title: Short-interfering RNAs induce retinal degeneration via TLR3 and IRF3.
abstract: The discovery of sequence-specific gene silencing by endogenous double-stranded RNAs (dsRNA) has propelled synthetic short-interfering RNAs (siRNAs) to the forefront of targeted pharmaceutical engineering. The first clinical trials utilized 21-nucleotide (nt) siRNAs for the treatment of neovascular age-related macular degeneration (AMD). Surprisingly, these compounds were not formulated for cell permeation, which is required for bona fide RNA interference (RNAi). We showed that these "naked" siRNAs suppress neovascularization in mice not via RNAi but via sequence-independent activation of cell surface Toll-like receptor-3 (TLR3). Here, we demonstrate that noninternalized siRNAs induce retinal degeneration in mice by activating surface TLR3 on retinal pigmented epithelial cells. Cholesterol conjugated siRNAs capable of cell permeation and triggering RNAi also induce the same phenotype. Retinal degeneration was not observed after treatment with siRNAs shorter than 21-nts. Other cytosolic dsRNA sensors are not critical to this response. TLR3 activation triggers caspase-3-mediated apoptotic death of the retinal pigment epithelium (RPE) via nuclear translocation of interferon regulatory factor-3. While this unexpected adverse effect of siRNAs has implications for future clinical trials, these findings also introduce a new preclinical model of geographic atrophy (GA), a late stage of dry AMD that causes blindness in millions worldwide.
authors: Department of Ophthalmology and Visual Sciences, University of Kentucky, Lexington, Kentucky 40536-0284, USA.
authors: Mark E
authors: ME
authors: Kleinman
authors: Hiroki
authors: H
authors: Kaneko
authors: Won Gil
authors: WG
authors: Cho
authors: Sami
authors: S
authors: Dridi
authors: Benjamin J
authors: BJ
authors: Fowler
authors: Alexander D
authors: AD
authors: Blandford
authors: Romulo J C
authors: RJ
authors: Albuquerque
authors: Yoshio
authors: Y
authors: Hirano
authors: Hiroko
authors: H
authors: Terasaki
authors: Mineo
authors: M
authors: Kondo
authors: Takashi
authors: T
authors: Fujita
authors: Balamurali K
authors: BK
authors: Ambati
authors: Valeria
authors: V
authors: Tarallo
authors: Bradley D
authors: BD
authors: Gelfand
authors: Sasha
authors: S
authors: Bogdanovich
authors: Judit Z
authors: JZ
authors: Baffi
authors: Jayakrishna
authors: J
authors: Ambati
mesh_terms: D000818:Animals; D053148:Caspase 3 / Q000378:metabolism; D016923:Cell Death / Q000235:genetics; D006801:Humans; D050838:Interferon Regulatory Factor-3 / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D011485:Protein Binding; D034741:RNA, Small Interfering / Q000378:metabolism / Q000633:toxicity*; D012162:Retinal Degeneration / Q000139:chemically induced* / Q000378:metabolism / Q000473:pathology; D055213:Retinal Pigment Epithelium / Q000378:metabolism; D015398:Signal Transduction; D051196:Toll-Like Receptor 3 / Q000378:metabolism*
chemical_list: D050838:Interferon Regulatory Factor-3; D008565:Membrane Proteins; D034741:RNA, Small Interfering; D051196:Toll-Like Receptor 3; D053148:Caspase 3