EpiSign complete

EpiSign is a test based on DNA methylation patterns. Various types of methylation abnormalities can be detected: Episignatures, Imprinting disorders and Methylation of promoter region.

Episignatures

Pathogenic variants in different genes (usually these are genes involved in the regulation of epigenetic processes and chromatin structure) can give a very specific aberrant genome-wide methylation profile. The specific profile is called an episignature. Such an episignature has now been established for more than 60 conditions.

Imprinting disorders

The analysis of imprinting disorders distinguishes the methylation status of the differentially methylated regions in/around the imprinted genes. If abnormal methylation is detected, further investigation may be necessary to determine the underlying cause, such as: UPD or deletion. There are currently 8 imprinting disorders detectable with EpiSign.

Methylation of promoter region of the FMR1 gene (Fragile X syndrome, FX)

In boys with FX, the FMR1 gene is inactivated by extension of a CGG repeat in the promoter region of the gene. With EpiSign, the consequence of the CGG repeat extension, inactivation of the FMR1 gene due to hypermethylation of the FMR1 promoter, can be detected (EpiSign is therefore not a test for determining the repeat length). In case of a positive result, an additional repeat analysis will have to be done to confirm the diagnosis of FX EpiSign is only suitable for diagnosing FX in males.

Indications for requesting EpiSign test:

• When the patient has a variant of unknown clinical relevance (VUS) in one of the genes for which an Episignature is available. A methylation profile similar to that of the associated syndrome supports pathogenicity of the variant.
• When previous research has not resulted in a diagnosis. A positive episignature could provide a direction for further research, or targeted (re)analysis of existing data. EpiSign can also detect imprinting disorders and FX, which cannot be detected with WES.

Please note: if Silver Russell Syndrome (SRS) or Beckwith-Wiedemann Syndrome (BWS) is suspected, we advise to request (first) a targeted MLPA analysis for these syndromes. The methylation aberrations on chromosome 11 are often mosaic and can be very subtle. The sensitivity of the MLPA is greater than EpiSign. The methylation aberrations on chromosome 11 are often mosaic and can be very subtle. The sensitivity of the MLPA is greater than EpiSign.

Abnormalities detected using this initial screen may require additional targeted testing to confirm and further characterize the underlying genomic abnormality.

Imprinting and Trinucleotide Repeat Disorders:
Angelman syndrome 15q11.2-q13 (SNRPN promoter, SNURF)
Beckwith-Wiedemann syndrome 11p15 (ICR1, KCNQ1OT1, CDKN1C)
Diabetes mellitus, transient neonatal 1 6q24 (PLAG1)
Fragile X syndrome FMR1
Kagami-Ogatta syndrome 14q32 (MEG3 promoter)
Mulchandani-Bhoj-Conlin syndrome 20q11-q13 (GNAS)
Prader-Willi syndrome 15q11.2 (SNRPN promoter, SNURF)
Pseudohypoparathyroidism, Type IA, IB 20q13.32 (GNAS)
Silver Russel syndrome 1 11p15 (ICR1)
Silver Russel syndrome 2 7p13-q32
Temple syndrome 14q32 (MEG3 promoter)

Episignatures:
Alpha-thalassemia mental retardation syndrome (ATRX)
Arboleda-Tham syndrome (KAT6A)
Autism, susceptibility to, 18 (CHD8)
BAFopathies: Coffin-Siris 1-4 & Nicolaides-Baraitser syndromes ¹ (BAFopathy) (ARID1B, ARID1A, SMARCB1, SMARCA4, SMARCA2)
Beck-Fahrner syndrome ^2,3 (TET3)
Blepharophimosis Intellectual disability SMARCA2 syndrome (SMARCA2)
Börjeson-Forssman-Lehmann syndrome (PHF6)
Cerebellar ataxia, deafness, and narcolepsy, autosomal dominant (DNMT1)
CHARGE syndrome (CHD7)
Chr1p36 deletion syndrome, distal (Chr1p36 deletion)
BAFopathies: Coffin-Siris syndrome 1 and 2 ⁴ (ARID1B, ARID1A)
Coffin-Siris syndrome-1 ⁵ (ARID1B)
Coffin-Siris syndrome-2 ⁵ (ARID1A)
Coffin-Siris syndrome-3 ⁵ (SMARCB1)
Coffin-Siris syndrome-4 ⁵ (SMARCA4)
Coffin-Siris syndrome-4 ⁶ (CSS4_c.2656) (SMARCA4)
Coffin-Siris syndrome-9 (SOX11)
Congenital Heart Defects, Dysmorphic Facial Features, and Intellectual Developmental Disorder (CDK13, CCNK)
Cornelia de Lange syndromes 1-4 ⁷ (NIPBL, SMC1A, SMC3, RAD21)
Down syndrome (Chr21 trisomy)
Dystonia 28 (KMT2B)
Epileptic encephalopathy, childhood-onset (CHD2)
Floating Harbour syndrome (SRCAP)
Gabriele-de Vries syndrome (YY1)
Genitopatellar syndrome (see also Ohdo syndrome) ⁸ (KAT6B)
Helsmoortel-van der Aa syndrome ⁹ (ADNP)
Hunter McAlpine craniosynostosis syndrome (Chr5q35-qter duplication)
Immunodeficiency-centromeric instabilityfacial anomalies syndromes 1-4 ¹⁰ (DNMT3B, CDCA7, ZBTB24, HELLS (242860, 614069, 616910, 616911)
Intellectual developmental disorder with seizures and language delay (SETD1B)
Intellectual developmental disorder, X-linked 93 ² (BRWD3)
Intellectual developmental disorder, X-linked 97 (ZNF711)
Intellectual developmental disorder, X-linked, syndromic, Armfield type (FAM50A)
Intellectual developmental disorder, X-linked, syndromic, Claes-Jensen type ^2,11 (KDM5C)
Intellectual developmental disorder, X-linked syndromic, Nascimento-type ¹² (UBE2A)
Intellectual developmental disorder, X-linked, Snyder-Robinson type (SMS)
Kabuki syndrome 1 and 2 (KMT2D, KDM6A)
Kabuki syndrome 1 ¹³ (KMT2D)
Kabuki syndrome 2 ¹³ (KDM6A)
KBG Syndrome ¹⁴ (ANKRD11)
KDM2B-related syndrome (KDM2B)
Kleefstra syndrome 1 (EHMT1)
Klinefelter Syndrome (XXY)
Koolen de Vreis syndrome (KANSL1)
Luscan-Lumish syndrome (SETD2)
Menke-Hennekam syndrome 1 and 2 ¹⁵ (CREBBP, EP300) ID4 domains
Mental retardation, autosomal dominant 23 ¹⁴ (SETD5)
Mental retardation, autosomal dominant 51 ² (KMT5B)
Nicolaides-Baraitser syndrome ⁵ (SMARCA2)
Ohdo syndrome, SBBYSS variant (see also Genitopatellar syndrome) ⁸ (KAT6B)
Phelan-McDermid syndrome ¹⁶ (Chr22q13.3 deletion)
Potocki-Lupski syndrome (Chr17p11.2 duplication)
PRC2 Complex (Weaver syndrome and Cohen-Gibson syndrome) ¹⁷ (EED, EZH2)
Rahman syndrome (HIST1H1E)
Renpenning syndrome (PQBP1)
Rubinstein-Taybi syndrome 1 and 2 (CREBBP, EP300)
Rubinstein-Taybi syndrome 1 ¹⁸ (CREBBP)
Rubinstein-Taybi syndrome 2 ¹⁸ (EP300)
Sifrim-Hitz-Weiss yndrome (CHD4)
SLC32A1 related disorder (SLC32A1)
Smith-Magenis syndrome ¹⁹ (Chr17p11.2 deletion)
Sotos syndrome 1 (NSD1)
Tatton-Brown-Rahman syndrome (DNMT3A)
Velocardiofacial syndrome (Chr22q11.2 deletion)
White-Sutton syndrome (POGZ)
Wieacker-Wolff Syndrome (ZC4H2)
Wiedemann-Steiner syndrome (KMT2A)
Williams-Beuren syndrome (Chr7q11.23 deletion)
Williams-Beuren region duplication syndrome (Chr7q11.23 duplication)
Witteveen-Kolk syndrome (SIN3A)
Wolf-Hirschhorn syndrome ²⁰ (Chr4p16.13 deletion, NSD2)

