A genomic screen for genes upregulated by demethylation and histone deacetylase inhibition in human colorectal cancer

Aberrant hypermethylation of gene promoters is a major mechanism associated with inactivation of tumor-suppressor genes in cancer. We previously showed this transcriptional silencing to be mediated by both methylation and histone deacetylase activity, with methylation being dominant. Here, we have used cDNA microarray analysis to screen for genes that are epigenetically silenced in human colorectal cancer. By screening over 10,000 genes, we show that our approach can identify a substantial number of genes with promoter hypermethylation in a given cancer; these are distinct from genes with unmethylated promoters, for which increased expression is produced by histone deacetylase inhibition alone. Many of the hypermethylated genes we identified have high potential for roles in tumorigenesis by virtue of their predicted function and chromosome position.We also identified a group of genes that are preferentially hypermethylated in colorectal cancer and gastric cancer. One of these genes, SFRP1, belongs to a gene family; we show that hypermethylation of four genes in this family occurs very frequently in colorectal cancer, providing for (i) a unique potential mechanism for loss of tumor-suppressor gene function and (ii) construction of a molecular marker panel that could detect virtually all colorectal cancer.

[1]  H Glatt,et al.  Characterization of two novel protocadherins (PCDH8 and PCDH9) localized on human chromosome 13 and mouse chromosome 14. , 1998, Genomics.

[2]  J. Nathans,et al.  A family of secreted proteins contains homology to the cysteine-rich ligand-binding domain of frizzled receptors. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[3]  M. Caligiuri,et al.  Aberrant CpG-island methylation has non-random and tumour-type–specific patterns , 2000, Nature Genetics.

[4]  S. Clark,et al.  Loss of annexin II heavy and light chains in prostate cancer and its precursors. , 2001, Cancer research.

[5]  松尾 恵太郎.  Association between polymorphisms of folate- and methionine-metabolizing enzymes and susceptibility to malignant lymphoma , 2002 .

[6]  J. Herman,et al.  Synergy of demethylation and histone deacetylase inhibition in the re-expression of genes silenced in cancer , 1999, Nature Genetics.

[7]  S. Saitoh,et al.  Imprinting in Prader-Willi and Angelman syndromes. , 1998, Trends in genetics : TIG.

[8]  S. Baylin,et al.  Dnmt3a and Dnmt3b Are Transcriptional Repressors That Exhibit Unique Localization Properties to Heterochromatin* , 2001, The Journal of Biological Chemistry.

[9]  S. Baylin,et al.  Identification of differentially methylated sequences in colorectal cancer by methylated CpG island amplification. , 1999, Cancer research.

[10]  J. Varley,et al.  Isolation and characterization of a human homologue of the latrophilin gene from a region of 1p31.1 implicated in breast cancer , 1998, Oncogene.

[11]  J. Yokota,et al.  Identification of a 428-kb homozygously deleted region disrupting the SEZ6L gene at 22q12.1 in a lung cancer cell line , 2000, Oncogene.

[12]  M. Mahadevappa,et al.  SARPs: a family of secreted apoptosis-related proteins. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[13]  O. Kocher,et al.  Differential expression of frpHE: a novel human stromal protein of the secreted frizzled gene family, during the endometrial cycle and malignancy. , 1999, Laboratory investigation; a journal of technical methods and pathology.

[14]  P. Vezzoni,et al.  A low-copy repeat in Xq26 represents a novel putatively prenylated protein gene (CXX1) and its pseudogenes (DXS9914, DXS9915, and DXS9916). , 1997, Genomics.

[15]  J. Herman,et al.  Methylation-specific PCR: a novel PCR assay for methylation status of CpG islands. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[16]  M. Frommer,et al.  CpG islands in vertebrate genomes. , 1987, Journal of molecular biology.

[17]  J. Herman,et al.  Methylation-associated silencing of the tissue inhibitor of metalloproteinase-3 gene suggest a suppressor role in kidney, brain, and other human cancers. , 1999, Cancer research.

