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Silencing of H-ras gene expression by retrovirus-mediated siRNA decreases transformation efficiency and tumorgrowth in a model of human ovarian cancer

Oncogene volume 22, pages 5694–5701 (2003)Cite this article

Abstract

To examine the role of H-ras in the development of human ovarian cancer, we used small inhibitory RNA (siRNA) to silence its expression in human ovarian cancer cell lines and assessed the effects of its silencing on proliferation, apoptosis, and tumorgrowth. First, we developed a retrovirus-based delivery system that allowed long-term stable expression of the desired siRNA. Retrovirus-mediated expression of siRNA against green fluorescence protein (GFP) reduced its expression more than 90% in four cancer cell lines. We then constructed three retroviruses that expressed siRNAs targeting the H-rasV12 mutation (H1/siRNA) or either of two wild-type sequences of the H-ras gene (H2/siRNA and H3/siRNA) and used these retroviruses to infect T80H and SKOV-3 cells. In T80H cells (a genetically transformed human ovarian surface epithelial cell line whose tumorigenicity depends on H-rasV12 expression), infection with the H1/siRNA and H2/siRNA, but not with H3/siRNA, decreased T80H proliferation, increased G0/G1 arrest and apoptosis, blocked transformation in vitro, and suppressed tumor growth in nude mice. In SKOV-3 cells (a human ovarian cancer cell line that contains high levels of wild-type H-ras protein but no H-rasV12 mutation), introduction of the H2/siRNA construct, but not H1/siRNA or H3/siRNA, produced similar effects, demonstrating that the suppression of tumorgrowth by siRNA was sequence-specific. We conclude that H-ras is involved in maintenance of tumorgrowth of human ovarian cancer, and that retrovirus-mediated siRNA expression against H-ras expression is a powerful tool to dissect ras-signaling pathways and may be used therapeutically against ovarian cancer.

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Acknowledgements

This work was supported in part by institutional start-up funds, an institutional research grant, and a Career Development Award from The University of Texas MD. Anderson Cancer Center SPORE on Ovarian Cancer to JL. We thank Dr Yang Shi for kindly providing the pBS/U6 and pBS/U6/gfp/siRNA plasmids. We thank Christine Wogan for excellent editorial assistance.

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Authors and Affiliations

  1. Department of Pathology, The University of Texas MD Anderson Cancer Center, 1515 Holcombe Boulevard, Houston, 77030-4095, TX, USA

    Gong Yang, Jennifer Anne Thompson & Jinsong Liu

  2. Department of Thoracic and Cardiovascular Surgery, The University of Texas MD Anderson Cancer Center, 1515 Holcombe Boulevard, Houston, 77030-4095, TX, USA

    Bingliang Fang

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  1. Gong Yang
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Correspondence to Jinsong Liu.

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Yang, G., Thompson, J., Fang, B. et al. Silencing of H-ras gene expression by retrovirus-mediated siRNA decreases transformation efficiency and tumorgrowth in a model of human ovarian cancer. Oncogene 22, 5694–5701 (2003). https://doi.org/10.1038/sj.onc.1206858

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