Intramuscular prime/intranasal boost vaccination to induce sterilizing immunity against influenza A virus infection

The most commonly used influenza vaccines are made from inactivated viruses and are administered via the intramuscular route. Although these vaccines can protect from severe lower respiratory tract disease, they do not completely prevent virus replication in the upper respiratory tract, and this may lead to virus excretion and dissemination. Therefore, nasally administered live-attenuated influenza vaccines (LAIV) that induce mucosal immunity have been developed, but finding an optimal balance between sufficient attenuation and immunogenicity remained challenging. These problems apply to both human and swine influenza vaccines. We have recently developed an LAIV candidate based on the 2009 pandemic H1N1 virus which encodes a truncated NS1 protein and lacks PA-X protein expression (NS1(1-126)-{Delta}PAX). This virus showed a blunted replication and elicited a strong innate immune response. In the present study, we took advantage of the pig animal model to evaluate this vaccine candidate in vivo and to identify a strategy for its improvement. Nasal infection of pigs with the NS1(1-126)-{Delta}PAX LAIV candidate did not cause disease but was associated with prolonged virus shedding from the upper respiratory tract. To increase safety of the vaccine candidate, we developed a novel prime/boost vaccination strategy consisting of a haemagglutinin-encoding propagation-defective vesicular stomatitis virus replicon vaccine for primary immunization via the intramuscular route, and the NS1(1-126)-{Delta}PAX LAIV for secondary immunization via the nasal route. This immunization strategy significantly reduced LAIV shedding, increased the production of specific serum IgG, neutralizing antibodies, Th1 memory cells, and induced virus-specific mucosal IgG and IgA. Of particular note, the immune response induced by this vaccination strategy completely blocked replication of the homologous challenge virus in the respiratory tract, indicating that sterilizing immunity was achieved. In summary, our novel intramuscular prime/intranasal boost vaccine combines the features of high efficacy and safety which are urgently needed to combat influenza epidemics and pandemics. Author summaryInactivated influenza vaccines which are administered intramuscularly are safe but offer only limited protection. In addition, they do not adequately prevent virus transmission by infected individuals. On the other hand, nasally administered live-attenuated influenza vaccines induce a mucosal immune response, which can effectively prevent primary infection and virus excretion. However, live-attenuated vaccines might not be sufficiently immunogenic if they are too attenuated or they trigger a robust immune response but are still too virulent. To overcome this challenge, we have developed a novel prime/boost vaccination strategy consisting of an initial intramuscular immunization with a propagation-defective RNA virus vector and a subsequent nasal immunization with a modified influenza virus that has lost its ability to counteract the hosts innate immune response. Using the pig model, we demonstrate that this approach elicited a more robust immune response both systemically and at mucosal surfaces. Importantly, replication of the vaccine virus in the respiratory tract was reduced, and challenge virus remained undetectable. In summary, our innovative vaccine, which combines intramuscular and intranasal routes of application, demonstrates high efficacy and safety and represents a valuable tool to control influenza epidemics and pandemics.