Frequent epigenetic inactivation of cystatin M in breast carcinoma

Schagdarsurengin, Undraga; Pfeifer, Gerd P.; Dammann, Reinhard

doi:10.1038/sj.onc.1210107

Cited by 35 publications

(33 citation statements)

References 25 publications

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“…In a subsequent study, methylation analysis of CST6 in breast cancer cell lines revealed an inverse correlation between CpG island hypermethylation and CST6 gene expression, and the extent of regional methylation in the proximal promoter was strongly correlated with the lack of CST6 expression (Rivenbark et al, 2006a). Consistent with these findings, several other studies have shown that CST6 is epigenetically regulated by DNA methylation-dependent silencing in breast cancer cell lines and primary invasive ductal carcinomas (Ai et al, 2006;Schagdarsurengin et al, 2006).…”

Section: Introductionsupporting

confidence: 55%

“…Ai et al showed that 12/20 (60%) primary breast tumors exhibit CST6 promoter hypermethylation, and microdissection of individual cells from select tumors revealed that methylation occurs in both DCIS and IDC cells (Ai et al, 2006). In a similar study, Schagdarsurengin et al showed that 24/40 (60%) breast carcinomas exhibited CST6 promoter hypermethylation, and that estrogen-receptor positive tumors were more frequently methylated than estrogen-receptor negative tumors (Schagdarsurengin et al, 2006). While CST6 is suggested to be epigenetically regulated through DNA methylation-dependent mechanisms in breast cancer cell lines and primary breast tumors that lack cystatin M protein expression, tumor metastases have not been examined for cystatin M expression or methylation status.…”

Section: Discussionmentioning

confidence: 98%

“…Furthermore, expression of CST6 in MDA-MB-435S breast cancer cells delays tumorigenesis by transplanted cells and suppresses spontaneous formation of liver and lung metastases (Zhang et al, 2004). More recently, it has been shown that CST6 is epigenetically regulated by DNA methylation-dependent silencing in breast cancer cell lines (Ai et al, 2006;Rivenbark et al, 2006a;Schagdarsurengin et al, 2006;Shridhar et al, 2004) and primary invasive ductal carcinomas (Ai et al, 2006;Schagdarsurengin et al, 2006). Ai et al showed that 12/20 (60%) primary breast tumors exhibit CST6 promoter hypermethylation, and microdissection of individual cells from select tumors revealed that methylation occurs in both DCIS and IDC cells (Ai et al, 2006).…”

Section: Discussionmentioning

confidence: 99%

“…Cystatin M is involved in regulating the activity of cathepsin B and cathepsin L, and imbalances between these proteases and cystatin M may lead to the metastatic phenotype in tumor cells (Frosch et al, 1999;Kos et al, 2000;Lah and Kos, 1998). Cystatin M expression has been reported to be diminished or lost in various forms of cancer including, (i) basal and squamous cell carcinomas of the skin (Zeeuwen, 2004), (ii) squamous cell carcinomas of the head/neck and lung (Zeeuwen et al, 2002), (iii) non-small cell lung cancer (Zhong et al, 2006), (iv) metastatic oral cancer cell lines (Vigneswaran et al, 2006), (v) malignant glioma (Kim et al, 2006), and (vi) breast cancer (Ai et al, 2006;Rivenbark et al, 2006a;Schagdarsurengin et al, 2006;Shridhar et al, 2004;Song et al, 2006;Sotiropoulou et al, 1997;Zhang et al, 2004). Cystatin M contains a large CpG island that flanks the transcription start site and spans the proximal promoter and exon 1 regions.…”

Section: Introductionmentioning

confidence: 99%

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Methylation-dependent silencing of CST6 in primary human breast tumors and metastatic lesions

Rivenbark

Livasy

Boyd

et al. 2007

Experimental and Molecular Pathology

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CST6 is a breast tumor suppressor gene that is expressed in normal breast epithelium, but is epigenetically silenced as a consequence of promoter hypermethylation in metastatic breast cancer cell lines. In the current study, we investigated the expression and methylation status of CST6 in primary breast tumors and lymph node metastases. 25/45 (56%) primary tumors and 17/20 (85%) lymph node metastases expressed significantly lower levels of cystatin M compared to normal breast tissue. Bisulfite sequencing demonstrated CST6 promoter hypermethylation in 11/23 (48%) neoplastic lesions analyzed, including 3/11 (27%) primary tumors and 8/12 (67%) lymph node metastases. In most cases (12/23, 52%), the expression of cystatin M directly reflected CST6 promoter methylation status. In remaining lesions (8/23, 35%) loss of cystatin M was not associated with CST6 promoter hypermethylation, indicating that other mechanisms can account for loss of CST6 expression. These results show that methylation-dependent silencing of CST6 occurs in a subset of primary breast cancers, but more frequently in metastatic lesions, possibly reflecting progressionrelated genomic events. To examine this possibility, primary breast tumors and matched lymph node metastases were analyzed. In 2/3 (67%) patients, primary tumors were positive for cystatin M and negative for CST6 promoter methylation, and matched metastatic lesions lacked cystatin M expression and CST6 was hypermethylated. This observation suggests that progression-related epigenetic alterations in CST6 gene expression can accompany metastatic spread from a primary tumor site. Overall, the results of the current investigation suggest that methylation-dependent epigenetic silencing of CST6 represents an important mechanism for loss of CST6 during breast tumorigenesis and/or progression to metastasis.

