Defining determinants of viral membrane protein immunogenicity

Abstract

Acquired immunity to the delivery vector for in vivo gene transfer or expression applications limits therapy efficacy upon repeat vector administration. However, a controlled comparison of vector immunogenicity, comparing immune responses to different viral vectors at comparable doses through the same delivery route, is yet to be carried out. In this thesis, antibody assays were used to study the humoral response to repeat dosing of lentiviral (LV) vectors of variable pseudotype (Sendai Virus Fusion protein/Hemagglutinin Neuraminidase protein and Vesicular Stomatitis Virus glycoprotein). The effects of antigen dose and delivery method were assessed. Through this, the envelope:gag ratio was identified as an important determinant for vector immunogenicity. Upon the emergence of SARS-CoV-2 in the United Kingdom in early 2020, the focus of this thesis shifted to studying the human antibody response to this pandemic virus. Antibody assays previously used to study anti-vector pseudotype antibodies to gene transfer vectors were used to study the kinetics and magnitude of antibody responses to SARS-CoV-2 variants. Interestingly, SARSCoV-2 Spike (S) S2 domain-specific IgG antibodies, derived from previous heterotypic human coronavirus (HCoV) infection, were identified in individuals uninfected and unexposed to SARS-CoV-2, and these S2-specific cross-reactive antibodies showed neutralising activity to SARS-CoV-2 S-pseudotyped LV vectors in vitro with comparable efficacy to sera from COVID-19 patients. Sera from patients infected with SARS-CoV-2 variants D614G or Alpha were able to recognise and neutralise homotypic and heterotypic variant S proteins with variable efficacy. Surprisingly, in some cases, a reduction in efficacy of binding and neutralisation occurred in a unidirectional and asymmetric pattern. Together, this thesis explores the many factors which determine antibody induction and duration, and suggests ways in which we can use this information to design better therapeutics involving LV gene transfer, such as gene therapy and vaccination.