Marzia Margotti

Genome-wide meta-analyses identify three loci associated with primary biliary cirrhosis

A genome-wide association screen for primary biliary cirrhosis risk alleles was performed in an Italian cohort. The results from the Italian cohort replicated IL12A and IL12RB associations, and a combined meta-analysis using a Canadian dataset identified newly associated loci at SPIB (P = 7.9 × 10−11, odds ratio (OR) = 1.46), IRF5-TNPO3 (P = 2.8 × 10−10, OR = 1.63) and 17q12-21 (P = 1.7 × 10−10, OR = 1.38).

Silybin combined with phosphatidylcholine and vitamin E in patients with nonalcoholic fatty liver disease: A randomized controlled trial

The only currently recommended treatment for nonalcoholic fatty liver disease (NAFLD) is lifestyle modification. Preliminary studies of silybin showed beneficial effects on liver function. Realsil (RA) comprises the silybin phytosome complex (silybin plus phosphatidylcholine) coformulated with vitamin E. We report on a multicenter, phase III, double-blind clinical trial to assess RA in patients with histologically documented NAFLD. Patients were randomized 1:1 to RA or placebo (P) orally twice daily for 12 months. Prespecified primary outcomes were improvement over time in clinical condition, normalization of liver enzyme plasma levels, and improvement of ultrasonographic liver steatosis, homeostatic model assessment (HOMA), and quality of life. Secondary outcomes were improvement in liver histologic score and/or decrease in NAFLD score without worsening of fibrosis and plasma changes in cytokines, ferritin, and liver fibrosis markers. We treated 179 patients with NAFLD; 36 were also HCV positive. Forty-one patients were prematurely withdrawn and 138 patients analyzed per protocol (69 per group). Baseline patient characteristics were generally well balanced between groups, except for steatosis, portal infiltration, and fibrosis. Adverse events (AEs) were generally transient and included diarrhea, dysgeusia, and pruritus; no serious AEs were recorded. Patients receiving RA but not P showed significant improvements in liver enzyme plasma levels, HOMA, and liver histology. Body mass index normalized in 15% of RA patients (2.1% with P). HCV-positive patients in the RA but not the P group showed improvements in fibrogenesis markers. This is the first study to systematically assess silybin in NAFLD patients. Treatment with RA but not P for 12 months was associated with improvement in liver enzymes, insulin resistance, and liver histology, without increases in body weight. These findings warrant further investigation.

Cytokine profile of peripheral blood mononuclear cells from patients with different outcomes of hepatitis C virus infection

Summary.  The relationship between the balance of helper T‐cell type 1 (Th1) or type 2 (Th2) cytokines and the clinical course of hepatitis C virus (HCV) infection is unclear. We evaluated Th1 [interleukin (IL)‐2, interferon‐gamma (IFN‐γ)] and Th2 cytokine (IL‐4, IL‐10) and 2,5‐oligoadenylate synthetase (OAS, an IFN‐induced antiviral protein) production by peripheral blood mononuclear cells from 10 healthy anti‐HCV‐positive individuals (group A), 10 HCV‐RNA‐positive with persistently normal alanine aminotransferase (ALT) levels (group B), 10 HCV‐RNA‐positive with abnormal ALT (group C) and 10 uninfected healthy controls. IL‐2 production was significantly increased in group B when compared with all the other groups. No difference was found for IFN‐γ. IL‐4 was significantly higher in group C than in both group B (P = 0.0006) and controls (P = 0.004). Compared with controls, IL‐10 was significantly decreased in group A (P = 0.013) and B (P = 0.004). The production of 2,5‐OAS was significantly higher in group B than in A (P = 0.04) and in C (P = 0.004). Finally, in all HCV‐RNA‐positive patients, a significant correlation was found between ALT and both IL‐2 (r = −0.78; P = 0.0008) and IL‐4 (r = 0.75; P = 0.0008). In conclusion: (i) subjects who cleared HCV showed a cytokine profile similar to controls; (ii) a preferential shift towards a Th1 profile seems associated with a more favourable clinical outcome in chronic hepatitis C; and (iii) a prevalent Th2 profile seems implicated in HCV pathogenesis and severity of liver disease.

