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OBJECTIVE.-Overexpression of EGF receptors and constitutive cyclin D1 expression are frequently associated with human squamous carcinomas. One mechanism for mitogenic activation mediated by EGF is through induction of cyclin D1 . Besides their known growth promoting role , EGF signalling and cyclin D1 expression may prevent tumor cell apoptosis . We now investigated whether A431 carcinoma known to overexpress EGF receptors become more resistant to apoptosis ,when engineered to express a cyclin D1 transgene. RESULTS.- exposure of A431 cells to 20 nM okadaic acid induced apoptosis-associated caspase 3 activation , DNA fragmentation , cleavage of Poly ADP-Ribose Polymerase (PARP), p53-independent expression of pro-apoptotic bax , and loss of proliferation-promoting cyclin D1. Allthese alterations were antagonized by concurrent addition of exogenous EGF. Ectopic overexpression of the cyclin D1 gene conferred resistance to 20 nM okadaic acid irrespective of exogenous EGF, associated with a parallel induction of anti-apoptotic bcl-2. Treatment with a subtoxic concentration of a bispecific bcl-2/bcl xL antisense oligonucleotide cooperated with okadaic acid to down-regulate bcl-2 and sensitize cyclin D1-overexpressing cells to okadaic acid-induced cell death . CONCLUSIONS.-Although EGF protects EGF-Receptor proficient epithelial cells from diverse apoptotic stimuli through induction of anti-apoptotic Mcl-1, this is the first report demonstrating that cyclin D1 overexpression provides an EGF independent protection from okadaic acid- induced cell death in A431 squamous carcinoma through induction of bcl-2 . We also show that this anti- apoptotic effect of cyclin D1 overexpression ,can be partl antagonized with antisense strategies that down-regulate anti-apoptotic bcl-2 family members. e-mail mrieber@ivic.ve

[Proc Amer Assoc Cancer Res, Volume 47, 2006]