Characterization and Contribution of Genome-wide DNA Methylation (DNA Methylation Episignatures) in Rare Diseases With Prenatal Onset (FOETEPISIGN)

It is necessary to define reference DNA Methylation Episignatures from fetal DNA. The hypotheses are:

• It is possible to define reference DNA Methylation Episignatures from fetal DNA extracted from amniotic fluid or frozen tissues collected during the postmortem examination
• Fetal DNA Methylation Episignatures may be different to postanal DNA Methylation Episignatures defined on DNA extracted from blood

Study Overview

• Status: Recruiting
• Conditions: Congenital Malformation; Rare Fetal Genetic Diseases
• Intervention / Treatment: Genetic: Methylation analysis

Detailed Description

Congenital anomalies (CA) complicate 3 to 5% of pregnancies and may be associated with genetic disorders. Diagnosis of genetic diseases is a major medical challenge, especially during pregnancy.

Over the past two decades, next-generation sequencing (NGS) has revolutionized our ability to identify the genetic condition associated with CA. During pregnancy, prenatal exome sequencing identified an additional diagnosis in around 30% of fetuses with CA when standard chromosomal investigations (karyotype and chromosomal microarray analysis, CMA) fail to provide a diagnosis.

Despite these major advances, around 40% of rare diseases remain unsolved, including 10-15% of patients harboring variants of uncertain significance (VUS).

After birth, additional functional analyses ("multi-OMICS"), including genome-wide DNA methylation studies, may be offered to reclassify VUS.

DNA methylation anomalies play an important role in pathologies (developmental disorders and oncology).

DNA methylation Episignatures, defined as the cumulative DNA methylation patterns occurring at multiple CpG dinucleotides across the genome, have been recognized to be intricately associated with many human traits, including age, sex, and disease status. Recently, DNA Methylation Episignatures have been identified in the blood of children or adults for several well-characterized genetic diseases. However, these postnatal DNA Methylation Episignatures cannot be used during pregnancy, because DNA methylation changes from one tissue to another and during time, especially during fetal developpement. In addition, the tissues available during pregnancy are different from those analyzed postnatally (blood).

• Study Type: Observational
• Enrollment (Estimated): 63

Contacts and Locations

• Study Contact: Name: Nelly BRIAND, PhD; Phone Number: 0144381862; Email: nelly.briand@aphp.fr
• Study Contact Backup: Name: Nicolas BOURGON, MD, PhD; Phone Number: +33 1 42 19 27 96; Email: nicolas.bourgon@aphp.fr

Study Locations

• France
  • Paris, France, 75015
    • Recruiting
    • Department of Genomic Medicine for Rare Diseases and the Multidisciplinary Center for Prenatal Diagnosis of the Necker-Enfants malades Hospital
    • Contact: Nicolas BOURGON, MD, PhD; Phone Number: : +33 1 42 19 27 96; Email: nicolas.bourgon@aphp.fr

Participation Criteria

Eligibility Criteria

• Ages Eligible for Study: Child; Adult
• Accepts Healthy Volunteers: Yes

• Sampling Method: Non-Probability Sample

Study Population

The study population is derived from the Department of Genomic Medicine for Rare Diseases and the Multidisciplinary Center for Prenatal Diagnosis of the Necker-Enfants malades Hospital - APHP - GHU Paris Cité.

The parents signed a consent to the storage and use of biological samples, stating that these samples taken as part of the treatment could be reused for medical research.

Description

Inclusion Criteria:

• Patient Inclusion Criteria:

  • Fetuses with a postmortem examination as part of the etiological diagnosis of developmental abnormality within the Genomic Medicine of Rare Diseases department of the Necker Children's Hospital, and whose DNA extracted from lung and amniotic fluid is available
  • OR a child cared for in the Genomic Medicine for Rare Diseases department of the Necker Children's Hospital, and whose DNA extracted from whole blood is available
  • with pathogenic or probably pathogenic variation in a gene following CHD7, KMT2D, HYLS1, TCTN3 or FLVCR2
  • whose parents have consented to molecular genetic testing as part of diagnosis and research

• Negative Controls :

  • Fetuses with a postmortem examination as part of the etiological diagnosis of developmental abnormality within the Genomic Medicine of Rare Diseases department of the Necker Children's Hospital, and whose DNA extracted from lung and amniotic fluid are available
  • OR a child cared for in the Genomic Medicine for Rare Diseases department of the Necker Children's Hospital, and whose DNA extracted from whole blood is available
  • does not have pathogenic or probably pathogenic variation in a gene following CHD7, KMT2D, HYLS1, TCTN3 or FLVCR2
  • whose parents have consented to molecular genetic testing as part of diagnosis and research

• For everyone:

  • For living participants: Non-objection by holders of parental authority to the reuse of clinical data and biological samples collected and stored in the context of care (consent of care).

  • For deceased participants:

• Consent of the holders of parental authority to the use of the samples kept for research purposes, signed as part of the treatment
• No mention of opposition to the reuse of clinical data from the treatment in the patient's medical record

Exclusion Criteria:

• Refusal of postmortem examination in case of fetal loss
• Parents' refusal of molecular investigations

Study Plan

How is the study designed?

