Early Translational I
$5 503 069
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.
This proposal focuses on the development of novel cell therapies for the treatment of age-related macular degeneration (AMD). AMD is a common blinding disease of the elderly caused by progressive accumulation of degenerative and vascular changes in the macula of the eye, particularly in the retinal pigment epithelium (RPE). The applicant proposes a cell replacement therapy in which RPE cells would be transplanted into the subretinal space of the patient’s eye. First, the applicant proposes to differentiate RPE cells from two autologous stem cells sources: ciliary margin zone (CMZ) cells obtained by biopsy of the eye, and induced pluripotent stem cells (iPSCs) from skin fibroblasts. The applicant will test these cells for function first in vitro and then in vivo by transplantation into several transgenic mouse models of eye disease. To address the pathophysiology of AMD in which inflammation is believed to be due to inappropriate attack of RPE cells by the complement system, the applicant proposes to test stem cell-derived RPE cells that have been genetically modified to overexpress negative regulators of the complement cascade. Finally, the applicant will monitor the potential tumorigenicity of iPSC-derived RPE cells by transplantation into immunodeficient mice. The reviewers agreed that this proposal addresses an unmet medical need and could have significant impact. They noted that AMD is a common and severe disease lacking effective therapies and an attractive target for stem cell derived therapies. The eye confers several advantages as a transplantation site, including the ability to monitor transplanted cells non-invasively and the option of removing an eye in the event of tumorigenesis (to prevent life threatening spread). A reviewer noted that surgical methods exist for the replacement of diseased RPE but allogeneic transplanted RPE eventually undergo immune rejection. This proposal circumvents this issue by using autologous cell sources to derive RPE cells. Several forms of autologous transplantation therapies for AMD have generated encouraging results, including macular translocation, iris pigment epithelial cell transplant and peripheral retinal RPE cell transplant. But all of these methods have limitations in either cell functionality or the number of cells available for collection and transplantation. This proposal addresses those limitations by using a potentially unlimited source of autologous cells (iPSCs) and several strategies for functional testing and optimization. The reviewers described the research plan as well laid-out and carefully designed. They found the proposal to be feasible and praised the preliminary data showing differentiation and in vitro characterization of RPE cells. Some concerns were expressed about the lack of data describing transduction of RPEs with the viral vectors, whether replenishment of RPEs will be therapeutic for AMD, and whether transplanted cells will suffer the same fate as the original cells. However, reviewers agreed that the assays for RPE function are well described and praised the elegant use of in vitro and in vivo functional tests. Reviewers appreciated the applicant’s appropriate consideration of the need for immune modulation and the risk of tumorigenesis but expressed some concern about the validity of the mouse model for preclinical assessment and cautioned about issues surrounding xenotransplantation in the immune modulation experiments. The reviewers praised the qualifications of the applicant and assembled research team. One reviewer noted that the team is particularly strong in RPE physiology and assays of visual function. Although the team lacks extensive experience in stem cell biology, a core facility is readily available to supply expertise and help in the generation of clinical-grade cells. Reviewers raised significant concerns with the budget, finding it excessive in the number of FTEs, supplies, requested equipment purchases, and travel. Reviewers noted that the available resources and research environment for the proposed studies are excellent. Overall, reviewers were highly enthusiastic about the potential impact of the proposed research and felt that AMD represents one of the best opportunities for advancing stem cell-based therapies to the clinic. Reviewers praised the careful, logical design of the research plan and were optimistic about its feasibility based on impressive and relevant preliminary data.