Submission
| Title: | Loss of NF2 Disrupts Differentiation in Neuroepithelial Stem Cells |
| Presenter: | Noah Burket |
| Institution: | Indiana University School of Medicine |
| Authors: | Noah Burket1; Scott Cooper1; Victoria Dershem1; Jignesh Tailor, MD, PhD1,2,3
1. Department of Neurosurgery, Indiana University School of Medicine, Indiana 2. Indiana University Melvin and Bren Simon Comprehensive Cancer Center 3. Department of Pediatrics and Herman B Wells Center for Pediatric Research, Indiana University School of Medicine |
Abstract
| Background/Significance/Rationale: | NF2-related schwannomatosis is a tumor predisposition syndrome caused by mutations in the NF2 gene and associated with spinal ependymomas (SP-EPN). These tumors are suspected to originate from mutations in the radial glia (RG) cell lineage. They are only effectively treated through high-risk surgical resection, emphasizing the critical need for identification of targets for medical therapy. Yet, the role of NF2-dependent disruption in RG cell development is poorly understood. We hypothesize that a loss of the NF2 gene in NES cells will prevent normal differentiation and promote a RG-like progenitor state. |
| Methods: | An NF2-knockout was generated in neuroepithelial stem (NES) cells using CRISPR/Cas9. Knockouts were validated using Western blot and Sanger sequencing. In vitro differentiation was induced with removal of growth factors. NF2-knockout phenotypes were assessed with polymerase chain reaction and compared with wildtype NES cells. |
| Results/Findings: | Preliminary data has shown that NF2-knockout cells express similar levels of early pan-neural and neural stem cell genes compared to wildtype after CRISPR editing. The NF2-knockodown cells retain this stem cell-like gene expression following attempted differentiation, whereas wildtype cells take on a primarily neural phenotype. The knockout cells also form what appears to be pre-neoplastic spheres when allowed to differentiate. Two clones that were identified as having NF2 mutations on each allele still retain NF2 protein expression. |
| Conclusions/Discussion: | NF2-knockout NES cells fail to differentiate normally compared to wildtype NES cells. They retain early stem cell-like markers, including DACH1, HES1, and PLZF. Furthermore, formation of spheres when growth factors were removed hints at NF2 loss being relevant to formation of pre-neoplastic growths. Future work will include investigating downstream effects of NF2 loss in this model. |
| Translational/Human Health Impact: | The long-term goal of this project is to use this model to study potential therapies for medical treatment of SP-EPN. |
