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Detection and interpretation of altered methylation patterns in cancer cells

Nature Reviews Cancer volume 5, pages 223–231 (2005)Cite this article

Abstract

Epigenetic alterations, such as abnormal DNA-methylation patterns, are associated with many human tumour types. New techniques have been developed to perform genome-wide screening for alterations in DNA-methylation patterns, not only to identify tumour-suppressor genes, but also to find patterns that can be used in diagnosis and prognosis. However, interpretation of differential methylation has proven difficult because the significance of methylation alterations depends on the genomic region, and functions of CpG islands at specific sites have not been fully clarified. What techniques can be used to identify new tumour suppressors and diagnostic markers?

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Figure 1: CpG sites and CpG islands.
Figure 2: Regions within a promoter CpG island and a core region.
Figure 3: Distribution of methylation and its effect on gene expression.

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References

  1. Jones, P. A. & Baylin, S. B. The fundamental role of epigenetic events in cancer. Nature Rev. Genet. 3, 415–428 (2002).

    Article  CAS  PubMed  Google Scholar 

  2. Ohlsson, R., Renkawitz, R. & Lobanenkov, V. CTCF is a uniquely versatile transcription regulator linked to epigenetics and disease. Trends Genet. 17, 520–527 (2001).

    Article  CAS  PubMed  Google Scholar 

  3. Hark, A. T. et al. CTCF mediates methylation-sensitive enhancer-blocking activity at the H19/Igf2 locus. Nature 405, 486–489 (2000).

    Article  CAS  PubMed  Google Scholar 

  4. Bell, A. C. & Felsenfeld, G. Methylation of a CTCF-dependent boundary controls imprinted expression of the Igf2 gene. Nature 405, 482–485 (2000).

    Article  CAS  PubMed  Google Scholar 

  5. Jaenisch, R. & Bird, A. Epigenetic regulation of gene expression: how the genome integrates intrinsic and environmental signals. Nature Genet. 33 (Suppl.), 245–254 (2003).

    Article  CAS  PubMed  Google Scholar 

  6. Egger, G., Liang, G., Aparicio, A. & Jones, P. A. Epigenetics in human disease and prospects for epigenetic therapy. Nature 429, 457–463 (2004).

    Article  CAS  PubMed  Google Scholar 

  7. Takai, D. & Jones, P. A. Comprehensive analysis of CpG islands in human chromosomes 21 and 22. Proc. Natl Acad. Sci. USA 99, 3740–3745 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. De Smet, C., Lurquin, C., Lethe, B., Martelange, V. & Boon, T. DNA methylation is the primary silencing mechanism for a set of germ line- and tumor-specific genes with a CpG-rich promoter. Mol. Cell. Biol. 19, 7327–7335 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Futscher, B. W. et al. Role for DNA methylation in the control of cell type specific maspin expression. Nature Genet. 31, 175–179 (2002).

    Article  CAS  PubMed  Google Scholar 

  10. Bird, A. DNA methylation patterns and epigenetic memory. Genes Dev. 16, 6–21 (2002).

    Article  CAS  PubMed  Google Scholar 

  11. Feinberg, A. P. & Vogelstein, B. Hypomethylation distinguishes genes of some human cancers from their normal counterparts. Nature 301, 89–92 (1983).

    Article  CAS  PubMed  Google Scholar 

  12. Kaneda, A. et al. Frequent hypomethylation in multiple promoter CpG islands is associated with global hypomethylation, but not with frequent promoter hypermethylation. Cancer Sci. 95, 58–64 (2004).

    Article  CAS  PubMed  Google Scholar 

  13. Chen, R. Z., Pettersson, U., Beard, C., Jackson-Grusby, L. & Jaenisch, R. DNA hypomethylation leads to elevated mutation rates. Nature 395, 89–93 (1998).

    Article  CAS  PubMed  Google Scholar 

  14. Gaudet, F. et al. Induction of tumors in mice by genomic hypomethylation. Science 300, 489–492 (2003).

    Article  CAS  PubMed  Google Scholar 

  15. Kanai, Y. et al. Aberrant DNA methylation on chromosome 16 is an early event in hepatocarcinogenesis. Jpn J. Cancer Res. 87, 1210–1217 (1996).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Yoshikawa, H. et al. SOCS-1, a negative regulator of the JAK/STAT pathway, is silenced by methylation in human hepatocellular carcinoma and shows growth-suppression activity. Nature Genet. 28, 29–35 (2001).

