TGF-β Regulates DNA Methyltransferase Expression in Prostate Cancer, Correlates with Aggressive Capabilities, and Predicts Disease Recurrence

Background DNA methyltransferase (DNMT) is one of the major factors mediating the methylation of cancer related genes such as TGF-β receptors (TβRs). This in turn may result in a loss of sensitivity to physiologic levels of TGF-β in aggressive prostate cancer (CaP). The specific mechanisms of DNMT's role in CaP remain undetermined. In this study, we describe the mechanism of TGF-β-mediated DNMT in CaP and its association with clinical outcomes following radical prostatectomy. Methodology/Principal Findings We used human CaP cell lines with varying degrees of invasive capability to describe how TGF-β mediates the expression of DNMT in CaP, and its effects on methylation status of TGF-β receptors and the invasive capability of CaP in vitro and in vivo. Furthermore, we determined the association between DNMT expression and clinical outcome after radical prostatectomy. We found that more aggressive CaP cells had significantly higher TGF-β levels, increased expression of DNMT, but reduced TβRs when compared to benign prostate cells and less aggressive prostate cancer cells. Blockade of TGF-β signaling or ERK activation (p-ERK) was associated with a dramatic decrease in the expression of DNMT, which results in a coincident increase in the expression of TβRs. Blockade of either TGF-β signaling or DNMT dramatically decreased the invasive capabilities of CaP. Inhibition of TGF-β in an TRAMP-C2 CaP model in C57BL/6 mice using 1D11 was associated with downregulation of DNMTs and p-ERK and impairment in tumor growth. Finally, independent of Gleason grade, increased DNMT1 expression was associated with biochemical recurrence following surgical treatment for prostate cancer. Conclusions and Significance Our findings demonstrate that CaP derived TGF-β may induce the expression of DNMTs in CaP which is associated with methylation of its receptors and the aggressive potential of CaP. In addition, DNMTs is an independent predictor for disease recurrence after prostatectomy, and may have clinical implications for CaP prognostication and therapy.

[1]  Ximing J. Yang,et al.  Tumor Evasion of the Immune System by Converting CD4+CD25− T Cells into CD4+CD25+ T Regulatory Cells: Role of Tumor-Derived TGF-β , 2007, The Journal of Immunology.

[2]  Martin R. Johnson,et al.  Hedgehog Pathway Expression in Heterogeneous Pancreatic Adenocarcinoma: Implications for the Molecular Analysis of Clinically Available Biopsies , 2007, Diagnostic molecular pathology : the American journal of surgical pathology, part B.

[3]  Miao-Fen Chen,et al.  Role of DNA methyltransferase 1 in hormone-resistant prostate cancer , 2010, Journal of Molecular Medicine.

[4]  K. Amara,et al.  DNA methyltransferase DNMT3b protein overexpression as a prognostic factor in patients with diffuse large B‐cell lymphomas , 2010, Cancer science.

[5]  R. Dahiya,et al.  DNA methyltransferase and demethylase in human prostate cancer , 2002, Molecular carcinogenesis.

[6]  Mike Wilson,et al.  Selection of highly metastatic variants of different human prostatic carcinomas using orthotopic implantation in nude mice. , 1996, Clinical cancer research : an official journal of the American Association for Cancer Research.

[7]  T. Fandy.  Development of DNA methyltransferase inhibitors for the treatment of neoplastic diseases. , 2009, Current medicinal chemistry.

[8]  Keith D Robertson,et al.  DNA methylation, methyltransferases, and cancer , 2001, Oncogene.

[9]  Chung Lee,et al.  Genetic change in transforming growth factor β (TGF-β) receptor type I gene correlates with insensitivity to TGF-β1 in human prostate cancer cells , 1996 .

[10]  S. Hirohashi,et al.  Increased protein expression of DNA methyltransferase (DNMT) 1 is significantly correlated with the malignant potential and poor prognosis of human hepatocellular carcinomas , 2003, International journal of cancer.

[11]  M. Spitz,et al.  Expression of methylation-related genes is associated with overall survival in patients with non-small cell lung cancer , 2008, British Journal of Cancer.

[12]  A. Moustakas,et al.  Actions of TGF-beta as tumor suppressor and pro-metastatic factor in human cancer. , 2007, Biochimica et biophysica acta.

[13]  B. Teicher,et al.  Treatment of transforming growth factor-beta-insensitive mouse Renca tumor by transforming growth factor-beta elimination. , 2008, Urology.

[14]  P. Jones,et al.  Inhibition of DNA methylation by 5-aza-2'-deoxycytidine suppresses the growth of human tumor cell lines. , 1998, Cancer research.

[15]  Jing-Yuan Fang,et al.  Inhibition of the Extracellular Signal-regulated Kinase/Mitogen-activated Protein Kinase Pathway Decreases DNA Methylation in Colon Cancer Cells* , 2007, Journal of Biological Chemistry.

[16]  C. Lee,et al.  Growth requirements of human prostate cancers in vitro and in vivo , 1991 .

[17]  S. Kitano,et al.  Increased DNA methyltransferase 1 (DNMT1) protein expression correlates significantly with poorer tumor differentiation and frequent DNA hypermethylation of multiple CpG islands in gastric cancers. , 2004, The American journal of pathology.

[18]  J. Herman,et al.  Alterations in DNA methylation: a fundamental aspect of neoplasia. , 1998, Advances in cancer research.

