title: Selective modification of HK peptides enhances siRNA silencing of tumor targets in vivo.
abstract: Our research has focused on systemic delivery of small interference RNA (siRNA) by branched peptides composed of histidine and lysine. After studying several histidine-lysine (HK) peptides, one four-branched peptide, H3K(+H)4b, with a predominant repeating pattern of -HHHK-, was found to be an effective carrier of siRNA. Although the unmodified H3K(+H)4b carrier of siRNA targeting an oncogene was previously shown to have promise in a tumor-bearing mouse model, we sought to develop a more effective HK carrier of siRNA in this study. Our primary goal was to determine whether different ligand (cyclic RGD)-pegylation patterns on the H3K(+H)4b peptide affect siRNA delivery in vitro and in vivo. We compared the unmodified H3K(+H)4b with two modified H3K(+H)4b peptides for their ability to deliver siRNA in a tumor-bearing mouse model; one modified HK peptide, (RGD-PEG)(4)-H3K(+H)4b, had four cyclic RGD-polyethylene glycol (cRGD-PEG) conjugates per molecule, whereas the other peptide, (RGD-PEG)-H3K(+H)4b, had one cRGD-PEG per molecule. Although the modified HK peptides by themselves did not form stable nanoplexes with siRNA, combination of a highly charged unmodified HK peptide, H2K4b, with either of the modified HK peptides did form stable siRNA nanoparticles. For in vitro experiments with MDA-MB-435 cells that expressed luciferase (Luc), the H3K(+H)4b siRNA nanoplexes targeting Luc decreased its activity by 90% compared with negligible downregulation by the modified H3K(+H)4b nanoplexes (P<0.01). In contrast, the two modified H3K(+H)4b siRNA nanoplexes administered intravenously were more effective than the H3K(+H)4b nanoplexes in silencing Luc in a tumor xenograft model. The Luc activity in tumor lysates of mice administered H3K(+H)4b, (RGD-PEG)-H3K(+H)4b and (RGD-PEG)(4)-H3K(+H)4b nanoplexes decreased by 18, 35 and 75%, respectively. Thus, the siRNA nanoplex incorporating the highly modified peptide, (RGD-PEG)(4)-H3K(+H)4b, was the most effective at silencing its target in vivo (P<0.01). These studies demonstrate that selectively modified HK polymers are promising candidates for targeting oncogenes with siRNA.
authors.affiliation: Department of Pathology, University of Maryland Baltimore, Baltimore, MD, USA.
authors.forename: S-T
authors.initials: ST
authors.lastname: Chou
authors.forename: Q
authors.initials: Q
authors.lastname: Leng
authors.forename: P
authors.initials: P
authors.lastname: Scaria
authors.forename: M
authors.initials: M
authors.lastname: Woodle
authors.forename: A J
authors.initials: AJ
authors.lastname: Mixson
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor; D016207:Cytokines; D005260:Female; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D006801:Humans; D051379:Mice; D008819:Mice, Nude; D053758:Nanoparticles / Q000008:administration & dosage / Q000737:chemistry; D009369:Neoplasms / Q000235:genetics* / Q000378:metabolism; D010455:Peptides / Q000737:chemistry* / Q000378:metabolism; D034622:RNA Interference; D034741:RNA, Small Interfering; D023041:Xenograft Model Antitumor Assays
chemical_list: D016207:Cytokines; D010455:Peptides; D034741:RNA, Small Interfering