In vitro effect of thymosin-α1 and interferon-α on Th1 and Th2 cytokine synthesis in patients with chronic hepatitis C

Current evidence suggests that increased expression of Th1‐associated cytokines is important for immune‐mediated eradication of hepatitis C infection, while an increase in Th2‐associated cytokines is associated with persistence of infection. In this study we evaluated the effects of thymosin‐α1 (TA1), a naturally occurring thymic peptide, and interferon‐α (IFN‐α) on cytokine production in peripheral blood mononuclear cells from untreated patients with chronic hepatitis C. We examined the effect of incubation with TA1, IFN‐α, or both, on production of Th1‐associated cytokines (IL‐2, IFN‐γ), Th2‐associated cytokines (IL‐4, IL‐10), and synthesis of the antiviral protein 2′,5′‐oligoadenylate synthetase. TA1 treatment induced a significant increase in production of IL‐2 and 2′,5′‐oligoadenylate synthetase. Smaller increases were also seen after treatment with IFN‐α, while incubation with TA1 and IFN‐α together led to an additive or synergistic effect. Incubation with TA1 resulted in a decrease in IL‐4 and IL‐10, whereas IFN‐α increased these cytokines. The addition of TA1 to IFN‐α significantly reversed this IFN‐α‐induced increase. Hence, TA1 treatment could benefit patients with hepatitis C infection by increasing the Th1‐type response, fundamental for sustained clearance of hepatitis C; and by decreasing the Th2‐type response, associated with persistence of viraemia.

Vitamin E as treatment for chronic hepatitis B: results of a randomized controlled pilot trial.

BACKGROUND AND AIMS Interferon-alpha treatment has been the treatment of choice for chronic hepatitis with unpredictable results. Recently, Lamivudine has been licensed for use against HBV infection with good results. Unfortunately, recurrence of viremia after lamivudine withdrawal is common and prolonged treatment can induce the emergence of resistant mutant strains. It has been shown that vitamin E can increase the host immune response, and this may provide protection against infectious diseases. METHODS We evaluated vitamin E supplementation as therapy for chronic hepatitis B in a pilot study including 32 patients. Patients were randomly allocated to receive vitamin E at the dose of 300 mg twice daily for 3 months (15 patients) or no treatment (17 patients). They were seen monthly during the first 3 months and thereafter quarterly for additional 12 months. RESULTS The two groups were comparable at enrollment. At the end of the study period, alanine aminotransferase (ALT) normalization was observed in 7 (47%) patients in vitamin E group and only in 1 (6%) of the controls (P=0.011); HBV-DNA negativization was observed in 8 (53%) patients in the vitamin E group as compared to 3 (18%) in the control group, respectively (P=0.039). A complete response (normal ALT and negative HBV-DNA) was obtained in 7 (47%) patients taking vitamin E and in none of the controls (P=0.0019). CONCLUSION Vitamin E supplementation might be effective in the treatment of chronic hepatitis B.

Laboratory signs of acute or recent cytomegalovirus infection are common in cirrhosis of the liver

Human cytomegalovirus (CMV) is an ubiquitous pathogen that can cause severe and often fatal infections in immunocompromised patients. Patients with cirrhosis often show various degrees of impaired cellular immunity that could lead to acute CMV reactivation. The aim of the present study was to determine whether laboratory findings of active CMV infections are common in patients with cirrhosis. Fifty‐five patients with cirrhosis were studied for acute CMV infection by virological (antigenemia and quantitative polymerase chain reaction in polymorphonuclear leukocytes) and serological (detection of anti‐CMV IgM by immunoblot) methods. The same tests were carried out on 50 blood donors and on 20 chronic hepatitis patients, considered as control populations. Acute or recent CMV infection had occurred in 31 (56%) of 55 patients with cirrhosis, whereas only 1 out of 20 (5%) patients with chronic non‐cirrhotic liver disease and none of the 50 blood donors had laboratory signs of active CMV infection. The difference between patients with cirrhosis and the control groups was significant (P < 0.001, χ2 test). CMV in patients with cirrhosis was not related to age, gender, hepatitis C virus infection or hepatocellular carcinoma. There was no significant correlation between impairment of liver function and the presence of active CMV infection. Patients with cirrhosis should be considered at risk for CMV infection, that seems to be mild and asymptomatic. J. Med. Virol. 62:25–28, 2000.

