Frequent LOH at Chromosome 12q22‐23 and Apaf‐1 Inactivation in Glioblastoma

Abstract

Glioblastoma (GB) often has loss of heterozygosity on the chromosomes, 1p, 10p, 10q, 11p, 17p, 19q, 22q, and several others. In the case of chromosome 12q; however, it remains to be seen whether LOH occurs. Apaf‐1, the apoptotic protease activating factor‐1, located at chromosome 12q22‐23, is a major effecter of the p53 mediated apoptosis pathway, and Apaf‐1 inactivation due to chromosome 12q22‐23 LOH and hypermethylation may be involved in some of the neoplasms in malignancy. However, little is known about the frequency of the 12q22‐23 LOH or the state of Apaf‐1 in GB. To elucidate their involvement in GB, we analyzed a series of 33 GBs for chromosome 12q22‐23 LOH, Apaf‐1 mRNA expression, and Apaf‐1 protein expression, using microsatellite analysis, reverse transcription (RT) ‐PCR analysis, and immunohistochemical (IHC) analysis, respectively. We also evaluated if and how the 12q22‐23 LOH correlated with the p53 gene mutation and EGFR gene amplification. Chromosome 12q22‐23 LOH was detected in 14 (42%) of 33 cases. Among the examined cases with LOH at 12q22‐23, a low expression of Apaf‐1 mRNA was detected in 9 (69%) of 13 cases, and a low expression of Apaf‐1 protein was detected in 12 (86%) of 14 cases. The 12q22‐23 LOH was significantly correlated with low expression of mRNA and protein (p<0.05, p<0.001 respectively). The p53 gene mutation and EGFR gene amplification were found in 13 cases (39%) and 8 cases (24%), respectively, and these gene alterations were inversely correlated. However, 12q22‐23 LOH had no correlations with the p53 gene mutation or EGFR gene amplification. Six of 9 GBs (67%) with neither p53 gene mutation nor EGFR gene amplification tested positive for 12q22‐23 LOH. These GBs are likely to belong to another subset independent from the 2 common genetic subsets in GB (one with p53 gene mutation and without EGFR gene amplification, and the other with EGFR gene amplification and without p53 gene mutation). Twenty‐three (70%) out of the 33 GBs with the 12q22‐23 LOH also tested positive for Apaf‐1 inactivation or p53 gene mutation. This high frequency of alterations in the apoptosis‐associated factors prompts a speculation that abrogation of the Apaf‐1 and p53 mediated apoptosis pathway may play an important role in the tumorigenesis of GB.

Methylation Silencing of the Apaf-1 Gene in Acute Leukemia

“…Aberrant expression of Apaf-1 with defects in dATP-mediated caspase-3 activation is also detected in male germ cell tumors (7). Recently, Watanabe et al (8) reported frequent inactivation of Apaf-1 due to loss of heterozygosity in glioblastoma. Furthermore, a frame-shift mutation of the mononucleotide tract in the Apaf-1 gene is associated with gastrointestinal cancer of the microsatellite mutator phenotype (9).…”

Apaf-1 expression in malignant melanoma

“…16 Alternatively, the mechanisms repressing Apaf-1 in primary tumor specimens can be occasionally altered through genetic or epigenetic changes that accompany passaging in culture (e.g., by potentiation of the Rb/E2F-1 pathway, which can control Apaf-1 mRNA levels 17 ). The fact that certain leukemia cells 18,19 as well as AML, ALL, and CML blasts 18 and cells from glioblastoma 20 and cervical cancer 21 may also inactivate Apaf-1 expression, in part by methylation, suggests a broader impact of tumorrelated events modulating Apaf-1 expression.…”

Analysis of apoptosome dysregulation in pancreatic cancer and of its role in chemoresistance

“…6 In this regard, the two proteins seem to act as tumor suppressors. In fact, the APAF1 gene does not appear to be functional in melanoma, 7-9 glioblastoma, 10,11 gastrointestinal tumor, 12 germ line tumor. 13 More recently, a poor prognosis was observed in patients suffering from B-type chronic lymphocytic leukemia with loss of APAF1 expression and additional p53 mutation.…”