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Alzheimer Disease, Familial, Sequencing Panel

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

NGS Sequencing

Test Code Test Copy GenesCPT Code Copy CPT Codes
3409 APP 81406 Add to Order
PSEN1 81405
PSEN2 81406
Full Panel Price* $990.00
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

Pathogenic variants in PSEN1 gene accounts for 30%-70% of early onset familial Alzheimer disease while pathogenic variants in the APP gene are identified in ~15% of familial Alzheimer disease cases. Fewer pathogenic variants are found in PSEN2 gene which account for about 5% of all early onset familial Alzheimer disease (Marcon et al. 2009; Campion et al 1999; Wallon et al. 2012; Janssen et al. 2003).

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Deletion/Duplication Testing via aCGH

Test Code Test Copy GenesIndividual Gene PriceCPT Code Copy CPT Codes
600 APP$690.00 81479 Add to Order
PSEN1$690.00 81479
PSEN2$690.00 81479
Full Panel Price* $770.00
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

In cohorts of autosomal dominant early-onset Alzheimer disease, the clinical sensitivity of the APP locus duplications can be roughly estimated to be 8% (Rovelet-Lecrux A. et al. 2006). Large deletions in PSEN1 in a large cohort of patients with autosomal dominant early-onset Alzheimer disease is unavailable in the literature because large deletions have only been reported in individual cases. No large deletions/duplications in PSEN2 have been reported.

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Clinical Features

Familial Alzheimer disease is a neurodegenerative disorder characterized by onset of dementia at a relatively young age. Dementia initially presents in familial Alzheimer disease patients as short term memory problems or disorientation between 30 and 60 years of age. Cognitive decline is observed over the next 10-20 years with apraxia, progressive memory loss and impaired spatial skills being common presentations (Wallon et al. 2012). Motor disturbances such as cerebellar ataxia and spastic paraparesis are also observed in a subset of patients. The key neuropathology of familial Alzheimer disease includes: amyloid plaques, neurofibrillary tangles, neuronal loss and brain atrophy (Wu et al. 2012). An important feature of the familial Alzheimer disease diagnosis is that an individual has at least one affected family member. Patients with familial Alzheimer disease caused by PSEN2 pathogenic variants typically show a later age of onset in the 50s or 60s as compared to onset in the 30s or 40s as seen in familial Alzheimer disease caused by APP or PSEN1 variants (Jayadev et al. 2010). In addition, PSEN2-related familial Alzheimer disease patients have a higher frequency of behavioral and psychotic symptoms of dementia, such as hallucinations or delusions (Canevelli et al. 2014).

Genetics

Familial Alzheimer disease is inherited in an autosomal dominant manner and can be caused by pathogenic variants in the APP, PSEN1 and PSEN2 genes. The reported APP pathogenic variants are missense, frameshift and small deletion variants mainly contained within exons 16 and 17 of the APP gene. These variants disrupt processing of APP into mature amyloid-beta protein (Janssen et al. 2003; Mullan et al. 1992; Tomiyama et al. 2008). Large duplications and large deletion encompassing the entire APP gene have also been identified in familial Alzheimer disease patients (McNaughton et al. 2012). A rare recessive pathogenic variant in the APP gene was reported to cause familial Alzheimer disease (Di Fede et al. 2009). APP encodes amyloid-beta precursor protein (APP). APP is post-translationally processed into two amyloid-beta isoforms: AB40 and AB42. The gamma-secretase complex that processes APP contains the proteins PSEN1 and PSEN2, which play a role in pathogenesis of familial Alzheimer disease (Sua'rez-Calvet et al. 2014). Most causative PSEN1 variants are missense variants distributed throughout the gene, though frameshifts, splice site variants and large deletions in PSEN1 have also been identified in familial Alzheimer disease patients (Rogaeva et al. 2001; Janssen et al. 2003). PSEN1 encodes the presenilin-1 (PS1) protein. PS1 is the catalytic subunit of the gamma-secretase complex which cleaves the Alzheimer's-associated alpha-beta precursor protein (APP). PSEN1 variants are believed to act in a dominant negative manner to interfere with wildtype PS1 activity and to cause Familial Alzheimer disease via impaired APP processing by gamma-secretase (Nornes et al. 2007; Heilig et al. 2013). The major causative PSEN2 variants are missense variants. Two variants, N141I and M239V, account for 74% of all PSEN2-related familial Alzheimer disease cases (Canevelli et al. 2014). No large deletions or duplications have been reported to date. PSEN2 encodes the presenilin-2 (PS2) protein. PS2 is a subunit of the gamma-secretase complex which cleaves the Alzheimer's-associated alpha-beta precursor protein (APP). Although PSEN2 knockout mice do not show amyloid-beta processing defects, pathogenic PSEN2 variants were shown to alter AB40:AB42 ratios in in vitro cell-based assays (Herreman et al. 1999; Walker et al. 2005).

