Dataset Information

Oncolytic adenovirus mediated Survivin knockdown by RNA interference suppresses human colorectal carcinoma growth in vitro and in vivo.

ABSTRACT:

Background

Colorectal cancer is a one of the most common alimentary malignancies. Survivin has been proved by many studies to be an ideal target for cancer gene therapy because of its strong anti-apoptotic effect. The reduction of Survivin expression by means of chemically synthesized small interfering RNA or small hairpin RNA expressed from plasmid and resulted growth inhibition of cancer cells had been proved by many studies including ours, but the transfection efficiency was not encouraging. So for the first time we constructed the Survivin shRNA into an oncolytic adenovirus, tested its effects on colorectal cancer cell lines and nude mice xenograft model.

Methods

In this study, we constructed an oncolytic adenovirus with a Survivin targeted small hairpin RNA and a reporter gene (ZD55-Sur-EGFP). The expression of Survivin mRNA and protein were analyzed by RT-PCR and western blot. The cell growth and apoptosis were tested by in vitro cytopathic assay, MTT assay and flow cytometry respectively. The effect of the constructed virus on xenograft model was evaluated by tumor volume and western blot analysis.

Results

ZD55-Sur-EGFP replicated in cancer cells specifically, reduced the expression of Survivin mRNA and protein expression effectively (P < 0.0001), induced cancer cell apoptosis and inhibited SW480 cell growth both in vitro and in vivo significantly.

Conclusion

We conclude Survivin RNA interference combining with oncolytic adenovirus virotherapy to be a promising treatment for colorectal cancer.

SUBMITTER: Shen W

PROVIDER: S-EPMC2703625 | biostudies-literature | 2009 Jun

REPOSITORIES: biostudies-literature

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Publications

Oncolytic adenovirus mediated Survivin knockdown by RNA interference suppresses human colorectal carcinoma growth in vitro and in vivo.

Shen Wei W Wang Chun-Yi CY Wang Xue-Hu XH Fu Zhong-Xue ZX

Journal of experimental & clinical cancer research : CR 20090615

<h4>Background</h4>Colorectal cancer is a one of the most common alimentary malignancies. Survivin has been proved by many studies to be an ideal target for cancer gene therapy because of its strong anti-apoptotic effect. The reduction of Survivin expression by means of chemically synthesized small interfering RNA or small hairpin RNA expressed from plasmid and resulted growth inhibition of cancer cells had been proved by many studies including ours, but the transfection efficiency was not encou ...[more]

PMID: 19527508

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Project description:BackgroundSurvivin and octamer-binding transcription factor 4 (OCT4) are reportedly up-regulated in esophageal cancer (EC) and have been correlated with high tumor proliferative activity and poor prognosis. Oncolytic viruses encoding specific transgenes have been considered as therapeutic methods to increase therapeutic efficacy in a variety of solid tumors.MethodsIn this study, an oncolytic adenovirus carrying short hairpin RNA (shRNA) of survivin (shSRVN) and OCT4 (shOCT4) was constructed to achieve dual knockdown of survivin and OCT4 and to explore the potential effect of the oncolytic adenovirus in EC.ResultsThe oncolytic adenovirus replicated abundantly in human EC cells, with the replication multiplying by up to 192,085 and 620,055 times in esophageal carcinoma (Eca)-109 cells transfected with purified and completed recombinant adenoviruses called AdSProE1a-dual shRNA (shSRVN + shOCT4) and TE1 cells transfected with AdSProE1a-survivin shRNA (shSRVN) 96 hours after infection, respectively. The shRNAs targeting survivin and OCT4 significantly downregulated the expression levels of survivin and OCT4 in cells, thereby inhibiting the proliferative activity of cancer cells. Furthermore, E-cadherin and vimentin, which are both considered epithelial mesenchymal transition (EMT) markers, were found to be upregulated and downregulated, respectively, in cancer cells after exposure to the viral infection. The interference of survivin and OCT4 also contributed to cell cycle arrest and apoptosis, the half maximal inhibitory concentrations (IC50s) of oncolytic adenovirus loaded with AdSProE1a-shSRVN + shOCT4 in the Eca109 cells and the TE1 cells were 0.7271 and 0.1032 pfu/mL, respectively. Xenograft experiments in vivo showed that oncolytic adenovirus-mediated dual knockdown of survivin and OCT4 effectively inhibited the growth of xenografts and induced cancer cell apoptosis. We concluded that therapies targeting survivin and OCT4 have great potential for improving the therapeutic efficacy in EC.ConclusionsThe dual target design strategy ensured the efficacy and safety of the treatment system and provided a novel and effective adjuvant target therapy for EC.

