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.
Citation
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.