Signaling networks of convergence of nongenomic and genomic estrogen actions mediated by Erbb2 and Akt1 in MCF-7 cells
Authors: Adriana Stoica, Rebecca Riggins, Assya Abdallah, Emily Wang, Marissa Fox, Roxana Guerra, Lauren Mautner, Heather Summe, Cielo Zislis, Habtom Ressom
Journal: Cancer Research (2008)

5454   We have demonstrated that in MCF-7 cells, the nongenomic and genomic estradiol actions converge in the ErbB2/PI3-K/Akt1 pathway that plays an important role in the development of antiestrogen resistance. MCF-7 cells stably transfected with constitutively active Akt1 (myr-Akt1) increased estrogen receptor-a (ER-a) activity and hormone-independent cell growth  in vitro  and  in vivo  and this effect was blocked by the selective ErbB2 inhibitor, AG825, but only partially by tamoxifen. In contrast, in cells stably transfected with dominant negative Akt1 (K179M-Akt1 or R25C-Akt1), ER-a activity was inhibited and the  in vitro  and  in vivo  growth effects were reduced.  We used the same MCF-7/Akt1 breast cancer model to perform global gene expression profiling with the Affymetrix HG-U133A 2.0 GeneChip array and real time RT-PCR, to identify patterns of genes regulated by estradiol and Akt1 in the presence or absence of tamoxifen, ICI 182,780, and AG825. Of the 22,000 probe sets including 14,500 characterized human genes, over 1,500 showed a robust regulation. While 71 genes (87%) were up-regulated and 11 (13%) down-regulated by 20 min estradiol treatment, our results show an inverse regulation pattern with 24h estradiol treatment: 177 stimulated (48%) and 192 inhibited genes (52%). In myr-Akt1 cells, 464 genes (34.5%) were stimulated and 882 (62.5%) inhibited. Most of the stimulated or inhibited genes upon 20 min of estradiol treatment were involved in signaling, transcription, cancer, and apoptosis. In contrast, in the 24h treatment group, most genes’ functions were metabolism, immune function, signaling, cancer, transport, and gene regulation. From the myr-Akt1-regulated genes, most were involved in metabolism, signaling, immune function, degradation, cancer, biosynthesis, cell cycle, transcription, apoptosis, and gene regulation. Most of the estradiol- and myr-Akt1-regulated genes (82-96%) were mediated by ER-a, since both antiestrogens blocked their effect. AG825 also blocked the effect of 95-97% of estradiol- and myrAkt1-regulated genes. Only 2 genes were stimulated by both estradiol (20 min and 24h) and myr-Akt1: DUSP4 and GADD45A, but their effect was blocked by antiestrogens and AG825. 10 genes were regulated by estradiol 20 min and myr-Akt1 and 80 by estradiol 24h and myr-Akt1. 18 genes were regulated by 20 min and 24h estradiol treatment. All these genes were blocked in K179M-Akt1 cells and by tamoxifen, ICI 182,780 (except one) and AG825. Using a software program from Ingenuity Systems, we constructed a signaling network with estrogen- and Akt1-regulated genes downstream of ErbB2: SGK, EGR3, PDLIM3, DUSP4, MAFF, CYP51A1, DLG1, BIRC5, and SLC7A2.  Taken together, our results reveal the importance of the convergence of nongenomic and genomic estrogen actions in the ErbB2/PI3-K/Akt/ER-a pathway. The newly identified genes may provide new therapeutic targets for endocrine therapy.