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Evaluation of replication, immunogenicity and protective efficacy of a live attenuated cold-adapted pandemic H1N1 influenza virus vaccine in non-human primates.

ABSTRACT: We studied the replication of influenza A/California/07/09 (H1N1) wild type (CA09wt) virus in two non-human primate species and used one of these models to evaluate the immunogenicity and protective efficacy of a live attenuated cold-adapted vaccine, which contains the hemagglutinin and neuraminidase from the H1N1 wild type (wt) virus and six internal protein gene segments of the A/Ann Arbor/6/60 cold-adapted (ca) master donor virus. We infected African green monkeys (AGMs) and rhesus macaques with 2×10(6) TCID(50) of CA09wt and CA09ca influenza viruses. The virus CA09wt replicated in the upper respiratory tract of all animals but the titers in upper respiratory tract tissues of rhesus macaques were significant higher than in AGMs (mean peak titers 10(4.5) TCID(50)/g and 10(2.0) TCID(50)/g on days 4 and 2 post-infection, respectively; p<0.01). Virus replication was observed in the lungs of all rhesus macaques (10(2.0)-10(5.4) TCID(50)/g) whereas only 2 out of 4 AGMs had virus recovered from the lungs (10(2.5)-10(3.5) TCID(50)/g). The CA09ca vaccine virus was attenuated and highly restricted in replication in both AGMs and rhesus macaques. We evaluated the immunogenicity and protective efficacy of the CA09ca vaccine in rhesus macaques because CA09wt virus replicated more efficiently in this species. One or two doses of vaccine were administered intranasally and intratracheally to rhesus macaques. For the two-dose group, the vaccine was administered 4-weeks apart. Immunogenicity was assessed by measuring hemagglutination-inhibiting (HAI) antibodies in the serum and specific IgA antibodies to CA09wt virus in the nasal wash. One or two doses of the vaccine elicited a significant rise in HAI titers (range 40-320). Two doses of CA09ca elicited higher pH1N1-specific IgA titers than in the mock-immunized group (p<0.01). Vaccine efficacy was assessed by comparing titers of CA09wt challenge virus in the respiratory tract of mock-immunized and CA09ca vaccinated monkeys. Significantly lower virus titers were observed in the lungs of vaccinated animals than mock-immunized animals (p?0.01). Our results demonstrate that AGMs and rhesus macaques support the replication of pandemic H1N1 influenza virus to different degrees and a cold-adapted pH1N1 vaccine elicits protective immunity against pH1N1 virus infection in rhesus macaques.

SUBMITTER: Boonnak K

PROVIDER: S-EPMC3412900 | biostudies-literature | 2012 Aug

REPOSITORIES: biostudies-literature

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Evaluation of replication, immunogenicity and protective efficacy of a live attenuated cold-adapted pandemic H1N1 influenza virus vaccine in non-human primates.

Boonnak Kobporn K Paskel Myeisha M Matsuoka Yumiko Y Vogel Leatrice L Subbarao Kanta K

Vaccine 20120710 38

We studied the replication of influenza A/California/07/09 (H1N1) wild type (CA09wt) virus in two non-human primate species and used one of these models to evaluate the immunogenicity and protective efficacy of a live attenuated cold-adapted vaccine, which contains the hemagglutinin and neuraminidase from the H1N1 wild type (wt) virus and six internal protein gene segments of the A/Ann Arbor/6/60 cold-adapted (ca) master donor virus. We infected African green monkeys (AGMs) and rhesus macaques w ...[more]

