Pegylated interferon-alpha, ribavirin, and rituximab combined therapy of hepatitis C virus-related mixed cryoglobulinemia: a long-term study.

This study illustrates the use and efficacy of a combination of pegylated interferon-alpha (Peg-IFN-alpha) and ribavirin (RBV), with or without rituximab (RTX), in hepatitis C virus (HCV)-related mixed cryoglobulinemia (MC). Twenty-two patients with HCV-related MC received Peg-IFN-alpha (2a: 180 mug or 2b: 1.5 mug/kg) weekly plus RBV (1000 or 1200 mg) daily for 48 weeks, and RTX (375 mg/m(2)) once a week for 1 month followed by two 5-monthly infusions (termed PIRR). Fifteen additional patients received Peg-IFN-alpha/RBV with the same modalities as the PIRR schedule. Complete response was achieved in 54.5% (12/22) and in 33.3% (5/15) of patients who received PIRR and Peg-IFN-alpha/RBV, respectively (P < .05). Clearance of HCV RNA and conversion of B-cell populations from oligoclonal to polyclonal in liver, bone marrow, and peripheral blood was maintained for up to 3 years in 10 of 12 (83.3%) and in 2 of 5 (40%) patients receiving PIRR and Peg-IFN-alpha/RBV, respectively (P < .01). Cryoproteins in 22.7% (5/22) of patients with PIRR and in 33.3% (5/15) with Peg-IFN-alpha/RBV persisted despite sustained HCV RNA clearance. No response occurred in remaining 5 patients of both groups. PIRR therapy is well tolerated and more effective than Peg-IFN-alpha/RBV combination in HCV-related MC. Its effect may last for more than 3 years.

Properties of Two Enterovirus Antibodies that are Utilized in Diabetes Research.

Human enteroviruses (EVs) comprise >100 different types. Research suggests a non-chance association between EV infections and type 1 diabetes. Immunohistochemical studies with the anti-EV antibody 5D-8.1 have shown that the EV capsid antigen is present in pancreatic islet cells of diabetic subjects. When it was noticed that 5D-8.1 may cross-react with human proteins, doubt was casted on the significance of the above histopathologic findings. To address this issue, properties of EV antibodies 5D-8.1 and 9D5 have been investigated using peptide microarrays, peptide substitution scanning, immunofluorescence of EV-infected cells, EV neutralization assays, bioinformatics analysis. Evidence indicates that the two antibodies bind to distinct non-neutralizing linear epitopes in VP1 and are specific for a vast spectrum of EV types (not for other human viruses). However, their epitopes may align with a few human proteins at low expected values. When tested by immunofluorescence, high concentrations of 5D-8.1 yelded faint cytoplasmic staining in uninfected cells. At reduced concentrations, both antibodies produced dotted staining only in the cytoplasm of infected cells and recognized both acute and persistent EV infection. Thus, the two monoclonals represent distinct and independent probes for hunting EVs in tissues of patients with diabetes or other endocrine conditions.

Thymic stromal lymphopoietin in hepatitis C virus-related cryoglobulinemic vasculitis: gene expression level and protein distribution.

INTRODUCTION: Hepatitis C virus (HCV) infection can be detected in virtually all patients with cryoglobulinemic vasculitis (CV). Among its many effects, the virus is able to stimulate the production of thymic stromal lymphopoietin (TSLP) by infected hepatocytes. In this study, we assessed the systemic levels and tissue distribution of TSLP in 60 chronically HCV-infected patients, 36 with and 24 without CV. METHODS: Serum TSLP levels were measured by an enzyme-linked immunosorbent assay (ELISA) method. TSLP mRNA was assessed in patient samples by real-time reverse transcriptase-polymerase chain reaction (RT-PCR). TSLP protein in liver and skin biopsy samples was revealed by indirect immunofluorescence. All other methods were carried out according to standardized procedures. RESULTS: Serum TSLP levels were significantly higher in patients with than in those without CV and in healthy individuals. Higher TSLP levels paralleled specific mRNA expression and the up-regulation of TSLP protein in liver tissue. Compared with non-CV patients, higher TSLP levels in CV were accompanied by a higher frequency of circulating mono/oligoclonal B-cell expansions (8% vs. 92%, p<0.0001) and a higher number of peripheral CD20+ B-cells (10.3% vs. 15.5% p=0.04). In addition, TSLP mRNA expression in the liver of CV patients was lower than in their correspondent skin tissue and paralleled specific immune deposits of TSLP protein in keratinocytes. CONCLUSION: Overall, this study shows that TSLP secreted by hepatocytes and keratinocytes of HCV-infected patients with CV is involved in the pathogenesis of vasculitis and may possibly support the therapeutic use of TSLP-targeted monoclonal antibodies.

Assessment of total hepatitis C virus (HCV) core protein in HCV-related mixed cryoglobulinemia.