^1. Patients with other BAFopathy genes may be detected, but not confirmed in our experiments.
^2. Healthy carriers and those with incomplete penetrance are detectable.
^3. Patients with biallelic variants are distinguishable from those with monoallelic variants.
^4. Only for variants near c.6200. No separate episignature due small cohort size, however these samples cluster separately from other BAFopathy/CSS1&2 samples.
^5. This is a secondary signature; sample must also be positive for BAFopathy signature.
^6. Only for variants at c.2656. No separate episignature due small cohort size however these samples cluster separately from other BAFopathy/CSS4 samples.
^7. Male CdLS5 patients (HDAC8 mutations) may be detected, but not confirmed in our experiments.
^8. GTPTS and SBBYSS are both caused by KAT6B mutations. We will report both regardless of which one is requested.
^9. ADNP consists of two distinct episignatures dependent on variant location. HVDAS_T includes variants within the N- and C-terminus while HVDAS_C includes variants within the central region (approximately c.2054-2340).
^10. ICF1 exhibits a unique episignature while ICF 2, 3 and 4 exhibit a distinct, shared episignature.
^11. Heterozygotes have a distinct profile from hemizygotes
^12. Carriers have not been detected in our experiments.
^13. This is a secondary signature; sample must also be positive for combined Kabuki signature.
^14. KBGS and MRD23 share a common episignature. Separate KGBS and MRD23 episignatures will be used as secondary signatures, with sample positivity for the combined KBGS/MRD23 episignature required.
^15. Only for domain ID4. MKHK1/2 exhibit a shared ID4 domain episignature and therefore cannot distinguish between MKHK1 and MKHK2. Other domains of MKHK1/2 are not available for assessment.
^16. Only for copy number variants. Sequence variants in SHANK3 have been shown to not match the episignature.
^17. Shared episignatures between PRC2 complex syndromes WVS and COGIS.
^18. This is secondary signature; sample must also be positive for combined RSTS signature.
¹⁹. Only for copy number variants. Sequence variants in RAI1 have been shown to not match the episignature.
^20. WHS episignature can detect truncating variants in NSD2.

The following list of genes have been classified as having reduced sensitivity and more moderate episignatures based on
episignature strength, limited reference cohort size, or types of mutations that have been tested: ANKRD11, BRWD3,
CCNK, CDCA7, CDK13, CHD8, DNMT1, DNMT3A, DNMT3B, FAM50A, HELLS, KAT6A, KAT6B, KMT5B, PHF6, PQBP1, SETD5,
SIN3A, SLC32A1, SOX11, SMS, UBE2A, YY1, ZBTB24, ZC4H2, ZNF711, Chr1p36del, Chr17p11.2dup.

Pheripheral blood collected in an EDTA (lavender top) tube is the required specimen type. See Conditions application Genome Diagnostics.

EpiSign can only be performed on DNA isolated from peripheral blood.

The test CANNOT be performed on DNA isolated from umbilical cord blood, blood from premature babies or other tissues.

Extracted DNA or banked is also accepted for this test given the original specimen was whole blood collected in an EDTA tube, minumun required amount 5 ug.

Profile matches to a specific syndrome

• Variant present: This result supports the diagnosis and pathogenicity of the variant has become more likely.
• No variant present: This result supports the diagnosis and further investigation of the linked gene(s) is recommended. However, it cannot be completely ruled out that a mutation is located in another (yet to be discovered) gene that results in the same or largely overlapping episignature.

Profile does not match a specific syndrome or any of the syndromes in the EpiSign panel

• Variant present: pathogenicity of the variant is not completely ruled out. It is possible that a new variant results in a (yet unidentified) different methylation profile.

A number of profiles have been designated as less sensitive because they have either a less strong profile, or are based on a small number of patients or limited mutation type. For these syndromes (see list of disorders) the chance of an uncertain result or a false negative test is slightly higher. EpiSign is also less sensitive in patients with a mosaic variant, or in women with an X-linked disorder.