Although cadmium (Cd) is a proven carcinogen in rodents and may be a """"""""sufficient"""""""" carcinogen in men, the mechanism of Cd-induced carcinogenesis remains unknown. Additionally, while Cd is cytotoxic to a wide spectrum of tissues in the body, its carcinogenicity is limited to the prostate, testes and lungs. It has been postulated that the expression of the metal- binding protein, metallothionein (MT), plays a role in determining tissue susceptibility to Cd toxicity and carcinogenicity. We, and others, have shown that rat ventral prostate (VP) and testes, both target organs of Cd carcinogenesis, lack MT expression.
The aim of this proposal is to determine whether MT plays a protective role in Cd carcinogenesis. We hypothesize that """"""""induced"""""""" expression of MT in rat VP would render the gland resistant to Cd carcinogenicity. Using a transgenic animal approach, we shall attempt to induce MT expression in rat VP. A chimeric gene will be synthesized by fusing the rat probasin (PB) promotor and 5'-flanking region to a mouse MT coding region cDNA (PB-MT construct). The PB gene 5'- flanking region contains two androgen responsive elements and a sequence that targets a transgene expression specifically to the prostate of a transgenic animal. The expression of PB-MT will he induced in an androgen- receptor positive PC-3-cloned prostatic cancer cell line (AR+ PC-3) transfected with the chimeric gene and, later, in transgenic animals carrying the construct by androgen activation. Alterations in cellular or tissue susceptibility to Cd carcinogenicity after induction of MT expression will he assessed by monitoring a set of """"""""early or intermediate"""""""" biomarkers. Specifically, these biomarkers are: 1) cytotoxicity, as measured by the expression of a cell death-associated gene (TRPM-2) and """"""""internucleosomal DNA fragmentation"""""""", 2) proliferation, as quantitated in situ by levels of histone-3 transcripts and proliferation cell nuclear antigen (PCNA), 3) genotoxicity as assessed by the degree of nuclear DNA strand breaks and DNA-protein formation, and 4) epithelial atypical hyperplasia, as a morphological marker of neoplastic development. To measure tumorigenicity per se, tumor incidence will he enumerated histologically and malignancy of the primary tumors will be assessed by anchorage-independent growth, cellular ploidy and in vivo tumor formation efficiency in isogeneic hosts. Should our theory be proven, we expect to observe either abolition or delay in development of the early or intermediate biomarkers of carcinogenesis, reduction of tumor incidence, and/or reduced malignancy of the primary tumors formed in the VPs of transgenic rats expressing MT, as compared to those found in normal VPs. Data from these experiments are expected to yield important information concerning the role of MT in Cd tumorigenesis. They should illuminate the mechanisms of prostatic carcinogenesis and decipher the causes of heavy metal toxicity.
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