Number of groups / cohorts

9

Cohorts and Interventions

Group / Cohort	Intervention / Treatment
Workpackage 1, group 1; Patients (children) with CHARGE syndrome	Genetic: Methylation analysis; Genomic DNA will be treated with bisulfite. 500 ng of processed DNA is then hybrized on an EPICv2 array Infinium methylation (Illumina, San Diego, CA, USA). This microarray enables the analysis of approximately 865 000 methylation sites at promoters, enhancers, CpG islands, intergenic and intragenic regions. It is the most widely used chip in the literature, including almost all of the EPIGENETIC SIGNATURES reported in human pathology.
Workpackage 1, group 2; Healthy negative controls (children) matched for age and sex	Genetic: Methylation analysis; Genomic DNA will be treated with bisulfite. 500 ng of processed DNA is then hybrized on an EPICv2 array Infinium methylation (Illumina, San Diego, CA, USA). This microarray enables the analysis of approximately 865 000 methylation sites at promoters, enhancers, CpG islands, intergenic and intragenic regions. It is the most widely used chip in the literature, including almost all of the EPIGENETIC SIGNATURES reported in human pathology.
Workpackage 1, group 3; Fetuses with CHARGE syndrome	Genetic: Methylation analysis; Genomic DNA will be treated with bisulfite. 500 ng of processed DNA is then hybrized on an EPICv2 array Infinium methylation (Illumina, San Diego, CA, USA). This microarray enables the analysis of approximately 865 000 methylation sites at promoters, enhancers, CpG islands, intergenic and intragenic regions. It is the most widely used chip in the literature, including almost all of the EPIGENETIC SIGNATURES reported in human pathology.
Workpackage 1, group 4; Fetuses with no genetic pathology	Genetic: Methylation analysis; Genomic DNA will be treated with bisulfite. 500 ng of processed DNA is then hybrized on an EPICv2 array Infinium methylation (Illumina, San Diego, CA, USA). This microarray enables the analysis of approximately 865 000 methylation sites at promoters, enhancers, CpG islands, intergenic and intragenic regions. It is the most widely used chip in the literature, including almost all of the EPIGENETIC SIGNATURES reported in human pathology.
Workpackage 2, group 5; Patients (children) with KABUKI syndrome	Genetic: Methylation analysis; Genomic DNA will be treated with bisulfite. 500 ng of processed DNA is then hybrized on an EPICv2 array Infinium methylation (Illumina, San Diego, CA, USA). This microarray enables the analysis of approximately 865 000 methylation sites at promoters, enhancers, CpG islands, intergenic and intragenic regions. It is the most widely used chip in the literature, including almost all of the EPIGENETIC SIGNATURES reported in human pathology.
Workpackage 2, group 6; Fetuses with KABUKI syndrome	Genetic: Methylation analysis; Genomic DNA will be treated with bisulfite. 500 ng of processed DNA is then hybrized on an EPICv2 array Infinium methylation (Illumina, San Diego, CA, USA). This microarray enables the analysis of approximately 865 000 methylation sites at promoters, enhancers, CpG islands, intergenic and intragenic regions. It is the most widely used chip in the literature, including almost all of the EPIGENETIC SIGNATURES reported in human pathology.
Workpackage 3, group 7; Fetuses with hydrolethalus syndrome	Genetic: Methylation analysis; Genomic DNA will be treated with bisulfite. 500 ng of processed DNA is then hybrized on an EPICv2 array Infinium methylation (Illumina, San Diego, CA, USA). This microarray enables the analysis of approximately 865 000 methylation sites at promoters, enhancers, CpG islands, intergenic and intragenic regions. It is the most widely used chip in the literature, including almost all of the EPIGENETIC SIGNATURES reported in human pathology.
Workpackage 3, group 8; Fetuses with Meckel/OFD IV syndrome	Genetic: Methylation analysis; Genomic DNA will be treated with bisulfite. 500 ng of processed DNA is then hybrized on an EPICv2 array Infinium methylation (Illumina, San Diego, CA, USA). This microarray enables the analysis of approximately 865 000 methylation sites at promoters, enhancers, CpG islands, intergenic and intragenic regions. It is the most widely used chip in the literature, including almost all of the EPIGENETIC SIGNATURES reported in human pathology.
Workpackage 3, group 9; Fetuses with Fowler syndrome	Genetic: Methylation analysis; Genomic DNA will be treated with bisulfite. 500 ng of processed DNA is then hybrized on an EPICv2 array Infinium methylation (Illumina, San Diego, CA, USA). This microarray enables the analysis of approximately 865 000 methylation sites at promoters, enhancers, CpG islands, intergenic and intragenic regions. It is the most widely used chip in the literature, including almost all of the EPIGENETIC SIGNATURES reported in human pathology.

What is the study measuring?

