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Exploring functional mechanisms underlying tumor initiation, EMT and cell migration

Gruber, Dorothea Cornelia. Exploring functional mechanisms underlying tumor initiation, EMT and cell migration. 2012, PhD Thesis, University of Basel, Faculty of Science.

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Official URL: http://edoc.unibas.ch/diss/DissB_10210

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Abstract

Cancer is one of the leading causes of death. A primary tumor forms when cells start to proliferate in an uncontrolled way and stop reacting to restraining signals. Tumors that reach a critical volume induce angiogenesis, a vascular remodeling process that provides nutrients and oxygen to the degenerated cell mass. Upon further tumor progression to a malignant cancer, cells acquire the ability to invade the surrounding tissue. In order to do so, formerly epithelial tumor cells undergo an epithelial to mesenchymal transition (EMT). This process of cell-cell detachment, breaching through the basement membrane and gaining migratory capabilities is the first step of the metastatic cascade. Metastasis is a process that allows tumor cells to leave the primary lesion and disseminate via the vascular system to secondary sites. Metastases, as the fatal feature of cancer, lead in most of the cases to patients’ death.
Furthermore, metastasis, as the end stage of malignant disease, until now is incurable. This is because metastases are spread systemically throughout the body. Moreover, they even can establish from a disseminated tumor cell months after the primary tumor has already been surgically removed. Another barely controllable feature of cancer is tumor relapse. Even after a thorough surgery with the aim to completely remove the primary tumor and eventual draining lymph nodes combined with chemotherapy, patients relapse. Resistance to chemotherapy and establishment of metastases has both been accounted for to the abundance of cancer stem cells (CSC) within the tumor mass. These CSCs are endowed with the ability to evade chemotherapeutic drugs, they are mesenchymal in nature, migratory and invasive, and, most importantly, they are able to establish cancer and metastasis de novo.
The work of my thesis has been dedicated to investigate the process of metastasis and the function of cancer initiation. Tumor initiation has recently been associated with EMT. To understand the functional circumstances how EMT cells gain the ability to form tumors, I used cellular murine breast cancer models. These model systems allowed me to study cells’ behavior before and after EMT in vitro and in vivo. In vitro experiments validated the observation of others that EMT cells indeed resemble cancer stem cells by being able to form hollow spheres and being susceptible to the CSC-specific drug salinomycin. In vivo studies revealed that EMT cells initiate tumors with a much earlier onset and with a higher efficiency when limited amounts of cells were injected orthotopically into mice compared to their epithelial counterpart. Moreover, EMT cell-generated cancers are highly vascularized already in the early phase of tumor establishment. Knockdown studies of the main pro-angiogenic factor VEGF-A revealed that it is required for early tumor onset. Thus, tumor angiogenesis is not only an effect of early and fast EMT-tumor progression. Further supporting this notion are limiting dilution experiments, which suggest that tumor initiation in EMT cells is a multifactorial event. This concept was validated by the observation that 10 EMT cells could initiate a tumor, whereas VEGF‑A knockdown cells could not. Hence, EMT-induced tumor initiation is achieved by the ability of promoting angiogenesis.
EMT can be induced by the cytokine TGFβ. Normally, cells that experience TGFβ signaling become quiescent or die due to induction of apoptosis. Cancer cells can overcome these effects and react to TGFβ by undergoing EMT. As part of my thesis, I could show that the transcription factor Dlx2 is an important switch that allows cells to react to TGFβ by undergoing EMT without facing apoptosis induction. Dlx2 exerts its anti-apoptotic, pro-survival function by directly reducing TGFβRI expression and inducing the expression of the epidermal growth factor receptor ligand betacellulin.
Another feature of EMT is the gain of cell motility. Cell migration is extremely important for cancer cells in order to leave their primary site and to disseminate. I have found ephrinB2 expression upregulated during EMT. EphrinB2 is a member of the Eph-ephrin signaling network that is known to be crucial for cell-cell communication. Thereby, cells that do not belong to the same entity repulse each other, restricting intermingling of tissue. Furthermore, ephrinB2 is required for neuronal axon guidance and angiogenesis. In my thesis, I showed that EMT cells need ephrinB2 to efficiently migrate. As an explanation for this phenotype I described that a knockdown for ephrinB2 led to an over-stabilization of focal adhesions. Cells normally use focal adhesions to hold on to an extracellular matrix (ECM) surface. Over-stabilization of focal adhesions attaches cells too firmly to the ECM and, hence, cells cannot retract their rear-end anymore which decreases cell motility.
In summary, I succeeded in gathering further insights into tumor initiation, EMT, and with this, the metastatic process.
Advisors:Christofori, Gerhard
Committee Members:Affolter, Markus
Faculties and Departments:05 Faculty of Science > Departement Biozentrum > Growth & Development > Cell Biology (Affolter)
Item Type:Thesis
Thesis no:10210
Bibsysno:Link to catalogue
Number of Pages:141 S.
Language:English
Identification Number:
Last Modified:30 Jun 2016 10:51
Deposited On:18 Jan 2013 08:07

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