Compound Kushen Injection suppresses human breast cancer stem-like cells by down-regulating the canonical Wnt/β-catenin pathway

Journal of Experimental & Clinical Cancer Research

Liangliang Yang²  Zhizheng Zhao²  Xiaoyun Zhu³  Yingxia Pei²  Xin Qi²  Wei Hou²  Baojin Hua²  Xinyi Chen¹  Ying Zhang²  Hongsheng Lin²  Weiru Xu¹ 

[1] Department of Hematology and Oncology, Dong Zhi Men Hospital Affiliated to Beijing University of Chinese Medicine, No. 5, Haiyuncang, Dongcheng District, Beijing 100700, China;Oncology Department, Guang An Men Hospital, China Academy of Chinese Medical Sciences, No.5 Bei Xian Ge Street, Xicheng District, Beijing 100053, China;Endocrinology Department, Guang An Men Hospital, China Academy of Chinese Medical Sciences, No.5 Bei Xian Ge Street, Xicheng District, Beijing 100053, China

关键词: cisplatin;    Wnt/β-catenin signaling;    MCF-7;    Compound Kushen Injection;    side population;    cancer stem-like cells;   

Others  :  827097
DOI  :  10.1186/1756-9966-30-103

 received in 2011-08-11, accepted in 2011-10-28,  发布年份 2011

【 摘 要 】

Background

Cancer stem cells (CSCs) play an important role in cancer initiation, relapse and metastasis. To date, no specific medicine has been found to target CSCs as they are resistant to most conventional therapies and proliferate indefinitely. Compound Kushen Injection (CKI) has been widely used for cancer patients with remarkable therapeutic effects in Chinese clinical settings for many years. This study focused on whether CKI could inhibit MCF-7 SP cells in vitro and in vivo.

Methods

The analysis of CKI on SP population and the main genes of Wnt signaling pathway were studied first. Then we studied the tumorigenicity of SP cells and the effects of CKI on SP cells in vivo. The mice inoculated with 10,000 SP cells were randomly divided into three groups (6 in each group) and treated with CKI, cisplatin and saline (as a control) respectively for 7 weeks. The tumor formation rates of each group were compared. The main genes and proteins of the Wnt signaling pathway were analyzed by RT-PCR and western blot.

Results

CKI suppressed the size of SP population (approximately 90%), and down-regulated the main genes of Wnt signaling pathway. We also determined that MCF-7 SP cells were more tumorigenic than non-SP and unsorted cells. The Wnt signaling pathway was up-regulated in tumors derived from SP cells compared with that in tumors from non-SP cells. The tumor formation rate of the CKI Group was 33% (2/6, P < 0.05), and that of Cisplatin Group was 50%(3/6, P < 0.05), whereas that of the Control Group was 100% (6/6).The RT-PCR and western blot results indicated that CKI suppressed tumor growth by down-regulating the Wnt/β-catenin pathway, while cisplatin activated the Wnt/β-catenin pathway and might spare SP cells.

Conclusions

It suggested that CKI may serve as a novel drug targeting cancer stem-like cells, though further studies are recommended.

