Amyotrophic lateral sclerosis (ALS) is an adult-onset disorder characterized by progressive paralysis caused by the degeneration of motor neurons (MNs). Although most cases are sporadic, about 10 % are familial and caused by genetic mutations in genes such as SOD1, FUS, and C9ORF72. Transgenic mice expressing mutant SOD1 recapitulate hallmarks of ALS pathology including MN degeneration, mitochondrial dysfunction, aggregation of SOD1 protein, progressive paralysis, and shortened lifespan. Because mitochondria play a critical role in cell survival, many groups have sought to protect MNs from degeneration by protecting mitochondrial function from ALS pathology. However, paralysis in ALS patients is not the result of MNs degeneration, but, rather, degeneration of neuromuscular junctions (NMJs), which later progresses to MN loss. As a result, any therapies aiming only at preserving MN survival are likely to have no effect on ALS pathogenesis. Instead, it is essential to focus on preserving NMJ structure and function.
iPSCs are poised to revolutionize our understanding of ALS and to enable the identification of novel therapeutics. Through reprogramming, iPSCs can be derived from an ALS patient with a specific phenotype and genotype. ALS iPSC-derived MNs can then be used to recapitulate the disease pathogenesis of the donor patient. Using gene correction, isogenic iPSCs have been generated from ALS patients with mutant SOD1 and demonstrate that iPSC-derived MNs from ALS patients recapitulate ALS relevant phenotypes in vitro. Because NMJ loss is the initial cause of paralysis in ALS patients, we argue that a scalable platform for modeling NMJ dysfunction using iPSCs would be a powerful tool to identify novel ALS therapeutics.
The project aims at developing and validating a platform technology to enable NMJ-based models of ALS for compound screening. Together with the research group of PD Dr. Dr. Andreas Hermann we have designed and are manufacturing a prototype plate that connects MNs and myotubes in a highly reproducible pattern that is scalable and compatible with high – throughput screening (HTS). We propose to develop this tool and using it to generate a first – in – class model of ALS based on NMJ function. We aim at incorporating electrodes into our plates to enable HTS of NMJ function. We previously generated iPSC lines from ALS patients with mutations in FUS and C9ORF72 and are performing gene correction on these mutations to generate isogenic iPSC lines. We propose using these MNs differentiated from these isogenic iPSC lines to test, through use of our novel plates, the effects of ALS mutations on NMJ degeneration, which is directly comparable to the initial event causing paralysis in patients. Finally, we propose performing a pilot screen of 1000 known tool compounds to validate both the use of our plates and NMJ degeneration as a screening platform. The results of this project could facilitate ALS research and give insight into new therapies.