Ángel Hernández-Bartolomé

Genetic Variants of Interferon-Stimulated Genes and IL-28B as Host Prognostic Factors of Response to Combination Treatment for Chronic Hepatitis C

Chronic hepatitis C (CHC) is a worldwide health problem that is highly related to liver fibrosis, cirrhosis, and hepatocellular carcinoma. The achievement of response to the current standard of care—pegylated interferon plus ribavirin—has recently been described to be associated with single‐nucleotide polymorphisms (SNPs) near the IL‐28B gene. Additionally, baseline expression levels of genes involved in interferon (IFN)‐stimulated genes (ISGs) have been found to be related to treatment outcome. In the present study, 285 patients were genotyped for 63 SNPs within genes of the IFN signaling pathway (IPGs) and ISGs. Two ISG polymorphisms—OASL rs12819210 (odds ratio (OR) = 2.1, P = 0.03) and IFIT1 rs304478 (OR = 2.5, P = 0.01)—were found to be independent predictive factors of sustained virological response (SVR) after adjusting for other clinical covariates. Furthermore, the predictive value of IL‐28B SNP was notably improved by simultaneous genotyping of rs12819210 and rs304478, particularly in patients with the worst prognosis (viral genotype 1, area under the curve (AUC) = 0.74). In conclusion, ISG SNPs could constitute a valuable tool for individualizing CHC therapy.

Peripheral blood monocyte subsets predict antiviral response in chronic hepatitis C

Aliment Pharmacol Ther 2011; 34: 960–971

Angiopoietin-2 Serum Levels Improve Noninvasive Fibrosis Staging in Chronic Hepatitis C: A Fibrogenic-Angiogenic Link

Aims Accurate liver fibrosis staging is crucial for the management of chronic hepatitis C (CHC). The invasiveness and cost burden of liver biopsy have driven the search for new noninvasive biomarkers of fibrosis. Based on the link between serum angiopoietin-1 and 2 levels and CHC progression, we aimed to determine the value of these angiogenic factors as noninvasive biomarkers of liver fibrosis. Methods Serum levels of angiopoietin-1 and -2 were measured by ELISA in 108 CHC patients who underwent pretreatment liver biopsy. The correlation between angiopoietins and clinical and demographic variables with liver fibrosis was analyzed by univariate regression. Significant factors were then subjected to multivariate analysis, from which we constructed a novel noninvasive liver fibrosis index (AngioScore), whose performance was validated in an independent series of 71 CHC patients. The accuracy of this model was compared with other documented fibrosis algorithms by De Long test. Results Angiopoietins correlated significantly with hepatic fibrosis; however, only angiopoietin-2 was retained in the final model, which also included age, platelets, AST, INR, and GGT. The model was validated and behaved considerably better than other fibrosis indices in discriminating all, significant, moderate and severe liver fibrosis (0.886, 0.920, 0.923). Using clinically relevant cutoffs, we classified CHC patients by discarding significant fibrosis and diagnosing moderate and severe fibrosis with greater accuracy, sensitivity, and specificity. Conclusions Our novel noninvasive liver fibrosis model, based on serum angiopoietin-2 levels, outperforms other indices and should help substantially in managing CHC and monitoring long-term follow-up prognosis.

Polymorphisms in histone deacetylases improve the predictive value of IL-28B for chronic hepatitis C therapy

Histone deacetylases (HDACs) influence many cellular processes, including the modulation of signal transducer and activator of transcription activity (STAT) in response to interferon (IFN). To identify genetic markers that help optimize the IL-28B prediction of chronic hepatitis C (CHC) sustained virological response (SVR), we evaluated 27 single-nucleotide polymorphisms (SNPs) in HDAC1–11. Three SNPs, rs3778216, rs976552 and rs368328 in HDAC2, HDAC3 and HDAC5, respectively, were independently associated with SVR (P<0.05). The addition of these three HDAC’s SNPs to the IL-28B predictive model (area under the curve (AUC)=0.630) rendered an important improvement of AUC-receiver operating characteristic value (AUC=0.747, P=0.021). Chi-squared Automatic Interaction Detector (CHAID) analysis denoted the significance of the rs3778216 C/C genotype in identifying a group of good responders despite carrying IL-28B T allele (79.2% of SVR), whereas HDAC5 G allele characterized a subgroup with poor response rate (25.5%). However, HDAC3 rs976552 did not display a relevant role for the hierarchical classification of patients. Variables related to SVR in hepatitis C virus genotype 1 (HCV-1) cohort were the same of those obtained for the overall population. Interestingly, in non-HCV-1 patients (n=56) the HDAC2 C/C genotype was the unique predictive variable related to SVR (AUC=0.733, P<0.007). Thus, these preliminary results suggest the potential usefulness of combined IL-28B and HDAC genotyping for the CHC patients’ classification by likelihood of an SVR.

