Abstract
1,25-Dihydroxyvitamin D3 [1,25(OH)2D3], the active metabolite of vitamin D, is a potent inhibitor of breast cancer cell growth. Although it is evident that 1,25(OH)2D3 inhibits growth of both estrogen receptor alpha-positive [ERα(+)] and -negative [ERα(−)] breast cancer cells, the cellular pathways contributing to these effects remain unclear. We studied the gene expression patterns in ERα(+) MCF-7 and ERα(−) MDA MB 231 human breast cancer cells following 1,25(OH)2D3 treatment, using cDNA expression arrays. Both cell lines showed a significant induction of the 1,25(OH)2D3-dependent 24-hydroxylase gene, a marker for the actions of 1,25(OH)2D3. In MCF-7 cells, 51 genes were up-regulated and 19 genes were down-regulated. The up-regulated genes encoded cell adhesion molecules, growth factors/modulators, steroid receptors/co-activators, cytokines, kinases and transcription factors. Of the up-regulated genes, 40% were implicated in cell cycle regulation and apoptosis and included cyclin G1 and cyclin I, p21-activated kinase-1 (PAK-1), p53, retinoblastoma like-2 [Rb2 (p130)], insulin-like growth factor binding protein-5 (IGFBP5) and caspases. Among the down-regulated genes were ERα, growth factors, cytokines and several kinases. Some of these results were confirmed by real-time PCR. In MDA MB 231 cells, 20 genes were up-regulated and 13 genes were down-regulated. Very few genes directly implicated in cell cycle regulation were up-regulated. The matrix metalloproteinases formed a major class of genes that were down-regulated in the MDA MB 231 cells. Seven genes were commonly up-regulated in both cell lines and these included transforming growth factor (TGFβ2) and Rb2 (p130). In conclusion, the gene expression profiles of the two cell lines studied were different with a few overlapping genes suggesting that different cellular pathways might be regulated by 1,25(OH)2D3 to exert its growth inhibitory effects in ERα(+) and ERα(−) cells.
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Mangelsdorf D, Thummel C, Beato M, Herrlich P, Schutz G, Umesono K, Blumberg B, Kastner P, Mark M, Chambon P, Evans R: The nuclear receptor superfamily: the second decade. Cell 83: 835-839, 1995
Tsai MJ, O'Malley BW: Molecular mechanisms of action of steroid/thyroid receptor super family members. Annu Rev Biochem 63: 451-486, 1994
Osborne CK: Steroid hormone receptors in breast cancer management. Breast Cancer Res Treat 51: 227-238, 1998
Feldman D, Malloy PJ, Gross C: Vitamin D: Biology, Action and clinical implications. In: Marcus R, Feldman D, Kelsey J (eds) Osteoporosis. Academic Press, New York, 2001, pp 257-303
Colston KW, Hansen C: Mechanisms implicated in the growth regulatory effects of vitamin D in breast cancer. Endocr Relat Cancer 9: 45-59, 2002
Abe J, Nakano T, Nishii Y, Matsumoto T, Ogata E, Ikeda K: A novel vitamin D3 analog, 22-oxa-1,25-dihydroxyvitamin D3, inhibits the growth of human breast cancer in vitro and in vivo without causing hypercalcemia. Endocrinology 129: 832-837, 1991
Love-Schimenti CD, Gibson DFC, Ratnam AV, Bikle DD: Antiestrogens potentiation of antiproliferative effects of vitamin D3 analogs in breast cancer cells. Cancer Res 56: 2789-2794, 1996
Colston KW, Perks CM, Xie SP, Holly JM: Growth inhibition of both MCF-7 and Hs578T human breast cancer cell lines by vitamin D analogues is associated with increased expression of insulin-like growth factor binding protein-3. J Mol Endocrinol 20: 157-162, 1998
