Vision Loss

Coding Dimension ID: 
325
Coding Dimension path name: 
Vision Loss

Genetic analysis and modification of hES cells

Funding Type: 
SEED Grant
Grant Number: 
RS1-00222
ICOC Funds Committed: 
$0
Disease Focus: 
Aging
Vision Loss
Stem Cell Use: 
Embryonic Stem Cell
Public Abstract: 
Although some hESC lines have been reported to be stable in culture, we and others have found that the stability is variable for different lines. Furthermore, small DNA rearrangements that cannot be detected by chromosome analysis, which has been used as a key measure of hESC stability, exist in many cell lines. It is of crucial importance to detect any potential genetic abnormalities in hESC, and hESC genomes must be stable if they are to become useful clinical reagents. Our long-term goal is to generate genetically stable hESC lines that can be eventually used safely for therapeutic purposes. However, knowledge of long-term genetic stability of these cell lines is necessary to ensure the safe therapeutic use of these cells, and it is essential for the generation of new cell lines in the future. We believe that the groundwork for improving the genetic stability of the existing cell lines should be done before we derive new cell lines. Therefore, we would like to use the existing hESC lines to explore culture conditions for improving genomic stability of hESC upon long-term culture. We propose to follow the molecular features of existing cell lines, both federally- and non-federally-approved lines, over time to identify differences potentially caused by culturing methods and conditions. There is very little information on the non-federally approved cell lines due to the lack of funding. It is important to compare these cell lines with the federally approved cell lines because they are derived and cultured under different conditions and methods. The common genetic and epigenetic features of these cells may also provide insights into the characteristics and plasticity of embryonic stem cells. We also propose to study how genetic changes such as permanent removal or addition of a gene to the hESC will impact these cells, especially the genes involved in DNA repair and modification. We have already begun to design new ways to manipulate these cells for both the understanding of basic biology and the future therapeutic application.
Statement of Benefit to California: 
Similar to many other proposals, the work proposed here would help California in general ways such as biotechnology development and human health improvement. The work proposed here would also provide several specific benefits for the California community. The strength of the research team is genetics and functional analysis, and the approach is focused on fundamental understanding of stem cell genetics and is hypothesis-driven. Therefore, the work proposed will not only serve as a stepping stone for clinical application of embryonic cells, it will also provide a solid understanding of the biological function of these cells. Much work on embryonic stem cells has been focused on the potential clinical applications of these cells and not as much on genetic stability and fundamental biology. Also, most of the work has been done using the federally-approved cell lines due to funding constraints. The stability of these cells is of key importance for their clinical application, so improving genetic stability in culture and increasing the genetic adaptability of these cells are essential. Comparing federally-approved and non-federally-approved cell lines is also important since they were derived and cultured under different conditions. Therefore, we have begun work in these two directions with cell lines from both sources. Experiments designed to manipulate these cells based on critical genetic information may provide important insight into the function and genetic adaptability of human embryonic stem cells. The research reagents generated in this proposal will make many other experiments in normal human cells possible. The unique aspect of this proposal may move basic scientific understanding of normal cells forward and give California a leading edge in both clinical application and basic biology of human embryonic stem cells.
Progress Report: 
  • It is estimated that by 2020, over 450,000 Californians will suffer from vision loss or blindness due to the age-related macular degeneration (AMD), the most common cause of retinal degeneration in the elderly. A layer of cells at the back of the eye called the retinal pigment epithelium (RPE), provides support, protection, and nutrition to the light sensitive retina, and cooperates with other retinal cells to maintain normal visual function. The dysfunction and/or loss of these RPE cells play a critical role in the development of dry AMD, the most common form of that disease. RPE cells derived from human ES cells (hES-RPE) are a potentially unlimited resource for cell replacement therapy.
  • During this grant period we were able to develop methods to reproducibly derive RPE cells from established human ES (hES) cell lines.
  • Normal RPE cells have a polarized appearance, meaning that they have specialized appearances and functions on the top (retinal facing) and bottom of the cells. We were then able to induce the hES-derived RPE to develop a polarized appearance very similar to that found in the normal eye. These cells had specialized characteristics that were remarkable similar to normal RPE and showed ability to function like normal RPE cells. We then developed sheets of hES-derived RPE on substrates that could be used for surgical implantation into animal eyes.
  • We then performed studies to establish appropriate animal models in which to test the ability of hES-derived RPE to prevent or slow retinal degeneration in animal models that have some features of human AMD. We were not able to use standard mouse models of AMD because mouse eyes are too small to perform the surgical implantation of polarized RPE. We first looked at a rabbit model of RPE degeneration induced by a toxin (sodium iodate). We were able to establish reproducible RPE degeneration in these animals but whenever we tried to implant hES-derived RPE under the retinas of these animals, the RPE cells did not survive. We believe that the toxin that was used to induce this model was being taken up by the transplanted cells resulting in their death. We then tested a genetic model of retinal degeneration in rats (RCS rat). We were able to demonstrate reproducible retinal degeneration in these animals. When we transplanted hES-derived RPE under the retinas of these animals, we were able to show that the RPE were indeed human in origin and that they were able to survive for at least several weeks after transplantation. Importantly, we were able to show that in the areas where hES-derived RPE were transplanted, there was less retinal degeneration than in the areas in which there was no transplantation or "sham" transplantation.
  • Thus, in conclusion, we were able to show (1) the reproducible culture of RPE cells from human ES cell lines, (2) that these hES-derived RPE had the appearance and functional characterisitics of human RPE and could be grown on substrates as sheets or polarized cells, and (3) that when these sheets of polarized hES-derived RPE were implanted under the retinal of rats with retinal degeneration, that they were able to survive and showed the ability to slow the progression of retinal degeneration in these animals.
  • The results of this study suggest that hES cells may be an unlimited source of RPE cells suitable for transplantation into eyes with retinal degeneration and that subretinal implantation of polarized hES-derived RPE may represent a novel therapy for dry AMD.

Identification and Analysis of Genome-wide Intra- and Inter-chromosomal Associations in Human Embryonic Stem Cells

Funding Type: 
SEED Grant
Grant Number: 
RS1-00222
ICOC Funds Committed: 
$0
Disease Focus: 
Aging
Vision Loss
Stem Cell Use: 
Embryonic Stem Cell
Public Abstract: 
We used to think of genes as discrete segments of DNA that code for proteins and that are regulated or controlled by nearby or adjacent DNA sequences. Recently, it has been shown that DNA sequences on the same chromosome that are very distant from a gene may regulate its function, determining if the gene will be turned on or turned off. We have now discovered that DNA sequences that are even on different chromosomes from the protein-coding gene may control this gene. These interactions are often totally unexpected. For example, the NF1 gene, which causes neurofibromatosis, a genetic disease that may lead to the development of brain tumors, lies on chromosome 11 in the mouse, but the production of its protein product is partially under the control of a DNA segment near a growth factor gene(IGF2) on mouse chromosome 7; the same relationship appears to hold true in humans. No one had ever suspected such an interaction, and it is now logical to consider targeting the IGF2 gene as part of our hopes to treat neurofibromatosis. In other words, the discovery of a long-range interacting sequence of DNA provides us with totally new targets for drug development. Several other groups have also shown that inter-chromosomal gene regulation can control immune genes and the genes that regulate our sense of smell. We believe that many, if not most, genes are controlled, as least in part, by long range inter-chromosomal interactions, and we have proposed a new method to try to define all of these interactions in cultured human cells. It is highly likely that many of these long range inter-chromosomal interactions are important in the regulation of a gene, and therefore, we plan to catalog all of these interactions in human embryonic stem cells, and see whether the interactions remain the same or are altered as the cells differentiate into fat cells, and, in the future, into other kinds of cells. This catalog will provide us with numerous new clues to the regulation of disease-related genes, and will suggest new drug or gene-therapy related targets to treat these diseases. By comparing the list of interacting genes in normal embryonic cells with those in cells harboring a genetic disease, we may develop new diagnostic tools in addition to potential therapeutic avenues.
Statement of Benefit to California: 
Our goal is to characterize how genes are regulated by distant DNA segments on different chromosomes. We will develop a complete catalog of these long range interactions, which will provide a list of how one gene may regulate another. This catalog will provide the framework for determining how disease-related genes are controlled and will allow investigators to develop new diagnostic tools and/or new therapeutic targets to treat various genetic diseases. These are targets that had never heretofore been considered because the long range interactions among these genes had not been known. Clinical translation of these studies into new drugs will be of great benefit to the people of California.
Progress Report: 
  • It is estimated that by 2020, over 450,000 Californians will suffer from vision loss or blindness due to the age-related macular degeneration (AMD), the most common cause of retinal degeneration in the elderly. A layer of cells at the back of the eye called the retinal pigment epithelium (RPE), provides support, protection, and nutrition to the light sensitive retina, and cooperates with other retinal cells to maintain normal visual function. The dysfunction and/or loss of these RPE cells play a critical role in the development of dry AMD, the most common form of that disease. RPE cells derived from human ES cells (hES-RPE) are a potentially unlimited resource for cell replacement therapy.
  • During this grant period we were able to develop methods to reproducibly derive RPE cells from established human ES (hES) cell lines.
  • Normal RPE cells have a polarized appearance, meaning that they have specialized appearances and functions on the top (retinal facing) and bottom of the cells. We were then able to induce the hES-derived RPE to develop a polarized appearance very similar to that found in the normal eye. These cells had specialized characteristics that were remarkable similar to normal RPE and showed ability to function like normal RPE cells. We then developed sheets of hES-derived RPE on substrates that could be used for surgical implantation into animal eyes.
  • We then performed studies to establish appropriate animal models in which to test the ability of hES-derived RPE to prevent or slow retinal degeneration in animal models that have some features of human AMD. We were not able to use standard mouse models of AMD because mouse eyes are too small to perform the surgical implantation of polarized RPE. We first looked at a rabbit model of RPE degeneration induced by a toxin (sodium iodate). We were able to establish reproducible RPE degeneration in these animals but whenever we tried to implant hES-derived RPE under the retinas of these animals, the RPE cells did not survive. We believe that the toxin that was used to induce this model was being taken up by the transplanted cells resulting in their death. We then tested a genetic model of retinal degeneration in rats (RCS rat). We were able to demonstrate reproducible retinal degeneration in these animals. When we transplanted hES-derived RPE under the retinas of these animals, we were able to show that the RPE were indeed human in origin and that they were able to survive for at least several weeks after transplantation. Importantly, we were able to show that in the areas where hES-derived RPE were transplanted, there was less retinal degeneration than in the areas in which there was no transplantation or "sham" transplantation.
  • Thus, in conclusion, we were able to show (1) the reproducible culture of RPE cells from human ES cell lines, (2) that these hES-derived RPE had the appearance and functional characterisitics of human RPE and could be grown on substrates as sheets or polarized cells, and (3) that when these sheets of polarized hES-derived RPE were implanted under the retinal of rats with retinal degeneration, that they were able to survive and showed the ability to slow the progression of retinal degeneration in these animals.
  • The results of this study suggest that hES cells may be an unlimited source of RPE cells suitable for transplantation into eyes with retinal degeneration and that subretinal implantation of polarized hES-derived RPE may represent a novel therapy for dry AMD.

