Females with autism spectrum disorders show stronger DNA methylation signatures than males in perinatal tissues.

Autism spectrum disorder (ASD) comprises a group of neurodevelopmental conditions currently diagnosed by behavioral assessment in childhood, with reported underdiagnosis in females. Though diagnosis in early life is linked to improved outcomes, we currently lack objective screening tools for newborns. To address this gap, we sought to identify a sex-specific DNA methylation signature for ASD using perinatal tissues that reflect dysregulation in the brain. DNA methylation was assayed from ASD and typically developing (TD) newborn blood, umbilical cord blood, placenta, and post-mortem cortex samples using whole genome bisulfite sequencing (WGBS) in a total of 511 samples. We found that methylation levels of differentially methylated regions (DMRs) differentiated samples by ASD diagnosis in females more than males across the perinatal tissues. We tested three theories for ASD sex differences in newborn blood, finding epigenetic support for an X chromosome-related female protective effect, as well as a high replication rate of DMRs (48.1%) in females across two independent cohorts. In our pan-tissue analysis, three genes (X-linked BCOR , GALNT9 , OPCML ) mapped to ASD DMRs replicated in all four female tissues. ASD DMRs from all tissues were enriched for neuro-related processes (females) and SFARI ASD-risk genes (females and males). Overall, we found a highly replicated methylation signature of ASD in females across perinatal tissues that reflected dysregulation in the brain and involvement of X chromosome epigenetics. This comparative study of perinatal tissues shows the promise of newborn blood DNA methylation biomarkers for early detection of females at risk for ASD and emphasizes the importance of sex-stratification in ASD studies.