Using patient-specific iPSC derived dopaminergic neurons to overcome a major bottleneck in Parkinson's disease research and drug discovery
Early Translational I
The goals of this study are to develop patient-specific induced pluripotent cell lines (iPSCs) from patients with Parkinson’s disease (PD) with defined mutations and sporadic forms of the disease. Recent groundbreaking discoveries allow us now to use adult human skin cells, transduce them with specific genes, and generate cells that exhibit characteristics of embryonic stem cells, termed induced pluripotent stem cells (iPSCs). These lines will be used as an experimental pre-clinical model to study disease mechanisms unique to PD. We predict that these cells will not only serve an ‘authentic’ model for PD when further differentiated into the specific dopaminergic neurons, but that these cells are pathologically affected with PD. The specific objectives of these studies are to (1) establish a bank of iPSCs from patients with idiopathic PD and patients with defined mutations in genes associated with PD, (2) differentiate iPSCs into dopaminergic neurons and assess neurochemical and neuropathological characteristics of PD of these cells in vitro, and (3) test the hypothesis that specific pharmacologic agents can be used to block or reverse pathological phenotypes. The absence of cellular models of Parkinson’s disease represents a major bottleneck in the scientific field of PD, which, if solved in this collaborative effort, would be instantly translated into a wide range of clinical applications, including drug discovery. This research is highly translational, as the final component is aimed at testing lead compounds that could be neuroprotective, and ultimately at developing a high-throughput drug screening program to discover new disease modifying compounds. This is an essential avenue if we want to offer our patients a new therapeutic approach that can give them a near normal life after being diagnosed with this progressively disabling disease.
Statement of Benefit to California:
Approx. 36,000-60,000 people in the State of California are affected with Parkinson’s disease (PD), a common neurodegenerative disease that causes a high degree of disability and financial burden for our health care system. It is estimated that the number of PD cases will double by the year 2030. We have a critical need for novel therapies that will prevent or even reverse neuronal cell loss of specific neurons in the brain of patients. This collaborative proposal will provide real benefits and values to the state of California and its citizens in providing new approaches for understanding disease mechanisms, diagnostic tools and drug discovery of novel treatment for PD. Reprogramming of adult skin cells to a pluripotent state is the underlying mechanism upon which this application is built upon and offers an attractive avenue of research in this case to develop an ‘authentic’ pre-clinical model of PD. The rationale for the proposed research is that differentiated pluripotent stem cells from patients with known genetic forms of PD will recapitulate in vitro one or more of the key molecular aspects of neural degeneration associated with PD and thus provide an entirely novel human cellular system for investigation PD-related disease pathways and for drug discovery. The impact of this collaborative research project, if successful, is difficult to over-estimate. The scientific field has been struggling with the inability to directly access cells that are affected by the disease process that underlies PD and therefore all research and drug discovery has relied on ”best guess” models of the disease. Thus, the absence of cellular models of Parkinson’s disease represents a huge bottleneck in the field.
This proposal addresses a bottleneck in Parkinson’s disease (PD) research: the development of a predictive, in vitro model for studying disease mechanisms and performing drug discovery. In Aim 1, the applicant proposes to first expand their bank of cell samples from patients with idiopathic PD, patients with defined mutations associated with PD, and age-matched controls; the applicant will generate induced pluripotent stem cells (iPSCs) from this entire collection. In Aim 2, the applicant then proposes to differentiate these iPSC lines into dopaminergic (DA) neurons and assess their biochemical, phenotypic, and pathological characteristics. Finally, in Aim 3, the applicant plans to develop a high-throughput screening method to identify disease-modifying agents, focusing on reversal of pathological phenotypes discovered under Aim 2. The reviewers agreed that the impact of this proposal could be very high. They felt that an iPSC-derived model of PD would have tremendous potential for elucidating the pathogenesis of the disease and accelerating the development of new disease-modifying therapies through human cell-based drug discovery. However, reviewers cautioned that cells isolated from patients (and subjected to culture and manipulation) might not develop a phenotype that predicatively models the disease. However, the likelihood of finding a phenotype is enhanced by the richness of the patients with varieties of the disease available for the research, and by preliminary data suggesting a relevant phenotype in some of the cell lines. Reviewers noted that the proposal’s potential impact is further broadened by the plan to make cell lines available through a stem cell bank, which would permit widespread distribution for drug discovery efforts to scientists of many disciplines. The reviewers found the research plan to be clearly articulated and logically described. They appreciated its discussion of potential pitfalls and alternative approaches. Reviewers praised the considerable preliminary data supporting Aims 1 & 3, describing the data as strong and compelling. The applicant has demonstrated the creation of iPSC lines from one hereditary PD patient and three normal controls. The applicant has also identified 80 compounds effective in blocking pathogenic protein aggregation to various degrees, 11 of which actually disaggregate alpha-synuclein fibrils. Reviewers felt that Aim 2 was the least developed part of the proposal. One reviewer felt that the preliminary data supporting this aim were relatively weak and noted that entire proposal hinges on producing relatively large numbers of A9 DA neurons in a high-throughput fashion. However, this reviewer did appreciate the applicant’s rational and coherent plan for assessing the phenotype of DA neurons and demonstrating that they have the salient features of A9-specific ventral mesencephalic cells. Another reviewer commented that while DA neuron yields with their current methods may be low, strategies exist for increasing yield, and even small numbers of neurons will be useful. But this reviewer wondered how long a period of observation would be necessary to see signs of pathology without application of artificial stressors to push the phenotype, since PD is a disease of aging. This was a general concern of the reviewers, that the proposal’s feasibility was difficult to judge because it’s impossible to know whether iPSC-derived DA neurons will develop relevant hallmarks of PD in culture. However they also found that the PI clearly addressed this possibility and provided a cogent plan to accelerate the degeneration of cultured DA neurons with toxins and other known contributors to the pathologic phenotype. After this discussion reviewers ultimately found the plan to be a clever approach likely to result in novel translationally relevant findings. One reviewer cited the generation of cell lines with mutations in many different PD-associated genes as a strength and suggested increasing the number of cell lines/patients in order to confirm results between patients with similar mutations. The applicant is one of the world’s leading experts on PD and uniquely qualified to lead this research effort. Reviewers praised the formidable group of prominent neuroscientists assembled as collaborators, many of who have considerable expertise in translational research. Reviewers judged the resources and research environment to be excellent and commented that the applicant’s patient database is one of the best in the world. Overall, the reviewers were extremely enthusiastic about this proposal and felt that it has the potential to overcome a major bottleneck in PD research, accelerating drug discovery. They raised some concerns about feasibility but expressed a high level of confidence in the assembled research team’s ability to complete the proposed studies.