title: Stability and immunogenicity properties of the gene-silencing polypurine reverse Hoogsteen hairpins.
abstract: Gene silencing by either small-interference RNAs (siRNA) or antisense oligodeoxynucleotides (aODN) is widely used in biomedical research. However, their use as therapeutic agents is hindered by two important limitations: their low stability and the activation of the innate immune response. Recently, we developed a new type of molecule to decrease gene expression named polypurine reverse Hoogsteen hairpins (PPRHs) that bind to polypyrimidine targets in the DNA. Herein, stability experiments performed in mouse, human, and fetal calf serum and in PC3 cells revealed that the half-life of PPRHs is much longer than that of siRNAs in all cases. Usage of PPRHs with a nicked-circular structure increased the binding affinity to their target sequence and their half-life in FCS when bound to the target. Regarding the innate immune response, we determined that the levels of the transcription factors IRF3 and its phosphorylated form, as well as NF-κB were increased by siRNAs and not by PPRHs; that the expression levels of several proinflammatory cytokines including IL-6, TNF-α, IFN-α, IFN-ß, IL-1ß, and IL-18 were not significantly increased by PPRHs; and that the cleavage and activation of the proteolytic enzyme caspase-1 was not triggered by PPRHs. These determinations indicated that PPRHs, unlike siRNAs, do not activate the innate inflammatory response.
authors.affiliation: Department of Biochemistry and Molecular Biology, School of Pharmacy, University of Barcelona , Av. Diagonal 643, E-08028 Barcelona, Spain.
authors.forename: Xenia
authors.initials: X
authors.lastname: Villalobos
authors.forename: Laura
authors.initials: L
authors.lastname: Rodríguez
authors.forename: Jeanne
authors.initials: J
authors.lastname: Prévot
authors.forename: Carlota
authors.initials: C
authors.lastname: Oleaga
authors.forename: Carlos J
authors.initials: CJ
authors.lastname: Ciudad
authors.forename: Véronique
authors.initials: V
authors.lastname: Noé
mesh_terms: D000818:Animals; D015153:Blotting, Western; D001943:Breast Neoplasms / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D049109:Cell Proliferation; D016207:Cytokines / Q000235:genetics / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D005260:Female; D020868:Gene Silencing / Q000276:immunology*; D006207:Half-Life; D006801:Humans; D050838:Interferon Regulatory Factor-3 / Q000235:genetics / Q000378:metabolism; D008297:Male; D051379:Mice; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism; D011471:Prostatic Neoplasms / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D011685:Purine Nucleotides / Q000737:chemistry* / Q000276:immunology* / Q000494:pharmacology; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014407:Tumor Cells, Cultured
chemical_list: D016207:Cytokines; C494232:IRF3 protein, human; D050838:Interferon Regulatory Factor-3; D016328:NF-kappa B; D011685:Purine Nucleotides; D012333:RNA, Messenger; D034741:RNA, Small Interfering