Highly attenuated smallpox vaccine protects mice with and without immune deficiencies against pathogenic vaccinia virus challenge - PubMed (original) (raw)

Highly attenuated smallpox vaccine protects mice with and without immune deficiencies against pathogenic vaccinia virus challenge

Linda S Wyatt et al. Proc Natl Acad Sci U S A. 2004.

Abstract

Modified vaccinia virus Ankara (MVA), developed >30 years ago as a highly attenuated candidate smallpox vaccine, was recloned from a 1974 passage and evaluated for safety and immunogenicity. Replication of MVA is impaired in most mammalian cells, and we found that mice with severe combined immunodeficiency disease remained healthy when inoculated with MVA at 1,000 times the lethal dose of vaccinia virus derived from the licensed Dryvax vaccine seed. In BALB/c mice inoculated intramuscularly with MVA, virus-specific CD8+ T cells and antibodies to purified virions and membrane protein components of the intracellular and extracellular infectious forms of vaccinia virus were induced in a dose-dependent manner. After one or two inoculations of MVA, the T cell numbers and antibody titers equaled or exceeded those induced by percutaneous injection of Dryvax. Antibodies induced by MVA and Dryvax were neutralizing and inhibited virus spread in cultured cells. Furthermore, vaccinated mice were protected against lethal intranasal challenge with a pathogenic vaccinia virus. B cell-deficient mice unable to generate antibodies and beta2-microglobulin-deficient mice unable to express MHC class I molecules for a CD8+ T cell response were also protectively vaccinated by MVA. In contrast, mice with decreased CD4 or MHC class II expression and double-knockout mice deficient in MHC class I- and II-restricted activities were poorly protected or unprotected. This study confirmed the safety of MVA and demonstrated that the overlapping immune responses protected normal and partially immune-deficient animals, an encouraging result for this candidate attenuated smallpox vaccine.

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Figures

Fig. 1.

Fig. 1.

Safety of MVA. SCID mice were uninfected, infected i.p. with MVA (106 to 109 pfu) or Wyeth strain of vaccinia virus (105 to 106 pfu), or infected i.v. with 108 pfu of MVA. Mice were weighed individually, and the averages were plotted. †, died naturally or were killed because of 30% weight loss. Only mice inoculated with 106 pfu of the Wyeth strain of vaccinia virus showed persistent weight loss, disease, and death.

Fig. 2.

Fig. 2.

Anti-comet test. After BS-C-1 cells were infected with the IHD strain of vaccinia virus, pooled serum diluted 1:50 from mice immunized twice with MVA (106 to 108 pfu) or once with Dryvax was added to the liquid overlay medium. After 48 h, the monolayers were stained with crystal violet.

Fig. 3.

Fig. 3.

CD8+ T cell responses. BALB/c mice were unimmunized (controls) or immunized i.m. with MVA at 106, 107, or 108 pfu or percutaneously with Dryvax on day 0. On the indicated days, spleens were removed and mixed with P815 cells that had been stimulated overnight with MVA or WR. The percentage of vaccinia virus-specific IFN-γ expressing CD3+ CD8+ cells were determined for four animals in each group, and the values were plotted with standard deviations. On day 30, mice were challenged i.n. with 106 pfu of vaccinia virus WR.

Fig. 4.

Fig. 4.

Protective immunization of mice. (A) Groups of four BALB/c mice were inoculated i.m. with MVA (106 to 108 pfu) or percutaneously with Dryvax and challenged 4 weeks later by i.n. inoculation of 106 pfu of vaccinia virus WR. (B) BALB/c mice vaccinated with MVA as in A were revaccinated after 4 weeks and challenged 3 weeks after that. Dryvax-vaccinated mice were challenged at 7 weeks after the single vaccination. Mice were weighed individually and averages were plotted. †, Died naturally or were killed because of 30% weight loss. The mice used here were the same as those in Table 1.

Fig. 5.

Fig. 5.

Protective immunization of immune-deficient mice. Mice (n = 4) were vaccinated once i.m. with 108 pfu of MVA and challenged 3 weeks later with 106 pfu of vaccinia virus WR by the i.n. route. (A) BALB/c mice. (B) B cell-deficient mice. (C) C57BL/6 mice. (D) β2-microglobulin-deficient mice. (E) CD4-deficient mice. (F) MHC class II-deficient mice. (G) Double-knockout mice deficient in MHC class I and II. Inset numbers represent the IMV reciprocal endpoint ELISA titers at 3 weeks.

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