Preclinical Evaluation of a Replication-Deficient Intranasal ΔNS1 H5N1 Influenza Vaccine

Abstract

BackgroundWe developed a novel intranasal influenza vaccine approach that is based on the construction of replication-deficient vaccine viruses that lack the entire NS1 gene (ΔNS1 virus). We previously showed that these viruses undergo abortive replication in the respiratory tract of animals. The local release of type I interferons and other cytokines and chemokines in the upper respiratory tract may have a “self-adjuvant effect”, in turn increasing vaccine immunogenicity. As a result, ΔNS1 viruses elicit strong B- and T- cell mediated immune responses.Methodology/Principal FindingsWe applied this technology to the development of a pandemic H5N1 vaccine candidate. The vaccine virus was constructed by reverse genetics in Vero cells, as a 5∶3 reassortant, encoding four proteins HA, NA, M1, and M2 of the A/Vietnam/1203/04 virus while the remaining genes were derived from IVR-116. The HA cleavage site was modified in a trypsin dependent manner, serving as the second attenuation factor in addition to the deleted NS1 gene. The vaccine candidate was able to grow in the Vero cells that were cultivated in a serum free medium to titers exceeding 8 log10 TCID50/ml. The vaccine virus was replication deficient in interferon competent cells and did not lead to viral shedding in the vaccinated animals. The studies performed in three animal models confirmed the safety and immunogenicity of the vaccine. Intranasal immunization protected ferrets and mice from being infected with influenza H5 viruses of different clades. In a primate model (Macaca mulatta), one dose of vaccine delivered intranasally was sufficient for the induction of antibodies against homologous A/Vietnam/1203/04 and heterologous A/Indonesia/5/05 H5N1 strains.Conclusion/SignificanceOur findings show that intranasal immunization with the replication deficient H5N1 ΔNS1 vaccine candidate is sufficient to induce a protective immune response against H5N1 viruses. This approach might be attractive as an alternative to conventional influenza vaccines. Clinical evaluation of ΔNS1 pandemic and seasonal influenza vaccine candidates are currently in progress.

Response of BALB/c mice to a monovalent influenza A (H1N1) 2009 split vaccine

“…As for influenza-vaccine production, a high virus yield is a critical factor because it can reduce cost and sustain longer immune protection periods by providing sufficient antigens. 31,32 Importantly, the influenza (H1N1) split vaccine produced a satisfactory effect against influenza-virus infection at the lowest antigen dose tested (the vaccine at a dose of 7.5 mg with Al(OH) 3 and 15 mg of antigen alone were the optimal conditions). The promising results produced in this study concerning the safety and immunogenicity of the influenza H1N1 split vaccine support their careful evaluation in phase-1 clinical trials in humans.…”

Antimycotic-Antibiotic Amphotericin B Promotes Influenza Virus Replication in Cell Culture

“…All ⌬NS1 reassortant viruses were obtained by reverse genetics solely on Vero cells (10,14,27,34). The internal segments of influenza A viruses were derived from a modified IVR-116 vaccine strain (WHO) containing a deletion of the NS1 gene.…”

Mutations in the M-Gene Segment Can Substantially Increase Replication Efficiency of NS1 Deletion Influenza A Virus in MDCK Cells

“…These findings contradict the previous suggestion that the restricted replication of this virus in MDCK cells is primarily due to the inability to inhibit the IFN response. The mutations described may have direct applications, as they, for example, allow the development of delNS1-based viruses with improved replication efficiency, thereby making it possible to produce such a virus in cell lines other than Vero cells (35) or NS1-expressing MDCK cells (46). Moreover, these mutations may be combined with the G3A and C8U mutations in HA vRNA, which increased the HA expression level of a live, attenuated, NS1-truncated influenza vaccine strain (25).…”

“…The local release of IFN and other cytokines and chemokines appears to be an excellent adjuvant that enhances production of immunoglobulins and contributes to the activation of dendritic cells required for antigen presentation (22,31). DelNS1 candidate vaccines against influenza A and B viruses have been developed (35,51), and initial trials in humans showed successful induction of antibody responses (48). Apart from the use in vaccines, delNS1 viruses also show potential as oncolytic agents (27) and viral expression vectors (50).…”