Figure 5
Prolonged exposure of myeloma cells in culture to high-dose lenalidomide induces resistance to antiproliferative effect of lenalidomide correlating with decreases in CRBN. (a) Time course of acquisition of lenalidomide resistance in H929 cells and concurrent reduction in CRBN mRNA determined by reverse transcription--PCR (data for one clone R10-4 is shown, representative of generation of four independent lenalidomide resistant cells lines, H929 R10-1, R10-2, R10-3 and R10-4 ). H929 cells were treated with control (final 0.1% DMSO) (D) or lenalidomide (1 μM) (L1) for 2 months until the proliferation of cells was no longer inhibited by 1 μM lenalidomide. The lenalidomide treated cultures were subsequently treated with 10 μM lenalidomide for a further 4 months to fully establish resistance to high-dose lenalidomide (30 μM). Arbitary units for CRBN mRNA are shown. (b) Immunoblot of CRBN in H929 cells treated with DMSO for 6 months (H929/D1) and H929 lenalidomide-resistant cells lines (R10-1, R 10-2, R 10-3 and R 10-4). (c) Dose-dependent inhibition of proliferation of parental H929 cells (•) and H929/D1 cells grown continuously in 0.1% DMSO (▪) cells by lenalidomide. Lack of inhibition of proliferation by lenalidomide, up to 10 μM concentration, of four H929 cell lines (R10-1, R10-2, R10-3, R10-4; ○, □,∇, ◊) made resistant to high-dose lenalidomide over 6 months in culture. (d) Dose-dependent inhibition of proliferation of parental H929 cells (•) and H929/D1 cells grown continuously in 0.1% DMSO (▪) cells by pomalidomide. Dose-dependent inhibition of proliferation by pomalidomide of four H929 cell lines made resistant to high-dose lenalidomide over 6 months in culture.
