Melanoma Predisposition via the CDKN2A Gene

  • Summary and Pricing
  • Clinical Features and Genetics
  • Citations
  • Methods
  • Ordering/Specimens
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Test Code Test Copy GenesIndividual Gene PriceCPT Code Copy CPT Codes
1168 CDKN2A$540.00 81404 Add to Order
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 18 days.

Clinical Sensitivity

It has been reported that 25% to 50% of familial melanoma kindreds are affected by a CDKN2A causative mutation (Goldstein et al. J Med Genet 44(2):99-106, 2007). Individuals with multiple primary melanomas have a 1-3% chance of having a CDKN2A causative mutation (Berwick et al. Cancer Epidemiol Biomarkers Prev 15:1520-5, 2006).

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Deletion/Duplication Testing via aCGH

Test Code Test Copy GenesIndividual Gene PriceCPT Code Copy CPT Codes
600 CDKN2A$690.00 81479 Add to Order
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Turnaround Time

The great majority of tests are completed within 28 days.

Clinical Features

Melanoma is a malignant tumor that originated in melanocytes, a specialized cell type that produces melanin pigments that determine skin, hair and eye color (Lin and Fisher. Nature 445:843–850, 2007). Over the past few decades there has been a rise in the incidence of melanoma due both to improved awareness leading to additional diagnoses and to lifestyle changes that have resulted in an increase in sun exposure  (Mehnert and Kluger. Curr Oncol Rep 14:449–457, 2012). Most melanomas occur as sporadic cases with no recognized familial component; however melanoma has been reported to be twice as common in persons with an affected parent, three times as common if a sibling is affected, and nine times as common if both a parent and a sibling are affected (Hemminki et al. J Invest Dermatol 120:217–223, 2003). Familial clustering is likely the result of genetic and environmental factors. Heritable alleles for melanoma susceptibility range from high-risk, high-penetrance alleles that are rare, to low-risk, low-penetrance alleles that are common (Nelson et al. Clinics in Dermatology 27, 46–52, 2009). Mutations in the highly penetrant gene CDKN2A, and less frequently the CDK4 gene, are responsible for the majority of predisposition to melanoma cases. Individuals with genetic predisposition to melanoma have an earlier age of onset. For example, the median age at melanoma diagnosis is significantly lower in carriers of CDKN2A mutations (36 years) than in patients from families with wildtype CDKN2A (45 years) (Goldstein et al. Cancer Res 66:9818–9828, 2006). A family history of melanoma and pancreatic cancer may also suggest inherited mutations in CDKN2A (Goldstein et al. J Natl Cancer Inst 92(12):1006-10, 2000).


Melanoma predisposition is inherited in an autosomal dominant manner. The strongest genetic risk for the development of melanoma results from heritable alterations in the cyclin-dependent kinase inhibitor 2A (CDKN2A) gene, which encodes two separate but related proteins, p16/INK4a and p14/ARF, by using two different promoters. The CDKN2A gene is a tumor suppressor, and its protein products help regulate cell division and apoptosis (Nelson et al., 2009). The p16/INK4a protein is produced from a transcript generated from exons 1α, 2 and 3, whereas p14ARF is produced, using an alternative reading frame, from a transcript comprising exons 1β, 2 and 3 (Lin and Fisher, 2007). The function of the p16/INK4a protein is to inhibit CDK4/6 protein-mediated phosphorylation of the Rb (Retinoblastoma) tumor suppressor protein; dephosphorylated Rb is the active state.  Mutated p16/INK4a leads to phosphorylation of Rb, which in turn results in release of the bound transcription factor E2F, which then allows the cell to undergo unregulated cell division leading to the development of melanoma.

p14/ARF exerts its regulation of cell division through its indirect interaction with the p53 protein. p14/ARF binds to and inhibits the HDM2 protein. HDM2 functions to ubiquitinate proteins and target them for degradation, Mutations in p14/ARF abrogate binding to HDM2. As a result, HDM2 is released and increases ubiquitination of p53 leading to increased destruction of this tumor suppressor.  The main functions of p53 are to sense genetic damage to allow pause for DNA repair and to activate cellular apoptosis. Thus, the decreased levels of p53 associated with mutations in p14/ARF lead to genetic instability and to a higher risk of melanoma in individuals with CDKN2A mutations (Lin and Fisher, 2007). Families with mutated p14/ARF proteins also have an increase in neural system tumors in addition to melanoma (Nelson et al., 2009).

