MC1R variants predisposing to concomitant primary cutaneous melanoma in a monozygotic twin pair
© Pellegrini et al.; licensee BioMed Central Ltd. 2012
Received: 11 June 2012
Accepted: 3 September 2012
Published: 14 September 2012
Concomitant primary cutaneous melanoma in monozygotic twins has been reported in only two pairs but in neither of them genetic analysis was performed. Two high-penetrance susceptibility genes, CDKN2A and CDK4 and one low-penetrance gene, MC1R, are well-defined genetic risk factors for melanoma. MITF has been recently identified as a novel intermediate risk melanoma-predisposing gene.
We describe the extraordinary occurrence of a primary cutaneous invasive melanoma in two 44-year-old identical, female twins, on the same body site within 30 days of each other and report for the first time the genetic analysis of melanoma susceptibility genes in both twins. Data on characteristics of the twins were collected through a standardized questionnaire and skin examination. Exons 1α, 1β, 2 and 3 of CDKN2A, exon 2 of CDK4, the entire open reading frame of MC1R and the recently described MITF c.952 G > A (p.Glu318Lys) variant were investigated by direct sequencing. Sequencing analysis of the high-penetrance susceptibility genes showed no changes in CDKN2A and in exon 2 of the CDK4 gene. Both patients were heterozygous for the same CDKN2A UTR c.*29C > G variant. Interestingly, the same two heterozygous variants of the MC1R were identified in both twins: the c.451C > T (p.Arg151Cys) and the c.456C > A (p.Tyr152*) variants. Neither patient showed the c.952 G > A (p.Glu318Lys) substitution in the MITF gene.
Identification of two high-risk MC1R variants in our identical twins in the absence of CDKN2A and CDK4 mutations highlights the contribution of low penetrance genes, such as MC1R, in melanoma susceptibility.
KeywordsMelanoma Monozygotic twins CDKN2A CDK4 MC1R MITF Genetic susceptibility
Genetic, phenotypic and environmental factors are known to affect melanoma risk. Two high-penetrance susceptibility genes, CDKN2A (cyclin-dependent kinase inhibitor 2A, MIM 600160) and CDK4 (cyclin-dependent kinase 4, MIM 123829) and one low-penetrance gene, MC1R (melanocortin-1 receptor, MIM 155555), have been identified so far as genetic risk factors for melanoma [1–3]. CDKN2A is the major melanoma susceptibility gene, mutated in approximately 40 % of melanoma-prone families with at least three affected members. Predisposing mutations in CDK4 have been identified in a limited number of melanoma families worldwide. Allelic variants in the MC1R gene, important in the pigmentation process, have been associated with increased melanoma risk, with the strongest effect observed for red hair color (RHC) variants (p.ArgR151Cys, p.Arg160Trp, p.Asp294His). Recently, whole genome sequencing lead to the identification of MITF (microphtalmia-associated transcription factor, MIM 156845) as a novel melanoma-predisposing gene with the p.Glu318Lys variant defined as an intermediate risk variant .
We describe the extraordinary occurrence of a primary cutaneous invasive melanoma in two 44-year-old female, identical twins, on the same body site within 30 days of each other and report for the first time genetic analysis of melanoma susceptibility genes in both twins.
