122 represents an essential hepatocyte-specific host factor for HCV infection.
      Counter-intuitively, the beneficial role of miR-122 for the virus in vitro does not translate into a positive correlation between its expression and HCV load in patients. Particularly non-responders to IFN-based therapy have lower miR-122 pre-treatment levels [
      • Sarasin-Filipowicz M.
      • Krol J.
      • Markiewicz I.
      • Heim M.H.
      • Filipowicz W.
      Decreased levels of microRNA miR-122 in individuals with hepatitis C responding poorly to interferon therapy.
      ,
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      • Liu C.-J.
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      • et al.
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      ,
      • Kamo Y.
      • Ichikawa T.
      • Miyaaki H.
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      • Yamaguchi T.
      • Shibata H.
      • et al.
      Significance of miRNA-122 in chronic hepatitis C patients with serotype 1 on interferon therapy.
      ,
      • Estrabaud E.
      • Lapalus M.
      • Broët P.
      • Appourchaux K.
      • De Muynck S.
      • Lada O.
      • et al.
      Reduction of microRNA 122 expression in IFNL3 CT/TT carriers and during progression of fibrosis in patients with chronic hepatitis C.
      ], suggesting that pre-treatment miR-122 levels could be used as a biomarker to predict the therapeutic outcome. While it has been shown that IFN-based therapy does not appear to decrease intrahepatic miR-122 in patients [
      • Sarasin-Filipowicz M.
      • Krol J.
      • Markiewicz I.
      • Heim M.H.
      • Filipowicz W.
      Decreased levels of microRNA miR-122 in individuals with hepatitis C responding poorly to interferon therapy.
      ], another study reported that reduced serum miR-122 correlates with therapeutic success, probably by reflecting reduced liver damage [
      • Köberle V.
      • Waidmann O.
      • Kronenberger B.
      • Andrei A.
      • Susser S.
      • Füller C.
      • et al.
      Serum microRNA-122 kinetics in patients with chronic hepatitis C virus infection during antiviral therapy.
      ].
      Given its essential role in the HCV life cycle and its liver-enriched expression, miR-122 represents a target for antiviral therapy (Fig. 2). The first animal studies using antisense miR-122 oligonucleotides of different chemistry were encouraging as they indicated that targeting miR-122 did not result in liver toxicity in mice and African green monkeys [
      • Esau C.
      • Davis S.
      • Murray S.F.
      • Yu X.X.
      • Pandey S.K.
      • Pear M.
      • et al.
      MiR-122 regulation of lipid metabolism revealed by in vivo antisense targeting.
      ,
      • Hildebrandt-Eriksen E.S.
      • Aarup V.
      • Persson R.
      • Hansen H.F.
      • Munk M.E.
      • Ørum H.
      A locked nucleic acid oligonucleotide targeting microRNA 122 is well-tolerated in cynomolgus monkeys.
      ]. In addition, the treatment decreased their plasma cholesterol levels and this effect was sustained for several weeks but reversible following withdrawal of the inhibitor [
      • Esau C.
      • Davis S.
      • Murray S.F.
      • Yu X.X.
      • Pandey S.K.
      • Pear M.
      • et al.
      MiR-122 regulation of lipid metabolism revealed by in vivo antisense targeting.
      ,
      • Hildebrandt-Eriksen E.S.
      • Aarup V.
      • Persson R.
      • Hansen H.F.
      • Munk M.E.
      • Ørum H.
      A locked nucleic acid oligonucleotide targeting microRNA 122 is well-tolerated in cynomolgus monkeys.
      ], suggesting that targeting miR-122 might also be a potential therapeutic strategy for hypercholesterolemia (Fig. 2). A study using chronically HCV-infected chimpanzees then provided the first proof-of-concept for the potential of the miR-122 inhibitor SPC3649, now known as miravirsen, as an efficient antiviral. Indeed, the inhibitor reduced HCV RNA levels in the majority of treated animals and its effect was gradually lost once the inhibitor was withdrawn [
      • Lanford R.E.
      • Hildebrandt-Eriksen E.S.
      • Petri A.
      • Persson R.
      • Lindow M.
      • Munk M.E.
      • et al.
      Therapeutic silencing of microRNA-122 in primates with chronic hepatitis C virus infection.
      ], confirming a sustained but reversible inhibition of miR-122 in vivo. The potential of this inhibitor has recently been confirmed in a phase 2a clinical trial [
      • Janssen H.L.A.
      • Reesink H.W.
      • Lawitz E.J.
      • Zeuzem S.
      • Rodriguez-Torres M.
      • Patel K.
      • et al.
      Treatment of HCV infection by targeting microRNA.
      ]. Administration of this inhibitor for 5 weeks resulted in a dose-dependent and sustained reduction of HCV RNA levels up to 3 logs for the highest dose of 7 mg/kg with several patients transiently achieving undetectable HCV RNA levels. However, viral RNA levels rebounded in patients that did not start an IFN-based therapy at the end of the trial. No dose-limiting adverse events were observed but patients exhibited a sustained and reversible decrease in serum cholesterol levels. Nevertheless, the miR-122 inhibitor half-life and long-term implications of miR-122 inhibition in vivo may merit further studies. Very importantly, no adaptive mutations were detected with in the HCV miR-122 binding regions, indicating that miR-122 inhibitors have a high barrier to resistance [
      • Janssen H.L.A.
      • Reesink H.W.
      • Lawitz E.J.
      • Zeuzem S.
      • Rodriguez-Torres M.
      • Patel K.
      • et al.
      Treatment of HCV infection by targeting microRNA.
      ]. Despite these interesting results, given the recent tremendous advances in the treatment of chronic HCV infection with the approval of orally administered DAAs with pan-genotypic activity and high barrier to resistance (reviewed in [
      • Chung R.T.
      • Baumert T.F.
      Curing chronic hepatitis C – The arc of a medical triumph.
      ]) that enable very high rates of sustained virological response (SVR), it is likely that miR-122 inhibitors that require parenteral administration will not play a major role in the future antiviral therapy against HCV. However, since patients who cleared HCV remain at risk for HCC (reviewed in [
      • Chung R.T.
      • Baumert T.F.
      Curing chronic hepatitis C – The arc of a medical triumph.
      ]), a better understanding of the miRNA networks, modulated in the course of HCV infection and involved in development of HCC, will allow to ultimately uncover pathways that may represent potential therapeutic targets to prevent/treat HCC.

