Chang HC, Cho CY, Hung WC.. Downregulation of RECK by promoter methylation correlates with lymph node metastasis in non-small cell lung cancer. Cancer Sci 98: 169-173

Department of Chest Surgery, Kaohsiung Veterans General Hospital, 386, Ta-Chung 1st Road, Kaohsiung 813, Taiwan.
Cancer Science (Impact Factor: 3.52). 03/2007; 98(2):169-73. DOI: 10.1111/j.1349-7006.2006.00367.x
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In the present study, we addressed the molecular mechanism of the downregulation of reversion-inducing-cysteine-rich protein with Kazal motifs (RECK), a critical tumor suppressor that can potently inhibit angiogenesis and metastasis, in non-small cell lung cancer and its clinical significance. The methylation status of the RECK gene promoter was studied by methylation-specific polymerase chain reaction. RECK mRNA and protein levels were investigated by reverse transcription-polymerase chain reaction and western blot analysis. Downregulation of RECK was observed in 60% of the 55 tumors analyzed. Using methylation-specific polymerase chain reaction analysis methylation of the RECK promoter was detected in 63.6% (35/55) of the tumor tissues. A strong correlation between downregulation and promoter methylation was found in these tumors (P = 0.000005). More importantly, downregulation of RECK significantly correlated with lymph node metastasis (P = 0.038). Mutation of codon 12 of the K-ras gene was detected in 25.5% (14/55) of lung tumor tissues. Statistical analysis indicated that K-ras mutation was linked with RECK promoter methylation (P = 0.047) and downregulation (P = 0.023). Promoter methylation was also detected in human lung cancer cell lines, and the DNA methyltransferase inhibitor 5'-azacytidine reversed the expression of RECK and reduced the invasive ability of these cell lines. Collectively, our results suggest that downregulation of the metastasis suppressor RECK is caused by promoter methylation in non-small cell lung cancer and is associated with K-ras mutation and lymph node metastasis.

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Available from: Wen-Chun Hung, Jul 15, 2015
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    • "Germ line knockout of RECK induced the proliferation of mouse embryonic fibroblasts (MEFs) and enabled early escape from the cellular senescence induced by oncogenic insults, and RECK interferes with epidermal growth factor receptor (EGFR) signaling [5]. Epigenetics, such as, histone and DNA modifications, are involved in the silencing of RECK [6,7]. We previously reported that RECK is downregulated under hypoxic conditions through HDAC1 and hypoxia inducible factor (HIF)-1α [8]. "
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    ABSTRACT: Reversion-inducing cysteine-rich protein with Kazal motifs (RECK, a tumor suppressor) is down-regulated by the oncogenic signals and hypoxia, but the biological function of RECK in early tumorigenic hyperplastic phenotypes is largely unknown. Knockdown of RECK by small interfering RNA (siRECK) or hypoxia significantly promoted cell proliferation in various normal epithelial cells. Hypoxia as well as knockdown of RECK by siRNA increased the cell cycle progression, the levels of cyclin D1 and c-Myc, and the phosphorylation of Rb protein (p-pRb), but decreased the expression of p21(cip1), p27(kip1), and p16(ink4A). HIF-2α was upregulated by knockdown of RECK, indicating HIF-2α is a downstream target of RECK. As knockdown of RECK induced the activation of epidermal growth factor receptor (EGFR) and treatment of an EGFR kinase inhibitor, gefitinib, suppressed HIF-2α expression induced by the silencing of RECK, we can suggest that the RECK silenicng-EGFR-HIF-2α axis might be a key molecular mechanism to induce hyperplastic phenotype of epithelial cells. It was also found that shRNA of RECK induced larger and more numerous colonies than control cells in an anchorage-independent colony formation assay. Using a xenograft assay, epithelial cells with stably transfected with shRNA of RECK formed a solid mass earlier and larger than those with control cells in nude mice. In conclusion, the suppression of RECK may promote the development of early tumorigenic hyperplastic characteristics in hypoxic stress.
