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Autosomal Dominant Polycystic Kidney Disease Sequencing Panel

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

Sequencing

Test Code TestCPT Code Copy CPT Codes
1919 GANAB 81479 Add to Order
HNF1B 81405
PKD1 81407
PKD2 81406
Full Panel Price* $1990.00
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. If you would like to order a subset of these genes contact us to discuss pricing.

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

Given that documented large deletions and duplications account for less than 5% of all documented pathogenic variants in PKD1 and PKD2 (Human Gene Mutation Database; Rossetti et al. 2007; Audrézet et al. 2012), the detection rates of PKD1 and PKD2 pathogenic variants through sequencing are expected to be slightly lower than the reported overall detection rates, which are 75% for PKD1 and 15% for PKD2, respectively (Rossetti et al. 2007; Audrézet et al. 2012).

Defects in the newly reported third ADPKD causative gene GANAB account for another ~0.3% of total ADPKD (~3% of genetically unexplained ADPKD-affected families by PKD1 and PKD2 pathogenic variants) (Porath et al. 2016).

HNF1B defects explain approximately 1% of all MODY cases (McDonald et al. 2013). Considering that large deletions are common, the HNF1B mutation detection rate via sequencing is lower than 1% in MODY. HNF1B pathogenic variants were found via Sanger sequencing in up to 7% of patients/fetuses with renal hypodysplasia in three large cohort studies (Weber et al. 2006; Thomas et al. 2011; Madariaga et al. 2013).

Concurrent HNF1B and PKD1 pathogenic variants have been reported to have an aggravating effect on a patient's phenotypes and explain phenotypic variabilities within PKD-affected family members (Bergmann et al. 2011). However, this has been only reported in a limited number of cases and the prevalence of this effect is still unknown in a larger cohort of PKD patients.

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Deletion/Duplication Testing via aCGH

Test Code TestIndividual Gene PriceCPT Code Copy CPT Codes
600 HNF1B$690.00 81404 Add to Order
PKD1$540.00 81479
PKD2$690.00 81479
Full Panel Price* $960.00
Pricing Comment

PKD1 is analyzed by Multiplex Ligation-dependent Probe Amplification.

Turnaround Time

The great majority of tests are completed within 28 days.

Clinical Sensitivity

Large deletions and duplications are relatively rare in both PKD1 and PKD2, accounting for fewer than 5% of ADPKD patients (Ariyurek et al. 2004; Rossetti et al. 2007, Audrézet et al. 2012).

Large deletions and duplications are common in HNF1B (Raaijmakers et al. 2015).

GANAB is currently not available in our aCGH testing menu. No large deletions or duplications involving GANAB have been reported to date.

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Clinical Features

Autosomal dominant polycystic kidney disease (ADPKD) is a relatively common inherited kidney disease with multisystem involvement. ADPKD is characterized by bilateral renal cysts accompanied by cysts in other organs including the liver, seminal vesicles, pancreas, and arachnoid membrane (Harris et al. 2011). Renal symptoms include hypertension, renal pain, and renal insufficiency. Nearly half of all ADPKD patients have end-stage renal disease (ESRD) by age 60 years. The progressive growth of liver cysts is the most common extrarenal manifestation of ADPKD. The most important non-cystic manifestations of ADPKD are vascular and cardiac abnormalities including intracranial aneurysms, mitral valve prolapse, dilatation of the aortic root, dissection of the thoracic aorta, and abdominal wall hernias. The clinical spectrum of ADPKD is wide, and substantial variability of disease severity can occur even within the same family. Compared with PKD1-related ADPKD, patients with PKD2 defects have approximately 20 years longer renal survival (median age at onset of ESRD: 58 years vs 79 years) (Cornec-Le Gall et al. 2013). The phenotypes caused by defects in the GANAB gene are even milder than those of PKD2-associated patients (Porath et al. 2016). Sporadic cases account for about 10% of individuals with ADPKD in adulthood (Neumann et al. 2012).

Patients with ADPKD typically have onset of symptoms in adulthood. In some rare cases, however, patients with two causative PKD1 pathogenic variants in trans may have clinical features similar to those of patients with autosomal recessive polycystic kidney disease (caused by pathogenic variants in the PKHD1 gene) (Rossetti et al. 2009; Vujic et al. 2010). In these rare cases, symptoms may appear in early childhood or even in utero.

HNF1B defects alone cause renal cysts and diabetes syndrome (RCAD), also referred to as maturity-onset diabetes of the young type 5 (MODY5) (Horikawa et al. 1997; McDonald et al. 2013). In addition, HNF1B defects can cause phenotypes in the spectrum of congenital anomalies of the kidney and urinary tract (CAKUT) (Vivante et al. 2014). Concurrent HNF1B and PKD1 pathogenic variants have been reported to have an aggravating effect on a patient's phenotypes and explain phenotypic variabilities within PKD-affected family members (Bergmann et al. 2011).

Genetics

Pathogenic variants in PKD1 and PKD2 cause approximately 90% of ADPKD (Rossetti et al. 2007; Audrézet et al. 2012). Both genes encode members of the polycystin protein family, which together play an important role in renal tubular development. Defects in the newly reported third ADPKD causative gene GANAB account for another ~0.3% of total ADPKD (~3% of genetically unexplained ADPKD-affected families without PKD1 and PKD2 pathogenic variants) (Porath et al. 2016). The GANAB gene encodes glucosidase II subunit alpha, defects of which possibly results in disruption of the maturation of polycystin-1 (the PKD1-encoded protein).

