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A Role for the Tumor Microenvironment in Regulating Cytoskeletal Dynamics in the Setting of Breast Cancer Migration and Invasion

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2018, Doctor of Philosophy (PhD), University of Toledo, Biomedical Sciences (Cancer Biology).
The tumor microenvironment (TME) is a heterogeneous region that is comprised of tumor cells, stromal cells, and secreted factors and creates an environment favorable for tumor cell invasion and metastasis. An important step in the shift to a pro-cancerous microenvironment is the transformation of normal stromal fibroblasts to carcinoma-associated fibroblasts (CAFs). CAFs are present in a majority of solid tumors and can directly promote tumor cell motility via cytokine, chemokine and growth factor secretion into the TME, making them a critical TME component to understand. The exact effects that the TME has upon cytoskeletal regulation in motile tumor cells remain enigmatic. The conserved formin family of cytoskeleton regulating proteins plays essential roles in the assembly and/or bundling of unbranched actin filaments. Mammalian Diaphanous-related formin-2 (mDia2/DIAPH3/Drf3/Dia) assembles a dynamic F-actin cytoskeleton that underlies tumor cell migration and invasion. We previously showed that the chemokine CXCL12 can influence breast tumor cell migration and invasion through modification of the mDia2-directed actin cytoskeleton. The endogenous source of CXCL12 was unexplored. Therefore, the objective of this study was to understand whether CAF-derived chemokines from the TME impact breast tumor cell motility through modification of the formin-assembled F-actin cytoskeleton. In this study, we used CAF-derived conditioned media (CM) from WS19T fibroblasts, a transformed patient-derived tumor-adjacent breast CAF cell line, and studied its cell-autonomous impact upon tumor-cell motility. In triple-negative MDA-MB-231 human breast cancer cells, WS19T CAF-CM significantly and robustly increased wound closure and invasion relative to normal human mammary fibroblast (HMF)-CM. Unexpectedly, western blot analysis of WS19T-CM-treated MDA-MB-231 cells revealed a significant loss of mDia2 protein expression. WS19T-CM also promoted proteasome-mediated mDia2 degradation in MDA-MB-231s relative to control HMF-CM and WS21T CAF-CM, a breast CAF cell line that failed to reduced mDia2 expression, but significantly increased MDA-MB-231 migration. Cytokine array analysis of CM identified upregulated secreted factors in WS19T-CM relative to control WS21T CM, including the chemokine CXCL12 (SDF1¿). As CXCL12 was identified as a factor secreted by fibroblasts and heavily involved in cancer cell migration and invasion, we hypothesized that CXCL12 is a CM factor influencing mDia2 protein loss, while increasing MDA-MB-231 cell invasion and migration. Exogenous CXCL12 treatment resulted in increased wound closure and loss of mDia2 protein expression. Blocking CXCL12 signaling with AMD3100, a specific inhibitor of CXCR4, augmented motility and rescued mDia2 protein expression in the presence of WS19T-CM. Our data suggest a mechanism whereby CAFs promote tumor cell migration and invasion through CXCL12 secretion to regulate the mDia2-directed cytoskeleton in breast tumor cells, and highlights a novel therapeutic approach of treating highly invasive primary lesions through targeting the TME.
Kathryn Eisenmann (Committee Chair)
William Maltese (Committee Member)
Rafael Garcia-Mata (Committee Member)
Deborah Vestal (Committee Member)
Randall Ruch (Committee Member)
159 p.

Recommended Citations

Citations

  • Dvorak, K. (2018). A Role for the Tumor Microenvironment in Regulating Cytoskeletal Dynamics in the Setting of Breast Cancer Migration and Invasion [Doctoral dissertation, University of Toledo]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=mco1525433928091035

    APA Style (7th edition)

  • Dvorak, Kaitlyn. A Role for the Tumor Microenvironment in Regulating Cytoskeletal Dynamics in the Setting of Breast Cancer Migration and Invasion . 2018. University of Toledo, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=mco1525433928091035.

    MLA Style (8th edition)

  • Dvorak, Kaitlyn. "A Role for the Tumor Microenvironment in Regulating Cytoskeletal Dynamics in the Setting of Breast Cancer Migration and Invasion ." Doctoral dissertation, University of Toledo, 2018. http://rave.ohiolink.edu/etdc/view?acc_num=mco1525433928091035

    Chicago Manual of Style (17th edition)