Cyclin D1, Id1 and EMT in breast cancer

BMC Cancer

Göran Landberg³  Sophie Lehn³  Katja L Lundgren²  Andrew H Sims¹  Nicholas P Tobin⁴ 

[1]Applied Bioinformatics of Cancer, Breakthrough Breast Cancer Research Unit, Edinburgh Cancer Research Centre, Institute of Genetics and Molecular Medicine, Crewe Road South Edinburgh, EH4 2XR, UK

[2]Breakthrough Breast Cancer Research Unit, School of Cancer, Enabling Sciences and Technology, University of Manchester, Manchester Academic Health Science Centre, Paterson Institute for Cancer Research, The Christie NHS Foundation Trust, Wilmslow Road, Manchester, M20 4BX, UK

[3]Department of Laboratory Medicine, Center for Molecular Pathology, Lund University, Malmö University Hospital, Malmö, SE-205 02, Sweden

[4]Cancer Center Karolinska, Karolinska Institute and University Hospital, Stockholm, S-17176, Sweden

关键词: claudin-low;    recurrence-free survival;    migration;    breast cancer;    EMT;    Id1;    Cyclin D1;   

Others  :  1080718
DOI  :  10.1186/1471-2407-11-417

 received in 2011-04-04, accepted in 2011-09-28,  发布年份 2011

【 摘 要 】

Background

Cyclin D1 is a well-characterised cell cycle regulator with established oncogenic capabilities. Despite these properties, studies report contrasting links to tumour aggressiveness. It has previously been shown that silencing cyclin D1 increases the migratory capacity of MDA-MB-231 breast cancer cells with concomitant increase in 'inhibitor of differentiation 1' (ID1) gene expression. Id1 is known to be associated with more invasive features of cancer and with the epithelial-mesenchymal transition (EMT). Here, we sought to determine if the increase in cell motility following cyclin D1 silencing was mediated by Id1 and enhanced EMT-features. To further substantiate these findings we aimed to delineate the link between CCND1, ID1 and EMT, as well as clinical properties in primary breast cancer.

Methods

Protein and gene expression of ID1, CCND1 and EMT markers were determined in MDA-MB-231 and ZR75 cells by western blot and qPCR. Cell migration and promoter occupancy were monitored by transwell and ChIP assays, respectively. Gene expression was analysed from publicly available datasets.

Results

The increase in cell migration following cyclin D1 silencing in MDA-MB-231 cells was abolished by Id1 siRNA treatment and we observed cyclin D1 occupancy of the Id1 promoter region. Moreover, ID1 and SNAI2 gene expression was increased following cyclin D1 knock-down, an effect reversed with Id1 siRNA treatment. Similar migratory and SNAI2 increases were noted for the ER-positive ZR75-1 cell line, but in an Id1-independent manner. In a meta-analysis of 1107 breast cancer samples, CCND1^low/ID1^high tumours displayed increased expression of EMT markers and were associated with reduced recurrence free survival. Finally, a greater percentage of CCND1^low/ID1^high tumours were found in the EMT-like 'claudin-low' subtype of breast cancer than in other subtypes.

Conclusions

These results indicate that increased migration of MDA-MB-231 cells following cyclin D1 silencing can be mediated by Id1 and is linked to an increase in EMT markers. Moreover, we have confirmed a relationship between cyclin D1, Id1 and EMT in primary breast cancer, supporting our in vitro findings that low cyclin D1 expression can be linked to aggressive features in subgroups of breast cancer.

