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    Research Grants Awarded

    Targeted Therapy Of Chemoresistant And Metastatic Breast Cancer

    Grant Mechanism:
    Investigator Initiated Research

    Scientific Abstract:
    Rationale. Development of resistance to a wide variety of drugs and metastatic spread of cancer cells to distant organs pose major challenge in successful treatment of breast cancer. Virtually, all the roughly 40,000 annual breast cancer related deaths in the United States can be said to have occurred because the chemotherapy failed. Therefore, understanding the molecular factors responsible for drug resistance and metastasis of breast cancer is urgently needed to develop novel therapeutic approaches. On the basis of our strong preliminary data that breast cancer cells selected for resistance to drugs or isolated from metastatic sites express increased levels of TG2 and the observations that inhibition of TG2 by siRNA, ribozyme or antisense approach reversed drug resistance in breast cancer cells, we hypothesize that TG2 expression contributes to the development of these lethal phenotypes by constitutively activating cell survival and invasive signaling. Objective and hypothesis. The long-term goal of our research is to develop strategies for treating chemoresistant and metastatic tumors, which account for more than 90% cancer-related deaths. The objective of studies proposed in this grant application is to determine the significance of elevated TG2 expression in drug resistant and metastatic breast cancer cells. Based on our and others? published data, we believe that aberrant expression of TG2 in cancer cells promotes constitutive activation of anti-apoptotic and invasive signaling pathways and thus contributes to the development aggressive phenotype. We propose to study the cell survival signaling regulated by TG2. Using drug-resistant and metastatic breast cancer cell lines we also propose to determine the potential of TG2 as a therapeutic target for improving drug sensitivity and inhibiting metastatic growth of orthotopically growing breast tumors in a nude mouse model. We have recently developed a highly efficient in vivo method for delivery of siRNA by using neutral liposomes composed of 1,2-dioleyol-sn-glycero-3-phosphotidylcholine (DOPC). We will use these liposomes to deliver siRNA for silencing TG2 expression in tumors growing in mice. Specific aims. To address the involvement of TG2 in conferring drug resistance and metastatic phenotype on breast cancer cells, we propose the following specific aims. Aim 1, to determine whether expression of TG2 is necessary and/or sufficient to confer drug resistance and invasive functions. Aim 2, to investigate TG2 regulated signaling pathways that impact cell survival, cell migration and invasive functions. Aim 3, to determine whether expression of TG2 is necessary for in vivo growth, survival and spread of breast cancer disease in an orthotopic nude mouse model. It is our expectation that the results obtained under these aims will be critical for facilitating phase I study in patients with advanced breast cancer. Impact. As pointed out earlier, chemoresistance and metastasis are two major factors that account for majority of breast cancer related deaths in patients. One common feature between drug-resistant and metastatic cancer cells is that they both exhibit profound resistance to apoptosis. On the basis of our initial findings that chemoresistant and metastatic breast cancer cells express high basal levels of TG2, in conjunction with the observations that downregulation of TG2 rendered chemoresistant cells sensitive to drugs, we strongly feel that TG2 contributes to these phenotypes by promoting invasive and cell survival pathways. The proposed proof-of-concept studies involve validation of TG2 as therapeutic target for treating orthotopically growing drug-resistant and metastatic tumors in a nude mouse model. Validation of TG2 as therapeutic target for chemoresistant and metastatic tumors will facilitate our proposed phase I clinical trial with TG2 siRNA DOPC in patients with advanced tumors.

    Lay Abstract:
    Despite recent advances in early diagnosis, more than 40000 women still die of breast cancer every year in the USA. Two factors that impose major impediment in successful treatment of breast cancer include: 1) the dissemination and growth of cancer cells at distant sites (metastasis); and 2) their ability to develop resistance to anticancer therapies. According to a recent report (By Clifton Leaf in March 22, 2004 issue of the FORTUNE Magazine) more than 90% cancer-related deaths occur because the cancer has metastasized, yet > 92% grant applications submitted to major funding agencies, failed to even mention the word ?metastasis?. Therefore, it is imperative that more research efforts are geared to understand the intrinsic proteins and pathways that contribute to the development of drug resistance and metastasis in cancer cells. Precise understanding of these pathways is likely to yield immediate clinical benefit in terms of revealing novel/new targets for effective control and treatment of, at present hard to treat? metastatic and drug-resistant breast cancers. Our data have provided strong evidence documenting that progression of breast cancer cells to more aggressive phenotype is associated with an increased expression of the multifunctional protein, tissue transglutaminase (TG2). Importantly, downregulation of this protein by small-interfering RNA (siRNA), ribozyme or antisense RNA approach, diminished the invasiveness and restored sensitivity of drug-resistant breast cancer cells to chemotherapeutic drugs. Conversely, ectopic expression of TG2 in cancer cells protected them from drug-induced cell death and rendered them highly invasive. These results clearly suggest close association between TG2 expression progression of cancer cells to more aggressive phenotype. On the basis of these observations, we propose that TG2 expression contributes to the progression of breast cancer cells to more malignant drug-resistant and metastatic phenotypes. We believe that overexpression of TG2 can promote cell-survival, cell-growth, and invasive functions. Indeed, we have previously shown that high basal expression of TG2 in metastatic breast cancer cells leads to constitutive activation of cell survival and invasive functions. Therefore, understanding the pathways by which TG2 promotes these functions in chemoresistant and metastatic cancer cells should yield novel targets for paving the way to effective control and treatment of lethal forms of breast cancer. In view of these observations and our strong preliminary data suggesting that cancer cells selected for resistance to drugs or isolated from metastatic sites express elevated levels of TG2, we propose to investigate the significance of TG2 expression in conferring drug resistance and metastatic phenotypes. We will determine the signaling pathways that are regulated by TG2 and contributes to the development of these lethal phenotypes. Importantly, we propose to determine therapeutic potential of TG2 (in nude mouse model) for improving sensitivity of drug-resistant tumors to drugs that are commonly used for treating breast cancer. We also propose to determine the effect of TG2 inhibition on metastatic spread of breast tumors to distant sites in a nude mouse model. On the basis of our in vitro results and in vivo studies proposed this application, we plan to initiate a Phase I clinical trial with TG2 siRNA in breast cancer patients with advanced disease (patients with metastatic disease and who have failed the conventional chemotherapy).