Date of Award
2024
Document Type
Open Access Dissertation
Degree Name
Doctor of Philosophy in Chemical Engineering (PhD)
Administrative Home Department
Department of Chemical Engineering
Advisor 1
Caryn Heldt
Committee Member 1
Michael Betenbaugh
Committee Member 2
Timothy Eisele
Committee Member 3
Stephen Techtmann
Abstract
Cell and gene therapies, a ground-breaking class of drugs that can heal genetic diseases and cure cancer, often have treatment costs over $1 million. Most cell and gene therapies are delivered using virus-based delivery vectors, which are incredibly expensive to manufacture. Liquid chromatography is the favored method of viral vector purification since it is easily scalable and can be functionalized to purify by biospecificity, charge, hydrophobicity, or size. However, chromatography-based purification processes are highly expensive and usually only recover 20-50% of the viral vectors. There is currently no platform purification strategy for viral vectors due to differences in their surface chemistries. An alternative purification method that could deliver low-cost, low-shear, platformed purification could significantly simplify process development and manufacturing for these cutting-edge therapies.
Aqueous two-phase extraction (ATPS) is a liquid-liquid extraction method that has shown repeated success at virus purification. This dissertation outlines the economic motivation, chemical conditions, continuous processing adaption, and platformability of a polyethylene glycol and sodium citrate-based ATPS. The ATPS detailed here is a two-step process in which the first step partitions the virus into the polymer-rich phase to purify it from host cell contaminants. The second step back extracts the virus from the polymer-rich phase into fresh citrate. The key driving forces of this process are electrostatic interactions, which can be adjusted using pH and phase concentrations, and hydrophobic interactions, which can be adjusted using polymer molecular weight and phase concentrations. Economic modeling of our process showed that it delivers 90% reduction in capital costs and 50% reduction in operating costs at 11 kg per year production scale. Product recovery and DNA removal was similar for the continuous adaption of this process compared to initial batch-mode studies. Mixing and settling are the key operations in ATPS that must be controlled to ensure consistent purification performance. Finally, the two-step ATPS was tested with a diverse set of viral vectors to show its potential as a purification platform for both enveloped and non-enveloped vectors.
In all, this dissertation supports the economic and operational feasibility of ATPS for viral vector purification in the cell and gene therapy industry.
Creative Commons License
This work is licensed under a Creative Commons Attribution 4.0 License.
Recommended Citation
Nold, Natalie M., "CONTINUOUS, PLATFORM PURIFICATION OF VIRAL PRODUCTS USING AQUEOUS TWO-PHASE EXTRACTION", Open Access Dissertation, Michigan Technological University, 2024.