Abstract 4125: Targeting metastasis and cancer stem-like cells in triple negative breast cancer through inhibition of focal adhesion kinase

Abstract

Abstract Triple negative breast cancer (TNBC) is an aggressive subtype of breast cancer closely associated with an early pattern of metastasis and lack of targeted therapies. As such the prognosis for TNBC is poor. Whilst targeting signalling pathways known to promote metastasis-supporting behaviours may well provide an important therapeutic opportunity for such cancers, recent evidence points to an association between cancer stem-like cell (CSC) populations and the aggressive behaviours of TNBC, including their chemoresistance. Aberrant activity and/or expression of the non-receptor tyrosine kinase FAK (focal adhesion kinase) is known to contribute to the development and progression of cancer due to its central role as a mediator of cell attachment, migration and survival. More recently, FAK has been suggested to influence anoikis resistance, thus implying its activity may contribute to the phenotype of CSCs. Here we have explored the role of FAK in TNBC cells as a putative therapeutic target to suppress the metastatic phenotype. FAK activity was inhibited in MDA-MB-231 (TNBC) and MCF7 (luminal breast cancer) cells pharmacologically using PF-562,271 (PF271), a competitive-inhibitor of the ATP-kinase domain or through siRNA-mediated knockdown to probe scaffolding function of FAK. Functional assessment of TNBC proliferation, migration and invasion was performed using cell counting and Boyden chamber assays respectively, whilst stem-like behaviours (anchorage-independent growth, self-renewal) were investigated using stem-cell enrichment (mammosphere-based assays). The surface marker profile of CSCs was investigated using FACS, while changes in intracellular signalling pathways in response to FAK inhibition was revealed through Western blotting and immunofluorescent microscopy. MDA-MB-231 and MCF7 cells displayed comparable levels of total and activated (Y397/Y861) FAK which were depleted in response to PF271 in a dose dependent manner: however, TNBC cells were significantly more sensitive. Cellular migration in TNBC cell models occurred in a FAK-dependent manner with both PF271 and FAK siRNA suppressing endogenous and fibroblast-stimulated migration/invasion (p<0.001 versus vehicle control). Inhibition of FAK also caused a significant decrease in the growth rate of TNBC cells. FACS analysis of MDA-MB-231 cell cultures revealed a significant subpopulation of cells exhibiting the CSC surface marker profile CD24-/CD44+. Moreover, TNBC cells were able to form self-renewing mammospheres in non-adherent culture. PF271 treatment or siRNA-mediated knockdown of FAK significantly attenuated mammosphere-forming ability and self-renewal of these cultures. These data point to a role for FAK as a mediator of metastatic behaviour in TNBC cells, whilst also showing the capacity to influence self-renewal of a CSC population. FAK may therefore represent an emerging therapeutic target in such cases. Citation Format: Samuel Jones, Christopher Smith, Stephen Hiscox, Wen G. Jiang. Targeting metastasis and cancer stem-like cells in triple negative breast cancer through inhibition of focal adhesion kinase. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4125.