Aberrant promoter methylation of DLEC1, a critical 3p22 tumor suppressor for renal cell carcinoma, is associated with more advanced tumor stage.

PURPOSE Identifying tumor suppressor genes silenced by promoter CpG methylation uncovers mechanisms of tumorigenesis and identifies new epigenetic biomarkers for early cancer detection. DLEC1 is located at 3p22.3, a critical tumor suppressor gene locus for renal cell carcinoma. We explored its epigenetic alteration in renal cell carcinoma and possible clinicopathological association. MATERIALS AND METHODS We examined DLEC1 expression and methylation by semiquantitative reverse transcriptase and methylation specific polymerase chain reaction in 9 renal cell carcinoma cell lines and 81 primary tumors. We also analyzed the relationship between DLEC1 methylation and clinicopathological features in patients with renal cell carcinoma. We assessed DLEC1 inhibition of renal cell carcinoma cell growth by colony formation assay. RESULTS DLEC1 methylation and down-regulation were detected in all renal cell carcinoma cell lines. Treatment with 5-aza-2'-deoxycytidine (Sigma) and/or trichostatin A (Cayman Chemical, Ann Arbor, Michigan) reversed methylation and restored DLEC1 expression, indicating that methylation directly mediates its silencing. Aberrant methylation was further detected in 25 of 81 primary tumors (31%) but only 1 of 53 nonmalignant renal tissues (2%) showed methylation. DLEC1 methylation status was significantly associated with TNM classification and grade in patients with renal cell carcinoma (chi-square test p = 0.01 and 0.04, respectively). DLEC1 ectopic expression in silenced renal cell carcinoma cells resulted in substantial tumor cell clonogenicity inhibition. CONCLUSIONS To our knowledge we report for the first time that DLEC1 is often down-regulated by CpG methylation and shows tumor inhibitory function in renal cell carcinoma cells, indicating its role as a tumor suppressor. DLEC1 tumor specific methylation may serve as a biomarker for early detection and prognosis prediction of this tumor.

[1]  B. Teh,et al.  Tumor suppressor activity and epigenetic inactivation of hepatocyte growth factor activator inhibitor type 2/SPINT2 in papillary and clear cell renal cell carcinoma. , 2005, Cancer research.

[2]  R. Uzzo,et al.  Promoter Hypermethylation Profile of Kidney Cancer , 2004, Clinical Cancer Research.

[3]  R. Berkowitz,et al.  Candidate tumor-suppressor gene DLEC1 is frequently downregulated by promoter hypermethylation and histone hypoacetylation in human epithelial ovarian cancer. , 2006, Neoplasia.

[4]  Q. Tao,et al.  Aberrant methylation of the 8p22 tumor suppressor gene DLC1 in renal cell carcinoma. , 2007, Cancer letters.

[5]  Jun Yu,et al.  DLEC1 is a functional 3p22.3 tumour suppressor silenced by promoter CpG methylation in colon and gastric cancers , 2009, British Journal of Cancer.

[6]  P. Carroll,et al.  Methylation of the γ-Catenin Gene is Associated With Poor Prognosis of Renal Cell Carcinoma , 2005 .

[7]  J. Cheville,et al.  The Mayo Clinic experience with surgical management, complications and outcome for patients with renal cell carcinoma and venous tumour thrombus , 2004, BJU international.

[8]  A. Protopopov,et al.  The candidate tumor suppressor gene, RASSF1A, from human chromosome 3p21.3 is involved in kidney tumorigenesis , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[9]  E. Zabarovsky,et al.  Search for unknown tumor‐antagonizing genes , 2003, Genes, chromosomes & cancer.

[10]  Loren Lipworth,et al.  Epidemiologic aspects of renal cell carcinoma. , 2006, Seminars in oncology.

[11]  J. Califano,et al.  Quantitative Detection of Promoter Hypermethylation of Multiple Genes in the Tumor, Urine, and Serum DNA of Patients with Renal Cancer , 2004, Cancer Research.

[12]  G. Ito,et al.  Frequent inactivation of RASSF1A, BLU, and SEMA3B on 3p21.3 by promoter hypermethylation and allele loss in non-small cell lung cancer. , 2005, Cancer letters.

[13]  Q. Tao,et al.  Defective de novo methylation of viral and cellular DNA sequences in ICF syndrome cells. , 2002, Human molecular genetics.

[14]  Tamas Beothe,et al.  Deletion of chromosome 3p14.2-p25 involving the VHL and FHIT genes in conventional renal cell carcinoma. , 2003, Cancer research.

[15]  A. Knudson,et al.  Two genetic hits (more or less) to cancer , 2001, Nature Reviews Cancer.

[16]  Yusuke Nakamura,et al.  Epigenetic inactivation of the deleted in lung and esophageal cancer 1 gene in nasopharyngeal carcinoma , 2007, Genes, chromosomes & cancer.

[17]  M. Salto‐Tellez,et al.  The tumor suppressor gene DLEC1 is frequently silenced by DNA methylation in hepatocellular carcinoma and induces G1 arrest in cell cycle. , 2008, Journal of hepatology.

[18]  M. Toyota,et al.  Epigenetic inactivation of the candidate tumor suppressor gene HOXB13 in human renal cell carcinoma , 2006, Oncogene.

[19]  Qian Tao,et al.  The Stress-Responsive Gene GADD45G Is a Functional Tumor Suppressor, with Its Response to Environmental Stresses Frequently Disrupted Epigenetically in Multiple Tumors , 2005, Clinical Cancer Research.

[20]  Paul Cairns,et al.  Gene methylation and early detection of genitourinary cancer: the road ahead , 2007, Nature Reviews Cancer.

[21]  Peter A. Jones,et al.  The fundamental role of epigenetic events in cancer , 2002, Nature Reviews Genetics.

[22]  Y. Nakamura,et al.  Molecular cloning of a candidate tumor suppressor gene, DLC1, from chromosome 3p21.3. , 1999, Cancer research.

[23]  D. Gutmann,et al.  Promoter hypermethylation of the potential tumor suppressor DAL‐1/4.1B gene in renal clear cell carcinoma , 2006, International journal of cancer.

[24]  I. Zanna,et al.  Loss of heterozygosity and methylation of p16 in renal cell carcinoma , 2003, Urological Research.