Ovarian cancer (OC) is the highest-ranking cause of death from gynecological cancers among American women. Over 90 percent of OC are epithelial carcinoma derived from the human ovarian surface epithelium or HOSE. Epidemiological studies suggest that exposure of HOSE to pelvic inflammatory agents or to toxicants from the reproductive tract may lead to OC development. In addition, systemic exposure to environmental or dietary factors may contribute to the etiology since smoking and caffeine are linked to higher risks. Cytochrome P450-1 enzymes (CYP1 enzymes) are key enzymes responsible for activating procarcinogens to carcinogens. They are highly inducible by endogenous and exogenous factors such as inflammation, hypoxia, sex hormones, caffeine, polycyclic aromatic hydrocarbons, polychlorinated biphenols, and dioxins. Furthermore, their activities regulate estrogen inactivation and activation to genotoxic metabolites. Although overexpression of CYP1s has been linked to the development of several neoplasms an analogous knowledge base for OC is presently unavailable. Recently, we have demonstrated a) overexpression of CYP1A1 enzyme, but not CYP1B1 enzyme, in epithelial OC specimens, b) increased transcription of CYP1A1 in epithelial OC cells when compared to normal HOSE cells, c) existence of a novel CYP1A1 mRNA variant and its marked overexpression in epithelial OC cells, and d) differential growth response of normal and malignant HOSE cells to estrogen. Based on these preliminary data we hypothesize that overexpression of CYP1A1 enzyme is involved in the pathogenesis of epithelial OC, in part via modulation of estrogen metabolism, and that differential expression of wildtype and variant CYP1A1 transcript in vivo is a reflection of dissimilar roles played by these molecules during ovarian carcinogenesis. The goal of our proposed research is to elucidate 1) the role played by CYP1A1 in ovarian carcinogenesis, 2) the relationship between CYP1A1 activity and estrogen metabolism, and 3) the biological relevance of the newly identified CYP1A1 mRNA variant. Four aims have been put forth to test the hypothesis: Aim 1- Stable transfectants, with capabilities of conditional expression of wildtype- and/or variant CYP1A1 enzyme, will be used to determine whether their overexpression in immortalized non-tumorigenic HOSE cells leads to acquisition of malignant phenotypes. Aim 2- Epithelial OC cell lines will be inhibited from CYP1A1 enzyme expression, via a CYP1A1-specific antisense oligonucleotide (ODN), to determine if malignant phenotypes are diminished/abolished. Aim 3- Reverse phase HPLC will be used to determine whether HOSE cells and epithelial OC cells exhibit differential rates/patterns of estrogen metabolism, and if these differences could be explained by differential levels of CYP1A1 enzyme activity as measured by a functional assay (ethoxyresorufin O-deethylase activity). Aim 4- Using discarded human epithelial OC specimens, we will determine by real-time RT-PCR whether expression of CYP1A1 wildtype and/or variant transcript in laser-capture microscopy (LCM)-assisted microdissected carcinoma foci varies among different histopathological types and/or tumor grades of OC, and whether immuno-reactivity of the enzyme correlates with the relative abundance of one or both transcripts in the carcinoma foci.

R01CA094221

2002-07-16

2007-06-30