DNA Methylation Levels of Melanoma Risk Genes Are Associated with Clinical Characteristics of Melanoma Patients

de Araújo, Érica S. S.; Pramio, Dimitrius T.; Kashiwabara, André Y.; Pennacchi, Paula C.; Maria-Engler, Silvya S.; Achatz, Maria I.; Campos, Antonio H. J. F. M.; Duprat, João P.; Rosenberg, Carla; Carraro, Dirce M.; Krepischi, Ana C. V.

doi:https://doi.org/10.1155/2015/376423

BioMed Research International

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Abstract Introduction Methods Results Discussion Conclusion Acknowledgments Supplementary Materials References Copyright Related Articles

Research Article | Open Access

Volume 2015 | Article ID 376423 | https://doi.org/10.1155/2015/376423

DNA Methylation Levels of Melanoma Risk Genes Are Associated with Clinical Characteristics of Melanoma Patients

Érica S. S. de Araújo,¹Dimitrius T. Pramio,¹André Y. Kashiwabara,²Paula C. Pennacchi,³Silvya S. Maria-Engler,³Maria I. Achatz,^1,4Antonio H. J. F. M. Campos,⁵João P. Duprat,⁶Carla Rosenberg,⁷Dirce M. Carraro,¹and Ana C. V. Krepischi^1,7

Academic Editor: Rajiv Kumar

Received21 Jan 2015

Accepted23 Mar 2015

Published12 Apr 2015

Abstract

In melanoma development, oncogenic process is mediated by genetic and epigenetic mutations, and few studies have so far explored the role of DNA methylation either as predisposition factor or biomarker. We tested patient samples for germline CDKN2A methylation status and found no evidence of inactivation by promoter hypermethylation. We have also investigated the association of clinical characteristics of samples with the DNA methylation pattern of twelve genes relevant for melanomagenesis. Five genes (BAP1, MGMT, MITF, PALB2, and POT1) presented statistical association between blood DNA methylation levels and either CDKN2A-mutation status, number of lesions, or Breslow thickness. In tumors, five genes (KIT, MGMT, MITF, TERT, and TNF) exhibited methylation levels significantly different between tumor groups including acral compared to nonacral melanomas and matched primary lesions and metastases. Our data pinpoint that the methylation level of eight melanoma-associated genes could potentially represent markers for this disease both in peripheral blood and in tumor samples.

1. Introduction

Cutaneous melanoma has an increasing incidence rate worldwide [1]. The occurrence of this type of cancer is associated with skin color, geographic localization, and intermittent exposure to sunlight, and the former is probably the major cause of the increase in melanoma cases [2].

The risk of melanoma development rises 30- to 70-fold in individuals with family history of the disease [3]. Two genes harboring high-risk germline mutations have been recognized: CDKN2A [4–6] and CDK4 [7]. Germline mutations in additional genes were recently associated with melanoma occurrence: MITF [8], BAP1 [9], TERT promoter [10], POT1 [11], and MGMT [12]. In addition, some alterations in genes known to be related to melanin synthesis, such as MC1R [13, 14], have been reported as low-risk variants for melanoma development. The implication of other candidate genes in melanoma predisposition is supported by less conclusive evidence, such as PALB2, in which a deleterious mutation has been detected in a single individual affected by four kinds of cancer, including melanoma [15].

In addition to genetic mutations, early events of anomalous DNA methylation at specific loci (epimutations) can eventually result in cancer predisposition, as already reported for colorectal [16, 17], gastric [18], and breast cancers [19]. Although melanoma epimutations have not been identified so far [20, 21], abnormal methylation levels of TNF and TNFRSF10C genes have been associated with melanoma risk [22, 23]. Moreover, somatic DNA methylation changes are crucial for skin melanoma progression [24]. Recently, the occurrence of KIT epigenetic silencing in cutaneous melanomas has been demonstrated [25], supporting earlier studies that had shown decrease of KIT expression associated with melanoma progression [26].

In this work, we investigated the methylation pattern of germline CDKN2A and other 10 melanoma-related genes in leukocytes of melanoma patients (familial and sporadic cases), relating these findings to melanoma occurrence and clinical characteristics. The methylation profile of these genes and KIT was also investigated in a group of cutaneous melanomas.

2. Material and Methods

This was a retrospective study based on samples from melanoma patients retrieved from the A.C.Camargo Cancer Center Biobank, São Paulo, Brazil, and approved by the Institutional Ethics Committee Board (1695/12 and 1765/13).

