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Charcot Marie Tooth - Demyelinating Neuropathy Sequencing Panel

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

NGS Sequencing

Test Code Test Copy GenesCPT Code Copy CPT Codes
2643 COX6A1 81479 Add to Order
CTDP1 81479
DNM2 81479
EGR2 81404
FBLN5 81479
FGD4 81479
FIG4 81406
GDAP1 81405
GJB1 81403
GNB4 81479
HK1 81479
INF2 81406
KARS 81479
LITAF 81404
MPZ 81405
MTMR2 81479
NDRG1 81479
NEFL 81405
PLEKHG5 81479
PMP22 81325
PRX 81405
SBF1 81479
SBF2 81479
SH3TC2 81406
YARS 81479
Full Panel Price* $750.00
Test Code Test Copy Genes Total Price CPT Codes Copy CPT Codes
2643 Genes x (25) $750.00 81325, 81403, 81404(x2), 81405(x4), 81406(x3), 81479(x14) Add to Order
Pricing Comment

If you would like to order a subset of these genes contact us to discuss pricing.

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

A genetic etiology can be identified in approximately 50-70% of individuals with CMT (Saporta et al. 2011; Rossor et al. 2013). Specifically, a molecular diagnosis can be identified in approximately 80-85% of individuals with demyelinating neuropathy (CMT1), and a molecular diagnosis can be identified in approximately 25-35% of individuals with axonal neuropathy (CMT2) (Bird 2015; Bird 2015; Rossor et al. 2013). The sensitivity of this panel will vary based on the clinical phenotype of the patient. It is estimated that ~70% of all Charcot Marie Tooth Type 1 (CMT1) is due to the PMP22 1.5 Mb duplication, while only around 5% of CMT1 cases are due to point mutations (Bird 2015). This NGS sequencing panel will not detect the common 1.5 Mb duplication or deletion in the PMP22 gene.

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Deletion/Duplication Testing via aCGH

Test Code Test Copy GenesIndividual Gene PriceCPT Code Copy CPT Codes
600 COX6A1$690.00 81479 Add to Order
CTDP1$690.00 81479
DNM2$690.00 81479
EGR2$690.00 81479
FBLN5$690.00 81479
FGD4$690.00 81479
FIG4$690.00 81479
GDAP1$690.00 81479
GJB1$690.00 81479
GNB4$690.00 81479
HK1$690.00 81479
INF2$690.00 81479
KARS$690.00 81479
LITAF$690.00 81479
MPZ$690.00 81479
MTMR2$690.00 81479
NDRG1$690.00 81479
NEFL$690.00 81479
PLEKHG5$690.00 81479
PMP22$690.00 81324
PRX$690.00 81479
SBF1$690.00 81479
SBF2$690.00 81479
SH3TC2$690.00 81479
YARS$690.00 81479
Full Panel Price* $1290.00
Test Code Test Copy Genes Total Price CPT Codes Copy CPT Codes
600 Genes x (25) $1290.00 81324, 81479(x24) 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 Sensitivity

Large deletions/duplications have been reported in the FIG4 gene, but no large-scale study has been done (Human Gene Mutation Database).

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Clinical Features

Charcot Marie Tooth disease (CMT), also known as hereditary motor and sensory neuropathy (HMSN) is a large group of inherited disorders of the peripheral nerves. The progressive degeneration of motor nerves results in weakness and atrophy of the distal muscles. The degeneration of sensory nerves leads to decreased sensation, tingling and numbness in the legs, feet, arms and hands and neuropathic pain. The age of onset varies from childhood to mid adulthood. Symptoms usually begin with weakness and atrophy in the muscles of the legs and feet. As the disease progresses, weakness and atrophy of the muscles of the arms and hands may occur. CMT is heterogeneous in regards to symptoms, severity and progression rate. Although the disease may lead to disability and respiratory difficulty, life expectancy is usually unaffected. Most common symptoms include foot deformity, loss of balance, hammertoes, foot drop, frequent tripping and falls, and reduced manual dexterity (Bird 2015). Diagnosis is based on clinical features, family history, neurological examination, and electromyography (EMG) and nerve conduction velocity (NCV) findings (Rossor et al. 2013; Bird 2015). CMT affects approximately 1 in 3,300 people (Bird 2015; Saporta et al. 2011). Demyelinating forms of CMT primarily affect the myelin sheath of the peripheral nerve and are characterized by slow nerve conduction velocities (NCV) of less than 38 m/s in upper limbs. Axonal forms of CMT primarily affect the axons of the peripheral nerves and are characterized by normal or almost normal NCV of greater than 38 m/s. Intermediate NCV of 25-45 m/s can be difficult to classify as axonal or demyelinating. This panel includes genes that cause a strict CMT1 (demyelinating) phenotype, in addition to genes that cause intermediate forms with axonal and/or demyelinating conductions. Approximately 70% of CMT1 is caused by the recurrent PMP22 duplication (Bird 2015; Li et al. 2013; van Paassen et al. 2014). CMT1A is most commonly caused by a 1.5 Mb duplication of chromosome 17p11.2 which includes the PMP22 gene. PMP22 deletion/duplication testing via aCGH can be ordered through test code #600.

