Adenovirus type 5 E1A-induced apoptosis in COX-2-overexpressing breast cancer cells.
BACKGROUND
Suppression of Bcl-2 expression can overcome cellular resistance to apoptosis induced by the adenovirus type 5 gene E1A in models of ovarian and breast cancer. Celecoxib, a cyclooxygenase-2 (COX-2) inhibitor, is known to downregulate Bcl-2 expression. We hypothesized that celecoxib would enhance E1A-induced apoptosis by suppressing Bcl-2 through suppressing COX-2 expression. If successful, this strategy could represent a means of overcoming resistance to E1A gene therapy.


METHODS
We first established the cytotoxicity of celecoxib in two COX-2-overexpressing E1A-transfected breast cancer cell lines (MDA-MB-231 and MDA-MB-435) and in two low-COX-2-expressing E1A-transfected cell lines (MCF-7 (breast cancer) and SKOV3.ip1 (ovarian cancer)). We next tested whether higher sensitivity to celecoxib among these cell lines resulted from increased apoptosis by flow cytometry and western blotting. We further investigated whether suppression of Bcl-2 by celecoxib was involved in the apoptosis resulting from celecoxib treatment, and we explored whether the celecoxib-induced apoptosis in these cells depends on a COX-2 downstream pathway.


RESULTS
The two COX-2-overexpressing cell lines MDA-MB-231-E1A and MDA-MB-435-E1A were more sensitive to celecoxib than the corresponding control cells, but the two low-COX-2-expressing cell lines MCF-7-E1A and SKOV3.ip1-E1A were no more sensitive than control cells to celecoxib. Therefore, we used the MDA-MB-231-E1A and MDA-MB-435-E1A cells for all further experiments. In both cell lines, sub-G1 fraction was increased, or cleavage of PARP and caspase-9 were increased after 5 days of exposure to 40 microM celecoxib. However, Bcl-2 was suppressed only in the MDA-MB-435-E1A cells and not in the MDA-MB-231-E1A cells. Restoring Bcl-2 expression in the MDA-MB-435-E1A stable transfectants did not affect their sensitivity to celecoxib. However, adding prostaglandin E2 (PGE2) or PGF2alpha blunted the sensitivity to celecoxib of both E1A stable transfectants.


CONCLUSIONS
We speculate that one mechanism by which celecoxib enhances E1A-induced apoptosis in cells that express high levels of COX-2 is through blocking PGE2 or PGF2alpha.
Sugimoto
Takeshi
T
Breast Cancer Translational Research Laboratory, The University of Texas M, D, Anderson Cancer Center, Houston, TX, USA. tks_sugimoto@yahoo.co.jp.
Bartholomeusz
Chandra
C
Tari
Ana M
AM
Ueno
Naoto T
NT
D017500:Adenovirus E1A Proteins / Q000235:genetics* / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D001943:Breast Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D000068579:Celecoxib; D002470:Cell Survival / Q000187:drug effects; D051546:Cyclooxygenase 2 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D052246:Cyclooxygenase 2 Inhibitors / Q000494:pharmacology*; D015237:Dinoprost / Q000378:metabolism; D015232:Dinoprostone / Q000378:metabolism; D005260:Female; D005434:Flow Cytometry; D006801:Humans; D010051:Ovarian Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D011720:Pyrazoles / Q000494:pharmacology*; D013449:Sulfonamides / Q000494:pharmacology*; D014162:Transfection; D014407:Tumor Cells, Cultured / Q000187:drug effects
D017500:Adenovirus E1A Proteins; D052246:Cyclooxygenase 2 Inhibitors; D019253:Proto-Oncogene Proteins c-bcl-2; D011720:Pyrazoles; D013449:Sulfonamides; D015237:Dinoprost; D051546:Cyclooxygenase 2; C496540:PTGS2 protein, human; D000068579:Celecoxib; D015232:Dinoprostone