title: Celecoxib promotes c-FLIP degradation through Akt-independent inhibition of GSK3.
abstract: Celecoxib is a COX-2 inhibitor that reduces the risk of colon cancer. However, the basis for its cancer chemopreventive activity is not fully understood. In this study, we defined a mechanism of celecoxib action based on degradation of cellular FLICE-inhibitory protein (c-FLIP), a major regulator of the death receptor pathway of apoptosis. c-FLIP protein levels are regulated by ubiquitination and proteasome-mediated degradation. We found that celecoxib controlled c-FLIP ubiquitination through Akt-independent inhibition of glycogen synthase kinase-3 (GSK3), itself a candidate therapeutic target of interest in colon cancer. Celecoxib increased the levels of phosphorylated GSK3, including the α and β forms, even in cell lines, where phosphorylated Akt levels were not increased. Phosphoinositide 3-kinase inhibitors abrogated Akt phosphorylation as expected but had no effect on celecoxib-induced GSK3 phosphorylation. In contrast, protein kinase C (PKC) inhibitors abolished celecoxib-induced GSK3 phosphorylation, implying that celecoxib influenced GSK3 phosphorylation through a mechanism that relied upon PKC and not Akt. GSK3 blockade either by siRNA or kinase inhibitors was sufficient to attenuate c-FLIP levels. Combining celecoxib with GSK3 inhibition enhanced attenuation of c-FLIP and increased apoptosis. Proteasome inhibitor MG132 reversed the effects of GSK3 inhibition and increased c-FLIP ubiquitination, confirming that c-FLIP attenuation was mediated by proteasomal turnover as expected. Our findings reveal a novel mechanism through which the regulatory effects of c-FLIP on death receptor signaling are controlled by GSK3, which celecoxib acts at an upstream level to control independently of Akt.
authors.affiliation: Department of Hematology and Medical Oncology, Emory University School of Medicine and Winship Cancer Institute, Atlanta, Georgia 30322, USA.
authors.forename: Shuzhen
authors.initials: S
authors.lastname: Chen
authors.forename: Wei
authors.initials: W
authors.lastname: Cao
authors.forename: Ping
authors.initials: P
authors.lastname: Yue
authors.forename: Chunhai
authors.initials: C
authors.lastname: Hao
authors.forename: Fadlo R
authors.initials: FR
authors.lastname: Khuri
authors.forename: Shi-Yong
authors.initials: SY
authors.lastname: Sun
mesh_terms: D017209:Apoptosis / Q000187:drug effects; D053446:CASP8 and FADD-Like Apoptosis Regulating Protein / Q000378:metabolism*; D000068579:Celecoxib; D045744:Cell Line, Tumor; D052246:Cyclooxygenase 2 Inhibitors / Q000494:pharmacology*; D038362:Glycogen Synthase Kinase 3 / Q000037:antagonists & inhibitors / Q000378:metabolism; D006801:Humans; D007976:Leupeptins / Q000494:pharmacology; D009369:Neoplasms / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D010766:Phosphorylation / Q000187:drug effects; D046988:Proteasome Endopeptidase Complex / Q000187:drug effects / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism; D059748:Proteolysis / Q000187:drug effects; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D011720:Pyrazoles / Q000494:pharmacology*; D034741:RNA, Small Interfering; D053218:Receptors, Death Domain / Q000378:metabolism; D013449:Sulfonamides / Q000494:pharmacology*; D054875:Ubiquitination
chemical_list: D053446:CASP8 and FADD-Like Apoptosis Regulating Protein; D052246:Cyclooxygenase 2 Inhibitors; D007976:Leupeptins; D011720:Pyrazoles; D034741:RNA, Small Interfering; D053218:Receptors, Death Domain; D013449:Sulfonamides; D051057:Proto-Oncogene Proteins c-akt; D011493:Protein Kinase C; D038362:Glycogen Synthase Kinase 3; D046988:Proteasome Endopeptidase Complex; D000068579:Celecoxib; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde