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Effect of EBV LMP1 targeted DNAzymes on cell proliferation and apoptosis

Cancer Gene Therapy volume 12, pages 647–654 (2005)Cite this article

Abstract

The latent membrane protein (LMP1) encoded by Epstein–Barr virus (EBV) has been suggested to be one of the major oncogenic factors in EBV-mediated carcinogenesis. RNA-cleaving DNA enzymes are catalytic nucleic acids that bind and cleave a target RNA in a highly sequence-specific manner. In this study, we explore the potential of using DNAzymes as a therapeutic approach to EBV-associated carcinomas by targeting the LMP1 gene. In all, 13 different phosphorothioate-modified “10–23” deoxyribozymes (DNAzymes) were designed and synthesized against the LMP1 mRNA and transfected into B95-8 cells, which constitutively express the LMP1. Fluorescence microscopy was used to examine the cellular uptake and distribution in B95-8 cells. As demonstrated in Western blots, three out of 13 deoxyribozymes significantly downregulated the expression of LMP1 in B95-8 cells. These DNAzymes were shown to markedly inhibit B95-8 cell growth compared with a disabled DNAzyme and untreated controls, as determined by an alamarBlue Assay. It was further demonstrated that these DNAzymes arrested the B95-8 cells in G0/G1 using flow cytometry. Interestingly, the active DNAzymes could also downregulate the expression of Bcl-2 gene in treated cells, suggesting a close association between the LMP1 and Bcl-2 genes and their involvement in apoptosis. This was further confirmed with the result that the DNAzymes could induce the release of cytochrome c from mitochondria, which is the hallmark of the apoptosis. The present results suggest that the LMP1 may present a potential target for DNAzymes towards the EBV-associated carcinoma through cell proliferation and apoptosis pathways.

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Acknowledgements

This research was supported by the National Natural Science Foundation of China for Distinguished Young Scholars (No. 39525022), State Key Research Program, Fundamental Investigation on Human Carcinogenesis of China (No. G1998051201), and the National Natural Science Foundation of China (No. 30200009).

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Authors and Affiliations

  1. Cancer Research Institute, Xiangya School of Medicine, Central South University, Changsha, China

    Zhong-Xin Lu, Mao Ye, Guang-Rong Yan, Qun Li, Min Tang & Ya Cao

  2. Laboratory of Molecular Technology SAIC-Frederick, National Cancer Institute, Frederick, Maryland, USA

    Leo M Lee

  3. St Vincent's Clinical School, Faculty of Medicine, The University of New South Wales, Sydney, Australia

    Lun-Quan Sun

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  1. Zhong-Xin Lu
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Correspondence to Lun-Quan Sun or Ya Cao.

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Lu, ZX., Ye, M., Yan, GR. et al. Effect of EBV LMP1 targeted DNAzymes on cell proliferation and apoptosis. Cancer Gene Ther 12, 647–654 (2005). https://doi.org/10.1038/sj.cgt.7700833

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