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. 2018 Oct 15;9:4275. doi: 10.1038/s41467-018-06676-2

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© The Author(s) 2018

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 3 — Combinatorial MEK and CENPE inhibition results in synergistic cell death. a Each frame represents movie stills from time-lapse longer-term live-cell imaging as cells undergo mitosis starting from nuclear envelope break down (NEBD) to exit from mitosis. b Each bar graph show total time duration of each of the individually tracked cells spent in mitosis. The NEBD was taken as the beginning of mitosis. Individual cells (n = ~20/treatment group) were manually tracked from lime-lapse movies until they exit mitosis (anaphase) or died in mitosis for each of the treatment groups. c Flow cytometry profiles of DNA content in PDX366 PDAC cells treated with control, single-agent, or combinatorial MEK and CENPE inhibitors are shown. d Bar graphs represent treatment-mediated percent changes in cells with 2n (G1), 4n (G2), and >4n (polyploidy) DNA. e Immunofluorescent images of PDX366 PDAC cells treated with control and combinatorial (trametinib and CENPE inhibitors) are shown after DAPI and tubulin staining in the first two left columns respectively. f Representative hematoxylin and eosin (H&E)-stained tumor sections are shown. Tumors were harvested from mice that had undergone 72 h of treatment with 125 mg/kg CENPE inhibitor alone or in combination with 0.3 mg/kg trametinib. Arrows indicate mitotic figures. g Dot-plot showing the number of mitotic cells quantified from 10 different high-power imaging fields (HPFs) per tumor section. h Results show MRI-measured effects of single and combination of drugs on tumor formation. At 1 week after orthotopic implant, mice received control, MEK inhibitor (trametinib), CENPE inhibitor (GSK923295), or combined MEK and CENPE inhibitor. Results show MRI-measured tumor volumes after 4 weeks of treatment. i MRI-measured tumor volumes are shown in mice where tumors were allowed forming for 4 weeks and the treatments (as in e) were started. Beginning of treatment is marked with an arrow. Bounds of the box spans from 25 to 75% percentile, center line represents median, and whiskers visualize 5 and 95% of the data points. Individual data points are marked with circles or dots. Error bars represent the standard error of the mean value of at least three independent experiments. Symbols * and *** denote P < 0.05 and P < 0.001, respectively