High-throughput screening of ALS patient iPSC-derived spinal motor neurons identifies novel compounds that increase neurofilament light chain expression

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease primarily affecting motor neurons both in the spinal cord and brain. The cardinal pathology of ALS is motor neuron-selective inclusion of proteins such as TDP43, SOD1, C9orf72-derived dipeptide repeats, or FUS due to the mutations in the genes encoding them. Both familial and sporadic forms of ALS also show neurofilament (NF) aggregates, attributed to an imbalance in subunit expression, particularly a decrease in neurofilament light chain (NF-L) levels. Current FDA-approved treatments extend survival for only a few months, highlighting the urgent need for new therapies. In this study, we developed a cell-based reporter system for high-throughput screening by engineering induced pluripotent stem cells (iPSCs) derived from ALS patients and differentiating them into spinal motor neurons. We screened over 6000 compounds using these reporter iPSC-derived motor neurons and identified a novel compound that increases NF-L expression by >50 %. However, this novel compound also inhibits TGF-β signaling, prompting us to optimize its activity through a hit-to-lead chemistry analysis. In our subsequent investigations, we identified an additional compound that does not affect TGF-β signaling and outperforms the original compound in both in vitro and in vivo drug metabolism and pharmacokinetics assays. Our study highlights the utility of iPSC-derived neurons in disease modeling and illustrates how they can be employed to discover new compounds for therapeutic development through extensive screening in disease-relevant settings.