Testing Strategy

For this NextGen 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 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 and undocumented variants are confirmed by Sanger sequencing.

Indications for Test

Candidates for this panel include patients with symptoms suspected for familial Alzheimer disease.

Genes

Official Gene Symbol OMIM ID
APP 104760
PSEN1 104311
PSEN2 600759
Inheritance Abbreviation
Autosomal Dominant AD
Autosomal Recessive AR
X-Linked XL
Mitochondrial MT

Related Tests

Name
Alzheimer Disease, Familial or Cerebral Amyloid Angiopathy via the APP Gene
Alzheimer's Disease, Familial via the APP Gene, Exons 16 and 17
Alzheimer's Disease, Familial via the PSEN1 Gene
Alzheimer's Disease, Familial via the PSEN2 Gene
Amyotrophic Lateral Sclerosis and Frontotemporal Dementia Sequencing Panel
Autism Spectrum Disorders and Intellectual Disability (ASD-ID) Comprehensive Sequencing Panel with CNV Detection
Dementia Sequencing Panel

CONTACTS

Genetic Counselors
Geneticist
Citations
  • Campion D. et al. 1999. American Journal of Human Genetics. 65: 664-70. PubMed ID: 10441572
  • Canevelli M. et al. 2014. Neuroscience & Biobehavioral Reviews 42: 170–9. PubMed ID: 24594196
  • Di Fede G. et al. 2009. Science. 323: 1473-7. PubMed ID: 19286555
  • Heilig E.A. et al. 2013. The Journal of Neuroscience. 33: 11606-17. PubMed ID: 23843529
  • Herreman A. et al. 1999. Proceedings of the National Academy of Sciences of the United States of America. 96: 11872-7. PubMed ID: 10518543
  • Janssen J.C. et al. 2003. Neurology. 60: 235-9. PubMed ID: 12552037
  • Jayadev S. et al. 2010. Brain 133: 1143–54. PubMed ID: 23952003
  • Marcon G. et al. 2009. Journal of Alzheimer's Disease. 16: 509-11. PubMed ID: 19276543
  • Mullan M. et al. 1992. Nature Genetics. 1: 345-7. PubMed ID: 1302033
  • Nornes S. et al. 2008. Human Molecular Genetics. 17: 402-12. PubMed ID: 17981814
  • Rogaeva E.A. et al. 2001. Neurology. 57: 621-5. PubMed ID: 11524469
  • Rovelet-Lecrux A. et al. 2006. Nature Genetics. 38: 24-6. PubMed ID: 16369530
  • Suárez-Calvet M. et al. 2014. Journal of Neurochemistry. 128: 330-9. PubMed ID: 24117942
  • Tomiyama T. et al. 2008. Annals of Neurology. 63: 377-87. PubMed ID: 18300294
  • Walker E.S. et al. 2005. Journal of Neurochemistry. 92: 294-301. PubMed ID: 15663477
  • Wallon D. et al. 2012. Journal of Alzheimer's Disease. 30: 847-56. PubMed ID: 22475797
  • Wu L. et al. 2012. The Canadian Journal of Neurological Sciences. 39: 436–445. PubMed ID: 22728850
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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