Bidot, S

Blouin, L

Bouquet, C

Combal, JP

Fitoussi, S

Galy, A

Gilly, B

Girmens, JF

Katz, B

Meunier, S

Sahel, JA

Thomasson, N

Uretsky, S

Valero, S

Vignal, C

UCL Discovery

ReportsSafety of rAAV2/2-ND4 GeneTherapy for Leber HereditaryOptic NeuropathyLeber hereditary optic neuropathy (LHON) is the most commonlyrecognized mitochondrial disease. It typically occurs in young maleadults, causing painless, acute, and profound vision loss. It presentsasynchronously with the second eye almost always involved withinweeks or months, a phenotypic declaration nearly pathognomonicfor LHON. Visual prognosis is poor and therapy wanting.1Leber hereditary optic neuropathy is caused by mutations inmitochondrial genes encoding proteins of the respiratory chaincomplex I. Approximately 70% of subjects with LHON carry thepoint mutation G11778A in the ND4 gene encoding NADHdehydrogenase protein subunit 4 (ND4), accounting for the mostsevere phenotype.2 Retinal ganglion cells are primarily affected bythis mitochondrial dysfunction, leading to apoptotic cell death andensuing optic nerve atrophy.1Our therapy is based on a technology that demonstrably rescuedan induced defect in respiratory chain complex I in rat retinas andTable 1. Ocular Treatment-Emergent Adverse Events and OutcomrAA9  109 vg/eye(n ¼ 3)3  1010 vg/e(n ¼ 3)All Ocular TEAEs 4/3 10/3Anterior chamber inﬂammation 0 2/2Subconjunctival hemorrhage 0 0Allergic conjunctivitis 1/1 0Punctate serous detachment 0 0Eye pain 0 1/1Keratitis 0 1/1Ocular hypertension 2/2 3/3Vitreous hemorrhage 0 0Vitritis 1/1 3/2Positive Seidel test 0 0Cataract extraction (elective) 1/1 0Outcome of the 3 Most Common Ocular TEAEsAnterior chamber inﬂammationRecovered 0 2/2Ongoing 0 0VitritisRecovered 0 3/2Ongoing 1/1 0Ocular hypertensionRecovered 2/2 3/3Ongoing 0 0TEAE ¼ treatment-emergent adverse event; VG ¼ vector genomes.Results are presented as N events/N subjects.*Includes subjects of cohorts 3 and 5.yTwo events were considered unrelated to rAAV2/2-ND4 or the procedure; thrAAV2/2-ND4.zAll events were considered to be probably related to the procedure.xOf 10 events, 6 were considered to be related to rAAV2/2-ND4, and 4 werekSubject withdrew consent after week 48.#All events were considered to be probably related to rAAV2/2-ND4.prevented retinal ganglion cell degeneration.3 rAAV2/2-ND4(GS010) is a recombinant, replication-defective, adeno-associatedvirus, serotype 2, containing a modiﬁed cDNA encoding thehuman wild-type mitochondrial ND4 protein and supporting itsallotopic expression,3 that is, the expression of a mitochondrialgene by the nucleus and subsequent co-translation and importa-tion of the wild-type protein into the mitochondrial matrix. rAAV2/2-ND4 was investigated in an open-label single-center Phase I/IIclinical trial that included 4 dose-escalation cohorts (9  109 vectorgenomes [vg]/eye, 3  1010 vg/eye, 9  1010 vg/eye, 1.8  1011vg/eye) and an extension cohort (9  1010 vg/eye). Fifteen subjectswith LHON carrying the ND4-G11778A mutation were prospec-tively enrolled. Each subject received a single intravitreal injectionof rAAV2/2-ND4 in the worse-seeing eye. The study designincluded an initial follow-up period of 48 weeks, followed bylonger-term follow-up for an additional 4 years. The primaryobjective was the safety and tolerability of escalating doses ofrAAV2/2-ND4. Secondary objectives included bio-disseminationand immunogenicity of rAAV2/2-ND4 and evaluation of visualfunctions. The study received approval of the French Ethicses of the Most Common Treatment-Emergent Adverse EventsV2/2-ND4 dosesAll (N [ 15)ye 9  1010 vg/eye(n ¼ 6)*1.8  1011 vg/eye(n ¼ 3)29/6 11/3 56/1510/6 2/2 14/10y3/3 0 3/3z0 0 1/10 1/1 1/1#1/1 0 2/2#4/4 2/2 7/72/2 3/2 10/9x2/2 0 2/26/5 3/3 13/11#1/1 0 1/11/1 0 2/210/6 2/2 14/100 0 06/5 3/3 12/100 0 1/1k2/2 3/2 10/90 0 0e remaining 12 events were considered to be probably related torelated to the procedure.945Ophthalmology Volume 125, Number 6, June 2018Committee and adhered to the tenets of the Declaration of Helsinki;it was registered on Clinicaltrial.gov (NCT02064569).Subjects were mostly male (n ¼ 13) with an average age of 47years. Vision loss durationwas heterogeneous, ranging from8 to 271months at enrollment (Table S1, available at www.aaojournal.org).The incidence, type, severity, and presumed causality of