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Autosomal Dominant Optic Atrophy (ADOA) or Optic Atrophy, Kjer-Type (OAK) and DOA Plus Syndrome (DOA+) via the OPA1 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
3189 OPA1$690.00 81407 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 molecular screening of 980 cases of suspected hereditary optic neuropathy, 440 patients had molecular defects. Among these 440 patients, 295 had OPA1 mutations (67%) 131 patients had (30%) mtDNA mutations, and 14 patients (3%) had OPA3 mutations (Ferre, M., et al. Hum Mutat 30(7):E692-705, 2009). In another study done with ADOA patients, 75% of the cases had OPA1 mutations, whereas 1% of patients had OPA3 mutations (Lenaers, G. et al. Orphanet J Rare Dis 7:46, 2012).

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Deletion/Duplication Testing via aCGH

Test Code Test Copy GenesIndividual Gene PriceCPT Code Copy CPT Codes
600 OPA1$690.00 81406 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

Autosomal Dominant Optic Atrophy (ADOA; OMIM# 165500) is the most prevalent inherited optic neuropathy besides Leber’s hereditary optic neuropathy (LHON). Both share a common pathological hallmark, the preferential loss of retinal ganglion cells (RGCs) (Carelli, V., et al. Biochim Biophys Acta 1787(5):518–528, 2009; Yu-Wai-Man, P., et al. Ophthalmology. 117(8):1538-1546, 2010). ADOA is clinically characterized by bilateral reduction in visual acuity that progresses insidiously from early childhood onwards (Yu-Wai-Man, P., et al. Ophthalmology. 118(3):558-563, 2011). Other symptoms include central or near central scotomas, tritanopia, variable degree of ptosis, central visual field defects and/or ophthalmalgia and optic nerve pallor. However, there is a marked inter-and intrafamilial variation in disease severity (Votruba, M., et al. Hum. Genet. 102(1): 79-86, 1998). Phenotype-genotype studies of optic atrophies found that 20% of DOA patients develop a more severe phenotype called “DOA plus” (DOA+, OMIM# 125250), which is characterized by extraocular multi-systemic features, including neurosensory hearing loss, or less commonly chronic progressive external ophthalmoplegia, myopathy, peripheral neuropathy, multiple sclerosis-like illness, spastic paraplegia or cataracts (Yu-Wai-Man, P., et al. 2010; Amati-Bonneau, P. et al. Int J Biochem Cell Biol 41(10):1855-1865, 2009). Disease prevalence is estimated as 3/100,000 in most populations in the world, but in Denmark it can reach to 1/10,000 due to a founder effect (Kjer, B. et al. Acta Ophthalmol Scand 74(1):3-7, 1996; Thiselton, DL. et al. Hum Genet 109(5):498-502, 2001).

Genetics

Although heterogeneous, the majority of suspected hereditary optic neuropathy patients (>60%) harbor pathogenic mutations within OPA1; ~30% have mtDNA mutations; and ~3% have OPA3 mutations (Ferre et al. Hum Mutat 30(7): E692-705, 2009). OPA1 encodes mitochondrial dynamin-like GTPase, which localizes to the inner mitochondrial membrane and regulates several important cellular processes, which include the stability of the mitochondrial network, mitochondrial bioenergetic output, sequestration of proapoptotic cytochrome C oxidase molecules within the mitochondrial cristae spaces, control of apoptosis, maintenance of mitochondrial DNA and oxidative phosphorylation (Ferre et al., 2009; Yu-Wai-Man et al., 2010). OPA1 is composed of 30 coding exons that span ~90 kb of genomic DNA on chromosome 3q28-q29. Alternative splicing of exons 4, 4b, 5 and 5b leads to eight transcript isoforms shown to be expressed in a variety of tissues (Alavi, M.V. et al. Brain 130(Pt 4):1029-1042, 2007). Almost 50% of OPA1 mutations lead to haploinsufficiency due to complete deletion of the gene and nonsense mediated mRNA decay, which is believed to be the major pathomechanism underlying OPA1-associated ADOA (Pesch, U.E. et al. Hum Mol Genet 10(13):1359-1368, 2001). OPA1 genomic rearrangements account for ~13% cases of ADOA, suggesting that deletion and duplication analysis should be included in the routine genetic analysis of ADOA patients (Marchbank, N.J. et al. J Med Genet 39(8): e47, 2002 ; Fuhrmann, N. et al. J Med Genet 46(2):136-144, 2009 ; Almind, G.J. et al. BMC Med Genet 12:49, 2011). The haploinsufficiency has been shown to affect retinal ganglion cells, which have many mitochondria and especially high energy requirement due to their non-myelinated specialized extensions called axons that form the optic nerve. Based on Fuhrmann et al. (2009) genotyping results, ADOA penetrance was ~88%, which is consistent with previous reports (Toomes, C. et al. Hum Mol Genet 10(13):1369-1378, 2001; Cohn, A.C. et al. Am J Ophthalmol 143(4):656-662, 2007). Missense, nonsense, insertions, deletions and splicing mutations have been reported within OPA1 and are spread throughout the coding region, with most of them clustering over the cDNA region corresponding to the GTPase domain (Exons 8-16) and the 3' end of the coding region (exon 27-28) (Pesch, U.E. et al. 2001; Toomes, C. et al., 2001).

