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. 2020 Oct 30;40(2):322–333. doi: 10.1038/s41388-020-01530-6

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© The Author(s) 2020, corrected publication 2023

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. 6 — a Correlation between EMT transcription factors and NRP1 gene expression in the METABRIC dataset. b Phase-contrast microscopy showing the epithelial and mesenchymal phenotypes of control (YFP) and Twist-overexpressing HMLE, MCF-10A, and NMuMG cells. c RT-PCR showing NRP1 upregulation following Twist overexpression. Data are normalized to NRP1 levels in the control (YFP) cells for each cell type and are the mean ± SE of duplicate experiments completed in triplicate. d Immunoblot analyses showing expression of NRP1 and other EMT proteins in response to Twist overexpression. e RT-PCR showing NRP1 in control (YFP) and Twist-overexpressing cells following 7 days of treatment with the indicated concentrations of JQ1. Data are the mean ± SE of triplicate experiments resulting in the indicated P values. f Immunoblot analyses of NRP1 expression in control (YFP) and Twist-overexpressing HMLE and MCF-10A cells following 7 days of treatment with the indicated concentrations of JQ1. g RT-PCR analyses of NRP1 expression in afatinib-resistant (BMAR) cells following 7 days of treatment with JQ1 (50 nM). Data are the mean ± SE of triplicate experiments. h Immunoblot analyses of FGFR1 and NRP1 expression in BMAR cells following 7 days of treatment with JQ1 (50 nM). Expression of tubulin served as a loading control. i Phase-contrast images showing 3D spheroid growth of BMAR cells in response to FGF2 (20 ng/ml) in the presence or absence of JQ1 (50 nM). j Measurement of BMAR spheroid radius at the indicated time points, under the conditions described in i. Data are represented as a boxplot including the measurement of at least three images per condition and are representative of two independent experiments.