    CAS  PubMed  Google Scholar 

  17. Suzuki, H. et al. Epigenetic inactivation of SFRP genes allows constitutive WNT signaling in colorectal cancer. Nature Genet. 36, 417–422 (2004).

    Article  CAS  PubMed  Google Scholar 

  18. Kaneda, A. et al. Lysyl oxidase is a tumor suppressor gene inactivated by methylation and loss of heterozygosity in human gastric cancers. Cancer Res. 64, 6410–6415 (2004).

    Article  CAS  PubMed  Google Scholar 

  19. Esteller, M. et al. Detection of aberrant promoter hypermethylation of tumor suppressor genes in serum DNA from non-small cell lung cancer patients. Cancer Res. 59, 67–70 (1999).

    CAS  PubMed  Google Scholar 

  20. Wong, I. H. et al. Detection of aberrant p16 methylation in the plasma and serum of liver cancer patients. Cancer Res. 59, 71–73 (1999).

    CAS  PubMed  Google Scholar 

  21. Laird, P. W. The power and the promise of DNA methylation markers. Nature Rev. Cancer 3, 253–266 (2003).

    Article  CAS  Google Scholar 

  22. Belinsky, S. A. Gene-promoter hypermethylation as a biomarker in lung cancer. Nature Rev. Cancer 4, 707–717 (2004).

    Article  CAS  Google Scholar 

  23. Ohgane, J., Hattori, N., Oda, M., Tanaka, S. & Shiota, K. Differentiation of trophoblast lineage is associated with DNA methylation and demethylation. Biochem. Biophys Res. Commun. 290, 701–706 (2002).

    Article  CAS  PubMed  Google Scholar 

  24. Kaneda, A., Kaminishi, M., Yanagihara, K., Sugimura, T. & Ushijima, T. Identification of silencing of nine genes in human gastric cancers. Cancer Res. 62, 6645–6650 (2002).

    PubMed  Google Scholar 

  25. Hayatsu, H., Wataya, Y., Kai, K. & Iida, S. Reaction of sodium bisulfite with uracil, cytosine, and their derivatives. Biochemistry 9, 2858–2865 (1970).

    Article  CAS  PubMed  Google Scholar 

  26. Hatada, I., Hayashizaki, Y., Hirotsune, S., Komatsubara, H. & Mukai, T. A genomic scanning method for higher organisms using restriction sites as landmarks. Proc. Natl Acad. Sci. USA. 88, 9523–9527 (1991).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Lisitsyn, N., Lisitsyn, N. & Wigler, M. Cloning the differences between two complex genomes. Science 259, 946–951 (1993).

    Article  CAS  PubMed  Google Scholar 

  28. Welsh, J., Petersen, C. & McClelland, M. Polymorphisms generated by arbitrarily primed PCR in the mouse: application to strain identification and genetic mapping. Nucleic Acids Res. 19, 303–306 (1991).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Kawai, J. et al. Methylation profiles of genomic DNA of mouse developmental brain detected by restriction landmark genomic scanning (RLGS) method. Nucleic Acids Res. 21, 5604–5608 (1993).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Wimmer, K. et al. Combined restriction landmark genomic scanning and virtual genome scans identify a novel human homeobox gene, ALX3, that is hypermethylated in neuroblastoma. Genes Chromosom. Cancer 33, 285–294 (2002).

    Article  CAS  PubMed  Google Scholar 

  31. Ushijima, T. et al. Establishment of methylation-sensitive-representational difference analysis and isolation of hypo- and hypermethylated genomic fragments in mouse liver tumors. Proc. Natl Acad. Sci. USA 94, 2284–2289 (1997).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Kaneda, A., Takai, D., Kaminishi, M., Okochi, E. & Ushijima, T. Methylation-sensitive representational difference analysis and its application to cancer research. Ann. NY Acad. Sci. 983, 131–141 (2003).

    Article  CAS  PubMed  Google Scholar 

  33. Gonzalgo, M. L. et al. Identification and characterization of differentially methylated regions of genomic DNA by methylation-sensitive arbitrarily primed PCR. Cancer Res. 57, 594–599 (1997).

    CAS  PubMed  Google Scholar 

  34. Huang, T. H. et al. Identification of DNA methylation markers for human breast carcinomas using the methylation-sensitive restriction fingerprinting technique. Cancer Res. 57, 1030–1034 (1997).

    CAS  PubMed  Google Scholar 

  35. Toyota, M. et al. Identification of differentially methylated sequences in colorectal cancer by methylated CpG island amplification. Cancer Res. 59, 2307–2312 (1999).

