Abstract 4028: Identification of a novel combination therapy that prevents the metastatic outgrowth and reduces the viability of dormant breast cancer cells
Authors: Touny Lara H. El , Vieira Anthony , Mendoza Arnulfo , Khanna Chand , Hoenerhoff Mark J. , Green Jeffrey E. 
Published: May 2015

Abstract
Recurrent metastatic disease many years after initial therapy is a major cause of breast cancer (BC) mortality and strongly suggests that disseminated cells survive long periods in a growth-arrested state that is refractory to chemotherapy, known as “cellular dormancy”. Alterations in the components of the extracellular matrix (ECM) are associated with increased metastasis and poor prognosis for BC patients. This suggests that ECM alterations may have a role in influencing the behavior of dormant BC cells. We have previously shown both in vitro (using 3-dimensional Col-1-supplemented Matrigel culture) and in vivo (preclinical model of fibrosis at the lung metastatic site) that alterations in the ECM surrounding disseminated, dormant BC cells can induce signaling pathways requiring Src and ERK1/2 activation that switch dormant D2.0R cells into a proliferative state. We now demonstrate, using the Src inhibitor AZD0530 or Src shRNA, that the proliferative outbreak of dormant BC cells in Col-1-enriched microenvironments is prevented through the induction of a reversible G1-phase arrest and the upregulation of nuclear cyclin dependent kinase inhibitor p27, without affecting the viability of the dormant cells. Src inhibition, however, is not efficacious in regressing already established metastatic lesions, suggesting that Src signaling is critical to engage the dormant-to-proliferative switch but is not necessary for survival once the cells have begun to proliferate, which potentially explains its modest success in BC patients with advanced metastatic disease. Since ERK1/2 activation is also required for the dormant-to-proliferative switch, we determined the effect of the MEK1/2 inhibitor AZD6244 that prevents ERK1/2 activation on the proliferative switch alone and in combination with the Src inhibitor. Whereas AZD6244 is capable of reducing the proliferation of outbreaking cells, combination treatment with AZD0530 and AZD6244 of dormant cells undergoing the dormant-to-proliferative switch induces apoptotic cell death in a significant fraction of the dormant cells, which is not seen with either inhibitor alone, both in vitro and in vivo, and significantly delays metastatic outgrowth in the lung metastatic site. Induction of apoptotic cell death is also observed in a number of human and mouse cancer cell lines treated with AZD0530+AZD6244 but not with either inhibitor alone as they were induced to proliferate in 3-dimensional matrices, suggesting the potential applicability of this combination beyond BC. In conclusion, inhibition of Src alone prevents the proliferative response of dormant cells to external stimuli without affecting survival, but the addition of a MEK1/2 inhibitor suppresses survival of dormant cells, indicating that this combination may have clinical value in preventing breast cancer recurrence.
Citation Format: Lara H. El Touny, Anthony Vieira, Arnulfo Mendoza, Chand Khanna, Mark J. Hoenerhoff, Jeffrey E. Green. Identification of a novel combination therapy that prevents the metastatic outgrowth and reduces the viability of dormant breast cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4028. doi:10.1158/1538-7445.AM2014-4028