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Section: Introductionsupporting

confidence: 55%

Section: Discussionmentioning

confidence: 98%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Methylation-dependent silencing of CST6 in primary human breast tumors and metastatic lesions

Rivenbark

Livasy

Boyd

et al. 2007

Experimental and Molecular Pathology

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“…The availability of numerous breast carcinoma cell lines, with varying phenotypes (Soule et al, 1973;Cailleau et al, 1974;Thompson et al, 1992), has allowed for the identification of genes involved in the progression of breast cancer (Nagaraja and Kandpal, 2004;Nagaraja et al, 2006;Kagara et al, 2007;Schagdarsurengin et al, 2007). We have previously identified EphB6 as a molecule of interest in breast cancer by using representational difference analysis of MCF-7 and MDA-MB-231 cDNAs (Nagaraja et al, 2006) and semiquantitative RT-PCR (Fox and Kandpal, 2004).…”

Section: Introductionmentioning

confidence: 99%

EphB6 receptor significantly alters invasiveness and other phenotypic characteristics of human breast carcinoma cells

Fox

Kandpal

2009

Oncogene

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Breast cancer mortality in women is largely attributed to the metastasis of primary breast tumors. We have analysed the function of EphB6, a kinase-deficient receptor, in the invasive phenotype of breast cancer cell lines. We have demonstrated the loss of EphB6 protein in invasive breast carcinoma cell lines and absence of EphB6 transcript in a metastatic breast tumor specimen. The function of EphB6 in invasiveness was confirmed by the ability of EphB6 protein to decrease the in vitro invasiveness of MDA-MB-231, MDA-MB-435 and BT549 cells transfected with an EphB6 expression construct. In MDA-MB-231 cells, the decreased invasiveness appeared to be mediated by decreased transcript levels of matrix metalloproteinase (MMP)7 and MMP19, and increased transcript levels of tissue inhibitors of metalloproteinase 2. In addition to affecting invasiveness phenotype, EphB6 overexpression was also responsible for altering the growth rate and colony-forming efficiency of MCF-7 and MDA-MB-231 cells in a cell-line-specific manner. We suggest that the significant decrease in the invasiveness of MDA-MB-231 and other cell lines transfected with EphB6 is likely occurring by the ability of EphB6 to transduce signals to the nucleus and altering relevant gene expression.

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Inactivation of the cystatin E/M tumor suppressor gene in cervical cancer

Veena

Lee

Keppler

et al. 2008

Genes Chromosomes & Cancer

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We have previously localized a cervical cancer tumor suppressor gene to a 300 kb interval of 11q13. Analysis of candidate genes revealed loss of expression of cystatin E/M, a lysosomal cysteine protease inhibitor, in 6 cervical cancer cell lines and 9 of 11 primary cervical tumors. Examination of the three exons in four cervical cancer cell lines, 19 primary tumors, and 21 normal controls revealed homozygous deletion of exon 1 sequences in one tumor. Point mutations were observed in six other tumors. Two tumors contained mutations at the consensus binding sites for cathepsin L, a lysosomal protease over-expressed in cervical cancer. Introduction of these two point mutations using site directed mutagenesis resulted in reduced binding of mutated cystatin E/M to cathepsin L. Although mutations were not observed in any cell lines, four cell lines and 12 of 18 tumors contained promoter hypermethylation. Re-expression of cystatin E/M was observed after 5′aza 2-deoxycytidiene and/or Trichostatin A treatment of cervical cancer cell lines, HeLa and SiHa, confirming promoter hypermethylation. Ectopic expression of cystatin E/M in these two cell lines resulted in growth suppression. There was also suppression of soft agar colony formation by HeLa cells expressing the cystatin E/M gene. Re-expression of cystatin E/M resulted in decreased intracellular and extracellular expression of cathepsin L. Over-expression of cathepsin L resulted in increased cell growth which was inhibited by the reintroduction of cystatin E/M. We conclude, therefore, that cystatin E/M is a cervical cancer suppressor gene and that the gene is inactivated by somatic mutations and promoter hypermethylation.

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Frequent epigenetic inactivation of cystatin M in breast carcinoma

Cited by 35 publications

References 25 publications

Methylation-dependent silencing of CST6 in primary human breast tumors and metastatic lesions

Methylation-dependent silencing of CST6 in primary human breast tumors and metastatic lesions

EphB6 receptor significantly alters invasiveness and other phenotypic characteristics of human breast carcinoma cells

Inactivation of the cystatin E/M tumor suppressor gene in cervical cancer

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