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Date of Award

Fall 12-13-2021

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Biomedical Sciences

First Advisor

Sang-Moo Kang

Second Advisor

Timothy Denning

Third Advisor

Baozhong Wang

Abstract

Chapter1: Broad cross-protection by recombinant live attenuated influenza H3N2 seasonal virus expressing conserved M2 extracellular domain in a chimeric hemagglutinin.

The objective of project 1 was to determine whether intranasal immunization with live recombinant H3N2 virus expressing chimeric 4xM2e-HA would induce broadly cross-protective immunity against different subtypes of influenza A viruses in a mouse model. In recent years, antigenic drifts have severely limited the effectiveness of the H3N2 component of seasonal influenza vaccines. Here, using the reverse genetic (rg) technique, we generated reassortant seasonal influenza rgH3N2 4xM2e virus containing chimeric 4xM2e-HA in which the HA and NA genes were derived from A/Switzerland/9715293/2013 (H3N2) and the remaining 6 genes from the A/PR8 backbone. Reassortant rgH3N2 4xM2e virus containing chimeric 4xM2e-HA was found to retain comparable growth properties but displayed highly attenuated phenotypes in mice. Cross-protective efficacy against different subtypes (H1N1, H3N2, H5N1, H7N9, H9N2) of influenza A virus was tested in intranasally immunized BALB/c mice with rgH3N2 4xM2e. This study implicates a strategy of improving cross-protection by utilizing currently licensed recombinant influenza vaccine platforms.

Chapter2: Enhanced cross-protection by hetero prime-boost vaccination with recombinant influenza viruses containing chimeric hemagglutinin-M2e epitopes.

The goal of project 2 was to test whether a strategy of hetero prime-boost vaccination with recombinant influenza viruses expressing chimeric 4xM2e-HA would induce more effective cross-protection than homologous prime-boost vaccination. The impact of heterosubtypic vaccination with recombinant 4xM2e-HA influenza virus vaccines and pre-existing immunity on cross-protection against influenza viruses remains unknown. In this study, I investigated the efficacy of cross-protection by heterosubtypic prime- boost vaccination with live recombinant 4xM2e-HA H1N1 and H3N2 influenza virus vaccines in C57BL/6 mice known to be a low responder to immune-subdominant conserved epitopes. The experimental outcomes of project 2 demonstrated that hetero prime-boost strategies using recombinant 4xM2e-HA influenza virus vaccination induced more effective cross-protection against antigenically different viruses than homologous repeat vaccination in C57BL/6 mice. The roles of M2e and stalk immunity in conferring cross-protection were explored and discussed in this study.

Chapter 3: Hemagglutinin virus-like particle is immunogenic and provides heterologous protection against influenza virus in young adult and aged mice.

The goal of project 3 was to investigate immune responses and homo and cross-protective efficacy in aged mice after vaccination with a platform of virus-like particles (VLP) presenting H1 HA with and without molecularly anchored cytokine adjuvants incorporated, in comparison with those in young adult mice. Vaccine effectiveness is inferior in the aged population at high risk of severe illness from influenza virus infection. For the elderly, safe and highly immunogenic vaccines need to be developed. In project 3, host immune responses and homo and cross-protective efficacy was determined, after vaccination with a VLP vaccine platform which expresses H1 HA from A/PR8/34 (PR8 HA VLP) in young adults and aged (18-month-old) BALB/c mice. In addition, I investigated the adjuvant impact of cytokines (GM-CSF and IL-12) engineered to be incorporated into HA VLP vaccines on inducing IgG antibodies, hemagglutination inhibition titers, and homo and hetero protection in aged mice, compared to those in young adult mice. Higher doses of vaccination with H1 HA VLP and cytokines incorporated onto H1 HA VLP were found to be more effective in inducing protective immunity against homo and hetero viruses in aged mice.

DOI

https://doi.org/10.57709/26827412

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