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Chronic Pancreatitis Sequencing Panel

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

NGS Sequencing

Test Code Test Copy GenesCPT Code Copy CPT Codes
1395 CASR 81405 Add to Order
CFTR 81223
CTRC 81479
PRSS1 81404
SPINK1 81404
Full Panel Price* $1790.00
Test Code Test Copy Genes Total Price CPT Codes Copy CPT Codes
1395 Genes x (5) $1790.00 81223, 81404(x2), 81405, 81479 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. 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

About 48% of idiopathic chronic pancreatitis patients were found to display evidence of a genetic basis for their pancreatitis and carried at least one abnormal allele in one of the four genes, PRSS1, SPINK1, CFTR or CTRC (Masson et al. 2013).

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Deletion/Duplication Testing via aCGH

Test Code Test Copy GenesIndividual Gene PriceCPT Code Copy CPT Codes
600 CASR$690.00 81479 Add to Order
CFTR$690.00 81222
CTRC$690.00 81479
PRSS1$690.00 81479
SPINK1$690.00 81479
Full Panel Price* $840.00
Test Code Test Copy Genes Total Price CPT Codes Copy CPT Codes
600 Genes x (5) $840.00 81222, 81479(x4) 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 Sensitivity

Though rare, complete gene duplications have been reported for PRSS1-associated HP (Masson et al. 2008; Le Maréchal et al. 2006). To date, no gross duplications or deletions have been reported for CFTR-associated pancreatitis (Human Gene Mutation Database).

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Clinical Features

Pancreatitis is characterized by recurrent episodes of inflammation of the pancreas in both adults and children (Chen and Ferec 2009). Symptoms usually begin in late childhood with an episode of acute pancreatitis and include severe upper abdominal burning pain radiating to the back, nausea, and vomiting that is worsened with eating (acute pancreatitis). Recurrent acute pancreatitis leads to chronic pancreatitis due to persistent inflammation. Chronic pancreatitis usually develops by early adulthood in affected individuals and symptoms include occasional or frequent abdominal pain of varying severity, flatulence, and bloating. Unexplained weight loss may occur from a lack of pancreatic enzymes hindering digestion (Rebours et al. 2012). Episodes of pancreatitis can lead to permanent tissue damage and loss of pancreatic function. Chronic pancreatitis increases the risk for diabetes and pancreatic cancer, more so with smoking and use of alcohol (Yadav and Whitcomb 2010). Mutations in several genes have been identified to be causative for, or induce susceptibility to pancreatitis (Joergensen et al. 2010). Genetic testing can aid in differential diagnosis of chronic pancreatitis from other disorders such as Shwachman-Diamond syndrome and Johanson-Blizzard syndrome which also present with pancreatitis (LaRusch et al. 2014).

Genetics

Hereditary chronic pancreatitis (HP) occurs at an estimated incidence of 0.3/100,000 in western countries (Joergensen et al. 2010) and is inherited in an autosomal dominant manner through mutations in the PRSS1 gene in ~60% of cases. About 30% of HP results from inheritance with reduced penetrance through mutations in the SPINK1, CFTR, CTRC, and CASR genes (Masson et al. 2013; LaRusch et al. 2014).

PRSS1 encodes trypsinogen which becomes converted to active trypsin after secretion into the duodenum to aid in digestion. About 60% of HP cases are inherited in an autosomal dominant manner through mutations in the PRSS1 gene. Pathogenic mutations typically result in premature conversion of trypsinogen or resistance to degradation. Duplication in the PRSS1 gene resulting in elevated trypsinogen levels are also a reported cause to HP (Chen and Ferec et al. 2009; LaRusch et al. 2014). SPINK1 encodes a pancreatic secretory trypsin inhibitor which protects the pancreas from damage due to recurrent or persistent trypsin activation (Horii et al. 1987). HP can be inherited in an autosomal recessive manner through SPINK1 mutations or more commonly through a possible digenic mode together with mutations in the CFTR and CTRC genes. Splice site alterations, gross deletions, and frameshift mutations are considered severe alleles with the highest penetrance in the SPINK1 gene. However, a moderate penetrant missense mutation, c.101A>G (p.Asn34Ser), is most commonly found within SPINK1 in European and North American populations. The c.194+2T>C variant is commonly found in Asian populations (LaRusch et al. 2014). CFTR encodes a transmembrane conductance protein which functions to secrete bicarbonate fluid to flush pancreatic zymogens into the duodenum. HP can be inherited through an autosomal recessive manner through CFTR mutations but more commonly through a more complex mode together with mutations in the SPINK1 or CASR genes. CFTR mutations have been reported in the presence of PRSS1 mutations in a minor subset of HP patients (Masson et al. 2013). CTRC encodes chymotrypsin C which degrades prematurely activated trypsinogen within the pancreas. To date, CTRC mutations have primarily been reported in HP patients with co-occurring CFTR mutations, but may co-occur with SPINK1 mutations. Loss of function mutations are primarily found in the CTRC gene in patients with HP (Masson et al. 2013). CASR encodes a calcium sensing receptor that maintains calcium homeostasis. CASR mutation are low penetrant and are only found in HP patients together with CFTR or SPINK1 mutations (Whitcomb et al. 2013 Felderbauer et al. 2006).

For further information on PRSS1, SPINK1, CFTR, CTRC, or CASR genes please refer to the individual test descriptions.