Xie SP, Pirianov G, Colston KW: Vitamin D analogues suppress IGF-I signalling and promote apoptosis in breast cancer cells. Eur J Cancer 35: 1717-1723, 1999
Simboli-Campbell M, Narvaez CJ, van Weelden K, Tenniswood M, Welsh JE: Comparative effects of 1,25(OH)2D3 and EB1089 on cell cycle kinetics and apoptosis in MCF-7 breast cancer cells. Breast Cancer Res Treat 42: 31-41, 1997
James SY, Mackay AG, Colston KW: Effects of 1,25 dihydroxyvitamin D3 and its analogues on induction of apoptosis in breast cancer cells. J Steroid Biochem Mol Biol 58: 395-401, 1996
Hansen C, Rohde L, Madsen MW, Hansen D, Colston KW, Pirianov G, Holm PK, Binderup L: MCF-7/VD(R): a new vitamin D resistant cell line. J Cell Biochem 82: 422-436, 2001
Narvaez CJ, Welsh J: Role of mitochondria and caspases in vitamin D-mediated apoptosis of MCF-7 breast cancer cells. J Biol Chem 276: 9101-9107, 2001
James SY, Mackey AG, Binderup L, Colston KW: Effects of a new synthetic vitamin D analogue, EB1089, on the oestrogenresponsive growth of human breast cancer cells. J Endocrinol 141: 555-563, 1994
Stoica A, Saceda M, Fakhro A, Solomon HB, Fenster BD, Martin MB: Regulation of estrogen receptor-alpha gene expression by 1,25-dihydroxyvitamin D in MCF-7 cells. J Cell Biochem 75: 640-651, 1999
Swami S, Krishnan AV, Feldman D: 1alpha,25-Dihydroxyvitamin D3 down-regulates estrogen receptor abundance and suppresses estrogen actions in MCF-7 human breast cancer cells. Clin Cancer Res 6: 3371-3379, 2000
Elstner E, Linker-Israeli M, Said J, Umiel T, de Vos S, Shintaku IP, Heber D, Binderup L, Uskokovic M, Koeffler HP: 20-epi-vitamin D3 analogues: a novel class of potent inhibitors of proliferation and inducers of differentiation of human breast cancer cell lines. Cancer Res 55: 2822-2830, 1995
Wang Q, Lee D, Sysounthone V, Chandraratna RAS, Christakos S, Korah R, Wieder R: 1,25-dihydroxyvitamin D3 and retonic acid analogues induce differentiation in breast cancer cells with function-and cell-specific additive effects. Breast Cancer Res Treat 67: 157-168, 2001
Raghavachari N, Bao YP, Li G, Xie X, Muller UR: Reduction of auto-fluorescence on DNA microarrays and slide surfaces by treatment with sodium borohydride. Anal Biochem 312: 101-105, 2003
Bao P, Frutos AG, Greef C, Lahiri J, Muller U, Peterson TC, Warden L, Xie X: High-sensitivity detection of DNA hybridization on microarrays using resonance light scattering. Anal Chem 74: 1792-1797, 2002
Bieche I, Parfait B, Tozlu S, Lidereau R, Vidaud M: Quantitation of androgen receptor gene expression in sporadic breast tumors by real-time RT-PCR: evidence that MYC is an ARregulated gene. Carcinogenesis 22: 1521-1526, 2001
Narvaez CJ, Zinser G, Welsh J: Functions of 1alpha,25-dihydroxyvitamin D(3) in mammary gland: from normal development to breast cancer. Steroids 66: 301-308, 2001
Davoodi F, Brenner RV, Evans SRT, Schumaker LM, Shabahang M, Nauta RJ, Buras RR: Modulation of vitamin D receptor and estrogen receptor by 1,25(OH)2D2-vitamin D3 in cDNA microarray analysis in breast cancer cells after vitamin D treatment 61 T47D human breast cancer cells. J. Steroid Biochem Mol Biol 54: 147-153, 1995
Hall JM, McDonnell DP: The estrogen receptor beta-isoform (ERbeta) of the human estrogen receptor modulates ERalpha transcriptional activity and is a key regulator of the cellular response to estrogens and antiestrogens. Endocrinology 140: 5566-5578, 1999
Krishnan A, Feldman D: Regulation of vitamin D receptor abundance. In: Feldman D, Glorieux FH, Pike JW (eds) Vitamin D. Academic Press, San Diego, CA, 1997, pp 179-200
