Baseline anti-NS4a antibodies in combination with on-treatment quantitative HCV-RNA reliably identifies nonresponders to pegylated interferon-ribavirin combination therapy after 4 weeks of treatment

Background Early detection of nonresponders to hepatitis C therapy limits unnecessary exposure to treatment and its side-effects. A recent algorithm combining baseline anti-NS4a antibodies and on-treatment quantitative PCR identified nonresponders to a combination of interferon and ribavirin after 1 week of treatment. Aim To validate a stopping rule based on baseline anti-NS4a antibody levels and early on-treatment virological response in treatment-naive genotype 1 chronic hepatitis C patients treated with the current standard pegylated interferon and ribavirin combination therapy. Methods Eighty-nine genotype 1 patients from the Dynamically Individualized Treatment of hepatitis C Infection and Correlates of Viral/Host dynamics Study treated for 48 weeks with standard 180 mu g pegylated interferon (PEG-IFN)-alpha-2a (weekly) and ribavirin 1000-1200mg (daily) were analysed. Baseline anti-NS4a antibody enzyme-linked immunosorbent assay (NS4a AA 1687-1718) was performed on pretreatment serum. Hepatitis C virus-RNA was assessed at days 0, 1, 4, 7, 8, 15, 22, 29, weeks 6, 7, 8, 10, 12 and 6 weekly thereafter until end of treatment. Multiple regression logistic analysis was performed. Results Overall 54 of 89 (61%) patients achieved sustained virological response. A baseline anti-NS4a antibody titre less than 1/1250 correlated with absence of favourable initial viral decline according to variable response types (P=0.015). The optimal algorithm was developed using the combination of the absence of anti-NS4a Ab (= 100.000 IU/ml at week 4. This algorithm has a specificity of 43% and negative predictive value of 100% to detect nonresponse to standard PEG-IFN-alpha-2a and ribavirin therapy at fourth week of therapy (intention-to-treat analysis). Conclusion The decision to stop the therapy in genotype 1 chronic hepatitis C patients treated with PEG-IFN-alpha-2a and ribavirin can be confidently made after 4 weeks of treatment based on the absence of baseline anti-NS4a Ab and a week-4 hepatitis C virus-RNA above 100.000 IU/ml. Eur J Gastroenterol Hepatol 22:1443-1448 (C) 2010 Wolters Kluwer Health vertical bar Lippincott Williams & Wilkins

Validation of an enzyme-linked immunosorbent assay for the quantification of human IgG directed against the repeat region of the circumsporozoite protein of the parasite Plasmodium falciparum.

BACKGROUND: Several pre-erythrocytic malaria vaccines based on the circumsporozoite protein (CSP) antigen of Plasmodium falciparum are in clinical development. Vaccine immunogenicity is commonly evaluated by the determination of anti-CSP antibody levels using IgG-based assays, but no standard assay is available to allow comparison of the different vaccines. METHODS: The validation of an anti-CSP repeat region enzyme-linked immunosorbent assay (ELISA) is described. This assay is based on the binding of serum antibodies to R32LR, a recombinant protein composed of the repeat region of P. falciparum CSP. In addition to the original recombinant R32LR, an easy to purify recombinant His-tagged R32LR protein has been constructed to be used as solid phase antigen in the assay. Also, hybridoma cell lines have been generated producing human anti-R32LR monoclonal antibodies to be used as a potential inexhaustible source of anti-CSP repeats standard, instead of a reference serum. RESULTS: The anti-CSP repeats ELISA was shown to be robust, specific and linear within the analytical range, and adequately fulfilled all validation criteria as defined in the ICH guidelines. Furthermore, the coefficient of variation for repeatability and intermediate precision did not exceed 23%. Non-interference was demonstrated for R32LR-binding sera, and the assay was shown to be stable over time. CONCLUSIONS: This ELISA, specific for antibodies directed against the CSP repeat region, can be used as a standard assay for the determination of humoral immunogenicity in the development of any CSP-based P. falciparum malaria vaccine

Monoclonal anti-envelope antibody AP33 protects humanized mice against a patient-derived hepatitis C virus challenge

