'Validating the application of iPSC-derived Müller glia in a glaucoma therapy'

Glaucoma is the commonest cause of irreversible blindness in the world. It is characterized by damage to specialized neurons of the retina (ganglion cells) and the optic nerve (the part that connects the eye to the brain). The research aims to validate the use of Müller glia derived from induced pluripotent stem cells (iPSC), for designing a cell therapy to treat advanced glaucoma. This proposal stems from our published work that showed that a population of cells from the adult human retina, known as 'Müller glia', have stem cell characteristics. When transplanted into the eye of animals with ganglion cell damage, these cells induced a significant recovery of visual function. We ascribed this function to the ability of these cells to physically and metabolically support the repair of ganglion cell axons, causing restoration of visual function. However, sourcing Müller glia from the adult human eye could limit the development of a therapy for use in a large population due to scarcity of tissue availability and the immune response that these cells can induce in a host. To facilitate the development of a cell therapy that could benefit many individuals, induced pluripotent stem cells (iPSC) may constitute a better cell choice as it might be possible to produce well characterized and traceable cells (ie, semi-defined cell preparations) for retinal therapies. In addition, development of iPSC banks (haplobanks) to derive iPSC compatible with many individuals, may facilitate cell transplantation in a large population cohort. On this basis, the main objectives of our proposed research are:
1) To isolate and propagate Müller glia with stem cell characteristics from retinal organoids produced by iPSC in vitro, and to examine the genetic and physical properties of these cells, as well as their ability to produce nerve-protective factors in the laboratory: We will use iPSC and protocols provided by Prof Pete Coffey at the Institute of Ophthalmology, as well as protocols recently developed in our laboratory to isolate Müller cells from retinal organoids. These cells will be also examined for their ability to release factors that protect nerve cells.
2) To transplant iPSC-derived Müller cells into rat eyes with experimental glaucoma, and to assess visual function in these animals: To induce glaucoma, Brown Norway rats will be injected with magnetic beads into the anterior part of the eye to induce high intraocular pressure and consequent damage to the ganglion cells and the optic nerve (the main features of glaucoma). Animals will receive an intra-ocular injection of Müller cells (dead or alive) and their visual function will be assessed at various times (up to 24 weeks) after injection by measurement of their electrophysiological (ERG) response to light. We will also image in vivo the thickness of the nerve fibres that extend along the inner part of the retina towards the optic nerve.
3) To examine the ability of transplanted Müller cells to induce retinal ganglion cell/axon repair in the grafted eye using immuno-histochemical and electron-microscopy techniques, and to assess whether protective factors produced by Müller cells may be present in the transplanted retina: Histological and electron-microscopy examination of the transplanted retina will be undertaken at various times (up to 24 weeks) after transplantation to assess the integrity of host ganglion cells and to determine the presence of protective factors associated with the transplanted cells.
We expect that the results will provide the basis for the future formulation of a pre-clinical development plan for the use of iPSC-derived Müller glia as a therapy for end stage glaucoma. This project will benefit from the availability of GMP iPSC lines derived by our collaborator Prof P. Coffey. Although iPSC have been derived from UK and US donors, Prof Coffey has proved the safety of UK derived stem cell lines in collaboration with the MRC PRION Unit.

Glaucoma is a major cause of blindness characterized by retinal ganglion cell (RGC) loss and optic nerve damage. This study builds on our previous findings that a population of Müller glial with stem cell characteristics can be isolated from the human retina and can be induced to differentiate into the major types of retinal neurons in vitro. Upon transplantation onto the retina of RGC depleted animals, these cells induced a significant recovery of visual function. Sourcing Müller glia from donor eyes could limit therapy development due to limitations in obtaining tissue and compatibility issues. To develop a therapy that could benefit many patients, induced pluripotent stem cells (iPSC) may yield well characterized cells and may be potentially derived from cell banks to provide cells to a large number of individuals. On this basis, our objectives are to undertake a feasibility study and generate a proof of concept that iPSC-derived Müller glia may be used for development of therapy to treat advanced glaucoma. The main aims of the proposed research are: 1) To isolate and propagate Müller glia from retinal organoids derived from iPSC in vitro, and to examine their genetic and phenotypic characteristics as well as their ability to produce neurotrophins. 2) To transplant iPSC-derived Müller glia into the vitreous of the glaucomatous rat eye and to assess the thickness of the nerve fibre layer (NFL) in vivo, as well as the retinal function in transplanted animals using electrophysiological methods. 3) To examine the ability of transplanted cells to induce RGC/axon repair and RGC survival in the grafted eye using immuno-histochemical and electron-microscopy techniques, and to assess whether neurotrophins produced by MG may be present in the transplanted retina. We expect that the results will provide the basis for the future formulation of a pre-clinical development plan for the use of iPSC derived Müller glia as a therapy for end stage glaucoma.

Glaucoma is the commonest cause of irreversible blindness in the world. Although current management may control disease progression, many patients continue to develop visual impairment and eventually become blind. Globally there are 7 million people blinded by this condition. Glaucomatous visual loss is irreversible in nature and there are no current therapies to restore RGC or axonal function. Because only a small number of RGC are needed to maintain functional vision, stem cell transplantation to provide physical and metabolic support to RGC may potentially reverse some RGC-axonal damage, restore useful visual function and improve quality of life.
Blindness causes a major economic burden to society as people affected do not contribute financially and rather require support from it. Finding new treatments that prevent or delay blindness would have a significant impact on the economy, as individuals affected by end stage glaucoma may not only be active in the workforce for longer, but they will enjoy a better quality of life. The research has direct relevance to the exploration of innovative treatments for ocular conditions affecting a very large number of individuals worldwide. With recent advances in medical treatments, it is possible to prevent substantial visual loss in many patients affected by severe disease, but in a long term, it is impossible to prevent blindness finally occurring in many individuals.
Present treatments for end stage Glaucoma using stem cells are not yet available, and although much work is being undertaken by many Scientists in the retinal field, very little attention has been given to this condition. If a new cell based treatment could be developed for these patients to prolong or delay visual loss or even partially restore sight, this would significantly impact not only in the UK and the world economy, but also would improve the quality of life of many individual affected by retinal degenerative disease across developed and undeveloped countries.
Any new treatments for glaucoma that can be successfully applied could also benefit other scientists in the neuroscience field, as the principles and technologies used for cell based therapies to repair the inner neural retina could also be applied to a wide range of neural degenerative diseases of the brain. We are working with an International Venture Capitalist with a strong links to Moorfields Eye Hospital, who convened an international scientific panel in London last year to review our scientific approach to repairing RGC axon function by using MG cells. Amongst the panel were Drs Ron McKay (National Institute of Health, U.S.A), and Mahendra Rao (New York Stem Cell Foundation), who examined our research and supported our approach. The Venture Capitalist has expressed interest on creating a UK Start-Up company to develop the pre-clinical and clinical phases of the therapy once the experimental work to support the development of preclinical studies is completed. If a cell therapy for glaucoma can be established, it would not only benefit patients affected by terminal disease, but also the Biomedical Industry, as the work would pave the way for the generation of a new therapeutic modality to treat a large number of individual worldwide.
It is expected that in addition to the economic impact, the study will have educational impact. Knowledge acquired through the research will be disseminated across science, engineering, undergraduate and medical/graduate students. New knowledge in the development of cell based therapies to treat end stage glaucoma will become an integral part of specialised lectures, and research projects will provide expert training to postdoctoral fellows and postgraduate project students.