Dual developmental effects of ARX poly-alanine mutations on human cortical excitatory and inhibitory neurons

Infantile spasms (IS), a severe childhood epilepsy with an incidence of 1.6-4.5 per 10,000 live births, often lead to lifelong intellectual disability. Up to 5% of affected males carry mutations in the Aristaless-related homeobox (ARX) gene. The lack of human-specific models for developmental epilepsy limits progress, making organoids a promising alternative. We use human cortical organoids (COs) and ganglionic eminence organoids (GEOs) to model poly-alanine expansion (PAE) mutations in ARX. PAE mutations increase cortical progenitor proliferation and accelerate early cortical development. ARX expression is upregulated in patient-derived COs at 30 days in vitro (DIV), correlating with altered cell cycle gene expression. We observe enhanced, cell-autonomous interneuron migration, which is rescued by CXCR4 inhibition. ARX^PAE assembloids exhibit early network hyperactivity. These findings highlight the utility of human brain organoids in uncovering ARX^PAE-driven mechanisms and represent a critical step toward developing targeted therapies for IS and related developmental epilepsies.