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09 April 2020 by Sophie Lutter

Scalable manufacture

OXGENE’s recipe for success, and scalable gene therapy manufacture

Scalable manufacture – that is, producing enough of a gene therapy to meet the high-dose demands of a systemic disease, or one with a large patient population - is arguably one of the biggest challenges facing the cell and gene therapy industry today.

Many Contract Manufacturing Organisations (CMOs) work with adherent cells. This is the ‘standard’ approach to cell culture, allowing the cells to maintain contact with the bottom of the cell culture dish, and their neighbours. But to culture adherent cells at scale requires an enormous surface area, which quickly becomes a limiting factor. Therefore, the first step towards efficient scaling up is to transition from adherent to suspension cell culture.


Step one: switch to suspension culture

Suspension cell culture provides great advantage. It means adapting the cells so that they no longer rely on contact with the surface of the cell culture dish. Cell growth becomes limited only by the concentration of cells in the medium, so can be readily scaled up. But growing cells in suspension, especially at scale, requires careful – and expert – handling. Factors like cell growth, gas exchange and clumping must be monitored carefully and further optimised for scaling up. Optimising this process can significantly improve manufacturing efficiency by maximising the eventual viral yield.

Jakub Krakowiak, OXGENE’s Process Development Team group leader, explains:

“Moving from shake flasks to bioreactor can be challenging. Changes in the geometry of the vessels, as well as differences in aeration and in the way the systems are controlled all bring challenges to overcome.”


Step two: optimise scale-up in a transient expression platform

Jakub and his team do this by first optimising cell growth and viral vector production conditions in shake flasks. They then use an Ambr®15 high-throughput microscale bioreactor system to optimise cell growth in small scale bioreactors through a Design of Experiments (DoE) approach. Here, they test parameters such as cell density at seeding and transfection, DNA concentration and ratio relative to transfection reagent, as well as testing different transfection reagents.

Jakub adds, “From there, we move to 0.5 or 1L bioreactors, where we need to consider how the process will transfer to another different vessel type. This is where we need to optimise stirring, aeration and pH, both in terms of setpoints and control.”

Jakub’s team currently scale up to 10L, before introducing downstream processing. “ At this stage we combine filtration and chromatography techniques to remove impurities and concentrate the final material.”

The goal of OXGENE’s transient gene therapy manufacturing platform is to leave our customer with a recipe for success. “What makes OXGENE special is that we can partner with a customer from a very early stage of product development all the way through to manufacturing. We provide plasmids and cell lines as a basis for the process, depending on customer requirements, and then we work with our customers to develop and optimise their process. Once we transfer the process to a manufacturing facility, we continue to work with both the manufacturer and the customer to troubleshoot and tweak the process until it’s working perfectly in the manufacturing setting.”


Step three: transition to OXGENE’s fully scalable, stable expression platform

The major advantage of partnering with OXGENE however is gaining early access to our developing technology and solutions roadmap, focused on building future-proof technologies for scalable, stable gene therapy production.

Most journeys with our customers start with the optimisation of transient production, our goal is to further support their transition to a stable technology platform. Not only does this help reduce batch to batch variation, lower manufacturing costs and facilitate easier scale up, but it also provides the additional regulatory advantage of complete compatibility with previous data, as the stable platform retains the same expression cassettes and base cell line as the transient system. Process development is a major contributor to successful transitioning to stable gene therapy production.

Jakub expands on this, “When it comes to stable cell lines, process development can be quite different. Because some or all the DNA is already integrated in the cell line, we can do some things with the process that we're not able to do with a fully transient production. For example, we can try to optimise the different cell densities, timings and concentration of materials going into the process, which can have a big impact on the final titre. In fact, when it comes to improving titres for a stable cell line, the sky is the limit! But realistically we would hope to improve the titres by about ten-fold between the original method and final optimised process.”

Defining and optimising the process for efficient scale up is a key part of OXGENE’s ambition to pioneer the development of tightly controlled, and carefully optimised technologies for scalable, cost effective and high-quality gene therapy manufacture.

Ultimately, through increasing efficiency and reducing costs in the manufacturing process, we aim to lead the gene therapy industry towards the development of sustainable and accessible treatments for genetic disease.