Understanding Mechanisms of Beta-Cell Dysfunction using Genome Engineering in Human-Derived Induced Pluripotent Stem Cells

Type 2 diabetes (T2D) is a world-wide health concern, with its current prevalence increasing at a rate faster than the adult population growth rate. Our lack of progress in decreasing diabetes incidence reflects the complex interaction of a variety of genetic, environment and lifestyle factors that influence the risk of T2D. Regardless, the disease is typified by pancreatic insulin secretory failure and associated peripheral insulin resistance with the central hallmark being beta-cell dysfunction. This is demonstrated by the vast majority of T2D-associated genetic loci described as impacting upon insulin secretion rather than sensitivity. Monogenic forms of diabetes also primarily affect islet development and function. However, the development of our understanding in beta-cell biology is hampered by a shortage of appropriate cellular models: 1) rodent islets are substantially different to their human counterparts in features such as cell cycle regulation and ion channel composition 2) primary human islets can only be obtained via surgery and/or cadaveric donors, meaning a limited supply and 3) human beta-cell lines are in their infancy. Similarly, the inbuilt challenges of analysing gene function using over-expression and knockdown systems have made translation from novel genetic signals to biological insights and ultimately new therapeutics limited.

Recent progress has allowed the generation from induced pluripotent stem cells (iPSCs) to islet-like cells from any patient cell type, generating an important solution to the unmet requirement for more physiological-relevant diabetes cell models. This development, when combined with in vitro differentiation protocols, facilitates generation of endocrine pancreas lineage cells accurately recapitulating the dysfunction observed in patients. The parallel emergence of genome engineering tools such as CRISPR-Cas9 provides a tool for investigating gene function in an endogenous expression context, and a system for both introducing and correcting diabetes-causing genetic perturbations.

The aim of this studentship is to investigate mechanisms underlying beta cell failure via utilisation of iPSC-derived cell models of extreme diabetic phenotypes which contain both naturally occurring and artificially introduced mutations. In the first instance, monogenic forms of diabetes (both neonatal and maturity-onset diabetes of the young) caused by mutations in the insulin (INS) gene will be utilised as a means to probe the effect of aberrant gene regulation and defective protein processing on beta-cell function. The methods for investigating these pathologies are also directly translatable to more complex forms of diabetes, for example, the islet dysfunction observed in some INS-MODY individuals are also thought to occur in T2D.

The candidate will capitalise upon comprehensive techniques in genome engineering and cellular phenotyping, in addition to access to clinical data, diabetes patients, and primary human tissue at the University of Oxford, alongside expertise in iPSC culture and differentiation at Novo Nordisk. The candidate will gain skills such as:
1. Generation of iPSC lines harbouring monogenic diabetes mutations (affecting key features of islet function)
2. Utilisation of in vitro differentiation protocols to guide these mutated cells down the endocrine pancreas lineage
3. Characterise mutant cells using extensive phenotypic assays, and compare directly to data from wild-type cells and primary human tissue
4. Use the mutant cells in small molecule/compound screens with the goal to alleviate the observed pathophysiology. This element of the project will be in collaboration with the Target Discovery Institute (TDI) in Oxford, and use FDA-approved drugs
5. Translate these data and models into pipelines for studying more complex forms of beta-cell dysfunction, as occurs in T2D