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. 2020 Apr 24;10:7004. doi: 10.1038/s41598-020-64018-z

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© The Author(s) 2020

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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ADR-resistant (R) MDA-MB-231 cells are also resistant to SM-164, while SM-164-R MDA-MB-231 cells are sensitive to ADR. (A) Growth curve of the parental and ADR-R MDA-MB-231 cells, starting at 1 × 10⁴ cells/well, in a well of 6-well-plates treated with 30 ng/ml ADR or vehicle, and passaged to 60- and 100-mm dishes, respectively, when untreated cells grew to sub-confluence. 3 wells/group, **p < 0.01, one-way ANOVA +/Dunnett test. (B) Parental and ADR-R MDA-MB-231cells were treated with the indicated doses of SM-164 plus 1 ng/ml of TNFα overnight. Annexin V⁺PI^+/− apoptotic cells were analyzed by flow cytometry. (C) 0.5 × 10⁴ parental and SM-R MDA-MB-231 cells in 60 mm dishes were cultured overnight followed by treatment with 1 ng/ml TNFα plus the indicated doses of SM-164 for 3 days, and surviving cells were counted. (D) The parental and SM-164-R cells (0.5 × 10⁴ cells in 60 mm dishes) were treated with the indicated doses of ADR for 6 days and cell numbers were counted after digestion with 0.2% trypsin. **p < 0.01 vs. the respective parental cells, one-way ANOVA +/Dunnett test. The experiments were repeated 3 times with similar results.