2.1. Peripheral Blood Samples of Melanoma Patient and Control Groups

Melanoma patients were treated at the A.C.Camargo Cancer Center, São Paulo, Brazil. DNA samples were extracted from peripheral blood obtained from 69 unrelated cutaneous melanoma patients who had not previously received chemotherapy, most of them being stage I (58.0%). The patients were classified into three groups: hereditary melanoma carrying pathogenic CDKN2A-mutations (), familial history of melanoma without CDKN2A-mutations (), and sporadic cases (without family history of the disease; ). The 20 hereditary melanoma patients who had previously tested negative for CDKN2A-mutations [27] fulfilled at least one of the following criteria: (a) familial melanoma (family history of three or more relatives of two consecutive generations with melanomas, at least one case before 50 years old; ) or (b) multiple primary melanomas (≥3 primary lesions, ; or ≥2 primary melanomas, one of which before 35 years old, ).

The control group was composed of 12 individuals without cancer history, matched by age (range for each being 10 years) and gender to the subset of melanoma patients who were analyzed by DNA methylation array.

2.2. Cutaneous Melanoma and Melanocyte Samples

Frozen tissues from 18 unrelated primary skin melanomas (five acral and thirteen nonacral subtypes) were retrieved from the A.C.Camargo Tissue Bank. DNA extraction was processed at A.C.Camargo DNA and RNA Bank [28]. Paired metastases were available for seven tumors. The majority of tumors were classified as stage III (66.7%), but this information was missing for the remaining samples.

The primary cultures of melanocytes were provided by the School of Pharmaceutical Sciences (University of São Paulo). Primary skin cell cultures (melanocytes) were obtained from foreskin samples of three healthy donors [29] under approval of the local ethics committee (CEP HU/USP 943/09).

2.3. Pyrosequencing

Quantitative bisulfite pyrosequencing for CDKN2A promoter was performed using PyroMark Q96 CpG p16 (Qiagen), which analyzes seven CpG sites of p16 isoform (position +148 to +174 of the ENSG00000147889). A total of 68 peripheral blood samples from patients were analyzed: seven CDKN2A-mutation carriers, 20 hereditary melanoma patients without CDKN2A-mutations, and 41 sporadic melanoma patients. The methylation levels were obtained using PyroMark Q96-CpG Software (Qiagen), which provides the percentage of methylated cytosines relative to the sum of methylated and unmethylated cytosines.

2.4. HM450K Genome-Wide Methylation Analysis

DNA methylation array was performed in samples from 18 cutaneous melanomas, three melanocyte primary cultures, and leukocyte samples from 12 control individuals and from a subset of 39 melanoma patients (eight CDKN2A-mutation carriers, 19 hereditary melanoma patients without CDKN2A-mutations, and 12 sporadic melanoma patients).

Bisulfite modification was performed on 1 μg of DNA using EZ DNA methylation kit (Zymo Research). The DNA methylation levels of CpG sites were obtained using the Infinium HumanMethylation450K BeadChips (HM450K, Illumina), according to the manufacturer’s instructions. Bioconductor IMA package was applied for quality control of array methylation data [30]. We removed from further analyses probes that were lacking beta-values, containing SNPs, mapped at chromosomes X and Y, or with detection values > 0.01. Probe type bias adjustment was achieved by Beta Mixture Quantile dilation (BMIQ) using ChAMP package [31].

The level of methylation of each CpG probe was calculated as a beta-value ranging from 0 to 1 (0 for totally unmethylated CpG sites and 1 for fully methylated sites). Beta-values were retrieved for all CpG probes mapped within DNA sequences associated with the BAP1, CDK4, CDKN2A, KIT, MC1R, MGMT, MITF, PALB2, POT1, TERT, TNF, and TNFRSF10C genes (see Supplementary Tables 1 and 2 in Supplementary Material available online at http://dx.doi.org/10.1155/2015/376423). CpG sites were grouped into two categories, according to the annotation of the transcripts provided by Illumina, which is based on the UCSC database: “promoter” region (CpG probes mapped at TSS1500, TSS200, 5′UTR, or 1st exon) and “gene body” (remaining exons and intron probes). Methylation levels for promoters and bodies were obtained using the beta-value means of the corresponding CpG probes.

Microarray methylation data from peripheral blood were deposited into NCBI’s Gene Expression Omnibus under accession number GSE54939 (http://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE54939).

2.5. Statistical Analysis of HM450K Data

DNA methylation differences between groups were tested for significance using either Mann-Whitney test or Kruskal-Wallis with Dunn’s Multiple Comparison Post-test. The analysis comparing primary cutaneous melanoma and paired metastasis was performed using paired -test.

