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DOORS Syndrome and TBC1D24-related Epilepsy via the TBC1D24 Gene

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

NGS Sequencing

Test Code Test Copy GenesIndividual Gene PriceCPT Code Copy CPT Codes
4261 TBC1D24$690.00 81479 Add to Order
Pricing Comment

Our most cost-effective testing approach is NextGen sequencing with Sanger sequencing supplemented as needed to ensure sufficient coverage and to confirm NextGen calls that are pathogenic, likely pathogenic or of uncertain significance. If, however, full gene Sanger sequencing only is desired (for purposes of insurance billing or STAT turnaround time for example), please see link below for Test Code, pricing, and turnaround time information.

For Sanger Sequencing click here.
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 28 days.

Clinical Sensitivity

In a study of 26 families with DOORS syndrome, pathogenic TBC1D24 variants were identified in 9 (~35%) families (Campeau et al. 2014).

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Deletion/Duplication Testing via aCGH

Test Code Test Copy GenesIndividual Gene PriceCPT Code Copy CPT Codes
600 TBC1D24$690.00 81479 Add to Order
Pricing Comment

# of Genes Ordered

Total Price

1

$690

2

$730

3

$770

4-10

$840

11-30

$1,290

31-100

$1,670

Over 100

Call for quote

Turnaround Time

The great majority of tests are completed within 28 days.

Clinical Features

Variants in the TBC1D24 gene have recently been implicated in a number of neurological conditions with varied clinical presentations. TBC1D24 variants were identified in patients with focal epilepsy and intellectual disability. Seizure onset occurred at 2 months of age and consisted of focal seizures with eye blinking and facial and hand twitching (Corbett et al. 2010). These seizures persisted throughout life and were sensitive to fever, but were well controlled with medication (Afawi et al. 2013). Mild developmental delay was noted in early development. There was borderline to moderate intellectual disability seen in adulthood, dysarthria, and ataxia. MRI revealed abnormal cortical thickening. TBC1D24 has also been implicated in patients with progressive encephalopathy. Features included infantile onset myoclonic seizures, dystonia, and psychomotor delay (Guven and Tolun 2013). Variable features included postictal enduring hemiparesis, refractory seizures, bilateral optic atrophy, and macular degeneration. MRI revealed moderate cerebral atrophy and ventricular enlargement, both of which progressively declined as the patient aged. Death was reported in the first decade. Another study identified TBC1D24 variants in siblings with malignant migrating partial seizures of infancy (MMPSI)(Milh et al. 2013). These patients showed infantile onset of tonic-clonic seizures which increased in frequency until they approached status epilepticus. Psychomotor regression was seen after 3 months and was associated with hypotonia, loss of visual contact, and absence of prehension. More recently, whole exome sequencing studies have identified TBC1D24 as a cause of deafness, onychodystrophy, osteodystrophy, mental retardation, and seizures (DOORS) syndrome. The core features seen in TBC1D24 positive DOORS patients are intellectual disability, seizures, deafness, short distal phalanges and small or absent nails (Campeau et al. 2014). MRI in DOORS syndrome patients were heterogeneous, but revealed structural abnormalities in 50% of patients. In addition to these conditions which feature seizures as a primary symptom, TBC1D24 variants can also cause autosomal recessive and autosomal dominant non-syndromic hearing loss (ARNSHL, ADNSHL) in the absence of seizures (Rehman et al. 2014; Azaiez et al. 2014; Zhang et al. 2014).

