C. Cursaro

Diagnostic and prognostic role of alpha-fetoprotein in hepatocellular carcinoma: both or neither?

BACKGROUND:The clinical usefulness of α-fetoprotein (AFP) in hepatocellular carcinoma (HCC) management is debatable.OBJECTIVES:To assess, in a large multi-centric survey, diagnostic and prognostic reliability of AFP, predictive factors, and any correlation with the tumor immunophenotype.METHODS:A total of 1,158 patients with HCC were analyzed with reference to serum AFP levels at diagnosis. We evaluated: HCC grading, histotype, and size; Okuda, tumor–nodes–metastases (TNM), and Child-Pugh scores; liver function, symptoms, presence of metastases or portal thrombosis, etiology, survival, and treatment. In 66 patients with histological diagnosis, the pathologists evaluated p53 overexpression, MIB 1 labeling index, BCL-2 positive cells (index of apoptosis), and CD44 (adhesion molecule) positivity.RESULTS:Patients were divided into three AFP groups: normal (<20 ng/mL) [46%], elevated (21–400 ng/mL) [36%], and diagnostic (>400 ng/mL) [18%]. Statistical correlations were significant for: weight loss (P = 0.0056), pain (P = 0.0025), Child-Pugh score (P = 0.001), tumor size, Okudas and TNM stages, metastases, thrombosis, type of treatment (all p < 0.0001), and female sex (p < 0.004). AFP correlated with survival overall, in patients untreated, transplanted, or undergoing locoregional treatments; but not in those surgically treated. In the discriminant analysis, the related variables were size, female sex, Child-Pugh score, TNM staging (steps 1–4). When using the receiver operating characteristic curve, the prognostic reliability of AFP was limited with area under the curve of 0.59. Finally, patients with low expression of BCL2 had high AFP levels (p < 0.05). AFP positively correlated with Edmonson score (p < 0.0001).CONCLUSION:The evaluation of this large series of HCC patients allowed us to: confirm the low sensitivity (54%) of AFP in the diagnosis of HCC and its prognostic value, albeit limited, being tumor size, female sex (intriguingly enough), Child-Pugh score, and TNM staging independent predictors.

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.

Differential boosting of innate and adaptive antiviral responses during pegylated-interferon-alpha therapy of chronic hepatitis B

BACKGROUND & AIMS A better understanding of the immunomodulatory effects of PegIFNα therapy could allow more rational optimisation of future therapeutic approaches in chronic HBV infection. In this study, we evaluated dynamic changes in the innate and adaptive arms of the immune system induced by PegIFNα. METHODS PBMC were obtained from a cohort of patients with eAg-negative CHB before, during and after PegIFNα treatment. The number, phenotype and function of global and virus-specific T cells and NK cells were analyzed by flow cytometry and serum cytokines by ELISA or CBA. RESULTS The absolute number of CD8 T cells was strikingly reduced on PegIFNα therapy (p<0.001), with a predominant loss of end-stage effectors, including CMV-specific CD8 T cells. There was no significant recovery of the exhausted HBV-specific CD8 T cell response. By contrast, PegIFNα was able to potently and cumulatively drive the proliferation and expansion in absolute numbers of CD56(bright) NK cell numbers (p<0.001), with induction of the pro-proliferative cytokine IL-15. Expanded CD56(bright) NK cells showed enhanced expression of activation markers and the activating receptor NKp46, accompanied by augmentation of TRAIL and IFN-γ expression (p<0.001). Peak virological response (temporal within individual patients and cross-sectional within the cohort) correlated with the degree of expansion of functional CD56(bright) NK cells. CONCLUSIONS IFN-α mediates divergent effects on the innate and adaptive arms of the immune system in vivo. The efficacy of PegIFNα may be limited by its depleting effect on CD8 T cells; conversely, it can cumulatively drive proliferation, activation and antiviral potential of CD56(bright) NK cells.

Impact of interferon therapy on the natural history of hepatitis C virus related cirrhosis

