title: RNA interference-mediated suppression and replacement of human rhodopsin in vivo.
abstract: Mutational heterogeneity represents a significant barrier to development of therapies for many dominantly inherited diseases. For example, >100 mutations in the rhodopsin gene (RHO) have been identified in patients with retinitis pigmentosa (RP). The development of therapies for dominant disorders that correct the primary genetic lesion and overcome mutational heterogeneity is challenging. Hence, therapeutics comprising two elements--gene suppression in conjunction with gene replacement--have been investigated. Suppression is targeted to a site independent of the mutation; therefore, both mutant and wild-type alleles are suppressed. In parallel with suppression, a codon-modified replacement gene refractory to suppression is provided. Both in vitro and in vivo validation of suppression and replacement for RHO-linked RP has been undertaken in the current study. RNA interference (RNAi) has been used to achieve ~90% in vivo suppression of RHO in photoreceptors, with use of adeno-associated virus (AAV) for delivery. Demonstration that codon-modifed RHO genes express functional wild-type protein has been explored transgenically, together with in vivo expression of AAV-delivered RHO-replacement genes in the presence of targeting RNAi molecules. Observation of potential therapeutic benefit from AAV-delivered suppression and replacement therapies has been obtained in Pro23His mice. Results provide the first in vivo indication that suppression and replacement can provide a therapeutic solution for dominantly inherited disorders such as RHO-linked RP and can be employed to circumvent mutational heterogeneity.
authors.affiliation: Smurfit Institute of Genetics, Trinity College, Dublin, Ireland. oreillym@tcd.ie
authors.forename: Mary
authors.initials: M
authors.lastname: O'Reilly
authors.forename: Arpad
authors.initials: A
authors.lastname: Palfi
authors.forename: Naomi
authors.initials: N
authors.lastname: Chadderton
authors.forename: Sophia
authors.initials: S
authors.lastname: Millington-Ward
authors.forename: Marius
authors.initials: M
authors.lastname: Ader
authors.forename: Thérèse
authors.initials: T
authors.lastname: Cronin
authors.forename: Thérèse
authors.initials: T
authors.lastname: Tuohy
authors.forename: Alberto
authors.initials: A
authors.lastname: Auricchio
authors.forename: Markus
authors.initials: M
authors.lastname: Hildinger
authors.forename: Amanda
authors.initials: A
authors.lastname: Tivnan
authors.forename: Niamh
authors.initials: N
authors.lastname: McNally
authors.forename: Marian M
authors.initials: MM
authors.lastname: Humphries
authors.forename: Anna-Sophia
authors.initials: AS
authors.lastname: Kiang
authors.forename: Pete
authors.initials: P
authors.lastname: Humphries
authors.forename: Paul F
authors.initials: PF
authors.lastname: Kenna
authors.forename: G Jane
authors.initials: GJ
authors.lastname: Farrar
mesh_terms: D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D001483:Base Sequence; D015316:Genetic Therapy / Q000379:methods*; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics; D012160:Retina / Q000737:chemistry / Q000378:metabolism / Q000473:pathology; D012174:Retinitis Pigmentosa / Q000473:pathology / Q000628:therapy*; D012243:Rhodopsin / Q000032:analysis / Q000235:genetics*; D013489:Suppression, Genetic
chemical_list: D034741:RNA, Small Interfering; D012243:Rhodopsin