Pontocerebellar Hypoplasia Type 1B via the EXOSC3 Gene
- Summary and Pricing
- Clinical Features and Genetics
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The great majority of tests are completed within 18 days.
Pathogenic variants in the EXOSC3 gene were detected in 60% of patients with a clinical diagnosis of PCH1 (Wu et al. 2012).
Pontocerebellar hypoplasias (PCH) are a group of clinically and genetically heterogeneous neurodegenerative disorders characterized by abnormal development of the pons, cerebellum and cerebral cortex; progressive microcephaly; psychomotor developmental delay; and swallowing difficulties (Barth 1993; Namavar et al. 2011). Eight main types have been described (PCH1-8), based on the clinical presentation, progression, and pathological and molecular defects. PCH1, previously known as Norman’s disease, is distinguished by a loss of motor neurons in the anterior horn of the spinal cord similar to that observed in spinal muscular atrophy (SMA), and resulting in sensory and motor neuropathy. Additional features include severe hypotonia, fasciculation, ataxia, dysplasia, joint contractures, visual abnormalities and hyperventilation. Symptoms are usually apparent at birth, and death usually occurs within the first year of life. However, survival into childhood has been described. Based on the clinical features and electrophysiological and muscle biopsy findings, a number of patients with PCH1 were originally diagnosed with infantile SMA-PCH (Norman 1961; Rudnik-Schöneborn et al. 2003; Renbaum et al. 2009).
PCH1 is further divided into PCH1A and PCH1B based on the causative gene.
PCH1B is an autosomal recessive disorder. Pathogenic variants in the EXOSC3 gene result in PCH1B (Wan et al. 2012). A total of ten pathogenic variants have been reported to date. Half of these are missense; the other half consist of truncating variants and include one splicing and four small deletions or insertions. Defects in EXOSC3 were detected in patients of several ethnic and geographic origins such as American/European, Canadian/Cuban, German/Turkish, Spanish, Czech and Japanese (Wan et al. 2012; Ryan et al. 2000; Salman et al. 2003). They appear to be an important cause of PCH1. The current data suggest that patients with PCH1 and pathogenic variants in EXOSC3 have a milder phenotype and variable life span, ranging from a few months of age to late teens, compared to that observed in patients with no EXOSC3 pathogenic variants (Wan et al. 2012; Rudnik-Schöneborn et al. 2013).
The EXOSC3 gene encodes a core component of the human exosome complex, which is involved in RNA processing and degradation.
This test involves bidirectional Sanger DNA sequencing of all coding exons and splice sites of the EXOSC3 gene. 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 combination of pontocerebellar hypoplasia and infantile spinal muscular atrophy, a family history consistent with autosomal recessive mode of inheritance, and family members of patients who have known EXOSC3 pathogenic variants.
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- Genetic Counselor Team - email@example.com
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- Barth PG. 1993. Pontocerebellar hypoplasias. An overview of a group of inherited neurodegenerative disorders with fetal onset. Brain Dev 15:411-422. Review. PubMed ID: 8147499
- Namavar Y, Barth PG, Poll-The BT, Baas F. 2011. Classification, diagnosis and potential mechanisms in pontocerebellar hypoplasia. Orphanet J Rare Dis 6:50. Review. PubMed ID: 21749694
- Norman RM. 1961. Cerebellar hypoplasia in Werdnig-Hoffmann disease. Arch Dis Child 36:96-101. PubMed ID: 13729575
- Renbaum P, Kellerman E, Jaron R, Geiger D, Segel R, Lee M, King MC, Levy-Lahad E. 2009. Spinal muscular atrophy with pontocerebellar hypoplasia is caused by a mutation in the VRK1 gene. Am J Hum Genet 85:281-289. PubMed ID: 19646678
- Rudnik-Schöneborn S, Senderek J, Jen JC, Houge G, Seeman P, Puchmajerová A,Graul-Neumann L, Seidel U, Korinthenberg R, Kirschner J, Seeger J, Ryan MM, Muntoni F, Steinlin M, Sztriha L, Colomer J, Hübner C, Brockmann K, Van Maldergem L, Schiff M, Holzinger A, Barth P, Reardon W, Yourshaw M, Nelson SF, Eggermann T, Zerres K. 2013. Pontocerebellar hypoplasia type 1: clinical spectrum and relevance of EXOSC3 mutations. Neurology 80:438-446. PubMed ID: 23284067
- Rudnik-Schöneborn S, Sztriha L, Aithala GR, Houge G, Laegreid LM, Seeger J, Huppke M, Wirth B, Zerres K. 2003. Extended phenotype of pontocerebellar hypoplasia with infantile spinal muscular atrophy. Am J Med Genet A. 15;117A:10-17. PubMed ID: 12548734
- Ryan MM, Cooke-Yarborough CM, Procopis PG, Ouvrier RA. 2000. Anterior horn cell disease and olivopontocerebellar hypoplasia. Pediatr Neurol. 23:180-184. PubMed ID: 11020648
- Salman MS, Blaser S, Buncic JR, Westall CA, Héon E, Becker L. 2003. Pontocerebellar hypoplasia type 1: new leads for an earlier diagnosis. J Child Neurol.18:220-225. PubMed ID: 12731647
- Wan J, Yourshaw M, Mamsa H, Rudnik-Schöneborn S, Menezes MP, Hong JE, Leong DW, Senderek J, Salman MS, Chitayat D, Seeman P, von Moers A, Graul-Neumann L, Kornberg AJ, Castro-Gago M, Sobrido MJ, Sanefuji M, Shieh PB, Salamon N, Kim RC, Vinters HV, Chen Z, Zerres K, Ryan MM, Nelson SF, Jen JC. 2012. Mutations in the RNA exosome component gene EXOSC3 cause pontocerebellar hypoplasia and spinal motor neuron degeneration. Nat Genet 44:704-708. PubMed ID: 22544365
- Wu CH, Fallini C, Ticozzi N, Keagle PJ, Sapp PC, Piotrowska K, Lowe P, Koppers M, McKenna-Yasek D, Baron DM, Kost JE, Gonzalez-Perez P, Fox AD, Adams J, Taroni F, Tiloca C, Leclerc AL, Chafe SC, Mangroo D, Moore MJ, Zitzewitz JA, Xu ZS, van den Berg LH, Glass JD, Siciliano G, Cirulli ET, Goldstein DB, Salachas F, Meininger V, Rossoll W, Ratti A, Gellera C, Bosco DA, Bassell GJ, Silani V, Drory VE, Brown RH Jr, Landers JE. 2012. Mutations in the profilin 1 gene cause familial amyotrophic lateral sclerosis. Nature 488:499-503. PubMed ID: 22801503
Bi-Directional Sanger Sequencing
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.
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).
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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- The test can be added to your online orders in the Summary and Pricing section.
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- 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.
(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.
(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.
(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.