BACKGROUND The role of interferon treatment on the natural history of hepatitis C virus related cirrhosis is under debate. AIM To evaluate the effect of interferon on the clinical course of compensated hepatitis C virus related cirrhosis. PATIENTS AND METHODS Seventy two cirrhotic patients treated with interferon and 72 untreated controls matched treated patients with for quinquennia of age, sex, and Child-Pughs score were enrolled in a prospective non-randomised controlled trial. Treated patients received leucocytic interferon alfa, with an escalating schedule for 12 months. The incidence and risk (Cox regression analysis) of clinical complications (hepatocellular carcinoma, ascites, jaundice, variceal bleeding, and encephalopathy) and death were calculated. RESULTS Over median follow up periods of 55 months for treated and 58 for untreated subjects, seven and nine patients, respectively, died, and 20 and 32, respectively, developed at least one clinical complication (ns). Hepatocellular carcinoma developed in six treated and 19 untreated patients (p=0.018). Seven treated patients showed sustained aminotranferase normalisation and none died or developed complications. Clinical complications were significantly associated with low albumin, bilirubin, and prothrombin activity while hepatocellular carcinoma was significantly related to no treatment with interferon, oesophageal varices, and high α fetoprotein levels. By stratified analysis, the beneficial effect of interferon was statistically evident only in patients with baseline α fetoprotein levels ⩾20 ng/ml. CONCLUSIONS Interferon does not seem to affect overall or event free survival of patients with hepatitis C virus related cirrhosis while it seems to prevent the development of hepatocellular carcinoma. Patients who achieved sustained aminotransferase normalisation survived and did not develop any complications during follow up.

Sensitive line probe assay that simultaneously detects mutations conveying resistance to lamivudine and adefovir

ABSTRACT The INNO-LiPA HBV DR v2 assay is designed to detect hepatitis B virus mutations conveying resistance to lamivudine and adefovir. Our study confirms that this assay can simultaneously detect the presence of lamivudine and adefovir resistance mutations in clinical samples, has a high degree of concordance with sequencing, and can detect mutants earlier.

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.

High risk of hepatocellular carcinoma in anti-HBe positive liver cirrhosis patients developing lamivudine resistance

Summary.  The emergence of drug‐resistant virus in hepatitis B virus patients treated with lamivudine is well documented. However, its clinical impact in the long‐term treatment of anti‐HBe positive compensated cirrhotic patients is not well known. In this study, we treated 22 consecutive patients with anti‐HBe compensated cirrhosis with lamivudine for a median period of 42 months. All patients responded to lamivudine, but viral breakthrough occurred in 13 patients (59%) between 9 and 42 months of therapy due to the emergence of a mutant strain. During the follow‐up, 11 developed hepatocellular carcinoma. Of these, 10 occurred soon after the emergence of viral resistance, generally showing aggressive behaviour, and one in the nine long‐term responder patients (P = 0.013). Lamivudine resistance was the only independent predictor of hepatocellular carcinoma development (risk ratio: 10.4; 95% CI: 1.3–84.9). Our study suggests that the occurrence of lamivudine resistance increases the risk of hepatocellular carcinoma in anti‐HBe positive cirrhosis and warrants further research.

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.

Bed-rest-induced hypernatriuresis in cirrhotic patients without ascites: does it contribute to maintain 'compensation'?

Renal function, plasma renin activity, plasma aldosterone concentration and urine excretion of free norepinephrine were evaluated in 13 cirrhotics without previous or ongoing ascites and in 13 healthy subjects, after 6 days of controlled electrolyte intake (40 mmol of Na and 70 mmol of K per day) and during 24 h of recumbency. Plasma concentrations of the atrial natriuretic peptide (ANP) were also measured in 8 patients and 8 controls. Despite a low-normal filtered load of sodium (14.6 +/- 1.2 vs. 17.1 +/- 1.2 mmol/min), cirrhotic patients showed supernormal natriuresis (141.5 +/- 14.1 vs. 78.8 +/- 8.6 mmol/day; p < 0.001). Whereas the fractional excretion of sodium in these patients was twice that of controls (0.70 +/- 0.05 vs. 0.36 +/- 0.04%; p < 0.001), potassium excretion (42.5 +/- 2.7 vs. 43.1 +/- 2.7 mmol/day) and urine volume (1270 +/- 98 vs. 1452 +/- 148 ml/day) did not differ. In cirrhotics, plasma renin activity was reduced (0.50 +/- 0.12 vs. 1.39 +/- 0.33 ng/ml/h; p < 0.02), and plasma aldosterone concentration tended to be lower (66 +/- 10 vs. 86 +/- 9 pg/ml; p = 0.09), while urine norepinephrine excretion did not significantly differ from controls (961 +/- 120 vs. 782 +/- 43 ng/h). ANP was higher in patients than in controls (92 +/- 17 vs. 48 +/- 9 pg/ml; p < 0.05). Natriuresis was directly correlated with ANP (r = 0.69, p < 0.005) and ANP/plasma aldosterone ratio (r = 0.63; p < 0.01) in patients and healthy subjects taken together.(ABSTRACT TRUNCATED AT 250 WORDS)

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.