Identification and clinical characteristics of a novel missense ADGRG1 variant in bilateral Frontoparietal Polymicrogyria: The electroclinical change from infancy to adulthood after Callosotomy in three siblings

Abstract

Objective: Bilateral frontoparietal polymicrogyria (BFPP) is a rare genetic-related migration disorder. It has been attributed to loss-of-function of the ADGRG1 gene, which encodes an adhesion G protein-coupled receptor, ADGRG1/GPR56. We report the EEG findings of BFPP in three Asian patients, and confirmed that change in protein function was caused by the novel missense variant (p.Leu290Pro).

Methods: We reviewed the medical records of three siblings with BFPP including one elder girl and two identical twin boys from birth to adulthood. The clinical symptoms, electroencephalography (EEG), brain MRI, whole-exome sequencing, treatment including medications, neuromodulation, and epilepsy surgery, and clinical outcomes were reviewed. The protein structure of a novel missense variant (p.Leu290Pro) was predicted by in silico studies, and molecular analysis was performed via typical flow cytometry and Western blotting.

Results: The elder girl (Patient 1) was 22 years old and the twin boys (Patients 2 and 3) were 20 years old at the time of publication. All of them presented with typical clinical symptoms/signs and MRI findings of BFPP. Whole-exome sequencing followed by Sanger confirmation showed that all three patients had compound heterozygous variants in the ADGRG1 gene. The missense variant (p.Leu290Pro) was confirmed to be related to a reduction in cell surface GPR56 expression. High-amplitude rhythmic activity was noted in sleep EEG during infancy, which may have been due to excessive sleep spindle, and the rhythm disappeared when they were of pre-school age. Partial callosotomy provided short-term benefits in seizure control in Patients 1 and 2, and combined vagus nerve stimulation and partial callosotomy provided longer benefits in Patient 3.

Significance: Sleep EEG findings of high-amplitude rhythmic activity in our BFPP cases were only noted during infancy and childhood. We also confirmed that the missense variant (p.Leu290Pro) led to loss of function due to a reduction in cell surface GPR56 expression.

Keywords: congenital CNS malformation; epilepsy; genetic; polymicrogyria.

FIGURE 1

Identification of GPR56/ADGRG1 gene frameshift/missense mutations in a Taiwanese family. (A) the family pedigree of the three patients. Squares and circles represent male and female individuals, respectively. White and black represent normal and diseased individuals, respectively. & indicates the GPR56 wild‐type sequence, * indicates the L290P missense mutation, while # indicates the frameshift mutation. (B) Results of sanger DNA sequencing of the GPR56/ADGRG1 frameshift (left side) and missense mutations (right side) in diseased patients and their parents. (C) Schematic representation of the GPR56 protein with known BFPP‐associated missense point mutations (black arrow) and newly identified L290P mutation (red arrow)

FIGURE 2

Brain magnetic resonance imaging of bilateral frontoparietal polymicrogyria in adults A‐D refer to Patient 1; E‐H refer to Patient 2; I‐L refer to Patient 3. Absence of gyrus, sulcus, and fused, thick cortex distributed over bilateral frontoparietal areas, and ventriculomegaly in images a, E, and I. multiple T2 hyperintense nodules at subcortical white matter shown by the white arrows in images B,F,J. flattened pons and small cerebellar vermis in images C,G,K, cavum septum pellucidum and cavum verge were noted only in Patient 3 in image I. left cerebellar‐medullary cistern schwannoma, and right tumefactive perivascular space (1 x 1.4 cm) found at right inferior basal ganglion in Patient 2 in image H. linear enlarged perivascular space over bilateral white matter in image D and over right internal capsular in image L

FIGURE 3

Sleep EEG of Patient 1 at 1, 5, and 9 y of age (A‐C) with a montage of double banana, speed 30 mm, sensitivity 7(B,C), 10 (a) uV, high cut 15 Hz, low cut 3 Hz. Bilateral extremely high‐amplitude (300 ~ 400 uV) alpha activity was noted over the frontal, temporal, and parietal lobes (A,B). The duration evolved from 1 to 13 s to continuous at 1 and 5 y of age, respectively (A,B). At 9 y of age, the frequency and amplitude of the alpha activity became lower (C). This activity could have indicated extreme sleep spindles

FIGURE 4

Sleep EEG in Patient 2 at 10 y of age (A), and awake EEG in Patient 2 at 18 y of age before and after callosotomy (B,C) with a montage of double banana, speed 30 mm, sensitivity 10 (A,C), 7 (B) uV, high cut 40 (A), 50 (B,C) Hz, low cut 3 Hz. Figure a indicates 1 Hz generalized spike and wave during sleep, Figure B shows diffuse theta activity at 18 y of age with intermittent generalized spike and wave. Figure C shows that the abnormal theta activity almost completely disappeared after callosotomy, which might indicate that the theta activities were related to interictal epileptic discharges rather than cortical dysfunction

FIGURE 5

(A) Alignment and comparison of the amino acid sequences of ADGRG1/GPR56‐ECR from the following animal species: Human (Homo sapiens), Chimpanzee (Pan troglodytes), Rhesus macaque (Macaca mulatta), Western lowland gorilla (Gorilla gorilla gorilla), Bornean orangutan (Pongo pygmaeus), Rat (Rattus norvegicus), and Mouse (Mus musculus). The known BFPP‐associated mutant residues (R38, Y88, C91, C346, and W349) are highlighted in yellow. The newly identified L290 residue is highlighted in green. (B) Table summarizing the predicted ΔΔG by DynaMut, DynaMut2, and mCSM‐membrane; and ΔΔSvib for the GPR56‐C91S, ‐L290P, and ‐C346S mutants using the AlphaFold2‐, RoseTTAFold‐, or TrRosetta‐predicted structures compared to the WT structure. Negative ΔΔG denotes protein destabilization, while a positive value denotes protein stabilization. Positive ΔΔSvib denotes an increase in molecular flexibility, while a negative value denotes a decrease in molecular flexibility. (C) Flow cytometry analysis of cell surface (without saponin) and total cellular expression (with saponin) of the GPR56 receptors expressed in HEK293T transfectant cells as indicated using anti‐GPR56 CG2 ab. The gray area represents the mocktransfected control. The numbers shown indicate the percentage of receptor‐positive transfectant cells. (D) Western blot analysis of total cell lysates from HEK293T transfectant cells expressing GPR56‐WT and relevant BFPP‐associated mutants as indicated. Blots were probed with anti‐GPR56 CG4 (upper panel) and anti‐myc (lower panel) abs to detect the ECR and 7TM subunits, respectively. The monomeric, dimeric, and trimeric forms of the 7TM subunit are indicated by 1, 2, and 3 asterisks, respectively. The arrow denotes the uncleaved single‐chain L290P mutant