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. 2017 Dec 4;7:16863. doi: 10.1038/s41598-017-17319-9

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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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RR cells demonstrate no crizotinib—ALK binding and higher expression level of β-catenin compared to RU cells. (A) CETSA was performed to compare crizotinib—ALK binding ability between RU and RR cells. RU and RR cells derived from SK-N-SH were treated with DMSO or 500 nM crizotinib for 6 hours. Representative ALK western blots are shown on the upper panel. Vinculin level was blotted as a loading control. The densitometry quantification data from 3 independent experiments are shown on the lower panel. All data are presented as mean ± SD. Student’s t test was performed. (B) The expression of ALK and β-catenin protein in RU and RR cells were measured by western blots. Vinculin level was blotted as a loading control. (C) ALK pull-down was performed using co-immunoprecipitation assay and showed substantial ALK-β-catenin binding in RR cells compared to RU cells.