Sourcing Plasmid DNA From CDMOs Got More Challenging — Here's Why
By Antony Hitchcock, principal and owner, AGH Bioconsulting
It goes without saying that the last two years have been challenging for the whole of the biotechnology sector, with widespread layoffs and reduced investment in the sector. The cell and gene therapy sector is highly dependent on private investment in startups and clinical development companies, and funding shortfalls have led to many clinical development projects being canceled or put on hold until companies could obtain further funding.
Against this background, I attended the recent European Society of Gene & Cell Therapy (ESGCT) meeting in Rome to try to understand the current climate and challenges facing the CDMO industry in the gene therapy space and, more specifically, those engaged in the production of plasmid DNA and viral vectors.
Plasmid DNA Market
Market
While the vast majority of plasmid DNA produced by CDMOs is as a template product for production of viral vectors and, to a lesser extent at present, mRNA-based therapies, there are some exceptions, such as vaccines, including personalized cancer vaccines, and direct cell modification using electroporation.
While there are some licensed gene therapy products, the majority of the business is still focused on the production of materials to support clinical development rather than commercial supplies. Consequently, the demand for plasmid has been hit by the fall-off in demand for viral vectors as clinical development programs have been put on hold until developers are able to secure funding for their clinical development programs.
There has also been a growing market for plasmid DNA as a template for mRNA production, and while there are many mRNA products in clinical development, the only licensed products are COVID vaccines, including the updated version, which are now largely only being given to vulnerable patient groups. Until we see more licensed products, and their levels of adoption, it will be difficult to judge this market.
Consequently, the current demands within the plasmid DNA market are intrinsically linked to the demands and needs of these markets and funding environments for these products, and so understanding these needs and drivers is critical to understanding the plasmid DNA market.
Suppliers
Historically, product development companies looked to source plasmids from specialist plasmid producers and then provide these to a specialist vector producer, with a limited number of companies offering both services. In these cases, the customer would procure individual batches of the two to four plasmids required to construct their vectors. We are now seeing some changes in this model.
First, we have seen the concept of one-stop shops emerging where CDMOs look to provide both vector and plasmid suppliers. This model has some attraction for customers, but from the CDMO perspective it can be challenging with regard to business focus and being able to run both services as profitable operations, and until recently may of the specialist vector suppliers have remained focused on their own service offerings. However, some of these players are looking to establish in-house plasmid production capabilities, which seems to be focused on taking greater control of plasmid supply in terms of timelines and quality for their customers.
Second, we are seeing an increased number of plasmid producers selling “off the shelf” plasmids. This relates to the standard plasmids such as the Helper function and some RepCap plasmids used in the production of AAV vectors and the Rev, GagPol, and VSV-G plasmids used in the production of lentiviral vectors. This makes sense for the production of clinical materials in that it reduces cost and critically timelines for vector production. However, it is unclear how this option will play out for Phase 3 and commercial products, as priorities change with regard to control and ownership of materials.
Production Scale
While some CDMOs have established large production platforms of up to 1,000 L, the reality is that the majority of producers are operating at the 30 L to 50 L scale, producing < 10 g of plasmid, with most customers requiring less than 5 g to support early-phase clinical development programs, especially for lentiviral and RNA products, which have limited material requirements.
Some CDMOs have established larger-scale systems, usually based around 300 L single-use production vessels, targeting the anticipated needs of late-stage commercial AAV products. The low number of AAV products coming through to licensing and the high costs of these products have limited the number of CDMOs willing to invest in the scale-up and establishment of larger production facilities.
Production Grades
For a number of years there has been the concept of phase-dependent plasmids, where non-GMP-grade plasmids, which cost less and can be produced faster, may be used in the production of vectors for early-stage clinical studies. Meanwhile, GMP-grade plasmids are used for late-stage and commercial vector production. In response to this split demand, suppliers established dual supply chains to meet customers’ requests for both grades of plasmid.
Over the last few years both the EU and United States Pharmacopeia have issued guidelines detailing quality expectations for the non-GMP plasmids, which require significant investment in facilities and quality systems to meet these requirements, potentially decreasing the differential in costs and timelines between the non-GMP and GMP grade materials. This will inevitably affect the decision-making process for product developers going forward; however, it is clear that despite these challenges, there is still a demand for these materials, including the “off the shelf” plasmids made to non-GMP grades.
Technical Challenges
Plasmid production is based on the concept that production costs and timelines can be kept to a minimum using platform processes, which allows for the elimination of the majority of development studies and engineering runs. This approach also means that GMP documentation and supply chains can be retained with very limited changes between different production batches.