Genetics

DOORS syndrome and TBC1D24-related epilepsy are inherited in an autosomal recessive manner and are caused by variants in the TBC1D24 gene. Pathogenic missense, nonsense, splice site, and frameshift variants have been reported in TBC1D24 (Campeau et al. 2014). TBC1D24 encodes a protein with an N-terminal Tre2/Bub2/Cdc16 (TBC) domain which is shared by other Rab GTPase-activating proteins (Rab-GAPs). The TBC1D24 TBC domain lacks the arginine and glutamine residues required for GAP activity, suggesting it may not act as a Rab-GAP (Corbett et al. 2010). TBC1D24 is expressed in the brain and physically interacts with ADP ribosylation factor 6 (ARF6)- a GTP-binding protein involved in membrane exchange. Studies in rat neurons suggest that TBC1D24 positively regulates radial neuronal migration by maintaining ARF6 in an inactive state (Falace et al. 2014). Many reported variants in TBC1D24 disrupt its interaction with ARF6. Both loss of TBC1D24 function and expression of constitutively active ARF6 result in similar neuronal migration defects (Falace et al. 2014).

Testing Strategy

For this Next Generation Sequencing (NGS) test, sequencing is accomplished by capturing specific regions with an optimized solution-based hybridization kit, followed by massively parallel sequencing of the captured DNA fragments. Additional Sanger sequencing is performed for regions not captured or with insufficient number of sequence reads. All reported pathogenic, likely pathogenic, and variants of uncertain significance are confirmed by Sanger sequencing.

For Sanger sequencing, polymerase chain reaction (PCR) is used to amplify targeted regions. After purification of the PCR products, cycle sequencing is carried out using the ABI Big Dye Terminator v.3.0 kit. PCR products are resolved by electrophoresis on an ABI 3730xl capillary sequencer. In nearly all cases, cycle sequencing is performed separately in both the forward and reverse directions.

This test provides full coverage of all coding exons of the TBC1D24 gene, plus ~10 bases of flanking noncoding DNA. We define full coverage as >20X NGS reads or Sanger sequencing.

Indications for Test

TBC1D24 testing should be considered for patients with suspected DOORS syndrome and patients with infantile epilepsy and intellectual disability of unknown cause.

Gene

Official Gene Symbol OMIM ID
TBC1D24 613577
Inheritance Abbreviation
Autosomal Dominant AD
Autosomal Recessive AR
X-Linked XL
Mitochondrial MT

Related Tests

Name
Autism Spectrum Disorders and Intellectual Disability (ASD-ID) Comprehensive Sequencing Panel with CNV Detection
Early Infantile Epileptic Encephalopathy Sequencing Panel
Early Infantile Epileptic Encephalopathy, Recessive Sequencing Panel
Non-syndromic Intellectual Disability (NS-ID) Sequencing Panel with CNV Detection

