Development of universal influenza vaccine candidates

Recent characterizations of cross-neutralizing monoclonal antibodies against the influenza hemagglutinin (HA) have revealed conserved regions amongst divergent influenza virus subtypes. The mapping of these epitopes shows that these regions are located in the stem or stalk region of the influenza virus hemagglutinin. Preliminary studies in Dr. Palese`s laboratory have shown that immunogens, designated as headless HA, that resemble the stem or stalk region of the hemagglutinin, are able to protect mice against lethal challenge with influenza virus. Further, the group has also shown that vaccination with a peptide derived from the long alpha helix of the HA2 subunit, which forms the central core of the stem, was able to confer protection against homologous challenge and significantly delayed onset of disease against against heterologous challenge. Herein, the proposed project aims to further developing these approaches in generating a broad spectrum influenza virus vaccine. Since the expression of the headless HA immunogen in mammalian cells gives very low yields, we will optimize the production using the baculovirus expression system in order to provide enough material to perform extensive experiments. Additionally, we will produce the immunogen in different formats, as soluble protein or as virus-like particles (VLPs) for example. The soluble protein version will be helpful for biochemical characterization of the headless HA. We also aim to improve the stability of the protein by the introduction of different mutations. The second immunogen, the influenza HA long alpha helix will be expressed as a single peptide, as triple version or as fusion protein in a bacterial expression system or in the baculoviral expression system. Fusion partners will include toll-like receptor ligands as well as conserved influenza proteins with known T-cell epitopes like the influenza nucleoprotein. Both immunogens will be tested in homologous and heterologous challenge experiments in the mouse model. Immune responses against successful vaccine candidates will be analyzed and the particular mechanism behind protection will be revealed. Finally, we will test different prime boost regimes in order to test the potential of both immunogens in the context of a pre-existing immunity in the human population. The ultimate goal of this work is the development of a broad-spectrum influenza vaccine that induces long lasting immunity.