Scalable Gene Therapy Manufacturing

Scalable manufacture involves producing enough gene therapy to accommodate the high-dose demands of a systemic disease or one which has a substantial patient population. Today, this is perhaps one of the most challenging issues facing the cell and gene therapy industry.

A large number of Contract Manufacturing Organisations (CMOs) work with adherent cells. This is generally considered to be the standard approach to cell culture, enabling cells to maintain contact with the bottom of the cell culture dish, as well as their neighbours.

The culture of adherent cells at scale, however, involves a huge surface area – a consideration that rapidly turns into a limiting factor. In order to address this issue, the first step towards effective scaling up is to move from adherent to suspension cell culture.

Step one: Switch to suspension culture

Suspension cell culture offers a number of advantages, involving the adaption of cells so that they do not rely on contact with the cell culture dish surface. Here, cell growth is only limited by the concentration of cells in the medium, so this can be readily scaled up.

Growing cells in suspension - particularly at scale – involves experienced, careful handling. Factors such as gas exchange, cell growth, and clumping must be monitored carefully before being optimized for scaling up. Optimization of this process may increase manufacturing efficiency considerably, maximizing the eventual viral yield.

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.

Jakub Krakowiak, Group Leader, Process Development Team, OXGENE

Step Two: Optimize Scale-Up in a Transient Expression Platform

Jakub and his team approach this task in a measured, sequential manner. First, they optimize cell growth and viral vector production conditions within shake flasks. Next, they employ an Ambr^®15 high-throughput microscale bioreactor system to optimize cell growth via small scale bioreactors. They accomplish this by utilizing a Design of Experiments (DoE) approach.

This approach involves test parameters such as cell density at seeding and transfection, ratio relative to transfection reagent, DNA concentration, and the testing of transfection reagents.

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

Jakub Krakowiak, Group Leader, Process Development Team, OXGENE

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

OXGENE’s transient gene therapy manufacturing platform aims to provide its customers with the building blocks for successful applications.

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 optimize 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.

Jakub Krakowiak, Group Leader, Process Development Team, OXGENE

Step Three: Transition to OXGENE’s Fully Scalable, Stable Expression Platform

The primary advantage of working with OXGENE is early access to the company’s developing technology and solutions roadmap. This roadmap concentrates on developing future-proof technologies that are designed for scalable and stable gene therapy production.

Most of the company’s relationships with customers begin with the optimization of transient production, with the ultimate goal being the smooth transition to a stable technology platform.

This assists in the reduction of batch-to-batch variation while lowering manufacturing costs and enabling easier scale-up. It also provides the further regulatory advantage of total compatibility with previous data – an advantage that occurs due to the stable platform retaining the same base cell line and expression cassettes as the transient system.

Process development acts as a central contribution to the successful transition to stable gene therapy production.

Jakub expands on this concept, “When it comes to stable cell lines, process development can be quite different. Because some or all the DNA is already integrated into 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 optimize the different cell densities, timings and concentration of materials going into the process, which can have a big impact on the final titer. In fact, when it comes to improving titers for a stable cell line, the sky is the limit! But realistically we would hope to improve the titers by about ten-fold between the original method and final optimized process.”

The definition and optimization of efficient scale-up processes is a key element of OXGENE’s aspirations. The company is working to become a frontrunner in the development of strictly controlled, carefully optimized technologies for cost-effective, scalable, high-quality gene therapy manufacture.

Eventually, through increasing efficiency and cost reductions in the manufacturing process, OXGENE aims to lead the gene therapy industry in the development of accessible, sustainable solutions for the treatment of genetic disease.

Acknowledgments

Produced from materials originally authored by Sophie Lutter from OXGENE.

About OXGENE

OXGENE™ combines precision engineering and breakthrough science with advanced robotics and bioinformatics to accelerate the rational design, discovery and manufacture of cell and gene therapies across three core areas: gene therapy, gene editing and antibody therapeutics.

Gene therapy: We’re transforming the vision of truly scalable gene therapies into a reality; progressing our industry leading transient gene therapy systems towards alternative technologies for scalable, stable manufacturing solutions.

Gene editing: We have automated gene editing to deliver CRISPR engineered cell lines at unparalleled speed, scale and quality and generate complex disease models in mammalian cells.

Antibody therapeutics: We’re employing a novel proprietary mammalian display technology to discover antibodies against previously intractable membrane proteins.

OXGENE™ works at the edge of impossible in mammalian cell engineering. Our scientific expertise and technology solutions address industry bottlenecks. For more information, please visit www.oxgene.com

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