Legius Syndrome

Comprehensive Testing from Blood (SPD1)                                                           

DESCRIPTION

Mendelian Inheritance in Man number: 611431

Recently, a new autosomal dominant condition resembling NF1 was described, consisting of mainly multiple CAL-spots, freckling and relative macrocephaly (Brems et al, 2007). Some patients also have learning disabilities or hyperactivity. In none of the adult patients identified so far, neurofibromas or central nervous system tumors were observed Brems et al, 2007; Pasmant et al, 2009; Spurlock et al, 2009; Messiaen et al, 2009; Denayer et al, 2011. In addition, no Lisch nodules, optic pathway gliomas or NF1-associated osseous lesions have been found. A Noonan-like dysmorphology was observed in some individuals. The disorder is caused by germline loss-of-function mutations in SPRED1. ~50% of the individuals carrying a SPRED1 mutation fulfill the NIH diagnostic criteria for NF1, based on the pigmentary symptoms. So far, only SPRED1 point mutations had been identified as the cause of this syndrome. Using Multiplex Ligation-dependent Probe Amplification (MLPA) assay covering all SPRED1 exons and quantitative PCR in a cohort of 510 NF1-negative patients presenting with multiple CALMs with or without freckling, but no other NF1 diagnostic signs, 4 different deletions were identified (Spencer et al, AJMG In Press). Deletions account for ~10% of SPRED1 mutations in this cohort. These results indicate the need for dosage analysis to complement sequencing-based SPRED1 mutation analyses.

In individuals with CALMs with or without freckling and no other specific distinguishing features, the NIH criteria cannot reliably distinguish NF1 from Legius syndrome. In such patients, a correct diagnosis has important implications for prognosis, counseling, and potential prenatal genetic diagnosis.  Based on a cross-sectional study we estimate that patients presenting sporadically with these pigmentary signs only will carry a mutation in the NF1 gene in ~43% of cases and in the SPRED1 gene in ~1.3% of cases (Messiaen et al, 2009). When such patients have a family history of CALMs with or without freckling and no additional NF1-related criteria, an NF1 mutation will be found in ~73% of cases and in the SPRED1 gene in ~19% of cases.

SPRED1 is a member of the SPROUTY/SPRED family of proteins that act as negative regulators of RAS-RAF interaction and mitogen-activated protein kinase (MAPK) signaling.


INDICATIONS FOR DIRECT TESTING

  • Individuals with CAL-spots with or without skinfold freckling and no NF1 mutation after comprehensive NF1 mutation analysis
  • Individuals who seek confirmation of a clinical diagnosis


TESTING METHODOLOGY

We offer a direct test based on a RNA/cDNA based core assay resulting in the full characterization of the SPRED1 mutation (Test 1-cDNA based after NF1 testing). If NF1 testing was performed previously, the complete SPRED1 coding region is analyzed by sequencing exons 1 to 7 starting from the sample submitted for NF1 testing. In addition, DNA copy number analysis by MLPA is performed.  Mutations screened for include truncating mutations (nonsense, frameshift, splicing mutations), missense mutations, multi-exon deletions or duplications and total gene deletions.

If NF1 testing has not been performed previously, we offer a direct sequencing test starting from DNA (Test 1-gDNA based if NF1 testing was not previously done), complemented with MLPA to detect copy number changes.


SPECIMEN REQUIREMENTS

We require 10 milliliters of whole blood. Blood samples must be collected in EDTA (purple topped) tubes. For patients for whom venipuncture is very difficult, a minimum of 3 ml in EDTA suffices.

If NF1 testing was previously performed, but over one year ago, please contact us at medgenomics@uab.edu to verify whether residual sample is still available to perform SPRED1 testing.

For all samples submitted for clinical SPRED1 testing before April 2009, SPRED1 copy number analysis can be requested if previous analysis by sequencing was negative. This additional testing will be performed at no extra charge.


TRANSPORT

If specimen is from clinics within UAB or Kirklin Clinic, please call (205)-934-5562 for pick-up.

IMPORTANT!
Blood specimens must be kept at room temperature and received within 60-72 hours of collection.

