Stabilisation of therapeutic proteins using surface engineering and ionic liquids
File(s)
Author(s)
Bui-Le, Liem
Type
Thesis
Abstract
The temperature sensitivity of proteins hinders the bioavailability and safety of therapeutic antibodies and vaccines. These biomolecules require storage at 2 – 8 °C from manufacturing to administration to prevent thermal degradation, known as the ‘cold chain’. However, the cold chain cannot be implemented in resource-limited areas with inadequate infrastructure. Stabilising proteins at room temperature would break the ‘cold chain’ and allow life-saving treatments to be distributed to resource-limited areas, where infectious diseases can account for over half the mortality rate.
In this thesis, surface engineering and ionic liquids were used to improve the storage-lifetime of proteins (Chapter 3 – 5). Surface engineering can improve thermal stability whilst increasing solubility in ionic liquids; however, ionic liquid-protein interactions are misapprehended and often contradictory. Therefore, a holistic multi-technique framework — a combination of scattering and spectroscopic techniques — was developed on the green fluorescent protein and avidin as an archetypal probe to decipher the complex interactions (Chapter 3). The framework revealed the effect of ionic liquid anion nuances on protein structure and stability.
Surface engineering has been applied for the first time to ligand‐binding proteins (Chapter 4) and antibodies (Chapter 5) to create a thermally stable biofluid. Surface-engineering of avidin revealed a nanoconjugate with retention of activity and a half denaturation temperature of 139.0 °C and observed no denaturation after equivalent storage at 25 °C for 160 days. For the surface-engineering of antibodies, a maximum half denaturation temperature of 198.2 °C, while retaining binding activity, revealing that the high stability was transferable. However, a potential trade-off between thermal stability and bioavailability was observed. Regardless, the unprecedented transferable thermal stability presented herein provides a blueprint for stabilising vaccine candidates, avoiding the use of ultra-low temperature freezers. The stabilisation of these vaccines will be necessary for their dissemination to the unimmunised population to prevent or end a pandemic.
In this thesis, surface engineering and ionic liquids were used to improve the storage-lifetime of proteins (Chapter 3 – 5). Surface engineering can improve thermal stability whilst increasing solubility in ionic liquids; however, ionic liquid-protein interactions are misapprehended and often contradictory. Therefore, a holistic multi-technique framework — a combination of scattering and spectroscopic techniques — was developed on the green fluorescent protein and avidin as an archetypal probe to decipher the complex interactions (Chapter 3). The framework revealed the effect of ionic liquid anion nuances on protein structure and stability.
Surface engineering has been applied for the first time to ligand‐binding proteins (Chapter 4) and antibodies (Chapter 5) to create a thermally stable biofluid. Surface-engineering of avidin revealed a nanoconjugate with retention of activity and a half denaturation temperature of 139.0 °C and observed no denaturation after equivalent storage at 25 °C for 160 days. For the surface-engineering of antibodies, a maximum half denaturation temperature of 198.2 °C, while retaining binding activity, revealing that the high stability was transferable. However, a potential trade-off between thermal stability and bioavailability was observed. Regardless, the unprecedented transferable thermal stability presented herein provides a blueprint for stabilising vaccine candidates, avoiding the use of ultra-low temperature freezers. The stabilisation of these vaccines will be necessary for their dissemination to the unimmunised population to prevent or end a pandemic.
Version
Open Access
Date Issued
2020-12
Date Awarded
2021-03
Copyright Statement
Creative Commons Attribution NonCommercial Licence
License URL
Advisor
Hallett, Jason
Sponsor
Dr John Televantos
Engineering and Physical Sciences Research Council (EPSRC)
Grant Number
EP/N509486/1 (EPSRC DTP 2016-2021)
Publisher Department
Chemical Engineering
Publisher Institution
Imperial College London
Qualification Level
Doctoral
Qualification Name
Doctor of Philosophy (PhD)