Single Cell Transcriptome Profiling for Advanced Cell Therapy Manufacturing Process Monitoring

"Our project is a collaboration between ReNeuron Ltd, a well-established British stem cell therapy company and the Department of Advanced Therapies at the National Institute for Biological Standards and Control. ReNeuron has created an immortalised neural stem cell line called CTX, which is presently undergoing clinical trials as a transplant treatment for stroke and ischaemia (damage to the limbs similar to that caused by stroke in the brain) and also for exosomes (nano-scale particles produced by the CTX cells) as therapeutics in their own right. Stem cells such as CTX are capable of differentiating into other cell types found in the body; this is why there is so much interest in them as potential therapeutics for many otherwise hard-to-treat conditions. However, this presents difficulties: unlike a preparation of a standard small molecule drug, each preparation of stem cells is unique, being a constantly changing, dynamic living thing that responds to its environment. This presents challenges with confirming the efficacy of each cell population, and with ensuring that changes to manufacturing protocols, for example, when they are scaled up to produce large cell batches of a size suitable for treating significant numbers of patients, do not introduce untoward changes in the cell populations.

It is now possible, using a technique called scRNAseq (single cell RNA sequencing) to identify all of the genes expressed in a single cell, for large numbers (hundreds to thousands) of individual cells. This allows us to create the most fine-grained profile of a cell population, identify subpopulations in the stem cell population, and explore differences between different batches of nominally the same cell type prepared at different times or by different methods. We will compare our CTX neural stem cell line with similar sister cell lines, human ""pluripotent"" stem cell lines including reprogrammed CTX cells, and stem cells of different types such as mesenchymal stem cells (MSCs) derived in turn from CTX-derived pluripotent stem cells. Cross-comparing these results with current assays for therapeutic activity or phenotype will allow us to distinguish the therapeutic subpopulations from ""passenger"" cell types, to rationally optimise protocols for cell preparation, and most importantly confirm unequivocally that our stem cell therapies are safe and effective to the patient."