Identifying the Signaling Pathways Downstream of the Serotonin Receptor 5A in Breast Cancer

Abstract

Breast cancer therapy resistance and disease recurrence are driven by an infrequent population of stem-like tumor cells, termed breast cancer stem cells or tumor-initiating cells (BTIC). Whereas drugs that target BTIC could be combined with conventional therapies to provide durable remissions, identifying such agents has been difficult. To achieve the latter, our lab screened more than 35,000 compounds for their capacity to reduce the activity of BTIC-enriched mouse mammary tumorspheres, wherein we identified numerous antagonists of multiple serotonin receptors (HTRs). The serotonergic antagonist that prevented sphere formation with the highest potency is a highly selective antagonist of HTR5A, SB-699551. We subsequently demonstrated that this agent affects BTIC activity in breast tumor cell lines representative of all clinical and molecular subtypes of breast cancer. Whereas the primary target of SB-699551 is known, the downstream signaling pathways responsible for its anti-BTIC effect remains enigmatic. The goal of this thesis work was to elucidate the signaling pathways downstream of HTR5A in human breast tumor cell lines. We used a phospho-proteomic approach to establish that treatment of human SB-699551 affects the phosphorylation of proteins involved in the Gi-coupled and the PI3K/AKT/mTOR signaling axes. Moreover, we demonstrated that selective antagonists of PI3K, AKT, and mTOR phenocopied the effect of SB-699551 in tumorsphere forming assays. Taken together, our data suggests that SB-699551 elicits its effect through the PI3K/AKT/mTOR signaling pathways downstream of HTR5A.