Genetic and nutritional control of pancreatic beta cell identity.

Diabetes mellitus affects more than 20 m Europeans and 400 m individuals worldwide. The complications of the disease, including blindness, kidney failure, cardiovascular disease and cancer, drastically reduce quality of life, and consume almost 10 % of health care costs in most westernised nations. These figures are expected to increase further in coming years. The most common form, Type 2 diabetes (T2D), has both genetic and environmental causes, and is particularly prevalent in those affected by over-nutrition and obesity.
Therapeutic approaches towards T2D have relied in the past on enhancing the actions of insulin, responsible for lowering blood glucose levels, and on stimulating insulin secretion. However, none of the existing therapies reverse the progressive loss of normal beta cell identity and function and hence the gradual worsening of disease symptoms.
"Genome wide association studies" (GWAS) for T2D have now identified numerous genetic variants whose inheritance is associated with an increased risk of diabetes. The identification of these genes, most of which influence insulin production, provides both improved powers of prediction and, just as excitingly, potential new molecular targets for drug treatment.
More than 100 hundred genetic loci have now been identified which collectively harbour almost 500 genes. Our work seeks firstly to determine which of the genes in selected loci are responsible for increased disease risk. This involves both genetic studies in man, and functional analyses based on studying the impact of deleting a particular gene from the disease relevant tissue - usually the pancreatic beta cell.
We have shown that a changes in the expression of a gene termed STARD10, which is able to bind fat molecules (lipids) within the cell and carry them between discrete intracellular locations, is responsible for the increased diabetes risk observed in carriers with a specific set of genetic variants on chromosome 11. At present, however, we have very little idea how this gene affects the cell's metabolism to impair the release of insulin. Understanding this question is important since it may provide new ways in which to improve the production of the hormone in those individuals (more than 80 % of the population) who are at increased risk of diabetes thanks to carrying the risk variant of this gene.
We will therefore perform cellular analyses using human islets, human-derived beta and beta-like cells, the latter produced in the test tube from embryonic stem cells, to determine the impact of deleting STARD10, and to understand how the variants associated with disease risk alter the expression of this gene.
The second Aim of our studies is to understand how two gene products, LKB1 and AMPK, are able to regulate pancreatic beta cell function. We know that deleting either gene in the mouse beta cell leads to a change in cellular identity, leading to the up-regulation of other genes which are not normally expressed in the islet but present at high levels in nerve and liver cells. AMPK, which is itself regulated by LKB1, is of particular interest since this enzyme is controlled by nutrients including glucose. We will determine whether changes in the activity of either enzyme affect gene expression by prompting changes in the structure (opening or closing) of nuclear DNA. We will also determine the impact of small molecule AMPK activators, which hold therapeutic promise in diabetes, on beta cell function.
Our final Aim is to determine whether the role of STARD10 in controlling beta cell function may be altered in the absence of LKB1, a phenomenon we have recently described for another GWAS gene, TCF7L2, or by changes in nutritional status.
We will use novel and powerful technologies including genome editing, directed differentiation of human embryonic stem cells, mouse genetics, photopharmacology and imaging of the islet after engraftment within the mouse eye, to answer our questions.

Type 2 diabetes (T2D) is an epidemic of the 21 st century and consumes almost 10 % of the health care budgets in westernised societies. Progressive pancreatic beta cell failure, characterised by altered gene expression and cellular "identity", are central to disease development. GWAS for T2D have now identified more than 100 loci associated with disease risk, most of which affect insulin secretion rather than action. In an effort to provide new targets for disease treatment, our laboratories have taken a functional genomic approach to identity the causal gene(s) at selected loci, to dissect their mechanisms of action at the molecular level, and to understand how they interact with other genes and environmental risk factors including over-nutrition.
STARD10, located at a T2D locus on chromosome 11q, encodes an intracellular lipid binding and transfer protein whose expression is decreased in the beta cell by possession of risk alleles. Under Aim 1, we will explore the molecular mechanisms through which STARD10 affects insulin processing and secretion in human beta cells and human embryonic cell stem-derived beta cell lines deleted for the gene or nearby regulatory elements using CRISPR/Cas9-medated gene editing. In Aim 2 we will explore the mechanisms through which the protein kinase LKB1, and its downstream substrate AMPK, a nutrient-sensitive protein kinase, control beta cell identity, focussing on epigenetic changes and chromatin remodelling. We will also explore the effects of small molecule AMPK activators on insulin secretion in vivo, and develop a photactivatable AMPK regulator to achieve local control of the enzyme in the pancreas. Aim 3 will determine whether the actions of STARD10 on beta cell function are altered by deletion of LKB1, using both conventional mouse genetics and in vivo imaging of islet function after engraftment into the mouse eye.

