Date of Award

5-1-2012

Degree Name

Doctor of Philosophy

Department

Molecular Biology, Microbiology and Biochemistry

First Advisor

Watabe, Kounosuke

Abstract

Notch signaling is often and aberrantly activated by hypoxia during tumor progression; however, the exact pathological role of hypoxia-induced Notch signaling in tumor metastasis is as yet poorly understood. In the first part of this study, we aimed to define the mechanism of Notch ligand activation by hypoxia in both primary tumor and bone stromal cells in the metastatic niche and to clarify their roles in tumor progression. We have analyzed the expression profiles of various Notch liagnds in 779 breast cancer patients in GEO database and found that the expression of Jagged2 among all five ligands is most significantly correlated with the overall- and metastasis-free survival of breast cancer patients. The results of our immunohistochemical (IHC) analysis for Jagged2 in 61 clinical samples also revealed that both Jagged2 and Notch signaling were strongly up-regulated at the hypoxic invasive front. Activation of Jagged2 by hypoxia in tumor cells induced EMT and also promoted cell survival in vitro. Notably, a ã-secretase inhibitor significantly blocked Notch-mediated invasion and survival under hypoxia by promoting expression of E-cadherin and inhibiting Akt phosphorylation. Importantly, Jagged2 was also found to be up-regulated in bone marrow stroma under hypoxia and promoted the growth of cancer stem-like cells by activating their Notch signaling. Therefore, hypoxia-induced Jagged2 activation in both tumor invasive front and normal bone stroma plays a critical role in tumor progression and metastasis, and Jagged2 is considered to be a valuable prognostic marker and may serve as a novel therapeutic target for metastatic breast cancer. In the second part of this study, the role of Notch signaling in brain metastasis was investigated. Metastatic diseases are responsible for the majority of the deaths in breast cancer patients and the brain is one of the most common metastatic sites. The metastatic tumor in the brain profoundly affects the cognitive and sensory functions as well as morbidity of patients, and the one year survival rate among these patients remains less than 20%. However, the pathological mechanism of brain metastasis is as yet poorly understood. In this report, we found that metastatic breast tumor cells in the brain highly expressed IL-1â which can "activate" astrocytes. This activation significantly augmented the expression of JAG1 in the reactive astrocytes, which in turn activated Notch signaling pathway of cancer stem-like cells (CSCs) upon direct interaction. We also found that the activated Notch signaling in CSCs up-regulated Sox2 followed by promoting self-renewal of CSCs. Furthermore, we have shown that the blood-brain barrier permeable Notch inhibitor, Compound E, can significantly suppress the brain metastasis growth in our animal model. These results represent a novel paradigm for the understanding of how metastatic breast CSCs re-establish their niche for their self-renewal in a totally different microenvironment, which opens a new avenue to identify a novel and specific target for the brain metastatic disease

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