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The cytotoxic activity of the phage E protein suppress the growth of murine B16 melanomas in vitro and in vivo

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Abstract

Novel treatment modalities, including gene therapy, are needed for patients with advanced melanoma. The E gene from the phage ϕX174 encodes a 91-aa protein which lyses Escherichia coli by formation of a transmembrane tunnel structure. To evaluate whether this E gene has a cytotoxic impact on melanoma cells in vitro and in vivo, and could therefore be used as a new therapeutic strategy for this tumor type, we selected the B16-F10 murine melanoma cell line as a model. We used a nonviral gene delivery approach (pcDNA3.1/E plasmid) to study the inhibition of melanoma cells' proliferation in vitro and direct intratumoral injection of pcDNA3.1/E complexed with jetPEI to deliver E cDNA to rapidly growing murine melanomas, and found that the E gene has both a strong antiproliferative effect in B16-F10 cells in vitro and induces an efficient decrease in melanoma tumor volume in vivo (90% in 15 days). Interestingly, the GFP-E fusion protein expressed in melanoma cells was located in the mitochondria. In vitro and in vivo analysis demonstrated significant functional and morphological mitochondrial alterations accompanied by a significant increase of cytochrome c and active caspase-3 and -9 in transfected cells, which suggests that tumoral cell death is mediated by the mitochondrial apoptotic pathway. These results show that E gene expression in melanoma cells has an extraordinary antitumor effect, which means it may be a new candidate for an effective strategy for melanoma treatment.

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Acknowledgments

We thank Dr. Ana B. Martínez-Cruz and Dr. G. Ortiz Ferrón for excellent technical assistance with the TUNEL assay and for providing the GFP gene. We are also thankful to Dr. F. O’Valle and Dr. A. Soler (Department of Anatomopathology and Physiology, Granada University) for their help and suggestions. This study was supported by the Fondo Investigaciones Sanitarias Seguridad Social (FIS; project no. PI041372) and by Granada University (project no. UGR-30 B364 1101/2007) and MEC (FPU predoctoral fellowship).

Conflict of interest statement

The authors declare that they have no competing financial interests.

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Authors and Affiliations

  1. Instituto de Biopatología y Medicina Regenerativa (IBIMER), Depto. de Anatomía y Embriología, Facultad de Medicina, Universidad de Granada, 18071, Granada, Spain

    Raúl Ortiz, Jose Prados, Consolacion Melguizo, Ana R. Rama, Fernando Rodríguez-Serrano, Houria Boulaiz & Antonia Aranega

  2. Department of Environmental Protection, CSIC-Estación Experimental del Zaidín, Granada, Spain

    Ana Segura & Juan L. Ramos

  3. Department of Health Sciences, University of Jaén, Jaén, Spain

    Fidel Hita & Antonio Martinez-Amat

  4. Department of Biomedical Science, University of Sassari, Sassari, Italy

    Roberto Madeddu

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  1. Raúl Ortiz
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  2. Jose Prados
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Correspondence to Jose Prados.

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Ortiz, R., Prados, J., Melguizo, C. et al. The cytotoxic activity of the phage E protein suppress the growth of murine B16 melanomas in vitro and in vivo. J Mol Med 87, 899–911 (2009). https://doi.org/10.1007/s00109-009-0493-9

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  • DOI: https://doi.org/10.1007/s00109-009-0493-9

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