The Role of Crk Adaptor Proteins in Breast Tumorigenesis and Bone Metastasis

Principal Investigator's Career Goals: I aim to obtain my Ph.D. from McGill University before obtaining a postdoctoral position in the United States to continue my training in the field of breast cancer research. After my postdoctoral studies, I want to become an independent investigator working on development of targeted therapies for the treatment of breast cancer. My current Ph.D. project is a great first step towards reaching these goals. My project will lead me to a deep understanding of the mechanisms and cellular signaling pathways that are important in basal breast cancer progression. A solid foundation in understanding breast cancer at the molecular level will allow me to be a much more effective researcher later on in my career. Through my current project, I also have the opportunity to meet top researchers in the field of breast cancer, which will be very useful for setting up collaborations and sharing information as my career progresses. I am also lucky to be conducting my Ph.D. in a supportive and technologically advanced setting. My project will allow me to gain a lot of valuable knowledge about breast cancer and to learn a wide variety of techniques used in this field.

Project Proposal: The basal subtype of human breast cancer, which accounts for ~20% of breast cancer cases, is currently the most challenging form of the disease to treat, because cancer cells of this type do not express the standard therapeutic targets present in other subtypes. As a result, basal breast cancer has no form of personalized therapeutic intervention and is frequently associated with poor patient outcome. A clearer understanding of the molecular changes that are important to the development and metastasis of basal tumors will allow design of alternative therapeutic strategies for the treatment of patients with this disease. Metastasis is a leading cause of morbidity and mortality in breast cancer, and so is a main focus of this project.

I study the Crk adaptor protein family (CrkI, CrkII, and CrkL), which mediates the formation of protein complexes involved in transmitting cellular signals. Expression of Crk proteins has been shown to be increased in breast cancer. Interestingly, Crk proteins are key regulators of signaling pathways downstream of proteins known to drive the progression of breast cancer. Crk proteins have also been shown to be important in the migration and invasiveness of cells in culture, which are key processes contributing to metastasis. We have been working with a basal breast cancer cell line, MDA-231 1833TR, with an affinity to metastasize to bone, and we have generated knockdown of all three Crk proteins in these cells. This has allowed us to study the importance of Crk proteins in tumorigenesis and metastasis of basal breast cancer cells. Our preliminary work has shown that loss of Crk proteins decreases metastasis to bone in mice. Loss of Crk also decreases tumor growth of cells injected into the mouse mammary fat pad. These findings indicate that if the mechanistic role of Crk in these processes was better defined, then therapies targeting Crk could be effective in preventing both primary tumor growth and metastasis in basal breast cancer.

The initial stages of my project will be spent confirming my preliminary findings in other basal breast cancer cell lines. This is important in order to prove that the effects observed were due to an important role for Crk, and not just a phenomenon present in the cell line studied. After confirming our preliminary findings, we would like to find out which Crk protein is involved in metastasis and which is involved in tumorigenesis. This will be done by reintroducing each protein individually back into the MDA-231 knockdown cells in order to see which proteins can rescue the ability of the cells to metastasize and form tumors. When we know which protein(s) is/are important, we will determine which region of each protein is important. This involves reintroducing Crk proteins missing certain regions into the knockdown cells and studying metastasis and tumorigenesis. We will then know the regions of Crk protein that regulate signaling pathways involved in metastasis and tumorigenesis.

In order for our knowledge of signaling to translate into therapeutic benefit, the exact mechanism and protein interactions that are important in basal breast cancer progression must be determined. Our lab has a system in place called the tandem affinity purification (TAP) method, which we will use to purify and identify proteins that interact with the regions of Crk that regulate metastasis and tumorigenesis. This allows identification of key signaling events at the molecular level that lead to progression of basal breast cancer.

Clinical Applicability: Identification of the key protein-protein interactions that are necessary for tumor growth and metastasis of basal breast cancer will open the door to the development of new targeted therapies aimed at disrupting these interactions. We hope that this study can lead to new targeted therapies that will improve quality of life and clinical outcome of patients with basal breast cancer. Although the direct effects on patients will not occur until years after the completion of this study, we will contribute to the understanding of the signaling pathways that are important in basal breast cancer, which will further progress the field and benefit researchers working in both clinical and basic sciences.