[Identification of methylation related genes from laser capture microdissected colon samples during investigation of adenoma-carcinoma sequence].
Changes of the DNA methylation pattern are proven to be an important process during tumorigenesis. This event can occur in several manners in the tumor microenvironment and there are still not any effective and high-throughput methods for genome-wide analysis of this phenomenon.


OBJECTIVE
Our aim was to identify colorectal cancer development and progression specific marker genes regulated by DNA methylation using gene expression analysis. In this study we present a gene expression-based method combined with a cell culture model, which can be used for a genome-wide analysis of the methylation events during the colorectal tumorigenesis.


METHODS
Genes, which expression increased after the demethylation were determined in HT-29 colon adenocarcinoma cells treated with 10 microM 5-aza-2'-deoxycitidine. In parallel, 5000 epithelial cells were collected with laser microdissection (LCM) from normal, adenoma and tumorous colonic samples. The genes with gradually decreasing expression along the adenoma-carcinoma sequence were identified. By comparing the two groups, the transcripts, which are supposed to be regulated by methylation, could be determined. Finally, the identified gene set was validated on independent samples using RT-PCR.


CONCLUSIONS
The regulation of the identified genes showing decreased expression during the adenoma-carcinoma sequence, can be associated with DNA methylation. On the basis of our results, the set of genes including tumorsuppressors can be determined genome-widely, which can be key factors in the formation and the prognosis of the disease. The identified genes showing colorectal cancer specific methylation pattern can be potential therapeutic targets in the future.
Spisák
Sándor
S
Magyar Tudományos Akadémia, Molekuláris Medicina Kutatócsoport, Budapest. spisak@abc.hu
Kalmár
Alexandra
A
Galamb
Orsolya
O
Sipos
Ferenc
F
Wichmann
Barnabás
B
Molnár
Béla
B
Tulassay
Zsolt
Z
D000236:Adenoma / Q000175:diagnosis / Q000235:genetics*; D002277:Carcinoma / Q000175:diagnosis / Q000235:genetics*; D002471:Cell Transformation, Neoplastic / Q000235:genetics*; D003110:Colonic Neoplasms / Q000175:diagnosis* / Q000235:genetics*; D019175:DNA Methylation; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D007834:Lasers; D046228:Microarray Analysis; D042282:Microdissection / Q000379:methods; D011401:Promoter Regions, Genetic; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction