TGF-betas in breast cancer progression

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In FY20, we have continued to address the role of TGF-beta in regulating the cancer stem cell (CSC) compartment using a novel functional imaging approach that we developed to allow visualization of this minority cell population in real time and in situ. Our lentiviral-based CSC reporter uses a synthetic promoter in which expression of a fluorescent protein is driven by the stem cell master transcription factors Oct4 and Sox2. Using this approach, we have developed methods that allow extended cell fate mapping of individual CSCs in vitro and in vivo, including an ongoing collaboration with the lab of Dr. John Condeelis at the Albert Einstein College of Medicine to perform intravital imaging of the CSC population in primary breast tumor and at the lung metastatic site. The intravital imaging approach has allowed us to demonstrate that the CSCs are slow-moving, invasive cells in the primary tumor that are preferentially associated with the microanatomical structure (TMEM) that drives intravasation of tumor cells into the bloodstream. Importantly, we have shown induction of a stem phenotype in non-stem tumor cells on contact with macrophages, via a Notch-dependent mechanism in vitro and in vivo. We have characterized changes in CSC representation across the entire metastatic process, demonstrating peak CSC numbers on early arrival at the metastatic site. W have shown that TGF-beta effects and TGF-beta signal transduction are different in stem vs. non-stem cells, and also that TGF-beta regulates the CSC compartment differently depending on whether it is functioning as tumor suppressor or pro-progression factor. These results have important implications for the ongoing clinical development of TGF-beta pathway antagonists. Studies to understand the detailed mechanisms regulating the size of the cancer stem cell population are ongoing. We have identified the chromatin-modifying enzyme PADI4 as having novel tumor suppressive activity on the CSC population through epigenetic regulation of the expression of master transcription factors of stemness. We have developed fate-mapping approaches to address the effects of TGF-beta on phenotypic plasticity and on self-renewing vs differentiating cell divisions. We are also integrating these analyses with genomic and single cell approaches to address underlying molecular mechanisms. Understanding how CSCs are regulated in vivo will be critical to development of more effective cancer therapies, as these cells are largely resistant to existing therapeutic approaches.