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. Author manuscript; available in PMC: 2020 Mar 1.
Published in final edited form as: Cancer Res. 2019 Jul 4;79(17):4439–4452. doi: 10.1158/0008-5472.CAN-19-0024

Figure 5. HCC4006 Ge-R cells with mesenchymal phenotype are sensitive to MEK inhibition.

Figure 5.

A. Cell surface expression of CXCR7 in HCC4006 cells ectopically expressing LacZ-V5 or CXCR7-V5. Average of five independent assays with duplicate runs, ****p≤0.0001, ***p≤0.001. B. Lysates made from HCC4006 cells ectopically expressing LacZ-V5 or CXCR7-V5, treated with DMSO (DAY0) or 1 μmol/L osimertinib for 7 days or 14 days, were subject to immunoblots with antibodies indicted. Image is representative of three independent experiments. C. Lysates used in Fig.5B was subject to immunoblots with indicated antibodies. Image is representative of three independent experiments. D. HCC4006 cells ectopically expressing LacZ-V5 or CXCR7-V5 were treated with DMSO or 1 μmol/L osimertinib for the days indicated and surviving cells were stained with crystal violet. Image is representative of three independent assays. E. Additional set of cells were prepared and treated as described in Fig.5D. The surviving cells were counted. Average of three independent assays, p=0.0019. F. HCC4006 and Ge-R cells were treated with DMSO, 500nmol/L of gefitinib (EGFRi) or 500nmol/L trametinib (MEKi) or 500nmol/L buparlisib (PI3Ki) for 72 hours. Lysates were subjected to immunoblot with antibodies indicated. Image is representative of three independent experiments. G. HCC4006 and Ge-R cells were treated as in Fig.5F and Annexin V apoptosis assay was performed. Average of three independent assays. Bars, S.D. (***p≤0.001, **p≤0.01, *p≤0.05). H. HCC827 cells ectopically overexpressing constitutively active MEK (MEKDD) or pBabe control vector (Vector) were treated with DMSO (D), osimertinib (O), crizotinib (C) or combination (O/C) and phosphorylation of ERK1/2 for 48hours and EGFR and apoptotic markers were assessed. Image is representative of three independent experiments. I. Annexin V apoptosis assay was performed for the experiment described in Fig.5H. Results are average of three independent assays. (N.S. not significant, ****p≤0.0001). Average of three independent assays. Error Bars, S.D.