Stem Cells for Neuroprotection of Photoreceptors in Retinitis Pigmentosa

Funding Type: 
Disease Team Research I
Grant Number: 
DR1-01444
ICOC Funds Committed: 
$0
Disease Focus: 
Vision Loss
Stem Cell Use: 
Embryonic Stem Cell
Cell Line Generation: 
Embryonic Stem Cell
Public Abstract: 
The targeted disease is retinitis pigmentosa (RP), is a severe form of blindness that runs in families. This disease is not common, yet represents an attainable near term target for stem cell therapy for a number of reasons: 1) RP destroys the light detecting cells of the retina but generally leaves the rest of the visual system and body unharmed, so the clinical goal is circumscribed; 2) RP is prototypical of degenerations of the nervous system, so a cure for this less common disease would accelerate progress towards new therapies for a range of more familiar conditions; 3) scientific research has shown that the rods and cones can be spared in animals by transplanting particular types of stem cells, so the scientific feasibility of treating RP has already been established in principle. The therapeutic approach is to save the light sensing cells of the eye (rod and cone photoreceptors) in people going blind using a type of stem cell obtained from the immature retina, but not from early embryos. These particular stem cells from the retina, known as progenitor cells, are capable of rescuing photoreceptors from degeneration following transplantation to the eye. These same cells are also highly efficient at turning into rod photoreceptors and this provides another more sustained pathway by which they preserve the crucial cone photoreceptors. In addition, there is evidence that the stem cells themselves might become functional photoreceptors and thereby stabilize the retina by directly replacing the dying cells in the patient’s eye. Thus, transplanted stem cells could treat the targeted disease of RP in multiple ways simultaneously. Importantly, there are a host of reasons why clinical trials in the eye are easier and safer than most locations in the body. The eye is an important proving ground for stem cell-based therapies and provides a stepping stone to many incurable diseases of the brain and spinal cord.
Statement of Benefit to California: 
Benefits to the state of California and its citizens are both direct and indirect. The direct benefit is medical in that there is currently no cure or established treatment for the individuals and families that suffer from the dreadful hereditary blindness known as retinitis pigmentosa. In addition, there are many people in California and throughout the world that suffer from degenerative diseases of the retina and central nervous system that could benefit from the type of stem cell therapy proposed in the current application. The rapid progress that could be achieved via this proposal would help legitimize the use of stem cells and should thereby accelerate the development of stem cell-based therapeutics for a wide range of other diseases. In so doing there would be an indirect benefit to California by making our state a focal point for stem cell breakthroughs. This would increase medical capabilities, strengthen the {REDACTED}, and energize local biotechnology companies with outside investment and a payoff in jobs and tax revenues.
Progress Report: 
  • It is estimated that by 2020, over 450,000 Californians will suffer from vision loss or blindness due to the age-related macular degeneration (AMD), the most common retinal degenerative disease in the elderly. AMD is a progressive ocular disease of the part of the retina at the back of the eye, called the macula, which enables people to read, visualize faces, and drive. The disease initially causes distortion in central vision, and eventually leads to legal blindness.
  • A layer of cells at the back of the eye called the retinal pigment epithelium (RPE)provide support, protection, and nutrition to the light sensitive photoreceptors in the retina. The dysfunction and/or loss of these RPE cells play a critical role in the loss of the PR’s and hence the blindness in AMD. Effective treatment could be achieved by proper replacement of damaged RPE and retinal cells with healthy ones. RPE cells derived from human embryonic stem cells (hESC) are a potentially unlimited and robust source for regenerating RPE (hESC-RPE).
  • During the first year of our Disease Team Grant we have assembled a closely working team of interdisciplinary scientists and physician scientists at the University of Southern California (USC), Doheny Eye Institute (DEI), University of California Santa Barbara (UCSB), California Institute of Technology (Caltech), and City of Hope (COH). Our scientists work and meet together frequently to discuss progress and we have had 2 highly successful full-day retreats; one at USC and one at UCSB.
  • We are on track for each of the Year 1 proposed milestones. During the first year we have made a decision on final selection of hESC line; and have developed protocols for the generation and molecular and functional characterization of hESC-RPE and are transferring these protocols to our manufacturing and regulatory partners at City of Hope. COH has had onsite visits to learn protocols at UCSB and USC and we have had a day-long meeting at COH to further refine protocols and procedures. In collaboration with Caltech we have developed a non-biodegradable substrate on which these cells are grown and where they develop characteristics of mature RPE cells. Specialized surgical instruments have been developed at DEI and Caltech to implant the hESC cells grown on substrate under the retina. We have utilized sophisticated instrumentation to image the retina in live animals and have used these instruments to follow rats with progressive retinal degeneration before and after implantation of the hESC. We have demonstrated that hESC rescue the degeneration and integrate well into the host retina. Sections of these retinas are evaluated histologically at the microscopic level using sophisticated quantitative imaging techniques.
  • Our group is composed of unique multidisciplinary members who collectively have more than two decades of experience in efforts to restore sight to the blind as well as retinal cell transplantation and stem cell research. Our ultimate goal is to submit an Investigational New Drug (IND) Application to the Food and Drug Administration (FDA) by the end of the 4th year of the grant in order to get approval to conduct a clinical trial in patients at risk of vision loss due to AMD.
  • Age-Related Macular Degeneration (AMD) is a devastating disease that can lead to severe vision impairment and blindness. Vision loss in AMD usually is after age 55 and affects almost 2 million Americans.
  • Vision is initiated by light striking and activating specific cells called photoreceptor cells of the neural retina within the eye. There is a small, specialized region in the central portion of the retina called the macula that is particularly important for central, sharp and color vision. It has a high percentage of photoreceptor cells called "cone cells" and it is this region that is most affected by AMD. Thus, with cone cell degeneration in AMD, a person loses their central, sharp vision with color vision affected as well.
  • In the normal retina, photoreceptor cells are supported metabolically and structurally by a thin layer of cells next to them called Retinal Pigment Epithelial (RPE)Cells. Without these RPE cells, photoreceptor cells quickly degenerate and die. In AMD, it is thought that dysfunction of RPE cells is an early and critical sign of AMD. Thus, replacing dead or dying RPE cells in AMD could be a way to slow the disease process and even improve vision.
  • Our CIRM disease team grant which we call The California Project to Cure Blindness aims to treat AMD through replacement of these dysfunctional RPE cells with fresh RPE cells that will then keep photoreceptor cells alive and functioning. Because only very few RPE cells are present in a human eye, direct RPE transplantation would be very difficult so we rely on the use of human embryonic stem cells (hESCs). Through the work in our CIRM funded disease team grant we have been able to use a particular stem cell line and differentiate into adult-like RPE cells that exhibit many characteristics of normal mature human RPE cells. We believe that implanting these hESCs within the eye next to the retina could be of benefit in AMD in saving the photoreceptors.
  • Our technique is to not only implant the hESC derived RPE cells in the eye but to place them on a special ultrathin substrate platform that will maintain them in proper orientation next to the retina. Moreover, implantation can be done with hESC cells that had been allowed to differentiate in tissue culture prior to implantation, insuring that the implanted cells indeed express the characteristics of mature, functional RPE cells.
  • In this last year, substantial progress has been made in progressing to our goal of replenishing RPE cells in the AMD retina and restoring vision. First, a specific hESC line has been identified that will differentiate into cells that have many of the morphological and biochemical characteristics of normal adult RPE cells. Second, an appropriate material has been found that can act as a substratum (platform) that will support the RPE cells and allow them to function in a normal manner. Third, we have begun to demonstrated the safety of our procedure as well as its efficacy in slowing vision loss in a rodent model of retina degeneration. Fourth, we also have developed unique surgical techniques that will allow us to safely and effectively place the cells in the animal and ultimately human eyes.
  • In parallel to these basic studies, we are advancing our strategies that will let us treat human patients with AMD. Clinical protocols are being established that will be submitted to the US FDA to gain their approval in order to start phase 1 (early) Clinical Trial. In summary, our CIRM grant has allowed us to develop a procedure that should let us treat severe vision loss in AMD. Already, data at 6 months in animal studies has demonstrated the safety of the technique as well as the restoration of functional vision in a model system. Hopefully, this will lead us to a successful human clinical trial with sight restoration in currently untreatable cases of dry AMD.
  • Age-Related Macular Degeneration (AMD) is a devastating disease that can lead to severe vision impairment and blindness. Vision loss in AMD usually is after age 55 and affects almost 2 million Americans.
  • Vision is initiated by light striking and activating specific cells called photoreceptor cells of the neural retina within the eye. There is a small, specialized region in the central portion of the retina called the macula that is particularly important for central, sharp and color vision. It has a high percentage of photoreceptor cells called "cone cells" and it is this region that is most affected by AMD. Thus, with cone cell degeneration in AMD, a person loses their central, sharp vision with color vision affected as well.
  • In the normal retina, photoreceptor cells are supported metabolically and structurally by a thin layer of cells next to them called Retinal Pigment Epithelial (RPE)Cells. Without these RPE cells, photoreceptor cells quickly degenerate and die. In AMD, it is thought that dysfunction of RPE cells is an early and critical sign of AMD. Thus, replacing dead or dying RPE cells in AMD could be a way to slow the disease process and even improve vision.
  • Our CIRM disease team grant which we call The California Project to Cure Blindness aims to treat AMD through replacement of these dysfunctional RPE cells with fresh RPE cells that will then keep photoreceptor cells alive and functioning. Because only very few RPE cells are present in a human eye, direct RPE transplantation would be very difficult so we rely on the use of human embryonic stem cells (hESCs). Through the work in our CIRM funded disease team grant we have been able to use a particular stem cell line and differentiate into adult-like RPE cells that exhibit many characteristics of normal mature human RPE cells. We believe that implanting these hESCs within the eye next to the retina could be of benefit in AMD in saving the photoreceptors.
  • Our technique is to not only implant the hESC derived RPE cells in the eye but to place them on a special ultrathin substrate platform that will maintain them in proper orientation next to the retina. Moreover, implantation can be done with hESC cells that had been allowed to differentiate in tissue culture prior to implantation, insuring that the implanted cells indeed express the characteristics of mature, functional RPE cells.
  • In this last year, substantial progress has been made in progressing to our goal of replenishing RPE cells in the AMD retina and restoring vision. First, a specific hESC line has been identified that will differentiate into cells that have many of the morphological and biochemical characteristics of normal adult RPE cells. Second, an appropriate material has been found that can act as a substratum (platform) that will support the RPE cells and allow them to function in a normal manner. Third, we have begun to demonstrated the safety of our procedure as well as its efficacy in slowing vision loss in a rodent model of retina degeneration. Fourth, we also have developed unique surgical techniques that will allow us to safely and effectively place the cells in the animal and ultimately human eyes.
  • In parallel to these basic studies, we are advancing our strategies that will let us treat human patients with AMD. Clinical protocols are being established that will be submitted to the US FDA to gain their approval in order to start phase 1 (early) Clinical Trial. In summary, our CIRM grant has allowed us to develop a procedure that should let us treat severe vision loss in AMD. Already, data at 6 months in animal studies has demonstrated the safety of the technique as well as the restoration of functional vision in a model system. Hopefully, this will lead us to a successful human clinical trial with sight restoration in currently untreatable cases of dry AMD.
  • Age-Related Macular Degeneration (AMD) is a devastating disease that can lead to severe vision impairment and blindness. Vision loss in AMD usually is after age 55 and affects almost 2 million Americans.
  • Vision is initiated by light striking and activating specific cells called photoreceptor cells of the neural retina within the eye. There is a small, specialized region in the central portion of the retina called the macula that is particularly important for central, sharp and color vision. It has a high percentage of photoreceptor cells called "cone cells" and it is this region that is most affected by AMD. Thus, with cone cell degeneration in AMD, a person loses their central, sharp vision with color vision affected as well.
  • In the normal retina, photoreceptor cells are supported metabolically and structurally by a thin layer of cells next to them called Retinal Pigment Epithelial (RPE)Cells. Without these RPE cells, photoreceptor cells quickly degenerate and die. In AMD, it is thought that dysfunction of RPE cells is an early and critical sign of AMD. Thus, replacing dead or dying RPE cells in AMD could be a way to slow the disease process and even improve vision.
  • Our CIRM disease team grant which we call The California Project to Cure Blindness aims to treat AMD through replacement of these dysfunctional RPE cells with fresh RPE cells that will then keep photoreceptor cells alive and functioning. Because only very few RPE cells are present in a human eye, direct RPE transplantation would be very difficult so we rely on the use of human embryonic stem cells (hESCs). Through the work in our CIRM funded disease team grant we have been able to use a particular stem cell line and differentiate into adult-like RPE cells that exhibit many characteristics of normal mature human RPE cells. We believe that implanting these hESCs within the eye next to the retina could be of benefit in AMD in saving the photoreceptors.
  • Our technique is to not only implant the hESC derived RPE cells in the eye but to place them on a special ultrathin substrate platform that will maintain them in proper orientation next to the retina. Moreover, implantation can be done with hESC cells that had been allowed to differentiate in tissue culture prior to implantation, insuring that the implanted cells indeed express the characteristics of mature, functional RPE cells.
  • In this last year, substantial progress has been made in progressing to our goal of replenishing RPE cells in the AMD retina and restoring vision. First, a specific hESC line has been identified that will differentiate into cells that have many of the morphological and biochemical characteristics of normal adult RPE cells. Second, an appropriate material has been found that can act as a substratum (platform) that will support the RPE cells and allow them to function in a normal manner. Third, we have begun to demonstrated the safety of our procedure as well as its efficacy in slowing vision loss in a rodent model of retina degeneration. Fourth, we also have developed unique surgical techniques that will allow us to safely and effectively place the cells in the animal and ultimately human eyes.
  • In parallel to these basic studies, we are advancing our strategies that will let us treat human patients with AMD. Clinical protocols are being established that will be submitted to the US FDA to gain their approval in order to start phase 1 (early) Clinical Trial. In summary, our CIRM grant has allowed us to develop a procedure that should let us treat severe vision loss in AMD. Already, data at 6 months in animal studies has demonstrated the safety of the technique as well as the restoration of functional vision in a model system. Hopefully, this will lead us to a successful human clinical trial with sight restoration in currently untreatable cases of dry AMD.