While this approach works for the majority of plasmids, it does not work for all. This tends to apply to the gene of interest (GOI) plasmids used in the production of viral vectors and RNA vectors. This materializes in issues with plasmid stability, such that incomplete plasmids with low productivity are produced. These issues relate to elements such as ITRs, Poly A tails, and other sequence repeats.
While developers have sought approaches, including the use of alternative production strains and modification to fermentation strategies, what is clear is that there are no simple solutions to this problem, with little in the scientific literature as guidance. As a result, most of the approaches used to overcome this issue are largely trial-and-error-based, rather than knowledge-driven.
Production Approaches
While the majority of plasmid DNA for use in clinical products has been produced using bacterial systems, we are seeing an expansion of companies’ development capabilities for producing plasmid DNA through synthetic routes. While this is not entirely new technology, the technology is being applied for a number of applications in the gene therapy and vaccine segments.
What is not fully clear is the levels of success and adoption they will achieve, not least competing against alternative gene delivery technologies including RNA-based approaches and ultimately it will be the successes of these clinical applications that are likely to determine the further levels of investment and development in these technologies.
Future Developments
When looking at where the plasmid market is going and what might be the key areas of development, from a technical perspective, it is clear there is still room for process improvement.
We need improved approaches to the production of GOI plasmids with challenging sequence elements, such that producers have standardized approaches to this, which may include alternative non-E. coli-based approaches.
Additionally, there is a clear need for a systematic assessment to identify those sequences that create serious manufacturing issues, such that they can be designed out of plasmids. It may also be necessary for putative plasmid constructs to be screened for manufacturability before final constructs are selected.
The other area for technical development is the production of off-the-shelf cell lysis equipment. Currently, in-house systems are used, which limits production scale and scope for process intensifications, and there is a clear opportunity for suppliers to develop improved solutions for plasmid manufacturers.
Historically, the demand for plasmid was based on the large-scale production of AAV vectors at 2,000 L scale or greater and on the use of high dose treatments as high as 1E14vg/Kg for larger patient populations. However, as we see the emergence of improved vectors such as Pfizer’s recently-approved Beqvez for the treatment of hemophilia B with doses down to 5E11vg/kg, compared to the existing CSL Behring product dosed at 3E13vg/kg, some of these calculations relating to predicted AAV requirements and in plasmid demand may not hold true.
Additionally, when we look at the lentivirus, it has predominantly been used in ex vivo applications, with smaller amounts of vector required. Production scales have been in the region of 200 L. In terms of the demand for plasmids as templates for mRNA products, it is hard to judge at present. Demand may be relatively small for the majority of licensed products, other than prophylactic vaccine production for large patient populations.
There are also emerging threats and opportunities for the plasmid markets. A key threat is emergence of the producer and stable cell lines to produce both AAV and lentivirus, eliminating or reducing the need for plasmid. The development of these technologies has been challenging and taken significantly longer than developers would have hoped for; however, such technologies are likely to come into wider use, especially for therapies targeting larger patient populations. This again potentially negates the need for large-scale plasmid production.
Consequently, unless there are significant changes in the AAV market, the drivers for developing plasmid production scales beyond 300 L may be limited. What is more likely is that plasmid producers expand capabilities through increased numbers of production streams of existing scales, decreasing cost and risks.
Conclusions
Having met with a number of CDMOs during the ESGCT meeting, my perception is that less funding in the cell and gene therapy space has led to a very challenging environment for many of the companies offering plasmid DNA and viral vector production services.
While we are seeing an increase in licensed products, especially cell therapy/CAR-T rather than AAV-based gene therapy, this does not seem to always result in additional growth in the sector.
However, I did feel that there is still a level of confidence in the more established companies that the increases in funding we have seen during 2024 will eventually lead to increased numbers of entities entering clinical development as stalled and new development programs are brought forward.
The significant investments in the RNA-based therapies will also create potentially new markets outside of the COVID vaccines, which in turn creates new opportunities for GMP and high-quality plasmid suppliers. Alongside this, the increased interest and investment in synthetic plasmid production offers greater potential for the development of plasmid therapies and vaccines. While the industry is still facing a number of challenges, there is still a level of confidence that 2025 will be a year of improved growth within the sector.
About The Author:
Tony Hitchcock is the principal and owner of AGH Bioconsulting. He has spent his decades-long career in the in the biotechnology field, with over 30 years in the production of complex biologics for clinical trials in the European Union and U.S. He has worked in areas of process development and manufacturing with experience in engineering and process systems. He has worked on the development of more than 30 products for clinical trials, including plasmid DNA, viral and bacteriophage products, and recombinant proteins from microbial, mammalian, and insect cell sources. Contact him at tony@aghbioconsulting.com.