The research proposed here is likely to benefit both the general population, in terms of improvements in healthcare, as well as the UK and European Pharmaceutical industry.
1. The general population of the UK and the EU. Type 2 diabetes (T2D) affects ~4 m UK subjects and ~30 m Europeans (mean prevalence 9.1%; https://www.idf.org/our-activities/advocacy-awareness/resources-and-tools/13:diabetes-atlas-seventh-edition.html ). These values are predicted to grow further in a diabetes "epidemic" driven by increasingly sedentary lifestyles and obesity. The complications of the disease include stroke, retinopathy, neuropathy, renal failure, cardiovascular disease and cancer. The increased prevalence of this disease contributes to a ~10 year lowering in overall life expectancy in the UK (http://www.diabetes.co.uk/diabetes-life-expectancy.html). Treatment of diabetes is estimated to cost ~£8000 per year per patient, or £ 24 billion in total: diabetic patients are 3.5 times more likely to be admitted for hospital treatment than the rest of the population (http://www.physorg.com/news151077389.html). These direct economic costs, together account for 7-13 % of health care costs in most developed societies (IDF Diabetes Atlas, 2003), and are further aggravated by increased absenteeism and decreased individual productivity (ADA: Diabetic Care 31, 596, 2008).
Pancreatic beta-cell dysfunction and de-differentiation ("identity loss") are now thought to be cardinal elements of T2D, and strategies to rejuvenate or replace these cells, as explored here, are likely to be key to the development of new therapeutic approaches for the disease and its complications. Importantly, by examining gene-gene and gene-nutrient interactions the work is likely to facilitate the leveraging of GWAS data towards eventual therapeutic benefit. In addition, the proposed Programme address roadblocks in diabetes research as identified by the European Commission's Support Action "DIAMAP: A Road Map for Diabetes in Europe" (http://www.diamap.eu/roadmap/roadblock/?page=2) including "a lack of appropriate models that mimic the human condition (R3.07)" and "Lack of animal models to sufficiently mirror human disease (5.05)".
Finally, these studies directly address the recently-announced MRC PSMB priority for nutrition research (https://www.mrc.ac.uk/funding/science-areas/population-systems-medicine/mrc-priorities-for-nutrition-research/).

2. The UK Pharmaceutical Industry. The global market for anti-diabetes drugs is estimated to be worth ~$31 billion and is set to double to $58 bn (http://www.drug-dev.com/Main/Back-Issues/Global-Type-2-Diabetes-Market-Set-to-Almost-Double-1140.aspx). New drug targets and leads are desperately needed for the Pharmaceutical industry to produce novel diabetes treatments. By addressing highly promising new targets, including those identified by GWAS screens, and processes (e.g. control of cellular identity) the proposed study will enhance feeds of new Intellectual Property to this sector.

As a previous work package (WP) leader in the trans-European Initiative Medicines Initiative (IMI1)-funded diabetes research network "IMIDIA" (http://www.imidia.org/), and current WP leader in the IMI2/Horizon 2020-funded project "Rhapsody" (https://imi-rhapsody.eu/), GR has extensive collaborations with several UK, Europe, and US-based companies (eg Astra Zeneca, Servier, Janssen, Novo Nordisk, Sanofi Aventis, Boehringer-Ingelheim, and Novartis). The Rhapsody consortium also involves close interactions with major European and US prediabetes and T2D cohorts and multiple -omics platforms within the Pharmaceutical industry. These will facilitate the work on mouse tissues proposed in the CfS.
Each of the three research fellows and technician directly involved in the project will enhance their professional skills with training in basic biomedical research, and thus develop their skill set for application in both the academic and commercial sectors.