Gross C, Krishnan AV, Malloy PJ, Eccleshall TR, Zhao XY, Feldman D: The vitamin D receptor gene start codon polymorphism: a functional analysis of FokI variants. J Bone Miner Res 13: 1691-1699, 1998
Ly LH, Zhao XY, Holloway L, Feldman D: Liarozole actssynergistically with 1alpha,25-dihydroxyvitamin D3 to inhibit growth of DU 145 human prostate cancer cells by blocking 24-hydroxylase activity. Endocrinology 140: 2071-2076, 1999
Verlinden L, Verstuyf A, Convents R, Marcelis S, Van Camp M, Bouillon R: Action of 1,25(OH)2D3 on the cell cycle genes, cyclin D1, p21 and p27 in MCF-7 cells. Mol Cell Endocrinol 142: 57-65, 1998
Kobayashi T, Hashimoto K, Yoshikawa K: Growth inhibition of human keratinocytes by 1,25-dihydroxyvitamin D3 is linked to dephosphorylation of retinoblastoma gene product. Biochem Biophys Res Commun 196: 487-493, 1993
Hansen CM, Binderup L, Hamberg KJ, Carlberg C: Vitamin D and cancer: effects of 1,25(OH)2D3 and its analogs on growth control and tumorigenesis. Front Biosci 6: D820-D848, 2001
Jensen SS, Madsen MW, Lukas J, Binderup L, Bartek J: Inhibitory effects of 1alpha,25-dihydroxyvitamin D(3) on the G(1)-S phase-controlling machinery. Mol Endocrinol 15: 1370-1380, 2001
Verlinden L, Verstuyf A, Van Camp M, Marcelis S, Sabbe K, Zhao XY, De Clercq P, Vandewalle M, Bouillon R: Two novel 14-Epi-analogues of 1,25-dihydroxyvitamin D3 inhibit the growth of human breast cancer cells in vitro and in vivo. Cancer Res 60: 2673-2679, 2000
Mathiasen IS, Lademann U, Jaattela M: Apoptosis induced by vitamin D compounds in breast cancer cells is inhibited by Bcl-2 but does not involve known caspases or p53. Cancer Res 59: 4848-4856, 1999
Thomas A, Giesler T, White E: p53 mediates bcl-2 phosphorylation and apoptosis via activation of the Cdc42/JNK1 pathway. Oncogene 19: 5259-5269, 2000
Bates S, Rowan S, Vousden KH: Characterisation of human cyclin G1 and G2: DNA damage inducible genes. Oncogene 13: 1103-1109, 1996
Okamoto K, Prives C: A role of cyclin G in the process of apoptosis. Oncogene 18: 4606-4615, 1999
Kimura SH, Ikawa M, Ito A, Okabe M, Nojima H: Cyclin G1 is involved in G2/M arrest in response to DNA damage and in growth control after damage recovery. Oncogene 20: 3290-3300, 2001
Chen L, Shinde U, Ortolan TG, Madura K: Ubiquitinassociated (UBA) domains in Rad23 bind ubiquitin and promote inhibition of multi-ubiquitin chain assembly. EMBO Rep 2: 933-938, 2001
Bertolaet BL, Clarke DJ, Wolff M, Watson MH, Henze M, Divita G, Reed SI: UBA domains of DNA damage-inducible proteins interact with ubiquitin. Nat Struct Biol 8: 417-422, 2001
Takahashi T, Caviness Jr VS: PCNA-binding to DNA at the G1/S transition in proliferating cells of the developing cerebral wall. J Neurocytol 22: 1096-1102, 1993
Paunesku T, Mittal S, Protic M, Oryhon J, Korolev SV, Joachimiak A, Woloschak GE: Proliferating cell nuclear antigen (PCNA): ringmaster of the genome. Int J Radiat Biol 77: 1007-1021, 2001
Zamparelli A, Masciullo V, Bovicelli A, Santini D, Ferrandina G, Minimo C, Terzano P, Costa S, Cinti C, Ceccarelli C, Mancuso S, Scambia G, Bovicelli L, Giordano A: Expression of cell-cycle-associated proteins pRB2/p130 and p27kip in vulvar squamous cell carcinomas. Hum Pathol 32: 4-9, 2001
Ravid A, Rocker D, Machlenkin A, Rotem C, Hochman A, Kessler-Icekson G, Liberman UA, Koren R: 1,25-Dihydroxyvitamin D3 enhances the susceptibility of breast cancer cells to doxorubicin-induced oxidative damage. Cancer Res 59: 862-867, 1999