      miR-122 and HBV infection: A viral restriction factor?

      HBV is a DNA virus with a relaxed circular partially double-stranded genome (rcDNA) that is converted into a covalently closed circular DNA (cccDNA) in the host cell nucleus, following infection of human hepatocytes. The cccDNA serves as a template for the transcription of four viral RNAs that represent templates for the translation of the HBV proteins and for viral replication, involving reverse transcription [
      • Trépo C.
      • Chan H.L.Y.
      • Lok A.
      Hepatitis B virus infection.
      ]. In contrast to its role as a host-dependency factor for HCV, miR-122 appears to restrict HBV replication. Indeed, it has been shown that miR-122 directly targets a conserved region of the HBV pregenomic RNA that functions as a bicistronic mRNA, encoding the HBV polymerase and core protein [
      • Chen Y.
      • Shen A.
      • Rider P.J.
      • Yu Y.
      • Wu K.
      • Mu Y.
      • et al.
      A liver-specific microRNA binds to a highly conserved RNA sequence of hepatitis B virus and negatively regulates viral gene expression and replication.
      ] (Fig. 1). However, the exact mechanisms by which miR-122 binding to HBV RNA results in the inhibition of HBV protein expression, transcription and replication remain to be determined. Furthermore, miR-122 has been shown to indirectly interfere with HBV replication by decreasing expression of cyclin G1, which results in p53-mediated inhibition of HBV transcription [
      • Wang S.
      • Qiu L.
      • Yan X.
      • Jin W.
      • Wang Y.
      • Chen L.
      • et al.
      Loss of microRNA 122 expression in patients with hepatitis B enhances hepatitis B virus replication through cyclin G(1)-modulated P53 activity.
      ]. However, in human hepatoma cell lines, miR-122 was also observed to indirectly enhance HBV replication by repressing HMOX1, which in turn interfered with HBV replication by reducing the stability of the HBV core protein [
      • Qiu L.
      • Fan H.
      • Jin W.
      • Zhao B.
      • Wang Y.
      • Ju Y.
      • et al.
      MiR-122-induced down-regulation of HO-1 negatively affects miR-122-mediated suppression of HBV.
      ] (Fig. 1). In contrast to HCV, HBV infection downregulates miR-122 expression and viral load was shown to inversely correlate with miR-122 expression in HBV-infected patients [
      • Chen Y.
      • Shen A.
      • Rider P.J.
      • Yu Y.
      • Wu K.
      • Mu Y.
      • et al.
      A liver-specific microRNA binds to a highly conserved RNA sequence of hepatitis B virus and negatively regulates viral gene expression and replication.
      ,
      • Wang S.
      • Qiu L.
      • Yan X.
      • Jin W.
      • Wang Y.
      • Chen L.
      • et al.
      Loss of microRNA 122 expression in patients with hepatitis B enhances hepatitis B virus replication through cyclin G(1)-modulated P53 activity.
      ,
      • Li C.
      • Wang Y.
      • Wang S.
      • Wu B.
      • Hao J.
      • Fan H.
      • et al.
      Hepatitis B virus mRNA-mediated miR-122 inhibition upregulates PTTG1-binding protein, which promotes hepatocellular carcinoma tumor growth and cell invasion.
      ]. The exact underlying mechanisms are not fully understood, but one possibility could be that all HBV mRNAs contain a miR-122 binding site and could act as sponges to sequester miR-122 [
      • Li C.
      • Wang Y.
      • Wang S.
      • Wu B.
      • Hao J.
      • Fan H.
      • et al.
      Hepatitis B virus mRNA-mediated miR-122 inhibition upregulates PTTG1-binding protein, which promotes hepatocellular carcinoma tumor growth and cell invasion.
      ]. Moreover, a recent study demonstrated that the HBx protein could bind PPARγ, thereby leading to inhibition of miR-122 transcription [
      • Song K.
      • Han C.
      • Zhang J.
      • Lu D.
      • Dash S.
      • Feitelson M.
      • et al.
      Epigenetic regulation of MicroRNA-122 by peroxisome proliferator activated receptor-gamma and hepatitis b virus X protein in hepatocellular carcinoma cells.