INTRODUCTION: In hepatitis C virus (HCV)-related mixed cryoglobulinemia (MCG), the nonenveloped HCV core protein (HCV-Cp) is a constituent of the characteristic cold-precipitating immune complexes (ICs). A possible correlation between HCV-Cp, virologic, laboratory, and clinical parameters in both untreated MCG patients and those undergoing specific treatment was explored. METHODS: HCV-Cp was quantified by a fully automated immune assay. Correlations between HCV-Cp and HCV RNA, cryocrit, and virus genotype (gt) were investigated in 102 chronically HCV-infected MCG patients. RESULTS: HCV-Cp concentrations strongly correlated with HCV RNA levels in baseline samples. An average ratio of 1,425 IU and 12,850 IU HCV RNA per picogram HCV-Cp was estimated in HCV gt-1 and gt-2 patients, respectively. This equation allowed us to estimate that, on average, HCV-Cp was associated with the viral genome in only 3.4% of the former and in 35% of the latter group of patients. The direct relation between HCV-Cp and the cryocrit level suggests that the protein directly influences the amount of cryoprecipitate. Although the therapy with rituximab (RTX) as a single agent resulted in the enhancement of HCV-Cp levels, in patients treated with RTX in combination with a specific antiviral therapy (pegylated interferon-α plus ribavirin), the prompt and effective clearance of HCV-Cp was documented. CONCLUSIONS: Our data provide evidence that HCV-Cp has a direct effect on the cold-precipitation process in a virus genotype-dependence in HCV-related MCG patients.

Clonal CD27+ CD19+ B cell expansion through inhibition of FC gammaIIR in HCV(+) cryoglobulinemic patients.

Persistent HCV infection may be associated with extrahepatic manifestations such as type II mixed cryoglobulinemia (II-MC), a clonal B cell proliferative disorder. In persistent HCV infection without II-MC, an increase in serum immunoglobulins (Ig) is commonly observed. This increase is polyclonal and is determined primarily by increased levels of IgG which include both HCV-specific and nonspecific antibodies. Nonetheless, memory CD27(+) B cells do not accumulate. This paradoxical phenomenon depends on heightened sensitivity of memory B cells to BCR-independent noncognate T cell help, which speeds up their terminal differentiation into antibody-secreting cells and makes them more prone to apoptosis. In persistent HCV infection with II-MC, serum Ig elevation is also a general occurrence, and characteristically includes IgM antibodies with rheumatoid factor activity, which are essential for the development of circulating, cryoprecipitable immune complexes. Hypergammaglobulinemia is sustained by a peripheral expansion of IgM(+)k(+)IgD(low/neg)CD21(low)CD27(+) B cells. These cells exhibit marked V(H), J(H), and V(K) gene segment usage restriction, indicating that a limited number of antigens drive their proliferation through BCR interaction. Recently, two epitopes, one of the human IgG and the second of the HCV(NS3) protein, had been identified and demonstrated able to link the BCR exposed on II-MC subjects. Based on the above findings, we propose a model whereby BCR binding the IgM/IgG/HCV(NS3) immune complexes deprives Fc gammaIIR of its natural ligand. This takes the brake off RF(+)CD27(+) B cell proliferation and promotes their selective accumulation, which is otherwise prevented by increased apoptosis susceptibility in persistent HCV infection without II-MC.

B cells and HCV: an infection model of autoimmunity.

In addition to cause acute and chronic liver disease, hepatitis C virus (HCV) infection is frequently associated to autoimmune disorders, such as mixed cryoglobulinemia, primary glomerulonephritis, monoclonal gammopathy of undetermined significance and post-transplant proliferative disorders. Progression to malignant phenotype of B cells may be the consequence of additional genetic events or abnormal conditions resulting from modification of host cell genes involved in the control of oncogenes and oncoproteins. In this review, we will address the potential immune disregulatory mechanism(s) underlying HCV persistence. In addition, HCV/B-cell interaction that might explain defects in humoral immunity in individuals who develop chronic virus carriage and lymphoproliferative disorders will be emphasized.

Recent prognostic factors in diffuse large B-cell lymphoma indicate NF-kappaB pathway as a target for new therapeutic strategies.

The diffuse large B-cell lymphoma (DLBCL) is the most common type of lymphoid cancer. The classical chemotherapy regimen given to these patients was the CHOP (Cyclophosphamide, Hydroxydaunorubicin or Adriamycin, Oncovin or Vincristine, Prednisone or Prednisolone), but recently rituximab with CHOP (R-CHOP) increased the number of cases responding to first line therapy. DLBCL classification identified three principle subgroups. The first one, named germinal centre B cell-like (GCB), responds to both CHOP and R-CHOP treatment and it is mainly characterised by the expression of markers like Bcl-6 and CD10. The second, the activated B-cell like (ABC), has a worse prognosis in comparison with GCB, and is mainly characterised by the expression of IRF-4, PRDM1 and NF-kappaB. It is interesting to notice that IRF-4 and PRDM1 are under the transcriptional control of NF-kappaB, whose high activation level is associated with a worse prognosis. The third one, mediastinal large B-cell lymphoma (PMBCL) is an uncommon subtype characteristically found in young females. Gene expression profiling suggests that this disease resembles Hodgkin lymphoma more than other types of DLBCL. The impact of rituximab on the outcome of patients with PMBCL has still not been fully assessed. It was seen that rituximab inhibits NF-kappaB pathway in vitro. However, the clinical significance of this finding is still unknown, because both ABC and GCB DLBCL show a significant improvement of overall survival after R-CHOP treatment. In this review, the NF-kappaB pathway is suggested as a target for new chemotherapy strategies based on the association of CHOP with molecules more effective than rituximab in this pathway inhibition.