Recent discoveries have shown that loss-of-function variants of SCN2A are causal for a number of neurodevelopmental disorders, including autism spectrum disorder (ASD). Given this genetic linkage to disease, our team at the Broad Institute established a drug discovery program to develop small molecule activators of NaV1.2 as a therapeutic strategy for ASD. In this presentation, we will describe the design and establishment of a screening cascade for the identification of modulators of NaV1.2.
We first established a novel primary high-throughput screening (HTS) assay combining electric field stimulation with the detection of membrane potential changes in an NaV1.2 over-expressing cell line. Using this assay, we completed an HTS of nearly 80,000 compounds and identified 378 enhancers of NaV1.2 function. Comparison with other assay formats showed that this novel assay is substantially more sensitive in its ability to identify activators of NaV1.2 channel function relative to previous assay formats where the sodium channels have been forced to open via chemical means. Further, a number of the compounds were potent and efficacious activators with selectivity over the closely related NaV1.1 channel. Subsequent electrophysiological characterization showed that one hit, BRD4032, induces a shift in the voltage-dependence of NaV1.2 channel opening, causing it to open at lower membrane potentials. Further electrophysiology analysis of BRD4032 in mouse brain slices showed it elicits marked widening of the action potential waveform. Finally, in vivo EEG studies revealed that Scn2a+/- mice show significantly altered frequency bands, indicating that these alterations may be useful translational biomarkers.
Taken together, we have successfully implemented a robust screening strategy to identify NaV1.2 modulators based on a novel functional assay and are poised to advance compounds into our drug discovery pipeline that may eventually serve as leads towards treating ASD.
Thursday, November 19 @ 8 AM PST / 11 AM EST / 5 PM CET