Abstract 4577: Telomere oligonucleotides induce DNA damage responses in cancer cells: Mechanism of action and novel nanocomplex delivery system

Abstract

Abstract Telomeres, the ribonucleoprotein complexes found at the ends of eukaryotic chromosomes, protect genomic DNA from end-to-end fusions and degradation by nucleases. During each round of cell division, the ends of telomeres are incompletely replicated, resulting in a net loss of telomeric DNA. When critically shortened, telomeres lose their ability to protect chromosomes from sustaining damage, thereby triggering cellular senescence or apoptosis. In numerous cancer types, administration of an oligonucleotide homologous to 3′-telomere overhang (T-oligo) induces potent DNA damage responses, presumably by mimicking or inducing telomere DNA damage, resulting in cell cycle arrest, senescence, or apoptosis. Remarkably, T-oligo induces minimal side-effects in normal cells, and thus, is actively being investigated as a novel anticancer therapeutic. However, as an oligonucleotide, T-oligo's intrinsic instability makes it susceptible to degradation by intracellular and serum nucleases, which severely limits its clinical use. To develop T-oligo as an innovative, effective therapeutic drug, it was complexed with a novel cationic alpha helical peptide, PVBLG-8 (PVBLG-8). PVBLG has been shown to be highly effective in delivering plasmid DNA and siRNA to a variety of cell lines, however, it has not been investigated using oligonucleotides. Here, our studies focus on its mechanism of action, stabilization and improved delivery. We show that PVBLG effectively stabilizes T-oligo by forming a nanocomplex (TOP complex), and enhances its anticancer properties both in vitro and in vivo. In addition, we identify new proteins involved in T-oligo's mechanism of action. The uptake of T-oligo by MM-AN melanoma cells was confirmed by immunofluorescence, and fluorescence-activated cell sorting analysis indicated that the TOP complex increased uptake by 15-fold. In MM-AN cells, T-oligo complexed with the PVBLG (0.025-.25mg/ml) inhibited growth in vitro in a dose dependent manner by 4.5-8.3-fold, compared to 3-fold by T-oligo alone. Similar results were seen in H358 lung cancer cells. Preformed MM-AN tumors in immunodeficient mice treated with the T-oligo-PVBLG nanocomplex resulted in a 3-fold reduction in tumor volume compared with T-oligo alone. This reduction in tumor volume was associated with decreased vascular endothelial growth factor expression and induction of thrombospondin-1 expression and apoptosis. In addition, T-oligo treatment demonstrated downregulation of telomere repeat factor 1, a negative regulator of telomere length, in MU melanoma cells, indicating a role for TRF1 in T-oligo-induced DNA damage responses. In summary, our investigation further defines T-oligo's mechanism of action, and demonstrates the potential of a novel nanoparticle, the TOP complex, as an effective anticancer therapeutic. Citation Format: Luke M. Wojdyla, Srijayaprakash B. Uppada, Neelu Puri. Telomere oligonucleotides induce DNA damage responses in cancer cells: Mechanism of action and novel nanocomplex delivery system. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4577. doi:10.1158/1538-7445.AM2014-4577