Preliminary evidence of sustained expression of angiopoietin-2 during monocyte differentiation in chronic hepatitis C.

Monocytes are essential precursors of antigen‐presenting cells, such as macrophages and dendritic cells, and contribute to the pathogenesis of chronic inflammatory diseases and cancer.

Interferon-related genetic markers of necroinflammatory activity in chronic hepatitis C

Introduction Chronic hepatitis C (CHC) is a major cause of liver disease worldwide which often leads to progressive liver inflammation, fibrosis, cirrhosis and hepatocellular carcinoma (HCC). CHC displays heterogeneous progression depending on a broad set of factors, some of them intrinsic to each individual such as the patients genetic profile. This study aims to evaluate the contribution of certain genetic variants of crucial interferon alpha and lambda signaling pathways to the hepatic necroinflammatory activity (NIA) grade of CHC patients. Methods NIA was evaluated in 119 CHC patients by METAVIR scale and classified as low (NIA = 0–2, n = 80) or high grade (NIA = 3, n = 39). In a candidate gene approach, 64 SNPs located in 30 different genes related to interferon pathways (IL-28B, IFNAR1-2, JAK-STAT and OAS1-3, among others) were genotyped using the Illumina GoldenGate® Genotyping Assay. Statistical association was determined by logistic regression and expressed as OR and 95% CI. Those SNPs significantly associated were further adjusted by other covariates. Results Seven SNPs located in IL-28B (rs12979860), JAK1 (rs11576173 and rs1497056), TYK2 (rs280519), OAS1 (rs2057778), SOCS1 (rs33932899) and RNASEL (rs3738579) genes were significantly related to severe NIA grade (p<0.05). Regarding to clinical variables, elevated NIA was notably associated with aspartate aminotransferase (AST) serum levels >40 IU/L (p<0.05) but not with other clinical factors. Multivariate logistic regression analysis of these factors reflected that AST (>40 IU/L), TYK2 rs280519 (G allele) and RNASEL rs3738579 (G allele) were factors independently associated with elevated NIA (p<0.05). AST concentration showed a moderate AUC value (AUC = 0.63), similar to TYK2 (rs280519) and RNASEL (rs3738579) SNPs (AUC = 0.61, both) in the ROC_AUC analysis. Interestingly, the model including all significant variables reached a considerable predictive value (AUC = 0.74). Conclusion The identified genetic variants in interferon signaling pathways may constitute useful prognostic markers of CHC progression. Further validation in larger cohorts of patients is needed.

Inhibition of Tyrosine Kinase Receptor Tie2 Reverts HCV-Induced Hepatic Stellate Cell Activation

Background Hepatitis C virus (HCV) infection is a major cause of chronic liver disease (CLD) and is frequently linked to intrahepatic microvascular disorders. Activation of hepatic stellate cells (HSC) is a central event in liver damage, due to their contribution to hepatic renewal and to the development of fibrosis and hepatocarcinoma. During the progression of CLDs, HSC attempt to restore injured tissue by stimulating repair processes, such as fibrosis and angiogenesis. Because HSC express the key vascular receptor Tie2, among other angiogenic receptors and mediators, we analyzed its involvement in the development of CLD. Methods Tie2 expression was monitored in HSC cultures that were exposed to media from HCV-expressing cells (replicons). The effects of Tie2 blockade on HSC activation by either neutralizing antibody or specific signaling inhibitors were also examined. Results Media from HCV-replicons enhanced HSC activation and invasion and upregulated Tie2 expression. Notably, the blockade of Tie2 receptor (by a specific neutralizing antibody) or signaling (by selective AKT and MAPK inhibitors) significantly reduced alpha-smooth muscle actin (α-SMA) expression and the invasive potential of HCV-conditioned HSC. Conclusions These findings ascribe a novel profibrogenic function to Tie2 receptor in the progression of chronic hepatitis C, highlighting the significance of its dysregulation in the evolution of CLDs and its potential as a novel therapeutic target.

Intrahepatic angiopoietin-2 correlates with chronic hepatitis C progression and is induced in hepatitis C virus replicon systems.

Chronic hepatitis C (CHC) is a major cause of cirrhosis and hepatocellular carcinoma and angiogenesis is closely related to the pathogenesis and progression of different chronic liver diseases (CLD). Thus, the intrahepatic expression of angiopoietins 1 and 2 (Ang1 and Ang2), as relevant mediators of pathological angiogenesis in several CLD, was investigated. In addition, the differential influence of structural and non‐structural genomic regions of HCV on the expression of angiopoietins and the possible signalling involved were studied.

