Association of ultra-rare coding variants with genetic generalized epilepsy: A case-control whole exome sequencing study

Abstract

Objective: We aimed to identify genes associated with genetic generalized epilepsy (GGE) by combining large cohorts enriched with individuals with a positive family history. Secondarily, we set out to compare the association of genes independently with familial and sporadic GGE.

Methods: We performed a case-control whole exome sequencing study in unrelated individuals of European descent diagnosed with GGE (previously recruited and sequenced through multiple international collaborations) and ancestry-matched controls. The association of ultra-rare variants (URVs; in 18 834 protein-coding genes) with epilepsy was examined in 1928 individuals with GGE (vs. 8578 controls), then separately in 945 individuals with familial GGE (vs. 8626 controls), and finally in 1005 individuals with sporadic GGE (vs. 8621 controls). We additionally examined the association of URVs with familial and sporadic GGE in two gene sets important for inhibitory signaling (19 genes encoding γ-aminobutyric acid type A [GABA_A ] receptors, 113 genes representing the GABAergic pathway).

Results: GABRG2 was associated with GGE (p = 1.8 × 10^-5 ), approaching study-wide significance in familial GGE (p = 3.0 × 10^-6 ), whereas no gene approached a significant association with sporadic GGE. Deleterious URVs in the most intolerant subgenic regions in genes encoding GABA_A receptors were associated with familial GGE (odds ratio [OR] = 3.9, 95% confidence interval [CI] = 1.9-7.8, false discovery rate [FDR]-adjusted p = .0024), whereas their association with sporadic GGE had marginally lower odds (OR = 3.1, 95% CI = 1.3-6.7, FDR-adjusted p = .022). URVs in GABAergic pathway genes were associated with familial GGE (OR = 1.8, 95% CI = 1.3-2.5, FDR-adjusted p = .0024) but not with sporadic GGE (OR = 1.3, 95% CI = .9-1.9, FDR-adjusted p = .19).

Significance: URVs in GABRG2 are likely an important risk factor for familial GGE. The association of gene sets of GABAergic signaling with familial GGE is more prominent than with sporadic GGE.

Keywords: GABRG2; GABAA receptors; GGE; familial epilepsy; sporadic epilepsy.

Fig. 1:. Association of ultra-rare variation in protein coding genes with genetic generalized epilepsy.

The quantile-quantile plots compare the observed p values (Cochran-Mantel-Haenszel exact test) and the expected p values (drawn from a uniform distribution) in analyses of 1,928 individuals genetic generalized epilepsies (GGEs) in comparison to 8,578 matched controls (top panel) as well as subsequent analyses of familial GGEs (middle panel; 945 cases and 8,626 controls) or sporadic GGEs (bottom panel; 1,005 cases and 8,621 controls). These analyses focused on functional ultra-rare variants (URVs; with minor allele frequencies < 0.05% in the test dataset & not seen in DiscovEHR/gnomAD) that were annotated as predicted Loss-of-Function variants (pLoF), damaging missense variants with Polyphen2 (PPh2), or in-frame insertions and deletions. Study-wide significance after Bonferroni correction (dark red line) was defined by a p value < 2.9 x 10⁻⁷. λ: Genomic inflation factor. Among the five top-ranked genes, genes that had a higher carrier frequency in cases vs. controls in both study datasets are labeled (not labeled among top-ranked: enriched in controls or in cases in one dataset only).

Fig. 2:. Association of ultra-rare variation in genes encoding GABA_A receptors with familial and sporadic genetic generalized epilepsy.

The forest plots show the association of ultra-rare deleterious and intolerant variants with the phenotype in analyses of 1,928 individuals with GGE vs. 8,578 controls (A), 945 individuals with familial GGE vs. 8,626 controls (B), 1,005 individuals with sporadic GGE vs. 8,621 controls (C), and a direct comparison of 945 individuals with familial GGE vs. 1,005 individuals with sporadic GGE (D). Four (primary and control) ultra-rare variant models are shown (y axis). The association in each analysis is displayed as the natural logarithm of stratified odds ratio from a Cochran-Mantel-Haenszel exact test (x axis). Errors bars indicated the logarithm of the 95% confidence intervals (CI). The corresponding odds ratios and associated p values, and False Discovery Rate (FDR) adjusted p values are displayed on the side. The tests for synonymous variants were not adjusted for multiple testing.