Publications by authors named "Guylene Le Meur"

31 Publications

Thresholds of glycemia, insulin therapy, and risk for severe retinopathy in premature infants: A cohort study.

PLoS Med 2020 12 11;17(12):e1003477. Epub 2020 Dec 11.

Department of Neonatal Medicine, Nantes University Hospital, Nantes, France.

Background: Hyperglycemia in preterm infants may be associated with severe retinopathy of prematurity (ROP) and other morbidities. However, it is uncertain which concentration of blood glucose is associated with increased risk of tissue damage, with little consensus on the cutoff level to treat hyperglycemia. The objective of our study was to examine the association between hyperglycemia and severe ROP in premature infants.

Methods And Findings: In 2 independent, monocentric cohorts of preterm infants born at <30 weeks' gestation (Nantes University Hospital, 2006-2016, primary, and Lyon-HFME University Hospital, 2009-2017, validation), we first analyzed the association between severe (stage 3 or higher) ROP and 2 markers of glucose exposure between birth and day 21-maximum value of glycemia (MaxGly1-21) and mean of daily maximum values of glycemia (MeanMaxGly1-21)-using logistic regression models. In both the primary (n = 863 infants, mean gestational age 27.5 ± 1.4 weeks, boys 52.5%; 38 with severe ROP; 54,083 glucose measurements) and the validation cohort (n = 316 infants, mean gestational age 27.4 ± 1.4 weeks, boys 51.3%), MaxGly1-21 and MeanMaxGly1-21 were significantly associated with an increased risk of severe ROP: odds ratio (OR) 1.21 (95% CI 1.14-1.27, p < 0.001) and OR 1.70 (95% CI 1.48-1.94, p < 0.001), respectively, in the primary cohort and OR 1.17 (95% CI 1.05-1.32, p = 0.008) and OR 1.53 (95% CI 1.20-1.95, p < 0.001), respectively, in the validation cohort. These associations remained significant after adjustment for confounders in both cohorts. Second, we identified optimal cutoff values of duration of exposure above each concentration of glycemia between 7 and 13 mmol/l using receiver operating characteristic curve analyses in the primary cohort. Optimal cutoff values for predicting stage 3 or higher ROP were 9, 6, 5, 3, 2, 2, and 1 days above a glycemic threshold of 7, 8, 9, 10, 11, 12, and 13 mmol/l, respectively. Severe exposure was defined as at least 1 exposure above 1 of the optimal cutoffs. Severe ROP was significantly more common in infants with severe exposure in both the primary (10.9% versus 0.6%, p < 0.001) and validation (5.2% versus 0.9%, p = 0.030) cohorts. Finally, we analyzed the association between insulin therapy and severe ROP in a national population-based prospectively recruited cohort (EPIPAGE-2, 2011, n = 1,441, mean gestational age 27.3 ± 1.4, boys 52.5%) using propensity score weighting. Insulin use was significantly associated with severe ROP in overall cohort crude analyses (OR 2.51 [95% CI 1.13-5.58], p = 0.024). Adjustment for inverse propensity score (gestational age, sex, birth weight percentile, multiple birth, spontaneous preterm birth, main pregnancy complications, surfactant therapy, duration of oxygen exposure between birth and day 28, digestive state at day 7, caloric intake at day 7, and highest glycemia during the first week) and duration of oxygen therapy had a large but not significant effect on the association between insulin treatment and severe ROP (OR 0.40 [95% CI 0.13-1.24], p = 0.106). Limitations of this study include its observational nature and, despite the large number of patients included compared to earlier similar studies, the lack of power to analyze the association between insulin use and retinopathy.

Conclusions: In this study, we observed that exposure to high glucose concentration is an independent risk factor for severe ROP, and we identified cutoff levels that are significantly associated with increased risk. The clinical impact of avoiding exceeding these thresholds to prevent ROP deserves further evaluation.
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http://dx.doi.org/10.1371/journal.pmed.1003477DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7732100PMC
December 2020

Pathogenic variants in cause autosomal dominant and autosomal recessive retinitis pigmentosa.

J Med Genet 2020 Aug 17. Epub 2020 Aug 17.

Institute for Neurosciences of Montpellier, University of Montpellier, Montpellier, France

Background: Inherited retinal disorders are a clinically and genetically heterogeneous group of conditions and a major cause of visual impairment. Common disease subtypes include vitelliform macular dystrophy (VMD) and retinitis pigmentosa (RP). Despite the identification of over 90 genes associated with RP, conventional genetic testing fails to detect a molecular diagnosis in about one third of patients with RP.

Methods: Exome sequencing was carried out for identifying the disease-causing gene in a family with autosomal dominant RP. Gene panel testing and exome sequencing were performed in 596 RP and VMD families to identified additional variants. In vivo analysis in the medaka fish system by knockdown assays was performed to screen possible pathogenic role.

