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Development of a platform for somatic cell reprogramming to iPSCs in microfluidic environment

Supervisors:
Jane Sowden, Paolo De Coppi and Marco Pellegrini

Project Description:

Background: Induced Pluripotent Stem Cells (iPSCs) have become pivotal in translational medicine, offering exciting prospects for research and therapeutic applications. Their unique ability to differentiate into any cell type within the body positions them at the forefront of disease modelling, drug discovery and testing, and personalized cell-based therapies. Unfortunately, the generation of iPSCs is hindered by high costs and inefficiencies. Many protocols, particularly virus-based, are poorly suited for clinical translation. Moreover, the common practice of using fibroblasts as source of somatic cells for reprogramming involves invasive skin biopsy procedures in children and can pose additional and unnecessary risks for patients with conditions such as epidermolysis bullosa.

Aims/Objectives: Our project aims to demonstrate the effectiveness of microfluidic (μF) reprogramming technology in generating iPSCs from patient-derived somatic cells, extending beyond the conventional use of fibroblasts to include more challenging cells derived from urine and blood samples. This approach seeks to offer a cost-effective, patient-centric method to produce the necessary cells for innovative therapeutic applications. The research will lay the foundation to create an iPSC service for GOSH and a biobank of patient-specific iPSCs derived from patients with rare diseases.

Methods: In recent years, a novel protocol utilizing mRNA and a μF environment has emerged, offering a safer and more efficient pathway for iPSC generation1,2. This method circumvents the risks associated with viral vectors by using mRNA to deliver reprogramming factors, avoiding potential genomic integration issues. The μF system further enhances the interaction between cells and reprogramming factors, leading to a significant increase in efficiency compared to conventional culture conditions (CCC) (i.e., wells and plates). Compared to CCC, the μF technology has demonstrated a 50-fold improvement in reprogramming efficiency while simultaneously reducing the quantity of somatic cells required. This approach also shows a 100-fold decrease in the cost of raw materials. These features of μF technology could enable the use of cell sources obtained by less invasive procedures than fibroblasts, such as urine cells (UCs) and peripheral blood mononuclear cells (PBMCs). These cells can be collected through non-invasive methods3 or routine medical procedures, offering a patient-friendly approach to cell sourcing. However, the yields from these alternative sources currently don’t match those obtained from fibroblasts.

Timeline:
1.    Testing of different solutions of μF devices (months 0-12):
In collaboration with Prof Nicolas Szita (UCL) and the Making Lab (Crick), new μF solutions will be tested.
2.    μF device validation in comparison to benchmark reprogramming system (months 7-12):
In partnership with the Francis Crick Institute iPSC facility, μF reprogramming will be compared to mRNA reprogramming in CCC. 
3.    Derivation and reprogramming in μF of patient cells derived from low invasive sources (months 13-36):
A protocol for μF reprogramming of PBMCs and UCs from patients with rare diseases, as part of the NIHR GOSH BRC iPSC resource programme, will be developed and compared with CCC reprogramming.

References:
1. μF reprogramming for the generation of primed iPSCs: http://www.nature.com/articles/nmeth.3832
2. μF reprogramming for the production of naïve iPSCs: http://www.nature.com/articles/s41556-018-0254-5
3. Cell Catcher tool for efficient isolation of UCs: https://www.medrxiv.org/content/10.1101/2023.09.22.23295442v2


Contact Information:
Marco Pellegrini and Jane Sowden