p14^ARF Deletion and Methylation in Genetic Pathways to Glioblastomas

Abstract

The CDKN2A locus on chromosome 9p21 contains the p14 ARF and p16 INK4a genes, and is frequently deleted in human neoplasms, including brain tumors. In this study, we screened 34 primary (de novo) glioblastomas and 16 secondary glioblastomas that had progressed from low-grade diffuse astrocytomas for alterations of the p14 ARF and p16 INK4a genes, including homozygous deletion by differential PCR, promoter hypermethylation by methylationspecific PCR, and protein expression by immunohistochemistry. A total of 29 glioblastomas (58%) had a p14 ARF homozygous deletion or methylation, and 17 (34%) showed p16 INK4a homozygous deletion or methylation. Thirteen glioblastomas showed both p14 ARF and p16 INK4a homozygous deletion, while nine showed only a p14 ARF deletion. Immunohistochemistry revealed loss of p14 ARF expression in the majority of glioblastomas (38/50, 76%), and this correlated with the gene status, i.e. homozygous deletion or promoter hypermethylation. There was no significant difference in the overall frequency of p14 ARF and p16 INK4a alterations between primary and secondary glioblastomas. The analysis of multiple biopsies from the same patients revealed hypermethylation of p14 ARF (5/15 cases) and p16 INK4a (1/15 cases) already at the stage of low-grade diffuse astrocytoma but consistent absence of homozygous deletions. These results suggest that aberrant p14 ARF expression due to homozygous deletion or promoter hypermethylation is associated with the evolution of both primary and secondary glioblastomas, and that p14 ARF promoter methylation is an early event in subset of astrocytomas that undergo malignant progression to secondary glioblastoma.

The role of HRK gene in human cancer

“…It is also noteworthy that a majority of genes reported to be methylated in tumors also tend to be methylated in regions associated with lower-grade malignancy (Toyota et al, 1999;Nakamura et al, 2001aNakamura et al, , b, 2005a. In agreement with these reports, HRK methylation was detected less frequently in low-grade diffuse astrocytoma, suggesting that inactivation of the gene may be involved in a critical step in malignant progression.…”

Aberrant Hypermethylation of Non-Promoter Zygote Arrest 1 (Zar1) in Human Brain Tumors

“…[3][4][5][6][7][11][12][13] Based on our results, ZAR1 has emerged as the most frequently methylated gene in diffuse gliomas to date. Of particular interest is that frequent hypermethylation of ZAR1 was evident in low-grade diffuse astrocytomas (100%), anaplastic astrocytomas (94%), glioblastomas (93%), oligodendrogliomas (100%), and anaplastic oligodendrogliomas (100%), indicating that ZAR1 hypermethylation did not differ significantly between diffuse low-grade and high-grade gliomas.…”

“…In glioblastomas, for instance, frequent promoter hypermethylation has been noted for O 6 -methylguanine-DNA methyltransferase, p14 ARF , RB1, RUNX3, and Testin. [3][4][5][6][7][11][12][13] Most evaluations of DNA methylation have so far concentrated on genes primarily inactivated by hypermethylation of normally unmethylated CpG islands in the promoter regions and analyzed for only a restricted set of CpG sites in their target regions. Recent evaluation of the methylation status of global genomic regions in colorectal cancer has indicated that hypermethylation of some CpG islands located in non-promoter regions does not interfere with transcriptional repression and is even associated with up-regulation or ectopic gene expression.…”

Molecular Genetics of Secondary Glioblastoma

“…Either through homozygous deletion or promoter methylation, the alteration of p16INK4a is an important step in both primary and secondary GBMs (56). Conversely, methylation of the RB1 promoter, correlating with decreased RB1 expression, is more specific to secondary GBM (57).…”