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Neuronal Ceroid Lipofuscinosis 6 via the CLN6 Gene

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

Sequencing

Test Code TestIndividual Gene PriceCPT Code Copy CPT Codes
1534 CLN6$710.00 81479 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 CLN6 were identified in about 14% and 17% of clinically diagnosed NCL cases from Newfoundland and Italy, respectively (Moore et al. 2008; Santorelli et al. 2013).

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Deletion/Duplication Testing via aCGH

Test Code TestIndividual Gene PriceCPT Code Copy CPT Codes
600 CLN6$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

The neuronal ceroid lipofuscinoses are inherited neurodegenerative lysosomal storage disorders caused by the accumulation of ceroid and lipofuscin in various cell types, mainly cells of the cerebral cortex, cerebellar cortex, and retina (Dyken et al. 1988; Williams and Mole 2012; Bennett and Rakheja, 2013). Characteristic features at onset include clumsiness; deterioration of vision and psychomotor functions; seizures and behavioral changes. Progression of clinical features results ultimately in total disability, blindness and premature death. Although NCL affects primarily children, age of onset of symptoms varies from infancy to adulthood. The incidence of NCL is variable and ranges from 1.3 to 7 per 100,000 (Mole and Williams 2013). However, it is more common in northern European populations, particularly  Finland where the incidence may reach 1 in 12,500 individuals and a carrier frequency of 1 in 70 (Rider and Rider 1988; Vesa et al. 1995). NCLs are clinically and genetically heterogeneous. A nomenclature based both on the age of onset of symptoms and the disease-causing gene has been recently developed, which classifies NCLs into thirteen subtypes (CLN1-8, 10-14) (Williams and Mole 2012). The causative gene for the CLN9 phenotype has not been identified yet (Schulz et al. 2004).

CLN6 was previously further divided into a juvenile and late infantile type and an adult type known as Lake-Cavanagh Disease and Kufs Disease type A, respectively. It is now recognized that the age of onset for CLN6 is wide, ranging between 18 months and 51 years. Disease progression is slower compared to that of CLN2 (Kousi et al. 2012). 

Presenting features of CLN6 include seizures, motor difficulties, myoclonic jerks, speech impairment, ataxia and mental decline. Vision impairment appears later in life (Mole et al. 2005).  Electrophysiological findings include electroretinogram and electroencephalogram attenuation, cortical hyperexcitability, and enlarged somatosensory evoked potentials (Canafoglia et al. 2015).

Genetics

Most CLNs (including CLN6) are inherited in an autosomal recessive manner. Thirteen genes have to date been implicated in the disorder: PPT1, TPP1, CLN3, CLN5, CLN6, MFSD8, CLN8, CTSD, DNAJC5, CTSF, ATP13A2, GRN, and KCTD7 (Mole and Williams 2013).

CLN6 is caused by pathogenic variants in the CLN6 gene (Gao et al. 2002; Wheeler et al. 2002). Over 60 variants have been reported in various ethnic populations. The vast majority are missense. Other types include nonsense, splicing, and small insertions or deletions. Only one large pathogenic deletion was reported in patients from Saudi Arabia (Al-Muhaizea et al. 2009).

The CLN6 gene encodes a membrane protein localized in the endoplasmic reticulum  (Sharp et al. 2003; Heine et al. 2007). It is postulated that the encoded protein mediates the transport of proteins or lipids that are essential for lysosomal function and acidification (Mole et al. 2004). However, its precise function is unknown at this time.

Testing Strategy

This test involves bidirectional DNA Sanger sequencing of all coding exons and splice sites of CLN6. The full coding sequence of each exon plus ~ 20 bp of flanking DNA on either side are sequenced. We will also sequence any single exon (Test #100) or pair of exons (Test #200) in family members of patients with known mutations or to confirm research results.

Indications for Test

Candidates for this test are patients with a clinical diagnosis suggestive of neuronal ceroid lipofuscinosis, regardless to the age of onset of symptoms.

Gene

Official Gene Symbol OMIM ID
CLN6 606725
Inheritance Abbreviation
Autosomal Dominant AD
Autosomal Recessive AR
X-Linked XL
Mitochondrial MT

Related Tests

Name
Early Infantile Epileptic Encephalopathy, Recessive Sequencing Panel
Neuronal Ceroid Lipofuscinoses (Batten Disease) Sequencing Panel

