Abstract LB-100: Computational assessment of oncogenic pathways modulated by vorinostat in breast cancer stem cells

Abstract

Abstract Gene expression analysis predicted strong activation of HDACs, MYC and NF-kappaB with potential sensitivities to vorinostat in breast cancer stem cells. Vorinostat was subsequently confirmed to inhibit mammosphere formation of the ALDH+ subpopulation of SUM159. In the present study, we conducted microarray experiments with RNA extracts from stem cells (ALDH+) and non-stem cell (ALDH-) populations of SUM159 in the absence or presence of 1uM vorinostat for 12h and 24h. The action of vorinostat was modeled by a series of computational tools integrating chemical genomics, gene signatures, gene promoters, microRNAs, pathways and gene networks. In the stem-cell population (ALDH+ SUM159), promoter and gene signature analysis revealed that MYC and NF-kappaB were strongly down-regulated by vorinostat at 12h and more prominently at 24h. MicroRNAs predicted to be most likely down-regulated by vorinostat at 12h were known transcriptionally regulated by either MYC or NF-kappaB. Furthermore, Notch1 and its ligand JAG2 were identified as key factors in modulating MYC activities in response to vorinostat. The predicted MYC down-regulation by vorinostat was confirmed by western blot and RT-PCR and NF-kappaB down-regulation by luciferase reporter assay. Other pathways such as beta-catenin and SRC were not significantly modulated by vorinostat. Surprisingly, the RAS pathway was activated by vorinostat with up-regulation of a RAS-activation gene signature, over-expression of KRAS/MEK/ERK and potential down-regulation of let-7, a KRAS targeting microRNA. In addition, epithelial-to-mesenchymal transition (EMT) was predicted to be activated by vorinostat with up-regulation of an EMT gene signature (TGFB1, SNAIL, SLUG, TWIST, ZEB1, ZEB2, alpha-SMA, FN1, VIM, N-cadherin etc.) and potential down-regulation of miR-200, an EMT antagonizing microRNA. Activation of RAS and EMT could render drug resistance in bCSCs. In the non-stem cell population (ALDH- SUM159), vorinostat down-regulated MYC and NF-kappaB at 12h but to a lesser extent at 24h. Unlike the ALDH+ cells, the RAS pathway was down-regulated at both 12h and 24h with no significant change in EMT. In summary, vorinostat exhibited differential activities in the stem cell and non-stem cell populations of SUM159 with prominent down-regulation of MYC and NF-kappaB, and divergent activities in RAS and EMT. A gene network was constructed to model the dynamics of vorinostat action in bCSCs. Further investigations were warranted including potential drug combinations. The computational framework proved complementary to the wet-lab approaches and capable of providing guidance in therapeutic discovery in targeting breast cancer stem cells. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-100. doi:1538-7445.AM2012-LB-100