Safety and effects of the vector for the Leber hereditary optic neuropathy gene therapy clinical trial.
IMPORTANCE We developed a novel strategy for treatment of Leber hereditary optic neuropathy (LHON) caused by a mutation in the nicotinamide adenine dinucleotide dehydrogenase subunit IV (ND4) mitochondrial gene. OBJECTIVE To demonstrate the safety and effects of the gene therapy vector to be used in a proposed gene therapy clinical trial. DESIGN AND SETTING In a series of laboratory experiments, we modified the mitochondrial ND4 subunit of complex I in the nuclear genetic code for import into mitochondria. The protein was targeted into the organelle by agency of a targeting sequence (allotopic expression). The gene was packaged into adeno-associated viral vectors and then vitreally injected into rodent, nonhuman primate, and ex vivo human eyes that underwent testing for expression and integration by immunohistochemical analysis and blue native polyacrylamide gel electrophoresis. During serial follow-up, the animal eyes underwent fundus photography, optical coherence tomography, and multifocal or pattern electroretinography. We tested for rescue of visual loss in rodent eyes also injected with a mutant G11778A ND4 homologue responsible for most cases of LHON. EXPOSURE Ocular infection with recombinant adeno-associated viral vectors containing a wild-type allotopic human ND4 gene. MAIN OUTCOMES AND MEASURES Expression of human ND4 and rescue of optic neuropathy induced by mutant human ND4. RESULTS We found human ND4 expressed in almost all mouse retinal ganglion cells by 1 week after injection and ND4 integrated into the mouse complex I. In rodent eyes also injected with a mutant allotopic ND4, wild-type allotopic ND4 prevented defective adenosine triphosphate synthesis, suppressed visual loss, reduced apoptosis of retinal ganglion cells, and prevented demise of axons in the optic nerve. Injection of ND4 in the ex vivo human eye resulted in expression in most retinal ganglion cells. Primates undergoing vitreal injection with the ND4 test article and followed up for 3 months had no serious adverse reactions. CONCLUSIONS AND RELEVANCE Expression of our allotopic ND4 vector in the ex vivo human eye, safety of the test article, rescue of the LHON mouse model, and the severe irreversible loss of visual function in LHON support clinical testing with mutated G11778A mitochondrial DNA in our patients.
Bascom Palmer Eye Institute, Miller School of Medicine, University of Miami, Miami, Florida.
Rajeshwari D
RD
Koilkonda
Bascom Palmer Eye Institute, Miller School of Medicine, University of Miami, Miami, Florida.
Hong
H
Yu
Bascom Palmer Eye Institute, Miller School of Medicine, University of Miami, Miami, Florida.
Tsung-Han
TH
Chou
Bascom Palmer Eye Institute, Miller School of Medicine, University of Miami, Miami, Florida.
William J
WJ
Feuer
Bascom Palmer Eye Institute, Miller School of Medicine, University of Miami, Miami, Florida.
Marco
M
Ruggeri
Bascom Palmer Eye Institute, Miller School of Medicine, University of Miami, Miami, Florida.
Vittorio
V
Porciatti
Bascom Palmer Eye Institute, Miller School of Medicine, University of Miami, Miami, Florida.
David
D
Tse
Department of Ophthalmology, College of Medicine, University of Florida, Gainesville.
William W
WW
Hauswirth
Department of Ophthalmology, College of Medicine, University of Florida, Gainesville.
Vince
V
Chiodo
Department of Ophthalmology, College of Medicine, University of Florida, Gainesville.
Sanford L
SL
Boye
Department of Molecular Genetics and Microbiology, College of Medicine, University of Florida, Gainesville.
Alfred S
AS
Lewin
Division of Neuroscience, Oregon National Primate Research Center, Oregon Health and Science University, Beaverton, Oregon.
Martha
M
Neuringer
Division of Neuroscience, Oregon National Primate Research Center, Oregon Health and Science University, Beaverton, Oregon.
Lauren
L
Renner
Bascom Palmer Eye Institute, Miller School of Medicine, University of Miami, Miami, Florida.
John
J
Guy
D000818:Animals; D002986:Clinical Trials as Topic; D004272:DNA, Mitochondrial / Q000235:genetics; D000229:Dependovirus / Q000235:genetics*; D004195:Disease Models, Animal; D004596:Electroretinography; D005260:Female; D005786:Gene Expression Regulation / Q000502:physiology; D015316:Genetic Therapy / Q000009:adverse effects / Q000379:methods*; D005822:Genetic Vectors / Q000235:genetics*; D049452:Green Fluorescent Proteins / Q000235:genetics; D006801:Humans; D058449:Intravitreal Injections; D008253:Macaca mulatta; D008297:Male; D051379:Mice; D008811:Mice, Inbred DBA; D018613:Microscopy, Confocal; D008928:Mitochondria / Q000235:genetics / Q000473:pathology; D009245:NADH Dehydrogenase / Q000235:genetics* / Q000378:metabolism; D029242:Optic Atrophy, Hereditary, Leber / Q000235:genetics / Q000473:pathology / Q000628:therapy*; D020641:Polymorphism, Single Nucleotide; D051381:Rats; D012165:Retinal Ganglion Cells / Q000201:enzymology / Q000473:pathology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D041623:Tomography, Optical Coherence
D004272:DNA, Mitochondrial; C098483:NADH dehydrogenase subunit 4; D049452:Green Fluorescent Proteins; D009245:NADH Dehydrogenase