Identification of 5 novel genes methylated in breast and other epithelial cancers.
BACKGROUND
There are several high throughput approaches to identify methylated genes in cancer. We utilized one such recently developed approach, MIRA (methylated-CpG island recovery assay) combined with CpG island arrays to identify novel genes that are epigenetically inactivated in breast cancer.


RESULTS
Using this approach we identified numerous CpG islands that demonstrated aberrant DNA methylation in breast cancer cell lines. Using a combination of COBRA and sequencing of bisulphite modified DNA, we confirmed 5 novel genes frequently methylated in breast tumours; EMILIN2, SALL1, DBC1, FBLN2 and CIDE-A. Methylation frequencies ranged from between 25% and 63% in primary breast tumours, whilst matched normal breast tissue DNA was either unmethylated or demonstrated a much lower frequency of methylation compared to malignant breast tissue DNA. Furthermore expression of the above 5 genes was shown to be restored following treatment with a demethylating agent in methylated breast cancer cell lines. We have expanded this analysis across three other common epithelial cancers (lung, colorectal, prostate). We demonstrate that the above genes show varying levels of methylation in these cancers. Lastly and most importantly methylation of EMILIN2 was associated with poorer clinical outcome in breast cancer and was strongly associated with estrogen receptor as well as progesterone receptor positive breast cancers.


CONCLUSIONS
The combination of the MIRA assay with CpG island arrays is a very useful technique for identifying epigenetically inactivated genes in cancer genomes and can provide molecular markers for early cancer diagnosis, prognosis and epigenetic therapy.
Hill
Victoria K
VK
Department of Medical and Molecular Genetics, University of Birmingham, UK. f.latif@bham.ac.uk
Hesson
Luke B
LB
Dansranjavin
Temuujin
T
Dallol
Ashraf
A
Bieche
Ivan
I
Vacher
Sophie
S
Tommasi
Stella
S
Dobbins
Timothy
T
Gentle
Dean
D
Euhus
David
D
Lewis
Cheryl
C
Dammann
Reinhard
R
Ward
Robyn L
RL
Minna
John
J
Maher
Eammon R
ER
Pfeifer
Gerd P
GP
Latif
Farida
F
D001943:Breast Neoplasms / Q000235:genetics* / Q000473:pathology; D045744:Cell Line, Tumor; D015179:Colorectal Neoplasms / Q000235:genetics; D019175:DNA Methylation / Q000235:genetics*; D004848:Epithelium / Q000378:metabolism / Q000473:pathology*; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D052138:Genes, Neoplasm / Q000235:genetics*; D015894:Genome, Human / Q000235:genetics; D006023:Glycoproteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008297:Male; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D016019:Survival Analysis
C426245:EMILIN2 protein, human; D006023:Glycoproteins; D012333:RNA, Messenger