    PLoS ONE 12/2013; 8(12):e84520. DOI:10.1371/journal.pone.0084520 · 3.23 Impact Factor
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    • "DNMT inhibitor could reverse promoter methylation and restores related gene expression in human cancer cell Lines 20. DNMT inhibitor may restore RECK expression to inhibit cell invasion in colon cancer and lung cancer cell lines 11, 15. EGCG, a major component of green tea, also may enhance RECK expression by reversal of hypermethylation of RECK promoter and inhibit MMP activities as well as cancer cell invasion in OSCC cell lines 21. "
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    ABSTRACT: To evaluate the promoter methylation status of RECK gene and mRNA expression in patients with hepatocellular carcinoma (HCC). We analyzed RECK methylation by MSP, and RECK mRNA by real-time PCR in 74 HCC. The liver cell lines (7721, Chang and Hep-G2) were treated with 5-Aza-CdR and TSA. RECK mRNA were lower in HCC tissues (Mean (-∆Ct) = -3.29) than that in Non-Hcc tissues (Mean (-∆Ct) = -2.42). Expression of RECK was elevated in only 24 (32.43%) of the 74 HCC patients but decreased (-∆∆Ct<0) in 50 (67.57%) of the patients. RECK promoter was hypermethylated in 55.4% (41/74) of HCCs, and in only 17.6% (13/74) of Non-Hcc samples. RECK mRNA were lower in HCC patients with hypermethylation (∆MI>=0.5) (Mean (-∆∆Ct) = -1.75) than those with demethylation (∆MI<0.5) (Mean (-∆∆Ct) = 0.05), and there is a decreased tendency for RECK mRNA in HCC patients with promoter hypermethylation (p = 0.002). There was a significantly correlation found between RECK mRNA and poor survival after surgery. After treated by 5-Aza-CdR and TSA, we found that RECK mRNA induced different changes in 7721, Chang and Hep-G2 cells. And RECK demethylation also induced by epigenetic inhibitors. The results suggested that the hypermethylation may lead to promoter silencing of RECK mRNA and associated with poor survival in HCC.
    International journal of biological sciences 03/2012; 8(4):451-8. DOI:10.7150/ijbs.4038 · 4.51 Impact Factor
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    • "Specific genes linked to recurrence or survival include SMARCA2 (implicated in the regulation of gene expression cell cycle control and oncogenesis), MINK (linked to the JNK MAP kinase pathway) [47] and RECK, which has putative roles in the suppression of tumour growth, invasion, angiogenesis and metastasis [48]. KRAS mutation has been associated with reduced expression of RECK in NSCLC [49], consistent with the clinical observation of poor outcome in patients with KRAS mutation bearing NSCLC. Activation of the Ras pathway may reduce RECK expression and thereby increase tumour recurrence. "
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    ABSTRACT: The aim of this study was to identify critical genes involved in non-small cell lung cancer (NSCLC) pathogenesis that may lead to a more complete understanding of this disease and identify novel molecular targets for use in the development of more effective therapies. Both transcriptional and genomic profiling were performed on 69 resected NSCLC specimens and results correlated with mutational analyses and clinical data to identify genetic alterations associated with groups of interest. Combined analyses identified specific patterns of genetic alteration associated with adenocarcinoma vs. squamous differentiation; KRAS mutation; TP53 mutation, metastatic potential and disease recurrence and survival. Amplification of 3q was associated with mutations in TP53 in adenocarcinoma. A prognostic signature for disease recurrence, reflecting KRAS pathway activation, was validated in an independent test set. These results may provide the first steps in identifying new predictive biomarkers and targets for novel therapies, thus improving outcomes for patients with this deadly disease.
    BMC Cancer 03/2011; 11(1):93. DOI:10.1186/1471-2407-11-93 · 3.36 Impact Factor
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