RNase L, a ubiquitous endoribonuclease in the IFN system, has been harnessed for the selective and catalytic cleavage of RNA targets that function in signal transduction and oncogenesis. Chimeric oligonucleotides (ODNs) are chemically-synthesized with a 2',5'-tetraadenylate activator (2- 5A) of RNase L covalently coupled to antisense ODNs. In essence, the 2- 5A portion of the chimera binds and activates RNase L while the antisense arm of the ODN binds the RNA target thus directing RNase L to degrade the RNA specifically. The goal of this project is to determine the molecular mechanism for the tumoricidal activity of 2-5A-antisense directed against human telomerase RNA (hTR) and thus better understand the role of telomerase in cancer and cell immortality. The hypothesis to be tested is that the tumoricidal effect of 2-5A-anti-hTR is due to inhibition of telomerase function which directly or indirectly triggers an apoptotic response. Different chemical forms of 2-5A-antisense will be designed to enhance biostability, hybridization affinity, cellular uptake and RNase L activation ability. We will probe the signaling events which lead to apoptosis in tumor cells treated with 2-5A-antisense to telomerase RNA. The contributions of caspases, STAT 1, and bcl-2 family members will be determined. Effects of 2-5A-antisense on global gene expression patterns, telomere length and chromosome stability will be determined. Because human but not murine RNase L is activated by 2-5A-antisense, an animal model for studying the anti-tumor effects of 2-5A-antisense against telomerase RNA will be generated by expressing human RNase L in RNase L-/- mice. The effects of IFN induction of RNase L will be determined in 2-5A-antisense treated cells and mice. Because 2-5A-anti- hTR is a potent experimental therapeutic agent for cancer, this proposal has the potential to make an impact on intractable forms of cancer, such as malignant glioma.
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