treatment-emergent adverse events (TEAEs) were collected for each subjectat all study visits. Each event was recorded as a separate adverseevent even when occurring simultaneously (e.g., anterior and vitre-ous inﬂammation). Visual parameters measured included best-corrected visual acuity (BCVA), Pelli-Robson contrast sensitivity,15-hue color vision, Humphrey Visual Field 24-2, Octopus peri-metry, microperimetry, visual evoked potentials, and patternelectroretinogram.At week 96, 96 TEAEs were reported involving all 15 subjects,including 40 systemic TEAEs and 56 ocular TEAEs, consistentwith previous studies using intravitreal injections (Table 1).4,5There were no unexpected TEAEs, no serious adverse eventsrelated to the treatment or procedure, and no suspected unexpectedserious adverse reactions. No deaths and no TEAEs leading tostudy discontinuation were reported. Ninety of the 96 TEAEs(94%) were mild in intensity. Of the 56 ocular TEAEs, 2 eventsoccurred in 1 untreated eye and 2 were elective cataract extraction.Thirty-four (61%) and 18 (32%) TEAEs were considered treatmentand procedure related, respectively. Fifty-one ocular TEAEs (91%)were mild. One moderate event of intraocular pressure (IOP)elevation (34 mmHg) occurred in the only subject who did notreceive a pretreatment IOP-lowering agent. Another subjectconcomitantly experienced an event of ocular pain and moderateelevation of IOP (38 mmHg), followed by 2 severe events: anteriorchamber inﬂammation and vitritis.The most frequent ocular TEAEs were intraocular inﬂammationand IOP elevation. Twenty-seven events of intraocular inﬂamma-tion were reported in 13 subjects, starting from 7 to 541 daysposttreatment. All were mild, except 2 severe events that occurredin a single subject. All were deemed probably related to treatment,except for 2 TEAEs that occurred in 1 untreated eye (subject had ahistory of idiopathic uveitis). Most affected subjects were treatedfor their ocular inﬂammation with topical anti-inﬂammatoryagents. Two subjects were given oral steroids: the subject withsevere intraocular inﬂammation and another subject with mildvitritis.Ten events of IOP elevation following intravitreal injectionwere reported in 9 subjects, starting 4 hours to 30 days after in-jection. Most cases were mild, except for 2 subjects with moderateIOP elevation. Day-of-treatment IOP elevation was deemed to beprocedure related (due to the injection). Delayed IOP elevation wasdeemed treatment related, and such subjects developed subsequentintraocular inﬂammation, precipitating steroid use.All ocular events resolved spontaneously or after appropriatetherapy with IOP-lowering or anti-inﬂammatory therapy, exceptfor 1 subject with ongoing mild vitritis (0.5þ vitreous cell, novitreous haze, no treatment required at last visit), subsequently lostto follow-up but without documented worsening of TEAE. Atweek 96, no related TEAEs required ongoing treatment. Noanatomic sequelae were documented by fundus examination orspectral domain OCT, and no vision loss was reported due totreatment or TEAEs. A total of 40 systemic TEAEs were reportedover 96 weeks; none were related to rAAV2/2-ND4 or the studyprocedures (Table S2, available at www.aaojournal.org).946Although this study was neither designed nor powered toascertain efﬁcacy, a clinically signiﬁcant improvement in BCVAwas noted in the treated eyes of 6 of 14 subjects at week 96 (similarproportions at week 48 and week 78). A between-eye difference invisual acuity change from baseline favoring the treated eye wasobserved at week 96 in the subset of subjects with disease duration2 years and BCVA 20/12000 at inclusion (0.278 logarithm ofthe minimum angle of resolution [logMAR] [þ14 Early TreatmentDiabetic Retinopathy Study {ETDRS} letters] 95% conﬁdenceinterval [CI], 0.853 to þ0.297). This between-eye difference wasalso observed at week 48 and week 78 (0.338 logMAR [þ17ETDRS letters] 95% CI, 0.856 to þ0.180 and 0.398 logMAR[þ20 ETDRS letters] 95% CI, 1.021 to þ0.225, respectively).Our study demonstrates that rAAV2/2-ND4 is safe and welltolerated 2 years after a single unilateral intravitreal administration.Our 2 ongoing Phase III clinical studies in similar subjects,RESCUE (vision loss duration of 6 months) and REVERSE(vision loss duration of >6 months to 1 year), should help reﬁneand inform the trends