Frequent Inactivation of PTEN by Promoter Hypermethylation in Microsatellite Instability-High Sporadic Colorectal Cancers

Abstract

Loss of PTEN tumor suppressor function is observed in tumors of breast, prostate, thyroid, and endometrial origin. Allelic losses in the proximity of the PTEN locus (10q23) also occur in sporadic colorectal cancers (CRCs), but biallelic inactivation of this site has not been frequently demonstrated. We hypothesized that alternative mechanisms of PTEN allelic inactivation, such as promoter hypermethylation, might be operative in CRC and that PTEN inactivation may be related to recognized forms of genomic instability. We characterized a cohort of 273 sporadic CRCs by determining their microsatellite instability (MSI) status. Of these, 146 cancers were examined for PTEN promoter methylation by methylation-specific PCR. Mutations at the poly(A) 6 repeat sequences in PTEN exons 7 and 8 and deletions at the 10q23 locus were also identified using microsatellite analysis. The presence of PTEN protein was determined by immunostaining, and the results were correlated with the promoter methylation status. We observed that PTEN promoter hypermethylation was a frequent occurrence in MSI-high (MSI-H) tumors (19.1% of MSI-H versus 2.2% of MSI-low/microsatellite stable tumors; P ‫؍‬ 0.002). A PTEN mutation or a deletion event was present in 60% of the tumors with promoter region hypermethylation. Hypermethylation of the PTEN promoter correlated significantly with either decreased or complete loss of PTEN protein expression (P ‫؍‬ 0.004). This is the first demonstration of PTEN inactivation as a result of promoter hypermethylation in MSI-H sporadic CRCs. These data suggest that this silencing mechanism plays a major role in PTEN inactivation and, in colon cancer, may be more important than either allelic losses or inactivating mutations. The significant correlation of PTEN hypermethylation with MSI-H tumors further suggests that PTEN is an additional important "target" of methylation along with the hMLH1 gene in the evolution of MSI-H CRCs and also confers the "second hit" in the biallelic inactivation mechanism for some proportion of tumors.

APC promoter hypermethylation contributes to the loss of APC expression in colorectal cancers with allelic loss on 5q1

“…27 In addition, we have shown that PTEN expression in sporadic CRC is influenced by mutations or allelic loss of one allele and promoter hypermethylation of the other allele. 28 Promoter hypermethylation might occur at a very early state. Early forms of colorectal adenomas are frequently hypermethylated at various tumorrelated genes.…”

Prognostic vs predictive molecular biomarkers in colorectal cancer: is KRAS and BRAF wild type status required for anti-EGFR therapy?

“…Mutation constitutive activation of the PI3K signaling pathway has been reported to occur in~30% of colon tumors, primarily due to activating mutations in exons 9 and 20 of the PIK3CA gene 43,44 and, to a lesser extent, due to inactivating PTEN mutations or PTEN promoter methylation. 45 PTEN is a tumor suppressor that acts as a negative regulator of PI3K signaling by converting PIP3 to PIP2, and truncating mutations which result in loss of PTEN expression, reported in~20% of MSI colon cancers. [46][47][48][49][50][51] The molecular alteration of PTEN is often caused by epigenetic mechanisms, 45 supporting the detection of the intact protein by IHC as a better diagnostic tool than gene sequencing, as it potentially covers more mechanisms of alteration.…”

“…45 PTEN is a tumor suppressor that acts as a negative regulator of PI3K signaling by converting PIP3 to PIP2, and truncating mutations which result in loss of PTEN expression, reported in~20% of MSI colon cancers. [46][47][48][49][50][51] The molecular alteration of PTEN is often caused by epigenetic mechanisms, 45 supporting the detection of the intact protein by IHC as a better diagnostic tool than gene sequencing, as it potentially covers more mechanisms of alteration. PIK3CA mutation and PTEN expression status predicts response of colon cancer cells to the EGFR inhibitor cetuximab distinguishing drug sensitive and resistant cell lines.…”

RAB32 hypermethylation and microsatellite instability in gastric and endometrial adenocarcinomas

“…There is growing evidence to suggest that concurrent hypermethylation of multiple additional genes may represent an important component in the development and progression of colorectal cancers. [2][3][4][5][6][7][8] This propensity for methylation in such tumors has served as an interesting feature to exploit in the search for novel methylation targets.We have recently applied a global expression profiling-based technique to identify potential novel methylation targets in MSI-H colorectal cancers. One of the genes discovered to have significant differential methylation in microsatellite-unstable cancers was RAB32, a mitochondrial ras family member that encodes an A-kinase anchoring protein.…”

“…There is growing evidence to suggest that concurrent hypermethylation of multiple additional genes may represent an important component in the development and progression of colorectal cancers. [2][3][4][5][6][7][8] This propensity for methylation in such tumors has served as an interesting feature to exploit in the search for novel methylation targets.…”