[19]  Chung Lee,et al.  In vivo andin vitro approaches to study metastasis in human prostatic cancer , 1993, Cancer and Metastasis Reviews.

[20]  R. Blasberg,et al.  A novel triple-modality reporter gene for whole-body fluorescent, bioluminescent, and nuclear noninvasive imaging , 2004, European Journal of Nuclear Medicine and Molecular Imaging.

[21]  Ximing J. Yang,et al.  Blockade of transforming growth factor-β signaling in tumor-reactive CD8+ T cells activates the antitumor immune response cycle , 2006, Molecular Cancer Therapeutics.

[22]  Jing Qing,et al.  TGF-beta activates Erk MAP kinase signalling through direct phosphorylation of ShcA. , 2007, The EMBO journal.

[23]  Ximing J. Yang,et al.  Adoptive transfer of tumor-reactive transforming growth factor-beta-insensitive CD8+ T cells: eradication of autologous mouse prostate cancer. , 2005, Cancer research.

[24]  K. Miyazono,et al.  Loss of expression of transforming growth factor beta type I and type II receptors correlates with tumor grade in human prostate cancer tissues. , 1996, Clinical cancer research : an official journal of the American Association for Cancer Research.

[25]  Q. Lu,et al.  Demethylation of the Same Promoter Sequence Increases CD70 Expression in Lupus T Cells and T Cells Treated with Lupus-Inducing Drugs1 , 2005, The Journal of Immunology.

[26]  D. Haber,et al.  DNA Methyltransferases Dnmt3a and Dnmt3b Are Essential for De Novo Methylation and Mammalian Development , 1999, Cell.

[27]  Ximing J. Yang,et al.  Nuclear Factor-κB-Mediated Transforming Growth Factor-β-Induced Expression of Vimentin Is an Independent Predictor of Biochemical Recurrence after Radical Prostatectomy , 2009, Clinical Cancer Research.

[28]  Chung Lee,et al.  Dual regulation of proliferation and growth arrest in prostatic stromal cells by transforming growth factor-beta1. , 2003, Endocrinology.

[29]  P. Barr,et al.  On the mechanism of inhibition of DNA-cytosine methyltransferases by cytosine analogs , 1983, Cell.

[30]  A. Moustakas,et al.  Actions of TGF-β as tumor suppressor and pro-metastatic factor in human cancer , 2007 .

[31]  B. Helfand,et al.  Abstract LB-3: The recruitment of PP2A by TGF-β receptors mediates the response to TGF-β-induced activation of ERK in prostate cancer , 2011 .

[32]  M. Szyf,et al.  Regulation of DNA methylation by the Ras signaling pathway , 1995, The Journal of Biological Chemistry.

[33]  Z. Xia,et al.  Cutting Edge: TGF-β-Induced Expression of Foxp3 in T cells Is Mediated through Inactivation of ERK , 2008, The Journal of Immunology.

[34]  Ming-Rong Wang,et al.  Anticancer drug discovery targeting DNA hypermethylation. , 2008, Current medicinal chemistry.

[35]  Shoumei Bai,et al.  DNA methyltransferases as targets for cancer therapy. , 2007, Drugs of today.

[36]  T. Bestor,et al.  The DNA methyltransferases of mammals. , 2000, Human molecular genetics.

[37]  B. Richardson,et al.  Relative quantitation of DNA methyltransferase mRNA by real-time RT-PCR assay. , 2004, Methods in molecular biology.

[38]  J. Massagué,et al.  Multimodality imaging of TGFβ signaling in breast cancer metastases , 2009, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[39]  A. Jeltsch,et al.  Mechanism of inhibition of DNA methyltransferases by cytidine analogs in cancer therapy , 2004, Cancer biology & therapy.

[40]  R J Roberts,et al.  Recombinant Human DNA (Cytosine-5) Methyltransferase , 1999, The Journal of Biological Chemistry.

[41]  Zhiyong Zhang,et al.  Hydralazine may induce autoimmunity by inhibiting extracellular signal-regulated kinase pathway signaling. , 2003, Arthritis & Rheumatism.

[42]  G. Adler,et al.  TGFβ1 represses proliferation of pancreatic carcinoma cells which correlates with Smad4-independent inhibition of ERK activation , 2000, Oncogene.

[43]  A. Albini,et al.  Prostate cancer and the invasive phenotype: application of new in vivo and in vitro approaches , 1988 .

[44]  Jing Qing,et al.  TGF‐β activates Erk MAP kinase signalling through direct phosphorylation of ShcA , 2007 .

[45]  Ximing J. Yang,et al.  Insensitivity to transforming growth factor-beta results from promoter methylation of cognate receptors in human prostate cancer cells (LNCaP). , 2005, Molecular endocrinology.

[46]  D. Santi,et al.  Kinetic and catalytic mechanism of HhaI methyltransferase. , 1987, The Journal of biological chemistry.

[47]  Chih-Yi Chen,et al.  The tobacco-specific carcinogen NNK induces DNA methyltransferase 1 accumulation and tumor suppressor gene hypermethylation in mice and lung cancer patients. , 2010, The Journal of clinical investigation.

[48]  Chung Lee,et al.  Transforming Growth Factor-β1-Induced Proliferation of the Prostate Cancer Cell Line, TSU-Pr1: The Role of Platelet-Derived Growth Factor. , 1999, Endocrinology.

[49]  G. Liang,et al.  Relationship between transcription and DNA methylation. , 2000, Current topics in microbiology and immunology.