Epigenetic inactivation of mir-34b/c in addition to mir-34a and DAPK1 in chronic lymphocytic leukemia

Journal of Translational Medicine

Chor Sang Chim³  Anders Rosèn⁴  Eric Tse³  Dong Yan Jin¹  Chi Shan Bonnie Kho²  Kit Fai Wong⁵  Yok Lam Kwong³  Lu Qian Wang³ 

[1] Department of Biochemistry, The University of Hong Kong, Hong Kong, China;Department of Medicine, Pamela Youde Nethersole Hospital, Hong Kong, China;Department of Medicine, Queen Mary Hospital, The University of Hong Kong, Hong Kong, China;Department of Clinical & Experimental Medicine, Linköping University, Linköping, Sweden;Department of Pathology, Queen Elizabeth Hospital, Hong Kong, China

关键词: Chronic lymphocytic leukemia;    DNA methylation;    Tumor suppressor;    TP53 network;    MicroRNA;   

Others  :  817886
DOI  :  10.1186/1479-5876-12-52

 received in 2013-11-05, accepted in 2014-02-19,  发布年份 2014

【 摘 要 】

Background

TP53 mutation/deletion is uncommon in chronic lymphocytic leukemia (CLL). We postulated that components of TP53-centered tumor suppressor network, miR-34b/c, in addition to DAPK1 and miR-34a might be inactivated by DNA hypermethylation. Moreover, we tested if miR-34b/c methylation might correlate with miR-203 or miR-124-1 methylation in CLL.

Methods

miR-34b/c, miR-34a and DAPK1 methylation was studied in 11 normal controls, 7 CLL cell lines, and 78 diagnostic CLL samples by methylation-specific polymerase chain reaction. MEC-1 cells were treated with 5-Aza-2’-deoxycytidine for reversal of methylation-associated miRNA silencing. Tumor suppressor properties of miR-34b were demonstrated by over-expression of precursor miR-34b in MEC-1 cells.

Results

miR-34b/c promoter was unmethylated in normal controls, but completely methylated in 4 CLL cell lines. miR-34b/c expression was inversely correlated with miR-34b/c methylation. Different MSP statuses of miR-34b/c, including complete methylation and complete unmethylation, were verified by quantitative bisulfite pyrosequencing. 5-Aza-2’-deoxycytidine treatment resulted in promoter demethylation and miR-34b re-expression in MEC1 cells. Moreover, over-expression of miR-34b resulted in inhibition of cellular proliferation and increased cell death. In primary CLL samples, miR-34a, miR-34b/c and DAPK1 methylation was detected in 2.6%, 17.9% and 34.6% of patients at diagnosis respectively. Furthermore, 39.7%, 3.8% and 2.6% patients had methylation of one, two or all three genes respectively. Overall, 46.2% patients had methylation of at least one of these three genes. Besides, miR-34b/c methylation was associated with methylation of miR-34a (P = 0.03) and miR-203 (P = 0.012) in CLL.

Conclusions

Taken together, miR-34b/c is a tumor suppressor miRNA frequently methylated, and hence silenced in CLL. Together with DAPK1 methylation, miR-34b/c methylation is implicated in the disruption of the TP53-centered tumor suppressor network. Moreover, the association of miRNA methylation warrants further study.

