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The Role of TGF-ß Signaling in Mammary Tumor Metastasis
Transforming growth factor-beta (TGF-b) isoforms are growth factors that function physiologically to regulate development, cellular proliferation and immune responses. It is thought that TGF-ß functions as a growth inhibitor during early tumorigenesis, and later tumor cells become resistant to TGF-ß mediated growth inhibition and utilize TGF-ß signaling for tumor invasion and metastasis. We have shown that overexpression of TGF-b1 in mammary epithelium suppresses mammary tumor formation, while overexpression of a dominant-negative mutant form of the type II TGF-b receptor (DNIIR) in mammary epithelium results in lobular-alveolar hyperplasia and formation of mammary tumors after a long latency. Interestingly, the number of tumors showing evidence of invasion in bitransgenic animals co-expressing the DNIIR and TGF-a transgenes is significantly decreased when compared to the MMTV-TGF-a animals. To further elucidate the complex role of TGF-b signaling in mammary carcinogenesis, we developed several genetically modified mouse models. These unique models are powerful tools that allow us to precisely study the effects of impaired autocrine TGF-ß signaling specifically on the mammary epithelium and thus mammary tumor formation and metastasis.
Objective/Hypotheses: This proposal seeks to elucidate the complex role of TGF-ß signaling in mammary tumor metastasis. We hypothesize that autocrine TGF-ß signaling in the mammary gland functions to suppress mammary tumor formation and/or metastasis in the context of erbB-2/neu and PyVmT oncogene activation. We further hypothesize that tumor cells that are growth inhibited by TGF-ß and show no morphological change (epithelial to mesenchymal transition) upon TGF-ß treatment become more invasive and metastatic when TßRII expression is partially or completely abrogated. On the other hand, a partial or complete loss of TßRII expression will inhibit further tumor progression in tumor cells that are not growth inhibited by TGF-ß and exhibit a morphological change in response to TGF-ß. The following Specific Aims are designed to test these hypotheses:
Aim 1: Characterize spontaneous mammary tumor formation in mice co-expressing DNIIR and c-neu transgenes under the control of the MMTV promoter/enhancer (MMTV-neu/DNIIR). The tumor frequency, latency, degree of differentiation, invasiveness and metastatic spread will be determined. This model will provide partial loss of TßRII in the mammary epithelium in combination with stimulation of erbB/HER2 receptor signaling pathways, both of which are commonly found in human breast cancer. Aim 2: Characterize spontaneous mammary tumor formation in mice with a conditional knockout of TßRII in the mammary epithelium (Tgfbr2mgko) that have been crossed with MMTV-PyVmT or MMTV-c-neu transgenic mouse models. The characterization of the tumors will be the same as in Specific Aim 1. These models provide complete loss of TßRII expression in combination with stimulation of erbB/HER2 receptor and PyVmT signaling pathways. Aim 3: Determine the contribution of partial or complete loss of TßRII expression to growth inhibition, motility, conversion from an epithelial to mesenchymal morphology, and invasion on primary mammary tumor epithelial cell cultures from Specific Aims 1 and 2 and from previously established cultures from mammary tumors arising in MMTV-DNIIR and MMTV-TGF-a mice.
Potential Outcomes and Benefits of this Research: There is a great deal of evidence supporting both tumor suppressive and tumor permissive roles for TGF-ß signal transduction in the mammary gland. Recently, there have been several studies in different model systems indicating that blockade of TGF-ß signaling may inhibit formation of metastasis. The current model assumes that all tumor cells become refractory to TGF-ß’s growth inhibitory effects and that later in tumorigenesis all tumor cells will utilize TGF-ß signaling to promote tumor progression. However, as our data will illustrate, this may not be the case for all tumors, and if TGF-ß signaling is blocked by the use of TßRII antagonists while the tumor maintains its growth inhibitory response to TGF-ß this could prove to be extremely detrimental. These studies aim to restructure the current model and lead to the development of more targeted and effective therapies.
The Role of TGF-ß Signaling in Mammary Tumor Metastasis.
Growth factors are proteins that are involved in cell differentiation and growth. Growth factors are essential to the normal cell cycle, and are vital elements in the life of animals from conception to death. Among other things, they mediate fetal development, maintain and repair tissues, stimulate production of blood cells, and when misregulated they can participate in cancerous processes.
The mouse mammary gland is a dynamic organ, which unlike other organs, undergoes the majority of its development postnatally. Many different growth factors play key roles in the regulation of growth and development of the mammary gland. One growth factor that is critical to the normal development of the mammary gland is transforming growth factor-beta, or TGF-ß. The mammary gland is composed of an elaborate tree of ducts, which drain the lobes of the mammary gland and eventually lead to an opening at the nipple. Epithelial cells line the ducts of the mammary gland. These cells are the most common cell type to hyperproliferate and result in formation of mammary tumors.
TGF-ß activates many intracellular responses by binding to transmembrane receptors, designated type I (TßRI) and type II (TßRII). One cellular response to TGF-ß is growth inhibition. TGF-ß is able to arrest the growth of cells early in the cell cycle, and therefore tightly regulate cell proliferation and potentially have a tumor suppressive role. Normally, mammary epithelial cells are exquisitely sensitive to TGF-ß's growth inhibitory effects. However, many breast cancer cells eventually become resistant to TGF-ß-mediated growth arrest, and it is thought that they may utilize TGF-ß signaling for tumor invasion and metastasis. Metastasis is a complex process in which cancer cells migrate from their original site to other areas in the body and make a new home for themselves. It is metastasis of the breast cancer cells that most often leads to the death of the patient. TGF-ß is thought to play a key role in the process, however the exact role is not clearly understood.
We are investigating the role of TGF-ß in mammary tumor formation and metastasis. We have generated several transgenic mouse models. Transgenic mice are mice that are engineered to carry a foreign gene, or transgene, as part of its own genetic material. We are utilizing transgenic mouse models that overexpress genes know to be important in development breast cancer, and we are breeding these mice to mice that we have engineered to have partial or complete abrogation of TGF-ß signaling in the mammary epithelium.
While other model systems have affected not only the mammary epithelium but also the activity of the immune system, blood cells and many other systems, our unique models allow us to precisely study the effects of impaired TGF-ß signaling specifically on the mammary epithelium and thus mammary tumor formation and metastasis. This proposal seeks to elucidate TGF-ß’s complex role in mammary tumor formation and metastasis using innovative mouse models and assays. Novel information gained from these studies about the involvement of TGF-ß in mammary tumor formation and metastasis may lead to the discovery of new therapeutic targets and development of safer therapies.