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Reducing the Anti-apoptosis BRUCE Protein level Activates p53 Apoptosis Pathway and Sensitizes Breast Cancer Cells to Chemotherapy-induced Apoptosis
Tumor Cell Biology III
Background: BRUCE is an anti-apoptosis IAP protein and a ubiquitin-protein ligase. It has been clearly established that some human cancer cells such as SNB-78 human glioma cells possess an increased amount of BRUCE protein which contributes to apoptosis-resistance during chemotherapeutic drug treatment. Reducing BRUCE protein induces apoptosis and converts these cells from a chemo-resistant to a chemo-sensitive state. We recently discovered that reduction of the intracellular BRUCE protein level by siRNA triggers the stabilization of the tumor suppressor p53 protein, leading to p53-mediated apoptosis. This finding revealed a novel "BRUCE-p53-apoptosis pathway" associated with apoptosis in some cancer cells including MCF-7 breast cancer cells. Hypothesis: using modulators to reduce the levels of BRUCE and activate this novel BRUCE-p53-apoptosis pathway in combination with conventional chemotherapy could be a reasonable therapeutic strategy for killing MCF-7 human breast cancer cells in which BRUCE accounts for chemo-resistance. Specific Aims: (1) To investigate whether BRUCE directly ubiquitylates p53 protein for its degradation. (2) To identify and characterize BRUCE-interacting proteins that are BRUCE substrates or that regulate the BRUCE-p53-apoptosis pathway. (3) To investigate whether the BRUCE-p53-apoptosis pathway can sensitize MCF-7 breast cancer cells to chemotherapy-induced cell death. Study Design: In Aim 1, we will study whether BRUCE directly catalyzes p53 ubiquitylation. In Aim 2, we will use mass spectrometry to identify proteins that bind to BRUCE and regulate BRUCE-dependent p53-mediated apoptosis. We will also distinguish BRUCE substrates from regulators among those BRUCE-binding proteins and decipher their effect on modulating the BRUCE-p53-apoptosis in MCF-7 cells. In Aim 3, we will examine in MCF-7 cells whether directly altering BRUCE protein level or altering BRUCE's ability to interact with components of the pathway will enhance the killing efficacy of current chemo-treatments such as Etoposide. Potential Outcome and Benefits of the Research: The proposed studies will not only provide mechanistic insight in the roles of BRUCE in the control of breast cancer cell apoptosis, but will also provide therapeutic insight into new strategies for breast cancer intervention such as novel drug targets.
Background: Breast cancer is the most common form of cancer in women. Most chemotherapeutic drugs kill cancer cells by inducing apoptosis which is a type of cell suicide. BRUCE is an apoptosis-inhibitory protein that kills cells when its level is reduced or eliminated. But some breast cancer cells seem to be able to resist chemo-apoptosis by maintaining high levels of BRUCE protein. We have discovered a novel route to kill cancer cells: reducing the level of BRUCE protein can activate the tumor suppressor p53 which turns on apoptosis in breast cancer cells. Hypothesis: BRUCE contributes to apoptosis-resistance in breast cancer chemotherapy and reducing the level of BRUCE can sensitize apoptosis-resistant breast cancer cells to chemo-apoptosis and enhance chemotherapeutic efficacy. Specific Aims: (1) To characterize the functional relation between BRUCE and p53 in killing breast cancer cells. (2) To determine mechanisms that regulate the killing effects of the BRUCE-p53 pathway in breast cancer cells. (3) To evaluate the effect of the BRUCE-p53-apoptosis pathway in sensitizing chemotherapeutic drug treatments on breast cancer cells. Study Design: (1) BRUCE has an intrinsic activity to mark proteins for degradation. Whether p53 degradation is marked by BRUCE will be evaluated. (2) Other cellular components that bind to BRUCE and regulate the BRUCE-p53-apoptosis pathway will be identified and characterized from breast cancer cells. (3) Combined treatments of BRUCE reduction with chemotherapeutic drugs will be performed to evaluate if cell killing can be enhanced in apoptosis-resistant breast cancer cells. Potential Outcome and Benefits of the Research: This study will provide information on the regulation of the BRUCE-p53 killing pathway in breast cancer cells and insights into exploiting this killing pathway to accelerate breast cancer chemotherapy. Based on the findings of this study drugs may be designed to specifically decrease BRUCE protein or disrupt its interaction with p53 in breast cancer cells and this may result in a more effective therapeutic strategy to decrease the number of breast cancer deaths related to drug resistance.