EVALUATION OF G9A INHIBITION IN THE BTBR MOUSE MODEL OF AUTISM SPECTRUM DISORDER USING MULTI-OMICS INTEGRATION

Autism spectrum disorder (ASD) is a complex neurodevelopmental condition characterized by genetic, epigenetic, and neural circuit abnormalities. It affects approximately 1 in 100 children worldwide, and there are currently no pharmacological treatments targeting its core behavioral symptoms. Among the epigenetic mechanisms implicated in ASD, dysregulation of histone H3K9 dimethylation has emerged as a key factor. This modification is mediated by the methyltransferase G9a/EHMT2 and can negatively impact synaptic function and neurodevelopment. FLAV-27 is a novel SAM-competitive inhibitor of G9a designed to reduce pathological H3K9me2 levels and restore transcriptional pathways linked to ASD.

In this study, a multi-omics approach investigated the behavioral, molecular, and chromatin-level effects of FLAV-27 (5 and 10 mg/kg, intraperitoneal) in BTBR T+tf/J (BTBR) mice, a well-established model of idiopathic ASD. Behavioral assessments included marble burying, cotton shredding, and the three-chamber social interaction test to evaluate repetitive behaviors and sociability. These analyses were complemented by bulk RNA sequencing, ATAC-seq, and molecular validation techniques.

Acute FLAV-27 administration significantly reduced repetitive behaviors in BTBR mice dose-dependently, restoring wild-type performance and surpassing the effects of aripiprazole. Treatment also improved sociability and social novelty, though less than aripiprazole. Hippocampal transcriptomics showed increased expression of synaptic plasticity, GPCR signaling, and dopaminergic genes, with reduced stress-related and inflammatory expression. ATAC-seq revealed chromatin remodeling at neuronal regulatory regions, and Western blotting confirmed reduced H3K9me2 and modulation of ERK and SNAP25. Overall, these findings support G9a inhibition as a promising therapeutic strategy for ASD.