Testing Strategy

For this Next Generation (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, likely pathogenic, or variants of uncertain significance are confirmed by Sanger sequencing. This test also includes sequencing of CFTR regions within intron 11 and intron 21, along with a complete analysis of the compound (TG)n(T)n sequence (5T/TG tract) in intron 8. Variations in the 5T/TG tract have been reported to cause CBAVD (Chillon et al. 1995). Our 5T/TG tract analysis involves bidirectional sequencing along with allele length measurement. 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

Individuals with the following criteria should consider genetic testing (Ellis et al. 2001): recurrent unexplained attacks of acute pancreatitis and a positive family history, unexplained chronic pancreatitis and a positive family history, unexplained chronic pancreatitis without a positive family history after exclusion of other causes such as hyperlipidaemia type I, familiar hypercalciuric hypercalcemia (FBH), hereditary hyperthyroidism and autoimmune pancreatitis, and unexplained pancreatitis episodes in children.

Genes

Official Gene Symbol OMIM ID
CASR 601199
CFTR 602421
CTRC 601405
PRSS1 276000
SPINK1 167790
Inheritance Abbreviation
Autosomal Dominant AD
Autosomal Recessive AR
X-Linked XL
Mitochondrial MT

Disease

Name Inheritance OMIM ID
Pancreatitis, Chronic 167800

Related Tests

Name
Chronic Hereditary Pancreatitis via the PRSS1 Gene
Chronic Hereditary Pancreatitis via the SPINK1 Gene
Chronic Pancreatitis via the Chymotrypsin C (CTRC) Gene
Ciliopathy Sequencing Panel
Cystic Fibrosis and CF-Related Disorders via the CFTR Gene
Familial Hypocalciuric Hypercalcemia (FHH) Sequencing Panel
Hypercalcemic and Hypocalcemic Disorders via the CASR Gene
Hypomagnesemia Sequencing Panel
Hypoparathyroidism Sequencing Panel
Nephrolithiasis and Nephrocalcinosis Sequencing Panel

CONTACTS

Genetic Counselors
Geneticist
Citations
  • Chen J-M, Férec C. 2009. Chronic pancreatitis: genetics and pathogenesis. Annu Rev Genomics Hum Genet 10: 63–87. PubMed ID: 19453252
  • Chillon, M, Casals, T, Mercier, B, Bassas, L, Lissens, W, Silber, S, Romey, MC, Ruiz-Romero, J, Verlingue, C, Claustres, M, et al. 1995. Mutations in the cystic fibrosis gene in patients with congenital absence of the vas deferens. N. Engl. J. Med. 332: 1475-1480. PubMed ID: 7739684
  • Ellis, I, Lerch, MM, Whitcomb, DC. 2001. Genetic testing for hereditary pancreatitis: guidelines for indications, counselling, consent and privacy issues. Pancreatology 1:405-415. PubMed ID: 12120217
  • Felderbauer P, Klein W, Bulut K, Ansorge N, Dekomien G, Werner I, Epplen JT, Schmitz F, Schmidt WE. 2006. Mutations in the calcium-sensing receptor: a new genetic risk factor for chronic pancreatitis? Scand. J. Gastroenterol. 41: 343–348. PubMed ID: 16497624
  • Horii A, Kobayashi T, Tomita N, Yamamoto T, Fukushige S, Murotsu T, Ogawa M, Mori T, Matsubara K. 1987. Primary structure of human pancreatic secretory trypsin inhibitor (PSTI) gene. Biochem. Biophys. Res. Commun. 149: 635–641. PubMed ID: 3501289
  • Human Gene Mutation Database (Bio-base).
  • Joergensen, MT, Brusgaard, K, Crüger, DG, Gerdes, AM, Schaffalitzky de Muckadell, OB. 2010. Genetic, epidemiological, and clinical aspects of hereditary pancreatitis: a population-based cohort study in Denmark. Am. J. Gastroenterol. 105:1876-1883. PubMed ID: 20502448
  • LaRusch J, Solomon S, Whitcomb DC. 2014. Pancreatitis Overview. In: Pagon RA, Adam MP, Ardinger HH, Bird TD, Dolan CR, Fong C-T, Smith RJ, and Stephens K, editors. GeneReviews(®), Seattle (WA): University of Washington, Seattle. PubMed ID: 24624459
  • Le Maréchal C, Masson E, Chen J-M, Morel F, Ruszniewski P, Levy P, Férec C. 2006. Hereditary pancreatitis caused by triplication of the trypsinogen locus. Nat. Genet. 38: 1372–1374. PubMed ID: 17072318
  • Masson E, Maréchal C Le, Chandak GR, Lamoril J, Bezieau S, Mahurkar S, Bhaskar S, Reddy DN, Chen J-M, Férec C. 2008. Trypsinogen copy number mutations in patients with idiopathic chronic pancreatitis. Clin. Gastroenterol. Hepatol. 6: 82–88. PubMed ID: 18063422
  • Masson, E, Chen, JM, Audrézet, MP, Cooper, DN, Férec, C. 2013. A Conservative Assessment of the Major Genetic Causes of Idiopathic Chronic Pancreatitis: Data from a Comprehensive Analysis of PRSS1, SPINK1, CTRC and CFTR Genes in 253 Young French Patients. PLoS One 8:e73522. PubMed ID: 23951356
  • Rebours V, Lévy P, Ruszniewski P. 2012. An overview of hereditary pancreatitis. Dig Liver Dis 44: 8–15. PubMed ID: 21907651
  • Whitcomb DC. 2013. Genetic Risk Factors for Pancreatic Disorders. Gastroenterology 144: 1292–1302. PubMed ID: 23622139
  • Yadav D, Whitcomb DC. 2010. The role of alcohol and smoking in pancreatitis. Nat Rev Gastroenterol Hepatol 7: 131–145. PubMed ID: 20125091
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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.

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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