Abstract

Recent discoveries have shown that SCN2A genetic variants are causal for a number of neurodevelopmental disorders, including autism spectrum disorder (ASD), intellectual disability (ID), and infantile seizure disorders. SCN2A encodes the voltage-gated sodium channel Na_V1.2, with loss-of-function variants causing ASD and ID, and gain-of-function variants causing infantile seizure disorders. Given the genetic linkage, we set out to discover small molecule positive modulators of Na_V1.2 for the treatment of ASD. We first established a novel primary high-throughput screening (HTS) assay to allow the detection of voltage changes in Na_V1.2‑expressing HEK cells in response to electric filed stimulation (EFS). This physiologically relevant assay platform enabled completion of an HTS campaign of approximately 80,000 compounds, where we identified 378 Na_V1.2 activators and 1837 inhibitors. Follow up concentration-response testing yielded a number of potent and efficacious activators, and select compounds were followed up in a synaptic function model where a synaptically transmitted response to EFS was quantified in primary mouse cortical cultures to confirm the compound effect in native neuronal cultures. Subsequent electrophysiological characterization of one hit, BRD4032, revealed that it activates Na_V1.2 by changing both the voltage sensing mechanism and the inactivation kinetics of the channel. Taken together, we have successfully implemented a robust screening strategy to identify Na_V1.2 modulators and are poised to follow up on the selectivity, mechanism of action, and in vivo effects of these compounds.