Evaluation of the effects of human leukocyte IFN-α on the immune response to the HBV vaccine in healthy unvaccinated individuals

HBV vaccine needs 3 injections over 6 months to induce immunity. Thus, the use of adjuvants capable of inducing earlier immune protection would be highly desirable. Most adjuvants may act by inducing cytokines, and among them, type I interferons (IFNs), deserve a special attention in view of the potent immunomostimulatory activity observed in mouse models and on dendritic cell functions. The aim of the present trial was to evaluate the effects of IFN-alpha administered as an adjuvant of HBV vaccine in healthy unvaccinated individuals. No significant enhancing effect on the antibody response was observed, in spite of an early and transient upregulation of costimulatory molecule expression on peripheral blood mononuclear cells, which may be suggestive of an IFN-mediated activation of antigen presenting cells. We conclude that, under the conditions used in this trial, natural IFN-alpha does not act as an adjuvant of the HBV vaccine in healthy unvaccinated individuals.

Incidence of DAA failure and the clinical impact of retreatment in real-life patients treated in the advanced stage of liver disease: Interim evaluations from the PITER network

Background Few data are available on the virological and clinical outcomes of advanced liver disease patients retreated after first-line DAA failure. Aim To evaluate DAA failure incidence and the retreatment clinical impact in patients treated in the advanced liver disease stage. Methods Data on HCV genotype, liver disease severity, and first and second line DAA regimens were prospectively collected in consecutive patients who reached the 12-week post-treatment and retreatment evaluations from January 2015 to December 2016 in 23 of the PITER network centers. Results Among 3,830 patients with advanced fibrosis (F3) or cirrhosis, 139 (3.6%) failed to achieve SVR. Genotype 3, bilirubin levels >1.5mg/dl, platelet count <120,000/mm3 and the sofosbuvir+ribavirin regimen were independent predictors of failure by logistic regression analysis. The failure rate was 7.6% for patients treated with regimens that are no longer recommended or considered suboptimal (sofosbuvir+ribavirin or simeprevir+sofosbuvir±ribavirin), whereas 1.4% for regimens containing sofosbuvir combined with daclatasvir or ledipasvir or other DAAs. Of the patients who failed to achieve SVR, 72 (51.8%) were retreated with a second DAA regimen, specifically 38 (52.7%) with sofosbuvir+daclatasvir, 27 (37.5%) with sofosbuvir+ledipasvir, and 7 (9.7%) with other DAAs ±ribavirin. Among these, 69 (96%) patients achieved SVR12 and 3 (4%) failed. During a median time of 6 months (range: 5–14 months) between failure and the second DAA therapy, the Child-Pugh class worsened in 12 (16.7%) patients: from A to B in 10 patients (19.6%) and from B to C in 2 patients (10.5%), whereas it did not change in the remaining 60 patients. Following the retreatment SVR12 (median time of 6 months; range: 3–12 months), the Child-Pugh class improved in 17 (23.6%) patients: from B to A in 14 (19.4%) patients, from C to A in 1 patient (1.4%) and from C to B in 2 (2.9%) patients; it remained unchanged in 53 patients (73.6%) and worsened in 2 (2.8%) patients. Of patients who were retreated, 3 (4%) had undergone OLT before retreatment (all reached SVR12 following retreatment) and 2 (2.8%) underwent OLT after having achieved retreatment SVR12. Two (70%) of the 3 patients who failed to achieve SVR12 after retreatment, and 2 (2.8%) of the 69 patients who achieved retreatment SVR12 died from liver failure (Child-Pugh class deteriorated from B to C) or HCC complications. Conclusions Failure rate following the first DAA regimen in patients with advanced disease is similar to or lower than that reported in clinical trials, although the majority of patients were treated with suboptimal regimens. Interim findings showed that worsening of liver function after failure, in terms of Child Pugh class deterioration, was improved by successful retreatment in about one third of retreated patients within a short follow-up period; however, in some advanced liver disease patients, clinical outcomes (Child Pugh class, HCC development, liver failure and death) were independent of viral eradication.

