Gliomas are the most frequent primary intracranial neoplasm in adults and are invariably fatal, despite availability of conventional treatments. As such, there is a necessity to explore alternative therapies. We will develop proapoptotic nucleic acid agents that can activate Fas induced apoptosis or the intrinsic apoptotic pathway in gliomas. In preliminary studies, the Rzs showed activity against several novel proapoptotic mRNAs in prostate, colon and brain cancer cells. We have demonstrated that synthetic chimeric proapoptotic ribozymes (Rzs) delivered by cannula intracranially to normal rat brain or to brains of rats with established brain tumors are well tolerated. We subsequently engineered siRNAs against some of these same target genes and tested them in vitro. Our preliminary results show that the Rzs and siRNAs are efficacious in activating Fas induced apoptosis, as well as inducing apoptosis by the intrinsic pathway in colon carcinoma and gliomas in vitro and in vivo. We propose to optimize the in vitro efficacy of the proapoptotic nucleic acid agents by comparing a panel of proapoptotic Rzs and siRNAs targeting the same mRNAs of 3 genes. We will test the Rzs and siRNAs individually and then in combinations. We will first explore the stability of the Rz and siRNA in cerebrospinal fluid (CSF), serum, normal brain tissue homogenates, and tumor cell lysates. We will also assess the uptake and half-life of them when delivered to glioma cells under various transfection conditions. Delivery and cytotoxicity will first be explored in vitro culminating in the selection of an optimal transfection vehicle, if needed. Delivery and toxicity will then be examined in normal or tumor-bearing cannulated rat brains when given systemically and intracranially (IC). Enhancements to these delivery methods will be tested by intracranial convection enhanced delivery (IC- CED), and by intracarotid administration with blood-brain-barrier (BBB) modification. Finally, experiments comparing the efficacy of the optimized Rz and siRNA combination in a rat glioma model, by tumor volumetric and survival studies, and in a human xenotransplant model by tumor volumetric and imaging studies will be used to determine the most efficacious nucleic acid agents for testing in clinical trials. We hypothesize that proapoptotic nucleic acid agents will be effective and nontoxic adjuvants for glioma therapy. ? ? ?
