Discovery of a pyrazolopyridine alkaloid inhibitor of ERO1A that mitigates neuronal ER stress and age-related decline

Targeting Endoplasmic Reticulum Oxidoreductase 1 Alpha (ERO1A) offers therapeutic potential for ER stress-related conditions, including motor neurone diseases and congenital muscle disorders. However, selective ERO1A inhibitors remain unavailable. Here, we developed a multi-modal discovery pipeline combining molecular docking with in vitro and in vivo assays, screening 401,824 natural products from the COCONUT database. We identified two compounds, S88 and geniposide, that inhibited ERO1A in vitro , reduced tunicamycin-induced ER stress markers in human neurons, and improved locomotion and neuromuscular junctions in UBQLN2 ^ALS Drosophila . S88 maintained efficacy in late-stage disease and extended lifespan in a D-galactose aging model. In fly brains, S88 selectively reduced phosphorylated eIF2α without lowering its downstream effector ATF4, suggesting engagement of alternative mechanisms preserving ATF4 homeostasis. These findings highlight S88 as a promising lead for treating ER stress-associated neuromuscular disorders and demonstrate the utility of this integrative discovery pipeline for identifying bioactive natural compounds with disease-modifying potential.