      ]. HBx can also decrease the stability of miR-122 by downregulating germline development 2 (Gld2) that is involved in miR-122 adenylation [
      • Peng F.
      • Xiao X.
      • Jiang Y.
      • Luo K.
      • Tian Y.
      • Peng M.
      • et al.
      HBx down-regulated Gld2 plays a critical role in HBV-related dysregulation of miR-122.
      ].
      Given the important role of miR-122 in liver physiology, this virus-induced suppression of miR-122 may alter liver function and contribute to the development of liver disease including HCC. Indeed, it has been reported that the HBV-mediated downregulation of miR-122 increases the expression of the tumour promoter N-myc downstream regulated gene 3 (NDRG3) [
      • Fan C.-G.
      • Wang C.-M.
      • Tian C.
      • Wang Y.
      • Li L.
      • Sun W.-S.
      • et al.
      MiR-122 inhibits viral replication and cell proliferation in hepatitis B virus-related hepatocellular carcinoma and targets NDRG3.
      ]. Furthermore, this increases expression of the miR-122 target cyclin G1 (CCNG1) that results in enhanced Akt activation leading to epithelial-mesenchymal transition [
      • Wen W.
      • Ding J.
      • Sun W.
      • Fu J.
      • Chen Y.
      • Wu K.
      • et al.
      Cyclin G1–mediated epithelial-mesenchymal transition via phosphoinositide 3-kinase/Akt signaling facilitates liver cancer progression.
      ]. Moreover, HBV-induced inhibition of miR-122 also results in an increase in pituitary tumour-transforming gene 1-binding factor (PTTG1) that promotes tumour growth and cell invasion [
      • Li C.
      • Wang Y.
      • Wang S.
      • Wu B.
      • Hao J.
      • Fan H.
      • et al.
      Hepatitis B virus mRNA-mediated miR-122 inhibition upregulates PTTG1-binding protein, which promotes hepatocellular carcinoma tumor growth and cell invasion.
      ]. Taken together, these HBV-induced changes in regulatory networks may contribute to the development of HCC (Fig. 1). Given that restoration of miR-122 has been shown to reverse the tumourigenic properties of hepatoma cells and to prevent HCC development in vivo [
      • Hsu S.-H.
      • Wang B.
      • Kota J.
      • Yu J.
      • Costinean S.
      • Kutay H.
      • et al.
      Essential metabolic, anti-inflammatory, and anti-tumorigenic functions of miR-122 in liver.
      ,
      • Tsai W.-C.
      • Hsu S.-D.
      • Hsu C.-S.
      • Lai T.-C.
      • Chen S.-J.
      • Shen R.
      • et al.
      MicroRNA-122 plays a critical role in liver homeostasis and hepatocarcinogenesis.
      ,
      • Coulouarn C.
      • Corlu A.
      • Glaise D.
      • Guénon I.
      • Thorgeirsson S.S.
      • Clément B.
      Hepatocyte-stellate cell cross-talk in the liver engenders a permissive inflammatory microenvironment that drives progression in hepatocellular carcinoma.
      ,
      • Bai S.
      • Nasser M.W.
      • Wang B.
      • Hsu S.-H.
      • Datta J.
      • Kutay H.
      • et al.
      MicroRNA-122 inhibits tumorigenic properties of hepatocellular carcinoma cells and sensitizes these cells to sorafenib.
      ,
      • Fornari F.
      • Gramantieri L.
      • Giovannini C.
      • Veronese A.
      • Ferracin M.
      • Sabbioni S.
      • et al.
      MiR-122/cyclin G1 interaction modulates p53 activity and affects doxorubicin sensitivity of human hepatocarcinoma cells.
      ,
      • Xu Y.
      • Xia F.
      • Ma L.
      • Shan J.
      • Shen J.
      • Yang Z.
      • et al.
      MicroRNA-122 sensitizes HCC cancer cells to adriamycin and vincristine through modulating expression of MDR and inducing cell cycle arrest.
      ,
      • Hsu S.-H.
      • Yu B.
      • Wang X.
      • Lu Y.
      • Schmidt C.R.
      • Lee R.J.
      • et al.
      Cationic lipid nanoparticles for therapeutic delivery of siRNA and miRNA to murine liver tumor.
      ], potential future therapeutic strategies, aiming at restoring miR-122 to prevent/treat HCC in patients with reduced/absent miR-122 levels might be an interesting strategy for patients with HBV-induced HCC.