    CAS  PubMed  Google Scholar 

  36. Huang, T. H., Perry, M. R. & Laux, D. E. Methylation profiling of CpG islands in human breast cancer cells. Hum. Mol. Genet. 8, 459–470 (1999).

    Article  CAS  PubMed  Google Scholar 

  37. Yan, P. S. et al. CpG island arrays: an application toward deciphering epigenetic signatures of breast cancer. Clin. Cancer Res. 6, 1432–1438 (2000).

    CAS  PubMed  Google Scholar 

  38. Shi, H. et al. Expressed CpG island sequence tag microarray for dual screening of DNA hypermethylation and gene silencing in cancer cells. Cancer Res. 62, 3214–3220 (2002).

    CAS  PubMed  Google Scholar 

  39. Shi, H. et al. Triple analysis of the cancer epigenome: an integrated microarray system for assessing gene expression, DNA methylation, and histone acetylation. Cancer Res. 63, 2164–2171 (2003).

    CAS  PubMed  Google Scholar 

  40. Adorjan, P. et al. Tumour class prediction and discovery by microarray-based DNA methylation analysis. Nucleic Acids Res. 30, e21 (2002).

    Article  PubMed  PubMed Central  Google Scholar 

  41. Novik, K. L. et al. Epigenomics: genome-wide study of methylation phenomena. Curr. Issues Mol. Biol. 4, 111–128 (2002).

    CAS  PubMed  Google Scholar 

  42. Gitan, R. S., Shi, H., Chen, C. M., Yan, P. S. & Huang, T. H. Methylation-specific oligonucleotide microarray: a new potential for high-throughput methylation analysis. Genome Res. 12, 158–164 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Ballestar, E. et al. Methyl-CpG binding proteins identify novel sites of epigenetic inactivation in human cancer. EMBO J. 22, 6335–6345 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Kondo, Y., Shen, L., Yan, P. S., Huang, T. H. & Issa, J. P. Chromatin immunoprecipitation microarrays for identification of genes silenced by histone H3 lysine 9 methylation. Proc. Natl Acad. Sci. USA 101, 7398–7403 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Suzuki, H. et al. A genomic screen for genes upregulated by demethylation and histone deacetylase inhibition in human colorectal cancer. Nature Genet. 31, 141–149 (2002).

    Article  CAS  PubMed  Google Scholar 

  46. Gonzalgo, M. L. et al. The role of DNA methylation in expression of the p19/p16 locus in human bladder cancer cell lines. Cancer Res. 58, 1245–1252 (1998).

    CAS  PubMed  Google Scholar 

  47. Abe, M. et al. Cloning of the 5′ upstream region of the rat p16 gene and its role in silencing. Jpn J. Cancer Res. 93, 1100–1106 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  48. Deng, G., Chen, A., Hong, J., Chae, H. S. & Kim, Y. S. Methylation of CpG in a small region of the hMLH1 promoter invariably correlates with the absence of gene expression. Cancer Res. 59, 2029–2033 (1999).

    CAS  PubMed  Google Scholar 

  49. Miyakura, Y. et al. Extensive methylation of hMLH1 promoter region predominates in proximal colon cancer with microsatellite instability. Gastroenterology 121, 1300–1309 (2001).

    Article  CAS  PubMed  Google Scholar 

  50. Yan, P. S. et al. Differential distribution of DNA methylation within the RASSF1A CpG island in breast cancer. Cancer Res. 63, 6178–6186 (2003).

    CAS  PubMed  Google Scholar 

  51. Hagihara, A. et al. Identification of 27 5′ CpG islands aberrantly methylated and 13 genes silenced in human pancreatic cancers. Oncogene 23, 8705–8710 (2004).

    Article  CAS  PubMed  Google Scholar 

  52. Issa, J. P. et al. Methylation of the oestrogen receptor CpG island links ageing and neoplasia in human colon. Nature Genetics 7, 536–540 (1994).

    Article  CAS  PubMed  Google Scholar 

  53. Ahuja, N., Li, Q., Mohan, A. L., Baylin, S. B. & Issa, J. P. Aging and DNA methylation in colorectal mucosa and cancer. Cancer Res. 58, 5489–5494 (1998).

    CAS  PubMed  Google Scholar 

  54. Issa, J. P., Ahuja, N., Toyota, M., Bronner, M. P. & Brentnall, T. A. Accelerated age-related CpG island methylation in ulcerative colitis. Cancer Res. 61, 3573–3577 (2001).

    CAS  PubMed  Google Scholar 

  55. Waki, T., Tamura, G., Sato, M. & Motoyama, T. Age-related methylation of tumor suppressor and tumor-related genes: an analysis of autopsy samples. Oncogene 22, 4128–4133 (2003).

