Melanoma Panel

Malignant melanoma, also known as cutaneous malignant melanoma, results from new and abnormal growth of melanocytes most frequently occurring in the skin, but can also occur in the eyes, ears, and gastrointestinal tract. Malignant melanoma is characterized by atypical or numerous nevi (moles) often present in non-sun exposed areas. These moles are often larger than other moles and display an abnormal shape or color. Melanomas can develop anywhere on the skin, but often start on the chest and back in men and on the legs in women. A major risk factor for melanoma is UV exposure, which damages the DNA of skin cells. Unlike common skin cancers which are associated with total cumulative UV exposure, melanomas are associated with intense intermittent exposure. Other risk factors include having a weakened immune system, being male, and being older. In the United States, there is an overall higher rate of melanoma in males after age 50, and the risk of melanoma is higher in women before age 50.

Germline variants in the CDKN2A gene have been known to be most associated with high risk susceptibility to malignant melanoma; however, germline variant frequencies in this gene among melanoma families is variable with 46% in French familial melanoma (Soufir et al. 1998. PubMed ID: 9425228), 18% in American familial melanoma (Soufir et al. 1998. PubMed ID: 9425228; Fitzgerald et al. 1996. PubMed ID: 8710906) and 8% in Swedish familial melanoma (Platz et al. 1997. PubMed ID: 9168184; Platz et al. 1998. PubMed ID: 9724087). Changes in the CDK4 gene are also responsible for a small fraction of familial melanoma (Debniak. 2004. PubMed ID: 20233466).

In addition to germline variants in high penetrance genes, multiple studies have documented familial predisposition to malignant melanoma, noting that an individual has a higher risk of developing melanoma if an immediate family member has had the disease (Tucker et al. 2002. PubMed ID: 12115352; Debniak. 2004. PubMed ID: 20233466; Nelson et al. 2009. PubMed ID: 19095153). This increased risk might be due to shared lifestyle of sun exposure, fair skin, certain genetic variants, or a combination of these factors.

Melanoma is inherited in an autosomal dominant manner (Greene et al. 1983. PubMed ID: 6577466; Debniak. 2004. PubMed ID: 20233466). The locus for susceptibility for cutaneous malignant melanoma-1 (CMM1) was mapped to chromosome 1p36 (Cannon-Albright et al. 1990. PubMed ID: 2339690). To our knowledge, no gene of interest has been mapped to the CMM1 locus that is implicated in susceptibility to malignant melanoma-1. Subsequently, several other susceptibility loci were identified including CMM2 caused by variants in the CDKN2A gene; CMM3 caused by variants in CDK4; CMM4 mapped to chromosome 1p22; CMM5 caused by variants in MC1R gene; CMM6 caused by variants in XRCC3 gene; CMM7 mapped to chromosome 20q11; CMM8 caused by variants in MITF; CCM9 caused by variants in TERT; and CCM10 caused by variants in POT1. Though most familial cases of melanoma are a result of variants in CDKN2A, variants in CDK4 and RB1 have also been identified in familial melanoma families (Nelson et al. 2009. PubMed ID: 19095153).

The cyclin-dependent kinase inhibitor 2A (CDKN2A) gene encodes proteins that regulate two critical cell cycle regulatory pathways, namely TP53 and RB1. Two major separate but related proteins are produced from the CDKN2A gene as a result of shared coding regions and alternate reading frames: p16(INK4) which functions to inhibit CDK4/6-mediated phosphorylation of the Rb protein (active in dephosphorylated state), and p14(ARF) that binds MDM2, the p53-stabilizing protein, thereby inhibiting its function of marking proteins for degradation through ubiquitination (Nelson et al. 2009. PubMed ID: 19095153). TP53 functions to sense DNA damage and allow for repair, as well as to activate cellular apoptosis. Decreased levels of TP53 result in genetic instability and lead to a higher risk of melanoma in individuals with disruptions to the CDKN2A gene (Nelson et al. 2009. PubMed ID: 19095153). p16(INK4a) is formed by the fusion of exon 1α with exons 2 and 3, while p14(ARF) originated through alternate splicing of exon 1B with exons 2 and 3 (Nelson et al. 2009. PubMed ID: 19095153).

Tumor Protein p53 (TP53) encodes p53, the well-studied, ubiquitously expressed DNA-binding protein that plays significant roles in regulation of the cell cycle, DNA repair and programmed cells death. TP53 has 2 transcriptional start sites in exon 1, and alternative splicing occurs in intron 2 and between exon 9 and 10 (Bourdon et al. 2005. PubMed ID: 16131611). Germline TP53 pathogenic variants are found in patients with hereditary Li-Fraumeni syndrome, leading to early onset of several cancers, including melanoma (Platz et al. 1998. PubMed ID: 9724087).

Phosphatase and Tensin homolog deleted from chromosome 10 (PTEN) encodes a dual lipid and protein phosphatase. Loss of cancer suppressor genes located on chromosome 10 is highly implicated in melanoma tumorigenesis and is reported to contribute to 30-60% of non-inherited melanomas (Stahl et al. 2003. PubMed ID: 12782594). The 3’ end of exon 8 is subject to alternative splicing (Sharrard et al. 2000. PubMed ID: 11121587). PTEN also has a processed pseudogene PTENP1 that also exerts a tumor suppressive function (Poliseno et al. 2010. PubMed ID: 20577206).

BRCA2 is a tumor suppressor gene that is associated with a high risk for breast and ovarian cancer. Melanoma risk is elevated in some families with pathogenic variants in BRCA2 (Van Aperen et al. 2005. PubMed ID: 16141007). The most commonly reported cancers after breast and ovarian with BRCA2 variants are pancreas, prostate and melanoma (Mersch et al. 2015. PubMed ID: 25224030).

Microphthalmia-Associated Transcription Factor (MITF) is a basic helix-loop-helix-leucine zipper protein, and a melanocyte-specific transcription factor (Hartman and Czyz. 2015. PubMed ID: 25433395). It has been shown to play important developmental roles in various cell types including neural crest-derived melanocytes and optic cup-derived retinal epithelial cells (Fuse et al. 1999. PubMed ID: 10578055). It also helps to control the development and function of melanocytes which allow for the production of melanin that contributes to hair, eye and skin color. MITF acts as a melanoma oncogene, and controls the expression of CDKN2A/p16(INK4a) which has key roles in melanoma development. The germline missense variant c.952G>A (p.Glu318Lys) occurs at high frequency in patients affected with melanoma and renal cell carcinoma, and a >4 fold relative risk for melanoma observed in groups carrying the p.Glu318Lys variant compared to controls (Bertolotto et al. 2011. PubMed ID: 22012259; Yokoyama et al. 1999. PubMed ID: 22080950).

CDK4 is a protein-serine kinase that allows for orderly progression through the cell cycle. CDK4 encodes the cyclin dependent kinase 4 protein which is inhibited by p16(INK4a). Variants in CDK4 allow cells to skip the G1/S cell cycle checkpoint, thereby leading to uncontrolled cell proliferation (Ibrahim et al. 2009. PubMed ID: 19400696). Alterations in CDK4 are responsible for a very small proportion of familial melanoma (Debniak. 2004. PubMed ID: 20233466). The most common variants in CDK4 that are known to be involved in melanoma occur in codon 24 (p.Arg24Cys and p.Arg24His) (Soufir et al. 1998. PubMed ID: 9425228). When the amino acid substitutions occur, CDK4 is resistant to the inhibition of p16(INK4a) resulting in the phosphorylation of the Rb protein, allowing for the release of transcription factor E2F, and further resulting in uncontrolled cell proliferation (Ibrahim et al. 2009. PubMed ID: 19400696).

BRCA1-Associated Protein 1 (BAP1) gene encodes the deubiquitinating enzyme, nuclear ubiquitin carboxy-terminal hydrolase. It contains BRCA1 and BARD1 binding domains that together form a tumor suppressor heterodimeric complex. BAP1 has roles in chromatin dynamics, DNA damage response and cell cycle regulation (Goldstein et al. 2011. PubMed ID: 21956388). Germline variants in BAP1 have been reported to cause several types of cancer including uveal melanoma, cutaneous melanoma, non-melanoma skin, ovarian, breast, pancreatic, lung carcinoma and meningiomas.