【 授权许可】

   
2014 Wang et al.; licensee BioMed Central Ltd.

【 图 表 】

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【 参考文献 】

• [1]Esteller M: Epigenetics in cancer. N Engl J Med 2008, 358:1148-1159.
• [2]Chim CS, Liang R, Kwong YL: Hypermethylation of gene promoters in hematological neoplasia. Hematol Oncol 2002, 20:167-176.
• [3]Jones PA, Baylin SB: The epigenomics of cancer. Cell 2007, 128:683-692.
• [4]Rush LJ, Plass C: Alterations of DNA methylation in hematologic malignancies. Cancer Lett 2002, 185:1-12.
• [5]Gonzalez-Zulueta M, Bender CM, Yang AS, Nguyen TD, Beart RW, Van Tornout JM, Jones PA: Methylation of the 5′ CpG island of the p16/CDKN2 tumor suppressor gene in normal and transformed human tissues correlates with gene silencing. Cancer Res 1995, 55:4531-4535.
• [6]Chim CS, Fung TK, Cheung WC, Liang R, Kwong YL: SOCS1 and SHP1 hypermethylation in multiple myeloma: implications for epigenetic activation of the Jak/STAT pathway. Blood 2004, 103:4630-4635.
• [7]Chim CS, Liang R, Fung TK, Choi CL, Kwong YL: Epigenetic dysregulation of the death-associated protein kinase/p14/HDM2/p53/Apaf-1 apoptosis pathway in multiple myeloma. J Clin Pathol 2007, 60:664-669.
• [8]Xiang T, Li L, Yin X, Zhong L, Peng W, Qiu Z, Ren G, Tao Q: Epigenetic silencing of the WNT antagonist Dickkopf 3 disrupts normal Wnt/β-catenin signalling and apoptosis regulation in breast cancer cells. J Cell Mol Med 2013, 17:1236-1246.
• [9]Liu TH, Raval A, Chen SS, Matkovic JJ, Byrd JC, Plass C: CpG island methylation and expression of the secreted frizzled-related protein gene family in chronic lymphocytic leukemia. Cancer Res 2006, 66:653-658.
• [10]Raval A, Tanner SM, Byrd JC, Angerman EB, Perko JD, Chen S-S, Hackanson B, Grever MR, Lucas DM, Matkovic JJ, Lin TS, Kipps TJ, Murray F, Weisenburger D, Sanger W, Lynch J, Watson P, Jansen M, Yoshinaga Y, Rosenquist R, de Jong PJ, Coggill P, Beck S, Lynch H, de la Chapelle A, Plass C: Downregulation of death-associated protein kinase 1 (DAPK1) in chronic lymphocytic leukemia. Cell 2007, 129:879-890.
• [11]Chim CS, Pang R, Liang R: Epigenetic dysregulation of the Wnt signalling pathway in chronic lymphocytic leukaemia. J Clin Pathol 2008, 61:1214-1219.
• [12]Calin GA, Croce CM: MicroRNA signatures in human cancers. Nat Rev Cancer 2006, 6:857-866.
• [13]Bartel DP: MicroRNAs: target recognition and regulatory functions. Cell 2009, 136:215-233.
• [14]Sana J, Faltejskova P, Svoboda M, Slaby O: Novel classes of non-coding RNAs and cancer. J Transl Med 2012, 10:103. BioMed Central Full Text
• [15]Weinberg RA: Tumor suppressor genes. Science 1991, 254:1138-1146.
• [16]Harris CC, Hollstein M: Clinical implications of the p53 tumor-suppressor gene. N Engl J Med 1993, 329:1318-1327.
• [17]Zenz T, Eichhorst B, Busch R, Denzel T, Häbe S, Winkler D, Bühler A, Edelmann J, Bergmann M, Hopfinger G, Hensel M, Hallek M, Döhner H, Stilgenbauer S: TP53 Mutation and survival in chronic lymphocytic leukemia. J Clin Oncol 2010, 28:4473-4479.
• [18]Rossi D, Cerri M, Deambrogi C, Sozzi E, Cresta S, Rasi S, De Paoli L, Spina V, Gattei V, Capello D, Forconi F, Lauria F, Gaidano G: The prognostic value of TP53 mutations in chronic lymphocytic leukemia is independent of Del17p13: implications for overall survival and chemorefractoriness. Clin Cancer Res 2009, 15:995-1004.
• [19]Kohler M, Kerns B, Humphrey P, Marks J, Bast R Jr, Berchuck A: Mutation and overexpression of p53 in early-stage epithelial ovarian cancer. Obstet Gynecol 1993, 81:643.
• [20]Chim CS, Fung TK, Wong K, Lau J, Liang R: Frequent DAP kinase but not p14 or Apaf-1 hypermethylation in B-cell chronic lymphocytic leukemia. J Hum Genet 2006, 51:832-838.
• [21]Hallek M, Cheson BD, Catovsky D, Caligaris-Cappio F, Dighiero G, Döhner H, Hillmen P, Keating MJ, Montserrat E, Rai KR, Kipps TJ: Guidelines for the diagnosis and treatment of chronic lymphocytic leukemia: a report from the international workshop on chronic lymphocytic leukemia updating the national cancer institute–working group 1996 guidelines. Blood 2008, 111:5446-5456.