CONTACTS

Genetic Counselors
Geneticist
Citations
  • Azaiez H, Booth KT, Bu F, Huygen P, Shibata SB, Shearer AE, Kolbe D, Meyer N, Black-Ziegelbein EA, Smith RJH. 2014. TBC1D24 Mutation Causes Autosomal-Dominant Nonsyndromic Hearing Loss. Human Mutation 35: 819–823. PubMed ID: 24729539
  • Campeau PM, Hennekam RC, The DOORS syndrome collaborative group. 2014. DOORS syndrome: Phenotype, genotype and comparison with Coffin-Siris syndrome: AMERICAN JOURNAL OF MEDICINE PART C (SEMINARS IN MEDICAL GENETICS). American Journal of Medical Genetics Part C: Seminars in Medical Genetics 166: 327–332. PubMed ID: 25169651
  • Campeau PM, Kasperaviciute D, Lu JT, Burrage LC, Kim C, Hori M, Powell BR, Stewart F, Felix TM, Ende J van den, Wisniewska M, Kayserili H, et al. 2014. The genetic basis of DOORS syndrome: an exome-sequencing study. Lancet Neurol 13: 44–58. PubMed ID: 24291220
  • Corbett MA, Bahlo M, Jolly L, Afawi Z, Gardner AE, Oliver KL, Tan S, Coffey A, Mulley JC, Dibbens LM, Simri W, Shalata A, et al. 2010. A Focal Epilepsy and Intellectual Disability Syndrome Is Due to a Mutation in TBC1D24. The American Journal of Human Genetics 87: 371–375. PubMed ID: 20797691
  • Falace A, Buhler E, Fadda M, Watrin F, Lippiello P, Pallesi-Pocachard E, Baldelli P, Benfenati F, Zara F, Represa A, Fassio A, Cardoso C. 2014. TBC1D24 regulates neuronal migration and maturation through modulation of the ARF6-dependent pathway. Proceedings of the National Academy of Sciences 111: 2337–2342. PubMed ID: 24469796
  • Guven A, Tolun A. 2013. TBC1D24 truncating mutation resulting in severe neurodegeneration. Journal of Medical Genetics 50: 199–202. PubMed ID: 23343562
  • Milh M, Falace A, Villeneuve N, Vanni N, Cacciagli P, Assereto S, Nabbout R, Benfenati F, Zara F, Chabrol B, Villard L, Fassio A. 2013. Novel Compound Heterozygous Mutations in TBC 1 D 24 Cause Familial Malignant Migrating Partial Seizures of Infancy. Human Mutation 34: 869–872. PubMed ID: 23526554
  • Rehman AU, Santos-Cortez RLP, Morell RJ, Drummond MC, Ito T, Lee K, Khan AA, Basra MAR, Wasif N, Ayub M, Ali RA, Raza SI, et al. 2014. Mutations in TBC1D24, a Gene Associated With Epilepsy, Also Cause Nonsyndromic Deafness DFNB86. The American Journal of Human Genetics 94: 144–152. PubMed ID: 24387994
  • Zhang L, Hu L, Chai Y, Pang X, Yang T, Wu H. 2014. A Dominant Mutation in the Stereocilia-Expressing Gene TBC1D24 is a Probable Cause for Nonsyndromic Hearing Impairment. Human Mutation 35: 814–818. PubMed ID: 24729547
Order Kits
TEST METHODS

NextGen Sequencing using PG-Select Capture Probes

Test Procedure

We use a combination of Next Generation Sequencing (NGS) and Sanger sequencing technologies to cover the full coding regions of the listed genes plus ~20 bases of non-coding DNA flanking each exon.  As required, genomic DNA is extracted from the patient specimen.  For NGS, patient DNA corresponding to these regions is captured using an optimized set of DNA hybridization probes.  Captured DNA is sequenced using Illumina’s Reversible Dye Terminator (RDT) platform (Illumina, San Diego, CA, USA).  Regions with insufficient coverage by NGS are covered by Sanger sequencing.  All pathogenic, likely pathogenic, or variants of uncertain significance are confirmed by Sanger sequencing.

For Sanger sequencing, Polymerase Chain Reaction (PCR) is used to amplify targeted regions.  After purification of the PCR products, cycle sequencing is carried out using the ABI Big Dye Terminator v.3.0 kit.  PCR products are resolved by electrophoresis on an ABI 3730xl capillary sequencer.  In nearly all cases, cycle sequencing is performed separately in both the forward and reverse directions.

Patient DNA sequence is aligned to the genomic reference sequence for the indicated gene region(s). All differences from the reference sequences (sequence variants) are assigned to one of five interpretation categories, listed below, per ACMG Guidelines (Richards et al. 2015).

(1) Pathogenic Variants
(2) Likely Pathogenic Variants
(3) Variants of Uncertain Significance
(4) Likely Benign Variants
(5) Benign, Common Variants

Human Genome Variation Society (HGVS) recommendations are used to describe sequence variants (http://www.hgvs.org).  Rare variants and undocumented variants are nearly always classified as likely benign if there is no indication that they alter protein sequence or disrupt splicing.

Analytical Validity

As of March 2016, 6.36 Mb of sequence (83 genes, 1557 exons) generated in our lab was compared between Sanger and NextGen methodologies. We detected no differences between the two methods. The comparison involved 6400 total sequence variants (differences from the reference sequences). Of these, 6144 were nucleotide substitutions and 256 were insertions or deletions. About 65% of the variants were heterozygous and 35% homozygous. The insertions and deletions ranged in length from 1 to over 100 nucleotides.

