Human osteocalcin: a strong promoter for nitric oxide synthase gene therapy, with specificity for hormone refractory prostate cancer

Helen O. McCarthy, Jonathan A. Coulter, Jenny Worthington, Tracy Robson, David G. Hirst

    Research output: Contribution to journalArticle

    18 Citations (Scopus)

    Abstract

    Background Gene therapy has been identified as a promising treatment strategy for hormone refractory prostate cancer (HRPC). We report, for the first time, the use of the human osteocalcin (hOC) promoter to control inducible nitric oxide synthase (iNOS) transgene expression in HRPC. Methods Human prostate carcinoma cells (PC3, DU145, LNCaP), colon cancer cells (HT29) and human microvascular endothelial cells (HMEC-1) were transfected in vitro with constitutively driven CMV/iNOS or hOC/iNOS plasmid DNA by cationic lipid vector. End points of these experiments were Western blotting, NO generation using the Greiss test to measure accumulated nitrite, and clonogenic assay. Results Transfection of the hOC/iNOS plasmid increased iNOS protein and total nitrite levels in PC3 and DU145 cells, but not LNCaP or HT29. Transfection with CMV/iNOS or hOC/iNOS resulted in no additional cytotoxicity in androgen-dependent LNCaP cells or in the non-prostate cell lines. However, transfection with either construct resulted in a greatly reduced cell survival (to 10-20%) in the androgen-independent PC3 and DU145 cell lines. Conclusions Utilising the tumour-type specific properties of the hOC promoter in tandem with the iNOS gene, we have demonstrated target cell specificity, and transgene activation, in the androgen-independent prostate cancer cell lines (PC3 and DU145), an effect absent in normal and androgen-dependent cells. Furthermore, the levels of NO generated are comparable with those seen generated with constitutively (CMV)-driven iNOS. The data obtained from this study provide a basis for future development of hOC/iNOS gene therapy. Copyright (C) 2007 John Wiley & Sons, Ltd.
    LanguageEnglish
    Pages511-520
    JournalJournal of Gene Medicine
    Volume9
    Issue number6
    DOIs
    Publication statusPublished - Jun 2007

    Fingerprint

    Osteocalcin
    Nitric Oxide Synthase Type II
    Nitric Oxide Synthase
    Genetic Therapy
    Prostatic Neoplasms
    Hormones
    Androgens
    Transfection
    Nitrites
    Transgenes
    Cell Line
    Plasmids
    HT29 Cells
    Human Development
    Colonic Neoplasms
    Prostate
    Cell Survival
    Endothelial Cells
    Western Blotting
    Carcinoma

    Cite this

    McCarthy, Helen O. ; Coulter, Jonathan A. ; Worthington, Jenny ; Robson, Tracy ; Hirst, David G. / Human osteocalcin: a strong promoter for nitric oxide synthase gene therapy, with specificity for hormone refractory prostate cancer. In: Journal of Gene Medicine. 2007 ; Vol. 9, No. 6. pp. 511-520.
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    title = "Human osteocalcin: a strong promoter for nitric oxide synthase gene therapy, with specificity for hormone refractory prostate cancer",
    abstract = "Background Gene therapy has been identified as a promising treatment strategy for hormone refractory prostate cancer (HRPC). We report, for the first time, the use of the human osteocalcin (hOC) promoter to control inducible nitric oxide synthase (iNOS) transgene expression in HRPC. Methods Human prostate carcinoma cells (PC3, DU145, LNCaP), colon cancer cells (HT29) and human microvascular endothelial cells (HMEC-1) were transfected in vitro with constitutively driven CMV/iNOS or hOC/iNOS plasmid DNA by cationic lipid vector. End points of these experiments were Western blotting, NO generation using the Greiss test to measure accumulated nitrite, and clonogenic assay. Results Transfection of the hOC/iNOS plasmid increased iNOS protein and total nitrite levels in PC3 and DU145 cells, but not LNCaP or HT29. Transfection with CMV/iNOS or hOC/iNOS resulted in no additional cytotoxicity in androgen-dependent LNCaP cells or in the non-prostate cell lines. However, transfection with either construct resulted in a greatly reduced cell survival (to 10-20{\%}) in the androgen-independent PC3 and DU145 cell lines. Conclusions Utilising the tumour-type specific properties of the hOC promoter in tandem with the iNOS gene, we have demonstrated target cell specificity, and transgene activation, in the androgen-independent prostate cancer cell lines (PC3 and DU145), an effect absent in normal and androgen-dependent cells. Furthermore, the levels of NO generated are comparable with those seen generated with constitutively (CMV)-driven iNOS. The data obtained from this study provide a basis for future development of hOC/iNOS gene therapy. Copyright (C) 2007 John Wiley & Sons, Ltd.",
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    Human osteocalcin: a strong promoter for nitric oxide synthase gene therapy, with specificity for hormone refractory prostate cancer. / McCarthy, Helen O.; Coulter, Jonathan A.; Worthington, Jenny; Robson, Tracy; Hirst, David G.

