Investigating the interplay of protein expression and innate immunity on nucleic acid vaccine potency
Author(s)
Stirling, David
Type
Thesis
Abstract
After decades of research the COVID-19 pandemic accelerated the development of the nucleic acid vaccine platform and led to the first human RNA and DNA vaccines. However, despite their potential, the relatively weak immunogenicity of many nucleic acid vaccines has hindered their progress towards clinical application. To address the shortcomings of nucleic acid vaccines, this thesis focuses on unravelling the relationship between vaccine design, innate immune responses, and subsequent vaccine efficacy.
Using the systemic cytokine response as an indicator of innate stimulation, the acute immune response to nucleic acid vaccines were measured in vaccine challenge mouse models. Key cytokines were linked to the establishment of robust specific antibody responses by an influenza self amplifying RNA vaccine. Inflammation from lipid nanoparticle formulations was linked to improved outcomes compared to less inflammatory formulations. RIG I like receptor (RLR)signals were linked to immune dampening. Strategies to avoid RLR activation, namely inclusion of modified nucleotides and 2’O methylation of the 5’ cap, were linked to improved vaccine potency.
The protein expression of DNA vaccines was measured by in vitro flow cytometry assays and immunogenicity determined using mouse studies of immunisation. DNA vaccine immune responses to influenza were increased by the inclusion of regulatory sequences that drive protein expression but hindered by the inclusion of CpG motifs. Different approaches to modify the innate immune response to DNA vaccines were tested. CpG oligodeoxynucleotide stimulated a systemic cytokine response but did not measurably impact protein expression or vaccine immunogenicity. Delivery of immunomodulatory proteins that restrict DNA-PK or STING pathways were not found to impact protein expression from vaccines. However, drug-mediated blocking of the AIM2 inflammasome dampened cytokine release from transfected cells and increased vaccine induced protein expression...
Using the systemic cytokine response as an indicator of innate stimulation, the acute immune response to nucleic acid vaccines were measured in vaccine challenge mouse models. Key cytokines were linked to the establishment of robust specific antibody responses by an influenza self amplifying RNA vaccine. Inflammation from lipid nanoparticle formulations was linked to improved outcomes compared to less inflammatory formulations. RIG I like receptor (RLR)signals were linked to immune dampening. Strategies to avoid RLR activation, namely inclusion of modified nucleotides and 2’O methylation of the 5’ cap, were linked to improved vaccine potency.
The protein expression of DNA vaccines was measured by in vitro flow cytometry assays and immunogenicity determined using mouse studies of immunisation. DNA vaccine immune responses to influenza were increased by the inclusion of regulatory sequences that drive protein expression but hindered by the inclusion of CpG motifs. Different approaches to modify the innate immune response to DNA vaccines were tested. CpG oligodeoxynucleotide stimulated a systemic cytokine response but did not measurably impact protein expression or vaccine immunogenicity. Delivery of immunomodulatory proteins that restrict DNA-PK or STING pathways were not found to impact protein expression from vaccines. However, drug-mediated blocking of the AIM2 inflammasome dampened cytokine release from transfected cells and increased vaccine induced protein expression...
Version
Open Access
Date Issued
2023-11-30
Date Awarded
01/06/2024
License URL
Advisor
Tregoning, John
Caproni, Lisa
Sponsor
Medical Research Council (Great Britain)
Publisher Department
Department of Infectious Disease
Publisher Institution
Imperial College London
Qualification Level
Doctoral
Qualification Name
Doctor of Philosophy (PhD)