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Restoration of tumor suppressor miR-34 inhibits human p53-mutant gastric cancer tumorspheres
Ji, Qing Hao, Xinbao Meng, Yang Zhang, Min DeSano, Jeffrey Fan, Daiming Xu, Liang
Ji, Qing
Hao, Xinbao
Meng, Yang
Zhang, Min
DeSano, Jeffrey
Fan, Daiming
Xu, Liang
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Abstract
Background: MicroRNAs (miRNAs), some of which function as oncogenes or tumor suppressor genes,
are involved in carcinogenesis via regulating cell proliferation and/or cell death. MicroRNA miR-34 was
recently found to be a direct target of p53, functioning downstream of the p53 pathway as a tumor
suppressor. miR-34 targets Notch, HMGA2, and Bcl-2, genes involved in the self-renewal and survival of
cancer stem cells. The role of miR-34 in gastric cancer has not been reported previously. In this study, we
examined the effects of miR-34 restoration on p53-mutant human gastric cancer cells and potential target
gene expression.
Methods: Human gastric cancer cells were transfected with miR-34 mimics or infected with the lentiviral
miR-34-MIF expression system, and validated by miR-34 reporter assay using Bcl-2 3'UTR reporter.
Potential target gene expression was assessed by Western blot for proteins, and by quantitative real-time
RT-PCR for mRNAs. The effects of miR-34 restoration were assessed by cell growth assay, cell cycle
analysis, caspase-3 activation, and cytotoxicity assay, as well as by tumorsphere formation and growth.
Results: Human gastric cancer Kato III cells with miR-34 restoration reduced the expression of target
genes Bcl-2, Notch, and HMGA2. Bcl-2 3'UTR reporter assay showed that the transfected miR-34s were
functional and confirmed that Bcl-2 is a direct target of miR-34. Restoration of miR-34 chemosensitized
Kato III cells with a high level of Bcl-2, but not MKN-45 cells with a low level of Bcl-2. miR-34 impaired
cell growth, accumulated the cells in G1 phase, increased caspase-3 activation, and, more significantly,
inhibited tumorsphere formation and growth.
Conclusion: Our results demonstrate that in p53-deficient human gastric cancer cells, restoration of
functional miR-34 inhibits cell growth and induces chemosensitization and apoptosis, indicating that miR-
34 may restore p53 function. Restoration of miR-34 inhibits tumorsphere formation and growth, which is
reported to be correlated to the self-renewal of cancer stem cells. The mechanism of miR-34-mediated
suppression of self-renewal appears to be related to the direct modulation of downstream targets Bcl-2,
Notch, and HMGA2, indicating that miR-34 may be involved in gastric cancer stem cell self-renewal/
differentiation decision-making. Our study suggests that restoration of the tumor suppressor miR-34 may
provide a novel molecular therapy for p53-mutant gastric cancer.
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2008-09-21
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BioMed Central
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Ji, Q., Hao, X., Meng, Y., Zhang, M., DeSano, J., Fan, D., Xu, L. Restoration of tumor suppressor miR-34 inhibits human p53-mutant gastric cancer tumorspheres. BMC Cancer 2008, 8:266. http://dx.doi.org/10.1186/1471-2407-8-266.