    In: Journal of Gene Medicine, Vol. 9, No. 6, 06.2007, p. 511-520.

    Research output: Contribution to journalArticle

    TY - JOUR

    T1 - Human osteocalcin: a strong promoter for nitric oxide synthase gene therapy, with specificity for hormone refractory prostate cancer

    AU - McCarthy, Helen O.

    AU - Coulter, Jonathan A.

    AU - Worthington, Jenny

    AU - Robson, Tracy

    AU - Hirst, David G.

    PY - 2007/6

    Y1 - 2007/6

    N2 - Background Gene therapy has been identified as a promising treatment strategy for hormone refractory prostate cancer (HRPC). We report, for the first time, the use of the human osteocalcin (hOC) promoter to control inducible nitric oxide synthase (iNOS) transgene expression in HRPC. Methods Human prostate carcinoma cells (PC3, DU145, LNCaP), colon cancer cells (HT29) and human microvascular endothelial cells (HMEC-1) were transfected in vitro with constitutively driven CMV/iNOS or hOC/iNOS plasmid DNA by cationic lipid vector. End points of these experiments were Western blotting, NO generation using the Greiss test to measure accumulated nitrite, and clonogenic assay. Results Transfection of the hOC/iNOS plasmid increased iNOS protein and total nitrite levels in PC3 and DU145 cells, but not LNCaP or HT29. Transfection with CMV/iNOS or hOC/iNOS resulted in no additional cytotoxicity in androgen-dependent LNCaP cells or in the non-prostate cell lines. However, transfection with either construct resulted in a greatly reduced cell survival (to 10-20%) in the androgen-independent PC3 and DU145 cell lines. Conclusions Utilising the tumour-type specific properties of the hOC promoter in tandem with the iNOS gene, we have demonstrated target cell specificity, and transgene activation, in the androgen-independent prostate cancer cell lines (PC3 and DU145), an effect absent in normal and androgen-dependent cells. Furthermore, the levels of NO generated are comparable with those seen generated with constitutively (CMV)-driven iNOS. The data obtained from this study provide a basis for future development of hOC/iNOS gene therapy. Copyright (C) 2007 John Wiley & Sons, Ltd.

    AB - Background Gene therapy has been identified as a promising treatment strategy for hormone refractory prostate cancer (HRPC). We report, for the first time, the use of the human osteocalcin (hOC) promoter to control inducible nitric oxide synthase (iNOS) transgene expression in HRPC. Methods Human prostate carcinoma cells (PC3, DU145, LNCaP), colon cancer cells (HT29) and human microvascular endothelial cells (HMEC-1) were transfected in vitro with constitutively driven CMV/iNOS or hOC/iNOS plasmid DNA by cationic lipid vector. End points of these experiments were Western blotting, NO generation using the Greiss test to measure accumulated nitrite, and clonogenic assay. Results Transfection of the hOC/iNOS plasmid increased iNOS protein and total nitrite levels in PC3 and DU145 cells, but not LNCaP or HT29. Transfection with CMV/iNOS or hOC/iNOS resulted in no additional cytotoxicity in androgen-dependent LNCaP cells or in the non-prostate cell lines. However, transfection with either construct resulted in a greatly reduced cell survival (to 10-20%) in the androgen-independent PC3 and DU145 cell lines. Conclusions Utilising the tumour-type specific properties of the hOC promoter in tandem with the iNOS gene, we have demonstrated target cell specificity, and transgene activation, in the androgen-independent prostate cancer cell lines (PC3 and DU145), an effect absent in normal and androgen-dependent cells. Furthermore, the levels of NO generated are comparable with those seen generated with constitutively (CMV)-driven iNOS. The data obtained from this study provide a basis for future development of hOC/iNOS gene therapy. Copyright (C) 2007 John Wiley & Sons, Ltd.

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    SP - 511

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    JO - Journal of Gene Medicine

    T2 - Journal of Gene Medicine

    JF - Journal of Gene Medicine

    SN - 1099-498X

    IS - 6

    ER -