Cell-Killing Effect of Orphan Receptor TR3 in Breast Cancer

Institution: The Burnham Institute for Medical Research
Investigator(s): Nathalie Bruey-Sedano, Ph.D. -
Award Cycle: 2002 (Cycle VIII) Grant #: 8FB-0089 Award: $84,643
Award Type: Postdoctoral Fellowship
Research Priorities
Detection, Prognosis and Treatment>Innovative Treatment Modalities: search for a cure
Biology of the Breast Cell>Pathogenesis: understanding the disease



Initial Award Abstract (2002)
Vitamin A compounds, collectively called retinoids, show promise as anti-cancer agents for breast cancer prevention and treatment. However, clinical effects of classical retinoids are limited to hormone-dependent breast cancer. Recent studies have indicated that a retinoid derivative, called AHPN, is a promising agent against both hormone-dependent and independent breast cancer by causing their death (apoptosis). Dr. Xiao-kun Zhang's lab at the Burnham Institute recently discovered that the death-inducing effect of AHPN is mediated by a protein called TR3. TR3 was conventionally considered to function in the nucleus as a factor that controls expression of genes. Dr. Zhang's lab found that TR3 migrates from the nucleus to an intracellular organelle called mitochondria to exert killing activity; however, how TR3 migration is controlled is not known, and it is very important to know this in order to develop agents to induce TR3 killing activity to destroy breast cancer cells.

Previously several laboratories showed that TR3 can interact with another protein called RXR. Dr. Zhang's lab recently found that RXR and its regulators (a class of specific retinoids) may play a role in the control of TR3 migration and breast cancer killing activity. However, it is very difficult to show whether TR3 activity is regulated by RXR or by some other factors in breast cancer cells due to the complexity of human cells. Fortunately, scientists have found a much simpler system, the fruit fly Drosophila melanogaster, to study function and regulation of human proteins.

We propose to take a genetic approach to examine the interactions between TR3, RXR, and other components of the cell death pathways by using the fruit fly as a model. We hypothesize that the mechanism of TR3 control and killing activities are very similar in flies and in humans.

We plan to use the fruit fly to identify modulators of TR3 killing activity, including fly RXR. Drosophila melanogaster is a good model to use because genetic modification is relatively easy. Certain genes can be either "knocked out" or inserted, resulting in an altered fly genotype that is reflected by changes in visible characteristics and behaviors (phenotypes). Molecular cloning techniques will be used to make these genotypes, and histology techniques will be used to study the phenotype of each of the experimental genotype fly lines. The regulators identified in the fly will be further studied in human breast cancer cells.

Because RXR and TR3 compounds possess anti-cancer properties in breast cancer cells irrespective of hormone status, it is envisaged that these agents may be effective chemotherapeutic agents for breast cancer treatment. However, in order to identify a new generation of such compounds, it is essential to understand their mechanism of action. Our previous discovery that transcriptional factor TR3 migrates from the nucleus to mitochondria to induce killing activity is exciting. Our hypothesis that TR3 killing activities is regulated by RXR and its ligands is novel, and the proposed study of using fly system is also novel in nature.


Final Report (2006)
In the United States breast cancer is the most common cancer among women, despite significant progress in early detection and diagnosis during the last decade. New forms of therapies are urgently needed to treat breast cancer patients whose tumor does not respond to traditional therapy, such as surgery, radiation, chemotherapy and hormonal therapy. Combination therapy that enhances the anti-cancer activity of chemotherapeutic agents and minimizes some of their undesirable side effects represents one of the new approaches. However, the selection of optimal affect the therapeutic responses of chemotherapeutics agents.

Recent studies from our laboratory have demonstrated that ("orphan") receptor TR3 plays a critical role in mediating the apoptotic effect of various stimuli, including certain chemotherapeutic agents. During the two years of research funded by the BCRP, I have examined whether retinoids, a class of vitamin A derivatives, can sensitize breast cancer cells to the growth inhibitory effects of a variety of chemotherapeutics agents, based on the fact that TR3 can combine (heterodimerize) with RXR, a receptor for certain vitamin A derivatives. Our results demonstrate that combination of 9-cis-Retinoic Acid (9-cis-RA), a natural ligand of RXR, with certain chemotherapeutic agents including taxol, etoposide, (VP-16) and cis-platinum caused apoptosis, a programmed cell death, in breast cancer cells to a greater extent than when either the chemotherpeutic agent or the Retinoic acid was used alone. This synergistic effect on inducing cell death was observed in both hormone-dependent and -independent breast cancer cells. Furthermore, extensivecell death (apoptosis) of several commonly used breast cancer cell lines (MDA-MB231 and ZR-75-1) required a much shorter exposure time to the combination treatment compared to the treatment by individual agent alone.

In studying the molecular mechanism by which the combination treatment induces breast cancer cell death, we used microarray technology, which allows us to monitor possible change in expression of thousands of genes simultaneously, in response to the combination treatment. Our results show that expression of several genes related to cell death was altered in response to the combination treatment. The significance of these genes in mediating breast cancer cell death is currently under investigation.

