This proposal seeks to investigate the role of p53 abnormalities in the carcinogenesis and progression of CaP. The applicants found p53 alterations in 31% of 110 localized prostate cancers. Two key functions of the p53 gene are the control of the cell cycle and apoptosis. What now must be determined are the biological effects of different p53 mutations, such as those found by the applicants. The applicants will characterize the biological properties of the mutations they discovered. A yeast assay (FASAY) will be used to determine which p53 mutations are loss-of-function mutations. These will be tranfected into the p53-null SAOS-2 and murine 10(1) cell lines to identify gain of function activities as demonstrated by increased numbers of colonies in soft agar, transactivation of the MDR-1 promoter, and alteration of cell cycle progression as measured by flow cytometry. A subset of gain of function mutation will be analyzed in the p53-null prostate cancer cell line PC3. Whether dominant negative activity is present in these loss of function mutations will be tested in a modified yeast assay. If Class I mutations are found by this method, they will be transfected into the LNCaP cell line, and the response of the transfected cells to radiation will be determined by measuring alterations in cell survival, apoptosis and induction of p21. The applicants hypothesize that the gain of function and dominant negative mutations defined by these studies will play a role in the progression of CaP. The frequency of the different functional types of p53 mutations will be correlated with the stage and grade of CaP. Finally, the p53 functional data will be analyzed with respect to patient and tumor characteristics. The proposed studies are necessary to understand the role of p53 mutations in CaP carcinogenesis and progression. They also form the basis of future studies to investigate p53-based therapies.
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