Purification of Stem Cell Derived Insulin Producing Beta Cell
Creating tools for purification of insulin producing beta cells.
Definition of the problem: Despite advances in diabetes therapies, the leading treatment for people with type 1 and (occasionally) type 2 diabetes continues to be daily injections of exogenous insulin. Type 1 diabetes is the result of autoimmune destruction of β-cells, which are the insulin-producing endocrine cells of the pancreas. On the other hand, the more common type 2 diabetes results from insulin resistance and β-cell dysfunction. Loss and dysfunction of insulin-producing β-cell is the key pathological event in diabetes. Patients suffering from type 1 diabetes could be potentially cured through β-cell transplantation or even whole pancreas transplantation from a suitable donor. This method has the potential to bring down the glucose concentration to a normal level without insulin injection and has already been performed on patients with positive clinical outcomes.1, 2 However, this process is very limited due to the availability of suitable donors. Stem cell-derived β-cell formation has given a new hope to this problem. Human embryonic stem (ES) cells are pluripotent, which means they can be converted to any of the body’s cell types, including β-cells. It has been successfully demonstrated that human ES cells or induced pluripotent stem (iPS) cells can be converted to fully functional β -cells, which can release insulin upon glucose stimulation. These ES or iPS-derived β -cells have been successfully transplanted into experimental animals.3, 4 However, the major bottleneck of this process is the overall yield of β-cell production – the typical yield of β-cell formation is very low. Furthermore, this method also produces other undesired endocrine cells and also contains undifferentiated iPS cells which could be tumorigenic. There is no available method to isolate these β-cells from the other pancreatic cell types or non-transformed cells. This proposal seeks to develop a method to isolate β-cells from other cell types including untransformed iPS cells and other pancreatic cells. In this process, it would be possible to collect β-cells from multiple batches with high purity, which can then be used for clinical application.
Aim 1: Separation and purification of stem cell-derived β cells using magnetic nanoparticles. The core of our research strategy centers on our ability to chemically functionalize the magnetic nanoparticle Fe3O4 with GLP1-like peptides for direct labelling of β-cells. Fluorescence-activated cell sorting (FACS) is a widely used method for separating various types of cells from a mixture upon labelling with specific fluorescent antibodies. This low number of cells separated with this method may be useful only for laboratory purposes but cannot be translated to clinical application at a later stage. We intend to functionalize magnetic Fe3O4 magnetic nanoparticles with small molecule agonist of GLP1R. These small molecules will have much higher plasma stability as compared to the GLP1 and its analogues, which are prone to degradation by dipeptidyl dipeptidases-IV (DPP-IV). Some of the small molecule GLP1R agonists are having a comparable affinity with native GLP1. These agonists will be functionalized to the magnetic nanoparticles with a suitable photocleavable linker. After selective binding to the β cells, the cells can be detached from the magnetic nanoparticles after shining an appropriate wavelength of light.
Image courtesy of Debasish Manna.