Testing Strategy

For this Next Generation (NextGen) test, the full coding regions plus ~20 bp of non-coding DNA flanking each exon are sequenced for the gene listed below. 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 any regions not captured or with insufficient number of sequence reads. All pathogenic, likely pathogenic, or variants of uncertain significance are confirmed by Sanger sequencing.

Indications for Test

Ideal OPA1 test candidates are ADOA, DOA+ patients and patients undergoing a diagnostic evaluation of suspected hereditary optic neuropathy or a family history of optic neuropathy. Testing should begin with an affected family member. In the familial cases, OPA1 should be tested first, whenever autosomal dominant inheritance is obvious. If no OPA1 mutation is found, OPA3 should be considered.

Gene

Official Gene Symbol OMIM ID
OPA1 605290
Inheritance Abbreviation
Autosomal Dominant AD
Autosomal Recessive AR
X-Linked XL
Mitochondrial MT

Related Tests

Name
Metabolic Myopathies, Rhabdomyolysis and Exercise Intolerance Sequencing Panel
Mitochondrial Genome Maintenance/Integrity Nuclear Genes Sequencing Panel
Optic Atrophy Sequencing Panel

CONTACTS

Genetic Counselors
Geneticist
Citations
  • Alavi, M.V. et al. (2007). PubMed ID: 17314202
  • Almind GJ, Grønskov K, Milea D, Larsen M, Brøndum-Nielsen K, Ek J. 2011. Genomic deletions in OPA1 in Danish patients with autosomal dominant optic atrophy. BMC medical genetics 12: 49. PubMed ID: 21457585
  • Amati-Bonneau P, Milea D, Bonneau D, Chevrollier A, Ferré M, Guillet V, Gueguen N, Loiseau D, Crescenzo M-AP de, Verny C, Procaccio V, Lenaers G, et al. 2009. OPA1-associated disorders: phenotypes and pathophysiology. Int. J. Biochem. Cell Biol. 41: 1855–1865. PubMed ID: 19389487
  • Carelli V, Morgia C La, Valentino ML, Barboni P, Ross-Cisneros FN, Sadun AA. 2009. Retinal ganglion cell neurodegeneration in mitochondrial inherited disorders. Biochimica et Biophysica Acta (BBA) - Bioenergetics 1787: 518–528. PubMed ID: 19268652
  • Cohn, A.C. et al. (2007). PubMed ID: 17306754
  • Ferré M, Bonneau D, Milea D, Chevrollier A, Verny C, Dollfus H, Ayuso C, Defoort S, Vignal C, Zanlonghi X, Charlin J-F, Kaplan J, et al. 2009. Molecular screening of 980 cases of suspected hereditary optic neuropathy with a report on 77 novel OPA1 mutations. Human Mutation 30: E692–E705. PubMed ID: 19319978
  • Fuhrmann, N. et al. (2009). PubMed ID: 19181907
  • Kjer B, Eiberg H, Kjer P, Rosenberg T. 1996. Dominant optic atrophy mapped to chromosome 3q region. II. Clinical and epidemiological aspects. Acta Ophthalmol Scand 74: 3–7. PubMed ID: 8689476
  • Lenaers G, Hamel C, Delettre C, Amati-Bonneau P, Procaccio V, Bonneau D, Reynier P, Milea D. 2012. Dominant optic atrophy. Orphanet J Rare Dis 7: 46–46. PubMed ID: 22776096
  • Marchbank, N.J. etal. (2002). PubMed ID: 12161614
  • Pesch, U.E. et al. (2001). PubMed ID: 11440988
  • Thiselton DL, Alexander C, Morris A, Brooks S, Rosenberg T, Eiberg H, Kjer B, Kjer P, Bhattacharya SS, Votruba M. 2001. A frameshift mutation in exon 28 of the OPA1 gene explains the high prevalence of dominant optic atrophy in the Danish population: evidence for a founder effect. Human genetics 109: 498–502. PubMed ID: 11735024
  • Toomes, C. et al. (2001). PubMed ID: 11440989
  • Votruba, M. et al. (1998). PubMed ID: 9490303
  • Yu-Wai-Man P, Griffiths PG, Burke A, Sellar PW, Clarke MP, Gnanaraj L, Ah-Kine D, Hudson G, Czermin B, Taylor RW, Horvath R, Chinnery PF. 2010. The Prevalence and Natural History of Dominant Optic Atrophy Due to OPA1 Mutations. Ophthalmology 117: 1538–1546.e1. PubMed ID: 20417570
  • Yu-Wai-Man P, Shankar SP, Biousse V, Miller NR, Bean LJH, Coffee B, Hegde M, Newman NJ. 2011. Genetic Screening for OPA1 and OPA3 Mutations in Patients with Suspected Inherited Optic Neuropathies. Ophthalmology 118: 558–563. PubMed ID: 21036400
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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.

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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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