Modulating VCP to mitigate pathological mechanisms related to C9ORF72 mutation in ALS

Amyotrophic Lateral Sclerosis (ALS) is a fatal neurodegenerative disease characterized by the degeneration of upper and lower motor neurons. Various pathological mechanisms contribute to motor neuronal death, including the accumulation of toxic misfolded protein aggregates and lysosomal dysfunction. Many ALS cases are linked to a hexanucleotide repeat expansion (G4C2) in the C9ORF72 gene. The expansion produces five dipeptide repeat proteins (DPRs) that aggregate and lead to cell death. DPRs disrupt organelle function and the protein quality control (PQC) system. We investigated the biochemical behavior of each DPR in immortalized motoneurons, confirming their accumulation and localization, and examined their impact on lysosome stability. All DPRs caused lysosomal membrane damage and activity alterations, exacerbated by impaired autophagy induction. DPRs blocked the nuclear localization of transcription factors (TFEB and TFE3) essential for autophagy and lysosome biogenesis, preventing the removal of damaged lysosomes. Valosin-containing protein (VCP), an ATPase involved in multiple PQC pathways, was analyzed to rescue DPR toxicity. Overexpressing VCP in these models rescued the accumulation of the most toxic DPRs, enhancing clearance via the ubiquitin-proteasome system (UPS) and/or autophagy. VCP overexpression also reduced lysosomal alterations, likely due to increased DPR clearance. Chemical induction of VCP using SMER28 decreased DPR levels in both immortalized motoneurons and iPSC-derived motor neurons (iPSC-MNs). These findings suggest that VCP modulation could be a potential therapeutic target to reduce DPR-mediated toxicity and improve motoneuron survival in ALS.