Establishing iPSC-based models to investigate the mechanistic interaction between neurotrauma and ALS/FTD

Northwestern University At Chicago, Evanston IL

Investigators

Abstract

Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are devastating neurological disorders with limited treatment options. A hexanucleotide intronic (G4C2)n repeat expansion in C9orf72 is the most common genetic cause of ALS and FTD. However, patients with C9orf72 mutations are characterized by clinical heterogeneity and variable penetrance. The nature of this variability is not well understood but environmental factors have been proposed to act as an additional trigger beyond the pathophysiology associated with the C9orf72 repeat expansion. Moreover, 90% of ALS/FTD cases are classified as sporadic and likely arise from an interaction between genetic predisposition and environmental factors. One such environmental factor is neurotrauma, which results from traumatic brain or spinal cord injury and has been shown to substantially increase the risk of ALS/FTD in epidemiological studies. The association between traumatic injury and ALS/FTD is exemplified by the shared neuropathology TDP-43 aggregation. TDP-43 is a predominantly nuclear RNA binding protein involved in RNA transport and splicing, that forms cytoplasmic aggregations in more than 98% of ALS cases. Widespread TDP-43 aggregation is also seen in patients who have suffered traumatic brain injury (TBI). While these observations indicate a potential link between these diverse neurodegenerative diseases, it is unclear whether the mechanisms that drive TDP-43 pathology in ALS and TBI are shared or divergent. What also remains unclear is whether an ALS genetic background confers heightened vulnerability to neurotrauma. Here, we will use ALS/FTD patient-specific induced pluripotent stem cell (iPSC) derived neuronal cultures in combination with a custom-built device that applies biofidelic mechanical stretch-induced trauma in vitro, to establish novel human models that mimic the impact of traumatic injury. The innovative instrument can effectively recapitulate a key facet of TBI in diffuse axonal injury. In key preliminary work we found that C9orf72, but not isogenic control motor neurons exhibit selective degeneration and extended cytoplasmic accumulation of TDP- 43 in response to trauma. In Aim 1 we will interrogate several ALS C9orf72 iPSC lines and controls to test the hypothesis that stretch-trauma exacerbates C9orf72 pathology. We will measure both sense and antisense transcript and dipeptide repeat protein production in response to varying degrees of traumatic injury. We will investigate the ability of sense and antisense targeting ASOs to protect neurons from injury to dissect the relative contribution of these mechanisms to stress-induced mutant C9orf72 toxicity. In Aim 2 we will generate spinal and cortical neurons from 12 sporadic ALS and 12 age/gender matched control iPSC lines and interrogate their ability to cope with varying degrees of mechanical trauma. We will examine neuronal survival, the subcellular localization, solubility and functionality of TDP-43 protein using biochemical and RNA-Seq approaches. Our work will capture the interaction between mechanical trauma and an ALS/FTD genotype in a controlled environment that allows definitive hypothesis testing and mechanistic experiments