Primary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Epigenetic signature associated with pathogenic variations in the CHD7 gene (CHARGE Syndrome)	Evidence of epigenetic signature from fetal tissue DNA in fetuses with pathogenic or probably pathogenic variation	12 months
Epigenetic signature associated with pathogenic variations in the KMT2D gene (KABUKI syndrome)	Evidence of epigenetic signature from fetal tissue DNA in fetuses with pathogenic or probably pathogenic variation	12 months

Secondary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Differential methylation between fetal and postnatal epigenetic signature	Evidence of differential methylation between fetal an postnatal epigenetic signature	12 months
Differential methylation between tissue and amniotic fluid epigenetic signatures	Evidence of differential methylation between tissue and amniotic fluid epigenetic signatures	12 months
Statistical prediction parameter for each epigenetic signature	Measurement of the statistical prediction parameter for each epigenetic signature	12 months
Identification of a new epigenetic signature in foetal pathologies	Identification of news epigenetic signatures of exclusively pathologies associated with the HYLS1, TCTN3 and FLVCR2 genes	12 months

Collaborators and Investigators

• Sponsor: Assistance Publique - Hôpitaux de Paris
• Collaborators: URC-CIC Paris Descartes Necker Cochin
• Investigators: Study Director: Manon TESSIER, MD, PhD, Assistance Publique - Hopitaux de Paris

Publications and helpful links

General Publications

• Butcher DT, Cytrynbaum C, Turinsky AL, Siu MT, Inbar-Feigenberg M, Mendoza-Londono R, Chitayat D, Walker S, Machado J, Caluseriu O, Dupuis L, Grafodatskaya D, Reardon W, Gilbert-Dussardier B, Verloes A, Bilan F, Milunsky JM, Basran R, Papsin B, Stockley TL, Scherer SW, Choufani S, Brudno M, Weksberg R. CHARGE and Kabuki Syndromes: Gene-Specific DNA Methylation Signatures Identify Epigenetic Mechanisms Linking These Clinically Overlapping Conditions. Am J Hum Genet. 2017 May 4;100(5):773-788. doi: 10.1016/j.ajhg.2017.04.004.
• Aref-Eshghi E, Kerkhof J, Pedro VP; Groupe DI France; Barat-Houari M, Ruiz-Pallares N, Andrau JC, Lacombe D, Van-Gils J, Fergelot P, Dubourg C, Cormier-Daire V, Rondeau S, Lecoquierre F, Saugier-Veber P, Nicolas G, Lesca G, Chatron N, Sanlaville D, Vitobello A, Faivre L, Thauvin-Robinet C, Laumonnier F, Raynaud M, Alders M, Mannens M, Henneman P, Hennekam RC, Velasco G, Francastel C, Ulveling D, Ciolfi A, Pizzi S, Tartaglia M, Heide S, Heron D, Mignot C, Keren B, Whalen S, Afenjar A, Bienvenu T, Campeau PM, Rousseau J, Levy MA, Brick L, Kozenko M, Balci TB, Siu VM, Stuart A, Kadour M, Masters J, Takano K, Kleefstra T, de Leeuw N, Field M, Shaw M, Gecz J, Ainsworth PJ, Lin H, Rodenhiser DI, Friez MJ, Tedder M, Lee JA, DuPont BR, Stevenson RE, Skinner SA, Schwartz CE, Genevieve D, Sadikovic B. Evaluation of DNA Methylation Episignatures for Diagnosis and Phenotype Correlations in 42 Mendelian Neurodevelopmental Disorders. Am J Hum Genet. 2020 Mar 5;106(3):356-370. doi: 10.1016/j.ajhg.2020.01.019. Epub 2020 Feb 27.

Study record dates

Study Major Dates

• Study Start (Actual): February 26, 2026
• Primary Completion (Estimated): August 26, 2026
• Study Completion (Estimated): August 26, 2026

Study Registration Dates

• First Submitted: June 20, 2024
• First Submitted That Met QC Criteria: June 20, 2024
• First Posted (Actual): June 26, 2024

Study Record Updates

• Last Update Posted (Actual): March 27, 2026
• Last Update Submitted That Met QC Criteria: March 24, 2026
• Last Verified: March 1, 2026

More Information

Terms related to this study

• Keywords: CHARGE syndrome; Congenital malformation; DNA methylation abnormalities; Episignature; CHD7 gene; KMT2D gene; HYLS1 gene; TCTN3 gene; FLVCR2 gene; KABUKI syndrome
• Additional Relevant MeSH Terms: Neurologic Manifestations; Nervous System Diseases; Genetic Diseases, Inborn; Eye Diseases; Eye Diseases, Hereditary; Otorhinolaryngologic Diseases; Vision Disorders; Sensation Disorders; Abnormalities, Multiple; Ear Diseases; Deaf-Blind Disorders; Deafness; Hearing Loss; Hearing Disorders; Blindness; Coloboma; Congenital, Hereditary, and Neonatal Diseases and Abnormalities; Pathological Conditions, Signs and Symptoms; Signs and Symptoms; Congenital Abnormalities; CHARGE Syndrome; Hydrolethalus Syndrome 1; Kabuki syndrome
• Other Study ID Numbers: APHP231311

Plan for Individual participant data (IPD)

• Plan to Share Individual Participant Data (IPD)?: NO

Drug and device information, study documents

• Studies a U.S. FDA-regulated drug product: No
• Studies a U.S. FDA-regulated device product: No