W.P. Hofmann

Impact of donor and recipient IL28B rs12979860 genotypes on hepatitis C virus liver graft reinfection

BACKGROUND & AIMS Recent studies have described a major impact of genetic variations near the IL28B gene on the natural course and outcome of antiviral therapy in chronic hepatitis C. We therefore, aimed to explore the impact of donor and recipient genotypes of these polymorphisms on hepatitis C virus (HCV) liver graft reinfection. METHODS Donor and recipient genotypes of IL28B rs12979860C>T single nucleotide polymorphism were determined in 91 patients with HCV liver graft reinfection, 47 of whom were treated with pegylated interferon-α (PEG-IFN-α) and ribavirin. IL28B genetic polymorphisms were correlated with the natural course and treatment outcome of recurrent hepatitis C. RESULTS Patients requiring liver transplantation due to end-stage chronic hepatitis C appeared to be selected toward the adverse genotypes rs12979860 CT/TT compared to non-transplanted HCV-infected patients (p=0.046). Patients with the donor genotype rs12979860 CC had higher peak ALT and HCV RNA serum concentrations than those with CT/TT (p=0.04 and 0.06, respectively). No association was observed between ALT/HCV RNA serum concentrations and recipient genotypes (p>0.3). More important, donor IL28B rs12979860 CC vs. CT/TT genotypes were associated with rapid, complete early, and sustained virologic response (RVR, cEVR, SVR) to treatment with PEG-IFN-α and ribavirin (p=0.003, 0.0012, 0.008, respectively), but weaker associations of recipient genotypes with RVR, cEVR, and SVR were observed as well (p=0.0046, 0.115, 0.118, respectively). CONCLUSIONS We provide evidence for a dominant, but not exclusive impact of the donor rather than the recipient IL28B genetic background on the natural course and treatment outcome of HCV liver graft reinfection.

Comparison of acoustic radiation force impulse imaging with transient elastography for the detection of complications in patients with cirrhosis

Acoustic radiation force impulse (ARFI) imaging is a new non‐invasive, ultrasound‐based method for the evaluation of liver fibrosis and cirrhosis.

Prospective study of bone mineral density and metabolism in patients with chronic hepatitis C during pegylated interferon α and ribavirin therapy

Summary.  The importance of osteoporosis as a complication of end‐stage liver disease is well known. However, significant osteopenia may occur in earlier stages of chronic hepatitis C (CHC). Furthermore, antiviral therapy may influence bone metabolism. Thirty patients with CHC genotype 1 infection and without established cirrhosis were treated with peginterferon‐alfa and ribavirin. Dual‐energy x‐ray absorptiometry was performed at baseline, after 48 weeks of therapy, and by the end of a 24‐week follow‐up period. Bone mineral density (BMD), T‐scores, and Z‐scores were assessed. Serum C‐terminal propeptide of type I collagen (CICP) and osteocalcin levels were measured. Thirteen patients had osteopenia (43%) and osteoporosis was present in four patients (13%). Antiviral therapy led to significant on‐treatment increases of lumbar spine and hip BMD (P ≤ 0.05) as well as T‐scores (P ≤ 0.05) and Z‐scores (P ≤ 0.01) irrespective of subsequent treatment response. Further analyses showed that in patients with sustained virological response (n = 19) most parameters remained highly above baseline values by the end of the 24‐week follow‐up period, while patients with virological relapse (n = 11) had decreases of BMD, T‐scores and Z‐scores thereafter that did not differ from baseline. Serum CICP and osteocalcin levels decreased during therapy. Osteocalcin levels remained below baseline in sustained responder, but showed an increase in relapsers by the end of the 24‐week follow‐up (P ≤ 0.05). Osteopenia is detectable in a substantial proportion of CHC patients without established cirrhosis. Antiviral therapy leads to an on‐treatment increase of BMD, which may last in those patients who achieve a sustained virological response.

Portal vein thrombosis as complication of romiplostim treatment in a cirrhotic patient with hepatitis C-associated immune thrombocytopenic purpura

BACKGROUND & AIMS Thrombopoietin receptor agonists are a new class of compounds licenced for the treatment of immune thrombocytopenic purpura. They are currently being studied for patients with thrombopenia in advanced liver disease or under therapy for hepatitis C. There are indications that the risk for development of portal vein thrombosis in patients with advanced liver cirrhosis might be increased under therapy with thrombopoietin receptor agonists. We report a case of a patient with Child class B liver cirrhosis with concurrent immune thrombocytopenic purpura that developed portal vein thrombosis under therapy with the thrombopoietin receptor agonist romiplostim. METHODS A 50-year-old woman with hepatitis C virus associated immune thrombocytopenic purpura and Child class B liver cirrhosis presented in our emergency with rapidly evolving hydropic decompensation and general malaise. For immune thrombocytopenic purpura, the patient was started on the thrombopoietin receptor agonist romiplostim nine months ago. RESULTS During hospitalization, the platelet count was measured above 330,000/μl and partial portal vein thrombosis was diagnosed by imaging studies. The thrombotic event was assumed to be associated with the romiplostim treatment for immune thrombocytopenic purpura via excessive elevation of platelet count. After anticoagulation with heparin and cessation of romiplostim treatment, complete recanalisation of the portal vein was achieved. CONCLUSIONS We conclude that romiplostim should be used with precaution in patients with hepatitis C-associated immune thrombocytopenic purpura and advanced liver cirrhosis as the risk for thrombotic complications may increase significantly.

