ABSTRACT
Oligonucleotides homologous to 3'-telomere overhang (T-oligos) trigger inherent telomere-based DNA damage responses mediated by p53 and/or ATM and induce senescence or apoptosis in various cancerous cells. However, T-oligo has limited stability in vivo due to serum and intracellular nucleases. To develop T-oligo as an innovative, effective therapeutic drug and to understand its mechanism of action, we investigated the antitumor effects of T-oligo or T-oligo complexed with a novel cationic alpha helical peptide, PVBLG-8 (PVBLG), in a p53 null melanoma cell line both in vitro and in vivo. The uptake of T-oligo by MM-AN cells was confirmed by immunofluorescence, and fluorescence-activated cell sorting analysis indicated that the T-oligo-PVBLG nanocomplex increased uptake by 15-fold. In vitro results showed a 3-fold increase in MM-AN cell growth inhibition by the T-oligo-PVBLG nanocomplex compared with T-oligo alone. Treatment of preformed tumors in immunodeficient mice 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. Moreover, T-oligo treatment downregulated procaspase-3 and procaspase-7 and increased catalytic activity of caspase-3 by 4-fold in MM-AN cells. Furthermore, T-oligo induced a 10-fold increase of senescence and upregulated the melanoma tumor-associated antigens MART-1, tyrosinase, and thrombospondin-1 in MM-AN cells, which are currently being targeted for melanoma immunotherapy. Interestingly, siRNA-mediated knockdown of p73 (4-10-fold) abolished this upregulation of tumor-associated antigens. In summary, we suggest a key role of p73 in mediating the anticancer effects of T-oligo and introduce a novel nanoparticle, the T-oligo-PVBLG nanocomplex, as an effective anticancer therapeutic.
Subject(s)
Melanoma/drug therapy , Nanocapsules/administration & dosage , Nanocapsules/chemistry , Oligonucleotides/therapeutic use , Peptides/administration & dosage , Animals , Cell Line, Tumor , Drug Combinations , Male , Melanoma/genetics , Melanoma/pathology , Mice , Mice, Nude , Nanocapsules/ultrastructure , Oligonucleotides/genetics , Particle Size , Peptides/chemistry , Treatment OutcomeABSTRACT
Exposure of the telomere overhang acts as a DNA damage signal, and exogenous administration of an 11-base oligonucleotide homologous to the 3'-telomere overhang sequence (T-oligo) mimics the effects of overhang exposure by inducing senescence and cell death in non-small cell lung cancer (NSCLC) cells, but not in normal bronchial epithelial cells. T-oligo-induced decrease in cellular proliferation in NSCLC is likely directed through both p53 and its homolog, p73, with subsequent induction of senescence and expression of senescence-associated proteins, p21, p33(ING), and p27(Kip1) both in vivo and in vitro. Additionally, T-oligo decreases tumor size and inhibits angiogenesis through decreased VEGF signaling and increased TSP-1 expression.
Subject(s)
Carcinoma, Non-Small-Cell Lung/genetics , DNA Damage , Genetic Predisposition to Disease , Neovascularization, Pathologic , Telomere/ultrastructure , Animals , Basic Helix-Loop-Helix Leucine Zipper Transcription Factors/metabolism , Bronchi/pathology , Cell Line, Tumor , Cell Proliferation , Cellular Senescence , Cyclin-Dependent Kinase Inhibitor p21/metabolism , Cyclin-Dependent Kinase Inhibitor p27/metabolism , Epithelial Cells/cytology , Gene Expression Regulation, Neoplastic , Humans , Inhibitor of Growth Protein 1 , Intracellular Signaling Peptides and Proteins/metabolism , Male , Mice , Mice, Nude , Neoplasm Transplantation , Nuclear Proteins/metabolism , Oligonucleotides/genetics , Signal Transduction , Tumor Suppressor Proteins/metabolism , Vascular Endothelial Growth Factor A/metabolismABSTRACT
Effective drug delivery for many neurodegenerative diseases or tumors of the central nervous system is challenging. Targeted invasive delivery of large macromolecules such as trophic factors to desired locations inside the brain is difficult due to anisotropy and heterogeneity of the brain tissue. Despite much experimental research, prediction of bio-transport phenomena inside the brain remains unreliable. This article proposes a rigorous computational approach for accurately predicting the fate of infused therapeutic agents inside the brain. Geometric and physiological properties of anisotropic and heterogeneous brain tissue affecting drug transport are accounted for by in-vivo diffusion tensor magnetic resonance imaging data. The three-dimensional brain anatomy is reconstructed accurately from subject-specific medical images. Tissue anisotropy and heterogeneity are quantified with the help of diffusion tensor imaging (DTI). Rigorous first principles physical transport phenomena are applied to predict the fate of a high molecular weight trophic factor infused into the midbrain. Computer prediction of drug distribution in humans accounting for heterogeneous and anisotropic brain tissue properties have not been adequately researched in open literature before.
