Abstract
2296
The diffuse and highly invasive nature of malignant gliomas is believed to rely on the degradation of extracellular matrix components by several proteases. Our previous studies clearly demonstrated that uPA, uPAR, and cathepsin B levels are significantly increased during the progression of tumor growth in animal models and human brain tissue samples. Antisense stable clones for these molecules and ex vivo antisense Ad-constructs for these molecules do not form tumors in vivo in nude mice. Since antisense-mediated gene silencing does not completely inhibit the translation of target mRNA and high concentrations of antisense molecules are required to achieve gene silencing, we used the RNA interference (RNAi) approach to silence uPA, uPAR and cathepsin B in this study. We constructed a cytomegalovirus promoter-driven DNA template to induce hairpin RNA triggered RNAi to inhibit uPA, uPAR and cathepsin B gene expression. We then determined the levels of these molecules and their effects on invasiveness, angiogenesis and tumor regression in both in vitro and in vivo models. Western blot analysis of uPA, uPAR and cathepsin B were significantly decreased in glioblastoma cells transfected with a plasmid vector expressing dsRNA for uPA, uPAR and cathepsin B (pU2C). In comparison to parental and vector controls, pU2C-transfected cells retarded the invasion of glioblastoma cells as indicated by Matrigel and spheroid assays. In an in vitro model, pU2C significantly inhibited angiogenesis when compared to the parental and vector controls. Furthermore, pU2C treatment (directly at the tumor site or intraperitoneal injection) of pre-established intracranial tumors resulted in complete regression of the tumors in nude mice. Our results clearly support the therapeutic potential of RNAi-mediated targeting of uPA, uPAR and cathepsin B for the treatment of human gliomas.
- American Association for Cancer Research