Purchase this article with an account.
Rajeshwari D Koilkonda, Hong Yu, Huijun Yuan, Vittorio Porciatti, William W Hauswirth, Thomas J Conlon, Lauren Renner, Martha Neuringer, Carol Detrisac, John Guy; Gene Therapy For Mitochondrial Disease: Are We Ready?. Invest. Ophthalmol. Vis. Sci. 2014;55(13):836.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
To demonstrate safety and efficacy of allotopic human (h) ND4 (test article,TA) for treatment of LHON mouse model harboring G11778A mitochondrial (mt) mutation.
To induce optic neuropathy G11778A ND4 DNA was delivered to mouse retinal mitochondria by using modified scAAV, containing a second gene (mt encoded mCherry), both driven by single mt heavy strand promoter for expression visualization using CSLO. We then injected our TA, packaged in scAAV2-(Y444F+Y500F+Y730F) capsid mutant, into the vitreous. Control eyes received scAAV-GFP (n=20). RT PCR and confocal microscopy were perfomed at 2wks post injection (PI). Pattern electroretinograms (PERG), SD-OCT, histology and TEM were performed. We used the TA to infect cybrids (G11778A) to evaluate cell survival and ATP production relative to un-infected cells. Toxicology and Biodistribution. TA was administered intravitreally (iv) to rats at different doses (n=60) or to rhesus macaques at 2.46 x 1010 vg (n=3).
RT PCR analysis revealed the presence of hND4 transcripts exclusively in the ocular tissues. Confocal microscopy showed mt expression of hND4 in RGCs and mCherry was visualized in the retina of live mice using CSLO at 4mPI. PERG amplitudes showed a significant rescue in TA treated eyes at 1yr compared to controls, p=0.023, latencies showed less delay in rescued eyes compared to controls, p=0.009. SD-OCT showed significant rescue and preservation of inner retinal layers at 1yr, p=0.011. Light microscopy of retinal sections showed a significant rescue of RGCs in TA treated eyes compared to controls at 1yr PI, p=3.65x10-5. TEM revealed apoptotic RGCs with condensed chromatin in controls absent in TA treated eyes and ON axon counts for TA treated mice were 50% higher relative to control, p=0.00758. Cybrids infected with TA showed a significant increase in cell survival with greater rate of ATP synthesis relative to un-infected cells, p<0.05. Toxicology and Biodistribution. Showed no vector genomes in the ocular or non-ocular tissues except spleen and lymph nodes, no gross/ microscopic findings in the eye or any other body tissues of rodent and primates at 30 or 90d after iv injections indicates safety of the TA.
TA suppressed visual loss, demise of RGCs and ON axons induced by mutant ND4 delivered to mouse retinal mitochondria and improved ATP synthesis in cybrids. Rodent and primate toxicology studies suggests an acceptable profile of TA for human trials.
This PDF is available to Subscribers Only