Silencing Of Dna Repair Proteins With Eco/Sirna Nanoparticles For The Enhancement Of Radiation Response In Glioblastoma

Glioblastoma (GBM) is one of the most lethal human cancers due in large part to the presence of cancer stem cells which contribute to treatment resistance and disease recurrence. Radiation therapy remains a major component of the standard of care for GBM patients, fueling the development of modification and sensitization strategies that can treat cancer stem cell populations and enhance therapeutic outcome. One such strategy is targeting the DNA damage response (DDR) to interfere with DNA repair in response to ionizing radiation. Ataxia telangiectasia mutated (ATM) and DNA-dependent protein kinase (DNApk) play critical roles in the DDR through the detection and subsequent repair of double-strand breaks (DSBs) via homologous recombination and non-homologous end-joining. Therapeutic attempts to target these proteins have suffered from poor bioavailability and specificity, spurring investigation into the use of nanoparticle delivery of siRNA to more specifically inhibit the DDR pathways. Previously, (1-aminoethyl)iminobis[N-(oleicylcysteinyl-1-amino-ethyl)-propionamide (ECO) nanoparticles have been shown to effectively deliver siRNA targeting pathways involved with tumor growth and metastasis. In this study we investigate the use of these nanoparticles to deliver siRNA to inhibit ATM and DNApk activity and enhance radiation response in both glioma and glioma stem cell lines. Established glioma (U251) and glioma stem cell (NSC11) lines were used to evaluate the effectiveness of ECO nanoparticle delivery of siRNA in vitro . Cellular uptake of ECO nanoparticles loaded with fluorescent siRNA was assessed using flow cytometry and fluorescent microscopy, demonstrating the rapid uptake of ECO/siRNA nanoparticles in comparison to commercially available transfection agents. Protein and mRNA analyses revealed the kinetics and degree of gene silencing with ECO nanoparticle delivery. The effect of ECO delivery of ATM and DNApk siRNA on radiation response was evaluated using clonogenic cell survival and γH2AX foci formation assays. In comparison to untreated cells, cells treated with the ECO nanoparticle/siRNA complex exhibited a delayed DNA repair response following irradiation. Our results demonstrate the potential of ECO nanoparticles to efficiently deliver siRNA targeting ATM and DNApk to glioma and glioma stem cells, resulting in an impaired ability to repair DNA damage and an enhanced response to radiation treatment. These results provide the basis for our future investigation into the use of ECO/siRNA complexes targeting ATM and DNApk to improve the radiation response of GBM in vivo . Citation Format: Jennifer A. Lee, Nadia Ayat, Anita Tandle, Zheng-Rong Lu, Kevin Camphausen. Silencing of DNA repair proteins with ECO/siRNA nanoparticles for the enhancement of radiation response in glioblastoma. [abstract]. In: Proceedings of the AACR Special Conference on Engineering and Physical Sciences in Oncology; 2016 Jun 25-28; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2017;77(2 Suppl):Abstract nr B42.