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Key role for p27Kip1, retinoblastoma protein Rb, and MYCN in polyamine inhibitor-induced G1 cell cycle arrest in MYCN-amplified human neuroblastoma cells

Oncogene volume 24, pages 5606–5618 (2005)Cite this article

Abstract

Alpha-difluoromethylornithine (DFMO) inhibits the proto-oncogene ornithine decarboxylase (ODC) and is known to induce cell cycle arrest. However, the effect of DFMO on human neuroblastoma (NB) cells and the exact mechanism of DFMO-induced cell death are largely unknown. Treatment with DFMO in combination with SAM486A, an S-adenosylmethionine decarboxylase (AdoMetDC) inhibitor, has been shown to enhance polyamine pool depletion. Therefore, we analysed the mechanism of action of DFMO and/or SAM486A in two established MYCN-amplified human NB cell lines. DFMO and SAM486A caused rapid cell growth inhibition, polyamine depletion, and G1 cell cycle arrest without apoptosis in cell lines LAN-1 and NMB-7. These effects were enhanced with combined inhibitors and largely prevented by cotreatment with exogenous polyamines. The G1 cell cycle arrest was concomitant with an increase in cyclin-dependent kinase inhibitor p27Kip1. In a similar fashion, DFMO and DFMO/SAM486A inhibited the phosphorylation of the G1/S transition-regulating retinoblastoma protein Rb at residues Ser795 and Ser807/811. Moreover, we observed a dramatic decrease in MYCN protein levels. Overexpression of MYCN induces an aggressive NB phenotype with malignant behavior. We show for the first time that DFMO and SAM486A induce G1 cell cycle arrest in NB cells through p27Kip1 and Rb hypophosphorylation.

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Acknowledgements

We are grateful to Dr Randal Wada (Cancer Research Center of Hawaii) for providing us with the MYCN antibody in initial studies, and to Drs Carl-Wilhelm Vogel, Bonnie Warn-Cramer, Darren Park, Patricia Lorenzo, and Matt Tuthill (Cancer Research Center of Hawaii) for their support, advice, and stimulating discussions during the course of this work. Crystal Fo and Craig Coleman are thanked for their excellent technical support of this project. We give special thanks to Dr Patrick Woster (Wayne State University, MI, USA) for providing the ODC inhibitor DFMO. We also thank Novartis (Basel, Switzerland) for providing the AdoMetDC inhibitor SAM486A. This work was supported by the Hawaii Community Foundation, grants #20022113 and #20041684 to AS Bachmann. KY Takasaki received support from the Cell and Molecular Biology Graduate Program of the University of Hawaii at Manoa. SM Wilson was supported by the NIH grant R01CA79909 to CV Byus. Additional funds were provided by the NIH grant R01CA18138 to AE Pegg.

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Author notes

  1. Ivonne Gamper

    Present address: Institute for Biomedical Technology, Universitätsklinikum Aachen, RWTH, 52057, Aachen, Germany

  2. Mike Thorne

    Present address: Hawaii Biotech, Inc., Aiea, HI, 96701, USA

  3. Shannon M Wilson

    Present address: University of California, San Francisco, Lung Biology Center, San Francisco, CA, 94110, USA

Authors and Affiliations

  1. Cancer Research Center of Hawaii, University of Hawaii at Manoa, 1236 Lauhala Street, Honolulu, 96813, HI, USA

    Christopher J Wallick, Ivonne Gamper, Mike Thorne, Kelsie Y Takasaki, Jennifer A Seki & André S Bachmann

  2. Department of Cellular and Molecular Physiology, Pennsylvania State University College of Medicine, Hershey, 17033, PA, USA

    David J Feith & Anthony E Pegg

  3. Department of Biochemistry, University of California, Riverside, Riverside, 92521, CA, USA

    Shannon M Wilson & Craig V Byus

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Wallick, C., Gamper, I., Thorne, M. et al. Key role for p27Kip1, retinoblastoma protein Rb, and MYCN in polyamine inhibitor-induced G1 cell cycle arrest in MYCN-amplified human neuroblastoma cells. Oncogene 24, 5606–5618 (2005). https://doi.org/10.1038/sj.onc.1208808

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