    Article  CAS  PubMed  Google Scholar 

  56. Graff, J. R., Herman, J. G., Myohanen, S., Baylin, S. B. & Vertino, P. M. Mapping patterns of CpG island methylation in normal and neoplastic cells implicates both upstream and downstream regions in de novo methylation. J. Biol. Chem. 272, 22322–22329 (1997).

    Article  CAS  PubMed  Google Scholar 

  57. Song, J. Z., Stirzaker, C., Harrison, J., Melki, J. R. & Clark, S. J. Hypermethylation trigger of the glutathione-S-transferase gene (GSTP1) in prostate cancer cells. Oncogene 21, 1048–1061 (2002).

    Article  CAS  PubMed  Google Scholar 

  58. Stirzaker, C., Song, J. Z., Davidson, B. & Clark, S. J. Transcriptional gene silencing promotes DNA hypermethylation through a sequential change in chromatin modifications in cancer cells. Cancer Res. 64, 3871–3877 (2004).

    Article  CAS  PubMed  Google Scholar 

  59. De Smet, C., Loriot, A. & Boon, T. Promoter-dependent mechanism leading to selective hypomethylation within the 5′ region of gene MAGE-A1 in tumor cells. Mol. Cell. Biol. 24, 4781–4790 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  60. Ushijima, T. et al. Decreased fidelity in replicating CpG methylation patterns in cancer cells. Cancer Res. 65, 11–17 (2005).

    CAS  PubMed  Google Scholar 

  61. Nguyen, C. et al. Susceptibility of nonpromoter CpG islands to de novo methylation in normal and neoplastic cells. J. Natl Cancer Inst. 93, 1465–1472 (2001).

    Article  CAS  PubMed  Google Scholar 

  62. Ushijima, T. et al. Fidelity of the methylation pattern and its variation in the genome. Genome Res. 13, 868–874 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  63. Kaneda, A., Kaminishi, M., Nakanishi, Y., Sugimura, T. & Ushijima, T. Reduced expression of the insulin-induced protein 1 and p41 Arp2/3 complex genes in human gastric cancers. Int. J. Cancer 100, 57–62 (2002).

    Article  CAS  PubMed  Google Scholar 

  64. Bender, C. M. et al. Roles of cell division and gene transcription in the methylation of CpG islands. Mol. Cell. Biol. 19, 6690–6698 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  65. Abe, M. et al. CpG island methylator phenotype is a strong determinant of poor prognosis in neuroblastomas. Cancer Res. (in the press).

  66. Li, H. et al. SLC5A8, a sodium transporter, is a tumor suppressor gene silenced by methylation in human colon aberrant crypt foci and cancers. Proc. Natl Acad. Sci. USA 100, 8412–8417 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  67. Dai, Z. et al. Bone morphogenetic protein 3B silencing in non-small-cell lung cancer. Oncogene 23, 3521–3529 (2004).

    Article  CAS  PubMed  Google Scholar 

  68. Costello, J. F. et al. Aberrant CpG-island methylation has non-random and tumour-type-specific patterns. Nature Genet. 24, 132–138 (2000).

    Article  CAS  PubMed  Google Scholar 

  69. Takai, D. et al. Silencing of HTR1B and reduced expression of EDN1 in human lung cancers, revealed by methylation-sensitive representational difference analysis. Oncogene 20, 7505–7513 (2001).

    Article  CAS  PubMed  Google Scholar 

  70. Asada, K. et al. Reduced expression of GNA11 and silencing of MCT1 in human breast cancers. Oncology 64, 380–388 (2003).

    Article  CAS  PubMed  Google Scholar 

  71. Miyamoto, K. et al. Methylation-associated silencing of heparan sulfate D-glucosaminyl 3-O-sulfotransferase-2 (3-OST-2) in human breast, colon, lung and pancreatic cancers. Oncogene 22, 274–280 (2003).

    Article  CAS  PubMed  Google Scholar 

  72. Palmisano, W. A. et al. Aberrant promoter methylation of the transcription factor genes PAX5α and β in human cancers. Cancer Res. 63, 4620–4625 (2003).

    CAS  PubMed  Google Scholar 

  73. Toyota, M., Ho, C., Ohe-Toyota, M., Baylin, S. B. & Issa, J. P. Inactivation of CACNA1G, a T-type calcium channel gene, by aberrant methylation of its 5′ CpG island in human tumors. Cancer Res. 59, 4535–4541 (1999).

