Selective enhancement of hypoxic cell killing by tempol-regulated suicide gene expression

Kagiya,Go; Ogawa,Ryohei; Choudhuri,Rajani; Cook,John  A.; Hatashita,Masanori; Tanaka,Yoshikazu; Koda,Kana; Yamashita,Kei; Kubo,Makoto; Kawakami,Fumitaka; Mitchell,James  B.

doi:10.3892/or.2015.4020

Selective enhancement of hypoxic cell killing by tempol-regulated suicide gene expression

Authors:
- Go Kagiya
- Ryohei Ogawa
- Rajani Choudhuri
- John A. Cook
- Masanori Hatashita
- Yoshikazu Tanaka
- Kana Koda
- Kei Yamashita
- Makoto Kubo
- Fumitaka Kawakami
- James B. Mitchell
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Affiliations: School of Allied Health Sciences, Kitasato University, Minami-ku, Sagamihara, Kanagawa 228-8555, Japan, Department of Radiological Sciences, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama 930-0194, Japan, Radiation Biology Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA, Biology Group, The Wakasa Wan Energy Research Center, Tsuruga, Fukui 914-0192, Japan, Radiation Oncology, The Cancer Institute Hospital, Japanese Foundation for Cancer Research, Koto-ward, Tokyo 135-8550, Japan, Graduate School of Medical and Pharmaceutical Sciences, Chiba University, Inage-ku, Chiba 236-8555, Japan
Published online on: May 29, 2015 https://doi.org/10.3892/or.2015.4020
Pages: 1065-1073

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Abstract

The presence of hypoxic regions within solid tumors is caused by an imbalance between cell proliferation and angiogenesis. Such regions may facilitate the onset of recurrence after radiation therapy and chemotherapy, as hypoxic cells show resistance to these treatments. We found that tempol, a nitroxide, strongly induces the accumulation of hypoxia‑inducible factor (HIF)-1α, particularly under conditions of hypoxia. We, therefore, evaluated whether tempol enhances the gene expression via HIF-1α, potentially leading to various applications for cancer gene therapy targeting hypoxic cells. Consequently, following treatment with tempol under hypoxia, the luciferase (Luc) activity in the cells transfected with the plasmid containing the luc gene with the oxygen-dependent degradation domain and a promoter composed of hypoxia-responsive elements increased up to approximately 10-fold compared to that observed in cells treated identically with the exception of tempol. The plasmid constructed by replacing the luc gene with the fcy::fur fusion gene as a suicide gene, strongly induced the accumulation of the Fcy::Fur fusion protein, only when incubated in the presence of the hypoxic mimic CoCl2 and tempol. The transfected cells were successfully killed with the addition of 5-fluorocytosine to the cell culture according to the fcy::fur fusion gene expression. As similar but lesser enhancement of the Luc activity was also observed in solid tumor tissues in nude mice, this strategy may be applied for hypoxic cancer eradication.

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