In vitro effect of thymosin-alpha1 and interferon-alpha on Th1 and Th2 cytokine synthesis in patients with eAg-negative chronic hepatitis B

Summary.  Thymosin alpha‐1 (Tα1) has been shown to be effective in chronic hepatitis B treatment. This study investigated the effect of Tα1 and interferon‐alpha (IFNα) on cytokine production by peripheral blood mononuclear cells (PBMCs) of 12 patients with eAg‐negative chronic hepatitis B (HBV). We evaluated the effect of incubation with Tα1, IFNα or both on the synthesis of T‐helper 1 (Th1) cytokines [interleukin‐2 (IL‐2), IFNγ] and Th2 cytokines (IL‐4, IL‐10) and of antiviral protein 2′,5′‐oligoadenylate synthetase (2′,5′‐OAS) in patients and in a group of 10 healthy controls. Concerning Th1 profile, controls showed lower IL‐2 synthesis than HBV patients. In HBV setting, IFNα/Tα1 combination was able to increase IL‐2 production significantly, when compared with baseline condition. About the Th2‐cytokines, controls showed statistically lower synthesis of IL‐4 and higher production of IL‐10, than HBV patients. In these latter, IFNα increased the synthesis of IL‐10 compared with baseline. Interestingly, both Tα1 alone and the IFNα/Tα1 combination reversed this effect. Finally, compared with baseline, the synthesis of 2′,5′‐OAS was significantly higher in the presence of Tα1 and IFNα alone, and in the presence of IFNα/Tα1 association, while no differences were found between controls and HBV patients. In conclusion, in PBMCs from eAg‐negative HBV patients, Tα1 alone was able to increase the antiviral protein synthesis, while in association with IFNα, it stimulated the IL‐2 synthesis and inhibited the IFN‐induced IL‐10 production. These results need further investigations, but reinforce the idea of an immunotherapeutic approach for chronic hepatitis B.

Letter: calcineurin inhibitor level reduction during treatment with sofosbuvir in liver transplanted patients

SIRS, In liver transplant, during anti-viral therapy for hepatitis C virus (HCV) recurrence, the immunosuppressant levels should be monitored to prevent both toxicity and rejection. Sofosbuvir (SOF) has been used within compassionate programs for HCV recurrence and, according to pharmacokinetic analyses, is not supposed to have significant pharmacological interactions with tacrolimus (Tac) or ciclosporin. This was reported in the review article by Koff recently published. We treated eight transplant recipients with SOF/ribavirin (RBV) for a severe HCV recurrence, and observed unexpected Tac/ciclosporin reduction during SOF. The patients’ mean age was 48.1 years. They were all men and had mean time from liver transplant 4.2 years, mean MELD-score 13.4 (range, 7–22), mean Child-Turcotte-Pugh score 8 (6–11). There were six (75%) genotype 1, one (12.5%) was genotype 3, and one (12.5%) was genotype 4. All patients were on a stable Tac regimen except one who was on ciclosporin. Treatment with oral daily doses of SOF 400 mg + RBV (range, 200–1000 mg) was started after the SOF compassionate use approval by the Ethics Committee. Initial Tac/ciclosporin dose was not modified at the beginning of anti-viral treatment. The Tac/ciclosporin levels were monitored weekly (Figure 1). At week 4, mean Tac/ciclosporin reduction was 40.2% (range, 21.4–72.1%). Daily Tac/ciclosporin doses were increased in all, except one patient who had renal failure and was also receiving mycophenolate mofetil. All patients except one reached undetectable HCV-RNA at week 4. The mean CTP reduced from 8 at baseline to 6 at week 4. Currently available data report no significant reciprocal influence of absorption, distribution, metabolism or excretion of Tac/ciclosporin and SOF. No dose adjustment is recommended for immunosuppressants during SOF, as indicated in the recent review article (2). Nonetheless, this information arises from a single-dose study, while our data reveal a reduction in immunosuppressant levels. This might be a consequence of an improvement of liver function, even if a drug–drug interactions cannot be a priori excluded. Regardless of the underlying mechanism, our report is aimed to raise awareness to hepatologists who are familiarising themselves with SOF in the post-transplant HCV-recurrence setting as we are now moving towards SOF global availability.