Genetics

Demyelinating neuropathies can be inherited in an autosomal dominant, autosomal recessive or an X-linked manner. The MPZ, LITAF, NEFL, PMP22, FBLN5, YARS, INF2, GNB4 genes are involved in autosomal dominant CMT. Autosomal recessive forms of CMT involve the MTMR2, SBF2, SBF1, SH3TC2, PRX, FGD4, FIG4, NDRG1, HK1, KARS, CTDP1, PLEKHG5, and COX6A1 genes. Pathogenic variants in the EGR2, GDAP1, and DNM2 genes can exhibit both dominant and recessive inheritance. Pathogenic variants in the GJB1 gene are inherited in an X-linked dominant manner. See individual gene test descriptions for information on molecular biology of gene products. The most common genetic cause of CMT1 is a 1.5 Mb duplication of chromosome 17p11.2 which includes the PMP22 gene. This sequencing test will not detect this large copy number variant.

Testing Strategy

For this NGS panel, the full coding regions plus ~20 bp of non-coding DNA flanking each exon are sequenced for each of the genes listed below. Sequencing is accomplished by capturing specific regions with an optimized solution-based hybridization method, followed by massively parallel sequencing of the captured DNA fragments. Additional Sanger sequencing is performed for any regions not captured or with insufficient number of sequence reads. All pathogenic, undocumented and questionable variant calls are confirmed by Sanger sequencing. Please note that exon 8 of the INF2 gene is not currently included this panel. Thus far, only exons 2 to 6, especially exons 2 to 4 that encode the diaphanous inhibitory domain (DID), have been reported to harbor pathogenic INF2 variants (Boyer et al. 2011; Barua et al. 2013; Human Gene Mutation Database).

Indications for Test

Individuals with clinical symptoms consistent with a demyelinating neuropathy (nerve conduction velocities of less than 38 m/s).

Genes

Official Gene Symbol OMIM ID
COX6A1 602072
CTDP1 604927
DNM2 602378
EGR2 129010
FBLN5 604580
FGD4 611104
FIG4 609390
GDAP1 606598
GJB1 304040
GNB4 610863
HK1 142600
INF2 610982
KARS 601421
LITAF 603795
MPZ 159440
MTMR2 603557
NDRG1 605262
NEFL 162280
PLEKHG5 611101
PMP22 601097
PRX 605725
SBF1 603560
SBF2 607697
SH3TC2 608206
YARS 603623
Inheritance Abbreviation
Autosomal Dominant AD
Autosomal Recessive AR
X-Linked XL
Mitochondrial MT