End-stage liver disease caused by hepatitis C virus (HCV) infection is a major indication for liver transplantation. However, immediately after transplantation the liver graft of viremic patients universally becomes infected by circulating virus, resulting in accelerated liver disease progression. Currently available direct-acting antiviral therapies have reduced efficacy in patients with end-stage liver disease and prophylactic strategies to prevent HCV recurrence are still highly needed. In this study we compared the ability of two broadly reactive monoclonal antibodies (mAbs), designated 3/11 and AP33, recognizing a distinct but overlapping epitope in the viral E2 glycoprotein to protect humanized mice from a patient-derived HCV challenge. Their neutralizing activity was assessed using the HCVpp and HCVcc systems expressing multiple patient-derived envelopes and a human-liver chimeric mouse model. HCV RNA was readily detected in all control mice challenged with a patient-derived HCV genotype 1b isolate, while three out of four AP33-treated mice were completely protected. In contrast, only one out of four 3/11-treated mice remained HCV RNA negative throughout the observation period, while the other three had a viral load that was indistinguishable from that in the control group. The increased in vivo efficacy of AP33 was in line with its higher affinity and neutralizing capacity observed in vitro. Conclusion: Although mAbs AP33 and 3/11 target the same region in E2, only mAb AP33 can efficiently protect from challenge with a heterologous HCV population in vivo. Since mAb AP33 efficiently neutralizes viral variants that escaped the humoral immune response and re-infected the liver graft of transplant patients, it may be a valuable candidate to prevent HCV recurrence. In addition our data is valuable for the design of a prophylactic vaccine

Development and characterization of a human monoclonal antibody for prevention of HCV recurrence in liver transplant patients

More than 170 million people worldwide are chronically infected with hepatitis C virus (HCV) and are at risk of developing liver fibrosis, cirrhosis and hepatocellular carcinoma. Liver transplantation is the only option for patients with HCV-induced end-stage liver diseases. Nevertheless, infection of the newly grafted liver occurs immediately and universally after transplantation. Despite the recent progress in HCV therapy, a prophylactic vaccine is still not available. The role of neutralizing monoclonal antibodies (mAbs) in protection from different viral infections including HCV, HIV and Ebola has been reported. In the last few years, several mAbs with neutralizing activity have been described but only few mAbs have been evaluated in vivo. In the present study, we describe the development of a mAb, designated 2A5, isolated from HCV genotype 1b chronic patient. ELISA results indicated high affinity of mAb 2A5 towards HCV envelope glycoprotein (E1E2). The binding activity was completely lost against denatured E1E2 protein indicating that it targets a conformational epitope within the envelope region. Epitope mapping using alanine mutants of E1E2 proteins defined critical binding residues within the regions 419-447 and 612-617. Results of pseudoparticles (HCVpp) and cell culture produced virus (HCVcc) neutralization showed broad neutralizing activity of mAb 2A5 against all HCV genotypes. The efficacy study of mAb 2A5 in immune-deficient mice of which the liver is repopulated with human hepatocytes (humanized mice) showed complete protection from HCV challenge for genotypes 1a and 4a, while partial protection was achieved for genotypes 1b and 6a. Sequence analysis of E1E2 protein from non-protected mice did not revealed resistance mutations at interaction residues of mAb 2A5. In conclusion, mAb 2A5 shows potent anti-HCV neutralizing activity both in vitro and in vivo and could hence provide an effective strategy to prevent HCV recurrence in chronically infected HCV liver transplant patients. In addition, the broad neutralizing activity of this mAb presents a valuable epitope for the design of HCV vaccine with cross-protection activity

A novel neutralizing human monoclonal antibody broadly abrogates hepatitis C virus infection in vitro and in vivo

Infections with hepatitis C virus (HCV) represent a worldwide health burden and a prophylactic vaccine is still not available. Liver transplantation (LT) is often the only option for patients with HCV-induced end-stage liver disease. However, immediately after transplantation, the liver graft becomes infected by circulating virus, resulting in accelerated progression of liver disease. Although the effi cacy of HCV treatment using direct-acting antivirals has improved significantly, immune compromised LT-patients and patients with advanced liver disease remain difficult to treat. As an alternative approach, interfering with viral entry could prevent infection of the donor liver. We generated a human monoclonal antibody (mAb), designated 2A5, which targets the HCV envelope. The neutralizing activity of mAb 2A5 was assessed using multiple prototype and patient-derived HCV pseudoparticles (HCVpp), cell culture produced HCV (HCVcc), and a human-liver chimeric mouse model. Neutralization levels observed for mAb 2A5 were generally high and mostly superior to those obtained with AP33, a well-characterized HCV-neutralizing monoclonal antibody. Using humanized mice, complete protection was observed after genotype 1a and 4a HCV challenge, while only partial protection was achieved using gt1b and 6a isolates. Epitope mapping revealed that mAb 2A5 binding is conformation-dependent and identified the E2-region spanning amino acids 434 to 446 (epitope II) as the predominant contact domain. Conclusion : mAb 2A5 shows potent anti-HCV neutralizing activity both in vitro and in vivo and could hence represent a valuable candidate to prevent HCV recurrence in LT-patients. In addition, the detailed identification of the neutralizing epitope can be applied for the design of prophylactic HCV vaccines

