The human tissue kallikrein family of serine proteases (hK1-hK15 encoded by the genes KLK1-KLK15) is involved in several cancer-related processes. Accumulating evidence suggests that certain tissue kallikreins are part of an enzymatic cascade pathway that is activated in ovarian cancer and other malignant diseases. In the present study, OV-MZ-6 ovarian cancer cells were stably co-transfected with plasmids expressing hK4, hK5, hK6, and hK7. These cells displayed similar proliferative capacity as the vector-transfected control cells (which do not express any of the four tissue kallikreins), but showed significantly increased invasive behavior in an in vitro Matrigel invasion assay (p<0.01; Mann-Whitney U-test). For in vivo analysis, the cancer cells were inoculated into the peritoneum of nude mice. Simultaneous expression of hK4, hK5, hK6, and hK7 resulted in a remarkable 92% mean increase in 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 greater than 0.198, while this weight limit was not exceeded at all in the vector control group consisting of 13 mice (p=0.017; chi2 test). Our results strongly support the view that tumor-associated overexpression of tissue kallikreins contributes to ovarian cancer progression.

Recent evidence suggests that many tissue kallikreins are implicated in carcinogenesis. Kallikrein 8 (KLK8) plays a role in the physiology of the central nervous system. Kallikrein 7 (KLK7) takes part in skin desquamation. Both show altered expression in ovarian and breast cancer. In this study, we examined the level of mRNA expression of the KLK7 and KLK8 genes in 73 intracranial tumors using qualitative RT-PCR. The results were correlated with clinical and histomorphological variables and patient outcome. The expression of both genes was also examined in the brain cancer cell lines U-251 MG, D54 and SH-SY5Y and the invasive capacity of glioblastoma cells U-251 MG overexpressing hK7 or hK8 was also investigated in an in vitro Matrigel assay. Follow-up analysis revealed that expression of KLK7 mRNA was associated with shorter overall survival (OS) compared to patients with no KLK7 expression, as determined by Cox proportional hazard regression analysis. Overexpression of hK7 protein by cultivated brain tumor cells significantly enhanced the invasive potential in the Matrigel invasion assay, in contrast to cells overexpressing hK8 protein. Our data suggest that hK7 protein overexpression is associated with a more aggressive phenotype in brain cancer cells.

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.