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. 2014 Oct 20;124(11):5074–5084. doi: 10.1172/JCI76539

Figure 2. Impact of BRAF and MEK inhibitor treatment on signaling and survival of CLL cells.

Figure 2

Highly purified (>97%) CD19+CD5+ cells or CD14+ myeloid cells obtained from the patient were cultured in the presence of the BRAF inhibitors (vemurafenib, dabrafenib), the MEK inhibitor trametinib, BRAF inhibitor combined with MEK inhibitor, or DMSO only at the indicated concentrations. (A) OD as an indicator for viability and metabolic activity of CLL cells under different vemurafenib concentrations in an MTT assay. The experiment was performed twice using in dependent samples from the patient with similar results. Levels of pERK and tERK in the CLL cells (B) or CD14+ cells (C) derived from PBMC were measured by Western blot. We used vemurafenib, dabrafenib, and trametinib (0.07 μM) as indicated. One of 3 independent experiments with similar results is shown. (D) Western blot analysis for pERK and tERK of the protein lysate of highly purified (>97%) patient-derived CD19+CD5+ cells at the indicated concentrations of dabrafenib (Dab; 6 μM) and trametinib (MEK-i; 0.03, 0.07, 0.14 μM). Quantification of the protein amount of the described groups shown as a bar diagram. The experiment was performed 3 times with similar results. (E) The amount of CLL cells in the peripheral blood of Rag2–/–γC–/– mice relative to start of treatment with vehicle or vemurafenib (24 mg/kg/d). (F) The survival of the Rag2–/–γC–/– mice treated as described in panel E is shown (P = 0.0004). Data from 3 independent experiments were pooled.