The rapid spread and the transmission to humans of avian influenza virus (H5N1) has induced worldwide fears of a new pandemic and raised concerns over the ability of standard influenza vaccine production methods to rapidly supply sufficient amounts of an effective vaccine. We report here on a robust and flexible strategy which uses wild-type virus grown in a continuous cell culture (Vero) system to produce an inactivated whole virus vaccine. Candidate vaccines based on clade 1 and clade 2 influenza H5N1 strains were developed and demonstrated to be highly immunogenic in animal models. The vaccines induce cross-neutralising antibodies, highly cross-reactive T-cell responses and are protective in a mouse challenge model not only against the homologous virus but against other H5N1 strains, including those from another clade. These data indicate that cell culture-grown, whole virus vaccines, based on the wild-type virus, allow the rapid high yield production of a candidate pandemic vaccine.

The recent emergence and rapid spread of a novel swine-derived H1N1 influenza virus has resulted in the first influenza pandemic of this century. Monovalent vaccines have undergone preclinical and clinical development prior to initiation of mass immunization campaigns. We have carried out a series of immunogenicity and protection studies following active immunization of mice, which indicate that a whole virus, nonadjuvanted vaccine is immunogenic at low doses and protects against live virus challenge. The immunogenicity in this model was comparable to that of a whole virus H5N1 vaccine, which had previously been demonstrated to induce high levels of seroprotection in clinical studies. The efficacy of the H1N1 pandemic vaccine in protecting against live virus challenge was also seen to be equivalent to that of the H5N1 vaccine. The protective efficacy of the H1N1 vaccine was also confirmed using a severe combined immunodeficient (SCID) mouse model. It was demonstrated that mouse and guinea pig immune sera elicited following active H1N1 vaccination resulted in 100% protection of SCID mice following passive transfer of immune sera and lethal challenge. The immune responses to a whole virus pandemic H1N1 and a split seasonal H1N1 vaccine were also compared in this study. It was demonstrated that the whole virus vaccine induced a balanced Th-1 and Th-2 response in mice, whereas the split vaccine induced mainly a Th-2 response and only minimal levels of Th-1 responses. These data supported the initiation of clinical studies with the same low doses of whole virus vaccine that had previously been demonstrated to be immunogenic in clinical studies with a whole virus H5N1 vaccine.

This work demonstrates the feasibility of producing a H-IVIG preparation at large scale relatively rapidly, with a significant enrichment in antibodies to the H1N1 influenza, achieved by donor self-identification.

BackgroundThe development of novel influenza vaccines inducing a broad immune response is an important objective. The aim of this study was to evaluate live vaccines which induce both strong humoral and cell-mediated immune responses against the novel human pandemic H1N1 influenza virus, and to show protection in a lethal animal challenge model.Methodology/Principal FindingsFor this purpose, the hemagglutinin (HA) and neuraminidase (NA) genes of the influenza A/California/07/2009 (H1N1) strain (CA/07) were inserted into the replication-deficient modified vaccinia Ankara (MVA) virus - a safe poxviral live vector – resulting in MVA-H1-Ca and MVA-N1-Ca vectors. These live vaccines, together with an inactivated whole virus vaccine, were assessed in a lung infection model using immune competent Balb/c mice, and in a lethal challenge model using severe combined immunodeficient (SCID) mice after passive serum transfer from immunized mice. Balb/c mice vaccinated with the MVA-H1-Ca virus or the inactivated vaccine were fully protected from lung infection after challenge with the influenza H1N1 wild-type strain, while the neuraminidase virus MVA-N1-Ca induced only partial protection. The live vaccines were already protective after a single dose and induced substantial amounts of neutralizing antibodies and of interferon-γ-secreting (IFN-γ) CD4- and CD8 T-cells in lungs and spleens. In the lungs, a rapid increase of HA-specific CD4- and CD8 T cells was observed in vaccinated mice shortly after challenge with influenza swine flu virus, which probably contributes to the strong inhibition of pulmonary viral replication observed. In addition, passive transfer of antisera raised in MVA-H1-Ca vaccinated immune-competent mice protected SCID mice from lethal challenge with the CA/07 wild-type virus.Conclusions/SignificanceThe non-replicating MVA-based H1N1 live vaccines induce a broad protective immune response and are promising vaccine candidates for pandemic influenza.

BackgroundConvalescent plasma and fractionated immunoglobulins have been suggested as prophylactic or therapeutic interventions during an influenza pandemic.FindingsIntravenous immunoglobulin (IVIG) preparations manufactured from human plasma collected before the 2009 H1N1 influenza pandemic, and post-pandemic hyperimmune (H)-IVIG preparations were characterized with respect to hemagglutination inhibition (HI), microneutralization (MN) and neuraminidase-inhibiting (NAi) antibody titers against pandemic H1N1 (pH1N1) and seasonal H1N1 (sH1N1) viruses. The protective efficacy of the IVIG and H-IVIG preparations was evaluated in a SCID mouse challenge model.Substantial levels of HI, MN and NAi antibodies against pH1N1 (GMTs 1:45, 1:204 and 1: 727, respectively) and sH1N1 (GMTs 1:688, 1:4,946 and 1:312, respectively) were present in pre-pandemic IVIG preparations. In post-pandemic H-IVIG preparations, HI, MN and NAi antibody GMTs against pH1N1 were 1:1,280, 1:11,404 and 1:2,488 (28-, 56- and 3.4-fold enriched), respectively, compared to pre-pandemic IVIG preparations (p < 0.001). Post-pandemic H-IVIG (HI titer 1:1,280) provided complete protection from lethality of SCID mice against pH1N1 challenge (100% of mice survived for 29 days post-challenge). Pre-pandemic IVIG (HI titer 1:70) did not provide significant protection against pH1N1 challenge (50% of mice survived 29 days post-challenge compared to 40% survival in the buffer control group). There was a highly significant correlation between circulating in vivo HI and MN antibody titers and survival (p < 0001).ConclusionThe substantial enrichment of HA- and NA-specific antibodies in H-IVIG and the efficacious protection of SCID mice against challenge with pH1N1 suggests H-IVIG as a promising intervention against pandemic influenza for immunocompromised patients and other risk groups.