Small molecules targeting ARF1 interaction with C9orf72:SMCR8:WDR41 complexes suppress its overactivation implicated in ALS/FTD

The hexanucleotide repeat expansion in C9orf72 gene is the most common genetic cause of amyotrophic lateral sclerosis (ALS)/frontotemporal dementia (FTD). The C9orf72 protein forms a complex with SMCR8 and WDR41 (CSW), which functions as a GTPase-activating protein (GAP) regulating ARF1 and RAB small GTPases. While these findings implicated ARF1-GAP dysregulation in ALS/FTD and supported ARF1 suppression as potential intervention, small molecules that modulate ARF1-CSW interactions are lacking. In this study, we demonstrated upregulation of tyrosine-phosphorylated (Tyr-782) ASAP1 (also known as AMAP1, DDEF1, or Centaurin {beta}4), an ARF-GAP, in human motor cortex of both sporadic ALS and ALS with C9orf72 mutations. Ectopic C9orf72 expression partially mimicked the effects of a known ARF1 inhibitor brefeldin A to disperse Golgi apparatus. Computer-aided rational drug design with high-throughput in-silico screening identified MCULE-5095997944 (Named as SCC944) as a ARF1-CSW modulator. SCC944 binds directly to ARF1 and reduced GTP-bound ARF1 levels upon ARF1 activation. SCC944 demonstrated brefeldin A-like ARF1-dependent alteration of organelle organization including Golgi, microtubules, and mitochondria, but also a protein trafficking pattern that is distinct from brefeldin A mechanism. These studies identified the first small molecule targeting ARF1-CSW interaction and further support ARF1 modulation as a potential therapeutic approach for ALS/FTD.