Diseases

Name Inheritance OMIM ID
Charcot-Marie-Tooth Disease Dominant Intermediate 3 607791
Charcot-Marie-Tooth Disease Type 2E 607684
Charcot-Marie-Tooth Disease Type 2I 607677
Charcot-Marie-Tooth Disease Type 2J 607736
Charcot-Marie-Tooth Disease Type 2K 607831
Charcot-Marie-Tooth Disease, Axonal, With Vocal Cord Paresis, Autosomal Recessive 607706
Charcot-Marie-Tooth Disease, Dominant Intermediate B 606482
Charcot-Marie-Tooth Disease, Dominant Intermediate C 608323
Charcot-Marie-Tooth Disease, Dominant Intermediate E 614455
Charcot-Marie-Tooth Disease, Dominant Intermediate F 615185
Charcot-Marie-Tooth Disease, Recessive Intermediate A 608340
Charcot-Marie-Tooth Disease, Recessive Intermediate B 613641
Charcot-Marie-Tooth Disease, Recessive Intermediate C 615376
Charcot-Marie-Tooth Disease, Recessive Intermediate D 616039
Charcot-Marie-Tooth Disease, Type 1A 118220
Charcot-Marie-Tooth Disease, Type 1D 607678
Charcot-Marie-Tooth Disease, Type 1E 118300
Charcot-Marie-Tooth Disease, Type 1F 607734
Charcot-Marie-Tooth Disease, Type 3 145900
Charcot-Marie-Tooth Disease, Type 4A 214400
Charcot-Marie-Tooth Disease, Type 4B1 601382
Charcot-Marie-Tooth Disease, Type 4B2 604563
Charcot-Marie-Tooth Disease, Type 4B3 615284
Charcot-Marie-Tooth Disease, Type 4C 601596
Charcot-Marie-Tooth Disease, Type 4D 601455
Charcot-Marie-Tooth Disease, Type 4E 605253
Charcot-Marie-Tooth Disease, Type 4F 614895
Charcot-Marie-Tooth Disease, Type 4H 609311
Charcot-Marie-Tooth Disease, Type 4J 611228
Charcot-Marie-Tooth Disease, Type Ib 118200
Charcot-Marie-Tooth Disease, Type IC 601098
Charcot-Marie-Tooth Disease, X-Linked Dominant, 1 302800
Congenital Cataracts, Facial Dysmorphism, And Neuropathy 604168
Cutis Laxa, Autosomal Recessive, Type IA 219100
Neuropathy, Hereditary Motor and Sensory, Russe Type 605285

Related Tests

Name
KARS-Related Disorders via the KARS Gene
MPZ-Related Neuropathies
PMP22-Related Neuropathies
Autism Spectrum Disorders and Intellectual Disability (ASD-ID) Comprehensive Sequencing Panel with CNV Detection
Centronuclear Myopathy Sequencing Panel
Charcot Marie Tooth - Axonal Neuropathy Sequencing Panel
Charcot Marie Tooth - Comprehensive Sequencing Panel
Charcot Marie Tooth Disease via the COX6A1 Gene
Charcot Marie Tooth Type 4J via the FIG4 Gene
Charcot-Marie-Tooth 1C via the LITAF Gene
Charcot-Marie-Tooth Autosomal Dominant Intermediate C via the YARS Gene
Charcot-Marie-Tooth Autosomal Dominant Intermediate F via the GNB4 Gene
Charcot-Marie-Tooth Type 2E/1F via the NEFL Gene
Charcot-Marie-Tooth Type 4B1 via the MTMR2 Gene
Charcot-Marie-Tooth Type 4B2 via the SBF2 Gene
Charcot-Marie-Tooth Type 4B3 via the SBF1 Gene
Charcot-Marie-Tooth Type 4C via the SH3TC2 Gene
Charcot-Marie-Tooth Type 4D via the NDRG1 Gene
Comprehensive Neuromuscular Sequencing Panel
Comprehensive Neuropathy Sequencing Panel
Congenital Cataracts Facial Dysmorphism Neuropathy (CCFDN) Syndrome via the CTDP1 Gene
Congenital Cataracts Sequencing Panel
Congenital Myopathy Sequencing Panel
Cutis Laxa via the FBLN5 Gene
Dynamin-2 Related Disorders via the DNM2 Gene
Nephrotic Syndrome (NS)/Focal Segmental Glomerulosclerosis (FSGS) Sequencing Panel

CONTACTS

Genetic Counselors
Geneticist
Citations
  • Barua M. et al. 2013. Kidney International. 83: 316-22. PubMed ID: 23014460
  • Bird T.D. 2015. Charcot-Marie-Tooth Hereditary Neuropathy Overview. 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: 20301532
  • Bird T.D. 2015. Charcot-Marie-Tooth Neuropathy Type 1. 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: 20301384
  • Boyer O. et al. 2011. Journal of the American Society of Nephrology : Jasn. 22: 239-45. PubMed ID: 21258034
  • Human Gene Mutation Database (Bio-base).
  • Li J. et al. 2013. Molecular Neurobiology. 47: 673-98. PubMed ID: 23224996
  • Rossor Alexander M. et al. 2013. Nature Reviews Neurology. 9: 562-571. PubMed ID: 24018473
  • Saporta A.S. et al. 2011. Annals of Neurology. 69: 22-33. PubMed ID: 21280073
  • van Paassen B.W. et al. 2014. Orphanet Journal of Rare Diseases. 9: 38. PubMed ID: 24646194
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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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