mTOR inhibitors and sorafenib for recurrent heptocellular carcinoma after orthotopic liver transplantation

HBeAg and HBsAg loss after long-term follow-up of HBeAg positive patients treated with peginterferon alpha-2b. Gastroenterology 2008;135:459–467. [8] Chien RN, Yeh CT, Tsai SL, Chu CM, Liaw YF. Determinants for sustained HBeAg response to lamivudine therapy. Hepatology (Baltimore, Md.) 2003;38:1267–1273. [9] Buti M, Cotrina M, Valdes A, Jardi R, Rodriguez-Frias F, Esteban R. Is hepatitis B virus subtype testing useful in predicting virological response and resistance to lamivudine? J Hepatol 2002;36:445–446. [10] Lurie Y, Manns MP, Gish RG, Chang TT, Yurdaydin C, Lai CL, et al. The efficacy of entecavir is similar regardless of disease-related baseline subgroups in treatment of nucleoside-naive, HBeAg(+) and HBeAg( ) patients with chronic hepatitis B. J Hepatol 2005;42:184. [11] Yuen MF, Wong DK, Sablon E, Yuan HJ, Sum SM, Hui CK, et al. Hepatitis B virus genotypes B and C do not affect the antiviral response to lamivudine. Antivir Ther 2003;8:531–534. [12] Buster EH, Hansen BE, Verhey E, De Man RA, Janssen HL. HBV genotype is an important predictor of sustained off-treatment response to both peginterferon alpha-2b and entecavir in HBeAg positive chronic hepatitis B. Hepatology (Baltimore, MD) 2008;48:716A.

Correlation of amino acid variations within nonstructural 4B protein with initial viral kinetics during interferon-alpha-based therapy in HCV-1b-infected patients

Summary.  Chronic hepatitis C is a major cause of liver cirrhosis leading to chronic liver failure and hepatocellular carcinoma. Different hepatitis C virus (HCV) proteins have been associated with resistance to interferon‐alpha‐based therapy. However, the exact mechanisms of virus‐mediated interferon resistance are not completely understood. The importance of amino acid (aa) variations within the HCV nonstructural (NS)4B protein for replication efficiency and viral decline during the therapy is unknown. We investigated pretreatment sera from 42 patients with known outcome to interferon‐based therapy. The complete NS4B gene was amplified and sequenced. Mutational analyses of predicted conformational, functional, structural and phylogenetic properties of the deduced aa sequences were performed. The complete NS4B protein was highly conserved with a median frequency of 0.015 ± 0.009 aa exchanges (median ± SD, 4.00 ± 2.31). Especially within the predicted transmembranous domains of the NS4B protein, the mean number of aa variations was low (median frequency, 0.013 ± 0.013). Neither the number of aa variations nor specific aa exchanges were correlated with HCV RNA serum concentration at baseline. A rapid initial HCV RNA decline of ≥1.5 log10 IU/mL at week 2 of interferon‐based therapy was associated with a higher frequency of nonconservative aa exchanges within the complete NS4B protein in comparison with patients with a nonrapid HCV RNA decline (median frequency, 0.011 ± 0.005 vs 0.004 ± 0.003, P = 0.006). Overall, the aa sequence of the NS4B protein was highly conserved, indicating an important role for replication in vivo. Amino acid variations with relevant changes of physicochemical properties may influence replication efficiency, associated with a rapid early virological response.

Dynamics of hepatitis B virus quasispecies heterogeneity and virologic response in patients receiving low-to-moderate genetic barrier nucleoside analogs

We characterized the early dynamics of hepatitis B virus (HBV) quasispecies evolution during the first weeks of antiviral therapy with low‐to‐moderate genetic barrier antiviral drugs and associated these data with antiviral response patterns. Fifteen chronic hepatitis B patients (men, 10; mean age, 34; HBeAg positive, 6) who received lamivudine or telbivudine for at least 52 weeks were included. HBV DNA was extracted from serum, and a 910‐bp fragment covering domains A–F of the reverse transcriptase region was amplified, cloned and sequenced. Parameters of quasispecies heterogeneity, genetic diversity and complexity were calculated and were correlated with complete virologic response, defined as undetectable HBV DNA at week 52. Nine patients achieved complete virologic response during the observational period. While baseline HBV DNA levels and HBeAg status were associated with virologic response, baseline quasispecies complexity and diversity of responders showed no significant difference to those of nonresponders (P > 0.05). However, at week 4, quasispecies complexity of nonresponders was significantly higher compared with that of responders on the nucleotide level (P = 0.01) and the aa level (P = 0.04). The number of synonymous substitutions per synonymous site dropped significantly in responders at week 4 (P = 0.04), while there was no difference in nonresponders. The HBV quasispecies complexity at the early stage of antiviral therapy (week 4) with the low‐to‐moderate genetic barrier nucleoside analogs lamivudine or telbivudine was associated with subsequent virologic response. Further studies are needed to confirm HBV quasispecies evolution as additional predictive marker for beneficial treatment outcome.

