Infectious delivery and expression of a 135 kb human FRDA genomic DNA locus complements Friedreich's ataxia deficiency in human cells.
Friedreich's ataxia (FA) is the most common recessive ataxia, affecting 1-2 in 50,000 Caucasians, and there is currently no effective cure or treatment. FA results from a deficiency of the mitochondrial protein frataxin brought about by a repeat expansion in intron 1 of the FRDA gene. The main areas affected are the central nervous system (particularly the spinocerebellar system) and cardiac tissue. Therapies aimed at alleviating the neurological degeneration have proved unsuccessful to date. Here, we describe the construction and delivery of high capacity herpes simplex virus type 1 (HSV-1) amplicon vectors expressing the entire 80 kb FRDA genomic locus, driven by the endogenous FRDA promoter and including all introns and flanking regulatory sequences within a 135 kb genomic DNA insert. FA patient primary fibroblasts deficient in frataxin protein and exhibiting sensitivity to oxidative stress were transduced at high efficiency by FRDA genomic locus vectors. Following vector transduction, expression of FRDA protein by immunofluorescence was shown. Finally, functional complementation studies demonstrated restoration of the wild-type cellular phenotype in response to oxidative stress in transduced FA patient cells. These results suggest the potential of the infectious bacterial artificial chromosome-FRDA vectors for gene therapy of FA.
Gomez-Sebastian
Silvia
S
The Wellcome Trust Centre for Human Genetics, University of Oxford, Oxford, UK.
Gimenez-Cassina
Alfredo
A
Diaz-Nido
Javier
J
Lim
Filip
F
Wade-Martins
Richard
R
D000818:Animals; D002470:Cell Survival / Q000235:genetics / Q000502:physiology; D002478:Cells, Cultured; D022202:Chromosomes, Artificial, Bacterial / Q000235:genetics; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D005621:Friedreich Ataxia / Q000473:pathology / Q000628:therapy*; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000235:genetics; D018259:Herpesvirus 1, Human / Q000235:genetics; D006801:Humans; D033862:Iron-Binding Proteins / Q000737:chemistry / Q000235:genetics* / Q000502:physiology; D051379:Mice; D008957:Models, Genetic; D008970:Molecular Weight; D018384:Oxidative Stress; D000096923:Frataxin
D033862:Iron-Binding Proteins; D000096923:Frataxin