Human induced pluripotent stem cell ‐derived cortical neurons from monozygotic twins with the Gα o1 K278del mutation reveal disease mechanisms relevant to GNAO1 ‐related encephalopathy

Abstract Objective Rare de novo variants in GNAO1 cause a severe neurodevelopmental disorder characterized by a heterogenous display of early onset epilepsy, movement abnormalities, and global developmental delay. More than 100 pathogenic variants in GNAO1 have been identified and categorized into four groups according to their protein features. Most pathogenic variants affect both GNAO1 isoforms, whereas approximately 20% of patients have a GNAO1A ‐specific variant. Response to therapy is notoriously variable, even among patients harboring the same variant. Here, we set out to model GNAO1A ‐related encephalopathy using human induced pluripotent stem cells (hiPSCs) from monozygotic twins carrying the Gα o1 K278del variant and uncovered—while taking advantage of the identical genomic setting—phenotypic features that further our understanding of the defects underlying the disorder. Methods We generated hiPSCs expressing the c.832_834delAAG; p.K278del variant in GNAO1A obtained from monozygotic twins who both developed seizures 2 days after birth. For either twin, Gα o1 K278del and CRISPR‐Cas9‐repaired isogenic control hiPSC lines were differentiated into neural progenitor cells (NPCs) and subsequently into cortical neurons. Variant‐specific transcriptomic, morphological, and electrophysiological characteristics were analyzed. Results Gα o1 K278del mutated hiPSC‐derived NPCs showed disorganized neural rosette structures and revealed an altered transcriptome. In cortical neurons, the Gα o1 K278del variant caused a reduction in the size and complexity of dendrites that was accompanied by an increase in pre‐ and postsynaptic puncta. Multielectrode array (MEA) recordings revealed an abnormal cortical neuron bursting activity pattern with fewer bursts and more spikes per burst. The antiepileptic drug levetiracetam reduced the spike frequency and the number of spikes per burst in cortical neurons from both patients. Significance The developed GNAO1A hiPSC‐derived in vitro model uncovered key structural and functional features of GNAO1 ‐related encephalopathy. The comparable in vitro phenotypes in neurons from both twins underscore the utility of hiPSC‐derived models to investigate molecular mechanisms underlying GNAO1 ‐related encephalopathy as well as effects of therapeutic intervention.