Mutations in the putative HCV-E2 CD81 binding regions and correlation with cell surface CD81 expression

Summary.  The hepatitis C virus (HCV) envelope (E)2 protein interacts with the cellular receptor CD81 leading to modulation of B and T cell function. Recently, a higher binding affinity of subtype 1a in comparison with 1b derived E2 proteins for CD81 in vitro was described. The importance of mutations within the putative CD81 binding regions of different HCV geno‐/subtypes in correlation with CD81 expression is unknown. In the present study, CD81 expression on blood lymphocytes of patients with chronic hepatitis C infected with different HCV geno‐/subtypes were analysed by fluorescence activated cell sorter analyses. In addition, the putative CD81 binding regions on the E2 gene comprising the hypervariable region (HVR)2 were analysed by direct sequencing. CD81 expression on CD8(+) T‐lymphocytes from patients infected with subtype 1a (n = 6) was significantly higher in comparison with subtype 1b (n = 12) and 3 (n = 5) infected patients before and during antiviral therapy (P = 0.006; P = 0.021, respectively). Sequencing of the putative CD81 binding regions in the E2 protein comprising the HVR2 (codon 474–495 and 522–552 according to the HCV‐1a prototype HCV‐H) showed a highly conserved motif within HVR2 for subtype 1a isolates and an overall low number of mutations within the putative CD81 binding regions, whereas numerous mutations were detected for subtype 1b isolates (12.0 vs 23.6%). HCV‐3 isolates showed an intermediate number of mutations within the putative binding sites (19.2%; P = 0.022). In conclusion, the highly conserved sequence within HVR2 and putative CD81 binding sites of subtype 1a isolates previously associated with a high CD81 binding affinity in vitro is correlated with high CD81 expression on CD8(+) T‐lymphocytes in vivo.

Cognition in hepatitis C patients treated with pegylated interferon

Neuropsychiatric symptoms are frequently reported by patients with chronic hepatitis C during treatment with interferon (IFN)-α and may lead to treatment discontinuation. In order to assess the objective neuropsychiatric impairments we prospectively administered standardized neuropsychological testing before and 3 months after the initiation of antiviral treatment with pegylated IFN-α in 17 patients suffering from chronic hepatitis C. In addition depression and anxiety scores, social functioning and hepatological parameters were obtained. While depressive and anxiety symptoms increased significantly during treatment only subtle worsening in neurocognitive performance could be detected, indicating slight impairment in cognitive flexibility and psychomotor speed (Trail Making Test B; 69.6±23 s before vs. 80.7±31 s during therapy, P=0.011, not remaining significant after Bonferroni correction). We found no association between reduced neurocognitive performance and the severity of depression. Better neurocognitive performance in single domains was associated with better response to antiviral treatment measured as the decline of elevated liver enzymes (AST). We conclude that neurocognitive performance was influenced by antiviral IFN-α-based combination treatment only in single domains and not to a clinically relevant extent in contrast to the significant worsening of depression.

Somatic hypermutation and mRNA expression levels of the BCL-6 gene in patients with hepatitis C virus-associated lymphoproliferative diseases*

Summary.  Chronic hepatitis C virus (HCV) infection leads to mixed cryoglobulinaemia (MC) and B‐cell non‐Hodgkin lymphoma (B‐NHL). Aberrant somatic hypermutation and deregulation of the oncogene BCL‐6 is associated with lymphomagenesis. Recently, HCV was shown to induce BCL‐6 mutations in vitro. The BCL‐6 gene (area B) was cloned and sequenced from peripheral blood mononuclear cells (PBMC) of 21 chronically HCV‐infected patients with or without MC and B‐NHL, and six healthy controls. Mutational frequencies, genetic complexity and diversity were calculated. BCL‐6 mRNA from PBMC was quantified by real‐time polymerase chain reaction, and additional sustained virologic responders to antiviral therapy and HBV patients served as controls. The overall/recurrent mutational frequencies tended to be lower in MC and B‐NHL patients when compared with controls (P = 0.15 and 0.06, respectively). Genetic complexity was significantly lower in MC and B‐NHL patients (P = 0.025). BCL‐6 mRNA concentration was decreased in all HCV patients when compared with healthy controls, sustained virologic responder and HBV patients (P = 0.005). Although HCV can induce BCL‐6 mutations in vitro, lower mutational frequencies and decreased BCL‐6 mRNA expression in vivo suggest no major role of aberrant somatic hypermutation in HCV‐associated MC and B‐NHL.