3. Results

The quantitative bisulfite pyrosequencing of CDKN2A promoter was investigated to determine if any of the melanoma patients exhibited inactivation of this gene by hypermethylation. Data showed no evidence of CDKN2A hypermethylation in leukocytes of these patients. Among CDKN2A-mutation carriers, the methylation mean was 0.63% (ranging from 0 up to 1.5%), hereditary melanoma patients without CDKN2A-mutations showed a mean of 0.52% (0–3.0%), and sporadic melanoma patients presented a mean of 0.35% (0–1.5%). Statistical association between the clinical characteristics of melanoma patients and the methylation levels of p16 encode region was not detected (Breslow thickness ; number of melanomas ).

Next, we compared the levels of DNA methylation for CpG sites mapped in eleven melanoma-associated genes (BAP1, CDK4, CDKN2A, MC1R, MGMT, MITF, PALB2, POT1, TERT, TNF, and TNFRSF10C) between melanoma patients and controls. Only the gene body of MGMT showed a low level of methylation increase in CDKN2A-mutated patients compared to sporadic melanoma patients (2% increase in the methylation level, , Supplementary Table 3). Additionally, DNA methylation levels at the promoter regions of BAP1, MITF, and PALB2 genes were statistically associated with the number of melanomas presented by the individual and a hypermethylation of POT1 promoter was associated with Breslow thickness of the tumors (Table 1).

We also investigated the DNA methylation levels of twelve melanoma-associated genes (BAP1, CDK4, CDKN2A, KIT, MC1R, MGMT, MITF, PALB2, POT1, TERT, TNF, and TNFRSF10C) by comparing melanoma samples with cultured melanocytes. The methylation levels at the gene bodies of MGMT, TERT, and TNF were significantly lower in tumors, while both body and promoter of KIT were hypermethylated (Table 2). Acral melanomas exhibited differential methylation at the gene bodies of MGMT (4% hypomethylation in acral melanoma group, , Supplementary Table 4) and TERT (9% hypermethylation in acral melanoma group, , Supplementary Table 4) compared to nonacral tumors. We did not find significant differences comparing tumors from patients who did develop or not metastases (Supplementary Table 5); however, in the analysis of the seven paired primary-metastasis melanoma samples, the gene body of MITF was found to be hypomethylated in metastasis (8% of decreased methylation in relation to primary tumor, , Supplementary Table 6).

4. Discussion

Aberrant DNA methylation is usually an early and stable event in tumorigenesis and could be used for detecting and monitoring diseases [32]. Previous studies in melanoma and surrogate samples have revealed epigenetic markers based on methylation patterns of LINE-1 [33, 34], RASSF1A [35], TSLC1 [36],and MGMT [37]. In this study, we have investigated the methylation levels of twelve melanoma genes to evaluate their potential role as predisposition factors and/or biomarkers. In fact, control of gene expression by DNA methylation has already been described for half of these investigated genes in different types of cancer [38–43]. Considering that the functional impact of DNA methylation in gene expression is better understood for CpGs mapped at gene promoters, we analyzed separately the probes mapped at promoters and gene bodies. Most of the genes here analyzed exhibited CpG islands in their promoter sequences, except for MITF and TNF.

Independent cohorts of patient peripheral blood and melanoma samples were analyzed. Overall, the detected DNA methylation differences in blood were very small (<5%), while tumors exhibited higher (>10%) differences. Indeed, previous studies have shown that epigenetic biomarkers in peripheral blood exhibit very small differences [44–47]. Significant DNA methylation changes were identified in peripheral blood mainly at gene promoters, while in tumors they appeared to cluster at gene bodies. Also, melanomas were mainly hypomethylated when compared to the control melanocytes; however, we cannot exclude the possibility that the DNA methylation pattern of the melanocytes could be altered because of in vitro conditions [48].

MGMT encodes a repair protein which removes alkyl groups from the O₆-position of guanine residues [49] and its promoter methylation has been described as a biomarker in several studies, especially for glioblastoma [50] and colorectal cancer [51]. In melanoma, epigenetic silencing of this gene has been demonstrated in tumors and serum of patients [52, 53], suggesting that this event would be important for tumor development. In our study, we detected methylation differences at MGMT body both in peripheral blood and in tumor samples: CDKN2A-mutated patients showed a low-level hypermethylation in leukocytes compared to other melanoma patients, while tumors showed hypomethylation compared to melanocytes. Even though gene body methylation is a yet poorly explored mechanism, it has been reported in association with increased gene expression [54]. However, the relationship between intragenic methylation and gene expression appears not to be linear, since low body methylation levels have been associated with both low and high expression, and high body methylation levels have been already reported linked to intermediate expression [55]. Additional studies in melanomas are necessary to clarify the role of the observed methylation changes in the expression of the MGMT gene.

