Generating Biologically Relevant Subtypes of Autism Spectrum Disorder with differential responses to Acute Oxytocin Administration in a Randomized Trial using Random Forest Models and K-means Clustering

Autism Spectrum Disorder (ASD) is a heterogenous condition that has no biologically relevant subtypes yet. Here, we utilized a multidimensional approach considering social deficits in ASD alongside negative valence and empathy dysfunction to distinguish ASD from Neurotypicals (NT) and to generate ASD subtypes using machine learning approaches. 114 subjects were analyzed, with 70 being NT and 44 ASD, all male with an IQ greater than 70, with 5 domains of personality (NEO-PI-r) and Reading the Mind the Eyes Test (RMET) scores included in the main classifier. We then used a multitude of behavioral (such as IQ, Broader Autism Phenotype, Autism Quotient, Interpersonal Reactivity Index) and clinical measures such as Autism Diagnostic Interview-Revised (ADI-R) alongside biological methods including DNA methylation of <i>OXTR</i> gene and resting-state functional connectivity (rsFC) to validate the putative subtypes. 30 ASD who received IN-OXT in a randomized, placebo-controlled, within-subject design and 17 new NT were part of the rs-FC analysis. A random forest tree algorithm was used to classify NT and ASD and Shapley Additive Explanation Values were used to describe the model and to cluster ASD subtypes using K-Means clustering. Three subtypes were generated with two of them being highly distinctive in behavioral and brain functional traits. One subtype named NASA (or Negative Affect and Social Aloofness) was characterized by high Neuroticism and Low warmth alongside lower rsFC between networks involved in social cognition, self-awareness, and sensory processing, such as Superior Temporal Sulcus and Sensorimotor Network; or ACC/Insula with visual cortex, Posterior Cingulate Cortex and visual cortex. The second subtype NADR (Neurocognitive and Affect Dysregulation with Resistance to Change) was characterized by higher DNA methylation of <i>OXTR</i>, hyperconnectivity between default mode network, reward areas and inferior frontal and fusiform networks. NADR has more cognitive difficulties and higher ADI-R scores as well as higher Neuroticism, higher personal distress, higher rigidity and lower openness. In a mixed model analysis, we found that IN-OXT in a dose dependent manner impacted NASA subtype by modulating rsFC between PCC and cerebellum and between Brainstem/Cerebellum and Parietal cortex to probably enhance social cognition and to reduce negative valence in this subtype.