Pharmacologic inhibition of Sp1 by TMP decreases WM cell growth. (A) WM and lymphoma cell lines were treated with various concentrations of TMP (1 to 20 µM) for 48 hours, and cell growth was assessed by [³H]thymidine uptake. Data are presented as a percentage of untreated cell proliferation. (B) Flow cytometric analysis of bromodeoxyuridine (BrDU) incorporation was performed after treatment of BCWM1 cells with the inhibitor for 24 hours. Data shown are percentage of cells in the different phases of the cell cycle. (C) BCWM.1, MWCL1, and primary CD19⁺ WM cells (WM1 and WM2) were cultured with different concentrations of TMP for 24 hours. Cell survival was assessed by CTG, and presented as a percentage of growth of vehicle-treated cells. (D) BCWM.1 cells were left untreated or treated with 10 µM of TMP for different time points. Cells were then subjected to WB analysis using PARP and caspase-3 abs. GAPDH ab was used as loading control for WB analysis (right). Densitometric quantitation of band intensity was performed using ImageJ software. Data were normalized to GAPDH. Fold change compared with time = 0 is shown on graph (left). (E) BCWM.1, WSU-WM, MWCL1, and primary CD19⁺ WM cells (WM3 and WM4) were cultured in the absence (-) or presence (+) of BMSC from WM patients at different concentrations of TMP for 48 hours. Cell proliferation was assessed by [³H]thymidine uptake, and presented as percentage of growth of vehicle-treated cells cultured in the absence of BMSC (100% = control). (F) BCWM1 cells were injected s.c. in SCID mice. Treatment started following detection of tumor (approximately 3 weeks from cell injection). Mice were treated either with 50 mg/kg of TMP or placebo s.c. daily for 2 weeks. Tumors were measured in two perpendicular dimensions once every week.