Restoring vision by sheet transplants of retinal progenitors and retinal pigment epithelium (RPE) derived from human embryonic stem cells (hESCs)

Funding Type: 
Early Translational IV
Grant Number: 
TR4-06648
Investigator: 
Type: 
PI
Type: 
Co-PI
Award Value: 
$3,998,948
Disease Focus: 
Vision Loss
Stem Cell Use: 
Embryonic Stem Cell
Cell Line Generation: 
Embryonic Stem Cell
Status: 
Active
Public Abstract: 

There is currently no effective treatment to restore or improve vision for patients suffering from incurable blinding diseases such as dry age-related macular degeneration and retinitis pigmentosa, which need both new photoreceptors and retinal pigment epithelium. However, a unique method to transplant fetal retinal progenitor sheets together with its supporting retinal pigment epithelium (RPE) has been shown to improve vision in animal models of retinal degeneration and in patients. Differentiation of human embryonic stem cells (hESCs) into sheets of retinal progenitor tissue that contain photoreceptor progenitors and RPE cells could create an unlimited supply of donor tissue. Our lab has generated retinal progenitor tissue from hESCs in 3-D constructs (“layers”), and a new immunodeficient model of retinal degeneration. Recently, several laboratories have shown that hESCs can “self-assembly” into early stages of eye development and develop into laminated structures. The hypothesis of the proposed project then is that hESCs can be consistently differentiated into sheets of retinal tissue, which can restore visual responses after transplantation to a new immunodeficient rat model of retinal degeneration that does not reject human cells. In the final year, we will standardize methods to increase the production of these sheets in a way that complies to good manufacturing practice. This project will ultimately help to restore vision in patients suffering from retinal diseases.

Statement of Benefit to California: 

Retinal diseases reduce the quality of life of patients who suffer from vision loss and at significant cost to the health care system. Age-related macular degeneration (AMD) destroys the central vision and is the most common cause of blindness among people over 65. In 2010, AMD affected 2.1 % of the general population which means 2.1 million in the U.S. and about 240,000 in California, with these numbers projected to grow to 5 million (in the U.S.) in 2050 as the population ages. Ca. 20-35% of AMD cases develop irreversible geographic atrophy with local loss of RPE and photoreceptors in the macula. Another incurable disease, retinitis pigmentosa (RP) which is inherited (1:3500) and occurs in younger people, affects the light-sensing photoreceptors first, but also the supporting RPE layer beneath the retina following photoreceptor degeneration. Thus, both AMD and RP patients will need both new RPE and photoreceptors. The proposed replacement therapy is the only one that targets more mature disease stages of both AMD and RP, for which no other therapy exists. An effective treatment will keep afflicted individuals productive, enhance State tax revenues and defray the healthcare cost burden to taxpayers. It will also lead to robust industry developments in the fields of clinical transplantation, drug screening, and predictive toxicology, effectively leading to job creation and tax benefits to the State as a result of consumption of research and clinical goods and services.

Progress Report: 

There is currently no effective treatment to restore or improve vision for patients suffering from blinding diseases such as dry, age-related macular degeneration and retinitis pigmentosa. Alleviation of these diseases requires both new photoreceptors and retinal pigment epithelium (RPE). However, a unique method to transplant fetal retinal progenitor sheets together with its supporting retinal pigment epithelium has been shown to improve vision in animal models of retinal degeneration and in patients suffering these diseases. The goal of our research is to improve current procedures to the stage in which they can be used routinely to alleviate these causes of blindness in people.

We are differentiating human embryonic stem cells (hESCs) into sheets of retinal progenitor tissue that contain photoreceptor progenitors and RPE cells. We anticipate that the successful completion of our research will create an unlimited supply of donor tissue that can be transplanted into the eyes of blind people. Recently, several laboratories have shown that hESCs and induced pluripotent stem cells (iPSCs) can “self-assemble” into early stages of eye development and develop into laminated structures resembling the normal retina. The hypothesis of this project is that hESCs can be consistently differentiated into sheets of retinal tissue, which can restore visual responses in an immunodeficient rat model of retinal degeneration that does not reject human cells. In the third year of our research program, we will standardize methods to increase the production of these sheets in a way that complies with good manufacturing practice. This project will ultimately help to restore vision in patients suffering from retinal diseases.