In silico validation of insertions and deletions in 20 replicates of 5 genes was also performed. The validation included insertions and deletions of lengths between 1 and 100 nucleotides. Insertions tested in silico: 2200 between 1 and 5 nucleotides, 625 between 6 and 10 nucleotides, 29 between 11 and 20 nucleotides, 25 between 21 and 49 nucleotides, and 23 at or greater than 50 nucleotides, with the largest at 98 nucleotides. All insertions were detected. Deletions tested in silico: 1813 between 1 and 5 nucleotides, 97 between 6 and 10 nucleotides, 32 between 11 and 20 nucleotides, 20 between 21 and 49 nucleotides, and 39 at or greater than 50 nucleotides, with the largest at 96 nucleotides. All deletions less than 50 nucleotides in length were detected, 13 greater than 50 nucleotides in length were missed. Our standard NextGen sequence variant calling algorithms are generally not capable of detecting insertions (duplications) or heterozygous deletions greater than 100 nucleotides. Large homozygous deletions appear to be detectable.   

Analytical Limitations

Interpretation of the test results is limited by the information that is currently available.  Better interpretation should be possible in the future as more data and knowledge about human genetics and this specific disorder are accumulated.

When Sanger sequencing does not reveal any difference from the reference sequence, or when a sequence variant is homozygous, we cannot be certain that we were able to detect both patient alleles.  Occasionally, a patient may carry an allele which does not amplify, due to a large deletion or insertion.   In these cases, the report will contain no information about the second allele.  Our Sanger and NGS Sequencing tests are generally not capable of detecting Copy Number Variants (CNVs).

We sequence all coding exons for each given transcript, plus ~20 bp of flanking non-coding DNA for each exon.  Test reports contain no information about other portions of the gene, such as regulatory domains, deep intronic regions or any currently uncharacterized alternative exons.

In most 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 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.

Unless otherwise indicated, DNA sequence data is obtained from a specific cell-type (usually leukocytes from whole blood).   Test reports contain no information about the DNA sequence in other cell-types.

We cannot be certain that the reference sequences are correct.

Rare, low probability interpretations of sequencing results, such as for example the occurrence of de novo mutations in recessive disorders, are generally not included in the reports.

We have confidence in our ability to track a specimen once it has been received by PreventionGenetics.  However, we take no responsibility for any specimen labeling errors that occur before the sample arrives at PreventionGenetics.

Deletion/Duplication Testing Via Array Comparative Genomic Hybridization

Test Procedure

Equal amounts of genomic DNA from the patient and a gender matched reference sample are amplified and labeled with Cy3 and Cy5 dyes, respectively. To prevent any sample cross contamination, a unique sample tracking control is added into each patient sample. Each labeled patient product is then purified, quantified, and combined with the same amount of reference product. The combined sample is loaded onto the designed array and hybridized for at least 22-42 hours at 65°C. Arrays are then washed and scanned immediately with 2.5 µM resolution. Only data for the gene(s) of interest for each patient are extracted and analyzed.

Analytical Validity

PreventionGenetics' high density gene-centric custom designed aCGH enables the detection of relatively small deletions and duplications within a single exon of a given gene or deletions and duplications encompassing the entire gene. PreventionGenetics has established and verified this test's accuracy and precision.

Analytical Limitations

Our dense probe coverage may allow detection of deletions/duplications down to 100 bp; however due to limitations and probe spacing this cannot be guaranteed across all exons of all genes. Therefore, some copy number changes smaller than 100-300 bp within a targeted large exon may not be detected by our array.

This array may not detect deletions and duplications present at low levels of mosaicism or those present in genes that have pseudogene copies or repeats elsewhere in the genome.

aCGH will not detect balanced translocations, inversions, or point mutations that may be responsible for the clinical phenotype.

Breakpoints, if occurring outside the targeted gene, may be hard to define.

The sensitivity of this assay may be reduced when DNA is extracted by an outside laboratory.

Order Kits

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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