Early-infantile onset epilepsy and developmental delay caused by bi-allelic GAD1 variants

Abstract

Gamma-aminobutyric acid (GABA) and glutamate are the most abundant amino acid neurotransmitters in the brain. GABA, an inhibitory neurotransmitter, is synthesized by glutamic acid decarboxylase (GAD). Its predominant isoform GAD67, contributes up to ∼90% of base-level GABA in the CNS, and is encoded by the GAD1 gene. Disruption of GAD1 results in an imbalance of inhibitory and excitatory neurotransmitters, and as Gad1-/- mice die neonatally of severe cleft palate, it has not been possible to determine any potential neurological dysfunction. Furthermore, little is known about the consequence of GAD1 disruption in humans. Here we present six affected individuals from six unrelated families, carrying bi-allelic GAD1 variants, presenting with developmental and epileptic encephalopathy, characterized by early-infantile onset epilepsy and hypotonia with additional variable non-CNS manifestations such as skeletal abnormalities, dysmorphic features and cleft palate. Our findings highlight an important role for GAD1 in seizure induction, neuronal and extraneuronal development, and introduce GAD1 as a new gene associated with developmental and epileptic encephalopathy.

Keywords: GAD1; cleft palate; epilepsy; muscle weakness; neurodevelopmental delay.

Figure 1

Pedigrees of the reported families and clinical pictures of GAD1 patients. (A) The female patient from Family A (Patient II-2) carries the homozygous c.1691A>G p.(Asn564Ser) variant and shows dysmorphic features with thick eyebrows, protruding ears, scoliosis, and long fingers with clinodactyly. (B) Patient from Family B (Patient III-4) harbours the c.971T>G p.(Phe324Cys) variant. He has slight dysmorphic features (wide mouth, thin upper lips, bitemporal narrowing and retrognathia). (C) Pedigree showing the segregation of the compound heterozygous variants c.1591C>T, c.1591C>T p.(Arg531*) and c.670delC p.(Leu224Serfs*5) in Family C. (D) Pedigree of Family D shows the segregation of the c.1040C>T p.(Thr347Met) variant. Patient II-4 carries the variant in homozygous state. He is severely hypotonic and shows severe dysmorphic features (infra-orbital creases, severely depressed nasal bridge, anteverted nares, prominent nasolabial folds). In addition, significant diastasis recti can be observed. His sister (Patient II-2) is heterozygous for the same variant and suffers from a different neurodevelopmental condition without seizures. (E) Patient III-2 from Family E harbours the c.87C>G (Tyr29*) variant, severely affected with dysmorphic facial features and global atrophy on cardiovascular MRI, one similarly affected sibling passed away without any genetic testing being performed, another sibling passed away only a few hours after birth, no phenotypical or genetic assessment could be carried out, and one sibling is alive with a different phenotype (sensoneural hearing loss, Hirschsprung disease). (F) Patient II-1 from Family F harbours the c.568del (Gln190Serfs*11) variant. His parents are both heterozygous carriers of the same variant. Empty and full symbols represent healthy and affected individuals, respectively. The symbol with diagonal lines indicates carrier status/different phenotype. The double line indicates consanguinity.

Figure 2

Schematic and cartoon representation of GAD1. (A) Schematic representation of the GAD1 isoform GAD67 (NP_000808.2) with the pathogenic variants identified in this study. Of the six variants, four fall within the PLP-binding domain, a conserved region that is essential for the binding of the crucial cofactor pyridoxal 5′-phosphate (PLP). The remaining variants affect the C-terminal domain, which contains the catalytic site of the enzyme. Conservation status among different species is shown for the missense variants. (B) Cartoon representation of human GAD1 dimer (PDB: 2okj) with the two subunits in blue and yellow. Sites of the three missense mutations in this study are shown as red spheres, and close-up views of their nearby atomic environment are shown as insets. The PLP cofactor is shown in green.