【 授权许可】

   
2011 Xu et al; licensee BioMed Central Ltd.

【 图 表 】

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【 参考文献 】

• [1]Reya T, Morrison SJ, Clarke MF, Weissman IL: Stem cells, cancer, and cancer stem cells. Nature 2001, 414:105-111.
• [2]Gottesman MM: Mechanisms of cancer drug resistance. Annu Rev Med 2002, 53:615-627.
• [3]Zhou S, Schuetz JD, Bunting KD, Colapietro AM, Sampath J, Morris JJ, Lagutina I, Grosveld GC, Osawa M, Nakauchi H, Sorrentino BP: The ABC transporter Bcrp1/ABCG2 is expressed in a wide variety of stem cells and is a molecular determinant of the side-population phenotype. Nat Med 2001, 7:1028-1034.
• [4]Bao S, Wu Q, Mclendon RE, Hao Y, Shi Q, Hjelmeland AB, Dewhirst MW, Bigner DD, Rich JN: Glioma stem cells promote radioresistance by preferential activation of the DNA damage response. Nature 2006, 444:756-760.
• [5]Graham SM, Jorgensen HG, Allan E, Pearson C, Alcorn MJ, Richmond L, Holyoake TL: Primitive, quiescent, Philadelphia-positive stem cells from patients with chronic myeloid leukemia are insensitive to STI571 in vitro. Blood 2002, 99:319-325.
• [6]Reim F, Dombrowski Y, Ritter C, Buttmann M, Hausler S, Ossadnik M, Krockenberger M, Beier D, Beier CP, Dietl J, Becker JC, Honig A, Wischhusen J: Immunoselection of breast and ovarian cancer cells with trastuzumab and natural killer cells: selective escape of CD44high/CD24low/HER2low breast cancer stem cells. Cancer Res 2009, 69:8058-8066.
• [7]Bonnet D, Dick JE: Human acute myeloid leukemia is organized as a hierarchy that originates from a primitive hematopoietic cell. Nat Med 1997, 3:730-737.
• [8]Al-Hajj M, Wicha MS, Benito-Hernandez A, Morrison SJ, Clarke MF: Prospective identification of tumorigenic breast cancer cells. Proc Natl Acad Sci USA 2003, 100:3983-3988.
• [9]Singh SK, Hawkins C, Clarke ID, Squire JA, Bayani J, Hide T, Henkelman RM, Cusimano MD, Dirks PB: Identification of human brain tumour initiating cells. Nature 2004, 432:396-401.
• [10]Richardson GD, Robson CN, Lang SH, Neal DE, Maitland NJ, Collins AT: CD133, a novel marker for human prostatic epithelial stem cells. J Cell Sci 2004, 117:3539-3545.
• [11]Haraguchi N, Inoue H, Tanaka F, Mimori K, Utsunomiya T, Sasaki A, Mori M: Cancer stem cells in human gastrointestinal cancers. Hum Cell 2006, 19:24-29.
• [12]Kondo T, Setoguchi T, Taga T: Persistence of a small subpopulation of cancer stem-like cells in the C6 glioma cell line. Proc Natl Acad Sci USA 2004, 101:781-786.
• [13]Haraguchi N, Utsunomiya T, Inoue H, Tanaka F, Mimori K, Barnard GF, Mori M: Characterization of a side population of cancer cells from human gastrointestinal system. Stem Cells 2006, 24:506-513.
• [14]Patrawala L, Calhoun T, Schneider-Broussard R, Zhou J, Claypool K, Tang DG: Side population is enriched in tumorigenic, stem-like cancer cells, whereas ABCG2+ and ABCG2- cancer cells are similarly tumorigenic. Cancer Res 2005, 65:6207-6219.
• [15]Wang J, Guo LP, Chen LZ, Zeng YX, Lu SH: Identification of cancer stem cell-like side population cells in human nasopharyngeal carcinoma cell line. Cancer Res 2007, 67:3716-3724.
• [16]Brown MD, Gilmore PE, Hart CA, Samuel JD, Ramani VA, George NJ, Clarke NW: Characterization of benign and malignant prostate epithelial Hoechst 33342 side populations. Prostate 2007, 67:1384-1396.
• [17]Seigel GM, Campbell LM, Narayan M, Gonzalez-Fernandez F: Cancer stem cell characteristics in retinoblastoma. Mol Vis 2005, 11:729-737.
• [18]Tian J, Wang WH, Gao HM, Wang ZM: [Determination of matrine, sophoridine and oxymatrine in Compound Kushen Injection by HPLC]. Zhongguo Zhong Yao Za Zhi 2007, 32:222-224.
• [19]Wang ZY, Li GS, Huang HX: [Clinical observation on treatment of 75 mid-late stage cancer patients with yanshu Injection]. Zhongguo Zhong Xi Yi Jie He Za Zhi 2006, 26:681-684.
• [20]Chen J, Mei Q, Xu YC, Du J, Wei Y, Xu ZM: [Effects of Matrine Injection on T-lymphocyte subsets of patients with malignant tumor after gamma knife radiosurgery]. Zhong Xi Yi Jie He Xue Bao 2006, 4:78-79.
• [21]Dai ZJ, Gao J, Wang XJ, Ji ZZ, Wu WY, Liu XX, Kang HF, Guan HT, Ren HT: [Apoptotic mechanism of gastric carcinoma cells induced by matrine injection]. Zhonghua Wei Chang Wai Ke Za Zhi 2008, 11:261-265.
• [22]Dai ZJ, Gao J, Wu WY, Wang XJ, Li ZF, Kang HF, Liu XX, Ma XB: [Effect of matrine injections on invasion and metastasis of gastric carcinoma SGC-7901 cells in vitro]. Zhong Yao Cai 2007, 30:815-819.
