A bioengineered 3D ovarian cancer model for the assessment of peptidase–mediated enhancement of spheroid growth and intraperitoneal spread

Abstract

Cancer–associated proteases promote peritoneal dissemination and chemoresistance in malignant progression. In this study, kallikrein–related peptidases 4, 5, 6, and 7 (KLK4–7)–cotransfected OV–MZ–6 ovarian cancer cells were embedded in a bioengineered three–dimensional (3D) microenvironment that contains RGD motifs for integrin engagement to analyze their spheroid growth and survival after chemotreatment. KLK4–7–cotransfected cells formed larger spheroids and proliferated more than controls in 3D, particularly within RGD–functionalized matrices, which was reduced upon integrin inhibition. In contrast, KLK4–7–expressing cell monolayers proliferated less than controls, emphasizing the relevance of the 3D microenvironment and integrin engagement. In a spheroid–based animal model, KLK4–7–overexpression induced tumor growth after 4 weeks and intraperitoneal spread after 8 weeks. Upon paclitaxel administration, KLK4–7–expressing tumors declined in size by 91% (controls: 87%) and showed 90% less metastatic outgrowth (controls: 33%, P < 0.001). KLK4–7–expressing spheroids showed 53% survival upon paclitaxel treatment (controls: 51%), accompanied by enhanced chemoresistance–related factors, and their survival was further reduced by combination treatment of paclitaxel with KLK4/5/7 (22%, P = 0.007) or MAPK (6%, P = 0.006) inhibition. The concomitant presence of KLK4–7 in ovarian cancer cells together with integrin activation drives spheroid formation and proliferation. Combinatorial approaches of paclitaxel and KLK/MAPK inhibition may be more efficient for late–stage disease than chemotherapeutics alone as these inhibitory regimens reduced cancer spheroid growth to a greater extent than paclitaxel alone.