Sterility and immunogenicity of gamma-irradiated respiratory viruses

Abstract

Gamma (γ)-radiation is a method commonly applied to sterilise pathogens in the biomedical, food and pharmaceutical industries. γ-radiation inactivates pathogens by causing irreparable damage to genomes to prevent replication. However, proteins and other antigenic structures are left mostly intact. In recent years there has been increasing advocacy for highly immunogenic g-irradiated vaccines, several of which are currently in clinical or pre-clinical trials. Importantly, various methods of mathematical modelling and sterility testing are employed to ensure the safety of a given preparation. However, these methods are designed for materials with a low bioburden, such as food and pharmaceuticals. Consequently, current methods may not be reliable or applicable to estimate the irradiation dose required to sterilise microbiological preparations for vaccine purposes, where bioburden is deliberately high. In this study, I investigated different methods of modelling sterility and developed a new formula for calculating sterilising doses that is highly applicable to viruses and bacteria. Our research group has developed a whole-inactivated influenza A virus (IAV) vaccine using γ-radiation (γ-Flu). IAV presents a constant pandemic threat due to the mutagenic nature of the virus and the inadequacy of current vaccines to protect against emerging strains. Previous research has demonstrated the ability of γ-Flu to protect against not only vaccine-included strains but emerging strains as well. In this study, I compared γ-Flu irradiated at different temperatures and doses to meet internationally accepted sterility assurance levels. I found that, when using sterilising doses, the structural integrity and vaccine efficacy was well maintained regardless of irradiation temperature. In fact, using a higher temperature and lower radiation dose induced higher neutralising antibody responses and more effective cytotoxic T cell responses. These data provided new insights into optimal irradiation conditions as previously using frozen irradiation was considered the superior irradiation temperature. These concepts related to preparing γ-irradiated vaccines were applied to the development of a novel vaccine against Newcastle disease virus (NDV). NDV is an important poultry pathogen that is associated with widespread livestock losses and a large economic burden. Current vaccines are available but have limited efficacy so there exists a need for alternative NDV vaccines. In this study, NDV was inactivated by γ-irradiation and structural integrity was tested. I found overall virion structure and protein function of γ-NDV to be well maintained, however surprisingly I did not detect neutralising antibody responses after treatment in mice. Interestingly, previous studies from our group have demonstrated the ability of γ-Flu to adjuvant other γ-irradiated vaccines. In the current study, I expanded on the broader applicability of γ-Flu as an adjuvant by showing that γ-Flu can adjuvant the poorly immunogenic keyhole limpet hemocyanin. However, γ-Flu and other commonly used adjuvants were unable to enhance neutralising antibody responses to NDV. Overall, these data suggest that γ-irradiation may not be a suitable inactivation method for NDV vaccine development. NDV is used as an oncolytic virus and many clinical trials have demonstrated the ability of NDV to treat a range of different cancers. However, research with NDV is hindered by the biosecurity risk associated with live NDV. Given the high structural integrity and protein function of γ-NDV, I hypothesised that γ-NDV could be used as an alternative cancer treatment. Importantly, γ-NDV retained its ability to kill a range of different cancer cells in vitro. This suggests that γ-NDV can be used as a broadly applicable therapeutic agent. Furthermore, I tested γ-NDV in a murine melanoma model and found that γ-NDV was able to reduce tumour growth and enhance overall survival.