Date of Award

2014

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Graduate Group

Pharmacology

First Advisor

Emer M. Smyth

Abstract

Cyclooxygenase-2 (COX-2) expression is associated with poor prognosis across a range of human cancers, including breast. While the contributions of tumor cell-derived COX-2 are well studied, those of the stroma remain ill-defined. Macrophages, an essential component of the tumor microenvironment, exist within a range of two polar phenotypes, influenced by signals in their local environment: anti-tumorigenic M1 or pro-tumorigenic M2. M2-like tumor-associated macrophages (TAM) are positively associated with tumorigenesis. This thesis investigates the contribution of macrophage COX-2 in two models of HER2/neu-induced mammary tumorigenesis utilizing mice selectively lacking macrophage COX-2 (COX-2MÃ?KO) and the contribution of COX-2 derived products in modifying macrophage phenotype in vitro. Finally, a targeted macrophage COX-2 inhibitor is investigated in vitro and in vivo as a potential cancer therapeutic. Deletion of macrophage COX-2 led to reduced mammary tumorigenesis coincident with fewer TAMs and reduction in M2 characteristics of TAM. Further, depletion of CD8+ cytotoxic T lymphocytes (CTLs), but not CD4+ T helper and regulatory cells, restored tumor growth in COX-2MÃ?KO mice, suggesting enhanced CTL function caused by reduction in total and M2-like TAM. Investigation of COX-2-mediated polarization of bone marrow-derived macrophages (BMDM) in vitro revealed paracrine influences of prostaglandin (PG) E2 in modifying polarized macrophage phenotype to more closely resemble TAM. Interestingly, interference with macrophage COX-2 did not significantly modify BMDM polarization. This suggested that autocrine COX-2 minimally affects BMDM phenotype, and that polarization of COX-2MÃ?KO BMDM does not recapitulate reduced M2 characteristics observed in COX-2MÃ?KO TAM. Reconstituted high-density lipoprotein (rHDL) nanoparticles were utilized as a method to target macrophages in vitro and in vivo. rHDL conjugated to fluorescent dye DiR revealed efficient incorporation of rHDL nanoparticles with TAM. In preliminary experiments utilizing rHDL-celecoxib as a targeted macrophage COX-2 inhibitor, marked suppression of PGD2 and PGE2 generation was evident in lipopolysaccharide (LPS)-stimulated J774A.1 cells. Importantly, urinary prostaglandin levels were not altered in mice treated with rHDL-celecoxib, suggesting no systemic inhibition of COX-2 with this targeted approach. These studies provide rationale for targeting macrophage COX-2 in mammary tumorigenesis and provide essential preliminary experiments in translating these findings into a potential chemopreventative or chemotherapeutic agent.

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