U.S./CFP Country Airway Team: Tissue Engineered Airways for Children with Tracheal Obstruction

Funding Type: 
Disease Team Research I
Grant Number: 
DR1-01444
ICOC Funds Committed: 
$0
Disease Focus: 
Vision Loss
Stem Cell Use: 
Embryonic Stem Cell
Cell Line Generation: 
Embryonic Stem Cell
Public Abstract: 
The primary goal of our Airway Team is to bring a safe and effective therapy to children with severe large airway disease. Our intent is to implement all of the necessary steps for a successful new stem/progenitor cell-derived airway transplant for clinical trials in children within 4 years. Our team builds on a first-in-human surgical success with a stem cell-based tissue engineered airway implant in a compassionate care case in a young adult. To this end, we will perform the necessary preclinical studies to support a successful FDA application within 4 years. We propose to use stem/progenitor cells from the patient to treat an extraordinarily difficult-to-manage health problem in children, namely tracheobronchomalacia (TBM). TBM in children is a disease that leads to collapse of tracheal cartilage causing severe airway obstruction that is life-threatening. It occurs in approximately 200 California children each year and the morbidity and mortality associated with this disease is very high. Approximately 25% of these young patients die before their first birthday. Treatment costs for these children are very high, and the familial and societal investments are substantially higher, although outcomes are consistently poor. The endpoint desired is normal airway and lung function in these children and a normal quality of life. Our Airway Team aims to eliminate the need for repeated surgical interventions which are not necessarily successful, presently the standard of care for children with large airway obstruction. Bioengineered airway transplants that use the cells of the patients could be used in humans of all age groups and would not require lifelong, harmful anti-rejection medications. In 2008, a stem cell-based, tissue engineered tracheal implant was successfully used by our Airway Team partners to save a young woman’s life. This first-in-human study emphasizes that our goal is realistic and paves the way for Phase I/II trials in children. Stem/progenitor cell-derived airway transplants that use the patient's cells have the clinical advantage of not requiring anti-rejection medications long-term. Our experience to date indicates such medication is not needed and this finding represents a scientific and clinical breakthrough in organ transplantation. While clear medical benefit was demonstrated in this proof-of-principle, compassionate care case, there is substantial work that must be done before considering such transplants for pediatric patients, and on a large scale, for adults. We address this challenge with our Airway Team and emphasize the synergism resulting from linking Team members with expertise in a variety of related scientific and medical disciplines to address this critical need. This new therapeutic approach could offer a tremendous benefit to children and patients in other age groups in the State of California that are in desperate need of new treatment options.
Statement of Benefit to California: 
The citizens of California have generously invested in stem cell research and a return on their investment will include breakthroughs in medical treatments for diseases where there are currently limited options. Stem/progenitor cell-derived airway transplantation is a leading example of translational research in regenerative medicine that can be used for a host of diseases. Through this team effort scientists and physicians will lead the early promise of airway transplantation to clinical trials in California and beyond. The Airway Team proposes to use stem cells to cure an extraordinarily difficult-to-manage and life-threatening health problem in children. Tracheobronchomalacia (TBM) in children is a disease that leads to severe airway obstruction and occurs in approximately 200 California children each year. The morbidity and mortality associated with this disease is very high; approximately 25% of patients will die before their first birthday. TBM in children is our target disease, but the knowledge gained from the preclinical studies proposed will provide a new technology that can be applied to other disorders in California populations. We foresee that our stem cell-derived airway transplant could be applied to treat adults with severe tracheomalacia associated with lifelong bronchitis and severe chronic obstructive pulmonary disease (COPD) and the large number of children and adults with severe subglottic stenoses that have proven refractory to standard surgical interventions, and potentially even patients with debilitating laryngeal scarring. Given that the prevalence rate of COPD for California citizens >65 years of age approaches 8-10%, if even 0.1% of COPD patients in California were candidates, then >3,000 patients might benefit from this treatment. The methods and technology developed from this project can also be used as the basis for other similar health needs including esophageal, bladder, and bowel replacements for disorders where present treatments are very limited and impair quality of life. An additional benefit for California citizens will be the establishment of a scientific and clinical Center of Excellence in Airway Disorders in California. The Airway Team project will provide a cornerstone of a larger Airway Center that will serve a diverse population of California patients. Our Team will provide the impetus to better coordinate and share the considerable knowledge and resources that are presently used to treat adults and children with COPD, asthma, cystic fibrosis, and other upper airway disorders for which new treatment options are desperately needed.
Progress Report: 
  • It is estimated that by 2020, over 450,000 Californians will suffer from vision loss or blindness due to the age-related macular degeneration (AMD), the most common retinal degenerative disease in the elderly. AMD is a progressive ocular disease of the part of the retina at the back of the eye, called the macula, which enables people to read, visualize faces, and drive. The disease initially causes distortion in central vision, and eventually leads to legal blindness.
  • A layer of cells at the back of the eye called the retinal pigment epithelium (RPE)provide support, protection, and nutrition to the light sensitive photoreceptors in the retina. The dysfunction and/or loss of these RPE cells play a critical role in the loss of the PR’s and hence the blindness in AMD. Effective treatment could be achieved by proper replacement of damaged RPE and retinal cells with healthy ones. RPE cells derived from human embryonic stem cells (hESC) are a potentially unlimited and robust source for regenerating RPE (hESC-RPE).
  • During the first year of our Disease Team Grant we have assembled a closely working team of interdisciplinary scientists and physician scientists at the University of Southern California (USC), Doheny Eye Institute (DEI), University of California Santa Barbara (UCSB), California Institute of Technology (Caltech), and City of Hope (COH). Our scientists work and meet together frequently to discuss progress and we have had 2 highly successful full-day retreats; one at USC and one at UCSB.
  • We are on track for each of the Year 1 proposed milestones. During the first year we have made a decision on final selection of hESC line; and have developed protocols for the generation and molecular and functional characterization of hESC-RPE and are transferring these protocols to our manufacturing and regulatory partners at City of Hope. COH has had onsite visits to learn protocols at UCSB and USC and we have had a day-long meeting at COH to further refine protocols and procedures. In collaboration with Caltech we have developed a non-biodegradable substrate on which these cells are grown and where they develop characteristics of mature RPE cells. Specialized surgical instruments have been developed at DEI and Caltech to implant the hESC cells grown on substrate under the retina. We have utilized sophisticated instrumentation to image the retina in live animals and have used these instruments to follow rats with progressive retinal degeneration before and after implantation of the hESC. We have demonstrated that hESC rescue the degeneration and integrate well into the host retina. Sections of these retinas are evaluated histologically at the microscopic level using sophisticated quantitative imaging techniques.
  • Our group is composed of unique multidisciplinary members who collectively have more than two decades of experience in efforts to restore sight to the blind as well as retinal cell transplantation and stem cell research. Our ultimate goal is to submit an Investigational New Drug (IND) Application to the Food and Drug Administration (FDA) by the end of the 4th year of the grant in order to get approval to conduct a clinical trial in patients at risk of vision loss due to AMD.
  • Age-Related Macular Degeneration (AMD) is a devastating disease that can lead to severe vision impairment and blindness. Vision loss in AMD usually is after age 55 and affects almost 2 million Americans.
  • Vision is initiated by light striking and activating specific cells called photoreceptor cells of the neural retina within the eye. There is a small, specialized region in the central portion of the retina called the macula that is particularly important for central, sharp and color vision. It has a high percentage of photoreceptor cells called "cone cells" and it is this region that is most affected by AMD. Thus, with cone cell degeneration in AMD, a person loses their central, sharp vision with color vision affected as well.
  • In the normal retina, photoreceptor cells are supported metabolically and structurally by a thin layer of cells next to them called Retinal Pigment Epithelial (RPE)Cells. Without these RPE cells, photoreceptor cells quickly degenerate and die. In AMD, it is thought that dysfunction of RPE cells is an early and critical sign of AMD. Thus, replacing dead or dying RPE cells in AMD could be a way to slow the disease process and even improve vision.
