Dynamic changes in chemotactic and metabolic properties in tumor cells undergoing TGFb1-induced Epithelial-Mesenchymal Transition (EMT)

Epithelial-Mesenchymal Transition (EMT) is a process linked to several hallmarks of cancer including metabolic switching, inflammation, and chemotactic properties. Tumor cells that undergo EMT lose epithelial characteristics like cell-cell adhesion and apical basal polarity and gain expression of mesenchymal proteins including intermediate filaments and various cell surface receptors. As a result, they acquire invasive and metastatic properties. Inflammatory cytokines overexpressed in tumor microenvironment, like Transcription Growth Factor beta 1 (TGF-β1), can induce EMT and is linked to invasion and lymph metastasis in breast cancer. However, the mechanisms of how TGF-β1 promotes invasion and lymph metastasis are not clear.

The overall aim of this thesis was to study the role of TGF-β1-induced EMT in regulating metabolic activities and chemotactic properties of breast cancer cells. In Paper I, we investigated changes in glucose metabolism linked to TGF-β1-induced EMT. The results showed a link between reduction of the glucose transporter Glut1 and the anti-proliferative response of TGF-β1. They also showed that re-expression of Glut1 in breast cancer cells during chronic exposure to TGF-β1 allowed them to proliferate and develop a stable EMT on the same time. In Paper II, we demonstrate that TGF-β1-induced EMT is linked to translocation of the protein Syndecan-1 from the cell nucleus to the cell surface. In Paper III, we developed a new invasion method that enabled identification of chemotactic properties of breast cancer cells undergoing TGF-β1-induced EMT. The method involved using a cloning cylinder in the inner parts of the cell culture inserts, which eliminated problems with edge effects, and resulted in significantly improved reproducibility in the invasion assays. Using this method, we identified interleukin 7 (IL-7) and interleukin 15 (IL15) as novel chemotactic factors for EMT cells. In Paper IV, we studied whether TGF-β1 could promote lymph metastasis by promoting the production and secretion of factors that could function as chemo-attractants for breast cancer cells with EMT properties. Results from RNA-sequencing analysis showed the induction of an inflammatory cluster of genes in in lymphatic endothelial cells upon exposure to TGF-β1. Further analysis led to the identification of a number of pairs of chemokines induced by TGF-β1 in lymphatic endothelial cells and corresponding receptors specifically overexpressed in breast cancer cells with EMT properties.

In conclusion, the results presented in this thesis provide new insights into the capacity of TGF-β1-induced EMT to promote invasion of breast cancer cells by changing their metabolic activities and the intracellular localization of syndecan-1. The results also indicate that TGF-β1 plays dual roles in lymph metastasis by both promoting chemotactic properties of breast cancer cells and by inducing the expression of chemotactic factors in lymphatic endothelial cells. The results may lead to the development of new diagnostic tools and novel treatment strategies for cancer patients.

List of scientific papers

I. Different Regulation of Glut1 Expression and Glucose Uptake during the Induction and Chronic Stages of TGFβ1-Induced EMT in Breast Cancer Cells. Azadeh Nilchian*, Nikolina Giotopoulou*, Wenwen Sun and Jonas Fuxe. Biomolecules. 2020 Dec 1;10(12):1621. *These authors contributed equally to this paper.
https://doi.org/10.3390/biom10121621

II. Nuclear syndecan-1 regulates epithelial-mesenchymal plasticity in tumor cells. Ashish Kumar-Singh*, Malgorzata Maria Parniewska*, Nikolina Giotopoulou, Joman Javadi, Wenwen Sun, Tünde Szatmári, Katalin Dobra, Anders Hjerpe and Jonas Fuxe. Biology (Basel). 2021 Jun 11;10(6):521. *These authors contributed equally to this paper.
https://doi.org/10.3390/biology10060521

III. A cylinder-based invasion assay enables identification of chemotactic properties of breast cancer cells undergoing TGFβ1-Induced EMT. Nikolina Giotopoulou, Wenwen Sun and Jonas Fuxe. [Manuscript]

IV. TGFβ1-treated lymphatic endothelial cells secrete chemokines that promote invasion of breast cancer cells with EMT properties. Nikolina Giotopoulou, Malgorzata Maria Parniewska, Wenwen Sun and Jonas Fuxe. [Manuscript]