Koren R, Hadari-Naor I, Zuck E, Rotem C, Liberman UA, Ravid A: Vitamin D is a prooxidant in breast cancer cells. Cancer Res 61: 1439-1444, 2001
Lindner DJ, Hofmann ER, Karra S, Kalvakolanu DV: The interferon-beta and tamoxifen combination induces apoptosis using thioredoxin reductase. Biochim Biophys Acta 1496: 196-206, 2000
Rozen F, Yang XF, Huynh H, Pollak M: Antiproliferative action of vitamin D-related compounds and insulin-like growth factor-binding protein 5 accumulation. J Natl Cancer Inst 89: 652-656, 1997
Boyle BJ, Zhao XY, Cohen P, Feldman D: Insulinlike growth factor binding protein-3 mediates 1 alpha,25-dihydroxyvitamin d(3) growth inhibition in the LNCaP prostate cancer cell line through p21/WAF1. J Urol 165: 1319-1324, 2001
Zumkeller W: IGFs and IGFBPs: surrogate markers for diagnosis and surveillance of tumour growth? Mol Pathol 54: 285-288, 2001
Schmidt M, Lichtner RB: EGF receptor targeting in therapyresistant human tumors. Drug Resist Updat 5: 11-18, 2002
Zhang W, Wang H, Song SW, Fuller GN: Insulin-like growth factor binding protein 2: gene expression microarrays and the hypothesis-generation paradigm. Brain Pathol 12: 87-94, 2002
Wu G, Fan RS, Li W, Srinivas V, Brattain MG: Regulation of transforming growth factor-beta type II receptor expression in human breast cancer MCF-7 cells by vitamin D3 and its analogues. J Biol Chem 273: 7749-7756, 1998
Yang L, Yang J, Venkateswarlu S, Ko T, Brattain MG: Autocrine TGFbeta signaling mediates vitamin D3 analog-induced growth inhibition in breast cells. J Cell Physiol 188: 383-393, 2001
Hu X, Zuckerman KS: Transforming growth factor: signal transduction pathways, cell cycle mediation, and effects on hematopoiesis. J Hematother Stem Cell Res 10: 67-74, 2001
Miki Y, Swensen J, Shattuck-Eidens D, Futreal PA, Harshman K, Tavtigian S, Liu Q, Cochran C, Bennett LM, Ding W, et al.: A strong candidate for the breast and ovarian cancer susceptibility gene BRCA1. Science 266: 66-71, 1994
Wang SC, Lin SH, Su LK, Hung MC: Changes in BRCA2 expression during progression of the cell cycle. Biochem Biophys Res Commun 234: 247-251, 1997
MacDougall JR, Matrisian LM: Contributions of tumor and stromal matrix metalloproteinases to tumor progression, invasion and metastasis. Cancer Metastasis Rev 14: 351-362, 1995
Koli K, Keski-Oja J: 1alpha,25-dihydroxyvitamin D3 and its analogues down-regulate cell invasion-associated proteases in cultured malignant cells. Cell Growth Diff 11: 221-229, 2000
Akutsu N, Lin R, Bastien Y, Bestawros A, Enepekides DJ, Black MJ, White JH: Regulation of gene Expression by 1alpha,25-dihydroxyvitamin D3 and its analog EB1089 under growth-inhibitory conditions in squamous carcinoma Cells. Mol Endocrinol 15: 1127-1139, 2001
Lin R, Nagai Y, Sladek R, Bastien Y, Ho J, Petrecca K, Sotiropoulou G, Diamandis EP, Hudson TJ, White JH: Expression profiling in squamous carcinoma cells reveals pleiotropic effects of vitamin D3 analog EB1089 signaling on cell proliferation, differentiation, and immune system regulation. Mol Endocrinol 16: 1243-1256, 2002
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Swami, S., Raghavachari, N., Muller, U.R. et al. Vitamin D Growth Inhibition of Breast Cancer Cells: Gene Expression Patterns Assessed by cDNA Microarray. Breast Cancer Res Treat 80, 49–62 (2003). https://doi.org/10.1023/A:1024487118457
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DOI: https://doi.org/10.1023/A:1024487118457