  • Our CIRM disease team grant which we call The California Project to Cure Blindness aims to treat AMD through replacement of these dysfunctional RPE cells with fresh RPE cells that will then keep photoreceptor cells alive and functioning. Because only very few RPE cells are present in a human eye, direct RPE transplantation would be very difficult so we rely on the use of human embryonic stem cells (hESCs). Through the work in our CIRM funded disease team grant we have been able to use a particular stem cell line and differentiate into adult-like RPE cells that exhibit many characteristics of normal mature human RPE cells. We believe that implanting these hESCs within the eye next to the retina could be of benefit in AMD in saving the photoreceptors.
  • Our technique is to not only implant the hESC derived RPE cells in the eye but to place them on a special ultrathin substrate platform that will maintain them in proper orientation next to the retina. Moreover, implantation can be done with hESC cells that had been allowed to differentiate in tissue culture prior to implantation, insuring that the implanted cells indeed express the characteristics of mature, functional RPE cells.
  • In this last year, substantial progress has been made in progressing to our goal of replenishing RPE cells in the AMD retina and restoring vision. First, a specific hESC line has been identified that will differentiate into cells that have many of the morphological and biochemical characteristics of normal adult RPE cells. Second, an appropriate material has been found that can act as a substratum (platform) that will support the RPE cells and allow them to function in a normal manner. Third, we have begun to demonstrated the safety of our procedure as well as its efficacy in slowing vision loss in a rodent model of retina degeneration. Fourth, we also have developed unique surgical techniques that will allow us to safely and effectively place the cells in the animal and ultimately human eyes.
  • In parallel to these basic studies, we are advancing our strategies that will let us treat human patients with AMD. Clinical protocols are being established that will be submitted to the US FDA to gain their approval in order to start phase 1 (early) Clinical Trial. In summary, our CIRM grant has allowed us to develop a procedure that should let us treat severe vision loss in AMD. Already, data at 6 months in animal studies has demonstrated the safety of the technique as well as the restoration of functional vision in a model system. Hopefully, this will lead us to a successful human clinical trial with sight restoration in currently untreatable cases of dry AMD.
  • Age-Related Macular Degeneration (AMD) is a devastating disease that can lead to severe vision impairment and blindness. Vision loss in AMD usually is after age 55 and affects almost 2 million Americans.
  • Vision is initiated by light striking and activating specific cells called photoreceptor cells of the neural retina within the eye. There is a small, specialized region in the central portion of the retina called the macula that is particularly important for central, sharp and color vision. It has a high percentage of photoreceptor cells called "cone cells" and it is this region that is most affected by AMD. Thus, with cone cell degeneration in AMD, a person loses their central, sharp vision with color vision affected as well.
  • In the normal retina, photoreceptor cells are supported metabolically and structurally by a thin layer of cells next to them called Retinal Pigment Epithelial (RPE)Cells. Without these RPE cells, photoreceptor cells quickly degenerate and die. In AMD, it is thought that dysfunction of RPE cells is an early and critical sign of AMD. Thus, replacing dead or dying RPE cells in AMD could be a way to slow the disease process and even improve vision.
  • Our CIRM disease team grant which we call The California Project to Cure Blindness aims to treat AMD through replacement of these dysfunctional RPE cells with fresh RPE cells that will then keep photoreceptor cells alive and functioning. Because only very few RPE cells are present in a human eye, direct RPE transplantation would be very difficult so we rely on the use of human embryonic stem cells (hESCs). Through the work in our CIRM funded disease team grant we have been able to use a particular stem cell line and differentiate into adult-like RPE cells that exhibit many characteristics of normal mature human RPE cells. We believe that implanting these hESCs within the eye next to the retina could be of benefit in AMD in saving the photoreceptors.
  • Our technique is to not only implant the hESC derived RPE cells in the eye but to place them on a special ultrathin substrate platform that will maintain them in proper orientation next to the retina. Moreover, implantation can be done with hESC cells that had been allowed to differentiate in tissue culture prior to implantation, insuring that the implanted cells indeed express the characteristics of mature, functional RPE cells.
  • In this last year, substantial progress has been made in progressing to our goal of replenishing RPE cells in the AMD retina and restoring vision. First, a specific hESC line has been identified that will differentiate into cells that have many of the morphological and biochemical characteristics of normal adult RPE cells. Second, an appropriate material has been found that can act as a substratum (platform) that will support the RPE cells and allow them to function in a normal manner. Third, we have begun to demonstrated the safety of our procedure as well as its efficacy in slowing vision loss in a rodent model of retina degeneration. Fourth, we also have developed unique surgical techniques that will allow us to safely and effectively place the cells in the animal and ultimately human eyes.
  • In parallel to these basic studies, we are advancing our strategies that will let us treat human patients with AMD. Clinical protocols are being established that will be submitted to the US FDA to gain their approval in order to start phase 1 (early) Clinical Trial. In summary, our CIRM grant has allowed us to develop a procedure that should let us treat severe vision loss in AMD. Already, data at 6 months in animal studies has demonstrated the safety of the technique as well as the restoration of functional vision in a model system. Hopefully, this will lead us to a successful human clinical trial with sight restoration in currently untreatable cases of dry AMD.
  • Age-Related Macular Degeneration (AMD) is a devastating disease that can lead to severe vision impairment and blindness. Vision loss in AMD usually is after age 55 and affects almost 2 million Americans.
  • Vision is initiated by light striking and activating specific cells called photoreceptor cells of the neural retina within the eye. There is a small, specialized region in the central portion of the retina called the macula that is particularly important for central, sharp and color vision. It has a high percentage of photoreceptor cells called "cone cells" and it is this region that is most affected by AMD. Thus, with cone cell degeneration in AMD, a person loses their central, sharp vision with color vision affected as well.
  • In the normal retina, photoreceptor cells are supported metabolically and structurally by a thin layer of cells next to them called Retinal Pigment Epithelial (RPE)Cells. Without these RPE cells, photoreceptor cells quickly degenerate and die. In AMD, it is thought that dysfunction of RPE cells is an early and critical sign of AMD. Thus, replacing dead or dying RPE cells in AMD could be a way to slow the disease process and even improve vision.
  • Our CIRM disease team grant which we call The California Project to Cure Blindness aims to treat AMD through replacement of these dysfunctional RPE cells with fresh RPE cells that will then keep photoreceptor cells alive and functioning. Because only very few RPE cells are present in a human eye, direct RPE transplantation would be very difficult so we rely on the use of human embryonic stem cells (hESCs). Through the work in our CIRM funded disease team grant we have been able to use a particular stem cell line and differentiate into adult-like RPE cells that exhibit many characteristics of normal mature human RPE cells. We believe that implanting these hESCs within the eye next to the retina could be of benefit in AMD in saving the photoreceptors.
  • Our technique is to not only implant the hESC derived RPE cells in the eye but to place them on a special ultrathin substrate platform that will maintain them in proper orientation next to the retina. Moreover, implantation can be done with hESC cells that had been allowed to differentiate in tissue culture prior to implantation, insuring that the implanted cells indeed express the characteristics of mature, functional RPE cells.
  • In this last year, substantial progress has been made in progressing to our goal of replenishing RPE cells in the AMD retina and restoring vision. First, a specific hESC line has been identified that will differentiate into cells that have many of the morphological and biochemical characteristics of normal adult RPE cells. Second, an appropriate material has been found that can act as a substratum (platform) that will support the RPE cells and allow them to function in a normal manner. Third, we have begun to demonstrated the safety of our procedure as well as its efficacy in slowing vision loss in a rodent model of retina degeneration. Fourth, we also have developed unique surgical techniques that will allow us to safely and effectively place the cells in the animal and ultimately human eyes.
  • In parallel to these basic studies, we are advancing our strategies that will let us treat human patients with AMD. Clinical protocols are being established that will be submitted to the US FDA to gain their approval in order to start phase 1 (early) Clinical Trial. In summary, our CIRM grant has allowed us to develop a procedure that should let us treat severe vision loss in AMD. Already, data at 6 months in animal studies has demonstrated the safety of the technique as well as the restoration of functional vision in a model system. Hopefully, this will lead us to a successful human clinical trial with sight restoration in currently untreatable cases of dry AMD.

