Sugar rush - pig cells for diabetes treatment
Each day, more than 34 million people rely on regular insulin injections to stay alive. Peter Hosking, Head of Operations at Living Cell Technologies, unveils a new treatment product that could change the lives of diabetes sufferers worldwide.
Combine a herd of healthy pigs with a new cell-rejection prevention technology, and what do you get? While some might suggest a bacon sandwich minus the indigestion, these two components were in fact critical for the development of a breakthrough treatment for Type 1 diabetes by Living Cell Technologies (LCT), a cell implant company based in New Zealand. The company’s acquisition of a unique virus-free pig herd sourced from the remote sub-Antarctic Auckland Islands, along with the advancement of an encapsulation process that eliminates the need for immunosuppressant drugs, has put LCT at the forefront of cell transplantation research.
Diabetes is a chronic disease characterised by high blood-glucose levels resulting from the body’s inability to produce or respond appropriately to sufficient insulin. Type 1 diabetes occurs when the pancreas is unable to produce sufficient insulin for the body. Affecting around 34 million people worldwide, the disease requires insulin replacement therapy to stabilise blood glucose levels. LCT’s work to treat diabetes involves placing cells called islets – natural aggregates of cells that produce insulin as well as glucagon – into the body.
Saving your bacon
The pig breeding facility, from which the pancreatic islets for the new drug are obtained, runs a closed, designated pathogen-free (DPF) herd. Pigs from the Auckland Islands essentially had no contact with other pigs or humans for about 150 years, and have subsequently been bred in isolation in New Zealand. Their origin and isolation has protected them from porcine viruses.
Possible sources of zoonotic infection are closely monitored to ensure the pigs are free of all specified diseases. This also ensures that the production processes for the drug meet all safety standards for xenotransplantation – a procedure involving the transplantation, implantation or infusion into a human recipient of either a) live cells, tissues, or organs from a nonhuman animal source, or b) human body fluids, cells, tissues or organs that have had ex vivo contact with live nonhuman animal cells, tissues or organs.
Transplant without rejection
The vulnerability of transplanted islets to the recipient’s immune system has been a major scientific barrier to successful islet transplantation. The transplanted cells face not only rejection but also immune attack from a variety of cells, which may result in loss of function. Two approaches have generally been adopted to overcome this:
- the concurrent administration of immunosuppressive drugs, which are not always effective in altering the course and incidence of rejection episodes and can cause adverse effects, both in the recipient and the transplanted cells
- ‘immunoprotection’ of the transplanted cells via the use of a semipermeable membrane that acts as a protective barrier
The principle of the latter lies in the permeability of the membrane. While allowing smaller molecules such as glucose and other nutrients to penetrate it and reach the islets, and insulin to be released out into the bloodstream, the membrane inhibits the passage of large immune cells or antibodies that would cause rejection of the islets.
The means to achieving this immunoprotection has been advanced by LCT’s proprietary cell encapsulation technology. The procedure involves extruding a mixture of islets and an ultra-pure sodium alginate solution through a droplet-generating needle into a bath of gelling cations (calcium chloride). The islets entrapped in the calcium-alginate gel are then coated with poly-L-ornithine (PLO), followed by an outer coat of alginate. The central core of alginate is then liquefied by the addition of sodium citrate.
The encapsulated islets, measuring 650–750μm in diameter, are incubated in culture medium in cell-culture flasks with media changes every two to three days as required. The materials used in the encapsulation process are polyelectrolytes, which consist of linear or branched repeat units each carrying a negative (alginate) or positive (PLO) charge. Alginate is manufactured as a sodium salt, which is displaced during the crosslinking process. The formation of the alginatepolyornithine-alginate (APA) permselective membrane is achieved through the sequential exposure of oppositely charged materials, in a process designed to minimise exposure of stress to the islets within the capsule. This is accomplished by carefully selecting and purifying the bulk polyelectrolytes used to form the physical barrier, alongside an efficient and robust encapsulation process.
The role of calcium is an intermediate yet critical component of this process. Initially, crosslinking of alginate by divalent cations occurs between guluronate (G) residues due to their 1–4 linkages, which orient the monomers such that hydroxyl and carboxylic groups can participate in ionic bonding. The crosslinking serves to solidify the sphere and, while the majority of bound calcium is retained within it, cations at the surface undergo dynamic and reversible binding in the presence of coating and chelating agents, PLO and sodium citrate. Owing to the anisotropic nature of the crosslinking process, a theoretical gradient is established with more calcium present on the outside than within the capsule core.
The function of PLO is to impart diffusion limitations on the resulting APA membrane, and provide capsule strength and robustness. As a relatively small polymer, with a molecular weight of around 12kDa, and enhanced affinity relative to calcium, PLO binds to the alginate on the surface of the capsule by displacing small amounts of calcium.
Human clinical trials
An approval from the New Zealand Minister for Health in 2008 paved the way for a groundbreaking clinical trial involving cell transplantation between species. Now in phase two clinical trials in both Argentina and New Zealand, LCT’s new treatment has been shown to reduce episodes in patients lacking early warning signs of low blood sugar – a condition known as ‘unaware hypoglycaemia’.
In 2011, funding for the ongoing clinical development of the drug was secured through a AU$50m (US$50.8m) joint venture between LCT and Otsuka Pharmaceutical factory, with each owning a 50% share in Diatranz Otsuka Limited – a partnership providing opportunity for a fully-funded route to market. With phase two studies nearing completion, phase three clinical trials due to commence in 2013 and plans in place to file a new drug application in New Zealand soon after, the treatment could improve quality of life for millions worldwide as soon as 2015. Whoever said pigs weren’t clever?