Celecoxib downregulated COX-2 protein expression in all MDA-MB-231 and MDA-MB-435 variants, but celecoxib downregulated Bcl-2 expression in only the MDA-MB-435-E1A stable transfectants. (a) Western blots of MDA-MB-231 and MDA-MB-435 cells treated with 0 or 40 μM celecoxib for 5 days and tested for COX-2 and Bcl-2. Percentages indicate differences relative to the 0 μM control samples. Protein expression was considered to be downregulated if the treated condition was at least 20% less than the control (untreated) condition. (b) Time course of Bcl-2 expression after treatment with 0 or 40 μM celecoxib in MDA-MB-435-E1A and MDA-MB-231-E1A stable transfectants. Bcl-2 was suppressed at both 72 and 96 h in the MDA-MB-435-E1A stable transfectants but was not suppressed in the MDA-MB-231-E1A stable transfectants. (c) Transfection of MDA-MB-435 cells with Bcl-2 DNA (+) or a control DNA (-) led to overexpression of Bcl-2 in all variants. (d) MDA-MB-435-E1A cells made to overexpress Bcl-2 and non-Bcl-2-overexpressing cells were treated with 0 or 40 μM celecoxib for 5 days, and cell viability was determined with a trypan-blue assay. Bcl-2 overexpression did not restore sensitivity to celecoxib (P = 0.11).