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Spastic Paraplegia 4 via the SPAST Gene

  • Summary and Pricing
  • Clinical Features and Genetics
  • Citations
  • Methods
  • Ordering/Specimens
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TEST METHODS

Sequencing

Test Code Test Copy GenesIndividual Gene PriceCPT Code Copy CPT Codes
2165 SPAST$1100.00 81406 Add to Order
Targeted Testing

For ordering targeted known variants, please proceed to our Targeted Variants landing page.

Turnaround Time

The great majority of tests are completed within 18 days.

Clinical Sensitivity

Pathogenic variants in SPAST gene account for almost 40% of hereditary spastic paraplegia (HSP) and about 18% of sporadic or apparent sporadic HSP (Lo Giudice et al. 2014). 

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Clinical Features

Spastic paraplegia 4 (SPG4) is a type of hereditary spastic paraplegia (HSP) characterized by progressive bilateral lower-limb gait spasticity. Over 50% of SPG4 patients have weakness in the legs and abnormal vibration sensation at the ankles. About 30% have sphincter disturbances (Dürr et al. 2003). Onset in young adulthood is common, although in some rare cases symptoms may start as late as in the 70s (Depienne et al. 2007). The average age of onset is 34 (McDermott et al. 2006). SPG4 is the most prevalent type of HSP and accounts for almost 40% of familial HSP and about 18% of sporadic or apparent sporadic HSP (Lo Giudice et al. 2014). Typically, SPG4 is characterized by a pure HSP phenotype, i.e., the symptoms are limited to the lower limbs. A minority of SPG4 cases are complicated by some additional features such as seizures, cerebellar ataxia, and intellectual disability (Nielsen et al. 2004; Ribaï et al. 2008).

Genetics

SPG4 is caused by pathogenic variants in the SPAST gene and hence is also known as SPAST-associated HSP (Hazan et al. 1999). It is inherited in an autosomal dominant manner, though one unusual recessive pathogenic variant has been reported (Lindsey et al. 2000). The penetrance is age dependent, and is estimated to be about 85% by the age of 45 (Fonknechten et al. 2000). A few individuals with a pathogenic variant may remain lifelong asymptomatic (Dürr et al. 1996). In addition, intrafamilial symptomatic variability is considerable, suggesting the involvement of environmental factors and genetic modifiers (Tesson et al. 2015).

SPAST encodes the protein spastin, which is ubiquitously expressed and has a higher expression level in the fetal brain, playing roles in membrane trafficking and microtubule dynamics (Salinas et al. 2007). Spastin has two main domains, the MIT (microtubule interacting and transport) domain, and the AAA (ATPases associated with diverse cellular activities) domain. Over 300 different pathogenic variants have been identified in SPAST gene: nonsense, missense, splice site variants, small indels, and rare large deletions. With rare exceptions, most of the variants are located in the AAA domain, and act by a haploinsufficiency mechanism (Beetz et al. 2006).  

Testing Strategy

Testing is accomplished by amplifying the coding exons  of the SPAST gene and ~20 bp of adjacent noncoding sequence, then determining the nucleotide sequence using standard dideoxy Sanger sequencing methods and a capillary electrophoresis instrument. We will also sequence any single exon (Test #100) in family members of patients with a known mutation or to confirm research results.

Indications for Test

Individuals with symptoms consistent with autosomal dominant HSP, and family members of patients who have known SPAST mutations are ideal candidates for this test.  

Gene

Official Gene Symbol OMIM ID
SPAST 604277
Inheritance Abbreviation
Autosomal Dominant AD
Autosomal Recessive AR
X-Linked XL
Mitochondrial MT

Disease

Name Inheritance OMIM ID
Spastic Paraplegia 4 AD 182601

Related Tests

Name
Complex Hereditary Spastic Paraplegia Sequencing Panel
Hereditary Spastic Paraplegia Comprehensive Sequencing Panel
Pure Hereditary Spastic Paraplegia Sequencing Panel

CONTACTS

Genetic Counselors
Geneticist
Citations
  • Beetz C. et al. 2006. Neurology. 67: 1926-30. PubMed ID: 17035675
  • Dürr A. et al. 1996. Brain : a Journal of Neurology. 119 ( Pt 5): 1487-96. PubMed ID: 8931574
  • Dürr et al. 2003. Spastic Paraplegia 4. In: Pagon RA, Adam MP, Bird TD, Dolan CR, Fong C-T, Smith RJ, and Stephens K, editors. GeneReviews™, Seattle (WA): University of Washington, Seattle. PubMed ID: 20301339
  • Depienne C. et al. 2007. Current Opinion in Neurology. 20: 674-80. PubMed ID: 17992088
  • Fonknechten N. et al. 2000. Human Molecular Genetics. 9: 637-44. PubMed ID: 10699187
  • Hazan J. et al. 1999. Nature Genetics. 23: 296-303. PubMed ID: 10610178
  • Lindsey JC. et al. 2000. Journal of Medical Genetics. 37: 759-65. PubMed ID: 11015453
  • Lo Giudice T. et al. 2014. Experimental Neurology. 261: 518-39. PubMed ID: 24954637
  • McDermott CJ. et al. 2006. Neurology. 67: 45-51. PubMed ID: 16832076
  • Nielsen JE. et al. 2004. European Journal of Neurology. 11: 817-24. PubMed ID: 15667412
  • Ribaï P. et al. 2008. European Journal of Human Genetics : Ejhg. 16: 97-104. PubMed ID: 17957230
  • Salinas S. et al. 2007. Journal of Neuroscience Research. 85: 2778-82. PubMed ID: 17348041
  • Tesson C. et al. 2015. Human Genetics. 134: 511-38. PubMed ID: 25758904
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TEST METHODS

Bi-Directional Sanger Sequencing

Test Procedure

Nomenclature for sequence variants was from the Human Genome Variation Society (http://www.hgvs.org).  As required, DNA is extracted from the patient specimen.  PCR is used to amplify the indicated exons plus additional flanking non-coding sequence.  After cleaning of the PCR products, cycle sequencing is carried out using the ABI Big Dye Terminator v.3.0 kit.  Products are resolved by electrophoresis on an ABI 3730xl capillary sequencer.  In most cases, sequencing is performed in both forward and reverse directions; in some cases, sequencing is performed twice in either the forward or reverse directions.  In nearly all cases, the full coding region of each exon as well as 20 bases of non-coding DNA flanking the exon are sequenced.