• [22]Jaffe ES: Pathology and Genetics of Tumours of Haematopoietic and Lymphoid Tissues. Lyon: LARC Press; 2001.
• [23]Chim CS, Wong KY, Qi Y, Loong F, Lam WL, Wong LG, Jin D, Costello JF, Liang R: Epigenetic inactivation of the miR-34a in hematological malignancies. Carcinogenesis 2010, 31:745-750.
• [24]Stacchini A, Aragno M, Vallario A, Alfarano A, Circosta P, Gottardi D, Faldella A, Rege-Cambrin G, Thunberg U, Nilsson K: MEC1 and MEC2: two new cell lines derived from B-chronic lymphocytic leukaemia in prolymphocytoid transformation. Leuk Res 1999, 23:127-136.
• [25]Wendel-Hansen V, Sällström J, De Campos-Lima P, Kjellström G, Sandlund A, Siegbahn A, Carlsson M, Nilsson K, Rosen A: Epstein-barr virus (EBV) can immortalize B-cll cells activated by cytokines. Leukemia 1994, 8:476.
• [26]Rosén A, Bergh AC, Gogok P, Evaldsson C, Myhrinder AL, Hellqvist E, Rasul A, Björkholm M, Jansson M, Mansouri L: Lymphoblastoid cell line with B1 cell characteristics established from a chronic lymphocytic leukemia clone by in vitro EBV infection. Oncol Immunol 2012, 1:18-27.
• [27]Lujambio A, Calin GA, Villanueva A, Ropero S, Sánchez-Céspedes M, Blanco D, Montuenga LM, Rossi S, Nicoloso MS, Faller WJ: A microRNA DNA methylation signature for human cancer metastasis. Proc Natl Acad Sci USA 2008, 105:13556-13561.
• [28]Wong KY, Yim RLH, So CC, Jin DY, Liang R, Chim CS: Epigenetic inactivation of the MIR34B/C in multiple myeloma. Blood 2011, 118:5901-5904.
• [29]Chim CS, Wong KY, Leung CY, Chung LP, Hui PK, Chan SY, Yu L: Epigenetic inactivation of the hsa-miR-203 in haematological malignancies. J Cell Mol Med 2011, 15:2760-2767.
• [30]Livak KJ, Schmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2 ΔΔCT method. Methods 2001, 25:402-408.
• [31]Narayan G, Arias-Pulido H, Koul S, Vargas H, Zhang F, Villella J, Schneider A, Terry M, Mansukhani M, Murty V: Frequent promoter methylation of CDH1, DAPK, RARB, and HIC1 genes in carcinoma of cervix uteri: its relationship to clinical outcome. Mol Cancer 2003, 2:24. BioMed Central Full Text
• [32]Wong KY, So CC, Loong F, Chung LP, Lam WW, Liang R, Li GK, Jin DY, Chim CS: Epigenetic inactivation of the miR-124-1 in haematological malignancies. PLoS One 2011, 6:e19027.
• [33]Knudson AG: Karnofsky memorial lecture. Hereditary cancer: theme and variations. J Clin Oncol 1997, 15:3280-3287.
• [34]Haferlach C, Dicker F, Schnittger S, Kern W, Haferlach T: Comprehensive genetic characterization of CLL: a study on 506 cases analysed with chromosome banding analysis, interphase FISH, IgVH status and immunophenotyping. Leukemia 2007, 21:2442-2451.
• [35]Cardinaud B, Moreilhon C, Marcet B, Robbe-Sermesant K, LeBrigand K, Mari B, Eclache V, Cymbalista F, Raynaud S, Barbry P: miR-34b/miR-34c: a regulator of TCL1 expression in 11q- chronic lymphocytic leukaemia. Leukemia 2009, 23:2174-2177.
• [36]Lodygin D, Tarasov V, Epanchintsev A, Berking C, Knyazeva T, Korner H, Knyazev P, Diebold J, Hermeking H: Inactivation of miR-34a by aberrant CpG methylation in multiple types of cancer. Cell Cycle 2008, 7:2591-2600.
• [37]Berger AH, Knudson AG, Pandolfi PP: A continuum model for tumour suppression. Nature 2011, 476:163-169.
• [38]Wong KY, Yim RLH, Kwong YL, Leung CY, Hui PK, Cheung F, Liang R, Jin DY, Chim CS: Epigenetic inactivation of the MIR129-2 in hematological malignancies. J Hematol Oncol 2013, 6:16. BioMed Central Full Text
• [39]Pigazzi M, Manara E, Baron E, Basso G: miR-34b targets cyclic AMP–responsive element binding protein in acute myeloid leukemia. Cancer Res 2009, 69:2471-2478.
• [40]Wong KY, Liang R, So CC, Jin DY, Costello JF, Chim CS: Epigenetic silencing of MIR203 in multiple myeloma. Br J Haematol 2011, 154:569-578.