MITF is a transcription factor that controls genes of cell cycle and melanogenesis [56, 57]. Hypermethylation of the MITF promoter was detected in peripheral blood of those melanoma patients who developed more than one lesion (multiple primary melanomas), and MITF gene body was found to be somehow hypermethylated in primary tumors compared to metastasis. Interestingly, MITF expression was reported as variable among melanoma specimens and even intratumor [58], with high expression levels being associated with either differentiation or proliferation while low expression is related to an invasive potential [59]. This dual role of MITF according to its expression level could be involved in the different patterns of methylation observed in peripheral blood and tumors, suggesting that (a) high methylation levels at MITF promoter in leukocytes could be linked to a more aggressive disease and (b) higher methylation level at MITF gene body of primary tumors compared to metastasis would have a controlling role in cell cycle. Lauss and colleagues [60] have recently shown that MITF expression in melanomas is controlled by DNA methylation, but unfortunately RNA was unavailable to investigate whether the low DNA methylation changes here detected would impact MITF expression.

We also detected DNA methylation differences between acral and nonacral melanoma subtypes at MGMT and TERT bodies. Acral melanoma is an uncommon subtype not associated with sunburn and occurs more frequently in older and non-Caucasian individuals [61]. Additionally, acral driver mutations are distinct compared to cutaneous melanomas. For instance, KIT mutations are more common in acral than in the other melanomas [62, 63]. We did not observe significant difference in KIT methylation levels between acral and nonacral subtypes, but melanomas as a group were hypermethylated compared to melanocytes. The detection of KIT hypermethylation in the advanced tumors of this study (mostly stage III melanomas) was not unexpected and had already been described [25, 64].

The CDKN2A methylation level was analyzed by the use of microarray (HM450K) and pyrosequencing. This gene encodes p16 and p14 proteins, the p16 isoform harboring the majority of the germline mutations that confer a high melanoma risk [65]. Since the array platform contains CDKN2A CpGs not specifically associated with p16, we investigated additional CDKN2A promoter CpGs by pyrosequencing. The melanoma patients exhibited low methylation levels for CDKN2A, similar to the pattern already reported for healthy individuals [23]. Our data in this Brazilian cohort of patients are in accordance with previous studies that described absence of CDKN2A epimutation in hereditary melanoma patients [20, 21].

5. Conclusion

Here we investigated the potential role of twelve melanoma-associated genes as predisposition factors and epigenetic biomarkers for melanoma, using both peripheral blood and tumor samples. We did not find germline CDKN2A hypermethylation in our cohort of Brazilian melanoma patients. However, we detected low-level albeit significant DNA methylation differences in a gene subset both in leukocytes and in tumors (BAP1, KIT, MGMT, MITF, PALB2, POT1, TERT, and TNF). Additional studies in a large melanoma cohort are necessary to validate these candidate genes as epigenetic biomarkers in melanoma.

Conflict of Interests

The authors declare that there is no conflict of interests regarding the publication of this paper.

Acknowledgments

The authors acknowledge the A.C.Camargo Cancer Center Biobank for providing patient samples. This work was supported by grants from FAPESP (2012/13963-9, 2013/10785-5, and 2013/07480-8) and CNPq (470446_2013-7).

Supplementary Materials

The Supplementary Material files contain the list of the HM450K probes that were analyzed in peripheral blood samples (Supplementary Table 1) and in melanoma samples (Supplementary Table 2), the DNA methylation levels of melanoma-associated genes in peripheral blood samples (Supplementary Table 3) and in melanoma samples (Acral versus Non-acral melanomas, Supplementary Table 4; primary melanomas from patients who did develop or not metastasis, Supplementary Table 5; paired primary tumors and metastasis, Supplementary Table 6).

