Despite over 25 years of research, the goal of producing an HIV-1 vaccine able to
prevent infection has not been realised. Due to the limited immunogenicity of HIV-1
glycoproteins in vivo, this thesis explores the use of cholesterol depletion in increasing
the antigenicity and immunogenicity of an whole-inactivated HIV-1 immunogen.
The primary aim was to demonstrate that cholesterol depletion of HIV-1 could enhance
antibody binding and viral neutralisation. The laboratory-adapted strain, HIV-1MN, was
utilised to demonstrate effective cholesterol depletion of virus using the compound
methyl-β-cyclodextrin (MBCD). A range of MBCD concentrations were explored to
assess the effect of removing cholesterol on viral infectivity, morphology and protein
composition of the virus. A 1mM MBCD concentration, which effected a 50%
reduction in viral envelope cholesterol with little impact on viral structure and protein
composition whilst retaining viral infectivity, was chosen to explore the effect of
cholesterol depletion on virus antibody binding and neutralisation. Removal of
cholesterol from the viral envelope increased antibody binding and neutralisation using
a number of monoclonal antibodies, soluble CD4 (sCD4) and HIV-1 positive patient
antisera.
This concept was then extended to a pseudotyped primary isolate of HIV-1. Cholesterol
depletion of this isolate demonstrated a more restricted enhancement of antibody
binding and neutralisation of virus by monoclonal antibodies, sCD4 and homologous
and heterologous HIV-1 infected patient antisera.
A number of inactivating agents were then explored in the aim of creating a cholesteroldepleted,
whole inactivated HIV-1MN immunogen that retained key conformational
gp160 epitopes. This immunogen was then tested in a mouse model to investigate
whether cholesterol depletion could enhance the immunogenicity of whole-inactivated
HIV-1MN. Although measurement of antibody responses from pooled mouse sera
indicated significant enhancement of humoral responses to cholesterol-depleted, wholeinactivated
HIV-1MN, analysis of individual mouse responses to immunisation yielded
more variable results.