Rajeshwari Koilkonda, Hong Yu, Venu Talla, Vittorio Porciatti, William J. Feuer, William W. Hauswirth, Vince Chiodo, Kirsten E. Erger, Sanford L. Boye, Alfred S. Lewin, Thomas J. Conlon, Lauren Renner, Martha Neuringer, Carol Detrisac, John Guy, LHON Gene Therapy Vector Prevents Visual Loss and Optic Neuropathy Induced by G11778A Mutant Mitochondrial DNA: Biodistribution and Toxicology Profile. Invest. Ophthalmol. Vis. Sci. 2014,55(12):7739-7753. doi: 10.1167/iovs.14-15388. Purpose.: To demonstrate safety and efficacy of allotopic human ND4 for treatment of a Leber's hereditary optic neuropathy (LHON) mouse model harboring the G11778A mitochondrial mutation. Methods.: We induced LHON in mice by intravitreal injection of mutant (G11778A) human ND4 DNA, responsible for most cases of LHON, that was directed to mitochondria using an AAV2 vector to which we appended a mitochondrial targeting sequence to the VP2 capsid. We then attempted rescue of visual loss using our test article (ScAAV2-P1ND4v2) containing a synthetic nuclear encoded G11778G ND4 gene that was allotopically expressed. Control mice either were uninjected or received AAV2-GFP or AAV2-mCherry. We performed RT-PCR and confocal microscopy at 2 weeks post injection. Pattern electroretinograms (PERGs), spectral-domain optical coherence tomography (SD-OCT), histology, and transmission electron microscopy (TEM) were performed. For toxicology and biodistribution studies, the test article was administered intravitreally to rats and rhesus macaques at different doses. Results.: Mutant and wild-type ND4 were efficiently expressed in the mitochondria of retinal ganglion cells (RGCs). Visual function assessed by serial PERGs and retinal structure by serial SD-OCT showed a significant rescue by the test article. Histology and ultrastructural analysis confirmed that loss of RGCs and demise of axons was prevented by ScAAV2-P1ND4v2. Rat and nonhuman primate biodistribution studies showed that vector spread outside the injected eye into spleen and lymph nodes was minimal. Histopathology of tissues and organs including the eyes was comparable to that of uninfected and saline-injected eyes. Conclusions.: Allotopically expressed wild-type ND4 prevents the phenotype induced by G11778A mitochondrial DNA with a toxicology profile acceptable for testing in a phase I clinical trial. Source.