Supplementary Material

References

J. A. Usher-Smith, J. Emery, A. P. Kassianos, and F. M. Walter, “Risk prediction models for melanoma: a systematic review,” Cancer Epidemiology Biomarkers & Prevention, vol. 23, no. 8, pp. 1450–1463, 2014.
View at: Publisher Site | Google Scholar
U. Leiter, T. Eigentler, and C. Garbe, “Epidemiology of skin cancer,” Advances in Experimental Medicine and Biology, vol. 810, pp. 120–140, 2014.
View at: Google Scholar
R. F. Kefford, J. A. N. Bishop, W. Bergman, and M. A. Tucker, “Counseling and DNA testing for individuals perceived to be genetically predisposed to melanoma: a consensus statement of the melanoma genetics consortium,” Journal of Clinical Oncology, vol. 17, no. 10, pp. 3245–3251, 1999.
View at: Google Scholar
L. A. Cannon-Albright, D. E. Goldgar, L. J. Meyer et al., “Assignment of a locus for familial melanoma, MLM, to chromosome 9p13-p22,” Science, vol. 258, no. 5085, pp. 1148–1152, 1992.
View at: Publisher Site | Google Scholar
A. Kamb, D. Shattuck-Eidens, R. Eeles et al., “Analysis of the p16 gene (CDKN2) as a candidate for the chromosome 9p melanoma susceptibility locus,” Nature Genetics, vol. 8, no. 1, pp. 22–26, 1994.
View at: Publisher Site | Google Scholar
D. E. Quelle, F. Zindy, R. A. Ashmun, and C. J. Sherr, “Alternative reading frames of the INK4a tumor suppressor gene encode two unrelated proteins capable of inducing cell cycle arrest,” Cell, vol. 83, no. 6, pp. 993–1000, 1995.
View at: Publisher Site | Google Scholar
L. Zuo, J. Weger, Q. Yang et al., “Germline mutations in the p16INK4a binding domain of CDK4 in familial melanoma,” Nature Genetics, vol. 12, no. 1, pp. 97–99, 1996.
View at: Publisher Site | Google Scholar
S. Yokoyama, S. L. Woods, G. M. Boyle et al., “A novel recurrent mutation in MITF predisposes to familial and sporadic melanoma,” Nature, vol. 480, no. 7375, pp. 99–103, 2011.
View at: Publisher Site | Google Scholar
C.-N. J. Njauw, I. Kim, A. Piris et al., “Germline BAP1 inactivation is preferentially associated with metastatic ocular melanoma and cutaneous-ocular melanoma families,” PLoS ONE, vol. 7, no. 4, Article ID e35295, 2012.
View at: Publisher Site | Google Scholar
S. Horn, A. Figl, P. S. Rachakonda et al., “TERT promoter mutations in familial and sporadic melanoma,” Science, vol. 339, no. 6122, pp. 959–961, 2013.
View at: Publisher Site | Google Scholar
C. D. Robles-Espinoza, M. Harland, A. J. Ramsay et al., “POT1 loss-of-function variants predispose to familial melanoma,” Nature Genetics, vol. 46, no. 5, pp. 478–481, 2014.
View at: Publisher Site | Google Scholar
F. Appelqvist, M. Yhr, A. Erlandson, T. Martinsson, and C. Enerbäck, “Deletion of the MGMT gene in familial melanoma,” Genes, Chromosomes & Cancer, vol. 53, no. 8, pp. 703–711, 2014.
View at: Publisher Site | Google Scholar
J. S. Palmer, D. L. Duffy, N. F. Box et al., “Melanocortin-1 receptor polymorphisms and risk of melanoma: is the association explained solely by pigmentation phenotype?” American Journal of Human Genetics, vol. 66, no. 1, pp. 176–186, 2000.
View at: Publisher Site | Google Scholar
A. Marzuka-Alcalá, M. J. Gabree, and H. Tsao, “Melanoma susceptibility genes and risk assessment,” Methods in Molecular Biology, vol. 1102, pp. 381–393, 2014.
View at: Publisher Site | Google Scholar
L. G. Aoude, M. Xu, Z. Z. Zhao et al., “Assessment of PALB2 as a candidate melanoma susceptibility gene,” PLoS ONE, vol. 9, no. 6, 2014.
View at: Publisher Site | Google Scholar
I. Gazzoli, M. Loda, J. Garber, S. Syngal, and R. D. Kolodner, “A hereditary nonpolyposis colorectal carcinoma case associated with hypermethylation of the MLH1 gene in normal tissue and loss of heterozygosity of the unmethylated allele in the resulting microsatellite instability-high tumor,” Cancer Research, vol. 62, no. 14, pp. 3925–3928, 2002.
View at: Google Scholar
R. Venkatachalam, M. J. L. Ligtenberg, N. Hoogerbrugge et al., “Germline epigenetic silencing of the tumor suppressor gene PTPRJ in early-onset familial colorectal cancer,” Gastroenterology, vol. 139, no. 6, pp. 2221–2224, 2010.
View at: Publisher Site | Google Scholar
H. Pinheiro, R. Bordeira-Carriço, S. Seixas et al., “Allele-specific CDH1 downregulation and hereditary diffuse gastric cancer,” Human Molecular Genetics, vol. 19, no. 5, pp. 943–952, 2010.
View at: Publisher Site | Google Scholar
E. M. Wong, M. C. Southey, S. B. Fox et al., “Constitutional methylation of the BRCA1 promoter is specifically associated with BRCA1 mutation-associated pathology in early-onset breast cancer,” Cancer Prevention Research, vol. 4, no. 1, pp. 23–33, 2011.
View at: Publisher Site | Google Scholar
A. Erlandson, F. Appelqvist, and C. Enerbäck, “Epigenetic mutations in CDKN2A in western Swedish families with hereditary malignant melanoma,” Molecular Medicine Reports, vol. 1, no. 1, pp. 89–91, 2008.
View at: Google Scholar
R. van Doorn, W. H. Zoutman, and N. A. Gruis, “Absence of germline epimutation of the CDKN2A gene in familial melanoma,” Journal of Investigative Dermatology, vol. 129, no. 3, pp. 781–784, 2009.
View at: Publisher Site | Google Scholar
L. Pergoli, C. Favero, R. M. Pfeiffer et al., “Blood DNA methylation, nevi number, and the risk of melanoma,” Melanoma Research, vol. 24, no. 5, pp. 480–487, 2014.
View at: Publisher Site | Google Scholar
P. L. Hyland, L. S. Burke, R. M. Pfeiffer et al., “Constitutional promoter methylation and risk of familial melanoma,” Epigenetics, vol. 9, no. 5, pp. 685–692, 2014.
View at: Publisher Site | Google Scholar
S. Ecsedi, H. Hernandez-Vargas, S. C. Lima et al., “DNA methylation characteristics of primary melanomas with distinct biological behaviour,” PLoS ONE, vol. 9, no. 5, Article ID e96612, 2014.
View at: Publisher Site | Google Scholar
C. Dahl, C. Abildgaard, R. Riber-Hansen, T. Steiniche, J. Lade-Keller, and P. Guldberg, “KIT is a frequent target for epigenetic silencing in cutaneous melanoma,” Journal of Investigative Dermatology, vol. 135, no. 2, pp. 516–524, 2014.
View at: Publisher Site | Google Scholar
Y. Isabel Zhu and J. E. Fitzpatrick, “Expression of c-kit (CD117) in Spitz nevus and malignant melanoma,” Journal of Cutaneous Pathology, vol. 33, no. 1, pp. 33–37, 2006.
View at: Publisher Site | Google Scholar
A. L. R. de Ávila, A. C. V. Krepischi, L. F. Moredo et al., “Germline CDKN2A mutations in Brazilian patients of hereditary cutaneous melanoma,” Familial Cancer, vol. 13, no. 4, pp. 645–649, 2014.
View at: Publisher Site | Google Scholar
A. H. J. F. M. Campos, A. A. Silva, L. D. D. C. Mota et al., “The value of a tumor bank in the development of cancer research in Brazil: 13 years of experience at the A C Camargo Hospital,” Biopreservation and Biobanking, vol. 10, no. 2, pp. 168–173, 2012.
View at: Publisher Site | Google Scholar
C. A. Brohem, R. R. Massaro, M. Tiago et al., “Proteasome inhibition and ROS generation by 4-nerolidylcatechol induces melanoma cell death,” Pigment Cell & Melanoma Research, vol. 25, no. 3, pp. 354–369, 2012.
View at: Publisher Site | Google Scholar
D. Wang, L. Yan, Q. Hu et al., “IMA: an R package for high-throughput analysis of Illumina's 450K Infinium methylation data,” Bioinformatics, vol. 28, no. 5, pp. 729–730, 2012.
View at: Publisher Site | Google Scholar
T. J. Morris, L. M. Butcher, A. Feber et al., “ChAMP: 450k chip analysis methylation pipeline,” Bioinformatics, vol. 30, no. 3, pp. 428–430, 2014.
View at: Publisher Site | Google Scholar
H. Heyn and M. Esteller, “DNA methylation profiling in the clinic: applications and challenges,” Nature reviews. Genetics, vol. 13, no. 10, pp. 679–692, 2012.
View at: Publisher Site | Google Scholar
L. Sigalotti, E. Fratta, E. Bidoli et al., “Methylation levels of the ‘long interspersed nucleotide element-1’ repetitive sequences predict survival of melanoma patients,” Journal of Translational Medicine, vol. 9, article 78, 2011.
View at: Publisher Site | Google Scholar
É. S. de Araújo, A. Y. Kashiwabara, M. I. Achatz et al., “LINE-1 hypermethylation in peripheral blood of cutaneous melanoma patients is associated with metastasis,” Melanoma Research, vol. 25, no. 2, pp. 173–177, 2015.
View at: Publisher Site | Google Scholar
A. Tanemura, A. M. Terando, M.-S. Sim et al., “CpG Island methylator phenotype predicts progression of malignant melanoma,” Clinical Cancer Research, vol. 15, no. 5, pp. 1801–1807, 2009.
View at: Publisher Site | Google Scholar
Y. You, L. Ma, M. You et al., “TSLC1 gene silencing in cutaneous melanoma,” Melanoma Research, vol. 20, no. 3, pp. 179–183, 2010.
View at: Publisher Site | Google Scholar
P. Schraml, A. von Teichman, D. Mihic-Probst et al., “Predictive value of the MGMT promoter methylation status in metastatic melanoma patients receiving first-line temozolomide plus bevacizumab in the trial SAKK 50/07,” Oncology Reports, vol. 28, no. 2, pp. 654–658, 2012.
View at: Publisher Site | Google Scholar
Y. Cheng, J. W. Kim, W. Liu et al., “Genetic and epigenetic inactivation of TNFRSFI0C in human prostate cancer,” Prostate, vol. 69, no. 3, pp. 327–335, 2009.
View at: Publisher Site | Google Scholar
H. C. Pieper, B. O. Evert, O. Kaut, P. F. Riederer, A. Waha, and U. Wüllner, “Different methylation of the TNF-alpha promoter in cortex and substantia nigra: implications for selective neuronal vulnerability,” Neurobiology of Disease, vol. 32, no. 3, pp. 521–527, 2008.
View at: Publisher Site | Google Scholar
A. Potapova, A. M. Hoffman, A. K. Godwin, T. Al-Saleem, and P. Cairns, “Promoter hypermethylation of the PALB2 susceptibility gene in inherited and sporadic breast and ovarian cancer,” Cancer Research, vol. 68, no. 4, pp. 998–1002, 2008.
View at: Publisher Site | Google Scholar
R. Tuominen, R. Jewell, J. J. van den Oord et al., “MGMT promoter methylation is associated with temozolomide response and prolonged progression-free survival in disseminated cutaneous melanoma,” International Journal of Cancer, 2014.
View at: Publisher Site | Google Scholar
P. A. van der Veiden, J. A. W. Metzelaar-Blok, W. Bergman et al., “Promoter hypermethylation: a common cause of reduced p16INK4a expression in uveal melanoma,” Cancer Research, vol. 61, no. 13, pp. 5303–5306, 2001.
View at: Google Scholar
H. Zhang, X. Weng, J. Ye, L. He, D. Zhou, and Y. Liu, “Promoter hypermethylation of TERT is associated with hepatocellular carcinoma in the Han Chinese population,” Clinics and Research in Hepatology and Gastroenterology, pp. 1–10, 2015.
View at: Publisher Site | Google Scholar
L. E. Moore, R. M. Pfeiffer, C. Poscablo et al., “Genomic DNA hypomethylation as a biomarker for bladder cancer susceptibility in the Spanish Bladder Cancer Study: a case-control study,” The Lancet Oncology, vol. 9, no. 4, pp. 359–366, 2008.
View at: Publisher Site | Google Scholar
J. M. Flanagan, M. Munoz-Alegre, S. Henderson et al., “Gene-body hypermethylation of ATM in peripheral blood DNA of bilateral breast cancer patients,” Human Molecular Genetics, vol. 18, no. 7, pp. 1332–1342, 2009.
View at: Publisher Site | Google Scholar
A. E. Teschendorff, U. Menon, A. Gentry-Maharaj et al., “An epigenetic signature in peripheral blood predicts active ovarian cancer,” PLoS ONE, vol. 4, no. 12, Article ID e8274, 2009.
View at: Publisher Site | Google Scholar
K. S. Pedersen, W. R. Bamlet, A. L. Oberg et al., “Leukocyte DNA methylation signature differentiates pancreatic cancer patients from healthy controls,” PLoS ONE, vol. 6, no. 3, Article ID e18223, 2011.
View at: Publisher Site | Google Scholar
C. E. Nestor, R. Ottaviano, D. Reinhardt et al., “Rapid reprogramming of epigenetic and transcriptional profiles in mammalian culture systems,” Genome Biology, vol. 16, article 11, 2015.
View at: Publisher Site | Google Scholar
T. Mikeska, C. Bock, H. Do, and A. Dobrovic, “DNA methylation biomarkers in cancer: progress towards clinical implementation,” Expert Review of Molecular Diagnostics, vol. 12, no. 5, pp. 473–487, 2012.
