Pancreatic cancer is one of the deadliest in humans – with a lack of good therapeutic treatments – and Dr. Siim Pauklin, principal investigator in the Nuffield Department of Orthopaedics, Rheumatology and Musculoskeletal Science at the Botnar Research Center in the UK, is hoping to reverse this trend with the help of his awarded Asteria™ scRNA-seq kits. The focus of his research is on a specific subset of cells within the tumors, called cancer stem cells, which might hold the keys to the lethality of the disease.
“Those cancer stem cells are very important because they are very metastatic, they can give rise to new tumors in distant organs, such as the lungs and liver,” clarifies Dr. Pauklin. “They can also be more resistant to currently used chemotherapies.”
As the cancer stem cells survive the chemotherapies, they multiply and cause new tumors. Therefore, Dr. Pauklin and his team are concentrating their efforts on finding novel therapies to get rid of them. However, it was never going to be simple.
“Pancreatic cancer is quite special in terms of structure and how it develops,” adds Dr. Pauklin. “It has a very dense stroma [the supporting structure around the tumor] in which there is a lot of collagens, meaning therapeutic compounds can’t access the cancer cells. But there is also another specific characteristic: there are certain signaling pathways and transcription mechanisms which specifically regulate those cancer stem cells, and to understand them is to find a way to fight them.”
The team has already made progress toward finding molecules that might inhibit those pathways and thus prevent the growth of those cancer stem cells. However, to faithfully test the effect of those molecules on cancer stem cells and the many surrounding cells, they need to be able to measure the gene expression data down to the resolution of the individual cell. And to achieve this, a sleek and dependable single-cell RNA-seq workflow is critical.
Probing the impact of therapeutic compounds with single-cell insights
“We know that there is a large diversity of cells in pancreatic cancer: cancer cells of course, fibroblasts, immune cells, as well as a large amount of others.” Dr. Pauklin continues: “That’s why it would be really important to use single-cell techniques in the project, because we would be able to say how each of those cell types responds specifically to our compounds”.
The team has in mind a specific pathway in cancer stem cells, called TGF-β, that they hope to disrupt with these compounds. It has been hinted to be involved in both cancer cell proliferation and metastatic abilities. Within this pathway, some molecules have grasped the interest of the researchers, but their exact nature still eludes them.
“We only know some basic things about those transcription factors: they seem to be involved in regulating epigenetic mechanisms in the nucleus, binding chromatin, and regulating gene expression. But we do not know precisely how this occurs, which genes are concerned, and which other molecules are recruited in this process,” explains Pauklin.
Thanks to the Asteria scRNA-seq kit, the researchers might be able to uncover more information about this pathway, and if it gets disrupted by their promising therapeutic compounds. Moreover, they’ll be able to measure potential side effects on the surrounding cell populations.
Time is of the essence: a scRNA-seq workflow to stay close to patients
The issue with measuring the gene expression of a cell is that it is highly dynamic and very time-sensitive, even more so when under the influence of an external molecule such as a therapeutic compound. Therefore, the researchers were looking for a workflow that would enable them to process their cancer samples immediately.
“Ideally, we really want to understand what’s happening in human patients, which is the closest to the goal of developing new therapeutic strategies,” confirms Pauklin. “That is why we really want to use fresh patient samples to get the best source and understand how the cells really function in this situation and how we can impact them, as close to the in vivo situation as possible, and investigate our compounds further on those cells.”
The team has previous experience with single-cell techniques but was drawn to the accessibility of the Asteria technology and its lack of requirements for microfluidics, freezing, or fixing compromises coming from other methods. Moreover, its ability to handle fragile cells might come in very handy.
“Pancreatic cancer has this dense extracellular matrix, so we must go through drastic dissociation methods to get the sample into a single-cell suspension. During this process, cells may get potentially damaged, and come out quite fragile. Cells in pancreatic tumours also tend to have relatively high necrosis because there is hypoxia, inflammation, etc., so this makes it even more difficult for pancreatic cancer to be studied.”
With their new kits and the sequencing costs covered by the grant, Dr. Pauklin’s team is finally well-equipped to tackle the challenge at hand and test the success of their therapeutic compounds onto those resilient pancreatic cancer stem cells, one cell at a time!