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7B7: a novel antibody directed against the Ku70/Ku80 heterodimer blocks invasion in pancreatic and lung cancer cells

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Tumor Biology

Abstract

Development of more effective therapeutic strategies for cancers of high unmet need requires the continued discovery of disease-specific protein targets for therapeutic antibody targeting. In order to identify novel proteins associated with cancer cell invasion/metastasis, we present here an alternative to antibody targeting of cell surface proteins with an established role in invasion; our functional antibody screening approach involves the isolation and selection of MAbs that are primarily screened for their ability to inhibit tumour invasion. A clonal population of the Mia PaCa-2, a pancreatic ductal adenocarcinoma (PDAC) cell line, which displays a highly invasive phenotype, was used to generate MAbs with the objective of identifying membrane targets directly involved in cancer invasion. Selected MAb 7B7 can significantly reduce invasion in a dose-responsive manner in Mia PaCa-2 clone 3 and DLKP-M squamous lung carcinoma cells. Using immunoprecipitation and liquid chromatography-tandem mass spectrometry (LC-MS-MS) analysis, the target antigen of anti-invasive antibody, 7B7, was determined to be the heterodimeric Ku antigen, Ku70/80, a core protein composed of the Ku70 and Ku80 subunits which is involved in non-homologous end-joining (NHEJ) DNA repair. RNA interference-mediated knockdown of Ku70 and Ku80 resulted in a marked decrease in the invasive capacity of Mia PaCa-2 clone 3 and DLKP-M cells, indicating that Ku70/Ku80 is functionally involved in pancreatic and lung cancer invasion. Immunohistochemical analysis demonstrated Ku70/Ku80 immunoreactivity in 37 PDAC tumours, indicating that this heterodimer is highly expressed in this aggressive cancer type. This study demonstrates that a functional MAb screening approach coupled with immunoprecipitation/proteomic analyses can be successfully applied to identify functional anti-invasive MAbs and potential novel targets for therapeutic antibody targeting.

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Acknowledgments

This work was funded by Enterprise Ireland, project code TD/2009/0133. The authors wish to acknowledge the technical assistance provided by the RCSI Biomedical Facility, Beaumont Hospital, Dublin 9, Ireland.

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Author notes

  1. Dermot O’Sullivan

    Present address: Biomedical Diagnostics Institute, Dublin City University, Dublin 9, Ireland

Authors and Affiliations

  1. National Institute for Cellular Biotechnology (NICB), Dublin City University, Glasnevin, Dublin 9, Ireland

    Dermot O’Sullivan, Michael Henry, Helena Joyce, Naomi Walsh, Edel Mc Auley, Paul Dowling, Michael Moriarty, Martin Clynes & Annemarie Larkin

  2. Molecular Therapeutics Cancer Ireland (MTCI), National Institute for Cellular Biotechnology (NICB), Dublin City University, Glasnevin, Dublin 9, Ireland

    Helena Joyce

  3. National Surgical Centre for Pancreatic Cancer, St Vincent’s University Hospital, Elm Park, Dublin, Ireland

    Niall Swan

  4. Dublin City University, Biounite, Room JGO7, R & D Building, Glasnevin, Dublin 9, Ireland

    Paul Barnham

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  1. Dermot O’Sullivan
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  2. Michael Henry
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Correspondence to Annemarie Larkin.

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Dermot O’Sullivan and Michael Henry contributed equally to this study.

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O’Sullivan, D., Henry, M., Joyce, H. et al. 7B7: a novel antibody directed against the Ku70/Ku80 heterodimer blocks invasion in pancreatic and lung cancer cells. Tumor Biol. 35, 6983–6997 (2014). https://doi.org/10.1007/s13277-014-1857-5

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  • DOI: https://doi.org/10.1007/s13277-014-1857-5

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