Characterization of the functional and clinical impacts of CACNA1A missense variants found in neurodevelopmental...

CACNA1A encodes the α1A subunit of the Ca_V2.1 voltage-gated calcium channel essential for neuronal excitability, neurotransmitter release, and calcium signaling in neurons. Pathogenic CACNA1A variants are linked to diverse neurological and developmental disorders, including hemiplegic migraine, epilepsy, developmental delay, and ataxia. However, the contribution of CACNA1A variants to neurodevelopmental disorders has been less well established. Clinical heterogeneity and the complex impact of missense variants on channel function have made genotype-phenotype relationships difficult to resolve. Here, we characterized the biophysical properties of 42 de novo CACNA1A missense variants identified in a neurodevelopmental disorder cohort of over 31,000 individuals, alongside eight common variants from the general population. All but one de novo variant altered biophysical properties of the Ca_V2.1 channel, with 50% abolishing measurable currents. Among variants with detectable currents, nearly half disrupted voltage-dependent gating, whereas common variants had no measurable effect. Similar results were obtained using representative variants in the presence of β3 or β4 auxiliary subunits. The expression of two selected variants in CACNA1A-knockdown hiPSC-derived neurons confirmed similar functional alterations in human neurons. Simulations using the NEURON model of a Purkinje neuron demonstrated that these biophysical changes profoundly affected excitability. Coupling functional data with AlphaMissense predictions, we found that Ca_V2.1 missense variants contribute to the risk for developmental epileptic encephalopathy. Finally, correlating clinical features with molecular consequences of each variant revealed meaningful associations between specific channel dysfunctions and distinct clinical outcomes. These findings provide insights into CACNA1A molecular pathology and highlight the potential for precision medicine treatment of CACNA1A-associated disorders.