Genetic engineering of neural stem cells as a flexible model to evaluate functional impact of patient-specific variants in cognitive disorders.

Recent advances in the technology of sequencing have boosted the hunt for novel disease genes in various disorders beyond the throughput of classic functional follow-up. For neurodevelopmental disorders, and more specifically intellectual disability, sequencing both unaffected parents and the patient has been very successful in identifying de novo mutations as a major cause of disease. Using this approach, our group recently identified the ADNP gene as one of the more frequent causes of intellectual disability and autism. However, patients often harbour variants for which interpretation of the functional consequences is less straightforward. Functional investigation of such variants on relevant patient material is most often impossible as this would require brain biopsy. Recently, the technique of inducing pluripotent stem cells from skin biopsies and differentiating them into various cell types, has enabled the generation of patient-derived and disease-relevant in vitro cell cultures. Although this technique is very powerful, it requires large investments in both equipment and expertise to successfully set it up as part of a functional study, while scalability remains limited. Therefore, we propose a mid-throughput and cost-efficient approach for the functional validation of variant causality in disease-relevant tissues. Based on a wildtype culture of human Neural Stem Cells, specific variants can be introduced through Crispr/Cas9, followed by differentiation into neuronal cultures. The feasibility of the assay will be validated using the well defined AnkyrinG protein-protein interaction network for which aberrations have known and measurable outcomes.

Keywords:GENOTYPE-PHENOTYPE CORRELATION, GENOMICS, COGNITION, HUMAN GENETICS

Disciplines:Genetics, Systems biology, Laboratory medicine, Medical systems biology, Molecular and cell biology