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Test Code LAB1835 Craniosynostosis Sequencing Panel

Clinical System Name

Craniosynostosis Sequencing

Description

Craniosynostosis is the premature fusion of one or more of the cranial sutures. Although craniosynostosis has been associated with more than 150 different syndromes either as a major or minor feature, nearly 65-85% of patients present with nonsyndromic craniosynostosis with no additional birth defects.

 

The etiology of craniosynostosis is complex with both environmental and genetic risk factors reported. The Seattle Children's Hospital Molecular Laboratory offers different craniosynostosis panels based on clinical presentation with additional testing options:

 

  • Craniosynostosis Focused Sequencing Panel The focused gene panel  includes FGFR1, FGFR2, FGFR3, TWIST1, TCF12, EFNB1, and ERF. Pathogenic variants in these 7 genes account for approximately 75% of individuals with syndromic craniosynostosis.

 

  • Craniosynostosis Expanded Sequencing Panel: Clinically significant variants have also been identified in approximately 70 genes to date and the option to order or reflex to this expanded panel is available. See Methodology for complete gene list.

 

  • Additional testing options - we offer the following additional options for custom craniosynostosis testing:
    • Single-gene sequencing When there is strong clinical suspicion, any gene listed on our Craniosynostosis Expanded Panel is available as a single-gene sequencing test.
    • Reflex to Expanded Sequencing Panel The option to reflex to the expanded panel when single-gene or the focused panel test is non-diagnostic.

 

Please contact LabGC@seattlechildrens.org if you would like to order reflexive testing after the original testing report has been issued.

Sample Requirements

Note: For patients who have had a whole blood transfusion, wait 10 days post transfusion to draw for genetic testing. No wait time is necessary for blood or saliva collection if the patient received leuko-reduced red cells or plasma.

 

Specimen: Whole blood

Container(s): Lavender/EDTA

Preferred Vol: 3 mL

Minimum Vol: 1 mL

 

Specimen: Saliva collected using Oragene Dx OGD-575/675 collection kit.

Container: Oragene Dx OGD-575/675 collection kit

IMPORTANT NOTE: Manufacturer instructions must be followed. The Oragene Dx OGD575/675 kit is not for children under 6 months. Contact the lab directly for more information or to obtain a kit - 206-987-2617

 

Specimen: Extracted DNA (MUST specify source on requisition)

Preferred: 10µg

Minimum: 5µg

Note: Isolation of nucleic acids for clinical testing must be performed in a CLIA-certified
laboratory or a laboratory meeting equivalent requirements as determined by the CAP
and/or the CMS. DNA concentration minimum 50 µg/mL; 260/280 ratio 1.70-2.00.

Processing Instructions

Reject due to: Heparin

Spin: No

Aliquot: No

Temp: 2 - 8 C

Storage location: Molecular Genetics box in CPA refrigerator #2

Off-site collection: Refrigerate blood samples until ready to ship.  Transport all sample types at room temperature via overnight shipping.

Stability

Specimen Type Temperature Time
Whole blood, extracted DNA RT 3-5 d
Whole blood, extracted DNA 2 - 8 C

7 d

Saliva, extracted from ORAgene Dx OGD-575/675

 room temp or refrigerated up to 2 weeks
Extracted DNA -20 C or -70 C  years

Note: Whole blood samples > 7days may be submitted to be assessed by our lab for acceptability for testing.

Availability

STAT Performed TAT
Contact lab Monday - Friday 4-6 weeks

Performing Laboratory

Seattle Children's Laboratory

Department

Department:  Molecular Genetics Laboratory

Phone: 206-987-3872

Lab Client Services: 206-987-2617

Lab Genetic Counselor: LabGC@seattlechildrens.org

CPT Codes

81479

Methodology

Method: Next Generation Sequencing technology using an Illumina NextSeq instrument. Target region includes coding exons and a minimum of 10 bp of flanking intron boundaries of the genes tested. Target enrichment performed using a custom Integrated DNA Technologies (IDT) panel. 

 

Average coverage is ~150x, depth of coverage for all target regions is at least 20x.

 

Reported gene set:

Craniosynostosis Focused Sequencing Panel: EFNB1, ERF, FGFR1, FGFR2, FGFR3, TWIST1, TCF12

 

Craniosynostosis Expanded Sequencing Panel: ACTB, ACTG1, ADAMTSL4, AHDC1, ALPL, ALX4, ASXL1, ASXL3, ATR, B3GAT3, CDC45, CHD5, CHD7, COLEC10, COLEC11, CTSK, CYP26B1, EFNB1, ERF, FAM20C, FGF9, FGFR1, FGFR2, FGFR3, FLNA, FREM1, GLI3, GPC3, HNRNPK, HUWE1, IFT122, IFT43, IGF1R, IL11RA, IRX5, JAG1, KAT6A, KAT6B, KMT2D, KRAS, MAN2B1, MASP1, MEGF8, MSX2, P4HB, PHEX, POR, PTCH1, RAB23, RECQL4, RUNX2, SIX1, SKI, SLC25A24, SMAD6, SOX6, SPECC1L, STAT3, TCF12, TFAP2B, TGFBR1, TGFBR2, TMCO1, TRAF7, TWIST1, WDR19, WDR35, ZEB2, ZIC1, ZNF462

 

 

Limitations:

This testing is performed on an exome backbone with analysis restricted to the panel genes. This method can detect single nucleotide variants (SNVs), small deletions, small insertions, and copy number variants in the regions targeted. Some regions cannot be efficiently captured due to sequence homology or sequence properties. This method will not detect large insertions and deletions, complex indels, structural variants (e.g. inversions, translocations), short tandem repeats, or other complex variants. Variants located outside of targeted regions will not be detected.

 

Based on validation studies, the bioinformatics pipeline showed precision and detection >99% for SNVs in regions with coverage greater than 20x and high mapping quality. Sensitivity for CNVs involving multiple genes is >99% and sensitivity for intragenic CNVs is >90%. Mosaic sequence variants present at <25% allele frequency may not be reliably detected, and detection sensitivity is dependent on the nature of the variant. The sensitivity of detection of mosaic copy number variants has not been evaluated.

Reference Range

Interpretive report will be provided. Variants are not reported if they are considered benign.

Requisition

Molecular Genetics

Clinical Utility

Approximately 80% of individuals with a syndromic form of craniosynostosis will have an underlying genetic etiology identified by this panel. This is not a complete list, but genetic testing may be considered in individuals when the clinical differential diagnosis includes:

  • 3MC syndrome
  • Apert syndrome
  • Cranioectodermal dysplasia
  • Crouzon syndrome
  • Jackson-Weiss syndrome
  • Loeys-Dietz syndrome
  • Mowat-Wilson syndrome
  • Muenke syndrome
  • Pfeiffer syndrome
  • Saethre-Chotzen syndrome