Selective cytotoxicity of 2ME2 against human MM cells.

(A) MM.1S cells were treated with 2ME2 (3 μM) for the indicated times and analyzed for apoptosis by DNA fragmentation (left panel is a representative of 3 separate experiments with similar results) and flow cytometric analysis of PI− and HO+ apoptotic cells (right panel is the mean ± SD from 3 independent experiments;P < .005). (B) MTT assay was performed after incubation of MM cell lines (MM.1S, ▵; RPMI-8226, ○; LR-5, ♦; Dox-6, ⋄; Dox-40, ■; MM.1R,) with the indicated doses of 2ME2 for 72 hours. Results are mean ± SD from 5 independent experiments; P < .0001 for all cell lines. (C) Effect of treatment with 2ME2 (0-20 μM) for 72 hours on normal lymphocyte viability, assessed by MTT assay. Results are the mean ± SD of 5 independent experiments; P = 0.23 from J-T test for trend. (D) MM cells (CD138+) from 5 patients (patients 1-5) were treated with 2ME2 (9 μM) for 72 hours, followed by BrdU assay. Values are the mean ± SD of triplicate samples (P = .06); experiments were repeated 3 times with similar results. (E) 2ME2 induces proteolytic cleavage of PARP in patient MM cells. CD138+ cell from 2 MM patients and normal lymphocytes from 2 healthy donors were treated with 2ME2 (9 μM) for 72 hours and harvested, and total protein lysates were subjected to SDS-PAGE analysis. Immunoblot analysis of the lysates was performed with anti-PARP antibody. FL indicates full length; CF, cleaved fragment.