Telomerase, a ribonucleoprotein enzyme, is detected in the vast majority of malignant gliomas, but not in normal brain tissues. In malignant gliomas, tumors with telomerase tend to have more malignant phenotype than those without telomerase. Therefore, our long-term goal is to explore a novel telomerase-targeting therapy for malignant gliomas. To inhibit telomerase function effectively, we have adopted the 2-5A (2', 5'-oligoadenylate) antisense system. 2-5A is a mediator of one pathway of interferon actions by activating RNase L, resulting in single-stranded RNA cleavage. By linking 2-5A to antisense, RNase L degrades the targeted RNA specifically and effectively. With the grant supported by NIH, we synthesized the antisense oligonucleotide against human telomerase RNA component (hTR) linked to 2-5A (2-5A-anti-hTR) and investigated its anti-tumor effect on malignant glioma cells. Treatment with 2-5A-anti-hTR for 4 days induced a massive apoptosis before a telomere length shortened critically. In contrast, normal cells such as astrocytes and fibroblasts lacking telomerase were insensitive to 2-5A-anti-hTR. Treatment of subcutaneous or intracerebral tumors in nude mice with intratumoral injections of 2-5A-anti-hTR was effective. Based on our previous results, we hypothesize that 2-5A-anti-hTR is a promising agent for the treatment of malignant gliomas expressing telomerase. However, the following questions remain to be answered. First, what molecular pathways play a key role in 2-5A-anti-hTR-induced apoptosis? Second, how can we enhance the effect of 2-5A-anti-hTR on intracerebral tumors? Third, which regimens based on 2-5A-anti-hTR show a significant combination effect on intracerebral tumors? The aim of this proposal is to address these issues.
The specific aims are to: 1: Characterize the molecular pathways of 2-5A-anti-hTR-induced apoptosis. Using the microarray assay, we will identify which genes are involved in 2-5A-anti-hTR-induced cell death. We will also determine (i) the involvement of the mitochondria-associated cell death signaling pathways and (ii) the effect of 2-5A-anti-hTR on telomere 3' overhang or telomerase-positive astrocytes with or without tumorigenicity. 2: Define the effect of convection-enhanced drug delivery (CEDD) of 2-5A-anti-hTR on intracerebral tumors in nude mice. We will optimize CEDD and then compare the effect of CEDD and bolus injections of 2-5A-anti-hTR on intracerebral tumors. 3: Define the in vitro and in vivo effect of combination therapy based on 2-5A-anti-hTR. We will determine (i) the molecular pathways underlying the combination effect of 2-5A-anti-hTR and apoptosis-inducing agent (cisplatin, paclitaxel, or BCNU) and (ii) the effect of combination therapy on intracerebral tumors.
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