Exosomes can modulate the early hyperexcitability in cortical neurons with ASD-associated Shank3 mutation.

Extracellular vesicles (EVs) are lipid membrane-bound structures that mediate intercellular communication by transferring diverse cargoes, including RNA and proteins. Shank3, a synaptic scaffolding protein critical for synapse structure and function, is implicated in autism spectrum disorder (ASD) and Phelan-McDermid Syndrome (PMS). Early hyperexcitability in cortical neurons is a recognized endophenotype in ASD. Here, we investigated EV-mediated effects in the context of Shank3 deficiency using human iPSC-derived cortical neurons and Shank3B-/- mice. Switching EVs between Shank3 mutant and control neurons revealed that Shank3 mutant-derived EVs transferred the hyperexcitability and accelerated maturation phenotypes to control neurons. This was driven by enriched synaptic proteins (e.g., ACTB, CFL1, AGRN, CLSTN1) in Shank3 mutant-derived EVs as confirmed by proteomic analysis. Conversely, control EVs failed to rescue mutant phenotypes consistent with their lower enrichment for synaptic proteins and related pathways. Further, EVs from mesenchymal stem cells (MSCs) and healthy donor iPSCs, containing synaptic modulators such as complement proteins (C1R, C1S), plasticity-related proteins (MDK, IGFBP3), and homeostatic regulators (FGF2, SFRP1), rescued the hyperexcitability and normalized the maturation in Shank3 mutant neurons. Moreover, intranasal administration of iPSC-derived EVs in Shank3B-/- mice significantly ameliorated ASD-like behavioral deficits, underscoring their therapeutic potential. Together, these findings reveal a novel EV-mediated mechanism for modulating dysregulated excitability and synaptic maturation, addressing a critical unmet need in ASD and related neurodevelopmental disorders treatment.