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. 2020 Mar 5;10:296. doi: 10.3389/fonc.2020.00296

Figure 1.

Figure 1

Generation and characterization of AZD1775-resistant cell lines. AZD1775 resistance was generated in Molm13, Jurkat and REH cell lines by culturing these cell lines in media containing 50–10,000 nM AZD1775 over a period of 3 months. (A–C) AZD1775-naive and -resistant Molm13 (A), Jurkat (B), and REH (C) cells were treated with 50–2,000 nM AZD1775 for 72 h. Cellular proliferation rates and viability was determined by flow cytometry. Viable cell numbers normalized to cells receiving no treatment (NT) are shown. Results are shown as mean ± SEM from three independent experiments. IC50 values for each cell line are displayed below the corresponding graph. (D–F) Molm13 (D), Jurkat (E), and REH (F) sensitive and resistant cells were treated with the indicated concentrations of AZD1775 for 24 h after which protein lysates were subjected to western blotting with antibodies specific to pCDK1 tyrosine 15, CDK1, and actin. (G–I) Molm13 (G), Jurkat (H), and REH (I) cells resistant to AZD1775 were cultured in drug-free media for 40 passages. Sensitive, resistant cells and resistant cells cultured in drug-free media were treated with 50–2,000 nM AZD1775 for 72 h. Viable cell counts normalized to cells receiving no treatment are shown. Results are shown as mean ± SEM from three independent experiments. ****P < 0.0001.