ID: 119 Effects of overexpression of the human tissue kallikrein genes KLK4, 5, 6, and 7 on the malignant phenotype of ovarian cancer cells

Abstract

The human tissue kallikrein family encompasses 15 serine proteases. Most of the tissue kallikrein genes (KLK1-15) or proteins (hK1-15) have been found to be either under-or overexpressed in certain carcinomas, especially in breast, prostate, testicular and ovarian cancer. In ovarian cancer tissue, several tissue kallikreins are strongly over-or even de novo expressed, compared to normal ovarian tissue. This may indicate that these tissue kallikreins participate in an enzymatic cascade pathway which is strongly activated in ovarian cancer. In the present study, full length cDNAs encoding hK4, hK5, hK6, and hK7 were amplified from ovarian cancer tissue and cloned into the eukaryotic expression vector pRcRSV. OV-MZ-6 ovarian cancer cells, which do not express any of the four tissue kallikreins, were stably co-transfected with these plasmids resulting in the tumor cell line OV-KLK4+5+6+7. Applying sensitive ELISA formats for hK4, hK5, hK6 and hK7, we demonstrated that the stably transfected OV-KLK4+5+6+7 cell line secreted detectable amounts of the respective tissue kallikreins into the medium, ranging from about 0.5 to 20 ng/ml after growth in culture for 48 h. OV-KLK4+5+6+7 cells displayed similar proliferative capacity as the vector-transfected control cells (OV-RSV), but showed a significantly increased invasive behavior in the in vitro Matrigel invasion assay (p<0.01; Mann-Whitney U-test). For in vivo analysis, cells expressing the four tissue kallikreins were inoculated into the peritoneum of nude mice. Simultaneous expression of hK4, hK5, hK6, and hK7 resulted in a remarkable 92% mean increase of tumor burden compared to the vector-control cell line. Five out of 14 mice in the "tissue kallikrein overexpressing" group displayed a tumor/situs ratio of greater than 0.198, while this weight limit was not at all exceeded in the vector control group consisting of 13 mice (p=0.017; chi-square-test). Our results strongly support the view that tumor-associated overexpression of tissue kallikreins contributes to ovarian cancer progression.