View at: Publisher Site | Google Scholar
M. Cankovic, M. N. Nikiforova, M. Snuderl et al., “The role of MGMT testing in clinical practice: a report of the association for molecular pathology,” The Journal of Molecular Diagnostics, vol. 15, no. 5, pp. 539–555, 2013.
View at: Publisher Site | Google Scholar
A. Inno, G. Fanetti, M. Di Bartolomeo et al., “Role of MGMT as biomarker in colorectal cancer,” World Journal of Clinical Cases, vol. 2, no. 12, pp. 835–839, 2014.
View at: Publisher Site | Google Scholar
A. Marini, A. Mirmohammadsadegh, S. Nambiar, A. Gustrau, T. Ruzicka, and U. R. Hengge, “Epigenetic inactivation of tumor suppressor genes in serum of patients with cutaneous melanoma,” Journal of Investigative Dermatology, vol. 126, no. 2, pp. 422–431, 2006.
View at: Publisher Site | Google Scholar
M. Rastetter, U. Schagdarsurengin, C. Lahtz et al., “Frequent intra-tumoural heterogeneity of promoter hypermethylation in malignant melanoma,” Histology and histopathology, vol. 22, no. 9, pp. 1005–1015, 2007.
View at: Google Scholar
M. Kulis, A. C. Queirós, R. Beekman, and J. I. Martín-Subero, “Intragenic DNA methylation in transcriptional regulation, normal differentiation and cancer,” Biochimica et Biophysica Acta, vol. 1829, no. 11, pp. 1161–1174, 2013.
View at: Publisher Site | Google Scholar
D. Jjingo, A. B. Conley, S. V. Yi, V. V. Lunyak, and I. King Jordan, “On the presence and role of human gene-body DNA methylation,” Oncotarget, vol. 3, no. 4, pp. 462–474, 2012.
View at: Google Scholar
Y. Cheli, M. Ohanna, R. Ballotti, and C. Bertolotto, “Fifteen-year quest for microphthalmia-associated transcription factor target genes,” Pigment Cell and Melanoma Research, vol. 23, no. 1, pp. 27–40, 2010.
View at: Publisher Site | Google Scholar
M. H. Law, S. MacGregor, and N. K. Hayward, “Melanoma genetics: recent findings take us beyond well-traveled pathways,” Journal of Investigative Dermatology, vol. 132, no. 7, pp. 1763–1774, 2012.
View at: Publisher Site | Google Scholar
M. Ennen, C. Keime, D. Kobi et al., “Single-cell gene expression signatures reveal melanoma cell heterogeneity,” Oncogene, 2014.
View at: Publisher Site | Google Scholar
M. L. Hartman and M. Czyz, “MITF in melanoma: mechanisms behind its expression and activity,” Cellular and Molecular Life Sciences, vol. 72, no. 7, pp. 1249–1260, 2015.
View at: Publisher Site | Google Scholar
M. Lauss, R. Haq, H. Cirenajwis et al., “Genome-wide DNA methylation analysis in melanoma reveals the importance of CpG methylation in MITF regulation,” Journal of Investigative Dermatology, 2015.
View at: Publisher Site | Google Scholar
J. Stubblefield and B. Kelly, “Melanoma in non-caucasian populations,” Surgical Clinics of North America, vol. 94, pp. 1115–1126, 2014.
View at: Google Scholar
S. J. Furney, S. Turajlic, G. Stamp et al., “The mutational burden of acral melanoma revealed by whole-genome sequencing and comparative analysis,” Pigment Cell & Melanoma Research, vol. 27, no. 5, pp. 835–838, 2014.
View at: Publisher Site | Google Scholar
C. Beadling, E. Jacobson-Dunlop, F. S. Hodi et al., “KIT gene mutations and copy number in melanoma subtypes,” Clinical Cancer Research, vol. 14, no. 21, pp. 6821–6828, 2008.
View at: Publisher Site | Google Scholar
J. A. Curtin, K. Busam, D. Pinkel, and B. C. Bastian, “Somatic activation of KIT in distinct subtypes of melanoma,” Journal of Clinical Oncology, vol. 24, no. 26, pp. 4340–4346, 2006.
View at: Publisher Site | Google Scholar
H. Tsao, L. Chin, L. A. Garraway, and D. E. Fisher, “Melanoma: from mutations to medicine,” Genes & Development, vol. 26, no. 11, pp. 1131–1155, 2012.
View at: Publisher Site | Google Scholar

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Copyright © 2015 Érica S. S. de Araújo et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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