【 授权许可】

   
2011 Tobin et al; licensee BioMed Central Ltd.

【 图 表 】

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【 参考文献 】

• [1]Kato J, Matsushime H, Hiebert SW, Ewen ME, Sherr CJ: Direct binding of cyclin D to the retinoblastoma gene product (pRb) and pRb phosphorylation by the cyclin D-dependent kinase CDK4. Genes Dev 1993, 7(3):331-342.
• [2]Lundberg AS, Weinberg RA: Functional inactivation of the retinoblastoma protein requires sequential modification by at least two distinct cyclin-cdk complexes. Mol Cell Biol 1998, 18(2):753-761.
• [3]Weinberg RA: The retinoblastoma protein and cell cycle control. Cell 1995, 81(3):323-330.
• [4]Jiang W, Kahn SM, Zhou P, Zhang YJ, Cacace AM, Infante AS, Doi S, Santella RM, Weinstein IB: Overexpression of cyclin D1 in rat fibroblasts causes abnormalities in growth control, cell cycle progression and gene expression. Oncogene 1993, 8(12):3447-3457.
• [5]Lammie GA, Fantl V, Smith R, Schuuring E, Brookes S, Michalides R, Dickson C, Arnold A, Peters G: D11S287, a putative oncogene on chromosome 11q13, is amplified and expressed in squamous cell and mammary carcinomas and linked to BCL-1. Oncogene 1991, 6(3):439-444.
• [6]Wang TC, Cardiff RD, Zukerberg L, Lees E, Arnold A, Schmidt EV: Mammary hyperplasia and carcinoma in MMTV-cyclin D1 transgenic mice. Nature 1994, 369(6482):669-671.
• [7]Zhou P, Jiang W, Zhang YJ, Kahn SM, Schieren I, Santella RM, Weinstein IB: Antisense to cyclin D1 inhibits growth and reverses the transformed phenotype of human esophageal cancer cells. Oncogene 1995, 11(3):571-580.
• [8]Jares P, Rey MJ, Fernandez PL, Campo E, Nadal A, Munoz M, Mallofre C, Muntane J, Nayach I, Estape J, et al.: Cyclin D1 and retinoblastoma gene expression in human breast carcinoma: correlation with tumour proliferation and oestrogen receptor status. J Pathol 1997, 182(2):160-166.
• [9]Sutter T, Doi S, Carnevale KA, Arber N, Weinstein IB: Expression of cyclins D1 and E in human colon adenocarcinomas. J Med 1997, 28(5-6):285-309.
• [10]Drobnjak M, Osman I, Scher HI, Fazzari M, Cordon-Cardo C: Overexpression of cyclin D1 is associated with metastatic prostate cancer to bone. Clin Cancer Res 2000, 6(5):1891-1895.
• [11]Coqueret O: Linking cyclins to transcriptional control. Gene 2002, 299(1-2):35-55.
• [12]Zwijsen RM, Wientjens E, Klompmaker R, van der Sman J, Bernards R, Michalides RJ: CDK-independent activation of estrogen receptor by cyclin D1. Cell 1997, 88(3):405-415.
• [13]Bienvenu F, Jirawatnotai S, Elias JE, Meyer CA, Mizeracka K, Marson A, Frampton GM, Cole MF, Odom DT, Odajima J, et al.: Transcriptional role of cyclin D1 in development revealed by a genetic-proteomic screen. Nature 2010, 463(7279):374-378.
• [14]Lehn S, Tobin NP, Berglund P, Nilsson K, Sims AH, Jirstrom K, Harkonen P, Lamb R, Landberg G: Downregulation of the Oncogene Cyclin D1 Increases Migratory Capacity in Breast Cancer and Is Linked to Unfavorable Prognostic Features. Am J Pathol 2010.
• [15]Ephrussi A, Church GM, Tonegawa S, Gilbert W: B lineage--specific interactions of an immunoglobulin enhancer with cellular factors in vivo. Science 1985, 227(4683):134-140.
• [16]Benezra R, Davis RL, Lockshon D, Turner DL, Weintraub H: The protein Id: a negative regulator of helix-loop-helix DNA binding proteins. Cell 1990, 61(1):49-59.
• [17]Singh J, Murata K, Itahana Y, Desprez PY: Constitutive expression of the Id-1 promoter in human metastatic breast cancer cells is linked with the loss of NF-1/Rb/HDAC-1 transcription repressor complex. Oncogene 2002, 21(12):1812-1822.
• [18]Lin CQ, Singh J, Murata K, Itahana Y, Parrinello S, Liang SH, Gillett CE, Campisi J, Desprez PY: A role for Id-1 in the aggressive phenotype and steroid hormone response of human breast cancer cells. Cancer Res 2000, 60(5):1332-1340.
