Abstract 2886: Carbon nanotube-mediated siRNA for gastrin-releasing peptide receptor silencing in neuroblastoma

Abstract

Abstract Small interfering RNA (siRNA) has the potential to influence gene expression with a high degree of target gene specificity, making siRNA-mediated therapeutic approaches very promising in biomedicine. However, the clinical applications of siRNA therapeutics have been impeded by its poor intracellular uptake, instability in vivo, and non-specific immune stimulations. In this study, we have aimed to resolve these difficulties in the applications of siRNA, we developed a magneto-fluorescent carbon nanotube (CNT) platform to deliver siRNA into neuroblastoma and down-regulate gastrin-releasing peptide receptor (GRP-R) expression in vitro and in vivo. METHODS. Human neuroblastoma BE(2)-C cells were used in this study. siRNA oligos were custom synthesized by Thermo Scientific (Dharmacon). CNTs were produced by a high-pressure CO (HiPco) at Rice University. PEG-N (1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy (polyethylene glycol)-5000]) was purchased from Avanti Polar Lipids Inc. (Alabaster, AL). Target gene expression was measured with Western blotting and immunohistochemical staining. siRNA stability was examined by agarose gel. The cytotoxicity of CNT was evaluated by measuring cell viability. RESULTS. We have developed a magneto-fluorescent carbon nanotube (CNT) platform to deliver siRNA into neuroblastoma and to down-regulate GRP-R expression in vitro and in vivo. We have systematically investigated the optical and magnetic properties of CNTs with different functional groups and have found that the functional groups on CNTs play an important role on the magnetic resonance imaging (MRI) and near-infrared fluorescence (NIRF) imaging of CNTs. Moreover, we have shown that CNT-mediated siRNA (CNT-siRNA) can silence GRP-R with efficiency up to 92%, which is much higher than that of siRNA mediated by the commercial transfection reagent, and effectively reduce downstream targets of GRP-R. This high efficiency may result from the enhanced stability of CNT-siRNA after transfection. We have also demonstrated the universal siRNA delivery efficiency by knocking down AKT2 in neuroblastoma cells and GRP-R in human breast cancer cells. In addition, we have found that CNT-siRNA can significantly silence GRP-R in in vivo subcutaneous mouse models. CONCLUSIONS. Our findings demonstrate that functionalized-CNTs mediated siRNA delivery caused high gene silencing efficiency of in vitro and in vivo. In addition, our results showed CNT produced MRI and NIRF imaging in cells. These fundamental studies offer a way to develop a multifunctional CNT delivery system with INTRINSIC MRI and NIRF imaging to evaluate the treatment effect simultaneously. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2886. doi:1538-7445.AM2012-2886