Susan G Komen  
I've Been Diagnosed With Breast Cancer Someone I Know Was Diagnosed Share Your Story Join Us And Stay Informed Donate To End Breast Cancer
    Home > Research & Grants > Grants Program > Research Grants > Research Grants Awarded > Abstract
    Awarded Grants
    A Novel Combinatorial Approach to Inhibit the Development of Mammary Cancer

    Scientific Abstract:
    Title: A novel combinatorial approach to inhibit the development of mammary cancer. ID Number: BCTR0402893 Ceramide is a major mediator of apoptosis induced by a wide variety of endogenous and exogenous agents including cancer chemotherapeutic drugs, radiation, hypoxia, just to mention a few. Cancer cells survive and grow by protecting themselves from ceramide-mediated apoptosis. We identified three targets P-glycoprotein (Pgp, product of mdr1), cyclooxygenase-2 (COX-2) and glucosylceramide synthase (GCS), which are often over-expressed in breast cancer cells and are uniquely positioned to decrease ceramide levels in a coordinated fashion. For example, Pgp decreases ceramide by flipping sphingomyelin from the inner leaflet of the plasma membrane to the outer leaflet thereby rendering it unavailable for neutral sphingomyelinase to generate ceramide. Induction of COX-2 is associated not only with up-regulation of Pgp but also with decreased arachidonic acid, which activates neutral sphingomyelinase. GCS depletes ceramide by converting it to glucosylceramide. Inhibition of these three uniquely over-expressed proteins should therefore increase the levels of free ceramide resulting in apoptosis/cell cycle arrest in tumor cells and inhibit tumor growth. The overall objective of this study therefore, is to design a combinatorial strategy of using inhibitors of Pgp, COX-2 and GCS to elevate the levels of free ceramide in mammary tumors and inhibit their development. Accordingly, 3 specific aims are proposed. In the first two specific aims, we will determine the inhibitory effects of PSC833 (PSC), an inhibitor of Pgp, Celecoxib (CXB), an inhibitor of COX-2 and D,L-threo-1-phenyl-2-decanoylamino-3-morpholino-1-propanol (PDMP), an inhibitor of GCS on mammary tumor development. We will use two models, a chemical model, wherein female Sprague-Dawley rats will be initiated with N-methyl-N-nitrosourea (MNU), and a transgenic model wherein mice carrying mouse mammary tumor virus-c-erbB-2 transgene develop spontaneously mammary tumors. These transgenic mice are particularly interesting because increased expression of erbB-2 is seen in many human breast cancers. Initiated rats and transgenic mice will be divided into several groups (30/group) and exposed to the inhibitors given either individually or in combinations of PDMP/PSC, PDMP/CXB and CXB/PSC. Animals will be palpated weekly for mammary tumors. In the MNU model the experiment will be terminated at 26 weeks. Because of the high cost of the transgenic mice ($55/mouse) only the combination that exerted highest inhibition in the rat model will be used in the mouse model. The mice will be euthanized at 32 weeks. At necropsy, the animals will be examined for tumors. The results of this study should indicate whether any combination of the inhibitors is better than any inhibitor given individually in inhibiting mammary tumor development. The parallel use of both the chemical induced and genetically manipulated mammary carcinogenesis models should be significantly informative. Specific aim 3 will determine the possible mechanism (s) by which PSC, CXB and PDMP inhibit the development of mammary tumors. For this purpose, rats will be initiated with MNU and exposed to the basal AIN 93G diet. 26 weeks later, at which time the rats exhibit mammary tumors, the inhibitors will be given intravenously either individually or in combinations as described above. After 24 and 48 hours, rats (5/ group) will be euthanized and mammary lesions and non-lesion mammary tissue will be processed for apoptosis and cell proliferation indices, Pgp, COX-2 and GCS and for the levels of ceramide, glucosylceramide, total sphingomyelin and sphingomyelin in the outer and in the inner leaflets of the membranes, arachidonic acid and neutral sphingomyelinase. Analysis of both the acute (Sp. Aim #3) and chronic (Sp. Aim #1) effects of the inhibitors on mammary tumors and non-lesion mammary tissue should provide a better understanding on the mechanism by which the combinatorial strategy inhibits mammary carcinogenesis. This novel combinatorial approach, we believe should be an effective strategy to inhibit the development and progression of mammary cancer. Further, this study will also open up a new hitherto unexplored roles for Pgp, COX-2 and GCS in the development and progression of mammary cancer.

    Lay Abstract:
    Title: A novel combinatorial approach to inhibit the development of mammary cancer. ID Number: BCTR0402893 Can ceramide hold the key for inhibiting breast cancer development? Ceramide, a normal cellular component is a major mediator of apoptosis (a type of cell death) induced by a wide variety of agents including cancer chemotherapeutic drugs, radiation, hypoxia, oxidative damage just to mention a few. Knowing how smart the cancer cells are, it is not surprising that they have developed mechanisms to survive and grow by protecting themselves from ceramide-mediated apoptosis. We identified three targets P-glycoprotein (Pgp, product of mdr1), cyclooxygenase-2 (COX-2) and glucosylceramide synthase (GCS), which are often over-expressed in breast cancer cells and are uniquely positioned to decrease ceramide levels in a coordinated fashion. If over-expression of Pgp, COX-2 and GCS is so intimately associated with the development of breast cancer, then inhibition of any one these targets should result in an inhibition of breast cancer development and a combinatorial strategy to inhibit all the three targets should only work better. The overall objective of the proposed study therefore, is to design a combinatorial strategy of using inhibitors of Pgp, COX-2 and GCS to elevate the levels of free ceramide in mammary tumors and inhibit their development. We have chosen three inhibitors and two experimental models to test the efficacy of the combinatorial strategy to inhibit mammary tumor development. The inhibitors are PSC833 (PSC), an inhibitor of Pgp, Celecoxib (CXB), an inhibitor of COX-2 and D,L-threo-1-phenyl-2-decanoylamino-3-morpholino-1-propanol (PDMP), an inhibitor of GCS. The experimental models are a chemical model, wherein female Sprague-Dawley rats will be initiated with N-methyl-N-nitrosourea (MNU) to develop mammary tumors, and a transgenic model wherein mice carrying mouse mammary tumor virus-c-erbB-2 transgene develop mammary tumors spontaneously. This transgenic model is particularly interesting because increased expression of erbB-2 is seen in many human breast cancers. The effect of the inhibitors given either individually or in combinations of PDMP/PSC, PDMP/CXB and CXB/PSC on mammary tumor development will be determined. The results of this study should indicate whether any combination of the inhibitors is better than any inhibitor given individually in inhibiting mammary tumor development. The parallel use of both the chemical induced and genetically manipulated mammary carcinogenesis models should be significantly informative. The combinatorial strategy has several advantages: For example, given the heterogeneity of cancers it is likely that not all mammary tumors equally over-express all the three targets (Pgp, COX-2 and GCS). The proposed combinatorial approach should take care of this un-predictability. One may ask why go through the combinatorial strategy. If ceramide is so important to kill cancer cells, why not administer ceramide directly. Unfortunately, there is no selectivity built into this approach and increased levels of ceramide may kill both the cancer as well as the normal cells! In the proposed combinatorial approach however, the chosen three targets (Pgp, COX-2 and GCS) are uniquely over-expressed in cancer cells and thereby introducing a degree of specificity in the killing. In addition, if GCS and Pgp are not inhibited the tumor cell can get rid of the exogenously administered ceramide – a situation similar to the fate of most cancer chemotherapeutic drugs. This potential problem is taken care in our combinatorial approach. Since inhibitors of Pgp and COX-2 are already in clinical trials, we feel confident that the translation of the results obtained in this study to a clinical setting should be rapid. Based on the uniqueness of the approach and exciting prospect, this novel combinatorial approach, we believe should be an effective strategy to inhibit the development and progression of breast cancer.