Project description:IntroductionIn human glioblastomas, glioma pathogenesis-related protein1 (GliPR1) is overexpressed and appears to be an oncoprotein. We investigated whether GliPR1 knockdown in glioma cells by RNA interference exerts anti-glioma effects.MethodsExperiments used human glioblastoma cell lines transduced with GliPR1 shRNA (sh#301, sh#258). Transduction produced stringent doxycycline-dependent GliPR1 knockdown in clones (via lentiviral "all-in-one" TetOn-shRNA vector) or stable GliPR1 knockdown in polyclonal cells (via constitutive retroviral-shRNA vector). In vitro assessments included cellular proliferation and clonogenic survival. In vivo assessments in tumor-bearing nude mice included tumor growth and survival.ResultsUsing doxycycline-dependent GliPR1 knockdown, shGliPR1-transduced U87-MG clones demonstrated reductions in cellular proliferation in the presence versus absence of doxycycline. Using stable GliPR1 knockdown, polyclonal shGliPR1-transduced U87-MG, A172, and U343-MG cells consistently showed decreased clonogenic survival and induced apoptosis (higher proportion of early apoptotic cells) compared to control shLuc-transduced cells. In tumor-bearing nude mice, using doxycycline-dependent GliPR1 knockdown, subcutaneous and cranial transplantation of the U87-MG clone 980-5 (transduced with GliPR1 sh#301) resulted in reduced subcutaneous tumor volume and cerebral tumor area in doxycycline-treated mice versus those left untreated. Using stable GliPR1 knockdown, nude mice cranially transplanted with polyclonal U87-MG cells transduced with GliPR1 sh#258 had significantly prolonged survival compared to mice cranially transplanted with control shLuc-transduced cells (41 versus 26 days; P < 0.001).ConclusionGliPR1 knockdown in glioma cells decreased cellular proliferation, decreased clonogenic survival, and induced apoptosis in vitro, and reduced glioblastoma tumor growth and prolonged survival in vivo. These findings support that GliPR1 may have potential value as a therapeutic target.

Project description:Gene therapy has become an important strategy for treatment of malignancies, but problems remains concerning the low gene transferring efficiency, poor transgene expression and limited targeting specific tumors, which have greatly hampered the clinical application of tumor gene therapy. Gallbladder cancer is characterized by rapid progress, poor prognosis, and aberrantly high expression of Survivin. In the present study, we used a human tumor-specific Survivin promoter-regulated oncolytic adenovirus vector carrying P53 gene, whose anti-cancer effect has been widely confirmed, to construct a wide spectrum, specific, safe, effective gene-viral therapy system, AdSurp-P53. Examining expression of enhanced green fluorecent protein (EGFP), E1A and the target gene P53 in the oncolytic adenovirus system validated that Survivin promoter-regulated oncolytic adenovirus had high proliferation activity and high P53 expression in Survivin-positive gallbladder cancer cells. Our in vitro cytotoxicity experiment demonstrated that AdSurp-P53 possessed a stronger cytotoxic effect against gallbladder cancer cells and hepatic cancer cells. The survival rate of EH-GB1 cells was lower than 40% after infection of AdSurp-P53 at multiplicity of infection (MOI) = 1 pfu/cell, while the rate was higher than 90% after infection of Ad-P53 at the same MOI, demonstrating that AdSurp-P53 has a potent cytotoxicity against EH-GB1 cells. The tumor growth was greatly inhibited in nude mice bearing EH-GB1 xenografts when the total dose of AdSurp-P53 was 1 × 10(9) pfu, and terminal dUTP nick end-labeling (TUNEL) revealed that the apoptotic rate of cancer cells was (33.4 ± 8.4)%. This oncolytic adenovirus system overcomes the long-standing shortcomings of gene therapy: poor transgene expression and targeting of only specific tumors, with its therapeutic effect better than the traditional Ad-P53 therapy regimen already on market; our system might be used for patients with advanced gallbladder cancer and other cancers, who are not sensitive to chemotherapy, radiotherapy, or who lost their chance for surgical treatment.

Dataset Information

Oncolytic adenovirus mediated Survivin knockdown by RNA interference suppresses human colorectal carcinoma growth in vitro and in vivo.

Background

Methods

Results

Conclusion

Publications

Oncolytic adenovirus mediated Survivin knockdown by RNA interference suppresses human colorectal carcinoma growth in vitro and in vivo.

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