• [19]Fong S, Itahana Y, Sumida T, Singh J, Coppe JP, Liu Y, Richards PC, Bennington JL, Lee NM, Debs RJ, et al.: Id-1 as a molecular target in therapy for breast cancer cell invasion and metastasis. Proc Natl Acad Sci USA 2003, 100(23):13543-13548.
• [20]Hay ED: The mesenchymal cell, its role in the embryo, and the remarkable signaling mechanisms that create it. Dev Dyn 2005, 233(3):706-720.
• [21]Trimboli AJ, Fukino K, de Bruin A, Wei G, Shen L, Tanner SM, Creasap N, Rosol TJ, Robinson ML, Eng C, et al.: Direct evidence for epithelial-mesenchymal transitions in breast cancer. Cancer Res 2008, 68(3):937-945.
• [22]Micalizzi DS, Farabaugh SM, Ford HL: Epithelial-mesenchymal transition in cancer: parallels between normal development and tumor progression. J Mammary Gland Biol Neoplasia 2010, 15(2):117-134.
• [23]Ikenouchi J, Matsuda M, Furuse M, Tsukita S: Regulation of tight junctions during the epithelium-mesenchyme transition: direct repression of the gene expression of claudins/occludin by Snail. J Cell Sci 2003, 116(Pt 10):1959-1967.
• [24]Ozdamar B, Bose R, Barrios-Rodiles M, Wang HR, Zhang Y, Wrana JL: Regulation of the polarity protein Par6 by TGFbeta receptors controls epithelial cell plasticity. Science 2005, 307(5715):1603-1609.
• [25]Hajra KM, Chen DY, Fearon ER: The SLUG zinc-finger protein represses E-cadherin in breast cancer. Cancer Res 2002, 62(6):1613-1618.
• [26]Li Y, Yang J, Luo JH, Dedhar S, Liu Y: Tubular epithelial cell dedifferentiation is driven by the helix-loop-helix transcriptional inhibitor Id1. J Am Soc Nephrol 2007, 18(2):449-460.
• [27]Gumireddy K, Li A, Gimotty PA, Klein-Szanto AJ, Showe LC, Katsaros D, Coukos G, Zhang L, Huang Q: KLF17 is a negative regulator of epithelial-mesenchymal transition and metastasis in breast cancer. Nat Cell Biol 2009, 11(11):1297-1304.
• [28]Berglund P, Stighall M, Jirstrom K, Borgquist S, Sjolander A, Hedenfalk I, Landberg G: Cyclin E overexpression obstructs infiltrative behavior in breast cancer: a novel role reflected in the growth pattern of medullary breast cancers. Cancer Res 2005, 65(21):9727-9734.
• [29]Sims AH, Smethurst GJ, Hey Y, Okoniewski MJ, Pepper SD, Howell A, Miller CJ, Clarke RB: The removal of multiplicative, systematic bias allows integration of breast cancer gene expression datasets - improving meta-analysis and prediction of prognosis. BMC Med Genomics 2008, 1:42. BioMed Central Full Text
• [30]Herschkowitz JI, Simin K, Weigman VJ, Mikaelian I, Usary J, Hu Z, Rasmussen KE, Jones LP, Assefnia S, Chandrasekharan S, et al.: Identification of conserved gene expression features between murine mammary carcinoma models and human breast tumors. Genome Biol 2007, 8(5):R76. BioMed Central Full Text
• [31]Neve RM, Chin K, Fridlyand J, Yeh J, Baehner FL, Fevr T, Clark L, Bayani N, Coppe JP, Tong F, et al.: A collection of breast cancer cell lines for the study of functionally distinct cancer subtypes. Cancer Cell 2006, 10(6):515-527.
• [32]Liang YY, Brunicardi FC, Lin X: Smad3 mediates immediate early induction of Id1 by TGF-beta. Cell research 2009, 19(1):140-148.
• [33]Gumireddy K, Li A, Gimotty PA, Klein-Szanto AJ, Showe LC, Katsaros D, Coukos G, Zhang L, Huang Q: KLF17 is a negative regulator of epithelial-mesenchymal transition and metastasis in breast cancer. Nature cell biology 2009, 11(11):1297-1304.
• [34]Gautschi O, Tepper CG, Purnell PR, Izumiya Y, Evans CP, Green TP, Desprez PY, Lara PN, Gandara DR, Mack PC, et al.: Regulation of Id1 expression by SRC: implications for targeting of the bone morphogenetic protein pathway in cancer. Cancer research 2008, 68(7):2250-2258.
