Prostate Cancer is the most frequently diagnosed malignancy and second leading cause of cancer mortality among American men. Maspin, a novel tumor suppressive serine protease inhibitor that is down-regulated in invasive prostate carcinoma, has been shown to inhibit prostate tumor invasion and metastasis. Emerging evidence also suggests that maspin may block tumor progression by sensitizing induced programmed cell death (apoptosis); thus, maspin may have a novel clinical application in enhancing apoptosis-based prostate cancer therapy. However, to explore this possibility, it is critical to understand the molecular mechanism of maspin in apoptosis.

My long-term goal is to investigate the maspin regulation of prostate epithelial cell apoptosis. In my preliminary study, I discovered that maspin protein overexpressed in prostate carcinoma cells, DU145 specifically, interacted with a molecule named heatshock protein-90 (Hsp90). Hsp90 has been shown to bind to numerous proteins involved in apoptosis regulation. I also found that when DU145 cells were treated with a potential apoptosis-inducing drug tumor necrosis factor-alpha (TNF-a), the interaction of maspin and Hsp90 was accompanied by one of these Hsp90-associated molecules.

Based on the existing evidence and literature, the goal of this application is to test a working hypothesis that maspin may sensitize TNF-a-induced apoptosis of DU145 cells by regulating Hsp90 and Hsp90-associated molecules both in the proliferative signaling pathway and in the caspase-mediated biochemical apoptotic cascade. I plan to focus on two specific objectives. Objective 1: To elucidate the specific molecular pathways regulated by maspin in TNF-a-induced apoptosis. I plan to carry out complementary cell biology, biochemical, and biophysical experiments to test two possibilities: (a) maspin may inhibit the activity of Hsp90-associated RIP (receptor-interacting protein) and block NF-kB (nuclear factor kB, a key transcription factor for cell proliferation)-mediated cell survival; and (b) maspin may facilitate the caspase (the major executing proteolytic enzymes in apoptosis)-mediated biochemical apoptotic cascade. In particular, I plan to test whether maspin regulates Hsp90/Apf1 (apoptotic protease-activating factor-1)-mediated caspase-9 pathway. The role of maspin will be evaluated by comparing DU145 cells and DU145-derived cells that re-express maspin. Objective 2: to date the exact biochemical activity of intracellular maspin is unclear. To elucidate the molecular mechanism underlying the sensitizing effect of maspin in prostate epithelial cell apoptosis, it is critical to investigate the structural and functional relationship of maspin; the wild-type maspin and three site-specific and domain-specific maspin mutants will be expressed by an adenovirus system in DU145 cells. The effect of maspin and maspin mutants will be analyzed in a similar manner as described in Objective 1.

The expected results of this study will provide (1) first hand knowledge regarding the specific apoptotic pathways regulated by maspin in TNF-a induced apoptosis, (2) molecular insights of critical sequence elements of maspin involved in apoptosis regulation, and (3) better understanding of the mode of maspin action. These results will lead to valuable experimental evidence for future application of maspin in apoptosis-based prostate cancer therapies.

In this project, Dr. Shijie Sheng will be my sponsoring supervisor. Dr. Sheng¿s laboratory is one of the most active research laboratories at Wayne State University (WSU) School of Medicine and the Barbara Ann Karmanos Cancer Institute (an NCI designated comprehensive cancer center) that offers an excellent training environment for basic and clinical cancer research. This project will allow me the growth as an independent research scientist with the expertise in apoptosis. After I complete my postdoctoral training, I wish to begin my independent career in prostate cancer research.