title: A serum microRNA signature is associated with the immune control of chronic hepatitis B virus infection.
abstract: OBJECTIVE
  The virus/host interplay mediates liver pathology in chronic HBV infection. MiRNAs play a pivotal role in virus/host interactions and are detected in both serum and HBsAg-particles, but studies of their dynamics during chronic infection and antiviral therapy are missing. We studied serum miRNAs during different phases of chronic HBV infection and antiviral treatment.
  
  
  METHODS
  MiRNAs were profiled by miRCURY-LNA-Universal-RT-miRNA-PCR (Exiqon-A/S) and qPCR-panels-I/II-739-miRNA-assays and single-RT-q-PCRs. Two cohorts of well-characterized HBsAg-carriers were studied (median follow-up 34-52 months): a) training-panel (141 sera) and HBsAg-particles (32 samples) from 61 HBsAg-carriers and b) validation-panel (136 sera) from 84 carriers.
  
  
  RESULTS
  Thirty-one miRNAs were differentially expressed in inactive-carriers (IC) and chronic-hepatitis-B (CHB) with the largest difference for miR-122-5p, miR-99a-5p and miR-192-5p (liver-specific-miRNAs), over-expressed in both sera and HBsAg-particles of CHB (ANOVA/U-test p-values: <0.000001/0.000001; <0.000001/0.000003; <0.000001/0.000005, respectively) and significantly down-regulated during- and after-treatment in sustained-virological-responders (SVR). MiRNA-profiles of IC and SVR clustered in the heatmap. Liver-miRNAs were combined with miR-335, miR-126 and miR-320a (internal controls) to build a MiR-B-Index with 100% sensitivity, 83.3% and 92.5% specificity (-1.7 cut-off) in both training and validation cohorts to identify IC. MiR-B-Index (-5.72, -20.43/14.38) correlated with ALT (49, 10/2056 U/l, ρ = -0.497, p<0.001), HBV-DNA (4.58, undetectable/>8.3 Log10 IU/mL, ρ = -0.732, p<0.001) and HBsAg (3.40, 0.11/5.49 Log10 IU/mL, ρ = -0.883, p<0.001). At multivariate analysis HBV-DNA (p = 0.002), HBsAg (p<0.001) and infection-phase (p<0.001), but not ALT (p = 0.360) correlated with MiR-B-Index. In SVR to Peg-IFN/NUCs MiR-B-Index improved during-therapy and post-treatment reaching IC-like values (5.32, -1.65/10.91 vs 6.68, 0.54/9.53, p = 0.324) beckoning sustained HBV-immune-control earlier than HBsAg-decline.
  