      Conclusions and perspectives

      Given its central role in liver biology and disease, miR-122 represents an interesting therapeutic target for the treatment of liver disease including viral hepatitis, fibrosis, steatosis and HCC. Proof-of-concept studies have elegantly demonstrated that a miR-122 inhibitor efficiently reduces viral load in chronically infected HCV patients without detectable resistance [
      • Janssen H.L.A.
      • Reesink H.W.
      • Lawitz E.J.
      • Zeuzem S.
      • Rodriguez-Torres M.
      • Patel K.
      • et al.
      Treatment of HCV infection by targeting microRNA.
      ] (Fig. 2). However, given the very high cure rates of orally administrated DAAs with a high genetic barrier for resistance (reviewed in [
      • Chung R.T.
      • Baumert T.F.
      Curing chronic hepatitis C – The arc of a medical triumph.
      ]), the need for parenteral administration of miRNA-122 inhibitors [
      • Janssen H.L.A.
      • Reesink H.W.
      • Lawitz E.J.
      • Zeuzem S.
      • Rodriguez-Torres M.
      • Patel K.
      • et al.
      Treatment of HCV infection by targeting microRNA.
      ], and a potential HCC/liver disease-promoting effect of miRNA depletion, the role of miR-122 inhibitors in the future treatment approaches for HCV infection remains to be determined. The exploration of miR-122 as a therapeutic target for HBV infection is ongoing. While experimental studies suggest that miR-122 plays a role in the HBV life cycle as a potential restriction factor, further studies are needed to assess whether targeting miR-122 would result in cccDNA eradication and viral cure – the ultimate goal for novel HBV therapeutic approaches. Given the limited or absent strategies to impair progression of liver disease and to prevent and treat HCC (reviewed in [
      • Forner A.
      • Gilabert M.
      • Bruix J.
      • Raoul J.-L.
      Treatment of intermediate-stage hepatocellular carcinoma.
      ]) and the association between loss of miR-122 and liver inflammation, fibrosis, steatosis and HCC, miR-122 mimics may provide a novel strategy to slow down liver disease progression and to prevent and treat HCC. Current and future randomized clinical trials with miRNA-based molecules will shed light on the perspective of this approach for advanced liver disease and HCC. Finally, given the major involvement of miR-122 in liver homeostasis, cholesterol biosynthesis and fatty acid metabolism, additional preclinical studies will be required to determine the optimal level of miRNA mimics in therapy and to assess the potential risks associated with miR-122 overexpression or depletion.

      Financial support

      The authors’ work was supported by Inserm , University of Strasbourg , the European Union (ERC-2008-AdG-233130-HEPCENT, ERC-StG-260767-ncRNAVIR, INTERREG-IV-Rhin Supérieur-FEDER-Hepato-Regio-Net 2012, EU FP7 HepaMab), ANRS ( 2012/239 , 2013/108 ), the Direction Générale de l’Offre de Soins ( A12027MS ) the Institut Hospitalo-Universitaire (IHU) Mix-Surg and ARC (TheraHCC, IHU201301187). This work has been published under the framework of the LABEX ANR-10-LABX-0028_HEPSYS and ANR-10-LABX-0036_NETRNA and benefits from a funding from the state, managed by the French National Research Agency as part of the investments for the future program.

      Conflict of interest

      The authors who have taken part in this study declared that they do not have anything to disclose regarding funding or conflict of interest with respect to this manuscript.

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