of improvement in visual outcome that wehave reported and further validate the safety and tolerability ofallotopic expression in mitochondrial disease.CATHERINE VIGNAL, MD1,2SCOTT URETSKY, MD3SERGE FITOUSSI, MD3ANNE GALY, PHD3LAURE BLOUIN, MSC3JEAN-FRANÇOIS GIRMENS, MD1SAMUEL BIDOT, MD1,2NITZA THOMASSON, PHD3CÉLINE BOUQUET, PHD3SONIA VALERO, PHARMD3SANDRINE MEUNIER, MSC3JEAN-PHILIPPE COMBAL, PHD3BERNARD GILLY, PHD3BARRETT KATZ, MD, MBA3,4JOSÉ-ALAIN SAHEL, MD1,2,5,61Centre d’Investigation Clinique 1423, Inserm-DGOS, CentreHospitalier National d’Ophtalmologie des Quinze-Vingts, Paris,France; 2Rothschild Ophthalmology Foundation Hospital, Paris,France; 3GenSight Biologics, Paris, France; 4GenSight Biologics Inc.,New York, New York; 5Sorbonne Université, INSERM, CNRS, Institutde la Vision, Paris, France; 6Department of Ophthalmology, Universityof Pittsburgh School of Medicine, University of Pittsburgh MedicalCenter, Pittsburgh, PennsylvaniaPresented in part at: North American Neuro-Ophthalmology Societyconference in Big Island, Hawaii, March 3e8, 2018; and the Associa-tion for Research in Vision and Ophthalmology conference to be held inHonolulu, Hawaii, April 29 to May 3, 2018.Financial Disclosure(s): The author(s) have made the following dis-closure(s): S.U., S.F., A.G., L.B., C.B., S.V., S.M., B.G., B.K.: Em-ployees e GenSight Biologics.C.V., J-A.S., S.B.: Consultants e GenSight Biologics.J-A.S.: Funding e LabEx LIFESENSES (ANR-10-LABX-65), ERCSynergy “HELMHOLTZ”, Banque publique d’Investissement, andFoundation Fighting Blindness; Cofounder e GenSight Biologics.C.V.: Grants and personal fees e GenSight Biologics.J-F.G.: Grants and personal fees e GenSight Biologics; Personal fees eNovartis, Bayer, Allergan, Sanoﬁ.ReportsS.B.: Consultant e Santhera Pharmaceuticals (Liesta, Switzerland).N.T. and J.P.C.: other e Former employee of GenSight Biologics.GenSight Biologics (Paris, France) fully funded and sponsored thestudy.The sponsor participated in the design, data analysis, and interpretationof the results, and in the preparation, review, and approval of themanuscript.HUMAN SUBJECTS: Human-derived materials and human medicalrecords were part of this study protocol. No animal subjects were used inthis study. Study protocol was approved by the French Ethics Committee.All studies adhered to the tenets of the Declaration of Helsinki.The data sets generated and analyzed during the current study are notpublicly available because the study is still ongoing, but are availablefrom the corresponding author on reasonable request.Author Contributions:Conception and design: Vignal, Sahel, Uretsky, Fitoussi, Galy, Combal,Valero, Meunier, Thomasson, Bouquet, GillyData collection: Vignal, Sahel, Girmens, BidotAnalysis and interpretation: Vignal, Sahel, Blouin, Uretsky, Katz, FitoussiObtained funding: N/AOverall responsibility: Blouin, Uretsky, Bidot, KatzCorrespondence:Barrett Katz, MD, MBA, GenSight Biologics, 74 rue du Faubourg Saint-Antoine, 75012 Paris, France. E-mail: bkatz@gensight-biologics.com.References1. Rasool N, Lessell S, Cestari DM. Leber hereditary optic neu-ropathy: bringing the lab to the clinic. Semin Ophthalmol.2016;31:107e116.2. Yu-Wai-Man P, Turnbull DM, Chinnery PF. Leber hereditaryoptic neuropathy. J Med Genet. 2002;39:162e169.3. Cwerman-Thibault H, Augustin S, Lechauve C, et al. Nuclearexpression of mitochondrial ND4 leads to the protein assem-bling in complex I and prevents optic atrophy and visual loss.Mol Ther Methods Clin Dev. 2015;2:15003.4. Koilkonda RD, Yu H, Chou TH, et al. Safety and effects of thevector for the Leber hereditary optic neuropathy gene therapyclinical trial. JAMA Ophthalmol. 2014;132:409e420.5. Koilkonda RD, Yu H, Talla V, et al. LHON gene therapy vectorprevents visual loss and optic neuropathy induced by G11778Amutant mitochondrial DNA: biodistribution and toxicologyproﬁle. Invest Ophthalmol Vis Sci. 2014;55:7739e7753.A Randomized Trial of a Short34-Gauge Needle forIntravitreal InjectionsIntravitreal injections of antievascular endothelial growth factoragents have achieved excellent results1,2 and have become one ofthe most frequently performed intraocular procedures. Althoughintravitreal drug delivery of antievascular endothelial growthfactor agents is effective, patients often report discomfort duringthe procedure. A literature review in a