In year 1 of this 3-year grant, the optimal procedure for differentiation of hESCs into laminated 3D-retina has been developed and tested. This method is based on selection of tissues developing into retinal progenitor cells, and culturing them in suspension (non-attached). We have shown by quantitative PCR and by immunohistochemistry that these tissues express retina-specific genes in a time sequence comparable to human retinal development. We have built up the capabilities for testing hESC-derived 3D retina in vivo. We have set up a colony of immunodeficient retinal degenerate (RD) rats (which do not require immunosuppression), and refined the transplantation, imaging and testing techniques. To hone our skills, we have started with transplants of rat and human fetal retinal tissue. We will start with transplantation of hESC-derived 3D retina in March 2015 and continue with testing the tissue through the remainder of the grant. The development of the transplants in vivo is monitored by high-resolution optical coherence tomography which can image retinal layers. Transplanted RD rats are tested by various methods (optokinetic tracking for visual acuity, electroretinograms and recording of visual responses from the superior colliculs, a visual brain center) to demonstrate the transplant effect on vision. In the first year of the grant, we have shown that these methods can demonstrate the improvement of vision by fetal-derived transplants

This work will be presented in several posters at this year’s ARVO meeting; several manuscripts are in preparation. The laboratory has attracted many students who are interested to learn about retinal development and vision restoration.

Currently, there is no effective treatment to restore or improve vision in patients suffering from blindness (retinal degeneration, RD) due to such diseases as age-related macular degeneration and retinitis pigmentosa. Alleviation of these diseases requires both new photoreceptors (the light-sensing cells in the back of the eye) and retinal pigment epithelium (RPE), which supports the photoreceptors. Proof of concept has been demonstrated by improved vision resulting from transplantation of fetal retinal progenitor sheets together with supporting RPE in both animals and patients with retinal disease. The ultimate goal of our research is to alleviate blindness using a renewable source of stem-cell derived retinal sheets.

Human embryonic stem cells (hESCs) are developed into sheets of immature retinal tissue that contains progenitors of photoreceptor as well as RPE cells. Optimization of this differentiation protocol will create an unlimited supply of immature retinal tissue that can be transplanted into the eyes of people with advanced blindness. hESCs and induced pluripotent stem cells (iPSCs) can be coaxed to “self-assemble” into early stages of eye development and develop into layers of cells that resemble the normal retina. The hypothesis of this research is that hESCs can be consistently differentiated into sheets of immature retinal tissue which can restore visual responses in a rat model of retinal degeneration. This particular model is immunodeficient and therefore does not reject human cells. In the third year of our research program, we will increase the production of effective retinal sheets in a way that complies with good manufacturing practice as outlined by the US Food and Drug Administration. This research program will ultimately help to restore vision in patients suffering from retinal diseases.

In year 2 of this 3-year grant, the procedures to obtain 3D retina from hESCs have been improved. The development of retinal structures in cell culture must follow closely its development within a living organism. These cellular products have been shown to express appropriate retinal markers using labeling methods called immunohistochemistry and quantitative polymerase chain reaction (qPCR) which determines which and how many genes are expressed. 3D- retinal progenitor sheets derived from hESCs and human fetal retina have been transplanted into the eyes of rats with fast retinal degeneration, which have been engineered not to reject human cells. In this 2nd year of the grant, such transplanted RD rats have been tested for visual acuity (how they can discriminate moving stripes) and electrical responses to light in the brain (superior colliculus recordings) after survival times of 4 – 8 months after transplant. Preliminary data indicate visual improvements after transplantation by both measures. In the 3rd year of the grant we will also use a second, newly developed rat model of retinal degeneration, immunodeficient RCS rats with a slower rate of retinal degeneration that may result in a higher degree of visual restoration.

This work will be presented at several meetings this year (ARVO, ICER, Neuroscience). One manuscript has been submitted, and several manuscripts are in preparation. The laboratory has attracted many students who are interested to learn about retinal development and vision restoration. The work has also led to new collaborations.