    CAS  PubMed  Google Scholar 

  74. Sato, N. et al. Discovery of novel targets for aberrant methylation in pancreatic carcinoma using high-throughput microarrays. Cancer Res. 63, 3735–3742 (2003).

    CAS  PubMed  Google Scholar 

  75. Kawakami, T., Okamoto, K., Ogawa, O. & Okada, Y. XISTunmethylated DNA fragments in male-derived plasma as a tumour marker for testicular cancer. Lancet 363, 40–42 (2004).

    Article  CAS  PubMed  Google Scholar 

  76. Herman, J. G., Graff, J. R., Myohanen, S., Nelkin, B. D. & Baylin, S. B. Methylation-specific PCR: a novel PCR assay for methylation status of CpG islands. Proc. Natl Acad. Sci. USA 93, 9821–9826 (1996).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  77. Liang, G. et al. DNA methylation differences associated with tumor tissues identified by genome scanning analysis. Genomics 53, 260–268 (1998).

    Article  CAS  PubMed  Google Scholar 

  78. Esteller, M. et al. Hypermethylation of the DNA repair gene O6-methylguanine DNA methyltransferase and survival of patients with diffuse large B-cell lymphoma. J. Natl Cancer Inst. 94, 26–32 (2002).

    Article  CAS  PubMed  Google Scholar 

  79. Esteller, M. et al. Inactivation of the DNA-repair gene MGMT and the clinical response of gliomas to alkylating agents. N. Engl. J. Med. 343, 1350–1354 (2000).

    Article  CAS  PubMed  Google Scholar 

  80. Esteller, M. & Herman, J. G. Generating mutations but providing chemosensitivity: the role of O6-methylguanine DNA methyltransferase in human cancer. Oncogene 23, 1–8 (2004).

    Article  CAS  PubMed  Google Scholar 

  81. Tada, Y. et al. The association of death-associated protein kinase hypermethylation with early recurrence in superficial bladder cancers. Cancer Res. 62, 4048–4053 (2002).

    CAS  PubMed  Google Scholar 

  82. Chan, A. O. et al. CpG island methylation in aberrant crypt foci of the colorectum. Am. J. Pathol. 160, 1823–1830 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  83. Waki, T. et al. Promoter methylation status of E-cadherin, hMLH1, and p16 genes in nonneoplastic gastric epithelia. Am. J. Pathol. 161, 399–403 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  84. Cui, H. et al. Loss of IGF2 imprinting: a potential marker of colorectal cancer risk. Science 299, 1753–1755 (2003).

    Article  CAS  PubMed  Google Scholar 

  85. Toyota, M. et al. CpG island methylator phenotype in colorectal cancer. Proc. Natl Acad. Sci. USA 96, 8681–8686 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  86. Brown, M. A. et al. The 5′ end of the BRCA1 gene lies within a duplicated region of human chromosome 17q21. Oncogene 12, 2507–2513 (1996).

    CAS  PubMed  Google Scholar 

  87. Tsuchiya, T. et al. Distinct methylation patterns of two APC gene promoters in normal and cancerous gastric epithelia. Oncogene 19, 3642–3646 (2000).

    Article  CAS  PubMed  Google Scholar 

  88. Li, W. -H., Gu, Z., Wang, H. & Nekrutenko, A. Evolutionary analyses of the human genome. Nature 409, 847–849 (2001).

    Article  CAS  PubMed  Google Scholar 

  89. Gardiner-Garden, M. & Frommer, M. CpG islands in vertebrate genomes. J. Mol. Biol. 196, 261–282 (1987).

    Article  CAS  PubMed  Google Scholar 

  90. Muller, H. M. et al. DNA methylation in serum of breast cancer patients: an independent prognostic marker. Cancer Res. 63, 7641–7645 (2003).

    PubMed  Google Scholar 

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Acknowledgements

I thank M. Abe, E. Okochi-Takada, S. Yamashita and J. Furuta at the National Cancer Center Research Institute for their critical reading of the manuscript.

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  1. the Carcinogenesis Division, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo, 104-0045, Japan

    Toshikazu Ushijima

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DATABASES

Entrez Gene

ABL

BMP3B

BRCA1

CDKN2A

CTCF

DAPK

MBD1

MBD2

MBD3

MBD4

MECP2

MGMT

MLH1

NEF3

p41ARC

PAX6

RASSF1A

RB

SFRP1

SLC5A8

SOCS1

VHL

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Ushijima, T. Detection and interpretation of altered methylation patterns in cancer cells. Nat Rev Cancer 5, 223–231 (2005). https://doi.org/10.1038/nrc1571

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