CDKN2A causative mutations reported to date include mostly missense mutations, however nonsense, splicing, small insertions and deletions, regulatory, and gross insertions and deletions have also been reported (Human Gene Mutation Database).

Testing Strategy

The cyclin-dependent kinase inhibitor 2A proteins are encoded by 3 exons (1-3) from the CDKN2A gene on chromosome 9p21.3. The two different proteins p16INK4a and p14 ARF are expressed from two distinct first exons of the CDKN2A gene which contain separate translational start codons.  Testing is accomplished by amplifying each coding exon, including the distinct first exons, and ~20 bp of adjacent noncoding sequence, then determining the nucleotide sequence using standard Sanger dideoxy sequencing methods and a capillary electrophoresis instrument. We will also sequence any single exon (Test #100) in family members of patients with a known mutation or to confirm research results.

Indications for Test

Candidates for this test are Individuals who have multiple family members with melanoma. Some members of the melanoma families may have other forms of cancer, particularly pancreatic cancer (Goldstein et al., 2000).  In such families, it is best to test an affected individual first.  Other candidates are patients with multiple primary melanomas.  This test is specifically designed for heritable germline mutations and is not appropriate for the detection of somatic mutations in tumor tissue.


Official Gene Symbol OMIM ID
CDKN2A 600160
Inheritance Abbreviation
Autosomal Dominant AD
Autosomal Recessive AR
X-Linked XL
Mitochondrial MT

Related Tests

Cancer Sequencing and Deletion/Duplication Panel
Pancreatic Cancer Sequencing Panel


Genetic Counselors
  • Berwick M. 2006. The Prevalence of CDKN2A Germ-Line Mutations and Relative Risk for Cutaneous Malignant Melanoma: An International Population-Based Study. Cancer Epidemiology Biomarkers & Prevention 15: 1520–1525. PubMed ID: 16896043
  • Goldstein AM, Chan M, Harland M, Hayward NK, Demenais F, Timothy Bishop D, Azizi E, Bergman W, Bianchi-Scarra G, Bruno W, Calista D, Cannon Albright LA, et al. 2006. Features associated with germline CDKN2A mutations: a GenoMEL study of melanoma-prone families from three continents. Journal of Medical Genetics 44: 99–106. PubMed ID: 16905682
  • Goldstein et al. (2000) "Genotype-phenotype relationships in U.S. melanoma-prone families with CDKN2A and CDK4 mutations." J Natl Cancer Inst 92(12):1006-10. PubMed ID: 10861313
  • Goldstein et al. (2006). "High-risk melanoma susceptibility genes and pancreatic cancer, neural system tumors, and uveal melanoma across GenoMEL." Cancer Res 66:9818–9828. PubMed ID: 17047042
  • Hemminki et al. (2003). "Familial and attributable risks in cutaneous melanoma: effects of proband and age." J Invest Dermatol 120:217–223. PubMed ID: 12542525
  • Human Gene Mutation Database (Bio-base).
  • Lin JY, Fisher DE. 2007. Melanocyte biology and skin pigmentation. Nature 445: 843–850. PubMed ID: 17314970
  • Mehnert and Kluger. (2012). "Driver mutations in melanoma: lessons learned from bench-to-bedside studies." Curr Oncol Rep 14:449–457. PubMed ID: 22723080
  • Nelson AA, Tsao H. 2009. Melanoma and genetics. Clinics in Dermatology 27: 46–52. PubMed ID: 19095153
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Bi-Directional Sanger Sequencing

Test Procedure

Nomenclature for sequence variants was from the Human Genome Variation Society (  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.

Analytical Validity

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).

Analytical Limitations

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.

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.
  • 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.
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