[18]  H. Varmus,et al.  Purification and molecular cloning of a secreted, Frizzled-related antagonist of Wnt action. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[19]  W. Birchmeier,et al.  Functional interaction of an axin homolog, conductin, with beta-catenin, APC, and GSK3beta. , 1998, Science.

[20]  S. Tsukita,et al.  Where is APC going? , 2001, The Journal of cell biology.

[21]  C. Nguyen,et al.  Differential expression assay of chromosome arm 8p genes identifies Frizzled-related (FRP1/FRZB) and Fibroblast Growth Factor Receptor 1 (FGFR1) as candidate breast cancer genes , 1999, Oncogene.

[22]  Peter L. Jones,et al.  DNMT1 forms a complex with Rb, E2F1 and HDAC1 and represses transcription from E2F-responsive promoters , 2000, Nature Genetics.

[23]  J. Herman,et al.  DNA hypermethylation in tumorigenesis: epigenetics joins genetics. , 2000, Trends in genetics : TIG.

[24]  Q. Wei,et al.  Methylenetetrahydrofolate reductase polymorphisms increase risk of esophageal squamous cell carcinoma in a Chinese population. , 2001, Cancer research.

[25]  C Eng,et al.  Excessive CpG island hypermethylation in cancer cell lines versus primary human malignancies. , 2001, Human molecular genetics.

[26]  P. Campochiaro,et al.  Cloning and characterization of a secreted frizzled-related protein that is expressed by the retinal pigment epithelium. , 1999, Human molecular genetics.

[27]  G. Liang,et al.  Identification and characterization of differentially methylated regions of genomic DNA by methylation-sensitive arbitrarily primed PCR. , 1997, Cancer research.

[28]  J. Herman,et al.  A gene hypermethylation profile of human cancer. , 2001, Cancer research.

[29]  T. Kouzarides,et al.  Dnmt3a binds deacetylases and is recruited by a sequence‐specific repressor to silence transcription , 2001, The EMBO journal.

[30]  P. Laird,et al.  COBRA: a sensitive and quantitative DNA methylation assay. , 1997, Nucleic acids research.

[31]  V. Gerke,et al.  Primary structure of human, chicken, and Xenopus laevis p11, a cellular ligand of the Src-kinase substrate, annexin II. , 1991, Gene.

[32]  W. Heston.  Characterization and glutamyl preferring carboxypeptidase function of prostate specific membrane antigen: a novel folate hydrolase. , 1997, Urology.

[33]  W. Gerald,et al.  Inactivation of the apoptosis effector Apaf-1 in malignant melanoma , 2001, Nature.

[34]  Luke Hughes-Davies,et al.  DNA methyltransferase Dnmt1 associates with histone deacetylase activity , 2000, Nature Genetics.

[35]  S. Baylin,et al.  DNMT1 binds HDAC2 and a new co-repressor, DMAP1, to form a complex at replication foci , 2000, Nature Genetics.

[36]  D. Birnbaum,et al.  WNT pathway and mammary carcinogenesis: Loss of expression of candidate tumor suppressor gene SFRP1 in most invasive carcinomas except of the medullary type , 2001, Oncogene.

[37]  J. Herman,et al.  CpG island methylator phenotype in colorectal cancer. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[38]  Hans Clevers,et al.  Activation of β-Catenin-Tcf Signaling in Colon Cancer by Mutations in β-Catenin or APC , 1997, Science.

[39]  S. Baylin,et al.  p53 activates expression of HIC-1, a new candidate tumour suppressor gene on 17p13.3 , 1995, Nature Genetics.

[40]  M. Toyota,et al.  Aberrant methylation of the Cyclooxygenase 2 CpG island in colorectal tumors. , 2000, Cancer research.

[41]  D. Ledbetter,et al.  Deletions of a differentially methylated CpG island at the SNRPN gene define a putative imprinting control region , 1994, Nature Genetics.