Background: Partial or an entire deletion of SHANK3 are considered as major drivers in the Phelan-McDermid syndrome, in which 75% of patients are diagnosed with autism spectrum disorder (ASD). During the recent years, there was an increasing interest in stem cell therapy in ASD, and specifically, mesenchymal stem cells (MSC). Moreover, it has been suggested that the therapeutic effect of the MSC is mediated mainly via the secretion of small extracellular vesicle that contains important molecular information of the cell and are used for cell-to-cell communication. Within the fraction of the extracellular vesicles, exosomes were highlighted as the most effective ones to convey the therapeutic effect. Methods: Exosomes derived from MSC (MSC-exo) were purified, characterized, and given via intranasal administration to Shank3B KO mice (in the concentration of 107 particles/ml). Three weeks post treatment, the mice were tested for behavioral scoring, and their results were compared with saline-treated control and their wild-type littermates. Results: Intranasal treatment with MSC-exo improves the social behavior deficit in multiple paradigms, increases vocalization, and reduces repetitive behaviors. We also observed an increase of GABARB1 in the prefrontal cortex. Conclusions: Herein, we hypothesized that MSC-exo would have a direct beneficial effect on the behavioral autistic-like phenotype of the genetically modified Shank3B KO mouse model of autism. Taken together, our data indicate that intranasal treatment with MSC-exo improves the core ASD-like deficits of this mouse model of autism and therefore has the potential to treat ASD patients carrying the Shank3 mutation.