1.    DO NOT SHIP ON ICE.
2. Be sure that the shipping air bill is marked “Priority”, either Domestic or International.
3. Specimens must be packaged to prevent breakage and absorbent material must be included in the package to absorb liquids in the event that breakage occurs.  Also, the package must be shipped in double watertight containers (e.g. a specimen pouch + the shipping companies Diagnostic Envelope).  You can use our collection kits, which we will send to physicians directly upon request.
4.   Please contact us (Email – medgenomics@uabmc.edu, Phone – 205-934-5562) prior to sample shipment and provide us with the date of shipment and the tracking number of the package, so that we can better ensure receipt of the samples within the 60-72 hour window. Please include the form for customs. This is especially important for samples sent from outside the US.


TURN AROUND TIME

10 working days if SPRED1 is requested after NF1 testing (and starting from the available sample)

15 working days if SPRED1 is requested first (starting from a new blood sample).


CPT CODES AND PRICES

Please find the most up to date prices and CPT codes for our testing services under the "Prices" tab of this website.


REQUIRED FORMS

SPRED1 Test Requisition including the phenotypic data form.

Form for customs (International shipment)

Note: Detailed and accurate completion of this document is necessary for reporting purposes. The Medical Genomics Laboratory issues its clinical reports based on the demographic data provided by the referring institution on the lab requisition form. It is the responsibility of the referring institution to provide accurate information. If an amended report is necessary due to inaccurate or illegible documentation, additional reports will be drafted with charge.

Requests for Molecular Genetic testing for SPRED1 will not be accepted for the following reasons:

  • No label (patient’s full name and date of collection) on the specimens
  • No referring physician’s or genetic counselor’s names and addresses
  • No billing information
  • No informed consent
  • No phenotypic checklist

 

For more information, test requisition forms, or sample collection and mailing kits, please call: 205-934-5562.


REFERENCES

Brems H, Chmara M, Sahbatou M, Denayer E, Taniguchi K, Kato R, Somers R, Messiaen L, De Schepper S, Fryns J-P, Cools J, Marynen P, Thomas G, Yoshimura A, Legius E – Germline loss-of function mutations in SPRED1 cause a neurofibromatosis 1-like phenotype.  Nat Genet 39: 1120-1126, 2007. (pubmed)

Messiaen L, Yao S, Brems H, Callens T, Sathienkijkanchai A, Denayer E, Spencer E, Arn P, Babovic-Vuksanovic D, Bay C, Bobele G, Cohen BH, Escobar L, Eunpu D, Grebe T, Greenstein R, Hachen R, Irons M, Kronn D, Lemire E, Leppig K, Lim C, McDonald M, Narayanan V, Pearn A, Pedersen R, Powell B, Shapiro LR, Skidmore D, Tegay D, Thiese H, Zackai EH, Vijzelaar R, Taniguchi K, Ayada T, Okamoto F, Yoshimura A, Parret A, Korf B, Legius E- Clinical and Mutational Spectrum of Neurofibromatosis type 1-like syndrome.  JAMA 2009: 302(19): 2111-8 (pubmed)

Muram-Zborovski TM, Stevenson DA, Viskochil DH, Dries DC, Wilson AR, Mao R. 2010. SPRED1 Mutations in a Neurofibromatosis Clinic. J Child Neurol. (pubmed)

Pasmant E, Sabbagh A, Hanna N, Masliah-Planchon J, Jolly E, Goussard P, Ballerini P, Cartault F, Barbarot S, Landman-Parker J, Soufir N, Parfait B, Vidaud M, Wolkenstein P, Vidaud D. 2009. SPRED1 germline mutations caused a neurofibromatosis type 1 overlapping phenotype. J Med Genet. (pubmed)

Spurlock G, Bennett E, Chuzhanova N, Thomas N, Jim H, Side L, Davies S, Haan E, Kerr B, Huson SM, Upadhyaya M. 2009. SPRED1 mutations (Legius syndrome): another clinically useful genotype for dissecting the NF1 phenotype. J Med Genet.  (pubmed)

Spencer E, Davis J, Mikhail F, Fu C, Vijzelaar R, Zackai E, Feret H, Meyn  S, Shugar A, Bellus G, Kocsis K, Kivirikko S, Poyhonen M, Messiaen L,.  Identification of SPRED1 Deletions using RT-PCR, Multiplex Ligation-dependent Probe Amplification and Quantitative PCR. American Journal of Medical Genetics. In Press