Optimising islet transplantation therapy by targeting the liver niche to promote the long-term engraftment and function of human islets.

Diabetes mellitus is one of the most common chronic conditions affecting 7% of the population. Approximately 15% of people with diabetes have Type 1 diabetes and are dependent on insulin injections to control blood glucose levels and due to inject insulin requirements, over 25 percent of people suffered repeated episodes of low blood glucose readings requiring help (severe hypoglycaemia) and at the time the warning symptoms associated with the hypoglycaemia (shakiness, dizziness sweating) are lost putting their lives at risk.

Islet transplantation whereby islets which contain insulin secreting cells are purified from donor pancreases and transplanted into subjects with Type 1 diabetes, consistently and effectively stabilises blood glucose control reducing hypoglycaemia iand saves lives. Our NHS funded islet transplant programme for Scotland was established in 2009 and since this time we have done 60% of the transplants in the UK. Our transplanted patients have had fewer episodes of hypoglycaemia, regained awareness of hypoglycaemia and experienced more stable blood glucose control With significant reductions the amount of insulin they inject and reported improved quality-of-life. Additionally there is evidence of delayed progression from diabetes related complications.

However patients require islets from at least 2 donor pancreases: islets do not have a blood supply when first transplanted and over 60% of islets are lost as they are susceptible to dying before a blood supply is established. There is also a gradual deterioration in islet graft function over time; pancreases with marginal numbers of islets cannot be used because they will not impact on blood glucose control and patients are currently awaiting more than one
the national waiting list. Strategies to improve the efficiency of islet transplantation are urgently needed.

Growth factors including vascular endothelial derived growth factor (VEGF) can be used to promote blood vessel formation and have been utilised in different ways to repair blood vessels experimentally and now programmes have taken these factors forward into clinical trials. Keratinocyte Growth Factor (KGF) is another growth factor, licensed for specific use in man.

Our laboratories at the University of Nottingham and University of Edinburgh have generated important experimental data. Firstly human KGF can be formulated into specific particles which target the liver. Using a mouse model we have shown that these particles can promote multiplication (=proliferation) of cells within the liver with no other cells in other organs demonstrating proliferation. Using a mouse model of Type 1 diabetes we have transplanted mouse islets with these particles loaded with KGF into the liver and have shown that this results in greater retention of the islets in the liver as well as better blood glucose control in the short and longer term with greater cure rate of diabetes versus those mice that received the islets alone. We are currently examining the islet transplant site to investigate if there are more blood vessels in it, which would support the concept that KGF promotes not only cells within the liver to proliferate but also that it causes blood vessel formation between the islets and the liver.

Significantly these particles can be used in man.

The aim of these experiments is to test these particles with human islets,(human islets isolated locally) and determine whether transplanting the particles loaded with KGF along with the islets into the liver of a mouse produces benefit in terms of controlling blood glucose levels in the long term leading to transplants which would function for longer. We will perform studies to understand how these particles work and will track the particles by sensitive imaging techniques to confirm where they target. These studies will potentially lead to first-in-man studies allowing more people with diabetes to be treated.

Islet transplantation consistently restores awareness of hypoglycaemia and stabilises blood glucose control. Unfortunately there is a limited supply of donor pancreases with most recipients requiring 2 donor pancreases. Islets are avascular when transplanted and rely on the process of angiogenesis for engraftment:>60% of islets are destroyed in this avascular state.
We have developed particles from the polymer poly(DL-lactide-co-glycolic acid)(PLGA): a biodegradable material used in pharmaceutical formulations in which growth factors can be loaded and their release controlled. The polymer has a galactose moiety attached to it and targets the liver through exploiting the asialoglycoprotein receptor(ASGPR), specific to liver. Keratinocyte growth factor (KGF) is a small polypeptide that binds to the KGF receptor and has proliferative and anti-apoptotic effects on epithelial cells eg. hepatocytes. We have determined the KGF particle dose which in mice promotes proliferation of cells within the liver, increases islet engraftment (histologically) and significantly improves glycaemic control when transplanted with islets via the hepatic portal vein. VEGF particles are also in development. In this proposal the objectives are to: 1. Refine/develop growth factor particles to target liver with appropriate growth factor release kinetics; 2. Demonstrate /confirm efficacy of human islets co-transplanted with these growth factor-particles; 3. Understand the mechanism of action of these growth factors.
To answer these objectives we plan to: 1. Refine the release kinetics of KGF from galactosylated KGF-PLGA particles already developed; 2. Further develop VEGF-PLGA particles to target the liver; 3. Assess fate/track particles; 4. Assess short term effects of particles on liver; 5. Assess long-term metabolic effects of particles on islet function; 6. long term negative or positive effect of particles on islet graft.
If successful, we will progress to first in man studies.

Below we describe who will benefit from this research and how:

Patients with Type 1 diabetes and potentially Type 2 diabetes: patients are enthusiastic about islet transplantation and this research will determine if co-transplantation with specialised biodegradable particles releasing growth factors for a short period of time, is feasible. It will determine if such formulated particles can impact on short term and also long term glycaemic control with the need of fewer transplants. Although islet transplantation is an effective therapy for people with Type 1 diabetes it is not efficient. Following islet isolation the islets do not have a blood supply and it takes over 7 days for blood vessels to form between the islets in the liver - the liver is the site of transplantation. Following transplant the islets are vulnerable to loss as they do not have a blood supply and over 60% of islets are therefore lost from apoptosis, necessitating the use of 2 to 3 donor pancreases per recipient. If the treatment of co-transplantation of growth factors with islets is successful, the societal impact would be large as it would mean that one donor pancreas may be transplanted into one recipient and that would be more donor pancreases to treat more people with Type 1 diabetes. At present people listed for islet transplantation are waiting over one year on our national waiting list. At present islet transplantation is limited to those people with Type 1 diabetes who have severely impaired awareness of hypoglycaemia. However if more pancreases were available for islet transplantation purposes, islet transplants may be applied to those adults with Type 2 diabetes. Furthermore if this approach of utilising growth factors targeted to the liver was successful then other approaches may be adopted using other pharmaceuticals directed to the liver including delivery of immunotherapy directed to the liver.

Islet transplantation researchers including diabetologists and transplant researchers and researchers is in the field of regenerative medicine. This project will answer the question as to whether growth factor releasing particles specifically targeting the liver are useful in improving islet graft function versus islet transplant alone. We will share techniques that will ensure that the particles are available for for use by other researchers including in the wider context of the UK RMP. These researchers may be in a variety of fields of disease. This will be of interest both the public and private sector researchers. (See letters from Professor Shakesheff and Professor Stuart Forbes).

Private sector companies e.g. pharmaceutical companies. Such companies may apply their pharmaceutical compounds to this field.

Local national and international policymakers for determining best use of funding: there is interest as to whether the addition of growth factors to particles in co-transplantation with islets are worth funding for future therapies for diabetes.