Methylation profiles of thirty four promoter-CpG islands and concordant methylation behaviours of sixteen genes that may contribute to carcinogenesis of astrocytoma.
BACKGROUND
Astrocytoma is a common aggressive intracranial tumor and presents a formidable challenge in the clinic. Association of altered DNA methylation patterns of the promoter CpG islands with the expression profile of cancer-related genes, has been found in many human tumors. Therefore, DNA methylation status as such may serve as an epigenetic biomarker for both diagnosis and prognosis of human tumors, including astrocytoma.


METHODS
We used the methylation specific PCR in conjunction with sequencing verification to establish the methylation profile of the promoter CpG island of thirty four genes in astrocytoma tissues from fifty three patients (The WHO grading: I: 14, II: 15, III: 12 and IV: 12 cases, respectively). In addition, compatible tissues (normal tissues distant from lesion) from three non-astrocytoma patients were included as the control.


RESULTS
Seventeen genes (ABL, APC, APAF1, BRCA1, CSPG2, DAPK1, hMLH1, LKB1, PTEN, p14ARF, p15INK4b, p27KIP1, p57KIP2, RASSF1C, RB1, SURVIVIN, and VHL) displayed a uniformly unmethylated pattern in all the astrocytoma and non-astrocytoma tissues examined. However, the MAGEA1 gene that was inactivated and hypermethylated in non-astrocytoma tissues, was partially demethylated in 24.5% of the astrocytoma tissues (co-existence of the hypermethylated and demethylated alleles). Of the astrocytoma associated hypermethylated genes, the methylation pattern of the CDH13, cyclin a1, DBCCR1, EPO, MYOD1, and p16INK4a genes changed in no more than 5.66% (3/53) of astrocytoma tissues compared to non-astrocytoma controls, while the RASSF1A, p73, AR, MGMT, CDH1, OCT6, MT1A, WT1, and IRF7 genes were more frequently hypermethylated in 69.8%, 47.2%, 41.5%, 35.8%, 32%, 30.2%, 30.2%, 30.2% and 26.4% of astrocytoma tissues, respectively. Demethylation mediated inducible expression of the CDH13, MAGEA1, MGMT, p73 and RASSF1A genes was established in an astrocytoma cell line (U251), demonstrating that expression of these genes is likely regulated by DNA methylation. AR gene hypermethylation was found exclusively in female patients (22/27, 81%, 0/26, 0%, P < 0.001), while the IRF7 gene hypermethylation preferentially occurred in the male counterparts (11/26, 42.3% to 3/27, 11%, P < 0.05). Applying the mathematic method "the Discovery of Association Rules", we have identified groups consisting of up to three genes that more likely display the altered methylation patterns in concert in astrocytoma.


CONCLUSIONS
Of the thirty four genes examined, sixteen genes exhibited astrocytoma associated changes in the methylation profile. In addition to the possible pathological significance, the established concordant methylation profiles of the subsets consisting of two to three target genes may provide useful clues to the development of the useful prognostic as well as diagnostic assays for astrocytoma.
Yu
Jian
J
Cancer Epigenetics and Gene Therapy, State-Key Laboratory for Oncogenes and Related Genes, Shanghai Cancer Institute, Shanghai Jiao Tong University, LN 2200/25, Xie-Tu Road, Shanghai 200032, China. gyujian@hotmail.com
Zhang
Hongyu
H
Gu
Jun
J
Lin
Song
S
Li
Junhua
J
Lu
Wei
W
Wang
Yifei
Y
Zhu
Jingde
J
D000293:Adolescent; D000328:Adult; D000951:Antigens, Neoplasm; D001254:Astrocytoma / Q000235:genetics*; D001374:Azacitidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D001932:Brain Neoplasms / Q000235:genetics*; D015820:Cadherins / Q000378:metabolism; D006528:Carcinoma, Hepatocellular / Q000235:genetics; D002648:Child; D018899:CpG Islands / Q000235:genetics*; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000378:metabolism; D019175:DNA Methylation / Q000187:drug effects; D004268:DNA-Binding Proteins / Q000378:metabolism; D000077209:Decitabine; D044127:Epigenesis, Genetic; D005260:Female; D016147:Genes, Tumor Suppressor; D005910:Glioma / Q000235:genetics; D006801:Humans; D008113:Liver Neoplasms / Q000235:genetics; D008297:Male; D058950:Melanoma-Specific Antigens; D008875:Middle Aged; D009363:Neoplasm Proteins / Q000378:metabolism*; D009687:Nuclear Proteins / Q000378:metabolism; D011401:Promoter Regions, Genetic; D012737:Sex Factors; D014407:Tumor Cells, Cultured; D000072160:Tumor Protein p73; D025521:Tumor Suppressor Proteins / Q000378:metabolism
D000951:Antigens, Neoplasm; C114483:CDH16 protein, human; D015820:Cadherins; D019941:Cyclin-Dependent Kinase Inhibitor p16; D004268:DNA-Binding Proteins; C072011:MAGEA1 protein, human; D058950:Melanoma-Specific Antigens; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; C411762:RASSF1 protein, human; C000606460:TP73 protein, human; D000072160:Tumor Protein p73; D025521:Tumor Suppressor Proteins; D000077209:Decitabine; D001374:Azacitidine