An interview with Professor Steven Pollard, University of Edinburgh

In this blog post, we spoke with Professor Steven Pollard, Group Leader at the Centre for Regenerative Medicine and Edinburgh Cancer Research Centre, at the University of Edinburgh. We discussed some of his latest work to understand the biology of Glioblastoma and explore how microfluidic technology can overcome current challenges in the genome editing workflow.

Current work using CRISPR/Cas editing technology

Our research focuses on understanding the fundamental mechanisms in stem cell biology and how they are deregulated in brain tumors, such as Glioblastoma. In order to uncover novel biology and reveal new therapeutic strategies to treat this complex and poorly understood disease.

Many of our studies are conducted on primary cell cultures using genome editing techniques, such as the CRISPR/Cas system. Leveraging this versatile technology, our lab can modify key genes and cellular pathways, perform functional experiments to examine their involvement in the protumorigenic processes and identify if they are a good therapeutic target.

In some of our latest research, we are engineering normal brain stem cells – without the brain tumor mutations – that drive the disease, then introducing the driver mutations stepwise, and in combinations, to watch the cancer program unfold. We are also performing live-cell imaging, using knock-in reporters to tag and track proteins of interest at endogenous levels. This knock-in gene targeting allows us to study where the protein is localized during the cell cycle, monitor the levels of a protein in live cells, and determine post-translational modifications.

The development of the CRISPR/Cas system has undoubtedly brought us closer to dissecting cell biology and advancing functional genomics. This multi-functional genome editing technology has also revolutionized biochemistry, allowing researchers to genetically manipulate cells to tag a protein, study the protein in vivo, identify their binding partners and learn about their biological roles. The ability to do this at scale using disease-relevant human tumors is having a profound impact on the field.

Overcoming challenges in genome editing

Alongside our research, our lab invests a lot of time in optimizing the genome editing process to overcome time-consuming technical bottlenecks and move away from the traditional "gene-by-gene" approach. Conventional genome editing methods require expensive instrumentation and highly trained technicians to perform the manual, multi-step process of growing up the cells, transfecting them to deliver the reagents, then screening, isolating, and expanding the edited cells.

Microfluidics offers many workflow advantages, such as improved throughput, speed, reduced reagent costs, and hands-on time. This expertise led to our interest in collaborating with Sphere Fluidics. Using microfluidics for high-throughput single cell screening technology, researchers can rigorously dissect a whole pathway or a gene family in parallel, rather than studying just one or two candidate genes at a time. This ability to gain richer information and more functional data is a massive benefit, providing greater confidence in the identified gene candidates while offering in an unbiased manner the potential for novel insights and discoveries as we comprehensively annotate the human genome.

Simplifying genome editing with microfluidics

Continuous innovation and collaboration are crucial to scaling this vision, as demonstrated in the Open Innovation Grant by the University of Edinburgh, Sphere Fluidics, Horizon Discovery, OXGENE, and Twist Bioscience. Funded by Innovate UK, this collaborative grant focused on making genome editing more efficient using microfluidic technology.

Sphere Fluidics has prototyped a benchtop device that integrates and miniaturizes the multi-step process for creating genome-edited cell lines, from transfection to validation of the edited cell. The convenient device provides a high-throughput, hands-off method to deliver the CRISPR reagents or the repair reagents, isolate healthy cells, confirm if individual cells have been edited, and automate single cell plating to grow colonies of clones. The microfluidic workflow also minimizes reagent usage and eliminates additional instrumentation, like flow cytometry, further reducing costs. Having reached the end of the grant, it will now be interesting to use the prototype for some pilot projects. If we can repeat some historically older studies in timeframes of a week, using the same reagents, the device will save us years of work, which will be a tremendous outcome.

The future potential and impact

While many believe CRISPR technology has democratized genetics, there are still limitations in throughput and costs that are restricting many applications. The development of integrated methods that circumvent these barriers, such as Sphere Fluidics' device, will improve the throughput, reliability, and reduce the cost of genome editing to facilitate wider adoption.

Over the next few years, these fully-integrated devices will enable a range of applications to be performed at the push of a button, empowering researchers and allowing them to focus on the biology and hypotheses. Aside from the new fundamental discoveries in genetic and cellular mechanisms, this will also impact biopharma research. Drug discovery is one important application, as these tools for cellular genetics can be deployed to confirm target identification, explore the mechanism of action and combine genetic and chemical screens to fast track drug discovery and development efforts.