Stem-cell mediated therapy is a promising candidate for an extensive array of central nervous system (CNS) diseases including, Parkinson’s disease, Alzheimer’s disease, multiple sclerosis, stroke, and spinal cord injuries. The ability of stem cells to potentially regenerate non-dividing cells such as neurons holds great promise for the development of new therapies to restore damages caused by such debilitating diseases. Recent development of induced pluripotent stem cells (IPSC) further increased the societal viability of such therapeutic strategies by removing the ethical concerns inherent with embryonic stem cells (ESC). IPSCs are derived from cells that form the bodies of organisms as opposed to sperm or egg cells. They also offer the advantage of the possibility of being derived from the patients receiving the treatment, and therefore remove the risk of rejection, requiring no immunosuppressive drugs.
However, despite such great potential and the urgency of the calls for cure, the actual number of approved stem cell therapies to date remains insignificant. Primary reasons for this under-utilized potential include the highly varying therapy efficacy, and the possibility of serious side effects resulting from interference with normal CNS functions. One of the main difficulty lies in the fact that the CNS, mainly consisting of the brain, is a highly complex structure which consists of approximately 100 billion neurons and 300 billion cells supporting the function of these neurons with various cell types and dense connections. Without a way to precisely monitor how these stem cell transplantations perturb the complex CNS system, proper placement of these regenerating elements (stem cells) in the brain and spinal cord becomes an extremely difficult task.
In this proposal, we seek to provide a break-through in a search for the development of stem cell mediated therapies by developing a method to directly visualize the effects of the stem cell transplantation. This will be achieved by combining genetic techniques that will introduce modulatory capability to neural cells based on its cell type. We will strategically introduce capabilities for the underlying neural circuit as well as transplanted stem cells to be modulated by light. For stem cells, we expect our proposed technique to allow modulatory capability from only those developing into specific cell types. This cell-specific modulation capability will then combined with a novel imaging technique called passband b-SSFP fMRI, permitting non-invasive and high-resolution visualization of the regeneration processes. This project, upon its success, will provide direct functional assessment capabilities for the regenerated nerve tissue. This in turn will provide key guidance for developing novel stem cell therapies for CNS diseases.
Statement of Benefit to California:
Stem cell mediated therapy is a highly promising technology that gives us hope for a cure of many debilitating central nervous system (CNS) diseases that do not have an alternative for cure. With recent outstanding developments in stem cell biology, we are accelerating towards the search for a viable therapy. However, one of the main road blocks in developing a full blown therapy utilizing these advances in stem cell biology is the lack of tools to quantitatively monitor the direct effects of the stem cell transplantation. We seek to be instrumental in this search for a cure by introducing a completely novel way of directly assessing the functionality of stem cell driven neural circuitry in vivo. This project, upon its success, will allow accelerated development of novel stem cell therapies for a wide range of brain and spinal cord diseases that include Parkinson’s and Alzheimer’s disease, multiple sclerosis, stroke, and spinal cord injuries.
This technology will function as a bridge between the many outstanding efforts in stem cell biology and the development of stem cell mediated therapy, accelerating the search for a cure. Such accelerated development will not only provide help for many patients in urgent need for care in California, it will also greatly help the California economy by placing the state at the leading edge of the therapeutic method development. This technology will lead to accelerated search for new reagents, protocols, and technologies associated with cell therapy, which will stimulate the economy through patents, royalties, licensing fees, formation of new businesses, and adaptation of the technology by existing biotech companies, increasing California tax revenue and creating new jobs for Californians. The high impact of the proposed research will provide the much needed care for the Californians with debilitating diseases while holding a great promise for placing California at the leading edge of stem cell therapy and care.
The goal of this proposal is to develop a tool to both modulate and monitor activity of transplanted pluripotent stem cell-derived neuronal populations and their effect upon host neuronal circuitry. It is hoped that this will enable the quantitative assessment of functional changes in the brain following cell transplantation. First, the applicants plan to genetically modify induced pluripotent stem cells (iPSC) from a patient with Parkinson’s disease (PD) and a normal sibling control with an inducible, light activated molecular switch. This tool enables researchers to use light to modulate the activity of specific neuronal cell types derived from these cells. The cells will be transplanted into an in vivo model of PD with and without light-mediated activation, and the effect on neuronal circuitry will be monitored using a novel, high spatial resolution functional magnetic resonance imaging technique (fMRI). Behavioral measures of PD will also be performed, and the applicants will correlate the behavioral data to the neuronal circuitry activation data with the ultimate goal of predicting therapeutic outcomes.
Overall, the panel appreciated the sophisticated genetic engineering aspects of the project and described the combination of the molecular switch and fMRI technologies as innovative, creative and synergistic. However, they also agreed this tool does not address a current translational bottleneck. They felt that the proposed technology, if successfully developed, could enable the identification of relevant transplant related events in animal models and ultimately have a beneficial impact on the field. Reviewers were uncertain of the rationale for using both PD and normal cells in these proof of concept studies, as the applicant did not clarify whether the phenotype of PD cells is expected to differ from those of normal controls when transplanted into the PD model.
Reviewers appreciated the convincing preliminary data supporting the applicants’ ability to perform fMRI in vivo, utilize the molecular switch in brain and generate iPSC. They expressed strong concern, however, that no preliminary data were provided to support the challenging cell biology and transplantation related elements of the project. Data substantiating the applicants’ ability to achieve dopaminergic neuron differentiation, transplant cells and perform behavioral testing in the animal model were all lacking, thus dampening reviewers’ confidence the applicants could successfully complete the proposed studies.
Beyond the genetic engineering aspects, reviewers found the experimental plan to be critically lacking in detail; they stressed the stem cell and transplantation portions of the project were particularly weak. Reviewers highlighted a lack of information regarding the stage at which dopaminergic neurons would be transplanted, or whether applicants would indeed differentiate the cells prior to transplantation. Specific Aim 4, correlating therapeutic outcomes in the PD model with changes in neuronal circuits, was noted as especially under developed and ill justified. The applicant fails to discuss expected outcomes, potential pitfalls and present alternative plans. For example, reviewers questioned how the applicants define a therapeutic outcome in the PD model and how they would interpret a subtle phenotype. One discussant asked how the applicants would assess effects of transplanted cells should they migrate away from the injection site and integrate in areas beyond the region interrogated by fMRI. Taken together, these weaknesses further eroded reviewers’ enthusiasm for the proposal.
The principal investigator (PI) is well trained and funded, and reviewers noted this promising individual has received recognition for innovative contributions to the imaging field. However, they raised serious issues regarding the rest of the team. The paucity of expertise in stem cells, dopaminergic differentiation, neural transplantation and the PD model decrease the likelihood that the applicants could achieve the goals of the project. Reviewers also noted the bulk of this technically demanding work would be performed by a small staff of junior scientists, and expressed concern regarding the logistics around sharing a student between the PI and co-investigator at different institutions. Finally, the application does not include letters of collaboration from the stem cell expert and the co-investigator, both of whom are listed at very low levels of effort at 1% and 2%, respectively.
In summary, this application describes the use of a method to control the activity of transplanted neurons and detect the effects using high-resolution fMRI methods in a model of PD. While reviewers appreciated the innovative method and the applicant’s expertise in imaging, they did not feel the application would resolve a translational bottleneck. Further, reviewers concluded the applicants would not be able to successfully complete the goals of the project owing to critical stem cell and transplantation expertise deficiencies of the team as well as flaws and omissions in the experimental design. Thus, the application was not recommended for funding.