Antitumor Activity of Systemically Delivered Ribozymes Targeting Murine Telomerase RNA

Abstract

Purpose: To test ribozymes targeting mouse telomerase RNA (mTER) for suppression of the progression of B16-F10 murine melanoma metastases in vivo.Experimental Design: Hammerhead ribozymes were designed to target mTER. The ribozyme sequences were cloned into a plasmid expression vector containing EBV genomic elements that substantially prolong expression of genes delivered in vivo. The activity of various antitelomerase ribozymes or control constructs was examined after i.v. injection of cationic liposome:DNA complexes containing control or ribozyme constructs. Expression of ribozymes and mTER at various time points were evaluated by quantitative real-time PCR. Telomerase activity was examined using the telomeric repeat amplification protocol.Results: Systemic administration of cationic liposome: DNA complexes containing a plasmid-expressed ribozyme specifically targeting a cleavage site at mTER nucleotide 180 significantly reduced the metastatic progression of B16-F10 murine melanoma. The antitumor activity of the anti-TER 180 ribozyme in mice was abolished by a single inactivating base mutation in the ribozyme catalytic core. The EBVbased expression plasmid produced sustained levels of ribozyme expression for the full duration of the antitumor studies. In addition to antitumor activity, cationic liposome: DNA complex-based ribozyme treatment also produced reductions in both TER levels and telomerase enzymatic activity in tumor-bearing mice.Conclusions: Systemic, plasmid-based ribozymes specifically targeting TER can reduce both telomerase activity and metastatic progression in tumor-bearing hosts. The work reported here demonstrates the potential utility of plasmid-based anti-TER ribozymes in the therapy of melanoma metastasis.

Genes and pathways downstream of telomerase in melanoma metastasis

“…This anti-TER 180 Rz was previously shown to down-regulate TER levels (by TaqMan assays; Applied Biosystems) and telomerase enzymatic activity, and to significantly suppress metastatic progression in vivo compared with vector-only and mutant ribozyme controls (16). Previously, a control inactive mutant anti-TER ribozyme was shown to be comparable to the empty vector control in its inability to suppress the lung metastasis of melanoma (16). Therefore, the effects of ribozyme expression were compared with B16 cells transfected with the empty vector as an appropriate control in these studies.…”

“…The well established roles for telomerase in tumor initiation and cellular immortalization (7,8) have led to the identification of telomerase as a potentially important molecular target in cancer therapeutics (9)(10)(11). To date, multiple studies have examined the utility of targeting telomerase to inhibit tumor cell proliferation (12)(13)(14)(15)(16).…”

“…In our recent study, we used a systemic injection model of an effective anti-telomerase ribozyme to reveal that inhibiting telomerase activity in tumorbearing mice significantly reduces metastatic progression (16). Here, we examine the direct role played by telomerase in the metastatic potential of murine melanoma and characterize cellular pathways altered by telomerase suppression in tumor cells.…”

TERT rs2736098 Polymorphism and Cancer Risk: Results of a Meta-analysis

“…Over-expression of the TERT gene and telomerase activity has been recognized in many types of cancers, and can possibly lead to unlimited cell division and carcinogenesis (Liu et al, 2010). In addition, telomerase may play a role in tumor progression and metastasis by activation of the glycolytic pathway and in suppression of tumor cell differentiation (Nosrati et al, 2004;Bagheri et al, 2006;Liu et al, 2010). The role of TERT rs2736098 expression has been well documented in the tumorigenesis of various cancers (Savage et al, 2007;Jin et al, 2009;Rafnar et al, 2009;Liu et al, 2010;Chen et al, 2011;Gago-Dominguez et al, 2011), but the influence of the TERT rs2736098 polymorphism in the development of cancer has been evaluated by only a few meta-analysis studies..…”

Telomerase reverse transcriptase expression elevated by avian leukosis virus integration in B cell lymphomas

“…TERT also seems to enhance the survival of ESC by increasing the expression of stress response and defense genes (32). Recent studies show telomerase may also play a role in tumor progression and metastasis by activation of the glycolytic pathway and in suppression of tumor-cell differentiation (33,34).…”