Cryo electron microscopy in pharma drug discovery
Cryo electron microscopy (cryo-EM) is a rapidly growing technique in the pharmaceutical industry due to its ability to accurately and rapidly visualize the intricate interactions between drug and receptor, enabling informed, accelerated drug discovery and design. Pfizer has been leading the charge, establishing one of the first cryo-EM facilities in the industry in 2016. We spoke with the head of Pfizer’s facility, Dr. Seungil Han, about their recent publication in Nature Communications and how cryo-EM has fundamentally impacted their research.
This interview has been edited for clarity and length.
How did you first get involved in pharmaceutical research?
I was trained as crystallographer, so I was always interested in structure-based drug design. At the time, X-ray crystallography was the primary tool, which is what I used to drive drug discovery.
How did you then transition to cryo-EM?
We were already interested in this technology when we established an academic collaboration more than seven years ago where we used negative-stain EM. It was then that we saw the value of EM. Additional academic collaborations and training/experience at multiple EM facilities ensured us that it would be a critical tool to have in-house.
These fruitful collaborations & experiences in other fee-for-service EM facilities opened our eyes to the possibility of electron microscopy to advance our drug discovery programs. It also provided us with the requisite training and we slowly built up our group and capabilities since then. Eventually, we established our cryo-EM facility in October 2016.
Can you talk a bit more about how cryo-EM has influenced your drug discovery process?
We’ve had impacts on small molecule, vaccine, gene therapy and protein degradation programs within Pfizer. Using this exciting technology platform, we have been successful in providing proprietary insights into some research programs and therapeutic modalities, especially when it comes to molecular binding sites. The cryo-EM method is well-suited for providing critical structural information to influence the discovery of biologics, vaccines and gene therapies. However, the most surprising aspect has been our ability to provide binding site and binding pose information for small molecules on large proteins or protein complexes.
Let’s talk about your article in Nature Communications. Could you give us a brief overview of the project?
Iron sulfur cluster proteins are very important; they are used in many biological systems, vitamins and other co-factors. Many diseases are also associated with iron sulfur protein from cancer to anemia and myopathies.
Our research focused on the frataxin protein associated with Friedreich ataxia, a devastating autosomal recessive genetic disease that lacks a cure or therapy. Frataxin deficiency leads to iron overload in mitochondria with consequent adverse effect on cellular proteins.
We were particularly interested in the major machinery of the iron sulfur assembly complex and solved the structure of this five-protein complex, which we called the SDAUF complex. This allowed us to elucidate how frataxin activates the iron-sulfur complex and essentially provided a new avenue for drug discovery.
Why was cryo-EM so essential to this study rather than traditional techniques like X-ray crystallography?
That’s really the essence of this paper. Two academic groups had previously solved the crystal structure of a portion of the SDAUF complex. The two groups arrived at two different structures, even though they used the same proteins. Additionally, based on these structures, it wasn’t clear how the fifth protein, frataxin, would turn the complex on.
So which X-ray structure was more appropriate? In the end, the cryo-EM technique came to the rescue and provided critical insights into the activation mechanism.
We tried crystallization but found that it didn’t work because frataxin’s binding affinity is very marginal. It has micromolar binding affinity for the rest of the complex. However, we managed to shift the equilibrium to the five-protein complex and, using cryo-EM, determined the structure of the complex. It turned out that one of the previously published crystal structures was more consistent with the topology of our five-protein complex.
What is the most important thing for scientists to know if they want to get started with cryo-EM?
That’s a tough question. In general, I think there are increasingly more core and national facilities, so microscope time is becoming more available. However, the most important thing is to understand your biological sample. If your sample is heavily aggregated and heterogenous and you don’t know much about it, it’s like garbage in and garbage out. Don’t expect cryo-EM to solve the problem of a poorly-behaved protein sample.
Second, there is still a learning curve for cryo-EM, even though it has been getting easier over last few years. You need to learn how to prepare the grid, how to prepare for data collection, how to process the data, and there’s no one established protocol. You still have to know all the fundamentals, just like any other scientific endeavor.
The trick is to find the ideal protein that behaves well in solution and on cryo-EM grids and is suitable for EM studies. In reality, each sample will be uniquely challenging and there is currently no automated, well-established protocol like X-ray crystallography. Even though cryo-EM is advancing rapidly, there’s still a learning curve.
What do you see in the future of cryo-EM? How does the technique need to adapt to get us there?
A couple of things. One is that the technology will become more automated, making it easier to use and providing more robust protocols from beginning to end. Hopefully that’s where it’s going, and that’s something Thermo Fisher could help with.
Second, cryo-EM has multiple applications: not just small molecule drug discovery but also biologics, vaccines, gene therapy, protein degradation and many others. I look forward to seeing cryo-EM applied to a variety of fields.
I’m very excited. Pfizer was the first to adopt this technology in-house, and we’re serious about using this technology for discovery. We have already seen the impact; some results are already in the clinic and even more publications are coming up in the future which I’m sure will excite the field.
Read more about Pfizer’s latest applications of cryo-EM in the following articles:
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Alex Ilitchev, PhD, is a science writer at Thermo Fisher Scientific. Seungil Han, PhD is an Associate Research Fellow at Pfizer Inc.