Human iPSC-derived motor neurons with TDP-43 nuclear loss present early ALS pathological phenotypes

Amyotrophic lateral sclerosis (ALS) is a complex neurodegenerative disorder characterized by progressive loss of motor neurons and presents a critical need for effective treatments. To investigate the impact of TAR DNA-binding protein 43 (TDP-43) nuclear depletion in ALS, we established and characterized a human-induced pluripotent stem cell-derived motor neuron model carrying a mutation that disrupts TDP-43 nuclear localization. The model successfully recapitulates functional deficits observed in ALS patients, including impaired neurite outgrowth, reduced axonal regeneration, and neuronal hyperexcitability.

Proteomic and metabolomic profiling revealed early mechanisms underlying ALS neuronal pathophysiology, involving altered oxidative stress and neurotransmitter release. TDP-43 nuclear depletion led to mis-splicing STMN2 and KCNQ2, defects characteristic of TDP-43-associated disease. Notably, a splice-switching antisense oligonucleotide successfully restored normal STMN2 splicing and function, demonstrating that directly correcting splicing defects can rescue motor neurons.

This work provides a comprehensive understanding of TDP-43 nuclear depletion in the context of ALS pathogenesis and offers potential targets for RNA-targeted therapeutic intervention, pointing towards the development of precision medicine approaches and early diagnostic markers for ALS. We thank Brian Henry, Jason Uslaner, Sean Smith and Simonne Longerich for their valuable support. We acknowledge Sophie Parmentier-Batteur and Yinghui Hu for generating the cell lines at Merck Sharp & Dohme LLC, a subsidiary of Merck & Co., Inc., Rahway, NJ, USA.

We would like to thank Philip Wong (Johns Hopkins Medicine) for an early characterization of TDP-43WT and TDP-43ΔNLS iMN, which formed the foundation of this study. The patch clamp electrophysiology work has been contributed by Dr Huang Hua from Electrophysiology Core Facility@NUS Medicine. We also appreciate Neha Raghavan and Jason E.

Miller for genetic analysis of STMN2, Yong Qing Victoria Ong for co-culture maintenance, Matthew S. Choo, Esther Sok Hwee Cheow and Yeun Li Koh for omics sample preparation and data analysis. The study was sponsored and funded by Merck Sharp & Dohme LLC, a subsidiary of Merck & Co., Inc., Rahway, NJ, USA.

The sponsor was involved in the study design and conceptualization, execution of the study, collection, analysis, and interpretation of the data. Quantitative Biosciences, MSD International GmbH (Singapore Branch), Singapore, Singapore Tze Khee Chan, U-Ming Lim, Emelyne Teo, Nikhil Kumar Tulsian & Aaron Zefrin Fernandis Neuroscience, Merck & Co., Inc., West Point, PA, USA Shahriar Niroomand, Michael J. Klein Precision Genetics, Data, AI & Genome Sciences, Merck & Co., Inc., Cambridge, MA, USA Discovery Chemistry, Merck & Co., Inc., Rahway, NJ, USA Discovery Chemistry, Merck & Co., Inc., Boston, MA, USA Department of Pharmacy, Faculty of Science, National University of Singapore, Singapore, Singapore Yong Loo Lin School of Medicine, National University of Singapore, Singapore, Singapore Former employee of Neuroscience, Merck & Co., Inc., West Point, PA, USA Early Discovery Genetics, Data and Genome Sciences, Merck & Co., Inc., Cambridge, MA, USA Correspondence to Aaron Zefrin Fernandis.