Angiopoietin-2/angiopoietin-1 as non-invasive biomarker of cirrhosis in chronic hepatitis C

AIM To evaluate the efficacy of peripheral blood concentrations of angiopoietins (Ang) as cirrhosis biomarkers of chronic hepatitis C (CHC). METHODS Ang1 and Ang2 serum levels were measured by enzyme-linked immunosorbent assays (ELISA) in samples from 179 cirrhotic and non-cirrhotic CHC patients, classified according to the METAVIR system. Groups were compared by non-parametric Mann-Whitney U test. Subsequently, the association of peripheral concentrations of angiopoietins with the stage of fibrosis was analyzed using Spearman correlation test. Finally, the accuracy, sensitivity and specificity of circulating angiopoietins for cirrhosis diagnosis were determined by the study of the respective area under the curve of receiver operator characteristics (AUC-ROC). RESULTS Peripheral blood concentrations of Ang1 and Ang2 in CHC patients were significantly related to fibrosis. While Ang1 was decreased in cirrhotic subjects compared to non-cirrhotic (P < 0.0001), Ang2 was significantly increased as CHC progressed to the end stage of liver disease (P < 0.0001). Consequently, Ang2/Ang1 ratio was notably amplified and significantly correlated with fibrosis (P < 0.0001). Interestingly, the individual performance of each angiopoietin for the diagnosis of cirrhosis reached notable AUC-ROC values (above 0.7, both), but the Ang2/Ang1 ratio was much better (AUC-ROC = 0.810) and displayed outstanding values of sensitivity (71%), specificity (84%) and accuracy (82.1%) at the optimal cut-off (10.33). Furthermore, Ang2/Ang1 ratio improved the performance of many other previously described biomarkers or scores of liver cirrhosis in CHC. CONCLUSION Ang2/Ang1 ratio might constitute a useful tool for monitoring the progression of chronic liver disease towards cirrhosis and play an important role as therapeutic target.

1162 ASSOCIATION OF SINGLE NUCLEOTIDE POLYMORPHISMS IN INTERFERON STIMULATED GENES WITH CHRONIC HEPATITIS C TREATMENT OUTCOME

insulin resistance and hepatic steatosis. Steatosis may increase susceptibility to apoptosis, inflammation and fibrosis by triggering hepatocytes to up-regulate CD95/Fas. We investigated this hypothesis and potential role of adipocytokines in modulating the progression of liver disease in patients with HCV-4. Methods: In 147 HCV patients and 89 controls we measured serum adiponectin, HMW adiponectin, leptin, TNF-a, IL-6 and CK-18. Liver biopsies were evaluated for steatosis/inflammation/ fibrosis, adiponectin mRNA/protein, AdipoR1/-R2 mRNA and phosphoenolpyruvate carboxykinase (PEPCK) gene expression; and adiponectin and CD95 immunoreactivity. The potential associations with hepatic steatosis and fibrosis were analyzed. Results: CD95 immunoreactivity and adiponectin immunoreactivity were readily detected in all biopsies examined and scored as grade 3 in 24 (16.3%) of patients, and as bright in 20 (13.6%) of patients, respectively. Adiponectin immunostaining within the liver correlated positively with the intensity of hepatic CD95/Fas immunostaining within the liver (r = 424; p =0.001). A significant association between high serum adiponectin and HMW adiponectin levels with CD95/Fas immunoreactivity (r = −0.16, p = 0.04, r = 0.21, p = 0.001; respectively), but not with adiponectin immunoreactivity in the liver was also identified. Adiponectin and HMW adiponectin were negatively correlated with the expression of AdipoR1, but positively correlated with the expression of AdipoR2. Hepatocyte CD95/Fas up-regulation correlated with steatosis, inflammation and fibrosis (p = 0.001). Significant correlation of serum adiponectin, HMW adiponectin and AdipoR1 and 2 mRNA expression, as well as liver adiponectin immunostaining within the liver were identified with steatosis. mRNA transcription for PEPCK was also significantly correlated with the amount of steatosis. A positive association between adiponectin and HMW adiponectin and hepatic inflammation was identified. This correlation remained significant even after following adjusting for age, gender and BMI (r = 0.17; p = 0.03; r = 0.45; p = 0.0001) respectively. Factors independently associated with the stage of fibrosis were HOMA-IR, inflammation score and age. Conclusions: Our findings in HCV-4 infection shows that adiponectin correlates with the different stages of liver injury. Steatosis up-regulates hepatocyte CD95/Fas and thus increases apoptosis, which facilitates inflammation and fibrosis. These findings may offer potential clues for future therapeutic intervention.