hESC-Derived Motor Neuron Progenitors for the Treatment of Spinal Muscular Atrophy

Funding Type: 
Disease Team Research I
Grant Number: 
DR1-01444
ICOC Funds Committed: 
$0
Disease Focus: 
Vision Loss
Stem Cell Use: 
Embryonic Stem Cell
Cell Line Generation: 
Embryonic Stem Cell
Public Abstract: 
Our goal is to obtain FDA-approval for a stem cell-based therapy to treat the leading genetic killer of infants under 2 years of age, spinal muscular atrophy (SMA) Type I. SMA is a disease that first causes dysfunction of and then destroys motor neurons controlling all movement, including swallowing and breathing. There is no known treatment. The diagnosis of SMA Type I is usually made before 3 months of age and >95 percent die or require full-time respiratory support in infancy. Of all motor neuron diseases, SMA is the easiest to obtain FDA approval for, and presents the greatest clinical need given the certain death of afflicted infants. The information that we obtain will be directly relevant to SMA Type II and III, ALS, chronic spinal cord injury, and polio, all of which are characterized by motor neuron loss. The treatment involves transplantation of high-purity populations of human motor neuron cells derived from human embryonic stem cells (hESC-MNPs). We have already completed many steps towards FDA approval, including 1) production of a motor neuron manufacturing facility to make the cells in a manner compatible with human use, which has been audited and shown to be compliant with FDA guidelines for cell production, 2) small animal proof-of-concept studies which show that transplantation of our cells, made under FDA-compliant conditions, restores lost motor neurons in models of motor neuron loss, 3) FDA-reviewed safety studies which show that transplantation of our cells, made under FDA-compliant conditions, does not cause tumors, pain or any adverse events, and 4) multiple meetings with expert researchers and doctors resulting in a final clinical plan. Our team has been preparing for this clinical trial for 3 years, and has already held formal meetings with the FDA, so we do not require all 4 years of support. In this proposal, we are requesting 18 months of support for, 1) scale up of the cell manufacturing facility to prepare for the large volumes required for human treatment, and confirm that the scaled up production process produces the same quality cells as the current production process, 2) replication of small animal proof-of-concept studies which show that transplantation of our cells, made under FDA-compliant conditions, restores lost motor neurons in models of motor neuron loss, 3) large animal safety studies to test the accuracy of our cell delivery with available instrumentation, determine survival and differentiation rate of transplanted cells, and make sure that transplantation does not result in tumor formation or any adverse event, and 4) hire the staff to complete the pre-IND and IND filings.
Statement of Benefit to California: 
The treatment and care of individuals with motor neuron diseases represents a significant social and economic burden to the State of California. Specific to the first target indication, there are approximately 1,500 individuals affected by SMA currently living in the State of California and nearly 1,000,000 carriers of the SMA gene. Additionally, the Families of SMA Research Director, Dr. Jill Jarecki, is headquartered in San Diego. If successful, our clinical strategy will improve the function and quality of life of individuals with motor neuron diseases, which will lessen the cost that citizens and the State bear in terms of patient care. This program will position California for international competitiveness in this emerging area of biotechnology, as our clinical trial development strategy addresses critical barriers limiting the development of this sector in California and abroad. High purity cultures of hESC-derivatives enable transplantation approaches to disease, drug discovery, and predictive toxicology. This research plan will lead to the development and thorough characterization of a renewable source of human motor neurons that enables these 3 strategies as they pertain to spinal muscular atrophy, amyotrophic lateral sclerosis, polio, and spinal cord injury. Our program is within 18 months of being presented to the FDA for an IND. Thus, it will provide precedent in California for subsequent stem cell-based clinical trials, {REDACTED}. Thus, California will benefit from ‘hosting’ the discovery of what will become the first and second human embryonic stem cell-based clinical trials in the world. This will result in California being a focus of the stem cell industry, including large pharmaceutical companies that will eventually participate in the latter stage stem cell clinical trials. Clinically relevant scientific advances lead to the development of biotechnology companies, creating jobs and taxation. Funding for this proposal will directly create 11 new jobs. In these challenging economic times, we feel that any award should be spent in a manner which enhances the local and state economy, in essence returning direct value to the citizens of California. To this end, we have purposely selected suppliers of equipment and services that are located in the state of California. In addition, due to the booming medical device and biotechnology industries in California, we feel that all new hires can be obtained from within the state of California.
Progress Report: 
  • It is estimated that by 2020, over 450,000 Californians will suffer from vision loss or blindness due to the age-related macular degeneration (AMD), the most common retinal degenerative disease in the elderly. AMD is a progressive ocular disease of the part of the retina at the back of the eye, called the macula, which enables people to read, visualize faces, and drive. The disease initially causes distortion in central vision, and eventually leads to legal blindness.
  • A layer of cells at the back of the eye called the retinal pigment epithelium (RPE)provide support, protection, and nutrition to the light sensitive photoreceptors in the retina. The dysfunction and/or loss of these RPE cells play a critical role in the loss of the PR’s and hence the blindness in AMD. Effective treatment could be achieved by proper replacement of damaged RPE and retinal cells with healthy ones. RPE cells derived from human embryonic stem cells (hESC) are a potentially unlimited and robust source for regenerating RPE (hESC-RPE).
  • During the first year of our Disease Team Grant we have assembled a closely working team of interdisciplinary scientists and physician scientists at the University of Southern California (USC), Doheny Eye Institute (DEI), University of California Santa Barbara (UCSB), California Institute of Technology (Caltech), and City of Hope (COH). Our scientists work and meet together frequently to discuss progress and we have had 2 highly successful full-day retreats; one at USC and one at UCSB.
  • We are on track for each of the Year 1 proposed milestones. During the first year we have made a decision on final selection of hESC line; and have developed protocols for the generation and molecular and functional characterization of hESC-RPE and are transferring these protocols to our manufacturing and regulatory partners at City of Hope. COH has had onsite visits to learn protocols at UCSB and USC and we have had a day-long meeting at COH to further refine protocols and procedures. In collaboration with Caltech we have developed a non-biodegradable substrate on which these cells are grown and where they develop characteristics of mature RPE cells. Specialized surgical instruments have been developed at DEI and Caltech to implant the hESC cells grown on substrate under the retina. We have utilized sophisticated instrumentation to image the retina in live animals and have used these instruments to follow rats with progressive retinal degeneration before and after implantation of the hESC. We have demonstrated that hESC rescue the degeneration and integrate well into the host retina. Sections of these retinas are evaluated histologically at the microscopic level using sophisticated quantitative imaging techniques.
  • Our group is composed of unique multidisciplinary members who collectively have more than two decades of experience in efforts to restore sight to the blind as well as retinal cell transplantation and stem cell research. Our ultimate goal is to submit an Investigational New Drug (IND) Application to the Food and Drug Administration (FDA) by the end of the 4th year of the grant in order to get approval to conduct a clinical trial in patients at risk of vision loss due to AMD.
  • Age-Related Macular Degeneration (AMD) is a devastating disease that can lead to severe vision impairment and blindness. Vision loss in AMD usually is after age 55 and affects almost 2 million Americans.
  • Vision is initiated by light striking and activating specific cells called photoreceptor cells of the neural retina within the eye. There is a small, specialized region in the central portion of the retina called the macula that is particularly important for central, sharp and color vision. It has a high percentage of photoreceptor cells called "cone cells" and it is this region that is most affected by AMD. Thus, with cone cell degeneration in AMD, a person loses their central, sharp vision with color vision affected as well.
  • In the normal retina, photoreceptor cells are supported metabolically and structurally by a thin layer of cells next to them called Retinal Pigment Epithelial (RPE)Cells. Without these RPE cells, photoreceptor cells quickly degenerate and die. In AMD, it is thought that dysfunction of RPE cells is an early and critical sign of AMD. Thus, replacing dead or dying RPE cells in AMD could be a way to slow the disease process and even improve vision.
  • Our CIRM disease team grant which we call The California Project to Cure Blindness aims to treat AMD through replacement of these dysfunctional RPE cells with fresh RPE cells that will then keep photoreceptor cells alive and functioning. Because only very few RPE cells are present in a human eye, direct RPE transplantation would be very difficult so we rely on the use of human embryonic stem cells (hESCs). Through the work in our CIRM funded disease team grant we have been able to use a particular stem cell line and differentiate into adult-like RPE cells that exhibit many characteristics of normal mature human RPE cells. We believe that implanting these hESCs within the eye next to the retina could be of benefit in AMD in saving the photoreceptors.
  • Our technique is to not only implant the hESC derived RPE cells in the eye but to place them on a special ultrathin substrate platform that will maintain them in proper orientation next to the retina. Moreover, implantation can be done with hESC cells that had been allowed to differentiate in tissue culture prior to implantation, insuring that the implanted cells indeed express the characteristics of mature, functional RPE cells.
  • In this last year, substantial progress has been made in progressing to our goal of replenishing RPE cells in the AMD retina and restoring vision. First, a specific hESC line has been identified that will differentiate into cells that have many of the morphological and biochemical characteristics of normal adult RPE cells. Second, an appropriate material has been found that can act as a substratum (platform) that will support the RPE cells and allow them to function in a normal manner. Third, we have begun to demonstrated the safety of our procedure as well as its efficacy in slowing vision loss in a rodent model of retina degeneration. Fourth, we also have developed unique surgical techniques that will allow us to safely and effectively place the cells in the animal and ultimately human eyes.
  • In parallel to these basic studies, we are advancing our strategies that will let us treat human patients with AMD. Clinical protocols are being established that will be submitted to the US FDA to gain their approval in order to start phase 1 (early) Clinical Trial. In summary, our CIRM grant has allowed us to develop a procedure that should let us treat severe vision loss in AMD. Already, data at 6 months in animal studies has demonstrated the safety of the technique as well as the restoration of functional vision in a model system. Hopefully, this will lead us to a successful human clinical trial with sight restoration in currently untreatable cases of dry AMD.
  • Age-Related Macular Degeneration (AMD) is a devastating disease that can lead to severe vision impairment and blindness. Vision loss in AMD usually is after age 55 and affects almost 2 million Americans.
  • Vision is initiated by light striking and activating specific cells called photoreceptor cells of the neural retina within the eye. There is a small, specialized region in the central portion of the retina called the macula that is particularly important for central, sharp and color vision. It has a high percentage of photoreceptor cells called "cone cells" and it is this region that is most affected by AMD. Thus, with cone cell degeneration in AMD, a person loses their central, sharp vision with color vision affected as well.
  • In the normal retina, photoreceptor cells are supported metabolically and structurally by a thin layer of cells next to them called Retinal Pigment Epithelial (RPE)Cells. Without these RPE cells, photoreceptor cells quickly degenerate and die. In AMD, it is thought that dysfunction of RPE cells is an early and critical sign of AMD. Thus, replacing dead or dying RPE cells in AMD could be a way to slow the disease process and even improve vision.
  • Our CIRM disease team grant which we call The California Project to Cure Blindness aims to treat AMD through replacement of these dysfunctional RPE cells with fresh RPE cells that will then keep photoreceptor cells alive and functioning. Because only very few RPE cells are present in a human eye, direct RPE transplantation would be very difficult so we rely on the use of human embryonic stem cells (hESCs). Through the work in our CIRM funded disease team grant we have been able to use a particular stem cell line and differentiate into adult-like RPE cells that exhibit many characteristics of normal mature human RPE cells. We believe that implanting these hESCs within the eye next to the retina could be of benefit in AMD in saving the photoreceptors.
  • Our technique is to not only implant the hESC derived RPE cells in the eye but to place them on a special ultrathin substrate platform that will maintain them in proper orientation next to the retina. Moreover, implantation can be done with hESC cells that had been allowed to differentiate in tissue culture prior to implantation, insuring that the implanted cells indeed express the characteristics of mature, functional RPE cells.
  • In this last year, substantial progress has been made in progressing to our goal of replenishing RPE cells in the AMD retina and restoring vision. First, a specific hESC line has been identified that will differentiate into cells that have many of the morphological and biochemical characteristics of normal adult RPE cells. Second, an appropriate material has been found that can act as a substratum (platform) that will support the RPE cells and allow them to function in a normal manner. Third, we have begun to demonstrated the safety of our procedure as well as its efficacy in slowing vision loss in a rodent model of retina degeneration. Fourth, we also have developed unique surgical techniques that will allow us to safely and effectively place the cells in the animal and ultimately human eyes.
  • In parallel to these basic studies, we are advancing our strategies that will let us treat human patients with AMD. Clinical protocols are being established that will be submitted to the US FDA to gain their approval in order to start phase 1 (early) Clinical Trial. In summary, our CIRM grant has allowed us to develop a procedure that should let us treat severe vision loss in AMD. Already, data at 6 months in animal studies has demonstrated the safety of the technique as well as the restoration of functional vision in a model system. Hopefully, this will lead us to a successful human clinical trial with sight restoration in currently untreatable cases of dry AMD.
  • Age-Related Macular Degeneration (AMD) is a devastating disease that can lead to severe vision impairment and blindness. Vision loss in AMD usually is after age 55 and affects almost 2 million Americans.
  • Vision is initiated by light striking and activating specific cells called photoreceptor cells of the neural retina within the eye. There is a small, specialized region in the central portion of the retina called the macula that is particularly important for central, sharp and color vision. It has a high percentage of photoreceptor cells called "cone cells" and it is this region that is most affected by AMD. Thus, with cone cell degeneration in AMD, a person loses their central, sharp vision with color vision affected as well.
  • In the normal retina, photoreceptor cells are supported metabolically and structurally by a thin layer of cells next to them called Retinal Pigment Epithelial (RPE)Cells. Without these RPE cells, photoreceptor cells quickly degenerate and die. In AMD, it is thought that dysfunction of RPE cells is an early and critical sign of AMD. Thus, replacing dead or dying RPE cells in AMD could be a way to slow the disease process and even improve vision.
  • Our CIRM disease team grant which we call The California Project to Cure Blindness aims to treat AMD through replacement of these dysfunctional RPE cells with fresh RPE cells that will then keep photoreceptor cells alive and functioning. Because only very few RPE cells are present in a human eye, direct RPE transplantation would be very difficult so we rely on the use of human embryonic stem cells (hESCs). Through the work in our CIRM funded disease team grant we have been able to use a particular stem cell line and differentiate into adult-like RPE cells that exhibit many characteristics of normal mature human RPE cells. We believe that implanting these hESCs within the eye next to the retina could be of benefit in AMD in saving the photoreceptors.
  • Our technique is to not only implant the hESC derived RPE cells in the eye but to place them on a special ultrathin substrate platform that will maintain them in proper orientation next to the retina. Moreover, implantation can be done with hESC cells that had been allowed to differentiate in tissue culture prior to implantation, insuring that the implanted cells indeed express the characteristics of mature, functional RPE cells.
  • In this last year, substantial progress has been made in progressing to our goal of replenishing RPE cells in the AMD retina and restoring vision. First, a specific hESC line has been identified that will differentiate into cells that have many of the morphological and biochemical characteristics of normal adult RPE cells. Second, an appropriate material has been found that can act as a substratum (platform) that will support the RPE cells and allow them to function in a normal manner. Third, we have begun to demonstrated the safety of our procedure as well as its efficacy in slowing vision loss in a rodent model of retina degeneration. Fourth, we also have developed unique surgical techniques that will allow us to safely and effectively place the cells in the animal and ultimately human eyes.
  • In parallel to these basic studies, we are advancing our strategies that will let us treat human patients with AMD. Clinical protocols are being established that will be submitted to the US FDA to gain their approval in order to start phase 1 (early) Clinical Trial. In summary, our CIRM grant has allowed us to develop a procedure that should let us treat severe vision loss in AMD. Already, data at 6 months in animal studies has demonstrated the safety of the technique as well as the restoration of functional vision in a model system. Hopefully, this will lead us to a successful human clinical trial with sight restoration in currently untreatable cases of dry AMD.

