480713.fig.006a
(a)
480713.fig.006b
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480713.fig.006c
(c)
Figure 6: Inhibition of ALDH with disulfiram inhibits metastatic properties of ALDH-high K7M2 cells. (a) Disulfiram (250 nM) was added to ALDH-high K7M2 cells and the cells were cultured for at least 24 hours in 10% FBS growth medium. RT-PCR was used to determine gene expression differences as a result of this treatment, with GADPH used as a control. Both mTOR and c-Myc expression were reduced as a result of the treatment with disulfiram. (b) Immunostaining with phospho-4E-BP1 (a mouse anti-mTOR antibody) was done to confirm that treatment with disulfiram reduced expression of mTOR As the concentration of disulfiram was increased (0 nm, 100 nm and 500 nm) cells displayed progressively less mTOR expression. (c) Morphologic differences after treatment with disulfriam were also present, with disulfiram treated cells (stained for actin) appearing less pleomorphic and with fewer filopodia.