City Research Online

Abstract

Rabies virus (RABV) causes possibly the oldest and most feared disease of mankind and is responsible for >59,000 human fatalities/year. Post exposure prophylaxis (PEP), the administration of vaccine and rabies immunoglobulin, is a highly effective tool but largely unavailable in areas where RABV is endemic. Furthermore, due to the constraints of the Blood-Brain Barrier, current PEP regimes are ineffective after the onset of clinical symptoms, which invariably results in death.

To circumvent this barrier, this thesis focuses on the design of a live attenuated recombinant RABV expressing a highly RABV-neutralising scFv antibody (62-71-3). 62-71-3 is a highly characterised mAb directed against antigenic site I (AS-I) of the RABV G protein. 62-71-3 has been shown to effectively neutralise RABV when expressed as either a full-length IgG or scFv. Using reverse genetics, 62-71-3 scFv was inserted at the front of the RABV genome for optimal expression and linked to the fluorophore mCHERRY for visualisation under UV illumination. Furthermore, to avoid self-neutralisation, AS-I was mutated to diminish 62-71-3 scFv binding. Once rescued, the resulting construct (named RABV-62scFv) was grown to high titres and its growth and cellular dissemination kinetics characterised. In addition, binding and neutralisation studies demonstrated that functional 62-71-3 scFv was effectively secreted in neutralising concentrations in cell culture.

RABV-62scFv was then taken forward to in vivo studies in mice where it was shown to be non-lethal when administered at high doses (104 FFU) intracranially. Intracranial administration was also shown to elicit high serum neutralisation titres. To assess whether RABV-62scFv treatment was able to prevent mortality, mice were treated with an intracranial injection of RABV-62scFv either concurrently, three, or five days after lethal RABV (RV437) challenge. This revealed that intracranial RABV-62scFv treatment significantly increased survival rate (33% survival when given 3 dpi) and delayed the progression of symptoms.

Publication Type:	Thesis (Doctoral)
Subjects:	Q Science > Q Science (General) R Medicine > R Medicine (General) R Medicine > RB Pathology
Departments:	School of Health & Medical Sciences > Infection and Immunity Research Institute School of Health & Medical Sciences > School of Health & Medical Sciences Doctoral Theses Doctoral Theses

Export

Downloads