Genetic defects in PKD1 (46 coding exons) and PKD2 (15 coding exons) explain approximately 85% and 15% of mutation-positive ADPKD cases, respectively (Rossetti et al. 2007; Audrézet et al. 2012). These pathogenic variants have been found across the whole coding region of both genes. Truncated variants (nonsense, typical splicing variants and frame-shifting small deletion/insertions) are the majority of PKD1 and PKD2 defects, although missense variants and small in-frame changes are also commonly found. Large deletions and duplications have been reported, but are relatively uncommon (Ariyurek et al. 2004; Rossetti et al. 2007; Audrézet et al. 2012). The majority of PKD1 and PKD2 defects were found in single patients (Audrézet et al. 2012). De novo pathogenic variants account for about 10% of individuals with ADPKD in adulthood (Neumann et al. 2012).

Genetic defects in GANAB (25 coding exons) reported to date include missense, nonsense, splicing variants, and small deletion/insertions (Human Gene Mutation Database; Porath et al. 2016). No large deletions or duplications involving GANAB have been reported tp date.

HNF1B-related diseases are inherited in an autosomal dominant manner. HNF1B has 9 coding exons that encode hepatocyte nuclear factor-1-beta (HNF1B), also known as transcription factor-2 (TCF2), which is a member of the homeodomain-containing superfamily of transcription factors and is an essential factor for embryogenesis of the kidney, pancreas, and liver. Genetic defects of HNF1B throughout the whole coding region include missense, nonsense, splicing variants, and small deletion/insertions. In addition, large deletions encompassing multiple exons or the whole HNF1B gene have been commonly reported (Human Gene Mutation Database; Bellanné-Chantelot et al. 2005). De novo HNF1B pathogenic variants are common, accounting for up to 50% of cases.

Testing Strategy

For this NextGen test, 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, likely pathogenic, or variants of uncertain significance are confirmed by Sanger sequencing. Of note, sequencing for exons 1-33 of PKD1 is performed via special Sanger sequencing due to close sequence similarity with other loci.

For deletion/duplication testing, PKD1 is analyzed by Multiplex Ligation-dependent Probe Amplification.

This panel provides 100% coverage of the aforementioned regions of the indicated genes. We define coverage as > 20X NGS reads for exons and 0-10 bases of flanking DNA, > 10X NGS reads for 11-20 bases of flanking DNA, or Sanger sequencing.

Indications for Test

Candidates for this test are patients with ADPKD. 

Genes

Official Gene Symbol OMIM ID
GANAB 104160
HNF1B 189907
PKD1 601313
PKD2 173910
Inheritance Abbreviation
Autosomal Dominant AD
Autosomal Recessive AR
X-Linked XL
Mitochondrial MT

Related Tests

Name
Autosomal Dominant Polycystic Kidney Disease via the PKD1 Gene
Autosomal Dominant Polycystic Kidney Disease via the PKD1 Gene
Autosomal Dominant Polycystic Kidney Disease via the PKD2 Gene
Congenital Abnormalities of the Kidney and Urinary Tract (CAKUT) Sequencing Panel
Hypomagnesemia Sequencing Panel
Maturity Onset Diabetes of the Young (MODY) Sequencing Panel
Renal Cysts and Diabetes Syndrome via the HNF1B Gene

CONTACTS

Genetic Counselors
Geneticist
Citations
  • Ariyurek Y. et al. 2004. Human Mutation. 23: 99. PubMed ID: 14695542
  • Audrézet M.P. et al. 2012. Human Mutation. 33: 1239-50. PubMed ID: 22508176
  • Bellanné-Chantelot C. et al. 2005. Diabetes. 54: 3126-32. PubMed ID: 16249435
  • Bergmann C. et al. 2011. Journal of the American Society of Nephrology. 22: 2047-56. PubMed ID: 22034641
  • Cornec-Le Gall E. et al. 2013. Journal of the American Society of Nephrology. 24: 1006-13. PubMed ID: 23431072
  • Harris P.C, Torres V.E. 2011. Polycystic Kidney Disease, Autosomal Dominant. 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: 20301424
  • Horikawa Y. et al. 1997. Nature Genetics. 17: 384-5. PubMed ID: 9398836
  • Human Gene Mutation Database (HGMD).
  • McDonald T.J., Ellard S. 2013. Annals of Clinical Biochemistry. 50: 403-15. PubMed ID: 23878349
  • Neumann H.P. et al. 2012. International Urology and Nephrology. 44: 1753-62. PubMed ID: 22367170
  • Porath B. et al. 2016. American Journal of Human Genetics. 98: 1193-207. PubMed ID: 27259053
  • Raaijmakers A. et al. 2015. Nephrology, Dialysis, Transplantation. 30: 835-42. PubMed ID: 25500806
  • Rossetti S. et al. 2007. Journal of the American Society of Nephrology. 18: 2143-60. PubMed ID: 17582161
  • Rossetti S. et al. 2009. Kidney International. 75: 848-55. PubMed ID: 19165178
  • Vivante A. et al. 2014. Pediatric Nephrology. 29: 695-704. PubMed ID: 24398540
  • Vujic M. et al. 2010. Journal of the American Society of Nephrology : Jasn. 21: 1097-102. PubMed ID: 20558538
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TEST METHODS

NextGen Sequencing

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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