previous study of the use ofa 33-gauge needle for intravitreal injections3 indicated that there isno consensus regarding the effect of the intravitreal needle size andperceived pain. Therefore, we conducted the current study toevaluate the effectiveness of the short 34-gauge needle on patientperceptions of pain during intravitreal injections.We performed a prospective, randomized, double-armed,single-blinded clinical trial to evaluate the usefulness of the short34-gauge needle for intravitreal injections compared with the30-gauge needle. This study was conducted according to the tenetsof the Declaration of Helsinki; the Aichi Medical UniversityHospital institutional review board approved the study, which isregistered at the University Hospital Medical Information NetworkClinical Trial site (http://www.umin.ac.jp/ctr/index-j.htm;UMIN000024903). After providing written informed consent, allpatients underwent ophthalmic examinations, including slit-lampand fundus examinations and intraocular pressure (IOP; TonorefII [Nidek Co., Ltd., Aichi, Japan]) and axial length (AL-Scan,Nidek Co., Ltd.) measurements.Patients were assigned randomly to an intravitreal injection eitherusing a 30-gauge needle (0.3 19 mm; Nipro, Inc., Osaka, Japan) or a34-gauge needle (0.18  8 mm; Pasny [Nanbu Plastics Co., Ltd., Shi-zuoka, Japan]; Fig S1, available at www.aaojournal.org). Eitherranibizumab (Lucentis; Genentech, South San Francisco, CA) 0.05 ml(0.5 mg) or aﬂibercept (Eylea; Regeneron Pharmaceuticals, Inc.,Tarrytown, NY) 0.05 ml (2 mg) was administered. We used a preﬁlledsyringe for ranibizumab and a 1-ml syringe for aﬂibercept; these wereattached to either a 30- or 34-gauge needle. Two surgeons (H.S. andH.O.) administered intravitreal injections in our outpatient ofﬁce.All eyes were anesthetized with a 2% lidocaine (Xylocaine;Aspen Japan K.K., Tokyo, Japan) eye drop and were sterilized with5% povidone iodine eye drops.4 A sterile lid speculum wasinserted, and all injections were performed at a straight angle viathe pars plana 3.5 to 4.0 mm posterior to the limbus in the uppertemporal or nasal quadrant. Immediately after injections, thesurgeons checked the hand movement visual acuity. The IOPwas measured while the patients were sitting before, immediatelyafter, and 20 minutes after the injection using an Icare PROtonometer (Tiolat Oy, Helsinki, Finland). If patients reportedvisual loss or the IOP became too elevated, a paracentesis wasconsidered to prevent central retinal artery occlusion.A research nurse asked the patients to rate their pain using anumeric rating scale ranging from 0 to 10,5 in which 0 indicated nopain and 10 indicated the most intense pain immediately after theinjection. The average of these scores was the primary outcome.The surgeons assessed the puncture resistance, reﬂux after injection,subconjunctival hemorrhage, and ocular movements during theintravitreal injections on a 0 (undetectable) to 1 (detectable) scale.Statistical analyses were performed using the SAS System soft-ware version 9.4 (SAS Institute, Cary, NC). A statistician (K.M.)performed all analyses and was blinded to the group assignments.TheManneWhitneyU test was used to analyze continuous variablesand the Fisher exact test was used for categorical variables.One hundred forty eyes of 110 consecutive patients wereenrolled. The baseline characteristics did not differ signiﬁcantlybetween the groups (Table S1, available at www.aaojournal.org).The short 34-gauge needle was associated with a signiﬁcantly(P < 0.0001) lower pain score than the 30-gauge needle (medianinterquartile range [IQR], 2 [3] vs. 4 [4], respectively; mean standard deviation, 1.91.64 vs. 42.53, respectively; Fig 2).The puncture resistance (0 vs. 45 times, respectively) and reﬂuxafter injection (1 vs. 22 times, respectively) were signiﬁcantly(P < 0.0001 for both comparisons) lower with the 34-gauge nee-dle; the subconjunctival hemorrhage (10 vs. 15 times, respectively;P ¼ 0.27) and ocular movement (3 vs. 7 times, respectively;P ¼ 0.20) scores did not differ signiﬁcantly between the groups.947

Safety of rAAV2/2-ND4 Gene Therapy for Leber Hereditary Optic Neuropathy

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Safety of rAAV2/2-ND4 Gene Therapy for Leber Hereditary Optic Neuropathy

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