ARID2-related disorder: further delineation of the clinical phenotype of 27 novel individuals and description of an epigenetic signature

Abstract

Rare genetic variants in ARID2 are responsible for a recently described neurodevelopmental condition called ARID2-related disorder (ARID2-RD). ARID2 belongs to PBAF, a unit of the SWI/SNF complex, which is a chromatin remodeling complex. This work aims to further delineate the phenotypic spectrum of ARID2-RD, providing clinicians with additional data for better care and aid in the future diagnosis of this condition. We obtained the genotypes and phenotypes of 27 previously unreported individuals with ARID2-RD and compared this series with findings in the literature. We also assessed peripheral blood DNA methylation profiles in individuals with ARID2-RD compared to episignatures of controls, unresolved cases, and other neurodevelopmental disorders. The main clinical features of ARID2-RD are developmental delay, speech disorders, intellectual disability (ID), behavior problems, short stature, and various dysmorphic and ectodermal features. Genome-wide differential methylation analysis revealed a global hypermethylated profile in ARID2-RD that could aid in reclassifying variants of uncertain significance. Our study doubles the number of reported individuals with ARID2 pathogenic variants to 53. It confirms loss-of-function as a pathomechanism and shows the absence of a clear genotype-phenotype correlation. We provide evidence for a unique DNA methylation episignature for ARID2-RD and further delineate the ARID2-associated phenotype.

Fig. 1. ARID2 pathogenic variants and associated clinical features.

A Position of frameshift (in red), nonsense (in orange), splice (in purple), and in-frame deletion (in blue) variants of the cohort and the literature along the ARID2 sequence. B (Micro)deletions identified in the cohort and the literature are represented by the red bars, all affecting ARID2. The balanced rearrangement of chromosome 12 in which one of the chromosomal breakpoints was predicted to split ARID2 is represented in salmon. Genomic positions are according to hg38; ARID2 is on the forward strand. C Front and side profile picture of individuals with a PTV or deletion affecting ARID2. D A table demonstrating the amino acid positions and the clinical findings of ARID2 variants detected in 27 cohort individuals. When the boxes are colored in gray (for SNV) or red (for CNV/SV), the clinical sign is present; when white, the clinical sign is absent; and when not available, N/A.

Fig. 2. Assessment of the Coffin-Siris syndrome-6 (ARID2) episignature.

A, B Euclidean hierarchical clustering heatmap, each column represents one ARID2 case or control, and each row represents one probe selected for this episignature. This heatmap and multidimensional scaling (MDS) plot shows a clear separation between ARID2 cases (red and purple) used for training and validation from controls (blue). The mosaic case (orange) is shown to map with control cases, one episignature-negative case (pink) is also shown to map with controls, three unresolved cases (brown) are mapping to cases, and four VUS cases (yellow) are mapping with controls. C Support Vector Machine (SVM) classifier model. The x-axis represents an episignature on the EpiSign™ classifier, and the y-axis a probability score referred to as a Methylation Variant Pathogenicity score (MVP). This model was trained using the selected ARID2 episignature probes, 75% of controls, and 75% of other neurodevelopmental disorder samples (blue). The remaining 25% of controls and 25% of other disorder samples were used for testing (grey). Plot shows the ARID2 mosaic, negative and VUS cases with a methylation variant pathogenicity (MVP) score close to 0, and the unresolved cases with an MVP score close to 1, similar to the ARID2 training cases, showing the specificity of the classifier and episignature. ADCADN Cerebellar ataxia deafness and narcolepsy syndrome, AUTS18 Susceptibility to autism 18, BEFAHRS Beck-Fahrner syndrome, BFLS Borjeson–Forssman– Lehmann syndrome, BISS Blepharophimosis intellectual disability SMARCA2 syndrome, CdLS Cornelia de Lange syndrome, CHARGE CHARGE syndrome, Chr16p11.2del Chromosome 16p11.2 deletion syndrome, CSS Coffin–Siris syndrome, CSS4 Coffin-Siris syndrome 4, CSS9 Coffin–Siris syndrome 9, Down Down syndrome, Dup7 7q11.23 duplication syndrome, DYT28 Dystonia 28, EEOC Epileptic encephalopathy-childhood onset, FLHS Floating Harbour syndrome, GTPTS Genitopatellar syndrome, HMA Hunter McAlpine craniosynostosis syndrome, HVDAS Helsmoortel–van der Aa syndrome, ICF Immunodeficiency-centromeric instability-facial anomalies syndrome, IDDSELD Intellectual developmental disorder with seizures and language delay, Kabuki Kabuki syndrome, KDVS Koolen-De Vries syndrome, Kleefstra Kleefstra syndrome, LLS Luscan-Lumish syndrome, MKHK Menke Hennekam syndrome, MLASA2 Myopathy lactic acidosis and sideroblastic anemia 2, MRD23 Intellectual developmental disorder 23, MRD51 Intellectual developmental disorder 51, MRX93 Intellectual developmental disorder X-linked 93, MRX97 Intellectual developmental disorder X-linked 97, MRXSA Intellectual developmental disorder X-linked syndromic Armfield type, MRXSCH Intellectual developmental disorder X-linked syndromic Christianson type, MRXSCJ Intellectual developmental disorder X-linked syndromic Claes-Jensen type, MRXSN Intellectual developmental disorder X-linked syndromic Nascimento type, MRXSSR Intellectual developmental disorder X-linked syndromic Snyder–Robinson type, PHMDS Phelan–McDermid syndrome, PRC2 PRC2 complex (Weaver and Cohen-Gibson) syndrome, RENS1 Renpenning syndrome, RMNS Rahman syndrome, RSTS Rubinstein–Taybi syndrome, SBBYSS Ohdo syndrome, Sotos Sotos syndrome, TBRS Tatton–Brown– Rahman syndrome, WDSTS Wiedemann–Steiner syndrome, WHS Wolf-Hirschhorn syndrome, Williams Williams syndrome.

Fig. 3. Differentially methylated probes (DMPs) annotated in the context of CpG islands and genes.

A DMRs in relation to genes. B DMPs in relation to genes. C DMRs in CpG islands and (D) DMPs in CpG island. Promoter, 0–1 kb upstream of the transcription start site (TSS); Promoter + , 1–5 kb upstream of the TSS; CDS, coding sequence; Intergenic, all other regions of the genome. Island, CpG islands; Shore, within 0–2 kb of a CpG island boundary; shelf, within 2-4 kb of a CpG island boundary; Inter_CGI, all other regions in the genome. The Probes column in both (b,d) represents the background distribution determined in the Levy et al. [9] study of all array probes after initial filtering and used as input for DMP analysis.

Fig. 4. Relationships between the ARID2 cohort and 56 other EpiSign™ disorders.

A Global methylation profiles of all differentially methylated probes (DMPs, FDR < 0.05) for each cohort, sorted by mean methylation. Each circle represents one probe, and red lines show the mean methylation. B Heatmap showing the percentage of probes shared between each paired cohort. Colors indicate the percentage of the y-axis cohort’s probes that are also found in the x-axis cohort’s probes.

Fig. 5. Tree and leaf visualization of Euclidean clustering of all 57 cohorts using the top n DMPs for each group, where n = min (# of DMPs, 500).

Cohort samples were aggregated using the median value of each probe within a group. A leaf node represents a cohort, with node sizes illustrating relative scales of the number of selected DMPs for the corresponding cohort, and node colors are indicative of the global mean methylation difference.