Modulation by decitabine of gene expression and growth of osteosarcoma U2OS cells in vitro and in xenografts: Identification of apoptotic genes as targets for demethylation

被引:38
作者
Al-Romaih, Khaldoun [1 ,2 ]
Somers, Gino R. [1 ,3 ]
Bayani, Jane [2 ]
Hughes, Simon [7 ]
Prasad, Mona [2 ]
Cutz, Jean-Claude [5 ,6 ]
Xue, Hui [4 ]
Zielenska, Maria [1 ,3 ]
Wang, Yuzhuo [4 ,8 ]
Squire, Jeremy A. [1 ,2 ,4 ]
机构
[1] Univ Toronto, Dept Lab Med & Pathobiol, Toronto, ON M5G 1L5, Canada
[2] Princess Margaret Hosp, Ontario Canc Inst, Toronto, ON M5G 2M9, Canada
[3] Hosp Sick Children, Dept Pediat Lab Med, Toronto, ON M5G 1X8, Canada
[4] British Columbia Canc Agcy, Dept Canc Endocrinol, Vancouver, BC V5Z 1L3, Canada
[5] McMaster Univ, Dept Pathol & Mol Med, St Josephs Healthcare Hamilton Reg Lab Med Progra, Hamilton, ON L8N 4A6, Canada
[6] McMaster Univ, Dept Lab Med, St Josephs Healthcare Hamilton Reg Lab Med Progra, Hamilton, ON L8N 4A6, Canada
[7] John Vane Sci Ctr, Div Tumor Biol, Inst Canc & Canc Res, UK Clin Ctr, London, England
[8] Vancouver Gen Hosp, Prostate Ctr, Vancouver, BC V6H 3Z6, Canada
关键词
Acute Myeloid Leukemia; Transcriptional Start Site; Decitabine; U2OS Cell; Apoptotic Gene;
D O I
10.1186/1475-2867-7-14
中图分类号
R73 [肿瘤学];
学科分类号
100214 ;
摘要
Background: Methylation-mediated silencing of genes is one epigenetic mechanism implicated in cancer. Studies regarding the role of modulation of gene expression utilizing inhibitors of DNA methylation, such as decitabine, in osteosarcoma (OS) have been limited. A biological understanding of the overall effects of decitabine in OS is important because this particular agent is currently undergoing clinical trials. The objective of this study was to measure the response of the OS cell line, U2OS, to decitabine treatment both in vitro and in vivo. Results: Microarray expression profiling was used to distinguish decitabine-dependent changes in gene expression in U2OS cells, and to identify responsive loci with demethylated CpG promoter regions. U2OS xenografts were established under the sub-renal capsule of immune-deficient mice to study the effect of decitabine in vivo on tumor growth and differentiation. Reduced nuclear methylation levels could be detected in xenografts derived from treated mice by immunohistochemistry utilizing a 5-methylcytidine antibody. Decitabine treatment reduced tumor xenograft size significantly (p < 0.05). Histological analysis of treated U2OS xenograft sections revealed a lower mitotic activity (p < 0.0001), increased bone matrix production (p < 0.0001), and a higher number of apoptotic cells (p = 0.0329). Microarray expression profiling of U2OS cultured cells showed that decitabine treatment caused a significant induction (p < 0.0025) in the expression of 88 genes. Thirteen had a >= 2-fold change, 11 of which had CpG-island-associated promoters. Interestingly, 6 of these 11 were pro-apoptotic genes and decitabine resulted in a significant induction of cell death in U2OS cells in vitro (p < 0.05). The 6 pro-apoptotic genes (GADD45A, HSPA9B, PAWR, PDCD5, NFKBIA, and TNFAIP3) were also induced to >= 2-fold in vivo. Quantitative methylation pyrosequencing confirmed that the tested pro-apoptotic genes had CpG-island DNA demethylationas a result of U2OS decitabine treatment both in vitro and in xenografts Conclusion: These data provide new insights regarding the use of epigenetic modifiers in OS, and have important implications for therapeutic trials involving demethylation drugs. Collectively, these data have provided biological evidence that one mode of action of decitabine may be the induction of apoptosis utilizing promoter-CpG demethylation of specific effectors in cell death pathways in OS.
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页数:16
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