Alterations in the expression of the DNA repair/redox enzyme APE/ref-1 in epithelial ovarian cancers

The DNA base excision repair pathway is responsible for the repair of alkyl ation and oxidative DNA damage. A crucial step in the base excision repair pathway involves the cleavage of an apurinic/apyrimidinic (AP) site in DNA by an AP endonuclease (APE), The major AP endonuclease in mammalian cells i s APE/ref-1 a multifunctional enzyme that acts not only as an AP endonuclea se but as a redox-modifying factor for a variety of transcription factors, The purpose of this study was to determine the expression of APE/redox fact or-1 (ref-l) in ovarian tissues, particularly ovarian cancers. Formalin-fix ed, paraffin-embedded specimens of ovarian tissues (normal, various benign conditions, and epithelial cancers) were studied using both polyclonal and monoclonal antibodies to APE/ref-1. The relationship between APE/ref-1 prot ein levels and DNA repair activity was studied in ovarian Hey and Hey-C2 ce ll Lines using Western blot and a specific AP-site oligonucleotide cleavage assay, Hey and Hey-C2 cells were fractionated, and the nuclear and cytopla smic extracts were quantitated for protein levels and assessed for APE/ref- 1 with Western blot. Normal ovarian tissues consistently demonstrated stron g nuclear staining of the surface epithelium, epithelial inclusions, corpor a lutea and albicantia, and stroma, Cytoplasmic staining was absent. A simi lar pattern was seen for benign conditions including endometriosis. Low mal ignant potential ovarian cancers stained in a pattern similar to normal ova rian and nonneoplastic tissues; however, two specimens also had areas of cy toplasmic staining. Epithelial ovarian cancers were remarkably different fr om all other ovarian tissues studied. Both nuclear and cytoplasmic staining of the malignant epithelium were seen and ranged from strong to weak, ofte n with considerable staining heterogeneity within the same tumor. The AP-si te oligonucleotide cleavage assay indicated that APE/ref-1 protein levels c orrelate well with DNA repair activity. The increased levels of APE/ref-1 i n the Hey-C2. cells was mainly attributable to increased cytoplasmic enzyme . APE/ref-1 immunoreactivity is altered in malignant ovarian tumors, furthe r studies will determine whether the altered expression and subcellular loc ation reflect changes in redox regulatory functions.