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The synergistic effect of dimethylamino benzoylphenylurea (NSC #639829) and X-irradiation on human lung carcinoma cell lines

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Abstract

Purpose

The present study was designed to investigate the ability of N-[4-(5-bromo-2-pyrimidyloxy)-3-methylphenyl]-(dimemethylamino)-benzoylphenylurea (dimemethylamino benzoylphenylurea; BPU) to sensitize cells to radiation and to examine the relationship between phenotype versus survival, DNA damage, apoptosis, or cell cycle progression in non-small cell lung cancer (NSCLC) cell lines.

Methods

Asynchronous cultures of three NSCLC (phenotype) lines, A549 (adenocarcinoma), NCI-H226 (squamous) and NCI-H596 (adenosquamous) were used. Cells were treated for 24 h with BPU at various concentrations (0–10 μM) to obtain drug doses for inhibiting cell survival by ∼50% (IC50). Cells were X-irradiated without BPU or after 24 h BPU treatment at IC50. Radiation doses ranged from 0 to10 Gy. Cell survival was determined by a colony-forming ability assay. The effect of BPU on the cell cycle distribution and induction of apoptosis were measured by flow cytometry-based assays. The effect of BPU on radiation-induced DNA damage and repair was analyzed according to nuclear γH2AX immunofluorescence of cells exposed to X-rays alone or after BPU. Anti-γH2AX antibody staining, a surrogate determinant of double stranded DNA breaks, was measured using flow cytometry.

Results

BPU (1.5 μM) for 24 h produced ∼50% cell survival. BPU and X-irradiation were synergistic in the three cell lines at survival levels of 20–50%. Flow cytometry analysis of replicate experiments with BPU (1.5 μM for 24 h) showed that BPU blocked cell progression at S and/or G2/M. The incidence of apoptosis in BPU-treated versus control cells ranged from ∼0.3 to ∼8%. Twenty-four hour after X-irradiation cells pre-treated with BPU and X-irradiated after drug exposure showed γH2AX levels approximately two times higher than did the cells exposed to X-rays only.

Conclusions

The study identified BPU as a novel radiation sensitizer. The analysis of phosphorylated histone H2AX as a surrogate marker of DNA double strand breaks suggested a positive association between radiosensitization and the inhibition of X-irradiation-induced DNA damage repair by BPU.

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Acknowledgments

The authors thank Angel Wood for technical assistance with cell culture and Wilfried Goetz for help with the γH2AX expression assay. Supported in part by the University of Maryland School of Medicine Research Initiative Grant.

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Authors and Affiliations

  1. Department of Radiation Oncology, Radiation Oncology Research Laboratory, Universtity of Maryland School of Medicine, 655 W. Baltimore Street, BRB 6-009, Baltimore, MD, 21201, USA

    Elizabeth K. Balcer-Kubiczek & Mona Attarpour

  2. University of Maryland Marlene and Stewart Greenebaum Cancer Center, Baltimore, MD, USA

    Elizabeth K. Balcer-Kubiczek & Martin J. Edelman

  3. Department of Medicine, University of Maryland School of Medicine, Baltimore, MD, USA

    Martin J. Edelman

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  1. Elizabeth K. Balcer-Kubiczek
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  2. Mona Attarpour
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  3. Martin J. Edelman
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Correspondence to Elizabeth K. Balcer-Kubiczek.

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Balcer-Kubiczek, E.K., Attarpour, M. & Edelman, M.J. The synergistic effect of dimethylamino benzoylphenylurea (NSC #639829) and X-irradiation on human lung carcinoma cell lines. Cancer Chemother Pharmacol 59, 781–787 (2007). https://doi.org/10.1007/s00280-006-0333-3

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  • DOI: https://doi.org/10.1007/s00280-006-0333-3

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