T- and B-cell responses to multivalent prime-boost DNA and viral vectored vaccine combinations against hepatitis C virus in non-human primates.

Immune responses against multiple epitopes are required for the prevention of hepatitis C virus (HCV) infection, and the progression to phase I trials of candidates may be guided by comparative immunogenicity studies in non-human primates. Four vectors, DNA, SFV, human serotype 5 adenovirus (HuAd5) and Modified Vaccinia Ankara (MVA) poxvirus, all expressing hepatitis C virus Core, E1, E2 and NS3, were combined in three prime-boost regimen, and their ability to elicit immune responses against HCV antigens in rhesus macaques was explored and compared. All combinations induced specific T-cell immune responses, including high IFN-γ production. The group immunized with the SFV+MVA regimen elicited higher E2-specific responses as compared with the two other modalities, while animals receiving HuAd5 injections elicited lower IL-4 responses as compared with those receiving MVA. The IFN-γ responses to NS3 were remarkably similar between groups. Only the adenovirus induced envelope-specific antibody responses, but these failed to show neutralizing activity. Therefore, the two novel regimens failed to induce superior responses as compared with already existing HCV vaccine candidates. Differences were found in response to envelope proteins, but the relevance of these remain uncertain given the surprisingly poor correlation with immunogenicity data in chimpanzees, underlining the difficulty to predict efficacy from immunology studies.This work was supported by European Union contract QLK2-CT-1999- 00356, by the Biomedical Primate Research Centre, The Netherlands, and by the Swedish Research Council. We are grateful to Alexander van den Berg for technical assistance with the ICS, to our colleagues from Animal Science Department for technical assistance and expert care of the macaques, to the participants of the European HCVacc Cluster who provided help and support, and to Thomas Darton (Oxford Vaccine Group, UK) for input and advice on the manuscript. Christine Rollier is an Oxford Martin fellow and a Jenner Insitute Investigator.This is the author accepted manuscript. The final version is available from Nature Publishing Group at https://doi.org/10.1038/gt.2016.55

Reduction of cPLA2alpha overexpression: an efficient anti-inflammatory therapy for collagen-induced arthritis.

Cytosolic phospholipase A2alpha (cPLA2) plays an important role in the development of several inflammatory diseases. The aim of the present study is to determine whether inhibition of cPLA2 expression, using specific antisense oligonucleotides against cPLA2 (antisense), is efficient in reducing inflammation after its development. Two mouse models of inflammation were included in the study: thioglicolate peritonitis and collagen-induced arthritis (CIA). The antisense was found to be specific and efficient in inhibiting cPLA2 expression and NADPH oxidase activity ex vivo in peritoneal phagocytes. Immunoblotting and immunohistochemistry analysis showed a significant elevation in cPLA2 expression in the inflamed joints of collagen-induced arthritis mice localized in cell infiltrate, chondrocytes and the surrounding skin and skeletal muscle. Similarly, the cPLA2 metabolite, leukotriene B4, accumulated in the peritoneal cavity of mice with peritonitis. Inhibition of elevated cPLA2 expression after development of inflammation by intravenous administration of antisense resulted in a dramatic reduction in inflammation and a significant reduction in neutrophils recruitment to the site of inflammation in both mouse models of inflammation. Our results demonstrate the critical role of cPLA2 for the duration of inflammation and suggest that inhibition of cPLA2 expression by antisense oligonucleotides may serve as an efficient treatment of inflammatory diseases

MONOCLONAL ANTIBODIES AND RECOMBINANT PROTEINS OF FILOVIRUSES: IMMUNOCHEMICAL PROPERTIES AND EVALUATION OF THEIR EFFICIENCY FOR IMMUNE DIAGNOSTICS OF MARBURG AND EBOLA VIRUSES

A panel of monoclonal antibodies (MAbs) against VP35, VP40 and NP viral proteins of Marburg and Ebola viruses, as well as recombinant VP35, VP40 and NP proteins were generated and tested for their capacity to specific immune reactions. Monoclonal antibodies to appropriate viral proteins effectively recognized the VP35, VP40 and NP recombinant proteins, thus allowing to develop a variant of a MAb-based ELISA analysis with different types of biotin-labeled MAbs, using these antibodies for capturing viral and recombinant antigens of Marburg and Ebola viruses. These techniques were able to detect viral and recombinant proteins in a concentration range between 1 and 150 ng/ml. We conclude that the recombinant VP35, VP40 and NP proteins of filoviruses, as well as MAbs against these viral proteins represent a promising tool for a new generation of immunodiagnostic kits and studying immunological features of filovirus infection

Obtaining and Immunologic Characterization of Filoviruses Recombinant Proteins VP40 and NP

Full-size genes coding matrix proteins VP40 of Ebola virus (VE) and Marburg virus (VM) and nucleoprotein (NP) of VE were cloned. These genes expression in Escherichia coli and purification of their products by affinity chromatography using Ni-NTA agarose permitted to obtain full-size recombinant VP40 and NP proteins of filoviruses. Immunological studies showed the recombinant polypeptides to possess antigenic properties similar to those of the native viral proteins. These proteins can be used for constructing of experimental vaccines against filoviruses and improving immunodiagnostic tests against filoviral infections