Trogocytosis and Dickkopf-1 Contribute To Triple Negative Breast Cancer Development and Progression

Abstract

The transformation, differentiation, and survival of tumors and their subsequent responsesto treatments are dependent on aberrant signaling of driver mutations and interactions within the tumor microenvironment. Triple negative breast cancers are said to be the most aggressive subtype of breast cancers. This is partially attributed to the lack of viable therapeutic targets. Current treatment of triple negative breast cancers relies on the use of nonspecific DNA-damaging agents. Thus, it is important to elucidate the mechanisms triple negative breast cancers use in their development and progression. This dissertation outlines two novel mechanisms found in triple negative breast cancers. The first is the contact-dependent cell-cell interaction trogocytosis, which is the transfer or swap of membrane and membrane proteins between two cells. I discovered that trogocytosis between triple negative breast cancer cells and lymphocytes results in the expression of lymphocyte-specific proteins. I then investigated the transfer of CD45, a leukocyte protein expressed on all immune cells, and found that the transfer of CD45 also increased tumor cell expression of PTPRC, the gene encoding CD45. Visualization of CD45 transfer resulted in the discovery of trogosomes, a spherical subcellular structure containing T cell membrane fragments, CD45 protein, and genomic DNA. Through trogocytosis, the acquisition of lymphocyte proteins sheds light on a novel mechanism by which triple negative breast cancer cells can utilize to dampen the immune response and promote genomic instability. The second mechanism characterizes the role of DKK1 in promoting genomic instability and chemoresistance. DKK1 is canonically known as an inhibitor of the Wnt/β-catenin transcriptional activity. However, DKK1 is often seen overexpressed in triple negative breast cancers despite elevated β-catenin transcriptional activity. I validated a candidate nuclear localization signal within the DKK1 amino acid sequence using immunoprecipitation mass spectrometry and coimmunoprecipitation western blotting. I then utilized the Importin β inhibitor Importazole to inhibit nuclear entry by DKK1. Knockdown of DKK1 resulted in the downregulation of nuclear DKK1 target ALDH1A1. Overexpression of DKK1 revealed reduced γH2AX foci formation and indicates a role of DKK1 in DNA repair. Analysis of patient samples showed that the incidence of nuclear DKK1 was highest in triple negative breast cancers compared to other subtypes of breast cancer. Further analysis revealed the expression of nuclear DKK1 after treatment with KEYNOTE-522 in case matched index and residual tumors of triple negative breast cancer samples. Taken together, both trogocytosis and DKK1 contribute to the role of development and progression in triple negative breast cancers by allowing tumor cells to further develop within their respective microenvironments through decreasing antitumor immunity and by promoting genomic instability.