CONTACTS

Genetic Counselors
Geneticist
Citations
  • Al-Muhaizea MA, Al-Hassnan ZN, Chedrawi A. 2009. Variant Late Infantile Neuronal Ceroid Lipofuscinosis (CLN6 Gene) in Saudi Arabia. Pediatric Neurology 41: 74–76. PubMed ID: 19520283
  • Bennett MJ, Rakheja D. 2013. The neuronal ceroid-lipofuscinoses: The Neuronal Ceroid-Lipofuscinoses. Developmental Disabilities Research Reviews 17: 254–259. PubMed ID: 23798013
  • Canafoglia L, Gilioli I, Invernizzi F, Sofia V, Fugnanesi V, Morbin M, Chiapparini L, Granata T, Binelli S, Scaioli V, Garavaglia B, Nardocci N, Berkovic SF, Franceschetti S. 2015. Electroclinical spectrum of the neuronal ceroid lipofuscinoses associated with CLN6 mutations. Neurology 10.1212/WNL.0000000000001784. PubMed ID: 26115733
  • Dyken PR, Opitz JM, Reynolds JF, Pullarkat RK. 1988. Reconsideration of the classification of the neuronal ceroid-lipofuscinoses. American Journal of Medical Genetics 31: 69–84. PubMed ID: 3146331
  • Gao H, Boustany R-MN, Espinola JA, Cotman SL, Srinidhi L, Antonellis KA, Gillis T, Qin X, Liu S, Donahue LR, Bronson RT, Faust JR, Stout D, Haines JL, Lerner TJ, MacDonald ME. 2002. Mutations in a Novel CLN6-Encoded Transmembrane Protein Cause Variant Neuronal Ceroid Lipofuscinosis in Man and Mouse. Am J Hum Genet 70: 324–335. PubMed ID: 11791207
  • Heine C, Quitsch A, Storch S, Martin Y, Lonka L, Lehesjoki A-E, Mole SE, Braulke T. 2007. Topology and endoplasmic reticulum retention signals of the lysosomal storage disease-related membrane protein CLN6. Mol. Membr. Biol. 24: 74–87. PubMed ID: 17453415
  • Kousi M, Lehesjoki A-E, Mole SE. 2012. Update of the mutation spectrum and clinical correlations of over 360 mutations in eight genes that underlie the neuronal ceroid lipofuscinoses. Human Mutation 33: 42–63. PubMed ID: 21990111
  • Mole S.E., Williams R.E. 2013. Neuronal Ceroid-Lipofuscinoses. In: Pagon RA, Adam MP, Bird TD, Dolan CR, Fong C-T, and Stephens K, editors. GeneReviews™, Seattle (WA): University of Washington, Seattle. PubMed ID: 20301601
  • Mole SE, Michaux G, Codlin S, Wheeler RB, Sharp JD, Cutler DF. 2004. CLN6, which is associated with a lysosomal storage disease, is an endoplasmic reticulum protein. Exp. Cell Res. 298: 399–406. PubMed ID: 15265688
  • Mole SE, Williams RE, Goebel HH. 2005. Correlations between genotype, ultrastructural morphology and clinical phenotype in the neuronal ceroid lipofuscinoses. Neurogenetics 6: 107–126. PubMed ID: 15965709
  • Moore SJ, Buckley DJ, MacMillan A, Marshall HD, Steele L, Ray PN, Nawaz Z, Baskin B, Frecker M, Carr SM, Ives E, Parfrey PS. 2008. The clinical and genetic epidemiology of neuronal ceroid lipofuscinosis in Newfoundland. Clin. Genet. 74: 213–222. PubMed ID: 18684116
  • Rider J.A., Rider D.L. 1988. American journal of medical genetics. Supplement. 5: 21-6. PubMed ID: 3146319
  • Santorelli FM. et al. 2013. Orphanet journal of rare diseases. 8: 19. PubMed ID: 23374165
  • Schulz A. et al. 2004. Annals of neurology. 56: 342-50. PubMed ID: 15349861
  • Sharp JD, Wheeler RB, Parker KA, Gardiner RM, Williams RE, Mole SE. 2003. Spectrum of CLN6 mutations in variant late infantile neuronal ceroid lipofuscinosis. Hum. Mutat. 22: 35–42. PubMed ID: 12815591
  • Vesa J, Hellsten E, Verkruyse LA, Camp LA, Rapola J, Santavuori P, Hofmann SL, Peltonen L. 1995. Mutations in the palmitoyl protein thioesterase gene causing infantile neuronal ceroid lipofuscinosis. Nature 376:584-587. PubMed ID: 7637805
  • Wheeler RB, Sharp JD, Schultz RA, Joslin JM, Williams RE, Mole SE. 2002. The Gene Mutated in Variant Late-Infantile Neuronal Ceroid Lipofuscinosis (CLN6) and in nclf Mutant Mice Encodes a Novel Predicted Transmembrane Protein. Am J Hum Genet 70: 537–542. PubMed ID: 11727201
  • Williams R.E., Mole S.E. 2012. Neurology. 79: 183-91. PubMed ID: 22778232
Order Kits
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.

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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