  
  CONCLUSIONS
  Serum miRNA profile change dynamically during the different phases of chronic HBV infection. We identified a miRNA signature associated with both natural-occurring and therapy-induced immune control of HBV infection. The MiR-B-Index might be a useful biomarker for the early identification of the sustained switch from CHB to inactive HBV-infection in patients treated with antivirals.
authors.affiliation: Laboratory of Molecular Genetics and Pathology of Hepatitis Viruses, Hepatology Unit, Reference Center of the Tuscany Region for Chronic Liver Disease and Cancer, University Hospital of Pisa, Pisa, Italy.
authors.forename: Maurizia Rossana
authors.initials: MR
authors.lastname: Brunetto
authors.affiliation: Laboratory of Molecular Genetics and Pathology of Hepatitis Viruses, Hepatology Unit, Reference Center of the Tuscany Region for Chronic Liver Disease and Cancer, University Hospital of Pisa, Pisa, Italy.
authors.forename: Daniela
authors.initials: D
authors.lastname: Cavallone
authors.affiliation: Laboratory of Molecular Genetics and Pathology of Hepatitis Viruses, Hepatology Unit, Reference Center of the Tuscany Region for Chronic Liver Disease and Cancer, University Hospital of Pisa, Pisa, Italy.
authors.forename: Filippo
authors.initials: F
authors.lastname: Oliveri
authors.affiliation: Laboratory of Molecular Genetics and Pathology of Hepatitis Viruses, Hepatology Unit, Reference Center of the Tuscany Region for Chronic Liver Disease and Cancer, University Hospital of Pisa, Pisa, Italy.
authors.forename: Francesco
authors.initials: F
authors.lastname: Moriconi
authors.affiliation: Laboratory of Molecular Genetics and Pathology of Hepatitis Viruses, Hepatology Unit, Reference Center of the Tuscany Region for Chronic Liver Disease and Cancer, University Hospital of Pisa, Pisa, Italy.
authors.forename: Piero
authors.initials: P
authors.lastname: Colombatto
authors.affiliation: Laboratory of Molecular Genetics and Pathology of Hepatitis Viruses, Hepatology Unit, Reference Center of the Tuscany Region for Chronic Liver Disease and Cancer, University Hospital of Pisa, Pisa, Italy.
authors.forename: Barbara
authors.initials: B
authors.lastname: Coco
authors.affiliation: Laboratory of Molecular Genetics and Pathology of Hepatitis Viruses, Hepatology Unit, Reference Center of the Tuscany Region for Chronic Liver Disease and Cancer, University Hospital of Pisa, Pisa, Italy.
authors.forename: Pietro
authors.initials: P
authors.lastname: Ciccorossi
authors.affiliation: Laboratory of Molecular Genetics and Pathology of Hepatitis Viruses, Hepatology Unit, Reference Center of the Tuscany Region for Chronic Liver Disease and Cancer, University Hospital of Pisa, Pisa, Italy.
authors.forename: Carlotta
authors.initials: C
authors.lastname: Rastelli
authors.affiliation: Laboratory of Molecular Genetics and Pathology of Hepatitis Viruses, Hepatology Unit, Reference Center of the Tuscany Region for Chronic Liver Disease and Cancer, University Hospital of Pisa, Pisa, Italy.
authors.forename: Veronica
authors.initials: V
authors.lastname: Romagnoli
authors.affiliation: Laboratory of Molecular Genetics and Pathology of Hepatitis Viruses, Hepatology Unit, Reference Center of the Tuscany Region for Chronic Liver Disease and Cancer, University Hospital of Pisa, Pisa, Italy.
authors.forename: Beatrice
authors.initials: B
authors.lastname: Cherubini
authors.affiliation: Division of Dx and Services, Exiqon A/S, Copenhagen, Denmark.
authors.forename: Maria Wrang
authors.initials: MW
authors.lastname: Teilum
authors.affiliation: Division of Dx and Services, Exiqon A/S, Copenhagen, Denmark.
authors.forename: Thorarinn
authors.initials: T
authors.lastname: Blondal
authors.affiliation: Digestive and Liver Disease, General Medicine II Unit, University Hospital of Pisa, Pisa, Italy.
authors.forename: Ferruccio
authors.initials: F
authors.lastname: Bonino
mesh_terms: D000328:Adult; D000368:Aged; D000998:Antiviral Agents / Q000627:therapeutic use; D016000:Cluster Analysis; D015331:Cohort Studies; D019295:Computational Biology; D005260:Female; D005500:Follow-Up Studies; D020869:Gene Expression Profiling; D006515:Hepatitis B virus / Q000276:immunology*; D019694:Hepatitis B, Chronic / Q000188:drug therapy / Q000235:genetics* / Q000276:immunology* / Q000821:virology; D054884:Host-Pathogen Interactions / Q000235:genetics / Q000276:immunology; D006801:Humans; D008297:Male; D035683:MicroRNAs / Q000097:blood / Q000235:genetics*; D008875:Middle Aged; D015203:Reproducibility of Results; D019562:Viral Load; D055815:Young Adult
chemical_list: D000998:Antiviral Agents; D035683:MicroRNAs