DNA methylation episignature and comparative epigenomic profiling for Pitt-Hopkins syndrome caused by TCF4 variants

Liselot van der Laan, Peter Lauffer, Kathleen Rooney, Ananília Silva, Sadegheh Haghshenas, Raissa Relator, Michael A Levy, Slavica Trajkova, Sylvia A Huisman, Emilia K Bijlsma, Tjitske Kleefstra, Bregje W van Bon, Özlem Baysal, Christiane Zweier, María Palomares-Bralo, Jan Fischer, Katalin Szakszon, Laurence Faivre, Amélie Piton, Simone MesmanRon Hochstenbach, Mariet W Elting, Johanna M van Hagen, Astrid S Plomp, Marcel M A M Mannens, Mariëlle Alders, Mieke M van Haelst, Giovanni B Ferrero, Alfredo Brusco, Peter Henneman, David A Sweetser, Bekim Sadikovic, Antonio Vitobello, Leonie A Menke

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Pitt-Hopkins syndrome (PTHS) is a neurodevelopmental disorder caused by pathogenic variants in TCF4, leading to intellectual disability, specific morphological features, and autonomic nervous system dysfunction. Epigenetic dysregulation has been implicated in PTHS, prompting the investigation of a DNA methylation (DNAm) “episignature” specific to PTHS for diagnostic purposes and variant reclassification and functional insights into the molecular pathophysiology of this disorder. A cohort of 67 individuals with genetically confirmed PTHS and three individuals with intellectual disability and a variant of uncertain significance (VUS) in TCF4 were studied. The DNAm episignature was developed with an Infinium Methylation EPIC BeadChip array analysis using peripheral blood cells. Support vector machine (SVM) modeling and clustering methods were employed to generate a DNAm classifier for PTHS. Validation was extended to an additional cohort of 11 individuals with PTHS. The episignature was assessed in relation to other neurodevelopmental disorders and its specificity was examined. A specific DNAm episignature for PTHS was established. The classifier exhibited high sensitivity for TCF4 haploinsufficiency and missense variants in the basic-helix-loop-helix domain. Notably, seven individuals with TCF4 variants exhibited negative episignatures, suggesting complexities related to mosaicism, genetic factors, and environmental influences. The episignature displayed degrees of overlap with other related disorders and biological pathways. This study defines a DNAm episignature for TCF4-related PTHS, enabling improved diagnostic accuracy and VUS reclassification. The finding that some cases scored negatively underscores the potential for multiple or nested episignatures and emphasizes the need for continued investigation to enhance specificity and coverage across PTHS-related variants.

Original languageEnglish
Article number100289
Pages (from-to)100289
JournalHuman Genetics and Genomics Advances
Issue number3
Early online date2 Apr 2024
Publication statusE-pub ahead of print - 2 Apr 2024


  • CNV
  • DNA methylation
  • PTHS
  • Pitt-Hopkins syndrome
  • TCF4
  • VUS
  • episignature
  • neurodevelopmental disorder

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