• [35]Fu J, Qin L, He T, Qin J, Hong J, Wong J, Liao L, Xu J: The TWIST/Mi2/NuRD protein complex and its essential role in cancer metastasis. Cell Res 2010.
• [36]Gilles C, Polette M, Zahm JM, Tournier JM, Volders L, Foidart JM, Birembaut P: Vimentin contributes to human mammary epithelial cell migration. J Cell Sci 1999, 112(Pt 24):4615-4625.
• [37]Charafe-Jauffret E, Ginestier C, Iovino F, Wicinski J, Cervera N, Finetti P, Hur MH, Diebel ME, Monville F, Dutcher J, et al.: Breast cancer cell lines contain functional cancer stem cells with metastatic capacity and a distinct molecular signature. Cancer Res 2009, 69(4):1302-1313.
• [38]Kao J, Salari K, Bocanegra M, Choi YL, Girard L, Gandhi J, Kwei KA, Hernandez-Boussard T, Wang P, Gazdar AF, et al.: Molecular profiling of breast cancer cell lines defines relevant tumor models and provides a resource for cancer gene discovery. PLoS One 2009, 4(7):e6146.
• [39]Lapuk A, Marr H, Jakkula L, Pedro H, Bhattacharya S, Purdom E, Hu Z, Simpson K, Pachter L, Durinck S, et al.: Exon-level microarray analyses identify alternative splicing programs in breast cancer. Mol Cancer Res 2010, 8(7):961-974.
• [40]Creighton CJ, Chang JC, Rosen JM: Epithelial-mesenchymal transition (EMT) in tumor-initiating cells and its clinical implications in breast cancer. J Mammary Gland Biol Neoplasia 2010, 15(2):253-260.
• [41]Prat A, Parker JS, Karginova O, Fan C, Livasy C, Herschkowitz JI, He X, Perou CM: Phenotypic and molecular characterization of the claudin-low intrinsic subtype of breast cancer. Breast Cancer Res 2010, 12(5):R68. BioMed Central Full Text
• [42]Creighton CJ, Li X, Landis M, Dixon JM, Neumeister VM, Sjolund A, Rimm DL, Wong H, Rodriguez A, Herschkowitz JI, et al.: Residual breast cancers after conventional therapy display mesenchymal as well as tumor-initiating features. Proc Natl Acad Sci USA 2009, 106(33):13820-13825.
• [43]Caldon CE, Swarbrick A, Lee CS, Sutherland RL, Musgrove EA: The helix-loop-helix protein Id1 requires cyclin D1 to promote the proliferation of mammary epithelial cell acini. Cancer Res 2008, 68(8):3026-3036.
• [44]Swarbrick A, Akerfeldt MC, Lee CS, Sergio CM, Caldon CE, Hunter LJ, Sutherland RL, Musgrove EA: Regulation of cyclin expression and cell cycle progression in breast epithelial cells by the helix-loop-helix protein Id1. Oncogene 2005, 24(3):381-389.
• [45]Lee JY, Kang MB, Jang SH, Qian T, Kim HJ, Kim CH, Kim Y, Kong G: Id-1 activates Akt-mediated Wnt signaling and p27(Kip1) phosphorylation through PTEN inhibition. Oncogene 2009, 28(6):824-831.
• [46]Chen H, Zhu G, Li Y, Padia RN, Dong Z, Pan ZK, Liu K, Huang S: Extracellular signal-regulated kinase signaling pathway regulates breast cancer cell migration by maintaining slug expression. Cancer Res 2009, 69(24):9228-9235.
• [47]Bolos V, Peinado H, Perez-Moreno MA, Fraga MF, Esteller M, Cano A: The transcription factor Slug represses E-cadherin expression and induces epithelial to mesenchymal transitions: a comparison with Snail and E47 repressors. J Cell Sci 2003, 116(Pt 3):499-511.
• [48]Come C, Magnino F, Bibeau F, De Santa Barbara P, Becker KF, Theillet C, Savagner P: Snail and slug play distinct roles during breast carcinoma progression. Clin Cancer Res 2006, 12(18):5395-5402.
• [49]Mironchik Y, Winnard PT Jr, Vesuna F, Kato Y, Wildes F, Pathak AP, Kominsky S, Artemov D, Bhujwalla Z, Van Diest P, et al.: Twist overexpression induces in vivo angiogenesis and correlates with chromosomal instability in breast cancer. Cancer Res 2005, 65(23):10801-10809.
• [50]Nakajima G, Patino-Garcia A, Bruheim S, Xi Y, San Julian M, Lecanda F, Sierrasesumaga L, Muller C, Fodstad O, Ju J: CDH11 expression is associated with survival in patients with osteosarcoma. Cancer Genomics Proteomics 2008, 5(1):37-42.