• [23]Brown AM: Wnt signaling in breast cancer: have we come full circle? Breast Cancer Res 2001, 3:351-355. BioMed Central Full Text
• [24]Yang W, Yan HX, Chen L, Liu Q, He YQ, Yu LX, Zhang SH, Huang DD, Tang L, Kong XN, Chen C, Liu SQ, Wu MC, Wang HY: Wnt/beta-catenin signaling contributes to activation of normal and tumorigenic liver progenitor cells. Cancer Res 2008, 68:4287-4295.
• [25]Ysebaert L, Chicanne G, Demur C, De Toni F, Prade-Houdellier N, Ruidavets JB, Mansat-De Mas V, Rigal-Huguet F, Laurent G, Payrastre B, Manenti S, Racaud-Sultan C: Expression of beta-catenin by acute myeloid leukemia cells predicts enhanced clonogenic capacities and poor prognosis. Leukemia 2006, 20:1211-1216.
• [26]Malanchi I, Peinado H, Kassen D, Hussenet T, Metzger D, Chambon P, Huber M, Hohl D, Cano A, Birchmeier W, Huelsken J: Cutaneous cancer stem cell maintenance is dependent on beta-catenin signalling. Nature 2008, 452:650-653.
• [27]Li X, Ren J: [Isolation of CD44+/CD24 -/low and side population cells from MDA-MB-453 cells and the analysis of their activation of Wnt and Notch pathway]. Beijing Da Xue Xue Bao 2008, 40:471-475.
• [28]Zhang Y, Piao B, Hua B, Hou W, Xu W, Qi X, Zhu X, Pei Y, Lin H: Oxymatrine diminishes the side population and inhibits the expression of beta-catenin in MCF-7 breast cancer cells. Med Oncol 2010, in press.
• [29]Goodell MA, Brose K, Paradis G, Conner AS, Mulligan RC: Isolation and functional properties of murine hematopoietic stem cells that are replicating in vivo. J Exp Med 1996, 183:1797-1806.
• [30]Hardman WE, Moyer MP, Cameron IL: Efficacy of treatment of colon, lung and breast human carcinoma xenografts with: doxorubicin, cisplatin, irinotecan or topotecan. Anticancer Res 1999, 19:2269-2274.
• [31]Livak KJ, Schmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. Methods 2001, 25:402-408.
• [32]He B, You L, Uematsu K, Xu Z, Lee AY, Matsangou M, Mccormick F, Jablons DM: A monoclonal antibody against Wnt-1 induces apoptosis in human cancer cells. Neoplasia 2004, 6:7-14.
• [33]Willert K, Brown JD, Danenberg E, Duncan AW, Weissman IL, Reya T, Yates JR, Nusse R: Wnt proteins are lipid-modified and can act as stem cell growth factors. Nature 2003, 423:448-452.
• [34]Behrens J, Von Kries JP, Kuhl M, Bruhn L, Wedlich D, Grosschedl R, Birchmeier W: Functional interaction of beta-catenin with the transcription factor LEF-1. Nature 1996, 382:638-642.
• [35]Cadigan KM, Nusse R: Wnt signaling: a common theme in animal development. Genes Dev 1997, 11:3286-3305.
• [36]Khan NI, Bradstock KF, Bendall LJ: Activation of Wnt/beta-catenin pathway mediates growth and survival in B-cell progenitor acute lymphoblastic leukaemia. Br J Haematol 2007, 138:338-348.
• [37]Woodward WA, Chen MS, Behbod F, Alfaro MP, Buchholz TA, Rosen JM: WNT/beta-catenin mediates radiation resistance of mouse mammary progenitor cells. Proc Natl Acad Sci USA 2007, 104:618-623.
• [38]Schulenburg A, Cech P, Herbacek I, Marian B, Wrba F, Valent P, Ulrich-Pur H: CD44-positive colorectal adenoma cells express the potential stem cell markers musashi antigen (msi1) and ephrin B2 receptor (EphB2). J Pathol 2007, 213:152-160.
• [39]Peifer M, Polakis P: Wnt signaling in oncogenesis and embryogenesis--a look outside the nucleus. Science 2000, 287:1606-1609.
• [40]Bienz M, Clevers H: Linking colorectal cancer to Wnt signaling. Cell 2000, 103:311-320.
• [41]Polakis P: Wnt signaling and cancer. Genes Dev 2000, 14:1837-1851.
• [42]Nelson WJ, Nusse R: Convergence of Wnt, beta-catenin, and cadherin pathways. Science 2004, 303:1483-1487.
• [43]Morrison SJ, Kimble J: Asymmetric and symmetric stem-cell divisions in development and cancer. Nature 2006, 441:1068-1074.
• [44]Bertolini G, Roz L, Perego P, Tortoreto M, Fontanella E, Gatti L, Pratesi G, Fabbri A, Andriani F, Tinelli S, Roz E, Caserini R, Lo Vullo S, Camerini T, Mariani L, Delia D, Calabro E, Pastorino U, Sozzi G: Highly tumorigenic lung cancer CD133+ cells display stem-like features and are spared by cisplatin treatment. Proc Natl Acad Sci USA 2009, 106:16281-16286.
• [45]Cortes-Dericks L, Carboni GL, Schmid RA, Karoubi G: Putative cancer stem cells in malignant pleural mesothelioma show resistance to cisplatin and pemetrexed. Int J Oncol 2010, 37:437-444.
• [46]Honoki K, Fujii H, Kubo A, Kido A, Mori T, Tanaka Y, Tsujiuchi T: Possible involvement of stem-like populations with elevated ALDH1 in sarcomas for chemotherapeutic drug resistance. Oncol Rep 2010, 24:501-505.