Part A: The stability of Adenoviral and mRNA based COVID-19 vaccines and the potential of HSA/ Part B: Characterisation of IgG A33 hFab’ and human serum albumin using hydrodynamic methods

Abu Hammad, Khalil (2021) Part A: The stability of Adenoviral and mRNA based COVID-19 vaccines and the potential of HSA/ Part B: Characterisation of IgG A33 hFab’ and human serum albumin using hydrodynamic methods. MRes thesis, University of Nottingham.

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Part A:

At the end of 2019, the SARS-COV-2 virus was observed to cause respiratory disease, and it is responsible for the high number of patients admitted to hospital. Many biopharmaceutical / biotechnology companies have started manufacturing vaccines that can prevent the SARS-COV-2 virus. An important issue is COVID-19 vaccine stability, one of the most frequently stated problems faced through vaccination programs particularly in the high temperatures low-income countries. Along with cost, availability, and delivery – stability makes delivery simple. This review analyses the characteristics of the two common groups of COVID-19 vaccines – Adenovirus based (such as the Oxford/AstraZeneca and Janssen vaccines) and mRNA based (such as Pfizer/BioNTech and Moderna) – and in particular the potential application of human serum albumin (HSA) as a stabiliser for the mRNA COVID-19 platform. From the stability point of view, while the Adenoviral vector-based vaccines can be stabilised and stored at 2–8 °C range yielding 6 months shelf-life, the mRNA-based vaccines have to be stored at much lower temperatures, limiting their distribution to the higher temperature regions. This emphasise the importance of research into formulation solutions, such as the use of excipients like human serum albumin, to improve these products stability and, therefore facilitate vaccination programs without the need for cold chain storage. The availability of these vaccines to lowincome countries would have a major role in lowering the cases of COVID-19 infections. In future investigations, it is possible to look at different unstable COVID-19 platforms by applying new technologies for making a stabilise vaccine.

Part B:

Background: Hydrodynamic methods such as analytical ultracentrifugation, dynamic light scattering, SDS-PAGE, viscosity analysis, dialysis tubing and UV spectroscopy were employed. These methods were to investigate the heterogeneity/homogeneity, molecular weights, purity, the concentration measurements, solution conformations and intrinsic viscosity of IgG A 33 hFab’ and human serum albumin (HSA). Both macromolecules relevant in cancer and biopharmaceutical therapies. We also investigate the properties of mixtures of the two, probing for a possible interaction, which will help in developing formulations of cancer and biopharmaceutical therapeutics in the future. Methods: The dialysis experiment was performed to produce pure samples of HSA and IgG A33hFab’ macromolecules and measuring the concentration via UV spectroscopy. Determination of the molecular weight was applied by using SDS-PAGE and sedimentation equilibrium (SE) for both macromolecules. To analyse the HSA and IgG A33 hFab’ purity and the presence of possible dimers and aggregates, the SV method was performed. Also, the SV was utilised to obtain more information about the possible interaction between the IgG A33 hFab’ and human serum albumin. The dynamic light scattering method was utilised to further evaluate the heterogeneity of the IgG A33 hFab’ macromolecule. Viscosity measurements were applied to determine IgG A33 hFab’ and human serum albumin macromolecules shape with their intrinsic viscosities, using Ostwald (U-tube) capillary viscometer. Results: DLS results reinforced by sedimentation velocity (single peak) showed that IgG A33 hFab’ is homogenous and free of aggregated particles. Its molecular weight is ~ 48- 50 kDa and monomeric state using the SDS-PAGE and SE methods. HAS molecular weight is also shown to be homogenous from sedimentation velocity with a molecular weight of 70.6 kDa using the absorbance optical system in SE method. A single sedimentation peak was also seen in mixtures of HSA with IgG A33 hFab’. This could either be due to an interaction or due to the Johnston-Ogston effect. Using ELLIPS (molecular shape) both IgG A33 hFab’ and HSA are globular with very similar axial ratios. These results will advance the knowledge in biotherapeutics for an effective formulation in manufacturing process.

Item Type: Thesis (University of Nottingham only) (MRes)
Supervisors: Harding, Stephen E
Wayte, Judith
Keywords: vaccines, biopharmaceutical therapeutics, macromolecules, albumin, antibodies
Subjects: Q Science > QR Microbiology > QR180 Immunology
R Medicine > RM Therapeutics. Pharmacology
Faculties/Schools: UK Campuses > Faculty of Science > School of Biosciences
Item ID: 67067
Depositing User: HARDING, Prof Stephen
Date Deposited: 08 Dec 2021 04:40
Last Modified: 08 Dec 2021 04:40

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