Although epilepsy has a large genetic contribution, causative genes are not known for most epilepsy patients. And while epilepsy can be inherited in families, it is more commonly 'sporadic'or spontaneous, without a clear family history, and causes of sporadic epilepsy cases are mostly unknown. Since the cause of epilepsy often directs the choice of treatments, categorizing patients is of great importance therapeutically. Many epilepsy syndromes appear to involve 'somatic'mutations, in which a subpopulation of brain cells shows a mutation not shared by all cells of the body, because the mutation occurred during mitosis of somatic cells of the embryo after fertilization. Our lab and other have reported on somatic mutations is several genes known to cause epilepsy, however until now, technological limitations have prevented a systematic search for somatic mutations in epilepsy. Whole genome or whole exome sequencing will not identify somatic mutations unless the study is designed to detect them: responsible mutations may not be present in most blood cells, but would instead be limited to cells in the brain. Even relevant mutations limited to neurons may be missed by bulk brain sequencing because 1] neurons are surrounded by glial cells with distinct embryological origins, and with whom they would not be expected to share mutations, and 2] neurons themselves are derived from two embryological origins, with pyramidal neurons derived from cortical proliferative regions, and inhibitory nonpyramidal neurons migrating into cortex from distant subcortical regions, so that bulk sequencing of postmortem brain tissue may also not detect mutations limited to specific neuronal populations. We have recently developed technology that allows sorting of single or small numbers of cerebral cortical neuronal nuclei, and amplification of their genomes in quantities sufficient for any next-generation sequencing. We are proposing to use this technology to examine cortical neurons from postmortem brains or cortical resections from epilepsy patients in order to 1) perform a systematic assessment of copy number variation (CNV) in cerebral neurons;and 2) perform whole exome sequencing to identify mutations in known epilepsy genes. Comparison of single neuron sequence to that from other cells of the body could then determine how frequently spontaneous mutations occur in cortical neurons, and identify and catalogue these mutations. Our proposed study thus enables a systematic analysis of all mechanisms of somatic mutation in the epileptic human brain.
Epilepsy is a major health problem, with lifetime risk of a seizure being 3%. Our work challenges major paradigms about epilepsy causation that suggest that epilepsy can be a 'complex trait'due to multiple interacting genetic mutations, and instead suggests that epilepsy can be caused by developmental genetic anomalies that occur in only some of the neurons of the brain. Our work will not only improve the specificity of epilepsy diagnosis, but also promises to provide a new paradigm for considering epilepsy treatment. Disclaimer: Note that reviewers were given the following special instructions for the review of these Exceptional Unconventional Research Enabling Knowledge Accelerations (EUREKA) grant applications: The purpose of the EUREKA initiative is to foster exceptionally innovative research that, if successful, will have an unusually high impact on the areas of science that are germane to the mission of one or more of the participating NIH institutes. EUREKA is for new projects. EUREKA is not for the continuation of existing projects. EUREKA is not for support of pilot projects (i.e., projects of limited scope that are designed primarily to generate data that wll enable the PI to seek other funding opportunities). Rather, it is anticipated that EUREKA projects will begin and be completed during the funding period. Please provide an opinion and assessment of the likelihood the project will exert a sustained, powerful influence on the research field(s) involved. Significance and innovation should be the major determinants of your overall impact score. The approach should be evaluated for general feasibility. An application should score poorly if it is clear to the reviewers that the proposed methodology has no probability at all of being successful, either because it is inherently illogical or because the sae approach has already been attempted and shown not to be feasible. Remember that unavoidable risk, which is intrinsic to novel and innovative approaches, is expected for these applications and reviewers are instructed that the presence or absence of preliminary data should not be taken into account when determining the score. Applications that are good science for standard research type investigation, but not likely to exert a sustained and powerful influence on the field, should be scored down. Also note that the following critiques were prepared by the reviewers prior to the Study Section meeting and are provided in an essentially unedited form. While there is opportunity for the reviewers to update or revise their written evaluation, based upon the group's discussion, there is no guarantee that individual critiques have been updated subsequent to the discussion at the meeting. Therefore, the critiques may not fully reflect the final opinions of the individual reviewers at the close of group discussion or the final majority opinion of the group. Thus the Resume and Summary of Discussion is the final word on what the reviewers actually considered critical at the meeting.
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