Journal: Cancer Letters

Article Title: TP53 mutations in head and neck cancer cells determine the Warburg phenotypic switch creating metabolic vulnerabilities and therapeutic opportunities for stratified therapies

doi: 10.1016/j.canlet.2020.02.032

Figure Lengend Snippet: The radiosensitisation of head and neck cancer cells with compromised p53 function correlates with radiation-induced apoptosis . A. Flow cytometry analysis of representative p53 wild-type (UM-SCC-17as) and mutant (UM-SCC-5) cell lines treated as indicated with either no treatment, staurosporine as a positive control for detection of apoptosis induction, ionising radiation (IR), 2-DG or a combination of IR plus 2-DG harvested 24 or 48 h later as indicated. In all plots the abscissa indicates Annexin V detection and the ordinate is propidium iodide detection. Percentages represent Annexin V positivity. B. Summarises the results for similar experiments to those described in A. performed on the indicated cell lines. As in , blue bars represent data from cell lines with wild-type p53, red bars represent data from cell lines that are either p53 null or harbouring mutant p53, orange bars represent data from a p53 null cell line manipulated to express wild-type p53 and green bars derive from a p53 knock-down wild-type cell line. In all cell lines with abrogation of wtp53 function (either through mutation or knock-down) addition of 2-DG to IR significantly increased the proportion of annexin V positive cells at either 24 or 48 h (* P < 0.01 in all cases), while in cell lines harbouring functional wtp53 no significant increase was observed. Results presented are typical results from an independent experiment that has been repeated on at least three occasions. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Article Snippet: HRP-conjugated secondary antibodies used were sheep anti-mouse antibody (NA931) at a 1/2500 dilution or donkey anti-rabbit antibody (NA934) at a 1/5000 dilution, both were supplied by GE Healthcare.Where indicated, cells were treated with or exposed to 1 μM staurosporine (LC Laboratories, Woburn, MA, USA), 25 mM 2-deoxy- d -glucose (2-DG), 25 mM N-acetylcysteine (NAC), 100 μM hydrogen peroxide (H 2 O 2 ), 2 μM UK5099 all from Sigma-Aldrich, Poole, UK, or ionizing radiation (IR) delivered at room temperature using a CellRad cabinet X-ray cell irradiator (Faxitron Bioptics LLC, Tucson, AZ, USA) calibrated to deliver 2 Gy/min of X-ray radiation.

Techniques: Flow Cytometry, Mutagenesis, Positive Control, Functional Assay