PMID: 22789506

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Project description:The only licensed live attenuated influenza A virus vaccines (LAIVs) in the United States (FluMist) are created using internal protein-coding gene segments from the cold-adapted temperature-sensitive master donor virus A/Ann Arbor/6/1960 and HA/NA gene segments from circulating viruses. During serial passage of A/Ann Arbor/6/1960 at low temperatures to select the desired attenuating phenotypes, multiple cold-adaptive mutations and temperature-sensitive mutations arose. A substantial amount of scientific and clinical evidence has proven that FluMist is safe and effective. Nevertheless, no study has been conducted specifically to determine if the attenuating temperature-sensitive phenotype can revert and, if so, the types of substitutions that will emerge (i.e., compensatory substitutions versus reversion of existing attenuating mutations). Serial passage of the monovalent FluMist 2009 H1N1 pandemic vaccine at increasing temperatures in vitro generated a variant that replicated efficiently at higher temperatures. Sequencing of the variant identified seven nonsynonymous mutations, PB1-E51K, PB1-I171V, PA-N350K, PA-L366I, NP-N125Y, NP-V186I, and NS2-G63E. None occurred at positions previously reported to affect the temperature sensitivity of influenza A viruses. Synthetic genomics technology was used to synthesize the whole genome of the virus, and the roles of individual mutations were characterized by assessing their effects on RNA polymerase activity and virus replication kinetics at various temperatures. The revertant also regained virulence and caused significant disease in mice, with severity comparable to that caused by a wild-type 2009 H1N1 pandemic virus.The live attenuated influenza vaccine FluMist has been proven safe and effective and is widely used in the United States. The phenotype and genotype of the vaccine virus are believed to be very stable, and mutants that cause disease in animals or humans have never been reported. By propagating the virus under well-controlled laboratory conditions, we found that the FluMist vaccine backbone could regain virulence to cause severe disease in mice. The identification of the responsible substitutions and elucidation of the underlying mechanisms provide unique insights into the attenuation of influenza virus, which is important to basic research on vaccines, attenuation reversion, and replication. In addition, this study suggests that the safety of LAIVs should be closely monitored after mass vaccination and that novel strategies to continue to improve LAIV vaccine safety should be investigated.

Project description:BACKGROUND:Development of live attenuated influenza vaccines (LAIV) against avian viruses with pandemic potential is an important public health strategy. METHODS AND FINDINGS:We performed open-label trials to evaluate the safety, infectivity, and immunogenicity of H5N1 VN 2004 AA ca and H5N1 HK 2003 AA ca. Each of these vaccines contains a modified H5 hemagglutinin and unmodified N1 neuraminidase from the respective wild-type (wt) parent virus and the six internal protein gene segments of the A/Ann Arbor/6/60 cold-adapted (ca) master donor virus. The H5N1 VN 2004 AA ca vaccine virus was evaluated at dosages of 10(6.7) TCID(50) and 10(7.5) TCID(50), and the H5N1 HK 2003 AA ca vaccine was evaluated at a dosage of 10(7.5) TCID(50). Two doses were administered intranasally to healthy adults in isolation at 4-8 week intervals. Vaccine safety was assessed through daily examinations and infectivity was assessed by viral culture and by realtime reverse transcription-polymerase chain reaction testing of nasal wash (NW) specimens. Immunogenicity was assessed by measuring hemagglutination-inhibition (HI) antibodies, neutralizing antibodies, and IgG or IgA antibodies to recombinant (r)H5 VN 2004 hemagglutinin (HA) in serum or NW. Fifty-nine participants were enrolled: 21 received 10(6.7) TCID(50) and 21 received 10(7.5) TCID(50) of H5N1 VN 2004 AA ca and 17 received H5N1 HK 2003 AA ca. Shedding of vaccine virus was minimal, as were HI and neutralizing antibody responses. Fifty-two percent of recipients of 10(7.5) TCID(50) of H5N1 VN 2004 AA ca developed a serum IgA response to rH5 VN 2004 HA. CONCLUSIONS:The live attenuated H5N1 VN 2004 and HK 2003 AA ca vaccines bearing avian H5 HA antigens were very restricted in replication and were more attenuated than seasonal LAIV bearing human H1, H3 or B HA antigens. The H5N1 AA ca LAIV elicited serum ELISA antibody but not HI or neutralizing antibody responses in healthy adults. (ClinicalTrials.gov Identifiers: NCT00347672 and NCT00488046).

Dataset Information

Evaluation of replication, immunogenicity and protective efficacy of a live attenuated cold-adapted pandemic H1N1 influenza virus vaccine in non-human primates.

Publications

Evaluation of replication, immunogenicity and protective efficacy of a live attenuated cold-adapted pandemic H1N1 influenza virus vaccine in non-human primates.

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