Figure 1
Sorafenib causes apoptosis of fresh and viably freeze-thawed CLL cells, in the presence of cells from the microenvironment. (A) CLL PBMCs were isolated from blood freshly drawn from CLL patients and exposed immediately after isolation to increasing doses of sorafenib. CLL cells were harvested after 48 h, stained with DiOC6/PI and analyzed by flow cytometry to determine viability. The fraction of viable cells, identified as PI-negative and DiOC6 bright cells, are represented, relative to d 0 (100%). Data shown are means ± SD from five different CLL samples. (B) Freshly isolated CLL PBMCs were exposed to 1 µmol/L sorafenib or equivalent volume of DMSO as vehicle control on d 0, 1, 2, 3, 6, 7 and 8, as indicated by the arrows. Cell viability was measured as above. Data shown are mean ± SD from eight different CLL samples. Statistically significant difference: *P < 0.05, **P < 0.01, ***P < 0.001 (paired Student t test). (C) CLL B cells were purified from freeze-thawed PBMCs and cultured in the presence of NLCs. Sorafenib (1 µmol/L) or DMSO was added to the co-cultures on d 0, 1, 2, 3, 6 and 7, as indicated by the arrows. CLL cell viability was measured as above. Data shown are mean ± SD from four CLL samples. (D) Representative flow cytometry data for (C) is shown for one patient sample in the presence of sorafenib or DMSO control and NLCs (d 3 and 8). (E) CLL B cells purified from freeze-thawed PBMCs were cultured in the presence of MSCs and a single dose of sorafenib (10 µmol/L) or the equivalent volume of DMSO was added at d 0. CLL cell viability was measured and analyzed as above. Data shown are from two CLL samples cocultured with MSCs derived from two different CLL patients, for a total of four independent experiments (mean ± SD). (F) Representative flow cytometry data for (E) is shown for one patient sample in the presence of sorafenib or DMSO control and MSCs (d 2 and 4).