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CHEMICAL BIOLOGY APPROACHES FOR TRACKING AND MANIPULATION OF MACROPHAGE PHENOTYPES

Abstract
Macrophages are white blood cells of the innate immune system that have the ability to change phenotypically depending on the stimuli present in their surroundings through a process commonly referred to as polarization. Macrophage phenotypes broadly range from pro-inflammatory, anti-tumor (M1) to immune-suppressing (M2). Of particular interest to this work, breast cancer progression and metastasis rely on the presence of M2-like tumor-associated macrophages (TAMs). While many studies have shown the involvement of macrophages in tumor progression and metastasis, there remains a need to further explore these interactions and the polarization process, including tracking of macrophage subtypes. Toward this end, I have sought to develop more reliable and efficient polarization-probing tools for the study of macrophage phenotypes, including in complex environments. In this thesis, I used novel chemical biology methods to track and manipulate macrophage phenotypes, including in the interactions between macrophages and breast cancer. First, I used an innovative sensor array that tracks macrophage polarization phenotypes in a time-efficient, high-throughput manner. I also studied the effects of phenotypic changes and cancerous environments on cellular circadian rhythms via luminometry. I found that different macrophage polarization states displayed differences in their circadian rhythms and that conditioned media derived from aggressive breast cancers affected macrophages’ circadian rhythms to a greater extent. The potential of re-programming macrophages as treatment strategies for diseases has gained a lot of attention in the immunology field. In cancers, the use of small molecules to activate macrophages to inflammatory states can induce specific immune responses. At the same time, there are other diseases where the reduction of inflammation is sought. Toward these ends, I evaluated the abilities of small molecules to modulate macrophage phenotypes. Specifically, I investigated the potential of curcumin-derivatives to reduce inflammation in macrophages and a TLR-4 agonist to induce immune-stimulating, anti-cancer responses. To overcome some of the limitations in studying macrophage phenotypes in complex, multi-cellular environments, I generated a promoter-based reporter cell line that is indicative of macrophage polarization state through fluorescence. This model allows for tracking of macrophage phenotypes in multi-cellular experimental formats and, therefore, facilitates the study of macrophage responses in the context of diseases, like cancer. In summary, this study provides new ways to probe macrophage phenotypes in real time and denotes the necessity of using more representative models of the tumor microenvironment (TME), when studying the interactions between cancer and the immune system.
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