Human Spinal Cord Neural Stem Cells for Treatment of Acute Traumatic Spinal Cord Injury

Funding Type: 
Disease Team Research I
Grant Number: 
DR1-01444
ICOC Funds Committed: 
$0
Disease Focus: 
Vision Loss
Stem Cell Use: 
Embryonic Stem Cell
Cell Line Generation: 
Embryonic Stem Cell
Public Abstract: 
Spinal cord injury is a devastating disease with no available treatments. It disproportionately affects young people who then lives many decades with the debilitation. In this project, we propose to develop a human neural stem cell line to treat spinal cord injury patients within 14 days after the injury. Administration of the cells into the injured tissue will limit the tissue from further damage and provide replacement neurons and glia that will promote self-repair and regeneration for the rest of the patient's life. During this project, we will assess the therapeutic as well as any toxicity potentials of the cells, using rats and pigs that will be injured in a manner similar to human spinal cord injury. These animal models will provide a rigorous testing grounds to prove or disprove potential clinical usefulness of the cells. If the cells can successfully treat rats or pigs with no major toxicity, then we will submit a IND application to the FDA to conduct a Phase 1 clinical trial with spinal cord injured patients.
Statement of Benefit to California: 
The proposed research is to develop a stem cell therapy to treat spinal cord injury patients in the acute phase after the initial injury event. By intervening early, benefits of the stem cell transplantation to the patients are likely to be greatest. Thus, the patients in the trials will require intensive care and thus only the local population within California could participate. The treatment requires surgery and invasive injection of the cells into the injury site, which will require a special training. Thus, the benefit to the State of California is first access to this therapy for about 2-3 years before it becomes available elsewhere. Secondly, California has two federally funded Regional Spinal Cord Injury Care Centers that research various aspects of rehabilitation after spinal cord injury. Conducting clinical trials in California will become further catalysts to strengthen statewide treatment and rehabilitation centers for spinal cord injury patients.
Progress Report: 
  • It is estimated that by 2020, over 450,000 Californians will suffer from vision loss or blindness due to the age-related macular degeneration (AMD), the most common retinal degenerative disease in the elderly. AMD is a progressive ocular disease of the part of the retina at the back of the eye, called the macula, which enables people to read, visualize faces, and drive. The disease initially causes distortion in central vision, and eventually leads to legal blindness.
  • A layer of cells at the back of the eye called the retinal pigment epithelium (RPE)provide support, protection, and nutrition to the light sensitive photoreceptors in the retina. The dysfunction and/or loss of these RPE cells play a critical role in the loss of the PR’s and hence the blindness in AMD. Effective treatment could be achieved by proper replacement of damaged RPE and retinal cells with healthy ones. RPE cells derived from human embryonic stem cells (hESC) are a potentially unlimited and robust source for regenerating RPE (hESC-RPE).
  • During the first year of our Disease Team Grant we have assembled a closely working team of interdisciplinary scientists and physician scientists at the University of Southern California (USC), Doheny Eye Institute (DEI), University of California Santa Barbara (UCSB), California Institute of Technology (Caltech), and City of Hope (COH). Our scientists work and meet together frequently to discuss progress and we have had 2 highly successful full-day retreats; one at USC and one at UCSB.
  • We are on track for each of the Year 1 proposed milestones. During the first year we have made a decision on final selection of hESC line; and have developed protocols for the generation and molecular and functional characterization of hESC-RPE and are transferring these protocols to our manufacturing and regulatory partners at City of Hope. COH has had onsite visits to learn protocols at UCSB and USC and we have had a day-long meeting at COH to further refine protocols and procedures. In collaboration with Caltech we have developed a non-biodegradable substrate on which these cells are grown and where they develop characteristics of mature RPE cells. Specialized surgical instruments have been developed at DEI and Caltech to implant the hESC cells grown on substrate under the retina. We have utilized sophisticated instrumentation to image the retina in live animals and have used these instruments to follow rats with progressive retinal degeneration before and after implantation of the hESC. We have demonstrated that hESC rescue the degeneration and integrate well into the host retina. Sections of these retinas are evaluated histologically at the microscopic level using sophisticated quantitative imaging techniques.
  • Our group is composed of unique multidisciplinary members who collectively have more than two decades of experience in efforts to restore sight to the blind as well as retinal cell transplantation and stem cell research. Our ultimate goal is to submit an Investigational New Drug (IND) Application to the Food and Drug Administration (FDA) by the end of the 4th year of the grant in order to get approval to conduct a clinical trial in patients at risk of vision loss due to AMD.
  • Age-Related Macular Degeneration (AMD) is a devastating disease that can lead to severe vision impairment and blindness. Vision loss in AMD usually is after age 55 and affects almost 2 million Americans.
  • Vision is initiated by light striking and activating specific cells called photoreceptor cells of the neural retina within the eye. There is a small, specialized region in the central portion of the retina called the macula that is particularly important for central, sharp and color vision. It has a high percentage of photoreceptor cells called "cone cells" and it is this region that is most affected by AMD. Thus, with cone cell degeneration in AMD, a person loses their central, sharp vision with color vision affected as well.
  • In the normal retina, photoreceptor cells are supported metabolically and structurally by a thin layer of cells next to them called Retinal Pigment Epithelial (RPE)Cells. Without these RPE cells, photoreceptor cells quickly degenerate and die. In AMD, it is thought that dysfunction of RPE cells is an early and critical sign of AMD. Thus, replacing dead or dying RPE cells in AMD could be a way to slow the disease process and even improve vision.
  • Our CIRM disease team grant which we call The California Project to Cure Blindness aims to treat AMD through replacement of these dysfunctional RPE cells with fresh RPE cells that will then keep photoreceptor cells alive and functioning. Because only very few RPE cells are present in a human eye, direct RPE transplantation would be very difficult so we rely on the use of human embryonic stem cells (hESCs). Through the work in our CIRM funded disease team grant we have been able to use a particular stem cell line and differentiate into adult-like RPE cells that exhibit many characteristics of normal mature human RPE cells. We believe that implanting these hESCs within the eye next to the retina could be of benefit in AMD in saving the photoreceptors.
  • Our technique is to not only implant the hESC derived RPE cells in the eye but to place them on a special ultrathin substrate platform that will maintain them in proper orientation next to the retina. Moreover, implantation can be done with hESC cells that had been allowed to differentiate in tissue culture prior to implantation, insuring that the implanted cells indeed express the characteristics of mature, functional RPE cells.
  • In this last year, substantial progress has been made in progressing to our goal of replenishing RPE cells in the AMD retina and restoring vision. First, a specific hESC line has been identified that will differentiate into cells that have many of the morphological and biochemical characteristics of normal adult RPE cells. Second, an appropriate material has been found that can act as a substratum (platform) that will support the RPE cells and allow them to function in a normal manner. Third, we have begun to demonstrated the safety of our procedure as well as its efficacy in slowing vision loss in a rodent model of retina degeneration. Fourth, we also have developed unique surgical techniques that will allow us to safely and effectively place the cells in the animal and ultimately human eyes.
  • In parallel to these basic studies, we are advancing our strategies that will let us treat human patients with AMD. Clinical protocols are being established that will be submitted to the US FDA to gain their approval in order to start phase 1 (early) Clinical Trial. In summary, our CIRM grant has allowed us to develop a procedure that should let us treat severe vision loss in AMD. Already, data at 6 months in animal studies has demonstrated the safety of the technique as well as the restoration of functional vision in a model system. Hopefully, this will lead us to a successful human clinical trial with sight restoration in currently untreatable cases of dry AMD.
  • Age-Related Macular Degeneration (AMD) is a devastating disease that can lead to severe vision impairment and blindness. Vision loss in AMD usually is after age 55 and affects almost 2 million Americans.
  • Vision is initiated by light striking and activating specific cells called photoreceptor cells of the neural retina within the eye. There is a small, specialized region in the central portion of the retina called the macula that is particularly important for central, sharp and color vision. It has a high percentage of photoreceptor cells called "cone cells" and it is this region that is most affected by AMD. Thus, with cone cell degeneration in AMD, a person loses their central, sharp vision with color vision affected as well.
  • In the normal retina, photoreceptor cells are supported metabolically and structurally by a thin layer of cells next to them called Retinal Pigment Epithelial (RPE)Cells. Without these RPE cells, photoreceptor cells quickly degenerate and die. In AMD, it is thought that dysfunction of RPE cells is an early and critical sign of AMD. Thus, replacing dead or dying RPE cells in AMD could be a way to slow the disease process and even improve vision.
  • Our CIRM disease team grant which we call The California Project to Cure Blindness aims to treat AMD through replacement of these dysfunctional RPE cells with fresh RPE cells that will then keep photoreceptor cells alive and functioning. Because only very few RPE cells are present in a human eye, direct RPE transplantation would be very difficult so we rely on the use of human embryonic stem cells (hESCs). Through the work in our CIRM funded disease team grant we have been able to use a particular stem cell line and differentiate into adult-like RPE cells that exhibit many characteristics of normal mature human RPE cells. We believe that implanting these hESCs within the eye next to the retina could be of benefit in AMD in saving the photoreceptors.
  • Our technique is to not only implant the hESC derived RPE cells in the eye but to place them on a special ultrathin substrate platform that will maintain them in proper orientation next to the retina. Moreover, implantation can be done with hESC cells that had been allowed to differentiate in tissue culture prior to implantation, insuring that the implanted cells indeed express the characteristics of mature, functional RPE cells.
  • In this last year, substantial progress has been made in progressing to our goal of replenishing RPE cells in the AMD retina and restoring vision. First, a specific hESC line has been identified that will differentiate into cells that have many of the morphological and biochemical characteristics of normal adult RPE cells. Second, an appropriate material has been found that can act as a substratum (platform) that will support the RPE cells and allow them to function in a normal manner. Third, we have begun to demonstrated the safety of our procedure as well as its efficacy in slowing vision loss in a rodent model of retina degeneration. Fourth, we also have developed unique surgical techniques that will allow us to safely and effectively place the cells in the animal and ultimately human eyes.
  • In parallel to these basic studies, we are advancing our strategies that will let us treat human patients with AMD. Clinical protocols are being established that will be submitted to the US FDA to gain their approval in order to start phase 1 (early) Clinical Trial. In summary, our CIRM grant has allowed us to develop a procedure that should let us treat severe vision loss in AMD. Already, data at 6 months in animal studies has demonstrated the safety of the technique as well as the restoration of functional vision in a model system. Hopefully, this will lead us to a successful human clinical trial with sight restoration in currently untreatable cases of dry AMD.
  • Age-Related Macular Degeneration (AMD) is a devastating disease that can lead to severe vision impairment and blindness. Vision loss in AMD usually is after age 55 and affects almost 2 million Americans.
  • Vision is initiated by light striking and activating specific cells called photoreceptor cells of the neural retina within the eye. There is a small, specialized region in the central portion of the retina called the macula that is particularly important for central, sharp and color vision. It has a high percentage of photoreceptor cells called "cone cells" and it is this region that is most affected by AMD. Thus, with cone cell degeneration in AMD, a person loses their central, sharp vision with color vision affected as well.
  • In the normal retina, photoreceptor cells are supported metabolically and structurally by a thin layer of cells next to them called Retinal Pigment Epithelial (RPE)Cells. Without these RPE cells, photoreceptor cells quickly degenerate and die. In AMD, it is thought that dysfunction of RPE cells is an early and critical sign of AMD. Thus, replacing dead or dying RPE cells in AMD could be a way to slow the disease process and even improve vision.
  • Our CIRM disease team grant which we call The California Project to Cure Blindness aims to treat AMD through replacement of these dysfunctional RPE cells with fresh RPE cells that will then keep photoreceptor cells alive and functioning. Because only very few RPE cells are present in a human eye, direct RPE transplantation would be very difficult so we rely on the use of human embryonic stem cells (hESCs). Through the work in our CIRM funded disease team grant we have been able to use a particular stem cell line and differentiate into adult-like RPE cells that exhibit many characteristics of normal mature human RPE cells. We believe that implanting these hESCs within the eye next to the retina could be of benefit in AMD in saving the photoreceptors.
  • Our technique is to not only implant the hESC derived RPE cells in the eye but to place them on a special ultrathin substrate platform that will maintain them in proper orientation next to the retina. Moreover, implantation can be done with hESC cells that had been allowed to differentiate in tissue culture prior to implantation, insuring that the implanted cells indeed express the characteristics of mature, functional RPE cells.
  • In this last year, substantial progress has been made in progressing to our goal of replenishing RPE cells in the AMD retina and restoring vision. First, a specific hESC line has been identified that will differentiate into cells that have many of the morphological and biochemical characteristics of normal adult RPE cells. Second, an appropriate material has been found that can act as a substratum (platform) that will support the RPE cells and allow them to function in a normal manner. Third, we have begun to demonstrated the safety of our procedure as well as its efficacy in slowing vision loss in a rodent model of retina degeneration. Fourth, we also have developed unique surgical techniques that will allow us to safely and effectively place the cells in the animal and ultimately human eyes.
  • In parallel to these basic studies, we are advancing our strategies that will let us treat human patients with AMD. Clinical protocols are being established that will be submitted to the US FDA to gain their approval in order to start phase 1 (early) Clinical Trial. In summary, our CIRM grant has allowed us to develop a procedure that should let us treat severe vision loss in AMD. Already, data at 6 months in animal studies has demonstrated the safety of the technique as well as the restoration of functional vision in a model system. Hopefully, this will lead us to a successful human clinical trial with sight restoration in currently untreatable cases of dry AMD.