Genomic DNA was isolated from whole blood of both patients using QIAamp® DNA Blood kit (Qiagen, Hilden, Germany). Exons 1α, 1β, 2 and 3, including the exon-intron boundaries of CDKN2A (NCBI accession NM_000077.3), exon 2 of CDK4 (NCBI accession NM_000075.3) and the entire open reading frame of MC1R (NCBI accession NM_002386.3) were amplified using PCR and directly sequenced on ABI Prism® 310 Genetic Analyzer (Applied Biosystems, Foster City, CA) as described previously [5, 6]. The recently described MITF c.952 G > A (p.Glu318Lys) variant (NCBI accession NM_000248.3) was screened by direct sequencing of a PCR-amplified 589 amplicon using the following primers: forward, 5'-TACATTCCCTCTGGTATTGT-3'; reverse, 5'-AGCAGTTTGTGCGAATGCA-3'. Primers for PCR amplification were designed from the MITF genomic nucleotide sequence (NCBI accession NG_011631.1) and were the followings: forward, 5'-TACATTCCCTCTGGTATTGT-3'; reverse 5'-AGCAGTTTGTGCGAATGCA-3'. An amplicon of 589 bp was obtained by standard PCR using 1.25 U of AmpliTaq Gold® 360 (Applied Biosystems) in a 50-μl volume, containing the 1X reaction buffer provided by the manufacturer, 1.6 mM of MgCl2, 200 μM of each deoxynucleoside triphosphate, 0.2 μM of each primer and 100 ng genomic DNA template. Five per cent dimethyl sulfoxide was added to the reaction solution. PCR amplification conditions were as follows: 95°C for 7 min, 35 cycles of 94°C for 1 min, 52.2°C for 1 min, and 72°C for 1 min, followed by a final extension step at 72°C for 7 min. Written informed consent was obtained from our patients under an Institutional Review Board-approved protocol.
Sequencing analysis showed no changes in the coding region and in the 5’UTR region of CDKN2A, although both patients were heterozygous for the same UTR c.*29C > G variant. No mutations were detected in exon 2 of the CDK4 gene. Interestingly, sequencing of the MC1R identified the same two heterozygous variants in both twins: the c.451C > T (p.Arg151Cys) and the c.456C > A (p.Tyr152*) variants (Figure 1D). Neither patient showed the c.1075 G > A (p.Glu318Lys) substitution in the MITF gene.
The occurrence of melanoma concordant for site and age of onset in monozygotic twins has been reported in only two pairs but in neither of them genetic analysis was performed. St-Arneault et al.  described two identical male twins (from a set of triplets) who developed melanoma at age 53, at the same site and within a time frame of 2 months, while the fraternal triplet showed no evidence of tumour. More recently, Rao et al. reported two 71-year-old, identical female twins, diagnosed with melanoma at the same time (within 10 days of each other) and location (right calf). Our patients were skin type 1 as the latter pair but they were younger than both pairs.
Studies in twin pairs are of great interest to estimate the relative effect of genetic and environmental influences on melanoma development. In a recent large population-based study of Australian twins, genetic influences were shown to be a significant source of variation in liability to melanoma with identical twins being more than four times more likely to be affected with melanoma if they had an affected co-twin than non-identical twins .
MC1R variants have been associated with a 1.2 to 2.4-fold increased risk of melanoma with an additive effect for multiple variants [10–12]. A double variation in the MC1R gene, the c.451C > T (p.Arg151Cys) and the c.456C > A (p.Tyr152*), was detected in our twins. The p.Arg151Cys has been shown to confer the highest melanoma risk with summary estimates ranging from 1.78 (95%CI 1.45–2.20) to 1.93 (95%CI 1.54–2.41) in two recent meta-analyses [10, 11]. We previously reported a significant association of the p.Arg151Cys allele with melanoma risk in a population from central Italy, with an OR of 2.94 (95%CI, 1.04–8.31) . In functional studies, the p.Arg151Cys mutant receptor showed a reduced cell surface expression with intracellular retention and a corresponding impairment in cAMP coupling . The nonsense p.Tyr152* is a rare RHC variant associated with a severe functional impairment of the MC1R due to the lack of the last four transmembrane fragments [14, 15]. Interestingly, MC1R-associated melanoma risk has been shown to increase with the number of variants in the genotype supporting the MC1R-associated susceptibility in our patients carrying two nonfunctional MC1R variants. Finally, carriers of MC1R RHC variants have been recently suggested to develop melanomas lacking significant pigmentation [16, 17] as also observed in the hypomelanotic melanomas of our twins.
