4-Phenylbutyrate Rescue in GABRA1 Variants Associated with Developmental Epileptic Encephalopathies: from Cell and Mouse Models to Human

Disease variants in GABR genes encoding γ-Aminobutyric acid type A receptor (GABA A R) subunits are major causes of developmental and epileptic encephalopathies (DEEs). There is no effective treatment for these DEEs although the GABA A R is a major target for antiseizure drugs. We previously identified the therapeutic effect of 4-phenyl-butyrate (PBA) in Gabrg2 +/Q390X knockin DEE mice and now test the effect of the drug in GABRA1 variants that encode the α1 subunit. We used a multidisciplinary approach including in silico structural modeling, flow cytometry, patch clamp recordings and bio-chemistry in conjunction with differential tagging of the wild-type and the mutant alleles to evaluate the effect of PBA on rescue of GABA A R subunit expression, surface trafficking, and function in vitro in heterologous HEK293T cell model and in vivo in Gabra1 +/A322D mice. We found that both total and cell surface α1 expression was reduced when the variant α1 protein was present; suggesting reduced functional receptor on the cell membrane and synapse. Patch clamp recordings identified α1 variants reduced GABA-evoked current amplitude. In silico prediction indicated reduced protein stability for GABRA1 variants indicated by negative ΔΔG values. PBA increased both total and surface expression of wildtype α1 and α1 variants; and improved expression of both wildtype and variant α1 alleles when these were co-expressed. Importantly, PBA also increased the GABA A R expression in the thalamus of the Gabra1 +/A322D mice. This study indicates that PBA is a promising treatment option for DEEs associated with GABRA1 mutations. Our previous work has demonstrated that PBA improves proteostasis by enhancing expression of the wildtype allele, repairing the mutant allele, and reducing endoplasmic reticulum stress. Therefore, it can mitigate seizures and improve neurobehavioral phenotypes at behavioral levels. Based on this and our previous work on GABRG2 and SLC6A1 mutations, we propose that PBA holds promise as a common medicine for multiple genetic neurologic disorders that share the proteostasis pathology with a broad clinical application in DEEs.