Figure 3

PPARα antagonist inhibits CLL proliferation. (A) To demonstrate CLL proliferation under the described culture conditions, CLL cells were CFSE-labeled and cocultured with activated allogeneic T cells as described in Materials and Methods. Five days after coculture, CLL cells were stained with CD19-APC and proliferation was assessed by flow cytometry. The histograms to the right show CFSE profile gated on CD19⁺ cells. (B) CLL cells were incubated with either PPARα antagonist NXT629 or the negative control compound NXT962 for 2 h. Subsequently, T cells were added and CLL proliferation was assessed after 5 d. The number of viable cells determined by gating on DiOC₆ bright and PI negative cells with the use of Accuri software is depicted. Data are mean ± SD from three independent experiments using CLL cells from the same donor. Significant difference: *p < 0.05, **p < 0.005, unpaired Student t test. (C) CLL cells were incubated with either PPARα antagonist NXT629 or vehicle control for 2 h. Subsequently, T cells were added, and CLL proliferation was assessed after 8 d. The number of viable cells (DiOC₆ bright and PI negative) was quantified by flow cytometry and normalized to the vehicle control, set as 100%. Data are mean ± SEM from 10 different CLL patient samples. (D) CLL cells were incubated as in (C) and, after 8 d, cell cycle analysis was performed using PI staining as described in Materials and Methods. Percentage of cells in G2/M and G0/1 phase was normalized to vehicle control. Data are mean ± SEM from four different CLL donors. Significant difference: *p < 0.05, **p < 0.005, unpaired Student t test.