There is a group of brain diseases that are caused by functional abnormalities. The brains of patients afflicted with these diseases which include autism spectrum disorders, schizophrenia, depression, and mania and other psychiatric diseases have a normal appearance and show no structural changes. Neurons, the cellular units of the brain, function by making connections (or synapses) with each other and exchanging information in form of electric activity. Thus, it is believed that in those diseases many of these connections are not working properly. However, using current technology, there is no way to investigate individual neuronal synapses in the human brain. This is because it is not ethical to biopsy the brain of a living person if it is not for the direct benefit to the patient. Therefore, scientists cannot study synaptic function in psychiatric diseases. Because of the limited knowledge about the functional consequences in the affected brains, there is no cure for these diseases and the few existing therapies are often associated with severe side effects and cannot restore the normal function of the brain. Therefore, it is of great importance to better study the disease processes. A better knowledge on what the defects are on the cellular level will enable us then in a second step to test existing drugs and measure its effect or screen for new therapeutic drugs that can improve the process and hopefully also the disease symptoms.
This proposal aims to develop a technology to overcome this limitation and ultimately provide neurons directly derived from affected patients. This will uniquely allow the study functional neuronal aspects in the patients' own neurons without the need to extract neurons from the brain. Our proposal has two steps, that we want to undertake in parallel with mouse and human cells. First, we want to find ways to optimally generate neurons from skin fibroblasts. Naturally, these artificial neurons will have to exhibit all functional properties that the neurons from the brain have. This includes their ability to form functional connections with each other that serve to exchange information between two cells. In the second step, we will generate such neuronal cells from a genetic form of a psychiatric disease and evaluate whether these cultured neuronal cells indeed exhibit changes in their functional behavior such as the formation of fewer connections or a decreased probability to activate a connection and thus limit the disease cells to communicate with other cells.
Our proposed research is to develop a cellular tool which will enable the research community to study human brain diseases that are caused by improperly functioning connections between brain cells rather than structural abnormalities of the brain such as degeneration of neurons or developmental abnormalities. These diseases, which are typically classified as psychiatric diseases, include schizophrenia, bipolar diseases (depression, mania) autism spectrum disorders, and others. There are many people in California and world-wide that suffer from these mentally debilitating diseases. Therefore, there is a great need to develop therapies for these diseases. However, currently drug development is largely restricted to animal models and very often drug candidates that are successful in e.g. rodent animals can not be applied to human. It would thus be much better to possess a model that reflects the human disease much closer, ideally using human cells.
We have experimental evidence that we can develop such a model. In particular, we will convert skin cells from patients suffering from psychiatric diseases into stem cells that are "pluripotent", which means they can differentiate into all cell types of the body including neurons. We want to explore whether these patient-derived neurons still contain the disease features that the neurons have in the brain. If we could indeed capture the disease in these cells, our technology would have a major impact on future work in this area. We believe that this approach could be applied to many neurological diseases including neurodegenerative diseases.
Our technology would not only provide a unique experimental basis to begin to understand how these diseases work, but it would allow to then interfere with the identified cellular abnormalities which would secondarily result in the development of new drugs that can counteract the diseases and would hopefully also work for the patients themselves.
Therefore, all those Californians that suffer from one of the above mentioned diseases will benefit from our research project, if it is successful.
The goal of this proposal is to develop cellular tools for studying neurological brain diseases affecting synaptic function. Many neuropsychiatric disorders are considered functional diseases exhibiting no histopathological abnormalities. The reason for our limited understanding is the lack of experimental access to the human brain. Such studies are limited to post-mortem analysis and brain imaging. Existing technology to derive functional neurons is difficult, time consuming, and variable from laboratory to laboratory. The team proposes to improve upon the existing technologies currently available to generate neurons from human embryonic stem cells (hESCs), induced pluripotent stem cells (hiPSCs), and from fibroblasts. In Aims 1 and 2, the applicant proposes to use neuronal transcription factors to optimize the induction of functional neurons from hESCs and hiPSCs and to directly convert human fibroblasts to neurons. The applicant then proposes in Aim 3 to perform proof of principle studies for the human work proposed in Aim 4 using a mutant mouse model for Rett syndrome. In this aim, the applicant will conduct morphological and electrophysiological studies on neurons converted directly from fibroblasts obtained from the mutant mouse. Finally, in Aim 4, the applicant will generate neurons from Rett syndrome patients using both iPSC technology and by converting neurons directly from patient fibroblasts. Finally, the PI will interrogate the synaptic function of these neurons in order to elucidate underlying pathologic abnormalities.
Reviewers considered the overall proposed research highly creative and important to the study of neuropsychiatric disorders that arise from functional disturbance and have no known pathological abnormalities. Reviewers commented that the proposal, if successful, would result in an innovative new technology that employs tools with a potential broad impact beyond the proposed application in Rett syndrome. The inclusion of electrophysiological functional validation studies was noted to be particularly innovative and added to the overall impact of the work proposed. Reviewers did note that the high impact is dependent upon the success of Aims 1 and 2 and a subset of reviewers thought that the focus on Rett syndrome in Aims 3 and 4 detracted from the overall innovative nature of the proposal as there are multiple laboratories working in this area.
In general, the experimental design and scientific approach was considered logical and feasible. The presented preliminary data was quite good, appropriate, and supported the likelihood that the PI would be successful in meeting the proposed milestones and timelines. However, reviewers were concerned that the description of alternative plans and methodologies was underdeveloped in the proposal. While potential pitfalls were addressed, reviewers thought that greater difficulties than suggested in the proposal would be encountered, and the investigator is likely to have considered this issue and should have articulated more fully the alternative plans. Reviewers agreed that the first two aims of the proposal are fabulous. The enthusiasm for Aims 3 and 4 was tempered by the concerns of some reviewers that the proposed studies in Rett syndrome might not be the best follow up to the neuron derivation studies. In addition to the possible derivative nature of these proposed studies, the PI did not mention the specific mutations that will be studied, nor did s/he consider how these different mutations might have phenotypes that are more or less easy to identify. Moreover, one reviewer felt that a discussion of how mutant cells will be identified in a mixed population was necessary but lacking in the proposal. Reviewers suggested that generation of a polycistronic construct composed of the identified transcription factors would provide a more useful tool for the research community and might alleviate some of the difficulties with optimizing conditions.
The reviewers universally praised the PI. The PI’s publication and track record is outstanding and this individual is considered a rising star in the field. The assembled team is small but consists of respected researchers who are extremely capable of performing the proposed research, including a leading researcher in synaptic biology and neurotransmission as well as a recognized expert in Rett syndrome. In addition, the environment is excellent and two postdoctoral fellows are already available for the project, which adds to the overall attractiveness of the team.
Overall, reviewers considered Aims 1 and 2 of this proposal highly innovative, impactful, and excellent and, although there were concerns regarding the scientific direction chosen in Aims 3 and 4 by some of the reviewers, this proposal from an outstanding young investigator was recommended for funding.
- A motion was made to move this application into Tier 1, Recommended for Funding. No specific programmatic reason was identified. The motion carried.