In addition to the well-defined association of MC1R with melanoma, other low-penetrance melanoma risk alleles have been described in genes related to pigmentation, nevus count, immune responses, DNA repair, metabolism and the vitamin D receptor . In this regard, we cannot rule out the possible involvement of other low-penetrance melanoma susceptibility alleles in our twins since they were not genotyped in our patients.
The concordance for cutaneous melanoma as well as the congruence for its location and identity of the age of onset in monozygotic twins support the role of genetic rather than environmental factors in the development of melanoma. Detection of two high-risk MC1R variants in our identical twins in the absence of CDKN2A and CDK4 mutations highlights the contribution of low penetrance genes, such as MC1R, in melanoma susceptibility, although additional known or as yet unknown genetic risk factors might be involved.
Written informed consent was obtained from both patients for publication of this case report and any accompanying images. A copy of the written consent is available for review by the Editor-in-Chief of this journal.
- CDKN2A :
Cyclin-dependent kinase inhibitor 2A
- CDK4 :
Cyclin-dependent kinase 4
- MC1R :
- MITF :
Microphtalmia-associated transcription factor
Red hair color
Superficial spreading melanoma.
- Fargnoli MC, Argenziano G, Zalaudek I, Peris K: High- and low-penetrance cutaneous melanoma susceptibility genes. Expert Rev Anticancer Ther. 2006, 6: 657-670. 10.1586/14737220.127.116.117.View ArticlePubMed
- Goldstein AM, Chan M, Harland M, Gillanders EM, Hayward NK, Avril MF, Azizi E, Bianchi-Scarra G, Bishop DT, Bressac-de Paillerets B, Bruno W, Calista D, Cannon Albright LA, Demenais F, Elder DE, Ghiorzo P, Gruis NA, Hansson J, Hogg D, Holland EA, Kanetsky PA, Kefford RF, Landi MT, Lang J, Leachman SA, Mackie RM, Magnusson V, Mann GJ, Niendorf K, Newton Bishop J, Palmer JM, Puig S, Puig-Butille JA, de Snoo FA, Stark M, Tsao H, Tucker MA, Whitaker L, Yakobson E, Melanoma Genetics Consortium (GenoMEL): High-risk melanoma susceptibility genes and pancreatic cancer, neural system tumors, and uveal melanoma across GenoMEL. Cancer Res. 2006, 66: 9818-9828. 10.1158/0008-5472.CAN-06-0494.View ArticlePubMed
- Meyle KD, Guldberg P: Genetic risk factors for melanoma. Hum Genet. 2009, 126: 499-510. 10.1007/s00439-009-0715-9.View ArticlePubMed
- Yokoyama S, Woods SL, Boyle GM, Aoude LG, MacGregor S, Zismann V, Gartside M, Cust AE, Haq R, Harland M, Taylor JC, Duffy DL, Holohan K, Dutton-Regester K, Palmer JM, Bonazzi V, Stark MS, Symmons J, Law MH, Schmidt C, Lanagan C, O'Connor L, Holland EA, Schmid H, Maskiell JA, Jetann J, Ferguson M, Jenkins MA, Kefford RF, Giles GG, Armstrong BK, Aitken JF, Hopper JL, Whiteman DC, Pharoah PD, Easton DF, Dunning AM, Newton-Bishop JA, Montgomery GW, Martin NG, Mann GJ, Bishop DT, Tsao H, Trent JM, Fisher DE, Hayward NK, Brown KM: A novel recurrent mutation in MITF predisposes to familial and sporadic melanoma. Nature. 2011, 480: 99-103. 10.1038/nature10630.PubMed CentralView ArticlePubMed