Regeneration of Functional Human Corneal Epithelial Progenitor Cells

Funding Type: 
Early Translational II
Grant Number: 
TR2-01768
ICOC Funds Committed: 
$3 054 024
Disease Focus: 
Vision Loss
Stem Cell Use: 
Adult Stem Cell
Cell Line Generation: 
Adult Stem Cell
oldStatus: 
Active
Public Abstract: 
Over 3.2 million people worldwide are bilateral blind from corneal diseases. Limbal stem cell deficiency (LSCD) has been recognized as a major cause, either primary or secondary, of significant visual loss and blindness in many common corneal disorders. A healthy, transparent ocular surface is made up of non-keratinized, stratified squamous epithelium that is highly differentiated. The corneal epithelium is constantly renewed and maintained by the corneal epithelial stem cells, or limbal stem cells (LSCs) that are presumed to reside at the limbus, the junction between the cornea and conjunctiva. When the LSCs are deficient and unable to repopulate the corneal surface, the cornea surface will become opaque. Corneal transplant can’t survive and is contraindicated in LSCD. LSC transplantation, in the form of keratolimbal allograft to restore a transparent corneal surface, has been the main therapy in the United States. The 5-year survival of these allografts is about 30%, largely due to immune rejection. Transplantation of autologous limbal epithelial stem cells that have been expanded on tissue culture has successfully restored vision and revolutionized the patient specific stem-cell based therapy as recently reported by an Italian LSC transplant team. They have achieved a 68% success rate during a mean follow up time of 3 years. The expansion process requires mouse 3T3 feeder cells to grow a sufficient amount of stem cells for transplantation. To reduce cross-contamination from animal products, LSCs that are expanded in a xenobiotic-free culture system has been established; however, the 3-year survival rate of these cells after transplantation is 50% and only 30% survive at 5 year, suggestive of inefficient expansion without the mouse feeders. Therefore, new cell engineering methods that can efficiently expand and regenerate autologous LSCs in a xenobiotic-free system are dearly needed to achieve acceptable clinical outcome and offer stem-cell based therapy to patients with this devastating blinding diseases in the United States. The first goal of this proposed translational research is to establish a xenobiotic-free culture system by replacing the mouse feeder cells with a human feeder system to expand sufficient amount of LSCs for transplantation. This will allow immediate initiation of clinical trial. We will then further optimized the expansion efficiency by modulating the Wnt and Notch signaling pathways based on our findings that Wnt and Notch signaling regulate the proliferation and differentiation of corneal epithelial cells. In parallel, transdifferentiation of human skin epithelial stem cells to corneal epithelial cells will be induced using a similar approach. The ability and safety of these regenerated human corneal epithelial stem cells to reconstruct the ocular surface will be tested in a LSCD animal model. The results of this proposed study will pave the way for preclinical development of this novel cell engineering technique.
Statement of Benefit to California: 
This proposal is to develop a stem-cell based transplantation therapy for treating a blinding corneal disorder, limbal stem cell deficiency (LSCD). Corneal diseases are the second leading cause of treatable blindness in the world and over 3.2 million people worldwide are bilateral blind from corneal diseases. LSCD has been recognized as a major cause, either primary or secondary, of significant visual loss and blindness in many common corneal disorders, such as chemical/thermal burn, keratopathy related to contact lens wear, and severe infection and inflammation. Due to visual impairment, LSCD patients lose the ability to drive, read, and watch TV. In addition, they would experience recurrent corneal erosion that causes severe pain and sensitivity to light. Frequent break down of the corneal surface increases the risk of infection that requires frequent medical interventions. All of these can also exert psychological impact to the patients and their family members. Therefore, LSCD imposes significant social and economical impact on our society. California is the most populated state in the US. There are more than 36 million people in the State of California and the population will increase to 46 million in 2030. Accordingly, the number of residents with limbal stem cell deficiency is likely disproportionately elevated due to the environmental risk factors. Thus, this disease affects a large population of patients in the state of California. A new treatment to restore vision would represent an important benefit to the people of California. Further, the project would train new stem-cell researchers and advance innovative technology in stem cell therapy. This technology has application to other stem-cell related diseases. When this project enters the clinical phase, it will bring together new physicians and scientists and attract funding by the federal government. In addition, it will undoubtedly attract biotechnology investment in California. The stem-cell based transplantation to treat a stem-cell related disease like limbal stem cell deficiency is well-aligned with the broad mission of CIRM and the objectives of the Early Translational Research Award program.
Progress Report: 
  • Over last year, our research team has made significant progress in achieving our research goals and reaching the appropriate milestones. We first have demonstrated that we are able to grow the human limbal stem cells under the standard method using mouse 3T3 cells as feeder at the same efficiency level as the leading group in the world. This is the first milestone that we have reached.
  • We then proceeded with the initial testing of all the proposed human feeder candidates for their ability to support the growth of human limbal stem cells. We found that the current standard culture method on 3T3 cells did not work for human feeder cells. We then investigated four new culture methods to maximize the growth of limbal epithelial cells on human feeder candidates. We also have included a new human feeder cell candidate, adipose-derived mesenchymal stem cells. We are very excited to find that two of the human feeder cell types could support the growth of limbal epithelial cells with a significantly higher efficiency than the 3T3 cells using our two new culture methods. We are in the process of further refining the culture methods and characterize the expanded limbal epithelial cells. We believe that we are able to establish a xenobiotic free culture system to efficiently expand limbal stem cells for transplantation.
  • The goal of the project is to establish a xenobiotic-free culture system that can efficiently expand human limbal stem cells (LSCs) for transplantation. We have met all of the milestones and have accomplished the following:
  • 1) We can grow LSCs on 3T3 feeder cells (the current gold standard culture method) in our laboratory as efficiently as those described in the literature.
  • 2) We have established two cell carrier systems for transplantation.
  • 3) We have established four 3T3 feeder-free culture methods, including a feeder-free system to expand human LSCs as efficiently as the 3T3 feeder cells do. Three types of human feeder cells— limbal fibroblasts, bone marrow-derived mesenchymal stem cells, and adipose-derived mesenchymal stem cells—support the growth of LSCs. All of the 3T3 feeder-free cultures contain more than 3% of p63bright cells.
  • 4) We have shown that two Wnt small molecule activators can increase the expansion efficiency of LSCs by more than 125%.
  • 5) A nude mouse model of LSC deficiency (LSCD) has been established to test the in vivo function of cultured human LSCs.
  • In addition to reaching these milestones, we have derived a novel method in which feeder cells are completely separated from LSCs during culture. This novel method eliminates contamination by the feeder cells but maintains close contact between them. We have found that trypsin has detrimental effects on LSCs during isolation. Limbal epithelial cells in clusters/sheets and limbal tissue explant culture are superior to single-cell culture for the expansion of LSCs. We will continue to investigate which of the four 3T3 feeder-free systems is the most efficient in expanding the LSC population, and we will continue to refine the LSCD animal model. Before the end of the award period, we will be able to select the most efficient and consistent xenobiotic-free method of cell culture and start regulatory tests that are necessary for submission of an investigational new drug (IND) application to the FDA so that we can begin the first clinical trial to treat patients with unilateral LSCD in California.
  • During the last six months, my laboratory has successfully established a xenobiotic-free and feeder-free culture method to efficiently expand human corneal epithelial stem cells in culture by removing the feeder cells and replacing the fetal bovine serum with human serum. The amount of stem cells produced using this method is comparable to the standard culture method using mouse 3T3 cells as feeder cells and fetal bovine serum. We also have tested the feasibility of using fibrin gel and amniotic membrane as cell carrier and found that amniotic membrane is superior to fibrin as cell carrier. We are establishing an animal model to test the disease modifying effects of these cultivated stem cells. Lastly, we can increase the limbal stem cell expansion efficiency by modulating the Wnt and Notch signaling pathway.

3D Modeling of Retina using Polymer Scaffolds for Understanding Disease Pathogenesis

Funding Type: 
Basic Biology IV
Grant Number: 
RB4-05785
ICOC Funds Committed: 
$1 526 319
Disease Focus: 
Vision Loss
Stem Cell Use: 
Embryonic Stem Cell
iPS Cell
Cell Line Generation: 
iPS Cell
oldStatus: 
Active
Public Abstract: 
Inherited retinal degenerations result in visual loss in patients as early as in their adolescence. Retinitis Pigmentosa includes a group of such degenerations which run in families and can result in legal blindness by 40 years of age. Even though we know by now a number of gene mutations which can cause these disorders, we do not understand how these mutations ultimately lead to loss of the cells. Recent advances in stem cell technologies now have provided us with the opportunity to gain a better understanding of the disorders. In this proposal, we plan to study the degenerative process by creating an eye-like structure in a dish using a combination of stem cell and bioengineering approaches. We plan to use 3D scaffolds to grow eye cells generated from pluripotent stem cells. We will directly compare normal retinal cells with cells derived from patients with Retinitis Pigmentosa. This approach will allow us to identify the processes that ultimately lead to the death of the photoreceptor cells int he eye and blindness. In the future, we could then extend this work to identify new drugs which will help halt or at least slow down the degeneration in these patients.
Statement of Benefit to California: 
Photoreceptor degenerations, including Retinitis Pigmentosa (RP), cause visual impairment for millions of patients in the United States and a number of patients in the state of California being one of the most populous states in the US. RP encompasses a group of retinal degeneration with a prevalence of 1 in 4000 and runs in families. Typical symptoms include night blindness followed by decreasing vision, and eventually legal blindness or, in many cases, complete blindness. RP is usually diagnosed in adolescents and young adults. Most people with RP are legally blind by age 40. There are no effective forms of treatment for a majority of these patients. Thus results in a tremendous stress on the state of CA's resources. In addition, there is both monetary and psychological stress on the families esp. in cases of Retinitis Pigmentosa as multiple family members are affected. The only way to potentially help these patients will be to either replace the dead cells with new photoreceptors or find ways to better understand the degenerative process in order to identify novel drugs to delay the progression of degeneration. The proposed research in this application is to generate eye tissue in a dish from patients with severe forms of Retinitis Pigmentosa in order to gain a better understanding of the disease. This will in turn help us in the future to identify new drugs to delay or stop the visual loss and this will attract new biotechnology partners in the state of CA.
Progress Report: 
  • Inherited retinal degenerations result in visual loss in patients early in life. Retinitis Pigmentosa, a form of inherited retinal degeneration, affects thousands of patients in US and in the state of California. The second most common gene whose mutation results in retinitis pigmentosa is RP1. How mutation in the gene result in loss of vision is not completely clear and we are working of gaining a better understanding of this by creating a 3D artificial retina in a dish. This invloved using stem-cell derived eye cells grown on patterned articifical scaffolds.
  • In our initial year of funding, we have improved our methodologies to make the various cells that make up the retina in the dish and are working on optimizing the generation of scaffolds. These scaffold will allow us to generated a retina, the light-sensing layer of the eye, in a dish. We have also over the course of the year generated stem cell lines using patient cells. Using the recent state-of-the-art technologies to convert patient cells to stem-cells, we have been able to convert blood cells to stem cells and then reconverted them to cells in the eye. We will now use these cells to better understand the degeneration and vision loss in the second year of the grant.

