377 Selective Inhibition of AKT Protein Isoforms Augments Gefitinib Response in EGFR Mutated NSCLC Cells

Abstract

Introduction: Genetic changes occurring in the triple negative (TN) subset of breast tumors (BT) contribute to different phenotypes in the tumor cells via deregulation of specific signaling-pathways. We identified a differential upregulation of the Wnt-beta-catenin pathway in TNBT (Barwick etal, 2008; Dey etal. 2009), and reported that in TNBT, the poor prognosis and the higher incidence of metastasis are associated with an activation of this pathway (Barwick etal., 2009). We also reported a role of the pathway in the control of migration and invasion of tumor cells (Dey etal. 2009, 2010). Since integrin-directed migration and invasion of the tumor cells are instrumental for the metastasis, and TN tumor cells are highly invasive in nature, we hypothesized that the upregulation of this pathway in TNBT is functionally connected to the control of metastasis-associated phenotypes. This hypothesis is further strengthened by our identification of MMP7 as a selective-biomarker of TNBT (Abramovitz etal, 2009). Purpose: We studied the involvement of the pathway in the control of metastasis-associated phenotypes in TN breast cancer (BC) cells. Methods: We tested the effects of (1) different chemical modulators of the Wnt signaling, and (2) downregulation of beta-catenin on the (a) cell signaling marker(s), (b) cell proliferation, (c) clonogenic growth, (d) migration, (e) invasion, and (f) vascular mimicry (VM) using a panel of six TNBT cells. Results: (1) CHIR99021 (selective GSK3b inhibitor which activates Wnt signaling) enhanced clonogenic growth, and VM while WntC59 (palmitylates Wnt proteins and blocks the Wnt secretion/activity), as well as XAV939 (tankyrase inhibitor that blocks beta-catenin-mediated transcription via axin stabilization) blocked clonogenic growth, cell proliferation, fibronectindirected migration, and VM in TNBT cells. (2) Sulindac sulfide (decreases cellular levels of total and active beta-catenin) inhibited migration, invasion, VM, actin cytoskeleton, podia-parameters, RAC1/Cdc42 activation, and MMP7 in TNBC cells. (3) SiRNA mediated downregulation of betacatenin protein in TNBC cells inhibited cell proliferation, clonogenic growth, migration, invasion, c-MYC, and VM. Conclusion: Wnt-beta-catenin pathway activates cellular signaling for the metastasis-associated phenotypes like proliferation, clonogenic growth, migration, invasion and VM in TNBT cells and this function is directly mediated by via cellular levels of beta-catenin. An inhibition of the Wntbeta-catenin pathway might be an effective combination treatment strategy in TNBC.