- Peris K, Fargnoli MC, Pacifico A, Surrenti T, Stolz W, Wolf P, Soyer HP, Chimenti S: CDKN2A and MC1R mutations in patients with sporadic multiple primary melanoma. J Invest Dermatol. 2004, 122: 1327-1330. 10.1111/j.0022-202X.2004.22532.x.View ArticlePubMed
- Fargnoli MC, Altobelli E, Keller G, Chimenti S, Höfler H, Peris K: Contribution of melanocortin-1 receptor gene variants to sporadic cutaneous melanoma risk in a population in central Italy: a case-control study. Melanoma Res. 2006, 16: 175-182. 10.1097/01.cmr.0000198454.11580.b5.View ArticlePubMed
- St-Arneault G, Nagel G, Kirkpatrick D, Kirkpatrick R, Holland JF: Melanoma in twins Cutaneous malignant melanoma in identical twins from a set of triplets. Cancer. 1970, 25: 672-677. 10.1002/1097-0142(197003)25:3<672::AID-CNCR2820250324>3.0.CO;2-A.View ArticlePubMed
- Rao BK, Noor O, Thosani MK: Identical twins with primary cutaneous melanoma presenting at the same time and location. Am J Dermatopathol. 2008, 30: 182-184. 10.1097/DAD.0b013e318164fc87.View ArticlePubMed
- Shekar SN, Duffy DL, Youl P, Baxter AJ, Kvaskoff M, Whiteman DC, Green AC, Hughes MC, Hayward NK, Coates M, Martin NG: A population-based study of Australian twins with melanoma suggests a strong genetic contribution to liability. J Invest Dermatol. 2009, 129: 2211-2219. 10.1038/jid.2009.48.PubMed CentralView ArticlePubMed
- Raimondi S, Sera F, Gandini S, Iodice S, Caini S, Maisonneuve P, Fargnoli MC: MC1R variants, melanoma and red hair color phenotype: a meta-analysis. Int J Cancer. 2008, 122: 2753-2760. 10.1002/ijc.23396.View ArticlePubMed
- Williams PF, Olsen CM, Hayward NK, Whiteman DC: Melanocortin 1 receptor and risk of cutaneous melanoma: a meta-analysis and estimates of population burden. Int J Cancer. 2011, 129: 1730-1740. 10.1002/ijc.25804.View ArticlePubMed
- Ward KA, Lazovich D, Hordinsky MK: Germline melanoma susceptibility and prognostic genes: A review of the literature. J Am Acad Dermatol. 2012, [Epub ahead of print] PMID: 22583682
- Beaumont KA, Shekar SN, Newton RA, James MR, Stow JL, Duffy DL, Sturm RA: Receptor function, dominant negative activity and phenotype correlations for MC1R variant alleles. Hum Mol Genet. 2007, 16: 2249-2260. 10.1093/hmg/ddm177.View ArticlePubMed
- García-Borrón JC, Sánchez-Laorden BL, Jiménez-Cervantes C: Melanocortin-1 receptor structure and functional regulation. Pigment Cell Res. 2005, 18: 393-410.PubMed
- Galore G, Azizi E, Scope A, Pavlotsky F, Yakobson E, Friedman E: The Y152X MC1R gene mutation: occurrence in ethnically diverse Jewish malignant melanoma patients. Melanoma Res. 2007, 17: 105-108. 10.1097/CMR.0b013e3280c31d81.View ArticlePubMed
- Cuéllar F, Puig S, Kolm I, Puig-Butille J, Zaballos P, Martí-Laborda R, Badenas C, Malvehy J: Dermoscopic features of melanomas associated with MC1R variants in Spanish CDKN2A mutation carriers. Br J Dermatol. 2009, 160: 48-53. 10.1111/j.1365-2133.2008.08826.x.View ArticlePubMed
- Zalaudek I, Meiklejohn W, Argenziano G, Thurber AE, Sturm RA: "White" nevi and "red" melanomas: association with the RHC phenotype of the MC1R gene. J Invest Dermatol. 2009, 129: 1305-1307. 10.1038/jid.2008.378.View ArticlePubMed
- The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2350/13/81/prepub
This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.