Analytical Validity

As of March 2016, we compared 17.37 Mb of Sanger DNA sequence generated at PreventionGenetics to NextGen sequence generated in other labs. We detected only 4 errors in our Sanger sequences, and these were all due to allele dropout during PCR. For Proficiency Testing, both external and internal, in the 12 years of our lab operation we have Sanger sequenced roughly 8,800 PCR amplicons. Only one error has been identified, and this was due to sequence analysis error.

Our Sanger sequencing is capable of detecting virtually all nucleotide substitutions within the PCR amplicons. Similarly, we detect essentially all heterozygous or homozygous deletions within the amplicons. Homozygous deletions which overlap one or more PCR primer annealing sites are detectable as PCR failure. Heterozygous deletions which overlap one or more PCR primer annealing sites are usually not detected (see Analytical Limitations). All heterozygous insertions within the amplicons up to about 100 nucleotides in length appear to be detectable. Larger heterozygous insertions may not be detected. All homozygous insertions within the amplicons up to about 300 nucleotides in length appear to be detectable. Larger homozygous insertions may masquerade as homozygous deletions (PCR failure).

Analytical Limitations

In exons where our sequencing did not reveal any variation between the two alleles, we cannot be certain that we were able to PCR amplify both of the patient’s alleles. Occasionally, a patient may carry an allele which does not amplify, due for example to a deletion or a large insertion. In these cases, the report contains no information about the second allele.

Similarly, our sequencing tests have almost no power to detect duplications, triplications, etc. of the gene sequences.

In most cases, only the indicated exons and roughly 20 bp of flanking non-coding sequence on each side are analyzed. Test reports contain little or no information about other portions of the gene, including many regulatory regions.

In nearly all cases, we are unable to determine the phase of sequence variants. In particular, when we find two likely causative mutations for recessive disorders, we cannot be certain that the mutations are on different alleles.

Our ability to detect minor sequence variants, due for example to somatic mosaicism is limited. Sequence variants that are present in less than 50% of the patient’s nucleated cells may not be detected.

Runs of mononucleotide repeats (eg (A)n or (T)n) with n >8 in the reference sequence are generally not analyzed because of strand slippage during PCR and cycle sequencing.

Unless otherwise indicated, the sequence data that we report are based on DNA isolated from a specific tissue (usually leukocytes). Test reports contain no information about gene sequences in other tissues.

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Ordering Options


myPrevent - Online Ordering
  • The test can be added to your online orders in the Summary and Pricing section.
  • Once the test has been added log in to myPrevent to fill out an online requisition form.
REQUISITION FORM
  • A completed requisition form must accompany all specimens.
  • Billing information along with specimen and shipping instructions are within the requisition form.
  • All testing must be ordered by a qualified healthcare provider.

SPECIMEN TYPES
WHOLE BLOOD

(Delivery accepted Monday - Saturday)

  • Collect 3 ml -5 ml (5 ml preferred) of whole blood in EDTA (purple top tube) or ACD (yellow top tube). For Test #500-DNA Banking only, collect 10 ml -20 ml of whole blood.
  • For small babies, we require a minimum of 1 ml of blood.
  • Only one blood tube is required for multiple tests.
  • Ship blood tubes at room temperature in an insulated container. Do not freeze blood.
  • During hot weather, include a frozen ice pack in the shipping container. Place a paper towel or other thin material between the ice pack and the blood tube.
  • In cold weather, include an unfrozen ice pack in the shipping container as insulation.
  • At room temperature, blood specimen is stable for up to 48 hours.
  • If refrigerated, blood specimen is stable for up to one week.
  • Label the tube with the patient name, date of birth and/or ID number.

DNA

(Delivery accepted Monday - Saturday)

  • Send in screw cap tube at least 5 µg -10 µg of purified DNA at a concentration of at least 20 µg/ml for NGS and Sanger tests and at least 5 µg of purified DNA at a concentration of at least 100 µg/ml for gene-centric aCGH, MLPA, and CMA tests, minimum 2 µg for limited specimens.
  • For requests requiring more than one test, send an additional 5 µg DNA per test ordered when possible.
  • DNA may be shipped at room temperature.
  • Label the tube with the composition of the solute, DNA concentration as well as the patient’s name, date of birth, and/or ID number.
  • We only accept genomic DNA for testing. We do NOT accept products of whole genome amplification reactions or other amplification reactions.

CELL CULTURE

(Delivery preferred Monday - Thursday)

  • PreventionGenetics should be notified in advance of arrival of a cell culture.
  • Culture and send at least two T25 flasks of confluent cells.
  • Some panels may require additional flasks (dependent on size of genes, amount of Sanger sequencing required, etc.). Multiple test requests may also require additional flasks. Please contact us for details.
  • Send specimens in insulated, shatterproof container overnight.
  • Cell cultures may be shipped at room temperature or refrigerated.
  • Label the flasks with the patient name, date of birth, and/or ID number.
  • We strongly recommend maintaining a local back-up culture. We do not culture cells.
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