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Profiling epigenetic inactivation of tumor suppressor genes in tumors and plasma from cutaneous melanoma patients

Oncogene volume 23, pages 4014–4022 (2004)Cite this article

Abstract

Aberrant methylation of CpG islands in promoter regions of tumor suppressor genes (TSG) has been demonstrated in epithelial origin tumors. However, the methylation profiling of tumor-related gene promoter regions in cutaneous melanoma tumors has not been reported. Seven known or candidate TSGs that are frequently hypermethylated in carcinomas were assessed by methylation-specific polymerase chain reaction (MSP) in 15 melanoma cell lines and 130 cutaneous melanoma tumors. Four TSGs were frequently hypermethylated in 86 metastatic tumor specimens: retinoic acid receptor-β2 (RAR-β2) (70%), RAS association domain family protein 1A (RASSF1A) (57%), and O6-methylguanine DNA methylatransferase (MGMT) (34%), and death-associated protein kinase (DAPK) (19%). Hypermethylation of MGMT, RASSF1A, and DAPK was significantly lower in primary melanomas (n=20) compared to metastatic melanomas. However, hypermethylation of RAR-β2 was 70% in both primary and metastatic melanomas. Cell lines had hypermethylation profiles similar to those of metastatic melanomas. The analysis of these four markers of metastatic tumors demonstrated that 97% had ⩾1 gene(s) and 59% had ⩾2 genes hypermethylated. The methylation of genes was verified by bisulfite sequencing. The mRNA transcripts could be re-expressed in melanoma cell lines having hypermethylated genes following treatment with 5′-aza 2′-deoxycytidine (5Aza-dC). Analysis of melanoma patients’ plasma (preoperative blood; n=31) demonstrated circulating hypermethylated MGMT, RAR-β2, and RASSF1A DNA for at least one of the markers in 29% of the patients. Our findings indicate that the incidence of TSG hypermethylation increases during tumor progression. Methylation of TSG may play a significant role in cutaneous melanoma progression.

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Abbreviations

ATRA:

all-trans-retinoic acid

5Aza-dC:

5′-aza 2′-deoxycytidine

GAPDH:

glyceraldehyde-3-phosphate dehydrogenase

DAPK:

death-associated protein kinase

GSTP1:

glutathione S-transferase P1

MGMT:

O6-methylguanine DNA methylatransferase

MSP:

methylation-specific polymerase chain reaction

RT–PCR:

reverse transcriptase-polymerase chain reaction

RAR-β2:

retinoic acid receptor-β2

RASSF1A:

RAS association domain family protein 1A

and TSG:

tumor suppressor gene

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Acknowledgements

This work was supported in part by NIH NCI R21 CA100314, PO CA 29605 Project II, and PO CA 13917 Project II and Roy E Coates Foundation.

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Authors and Affiliations

  1. Department Molecular Oncology, John Wayne Cancer Institute at Saint John's Health Center, 2200 Santa Monica Blvd, Santa Monica, 90404, CA, USA

    Dave S B Hoon, Mia Spugnardi, Christine Kuo, Sharon K Huang & Bret Taback

  2. Division of Surgical Oncology, John Wayne Cancer Institute at Saint John's Health Center, Santa Monica, 90404, CA, USA

    Donald L Morton

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  1. Dave S B Hoon
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Correspondence to Dave S B Hoon.

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Hoon, D., Spugnardi, M., Kuo, C. et al. Profiling epigenetic inactivation of tumor suppressor genes in tumors and plasma from cutaneous melanoma patients. Oncogene 23, 4014–4022 (2004). https://doi.org/10.1038/sj.onc.1207505

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