To further understand the TR3-mediated apoptotic pathways, we have developed an AHPN antagonist 3-A-AHPC in collaboration with Dr Marcia Dawson s group. Our result demonstrated that 3-A-AHPC could inhibit apoptosis by suppressing AHPN-induced TR3 expression. Thus, 3-A-AHPC represents an important tool to study the TR3-mediated apoptotic pathway in cancer cells. Together, our research demonstrate that retinoids can act to sensibilize breast cancer cells to treatments by chemotherapeutic agents, suggesting a possible new approach for treating breast cancer.


Progress Report 1 (2003)
In the United States breast cancer is the most common cancer among women, despite significant progress in early detection and diagnosis during the last decade. New forms of therapies are urgently needed to treat breast cancer patients whose tumor does not respond to traditional therapy, such as surgery, radiation, chemotherapy and hormonal therapy. Combination therapy that enhances the anti-cancer activity of chemotherapeutic agents and minimizes some of their undesirable side effects represents one of the new approaches. However, the selection of optimal combinations is largely depended on our understanding of the signal transduction pathways (the means by which cellular instructions are carried from the surface of a cell to its nucleus) that mediate therapeutic responses of chemotherapeutics agents.

Recent studies from our laboratory have demonstrated that orphan receptor TR3 plays a critical role in mediating an apoptotic (cell death) effect of various stimuli, including certain chemotherapeutic agents. In the first year of research funded by the BCRP, I have examined whether retinoids, a class of vitamin A derivatives, can sensitize breast cancer cells to the growth inhibitory effects of a variety of chemotherapeutics agents, based on the fact that TR3 can heterodimerize with RXR, a receptor for certain vitamin A derivatives.

Our results show that combination of 9-cis-Retinoic Acid (9-cis–RA), a natural derivative of vitamin A, with certain chemotherapeutic agents including taxol, etoposide, (VP-16) and cis-platinum synergistically induced apoptosis, a programmed cell death, in breast cancer cells. The synergistic effect on inducing cell death was observed in both hormone-dependent and –independent breast cancer cells. Furthermore, extensive apoptosis of MDA-MB231 and ZR-75-1 breast cancer cells required much shorter exposure of cells to the combination treatment compared to the treatment by individual agent alone.

In studying the molecular mechanism by which the combination treatment induces breast cancer cell death, we used microarray technology, which allows us to monitor possible change in expression of thousands of genes simultaneously, in response to the combination treatment. Our results show that expression of several genes related to cell death was altered in response to the combination treatment. The significance of these genes in mediating breast cancer cell death is currently under investigation. To further understand the TR3-mediated pathways that induce apoptosis, I have evaluated an AHPN antagonist SR11461 developed by Dr Marcia Dawson’s group. My result demonstrated that SR11461 could inhibit apoptosis by suppressing TR3 expression. Thus, SR11461 represents an important tool to study the TR3-mediated apoptotic pathway in cancer cells. Based on these exciting findings, I plan to further characterize the synergistic effect of the combination, with respect to their mechanism of action and activities in animal. Together, our research demonstrates that retinoids can act to sensitize breast cancer cells to treatments by chemotherapeutic agents, suggesting a possible new approach for treating breast cancer.


Symposium Abstract (2003)
In the United States breast cancer is the most common cancer among women, despite significant progress in early detection and diagnosis during the last decade. New forms of therapies are urgently needed to treat breast cancer patients whose tumor does not respond to traditional therapy, such as surgery, radiation, chemotherapy and hormonal therapy. Combination therapy that enhances the anti-cancer activity of chemotherapeutic agents and minimizes some of their undesirable side effects represents one of the new approaches. Epidemiological in vitro and in vivo studies have demonstrated that retinoids, vitamin A and its derivative are promising agents for cancer treatment. We have investigated the effect of combination of retinoids and other chemotherapeutic agents, including taxol, on the growth of breast cancer cells. Our results demonstrate that combination of 9-cis-retinoic acid (9-cis-RA), a natural derivative of vitamin A, with other agents currently used in the treatment of breast cancer synergistically induced apoptosis, a programmed cell death, in breast cancer cells. We measured this effect by analyzing specific markers of cell-death program (also called apoptosis) such as changes in cell shape, breakdown of DNA and activation of proteases. The synergistic effect on inducing cell death was observed in both hormone-dependent and –independent breast cancer cells. Furthermore, extensive apoptosis of MDA-MB231 and ZR-751 cells required much shorter exposure of cells to the combination treatment compared to the treatment by individual agent alone.

In studying the molecular mechanism by which the combination treatment induces breast cancer cell death, we used Microarray technology, which allows us to monitor possible change in expression of thousands of genes simultaneously in response to the combination treatment. Our results show that expression of several genes related to cell death was altered in response to the combination treatment. The significance of these genes in mediating breast cancer cell death is currently under investigation. Together, our research demonstrate that retinoids can act to sensitize breast cancer cells to treatments by chemotherapeutic agents, suggesting a possible new approach for treating breast cancer.