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. 2017 Jun 1;7:2602. doi: 10.1038/s41598-017-02483-9

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© The Author(s) 2017

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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ENb-binding to EGFR is critical for ENb-TRAIL activation of apoptosis. (A) Differential cell membrane EGFR, DR4, and DR5 expression levels in LN229, HT29 and Calu1 cells measured by Flow Cytometry. Left panel: cell membrane EGFR expression. Right panel: cell membrane DR4 and DR5 expression. (B–C) Cells were pretreated with Cetuximab for 30 min and then treated with ENb-TRAIL for 8 h and apoptosis markers were analyzed by Western blotting (B) and caspase 3/7 assay (C). *P < 0.05 determined by unpaired t test. Error bars indicate SD. (D) Co-immunoprecipitation and Western blot analysis showing EGFR and DR5 complex formation in the presence of ENb-TRAIL and the attenuation of complex by Cetuximab. Western blots were cropped to show specific bands only. For uncropped blots see Fig. S12.