Transient Protein Gene Expression and Biotherapeutics: Advances, Challenges, Solutions and Clinical Use

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Ann Nguyen:

Hello. I'm Ann Nguyen, Senior Associate Conference Producer with Cambridge Healthtech Institute, here for a podcast for the Optimising Expression Platforms conference at PEGS Europe 2018, happening the 14-15 November in Lisbon, Portugal.

Today, we're honored to chat with our keynote speaker, Dr. Florian Wurm, Professor Emeritus with the Swiss Federal Institute of Technology Lausanne and founder and chairman of ExcellGene SA.

Florian, welcome and thank you for being here.

Florian Wurm:

You're welcome. A pleasure.

Ann Nguyen:

Let's talk about transient protein expression. What advances do you foresee in this approach for enabling efficient engineering and expression of therapeutic proteins in the next decade?

Florian Wurm:

Well that depends on how you look at it. Obviously transient gene expression has been around for more than three decades and it has been instrumental in generating new protein variance, or fusions, or constructions, which nature never produced. So, it has been already a core of anything new in the business of making pharmaceutical products in such a long time.

The question I guess you are driving at is a little bit where does it go from here? Will we see maybe one day a transiently expressed protein be used in clinical trials. And while I would like to say yes, there's still a lot of barriers we have to overcome, and I was one of the first to using it, more than 30 years ago.

The yields at the time, when transfection technology was much poorer than it is today, were rather low and today we have transient expression technologies. Some of which my group has contributed to, where we can produce several hundred milligrams of a given protein per liter in a small- to moderate-scale bioreactor.

Now, obviously it's a very fast technology. It delivers you the protein essentially days after delivery of the DNA into the reactor. However, the scale-up issues are still very challenging because you need quite a lot of DNA. Actually, to transfect, let's say, a volume of cells in a one liter, or 10 liter, or even larger 100 liter reactors, then other things then, the pure expression technology become bottlenecks.

For example, making enough DNA at a quality which satisfies regulatory agencies and removal or prevention of introduction of endotoxins in the DNA because DNA's made in bacteria, which are known to produce endotoxins. So, there is a number of challenges, which are, let's say, both from a regulatory perspective, can the FDA accept a transiently produced protein? Whereas the entire industry is built on stable expression, where you pick a clone and scale that up and produce it in very large scale.

So, there are these, let's say, very much regulatory challenges, but there is also technical challenges, which are mostly related to scale-up and manufacturing.

Ann Nguyen:

What potential solutions seem promising for some of these challenges?

Florian Wurm:

This is a question which relates to each individual protein. If you're talking about antibody-based therapeutics, where we have to deliver large doses to patients – and we are talking literally grams per patient, which have to be delivered over a year and you treat hundreds and thousands and ten thousands of patients – you will always require a manufacturing approach, which is very much optimized and very much scalable to the very large scale of operation.

And you know, of course, that some of the leading products like Herceptin and Rituxan and Enbrel are produced in 10,000 liter reactors or larger and they're used year-round to deliver literally ton quantities of materials.

I don't think that products like these would be considered for transient expression in the near future. What I could however imagine, is that products which have a much higher potency, which can be used in much smaller doses, are absolutely thinkable in eventually converting into transient expression technology because it's so fast and so efficient. You don't need essentially create a clone, you just take a population of cells and transfect them with the DNA of interest.

So, if one could optimize this process and get the yields up a little bit better and also maybe reduce the quantities of DNA for operations in the, let's say, 100 to maximally 500 liter scale of operation, then I could imagine that transient gene expression could become, for protein delivery, very interesting model.

Ann Nguyen:

Are there promising glimpses into using transient protein gene expression for clinical use today?

Florian Wurm:

In the vaccine field, especially when it concerns virus vectors, transient expression is already used and is widely accepted already. So, we have a division, if you want, of our industry into the therapeutic field, where we treat patients with proteins and those other fields, where vaccines, which usually require very, very small doses, are readily made actually by transient gene expression because an AV particle, or a retrovirus, or an adenovirus can be essentially produced in the same way. Deliver DNA to a reactor and have the virus assemble itself and deliver the virus vector in this way.

Ann Nguyen:

You'll be discussing “Transient Protein (Gene) Expression: From R&D towards Pharmaceutical Manufacturing” during your keynote presentation on 14 November. What's the main theme you'd like to convey to your audience of protein engineers and researchers?

Florian Wurm:

Well, what I'd like to convey is that transient gene expression will continue to be a major driver for innovation. Clearly my own work many, many years back contributed to it. I was part of the first team to create a new molecule which was essentially designed based on transfecting hundreds of variants of a molecule into HEK293 cells and out of it came better thrombolytic, which was then rapidly approved. This happened already 30 years ago and all the new molecules we're looking at today, whether these are fusion molecules or really very uniquely structured molecules which take individual domains of proteins from different sources and put them together, will on the first level rely on transient gene expression to prove that actually a mammalian host can deliver such a product; because the key to everything is manufacturability.

If the biological system in the animal cell in the bioreactor cannot assemble the protein which you've designed in an efficient way, you will never have the product. So, the transient approaches give you the opportunity to look at these structures from all kinds of angles. Do all kinds of subtle modifications of your constructs. Minor, even single amino acid changes can have dramatic effects, and prove one thing which is most crucial for getting a drug to the market, which is can it be made in quantities which are sufficient to deliver the drug into the market and of course, into the patient first and deliver clinically relevant material for testing.

So, it will continue to be a major driver and maybe the only driver from an expression perspective because I don't think that any other system – bacterial or yeast or whatever plants – has this flexibility. The animal system and of course mostly now the CHO system has shown to be extremely modular, extremely responsive to all these new structures which we throw at them as the DNA molecule and has, so far, actually succeeded in most cases, to give us the protein molecules we wish.

Ann Nguyen:

Florian, thank you again for your thoughts.

That was Dr. Florian Wurm of the Swiss Federal Institute of Technology Lausanne and ExcellGene SA. He'll be speaking during the Optimising Expression Platforms conference at PEGS Europe, taking place 14-15 November in Lisbon.

To learn more from him, visit for registration info and enter the keycode “Podcast”.

This is Ann Nguyen. Thank you for listening.