Taking your adenoviral vector product from R&D to the clinic

Adenoviruses have been shown to be very effective vehicles for delivering genetic material to cells and are therefore in demand to develop gene therapies and vaccine products. The road from early-stage R&D to final production of an adenoviral vector product is well trodden, and there is a lot of detailed information out there to help you on your journey.

That said, taking an adenoviral product to the clinic always requires some process development prior to bringing the vector into GMP. This is based on the many examples in practice where the transgene had detrimental effects on vector yield or purity once inserted into the vector. For instance, the foreign gene may encode for a protein that interferes with HEK293 cell propagation, resulting in low yield of vector particles per cell. Likewise, the protein produced may bind to the Adenoviral vector capsid proteins, which are generally carry a net negative charge that can substantially affect the purification strategy.  A good way of mitigating risk is to seek expertise at the relevant stages of development to fill informational gaps and reduce the possibility of unexpected problems while moving from bench to clinic.

Working with a center of excellence contract development and manufacturing organization (CDMO) is an attractive option for many who embark on the adenoviral production journey. Other than a general vendor type CDMO, A center of excellence CDMO has in-depth experience and can help guide the process by providing practical solutions to challenges that may pop up.   If you are developing a therapy from the very beginning, a “center of excellence CDMO” can often complement your expertise and help you navigate the complexities of the scale-up process.

In this article, we highlight how a “center of excellence CDMO” can help you to smoothly navigate the transitions from R&D to scale-up and clinical manufacturing. The information here will provide you with a stable foundation when you start a conversation with a CDMO, allowing you to identify important traits that will give you a strong relationship going forward.

Common challenges that a CDMO can overcome

Concentration challenges

One of the most important considerations that need to be made early in the product development trajectory is the concentration of the vector needed to be an effective therapy in the clinic (part of Target Product Profile setting).

For adenovector based vaccines, a dose of 10¹⁰-10¹¹ virus particles per milliliter typically suffices for systemic administration i.e. intramuscular injection. For oncolytic therapies, either higher or lower doses could be required, depending on the route of administration and the target tissue. For instance, treatment of a wound-bed post cancer surgery may require doses of up to 10¹³ virus particles per milliliter to effectively deliver a high dose in a confined location with minimal spill-over to surrounding tissue.

At such high vector concentrations, aggregation of virus particles may occur, which reduces therapeutic efficacy. The high concentrations needed for many oncolytic and gene therapies exacerbate this problem. Aggregation can manifest itself in quite spectacular fashion, as concentrated adenovirus preparations become cloudy before “snow” appears on the bottom of the vial. Understanding how adenovirus particles behave and good forward planning are essential to reduce the chances of aggregation.

On the other end of the spectrum, there is also a risk that during the scale-up process the concentration step will not be sufficiently effective, leading to viral titers that are lower than expected. This is a critical issue that can arise due to the significant alterations that are often required during scale-up of concentration steps, such as switching from ultracentrifugation to chromatography.

Since the stakes and capital investment are always higher the further down the scale-up path you are, it pays to address any issues relating to concentration as early as possible.

Release of intracellular DNA

Eukaryotic cells have to be lysed in order to release the adenoviruses, and in the process the release of intracellular DNA is inevitable. The more viral particles needed, the greater the number of cells that must be lysed, and therefore the more host cell DNA is released. Unfortunately, DNA often forms an intricate and sticky web that easily traps adenovirus particles, causing them to aggregate. As discussed in the previous paragraph, adenoviruses are already prone to aggregation, so adding DNA into the mix only increases the risk. The DNA webs also are notorious for clogging filters, which further hampers the purification process. These problems can be circumvented in the process development stages by adding a DNA reduction step such as the introduction of benzonase. With careful quality control, interventions like this can mitigate the negative effects of extracellular DNA.

Process yield & scalability

Perhaps the biggest consideration when manufacturing Ad vectors is the process yield at large scales. In the R&D phase, simply ensuring there is “enough” virus to perform preclinical experiments is acceptable. However, production at larger scales inherently leads to larger losses. During the R&D phase it is therefore essential to prioritize yield. This can be done by reducing the number of empty capsids that are present in your viral preparation and ensuring that your process is ready for the switch to large scale production. This often requires changing from ultracentrifugation, which is a common way to concentrate adenovirus in the research lab but not appropriate for scaling. With technologies used for scale up such as chromatography, it is often difficult to achieve the same yield as ultracentrifugation which is why process development is so important.

How does a CDMO help?

Understanding the different applications for adenoviral vectors is a good way of highlighting how a good relationship with a center of excellence CDMO should work. The center of excellence will have industry experts who can save you a lot of time and expense in developing robust and scalable manufacturing processes for your vector. For example, at Batavia, our in-house experts have extensive experience in developing adenoviral vector-based therapies be it vaccines or oncolytic products. These experts are deployed to relevant projects to help guide the strategy and ensure the highest likelihood of manufacturing success.

Clear communication is a must for a good center of excellence CDMO. Clearly articulating the challenges and opportunities of a scale-up project is the only way to make effective progress. While good communication may seem less relevant than the technical capabilities of a prospective CDMO, it is nevertheless important and very easy to assess early on your interactions. Pay close attention to how your CDMO communicates. Make sure that misunderstandings are quickly resolved, and that communication is clear and timely. The way a CDMO communicates and structures their meeting and interactions is a reflection of how they operate, so if you are not satisfied, it’s best to move on.

A good CDMO should bring balance and clarity to the scale-up project. When you partner with a center of excellence CDMO, you should feel confident that you are in good hands. You should also feel that you acutely understand the process going forward, including the potential risks.

At some point when moving from R&D to scale-up and production, you will meet a whole range of different regulations that have to be thoroughly studied and strictly adhered to, if your adenovirus vector is to make it successfully into the clinic. A good center of excellence CDMO has experience with the relevant and most up-to-date regulations and requirements, and will help you navigate the complex landscape of rules and paperwork. Always ask about track record.

Choosing a CDMO

If you decide that working with a center of excellence CDMO is the right path for you, the next question is who should you choose? Fortunately, the CDMO field is competitive, and you do have a choice. It’s important that you exercise this choice and do your due diligence by thoroughly researching and interviewing your CDMO candidates. Pay close attention to the way they communicate with you and how they discuss your project and bring up challenges. Most importantly pay close attention to the quotation and make sure to read the small footnotes to understand what is included in price and what is excluded and needs to be paid additionally.

It’s never too early to begin discussions with a partner to help you scale your adenoviral vector product. Even informal conversations early on can give you insights and help you make decisions that will have a positive impact on the future of your project. To start with, why not book a call with one of our experts. We’re looking forward to hearing about your project and you can be sure that we’ll give you clear advice and an all-included costing overview for your project.