Date of Award

5-2011

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Program

Biomedical Sciences

Track

Microbial Pathogenesis, Immunology, and Inflammation

Research Advisor

Richard J. Webby, Ph.D.

Committee

Julia L. Hurwitz, Ph.D. Tony Marion, Ph.D. Mark Y. Sangster, Ph.D. Michael Whitt, Ph.D.

Keywords

Adjuvants, H5N1, Influenza, MF59, Vaccines

Abstract

Human influenza pandemics occur when influenza viruses to which the population has little or no immunity emerge and acquire the ability to transmit among humans. Since their emergence in 1996, human infections with highly pathogenic avian influenza A (H5N1) viruses presented a serious public health challenge. Additionally, H5N1 viruses caused significant agricultural and economic losses in the communities it has affected. Human infections with these viruses are rare but when they occur, these infections are highly fatal. A greater public health concern stems from the rapid evolution displayed by these viruses so far, which in turn might result in viruses able to cause sustained and widespread human‑to‑human transmission.

The development of H5N1 vaccines that can induce protective antibody responses is the cornerstone of the global efforts to address this pandemic threat. However, it was repeatedly shown in clinical trials that (at comparable antigen doses) candidate human H5N1 influenza vaccines generally elicit lower immune responses than seasonal human influenza vaccines. In addition, the evolution of H5N1 viruses into at least 10 antigenically distinct clades and multiple subclades suggests that an optimal H5N1 vaccine should confer cross-reactivity against H5N1 viruses from other clades. Therefore, the WHO has recommended the use of adjuvants especially the oil‑in‑water emulsions (such as MF59) in combination with H5N1 split or subunit vaccines. While the ability of these adjuvants to generally boost the vaccine-specific antibody titers has been well documented, the question of how adjuvants modulate the quality of such responses remains largely unanswered.

First, we studied the impact of two adjuvants that are licensed with human influenza vaccines, MF59 and alum on the kinetics of developing protective antibody responses to subunit H5N1 vaccines in the ferret model. With a single immunization regimen, we found that including adjuvants in the vaccine formulation was essential for protection against a lethal H5N1 virus challenge. Adjuvanted vaccines provided protection against lethality when administered as early as 7 days prior to challenge and protection against challenge-associated morbidity when administered 14 days or longer prior to challenge.

We also examined the breadth of the antibody responses to adjuvanted vs. unadjuvanted H5N1 vaccines. Previous studies have suggested that the oil‑in‑water emulsion adjuvanted vaccines did not only elicit higher antibody titers against homologous H5N1 strains, but also against representative isolates of different clades. Our data clearly showed that indeed MF59‑adjuvanted H5N1 vaccines elicited a quantitatively greater H5‑specific antibody response than the alum‑adjuvanted or the unadjuvanted vaccine. However, for the most part, the specificity of these antibodies as determined by binding to H5 antigen microarray and competitive ELISA assays was not different than induced by alum or vaccine alone.

Finally, we tested the contribution of the cytosolic innate immune sensing complex known as the NLRP3 (also known as cryopyrin, CIAS1, or Nalp3) inflammasome is in the adjuvant effect of MF59. It was recently shown that activation of the NLRP3 inflammasome is essential for the adjuvant effect of alum. Our data clearly demonstrated that while the NLRP3 is dispensable for the antibody responses to MF59‑adjuvanted H5N1 vaccines, ablation of the adapter molecule ASC abrogated these responses.

DOI

10.21007/etd.cghs.2011.0082

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