Results: Exome sequencing of a family with RP revealed a splice variant in . Subsequently, the same variant was identified in individuals from two families with either RP or VMD. A retrospective study of patients with RP or VMD revealed eight additional families with different missense or nonsense variants in . In addition, the clinical diagnosis of the retinopathy-associated variant, originally described as benign concentric annular macular dystrophy, was also revised to RP with early macular involvement. Using morpholino-mediated ablation of and its paralog in medaka fish, we confirmed a phenotype consistent with that observed in the families, including a decreased length of rod and cone photoreceptor outer segments.

Conclusion: This study discusses a previously unreported association between monoallelic or biallelic variants and RP. Notably, similar observations have been reported for .
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http://dx.doi.org/10.1136/jmedgenet-2020-107150DOI Listing
August 2020

Predictive values of initial semi-quantitative assessment of relative afferent pupillary defect for neovascularization in central retinal vein occlusion.

Acta Ophthalmol 2020 Jul 23. Epub 2020 Jul 23.

Department of Ophthalmology, University Hospital of Nantes, Nantes, France.

Purpose: To measure the predictive values of relative afferent pupillary defect (RAPD) assessed semi-quantitatively, and visual acuity (VA) at onset of central retinal vein occlusion (CRVO), for neovascularization.

Methods: Retrospective analysis of the TROXHEMO trial that included patients with CRVO within 30 days after the onset. Inclusion criteria were as follows: semi-quantitative RAPD assessment at diagnosis and/or at one month. RAPD was 'severe' if ≥ 0.9 log. Exclusion criteria were as follows: prophylactic panretinal photocoagulation (PRP) before neovascularization.

Results: Among the 119 patients enrolled in the main centre, 101 were analysed. 26 had a neovascular complication during the twelve months of follow-up: rubeosis (19), glaucoma (7) and posterior neovascularization (15). The mean time to onset of a neovascular complication was 4.7 months (1 to 12, median 3 months). All the patients who had a neovascular complication had RAPD at first examination or at one month (negative predictive value (NPV) = 100%) but the positive predictive value (PPV) was low (31%, 95% CI [21%; 42%]). The association 'severe RAPD or VA < 35 letters (ETDRS) at inclusion or at one month' was the best compromise between PPV (53%, [39%; 68%]) and NPV (96%, [92%; 100%]).

Conclusion: To predict neovascularization, RAPD should be routinely evaluated with filters: the risk of neovascular complication is (a) almost nil if there is no RAPD, (b) very low if there is no severe RAPD and if VA is higher than 35 letters, and (c) higher than 50% if RAPD is ≥ 0.9 log or if VA is less than 35 letters.
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http://dx.doi.org/10.1111/aos.14544DOI Listing
July 2020

[Gene therapy for retinitis pigmentosa].

Med Sci (Paris) 2020 Jun-Jul;36(6-7):607-615. Epub 2020 Jul 2.

Centre hospitalier universitaire de Nantes, Nantes Université, service d'ophtalmologie, 1 place Alexis Ricordeau, 44093 Nantes, France - Inserm UMR 1089, thérapie génique translationnelle des maladies génétiques, IRS 2 - Nantes Biotech, 22 boulevard Benoni Goullin, 44200 Nantes, France.

Retinitis pigmentosa is the most common blinding inherited retinal dystrophy. Gene therapy is a burgeoning revolutionary approach that paves the way to treatment of previously incurable diseases. At the end of 2017 and 2018, a gene therapy, Luxturna, obtained a marketing authorization by respectively the FDA (Food and Drug Administration) and the EMA (European Medicines Agency). This treatment, with proven efficacy, is available to patients with Leber congenital amaurosis and retinitis pigmentosa associated with bi-allelic mutations of the RPE 65 gene. In this paper, we present the current advances in gene therapy for retinitis pigmentosa and discuss the technological, economic and ethical challenges to overcome for gene therapy to improve medical practices.
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http://dx.doi.org/10.1051/medsci/2020095DOI Listing
October 2020

Mutations in the Kinesin-2 Motor KIF3B Cause an Autosomal-Dominant Ciliopathy.

Am J Hum Genet 2020 06 7;106(6):893-904. Epub 2020 May 7.

CHU Nantes, Service de Génétique Médicale, 9 quai Moncousu, 44093 Nantes Cedex 1, France; Université de Nantes, CNRS, INSERM, l'institut du thorax, 44000 Nantes, France. Electronic address:

Kinesin-2 enables ciliary assembly and maintenance as an anterograde intraflagellar transport (IFT) motor. Molecular motor activity is driven by a heterotrimeric complex comprised of KIF3A and KIF3B or KIF3C plus one non-motor subunit, KIFAP3. Using exome sequencing, we identified heterozygous KIF3B variants in two unrelated families with hallmark ciliopathy phenotypes. In the first family, the proband presents with hepatic fibrosis, retinitis pigmentosa, and postaxial polydactyly; he harbors a de novo c.748G>C (p.Glu250Gln) variant affecting the kinesin motor domain encoded by KIF3B. The second family is a six-generation pedigree affected predominantly by retinitis pigmentosa. Affected individuals carry a heterozygous c.1568T>C (p.Leu523Pro) KIF3B variant segregating in an autosomal-dominant pattern. We observed a significant increase in primary cilia length in vitro in the context of either of the two mutations while variant KIF3B proteins retained stability indistinguishable from wild type. Furthermore, we tested the effects of KIF3B mutant mRNA expression in the developing zebrafish retina. In the presence of either missense variant, rhodopsin was sequestered to the photoreceptor rod inner segment layer with a concomitant increase in photoreceptor cilia length. Notably, impaired rhodopsin trafficking is also characteristic of recessive KIF3B models as exemplified by an early-onset, autosomal-recessive, progressive retinal degeneration in Bengal cats; we identified a c.1000G>A (p.Ala334Thr) KIF3B variant by genome-wide association study and whole-genome sequencing. Together, our genetic, cell-based, and in vivo modeling data delineate an autosomal-dominant syndromic retinal ciliopathy in humans and suggest that multiple KIF3B pathomechanisms can impair kinesin-driven ciliary transport in the photoreceptor.
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http://dx.doi.org/10.1016/j.ajhg.2020.04.005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7273529PMC
June 2020

