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Targeting head and neck squamous cell carcinoma using a novel fusion toxin-diphtheria toxin/HN-1
Published in
2011
PMID: 20814829
Volume: 38
   
Issue: 2
Pages: 1389 - 1397
Abstract
The current treatment strategies, chemotherapy and radiation therapy being used for the management of cancer are deficient in targeted approach leading to treatment related toxicities and relapse. Contrarily, fusion toxins exhibit remarkable tumor specificity thus emerging as an alternative therapy for the treatment of cancer. Diphtheria toxin-HN-1 peptide (DT/HN-1) is a fusion toxin designed to target the head and neck squamous cell carcinoma (HNSCC). The aim of this study was to construct, characterize, and evaluate the cytotoxicity and specificity of DT/HN-1 fusion toxin against the HNSCC cells. The purified DT/HN-1 fusion toxin was characterized by SDS-PAGE and western blotting. Refolding of purified fusion toxins was monitored by fluorescence spectra and circular dichroism spectra. The activity of DT/HN-1 fusion toxin was demonstrated on various HNSCC cell lines by cell viability assay, cell proliferation assay, protein synthesis inhibition assay, apoptosis and cell cycle analysis. The fusion toxin DT/HN-1 demonstrated remarkably high degree of cytotoxicity specific to the HNSCC cells. The IC50 of DT/HN-1 fusion toxin was ~1 to 5 nM in all the three HNSCC cell lines. The percentage apoptotic cells in DT/HN-1 treated UMB-SCC-745 cells are 16% compared to 4% in untreated. To further demonstrate the specific toxicity of DT/HN-1 fusion toxin towards the HNSCC cells we constructed, characterized and evaluated the efficacy of DT protein. The DT protein coding for only a fragment of diphtheria toxin without its native receptor binding domain failed to exhibit any cytotoxicity on all the cell lines used in this study thus establishing the importance of a ligand in achieving targeted toxicity. To evaluate the translocation ability of HN-1 peptide, an additional construct DTΔT/HN-1 was constructed, characterized and evaluated for its cytotoxic activity. The fusion toxin DTΔT/HN-1 deficient of the translocation domain of diphtheria toxin showed no cytotoxicity on all the cell lines clearly indicating the inability of HN-1 peptide to translocate catalytic domain of the toxin into the cytosol. © 2010 Springer Science+Business Media B.V.
About the journal
JournalMolecular Biology Reports
ISSN03014851
Open AccessNo
Concepts (41)
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    CYTOTOXIC FACTOR
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    Diphtheria toxin
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    PROTEIN HN 1
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    Unclassified drug
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    Apoptosis
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    Article
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    Cancer cell culture
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    Cancer inhibition
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    CELL CYCLE REGULATION
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    Cell proliferation
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    Cell viability
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    Circular dichroism
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    Controlled study
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    Cytosol
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    Drug cytotoxicity
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    Drug efficacy
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    Drug receptor binding
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    DRUG SPECIFICITY
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    Drug targeting
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    Fluorescence spectroscopy
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    HEAD AND NECK CARCINOMA
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    Human
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    Human cell
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    IC 50
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    Polyacrylamide gel electrophoresis
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    Protein synthesis inhibition
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    Squamous cell carcinoma
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    Western blotting
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    Antineoplastic agents
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    CARCINOMA, SQUAMOUS CELL
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    Cell line, tumor
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    Cell survival
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    Cloning, molecular
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    Dose-response relationship, drug
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    HEAD AND NECK NEOPLASMS
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    Humans
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    Inhibitory concentration 50
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    Ligands
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    Mutagenesis, site-directed
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    Oligopeptides
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    Plasmids