A role for B cells to transmit hepatitis C virus infection

Hepatitis C virus (HCV) is highly variable and transmits through infected blood to establish a chronic liver infection in the majority of patients. Our knowledge on the infectivity of clinical HCV strains is hampered by the lack of in vitro cell culture systems that support efficient viral replication. We and others have reported that HCV can associate with and infect immune cells and may thereby evade host immune surveillance and elimination. To evaluate whether B cells play a role in HCV transmission, we assessed the ability of B cells and sera from recent (<2 years) or chronic (≥ 2 years) HCV patients to infect humanized liver chimeric mice. HCV was transmitted by B cells from chronic infected patients whereas the sera were non-infectious. In contrast, B cells from recently infected patients failed to transmit HCV to the mice, whereas all serum samples were infectious. We observed an association between circulating anti-glycoprotein E1E2 antibodies and B cell HCV transmission. Taken together, our studies provide evidence for HCV transmission by B cells, findings that have clinical implications for prophylactic and therapeutic antibody-based vaccine design

Insights From Early Clinical Trials Assessing Response to mRNA SARS-CoV-2 Vaccination in Immunocompromised Patients.

peer reviewedIt is critical to protect immunocompromised patients against COVID-19 with effective SARS-CoV-2 vaccination as they have an increased risk of developing severe disease. This is challenging, however, since effective mRNA vaccination requires the successful cooperation of several components of the innate and adaptive immune systems, both of which can be severely affected/deficient in immunocompromised people. In this article, we first review current knowledge on the immunobiology of SARS-COV-2 mRNA vaccination in animal models and in healthy humans. Next, we summarize data from early trials of SARS-COV-2 mRNA vaccination in patients with secondary or primary immunodeficiency. These early clinical trials identified common predictors of lower response to the vaccine such as anti-CD19, anti-CD20 or anti-CD38 therapies, low (naive) CD4+ T-cell counts, genetic or therapeutic Bruton tyrosine kinase deficiency, treatment with antimetabolites, CTLA4 agonists or JAK inhibitors, and vaccination with BNT162b2 versus mRNA1273 vaccine. Finally, we review the first data on third dose mRNA vaccine administration in immunocompromised patients and discuss recent strategies of temporarily holding/pausing immunosuppressive medication during vaccination

Prevalence and incidence of antibodies against SARS-CoV-2 among primary healthcare providers in Belgium during 1 year of the COVID-19 epidemic: prospective cohort study protocol.

peer reviewed[en] INTRODUCTION: National SARS-CoV-2 seroprevalence data provide essential information about population exposure to the virus and help predict the future course of the epidemic. Early cohort studies have suggested declines in levels of antibodies in individuals associated with, for example, illness severity, age and comorbidities. This protocol focuses on the seroprevalence among primary healthcare providers (PHCPs) in Belgium. PHCPs manage the vast majority of (COVID-19) patients and therefore play an essential role in the efficient organisation of healthcare. Currently, evidence is lacking on (1) how many PHCPs get infected with SARS-CoV-2 in Belgium, (2) the rate at which this happens, (3) their clinical spectrum, (4) their risk factors, (5) the effectiveness of the measures to prevent infection and (6) the accuracy of the serology-based point-of-care test (POCT) in a primary care setting. METHODS AND ANALYSIS: This study will be set up as a prospective cohort study. General practitioners (GPs) and other PHCPs (working in a GP practice) will be recruited via professional networks and professional media outlets to register online to participate. Registered GPs and other PHCPs will be asked at each testing point (n=9) to perform a capillary blood sample antibody POCT targeting IgM and IgG against the receptor-binding domain of SARS-CoV-2 and complete an online questionnaire. The primary outcomes are the prevalence and incidence of antibodies against SARS-CoV-2 in PHCPs during a 12-month follow-up period. Secondary outcomes include the longevity of antibodies against SARS-CoV-2. ETHICS AND DISSEMINATION: Ethical approval has been granted by the ethics committee of the University Hospital of Antwerp/University of Antwerp (Belgian registration number: 3002020000237). Alongside journal publications, dissemination activities include the publication of monthly reports to be shared with the participants and the general population through the publicly available website of the Belgian health authorities (Sciensano). TRIAL REGISTRATION NUMBER: NCT04779424

Development and characterization of a human monoclonal antibody targeting the N-terminal region of hepatitis C virus envelope glycoprotein E1

Monoclonal antibodies (mAbs) targeting the hepatitis C virus (HCV) envelope have been raised mainly against envelope protein 2 (E2), while the antigenic epitopes of envelope protein 1 (E1) are not fully identified. Here we describe the detailed characterization of a human mAb, designated A6, generated from an HCV genotype 1b infected patient. ELISA results showed reactivity of mAb A6 to full-length HCV E1E2 of genotypes 1a, 1b and 2a. Epitope mapping identified a region spanning amino acids 230-239 within the N-terminal region of E1 as critical for binding. Antibody binding to this epitope was not conformation dependent. Neutralization assays showed that mAb A6 lacks neutralizing capacity and does not interfere with the activity of known neutralizing antibodies. In summary, mAb A6 is an important tool to study the structure and function of E1 within the viral envelope, a crucial step in the development of an effective prophylactic HCV vaccine