Macrophage-Tumor Crosstalk in the Pathogenesis of Follicular Thyroid Cancer

Thyroid cancer is the most common endocrine malignancy and one of the fastest growing cancers in the United States. Follicular thyroid carcinoma (FTC) represents the second most common form of thyroid cancer diagnosed in the US and is most often tied to mutations in the RAS protein family of the MAP kinase pathway. In addition to driver mutations, FTC is characterized by a unique tumor microenvironment (TME) composed of cellular and non-cellular components that impact tumorigenesis and disease progression. Preliminary data from our lab has shown that CD45+ immune cells account for approximately 68% of all cells found in whole tumors collected from mouse models of RAS-driven disease. Macrophages account for the largest portion of known immune cell populations. Further experiments have demonstrated that tumor cell lines isolated from our RAS-driven models secrete cytokines known to impact the recruitment and activation state of cells of the myeloid lineage, particularly macrophages. However, it’s unclear what type of functional characteristics are induced by these secreted factors and how the resulting macrophage phenotype affects disease progression. Here, we sought to determine how bidirectional communication between macrophages and thyroid cancer cell lines could contribute to the development of a protumorigenic microenvironment. First, we began by defining how RAS-driven thyroid cancer cell lines affected the functional phenotype of previously unstimulated macrophages. Through gene expression analysis encompassing several markers of macrophage activation states, we determined that tumor cell-secreted factors induced the expression of multiple genes associated with tumor associated macrophages (TAM). In particular, we observed consistent upregulation of IL-10 and TNF-alpha, factors that have been associated with worsening disease. These results were further validated through quantification of protein secretion. In addition, we determined the role of activated macrophages in the progression of thyroid cancer, and specifically the effect of macrophage-secreted factors on tumor cell proliferation. Through direct and indirect assays of proliferation, we determined that factors secreted by classically-activated M1 macrophages inhibited cell proliferation. Surprisingly, secretions from alternatively-activated M2 macrophages reduced in vitro cell growth in some cell lines. Further analysis demonstrated that reduced cell proliferation was not associated with cell death, but rather was a result of delayed progression through the cell cycle. These results help to further define the macrophage phenotype within our model of FTC and will identify potential therapeutic targets to reduce the activity of protumorigenic cell populations.