Clinical Utility of a Unique Genome-Wide DNA Methylation Signature for KMT2A-Related Syndrome

Abstract

Wiedemann-Steiner syndrome (WDSTS) is a Mendelian syndromic intellectual disability (ID) condition associated with hypertrichosis cubiti, short stature, and characteristic facies caused by pathogenic variants in the KMT2A gene. Clinical features can be inconclusive in mild and unusual WDSTS presentations with variable ID (mild to severe), facies (typical or not) and other associated malformations (bone, cerebral, renal, cardiac and ophthalmological anomalies). Interpretation and classification of rare KMT2A variants can be challenging. A genome-wide DNA methylation episignature for KMT2A-related syndrome could allow functional classification of variants and provide insights into the pathophysiology of WDSTS. Therefore, we assessed genome-wide DNA methylation profiles in a cohort of 60 patients with clinical diagnosis for WDSTS or Kabuki and identified a unique highly sensitive and specific DNA methylation episignature as a molecular biomarker of WDSTS. WDSTS episignature enabled classification of variants of uncertain significance in the KMT2A gene as well as confirmation of diagnosis in patients with clinical presentation of WDSTS without known genetic variants. The changes in the methylation profile resulting from KMT2A mutations involve global reduction in methylation in various genes, including homeobox gene promoters. These findings provide novel insights into the molecular etiology of WDSTS and explain the broad phenotypic spectrum of the disease.

Keywords: DNA methylation; KMT2A gene; Wiedemann–Steiner syndrome; epigenetics; episignature; intellectual disability; neurodevelopmental disorders.

Figure 1

Volcano plot of methylation difference between 41 WDSTS samples and controls versus statistical significance (-log p-value) of individual probes. Red dots represent selected, significant differentially methylated probes (DMPs). Positive and negative mean methylation difference show hypermethylation and hypomethylation, respectively.

Figure 2

Identification of a WDSTS episignature. (a) Hierarchical clustering with Ward’s method on Euclidean distance was performed. In the heatmap plot, each row illustrates a selected CpG site, and each column is related to a sample. The heatmap color scale indicates the range of methylation level; from blue (no methylation or 0) to red (full methylation or 1). This plot conveys that the detected episignature clearly differentiates between 41 WDSTS samples and controls; (b) multidimensional scaling (MDS) plot using the selected probes. MDS plot illustrates power of the signature in separating the 41 WDSTS samples and control samples. Blue circles represent control subjects and red circles indicate subjects with pathogenic variants in the KMT2A gene and a confirmed diagnosis of the syndrome.

Figure 3

The methylation variant pathogenicity (MVP) scores plot. (a) The MVP scores were created by the SVM trained by comparing the 41 WDSTS samples against controls; (b) the MVP scores created by the SVM trained by comparing 41 WDSTS samples against controls and 38 neurodevelopmental disorders and congenital anomalies available in the EKD. The blue circles represent the training samples and the grey circles represent the testing samples.

Figure 4

Adding WDSTS (testing) and Kabuki1 samples to the WDSTS signature. (a) Hierarchical clustering; (b) multidimensional scaling.