Retinoblastoma 1 (RB1) is a tumor suppressor gene and a negative regulator of the cell cycle through repression of transcription of genes needed in S-phase. Retinoblastoma is an autosomal dominant disorder, and about 40% of cases are inherited or caused by germline variants in the RB1 gene. Families have also been reported with variants in the RB1 gene with an increased risk of melanoma. Germline variants in RB1 disrupt the ability of the RB1 protein to bind E2F and prevent unregulated cell division. It has been shown that though retinoblastoma is the most severe phenotype for carriers of RB1 variants, individuals who survive retinoblastoma tumors are at a 4-80 relative risk of developing melanoma later in life (Chin et al. 2006. PubMed ID: 16912270).

CHEK2 is involved in recognizing double-strand breaks in DNA, and allows for activation of downstream targets, such as p53 and BRCA2 (Weischer et al. 2012. PubMed ID: 2195612). Variants in CHEK2 have been frequently found in patients who have hereditary breast cancer, but do not have detectable variants in BRCA1 and BRCA2 (Desrichard et al. 2011. PubMed ID: 22114986), but has also been shown to associate with melanoma (Weischer et al. 2012. PubMed ID: 21956126). CHEK2 is upregulated in response to UV exposure, suggesting a role for the protein in control of UV damage (Yajima et al. 2009. PubMed ID: 19071136; Weischer et al. 2012. PubMed ID: 21956126). The CHEK2 c.1100delC variant has been shown to increase the risk of malignant melanoma ~2 fold in patients who are heterozygous for this variant, as these individuals have an impaired ability to repair DNA double-strand breaks induced by UV radiation (Weischer et al. 2012. PubMed ID: 21956126).

POT1 encodes a protein that is widely conserved across eukaryotic species and is associated with the telomere shelterin complex that regulates the protection of telomeres by preventing incorrect processing of chromosome ends. Studies have identified POT1 variants in individuals affected with malignant melanoma (Shi et al. 2014. PubMed ID: 24686846; Robles-Espinoza et al. 2014. PubMed ID: 24686849; Wilson et al. 2017. PubMed ID: 28389767). To date, approximately 15 missense variants have been reported to be associated with melanoma (Human Gene Mutation Database).

See individual gene test descriptions for further information on molecular biology of gene products and mutation spectra.

Genes tested in this panel have been associated with cutaneous malignant melanoma. Up to 50% of familial melanoma cases have causative germline variants in CDKN2A, and there is a 1-3% chance that an individual with primary melanoma has a causative germline variant in CDKN2A (Nelson et al. 2009. PubMed ID: 19095153; Ibrahim et al. 2009. PubMed ID: 19400696). CDKN2A variants were detected in 46%, 18%, and 8% of French, US and Swedish familial melanoma cases, respectively (Platz et al. 1997. PubMed ID: 9168184; Platz et al. 1998. PubMed ID: 9724087; Debiak. 2004. PubMed ID: 20233466). One study determined that 18% of men over age 50 with invasive melanoma are due to abnormal expression of p53 (Whiteman et al. 1998. PubMed ID: 9714052). Melanoma cell lines have been shown to contain approximately 10% of inactivating variants in PTEN (Stahl et al. 2003. PubMed ID: 12782594). The prevalence of causative variants in BRCA2 is 3% in patients with familial ocular melanoma (Scott et al. 2002. PubMed ID: 12385017; Debniak. 2004. PubMed ID: 20233466). MITF variant p.Glu318Lys was detected in 2.2% of melanoma prone families (n=270) (Yokoyama et al. 1999. PubMed ID: 22080950). The Geno-Mel study revealed that only 2% of families were carriers of CDK4 variants (Goldstein et al. 2006. PubMed ID: 17047042). Clinical sensitivity of BAP1 in malignant melanoma is unknown. Clinical sensitivity for detecting RB1 in retinoblastoma is ~95% (Rushlow et al. 2009. PubMed ID: 19280657), and is unknown at this time for malignant melanoma. Germline POT1 variants were identified in ~ 4% of melanoma patients who were negative for variants in CDKN2A or CDK4 (Robles-Espinoza. 2014. PubMed ID: 24686849). Therefore, a pathogenic variant in one of the ten genes in this panel would be expected in ~2-50% of individuals with a personal or family history of malignant melanoma.

This panel typically provides ≥98% coverage of all coding exons of the genes listed, plus ~10 bases of flanking noncoding DNA. We define coverage as ≥20X NGS reads or Sanger sequencing.