Human retinal progenitor cells as candidate therapy for retinitis pigmentosa

Funding Type: 
Early Translational II
Grant Number: 
TR2-01794
ICOC Funds Committed: 
$4 738 251
Disease Focus: 
Vision Loss
Stem Cell Use: 
Adult Stem Cell
oldStatus: 
Closed
Public Abstract: 
The targeted disease is retinitis pigmentosa (RP), is a severe form of blindness that runs in families. This disease is not overly common, yet represents an attainable near term target for stem cell therapy for a number of reasons: 1) RP destroys the light detecting cells of the retina but generally leaves the rest of the visual system and body unharmed, so the clinical goal is circumscribed; 2) RP is prototypical of degenerations of the nervous system, so a cure for this less common disease would accelerate progress towards new therapies for a range of more familiar conditions; 3) scientific research has shown that degenerating rods and cones can be spared in animals by transplanting particular types of stem cells, so the scientific feasibility of treating RP in this way has already been established in principle. The therapeutic approach is to save the light sensing cells of the eye (rod and cone photoreceptors) in people going blind using a type of stem cell obtained from the immature retina, but not from early embryos. These particular stem cells from the retina, known as progenitor cells, are capable of rescuing photoreceptors from degeneration following transplantation to the eye. These same cells are also highly efficient at becoming rod photoreceptors and this provides another more sustained pathway by which they preserve the crucial cone photoreceptors. In addition, there is evidence that the stem cells themselves might become functional photoreceptors and thereby stabilize the retina by directly replacing the dying cells in the patient’s eye. Thus, transplanted stem cells could treat the targeted disease of RP in multiple ways simultaneously. Importantly, there are a host of reasons why clinical trials in the eye are easier and safer than most locations in the body. The eye is an important proving ground for stem cell-based therapies and provides a stepping stone to many otherwise incurable diseases of the brain and spinal cord.
Statement of Benefit to California: 
Benefits to the state of California and its citizens are both direct and indirect. The direct benefit is medical in that there is currently no cure or established treatment for the individuals and families that suffer from the dreadful hereditary blindness known as retinitis pigmentosa. In addition, there are many people in California and throughout the world that suffer from degenerative diseases of the retina and central nervous system that could benefit from further development and alternate applications of the type of stem cell therapy proposed in the current application. The rapid progress that could be achieved via this proposal would help legitimize the use of stem cells and should thereby accelerate the development of stem cell-based therapeutics for a wide range of other diseases. In so doing there would be an indirect benefit to California by making our state a focal point for stem cell breakthroughs. This would increase medical capabilities, strengthen the [REDACTED], and energize local biotechnology companies with outside investment and a payoff in jobs and tax revenues.
Progress Report: 
  • At the end of Year 1 of the grant period, our team has made substantial progress and achieved quite a few of the milestones initially established for the entire 3 year period. A retinal cell bank has been established for research purposes and, in addition, initial clinical-grade cell production is now under way with the target of generating 3 clinical cell banks. We have shown that our human cells are able to help blind rats to see following transplantation to the eye. We have developed a range of methods to help prove the safety of our cells, both before and after transplantation, with definitive long term tests results to be obtained over the next few months. We have begun to unravel the molecular genetics of the cell and are specifically investigating what give them the ability to preserve vision in the face of progressive blindness. In summary, work is advancing rapidly and ahead of schedule, consistent with a realistic, achievable biological strategy. On behalf of the patients who we know are depending upon us, our team is committed to seeing this technology into the clinic in the shortest possible timeline.

Development of a Stem Cell-based Transplantation Strategy for Treating Age-related Macular Degeneration

Funding Type: 
Early Translational I
Grant Number: 
TR1-01272
ICOC Funds Committed: 
$5 503 069
Disease Focus: 
Vision Loss
Stem Cell Use: 
Adult Stem Cell
iPS Cell
oldStatus: 
Closed
Public Abstract: 
Age related macular degeneration (AMD) is a blinding disease of the elderly affecting nearly one in three individuals over the age of 75. Central vision is lost in AMD, severely impairing the ability to read, watch television, or drive. The epicenter of AMD is the retinal pigment epithelium (RPE), a single layer of cells in the retina adjacent to the photoreceptor cells. A recent breakthrough in AMD research showed that this disease is caused in about 50% of cases by the innate immune system (complement system) inappropriately attacking RPE cells. Specifically, AMD results when regulators of the complement system, which normally protect the RPE, are weakened by mutations. This sickens and later kills the RPE, causing secondary degeneration of photoreceptors in the central retina (macula). The goal of this proposal is to develop a strategy for transplanting stem-cell derived RPE cells into the eyes of patients with AMD. In the past, transplantation of RPE cells from postmortem donors yielded encouraging initial therapeutic effects that subsequently failed due to immune rejection. Current stem-cell technology offers the opportunity to avoid this complication. We plan to generate functional RPE cells from stem cells of the ciliary margin zone (CMZ) in the eye, or pluripotent stem cells induced from skin fibroblasts (iPS cells) taken from the same AMD patient who will later receive the induced RPE cells as a transplant. The study of inherited blindness has benefited greatly from mouse genetic models, where new potential therapies can be tested and developed. One aim of this proposal is to produce RPE cells from mouse and human CMZ- and iPS-cell precursors. To establish that these cells are functional, we will test for two hallmarks of a fully differentiated RPE: (i) the ability to convert vitamin A into visual chromophore for photoreceptor-opsin pigments, and (ii) the ability to phagocytose photoreceptor outer-segments. In a later aim we will transplant these induced RPE cells into the eyes of two genetic “knockout” mice that lack the ability to synthesize visual chromophore. We will test for rescue of the biochemical defects, and correction of the blindness in these mutant mice. In another experiment, we will add to the induced RPE cells a gene that protects from inappropriate complement activation. These cells will be transplanted into the eyes of two other knockout mouse-models of AMD that exhibit abnormal activation of the complement system. We will study these mice to establish correction of the immune defect. Finally, we will test the safety of CMZ- and iPS-derived RPE cells by transplanting them into immune-deficient mice to confirm no tumor formation. At the end of the grant period, we expect to have a new and well-tested stem-cell based transplantation strategy that will be ready for phase-one clinical trials in AMD patients.
Statement of Benefit to California: 
This proposal is to develop a stem-cell based transplantation approach for treating age-related macular degeneration (AMD). AMD is a severe and common disease of the elderly that causes central blindness. The prevalence of AMD increases with advancing age. By 75 years, approximately one in three individuals have some degree of visual loss due to AMD. Thus AMD is significantly more prevalent than Alzheimer disease. Patients with AMD lose the ability to drive, read, watch TV, and recognize faces. With advancing visual impairment, AMD patients lose the ability to care for themselves and others. Thus, AMD imposes a large social and economic burden on our society. As the population in California ages, this burden is expected to increase. The stem-cell based transplantation strategy in this proposal offers the real potential of slowing or arresting the progression of blindness in AMD patients. This alone would represent an important benefit to the people of California. Further, the project would advance innovative technology in stem cell therapy. This technology has application to other neurodegenerative diseases. The project will train new stem-cell researchers in California. As the project enters the clinical phase, it will engage new scientists and physicians and attract funding by the federal government. Further, the opportunity to treat a hugely prevalent disease like AMD will undoubtedly attract biotechnology investment in California. This stem-cell based transplantation approach to treat a major disease like AMD is well-aligned with the broad mission of CIRM and the objectives of the Early Translational Research Award program.
Progress Report: 
  • The broad goal of this project is to develop a stem-cell derived replacement for RPE cells in the eye that die in patients with age-related macular degeneration (AMD). Since RPE cells are often genetically defective in AMD, we will correct this defect in the stem-cell derived RPE cells before transplanting them into patients. These transplanted cells will express specific proteins that will protect them from being attacked by the innate immune system. During the first year of funding we made excellent progress toward this goal. We learned how to generate RPE cells from mouse and human stem cells of various sources. We prepared all the DNA constructs that will be required to protect the transplanted RPE cells from attack by the innate immune system. We also developed a new approach for injecting RPE cells into the correct part of the mouse eye (subretinal space) without damaging other parts of the retina. We are enthusiastic to continue our work on this exciting project.
  • The broad goal of this project is to develop a stem-cell derived replacement for retinal pigment epithelial (RPE) cells in the eye, which die in patients with age-related macular degeneration (AMD). Since RPE cells are often genetically defective in AMD, we will correct this defect in the stem-cell derived RPE cells before transplanting them into patients. These transplanted cells will express specific proteins that will protect them from being attacked by the innate immune system. During our second year of funding, we transplanted human stem-cell derived RPE cells into the eyes of mice that are blind due to a genetic defect in the rpe65 gene. We showed that the human cells integrated into the RPE layer of these mice. Further, we demonstrated that the transplanted cells partially rescued the blindness in rpe65 / mice. We also generated and tested the recombinant viruses that will protect RPE cells from attack by the innate immune system. We are on schedule to complete the planned studies for this project during the final year of funding.
  • This project is to develop a new treatment for age-related macular degeneration (AMD) based on transplantation of retinal pigment epithelial (RPE) cells into the subretinal space of a patient’s eyes. These RPE cells are induced from stem cells collected from the same patient, to avoid the problem of immune rejection. AMD is primarily an inflammatory disorder caused by inappropriate attack of RPE cells by the complement system. Accordingly, negative regulators of complement activation will be expressed in the stem-cell derived RPE cells by viral transduction. We explored two sources of stem cells that can be collected from a patient: embryonic stem cells and induced pluripotent stem (iPS) cells from skin fibroblasts. We successfully programed these stem cells into fully functional RPE cells. Our team has extensive experience with the biochemistry and cell biology of the RPE. We used “knockout” mouse models with mutations in RPE genes to test the efficiency of RPE-cell transplantation. Next, we planned to test the strategy of protecting RPE cells from complement attack by over-expressing complement negative-regulatory factors. We tested the long-term viability of induced RPE cells, and rule-out tumorgenicity by transplanting these cells into the eyes of severe-combined immunodeficient mice.

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