• [51]Berns EM, Foekens JA, van Staveren IL, van Putten WL, de Koning HY, Portengen H, Klijn JG: Oncogene amplification and prognosis in breast cancer: relationship with systemic treatment. Gene 1995, 159(1):11-18.
• [52]Bieche I, Olivi M, Nogues C, Vidaud M, Lidereau R: Prognostic value of CCND1 gene status in sporadic breast tumours, as determined by real-time quantitative PCR assays. Br J Cancer 2002, 86(4):580-586.
• [53]Gillett C, Smith P, Gregory W, Richards M, Millis R, Peters G, Barnes D: Cyclin D1 and prognosis in human breast cancer. Int J Cancer 1996, 69(2):92-99.
• [54]Hwang TS, Han HS, Hong YC, Lee HJ, Paik NS: Prognostic value of combined analysis of cyclin D1 and estrogen receptor status in breast cancer patients. Pathol Int 2003, 53(2):74-80.
• [55]Han S, Park K, Bae BN, Kim KH, Kim HJ, Kim YD, Kim HY: Cyclin D1 expression and patient outcome after tamoxifen therapy in estrogen receptor positive metastatic breast cancer. Oncol Rep 2003, 10(1):141-144.
• [56]Taneja P, Maglic D, Kai F, Zhu S, Kendig RD, Fry EA, Inoue K: Classical and Novel Prognostic Markers for Breast Cancer and their Clinical Significance. Clin Med Insights Oncol 2010, 4:15-34.
• [57]Kenny FS, Hui R, Musgrove EA, Gee JM, Blamey RW, Nicholson RI, Sutherland RL, Robertson JF: Overexpression of cyclin D1 messenger RNA predicts for poor prognosis in estrogen receptor-positive breast cancer. Clin Cancer Res 1999, 5(8):2069-2076.
• [58]Mimeault M, Batra SK: Functions of tumorigenic and migrating cancer progenitor cells in cancer progression and metastasis and their therapeutic implications. Cancer Metastasis Rev 2007, 26(1):203-214.
• [59]Mimeault M, Batra SK: Interplay of distinct growth factors during epithelial mesenchymal transition of cancer progenitor cells and molecular targeting as novel cancer therapies. Ann Oncol 2007, 18(10):1605-1619.
• [60]Blick T, Widodo E, Hugo H, Waltham M, Lenburg ME, Neve RM, Thompson EW: Epithelial mesenchymal transition traits in human breast cancer cell lines. Clin Exp Metastasis 2008, 25(6):629-642.
• [61]Uehara N, Chou YC, Galvez JJ, de-Candia P, Cardiff RD, Benezra R, Shyamala G: Id-1 is not expressed in the luminal epithelial cells of mammary glands. Breast Cancer Res 2003, 5(2):R25-29. BioMed Central Full Text
• [62]Perk J, Gil-Bazo I, Chin Y, de Candia P, Chen JJ, Zhao Y, Chao S, Cheong W, Ke Y, Al-Ahmadie H, et al.: Reassessment of id1 protein expression in human mammary, prostate, and bladder cancers using a monospecific rabbit monoclonal anti-id1 antibody. Cancer Res 2006, 66(22):10870-10877.
• [63]Hennessy BT, Gonzalez-Angulo AM, Stemke-Hale K, Gilcrease MZ, Krishnamurthy S, Lee JS, Fridlyand J, Sahin A, Agarwal R, Joy C, et al.: Characterization of a naturally occurring breast cancer subset enriched in epithelial-to-mesenchymal transition and stem cell characteristics. Cancer Res 2009, 69(10):4116-4124.
• [64]Weigelt B, Kreike B, Reis-Filho JS: Metaplastic breast carcinomas are basal-like breast cancers: a genomic profiling analysis. Breast Cancer Res Treat 2009, 117(2):273-280.
• [65]Luini A, Aguilar M, Gatti G, Fasani R, Botteri E, Brito JA, Maisonneuve P, Vento AR, Viale G: Metaplastic carcinoma of the breast, an unusual disease with worse prognosis: the experience of the European Institute of Oncology and review of the literature. Breast Cancer Res Treat 2007, 101(3):349-353.
• [66]Farmer P, Bonnefoi H, Anderle P, Cameron D, Wirapati P, Becette V, Andre S, Piccart M, Campone M, Brain E, et al.: A stroma-related gene signature predicts resistance to neoadjuvant chemotherapy in breast cancer. Nat Med 2009, 15(1):68-74.