Serotonergic Antagonists Affect the Activity of Breast Tumor Initiating Cells in Human and Mouse Models of Breast Cancer

Abstract

Breast cancer is the most frequently diagnosed cancer and the leading cause of cancer-related death amongst women worldwide. The relatively unchanging breast cancer-associated mortality rate is in part due to the existence of rare tumor cells (breast tumor initiating cells; BTIC) that possess stem-like properties permitting them to survive therapy and initiate disease recurrence. Hence, identifying agents capable of eradicating these cells would be a favourable therapeutic strategy to improve the durability of breast cancer remissions. To achieve the latter objective our lab screened over 35,000 small molecules for their capacity to inhibit the viability of BTIC-enriched mouse tumor cells. Unexpectedly, several antagonists of the serotonin (5-hydroxytryptamine; 5-HT) transporter and select receptors were among the hit compounds identified in the screen. This thesis aims to establish a connection between serotonergic activity and BTIC. We accomplished the latter by assessing whether components of the 5-HT signaling system are expressed in mouse and human breast tumor cells and whether inhibition of their activity affects BTIC frequency using multiple orthogonal assays. Our data suggest that breast tumor cells of both mouse and human origin express the components necessary for 5-HT synthesis, activity and metabolism and that inhibition of these proteins with selective antagonists reduces the capacity of these cells to form tumorspheres. We demonstrate that highly selective antagonists of SERT and HTR5A target BTIC as established ex vivo cell transplantation assays. We also discovered that these agents synergize with chemotherapy in vivo to affect the growth of mouse breast tumor allografts and human breast tumor xenografts. To validate the molecular targets of these agents, we attempted to phenocopy their effects in functional assays by knocking out their respective genes using CRISPR-Cas9 technology. Collectively, this thesis contributes to an understanding of how 5-HT signaling affects BTIC and identifies serotonergic antagonists as novel anticancer agents.