Macular Fold Complicating a Subretinal Injection of Voretigene Neparvovec.

Ophthalmol Retina 2020 04 13;4(4):456-458. Epub 2019 Dec 13.

Centre Hospitalier Universitaire de Nantes, Nantes Université, Service d'Ophtalmologie, Nantes, France.

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http://dx.doi.org/10.1016/j.oret.2019.12.005DOI Listing
April 2020

MOG-Ab prevalence in optic neuritis and clinical predictive factors for diagnosis.

Br J Ophthalmol 2020 06 3;104(6):842-845. Epub 2019 Oct 3.

Ophtalmologie, Centre Hospitalier Universitaire de Nantes, Nantes, France.

Objective: What is the proportion of antibodies to myelin oligodendrocyte glycoprotein (MOG-Ab) in optic neuritis (ON) in adults and what would be the ON presentation for which MOG-Ab should be tested?

Methods: Multicentric prospective study conducted during 1 year on all patients diagnosed with acute ON in all ophthalmological units in hospitals in a region in western France.

Results: Sixty-five patients were included. MOG-Ab prevalence was 14% (9/65) during an acute ON and 13% (7/55) after exclusion of patients already diagnosed with multiple sclerosis (MS) (8) or MOG+ON (2). Compared with MS and clinically isolated syndrome, MOG+ON had no female preponderance (67% of men in case of MOG+ON and 22% of men in case of MS and clinically isolated syndrome, p<0.05) were more often bilateral (44% vs 3%, p0.005) and associated with optic disc swelling (ODS) (78% vs 14%, p0.001). To predict MOG+ON, the positive predictive values (PPVs) of male sex, ODS and bilateral involvement were 29% (95% CI 9% to 48%), 41% (95% CI 18% to 65%) and 40% (95% CI 10% to 70%), respectively, while the negative predictive values (NPV) were 93% (95% CI 86% to 100%), 96% (95% CI 90% to 100%) and 91% (95% CI 83% to 99%), respectively. The combined factor 'ODS or bilateral or recurrent ON' was the best compromise between PPV (31% (95% CI 14% to 48%)) and NPV (100% (95% CI 100% to 100%)).

Conclusion: Among ON episodes, MOG-Ab were found in 14% of cases. MOG+ON occurred without female preponderance and was significantly associated with ODS and/or bilateral ON. Testing MOG-Ab only in patients presenting with ODS or bilateral or recurrent ON would limit MOG-Ab tests to fewer than half of all patients without the risk of missing any MOG+ON cases.
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http://dx.doi.org/10.1136/bjophthalmol-2019-314845DOI Listing
June 2020

Variable expressivity of syndromic BMP4-related eye, brain, and digital anomalies: A review of the literature and description of three new cases.

Eur J Hum Genet 2019 09 3;27(9):1379-1388. Epub 2019 May 3.

Center for Individualized Medicine, Mayo Clinic, Rochester, MN, USA.

Microphthalmia with brain and digital anomalies (MCOPS6, MIM# 607932) is an autosomal dominant disorder caused by loss-of-function variants or large deletions involving BMP4, which encodes bone morphogenetic protein 4, a member of the TGF-β protein superfamily. BMP4 has a number of roles in embryonic development including neurogenesis, lens induction, development of cartilage and bone, urogenital development, limb and digit patterning, hair follicle regeneration, as well as tooth formation. In addition to syndromic microphthalmia, BMP4 variants have been implicated in non-syndromic cleft lip with or without cleft palate and congenital healed cleft lip indicating different allelic presentations. MCOPS6 subjects may also lack some of the major phenotypic hallmarks of the disorder, including microphthalmia, indicating variable expressivity. As only a handful of individuals with MCOPS6 have been described, we review the clinical findings in previously reported cases with either deletions or loss-of-function variants in BMP4. We describe three new cases, including two subjects with novel deletions and one subject with a likely pathogenic de novo nonsense variant [c.1052C>G, p.(S351*)] in BMP4. One of the subjects had dual molecular diagnoses including a co-occurring microdeletion at 17q21.31 associated with Koolen de Vries syndrome, which has a partially overlapping disease phenotype. None of these individuals had clinically apparent microphthalmia or anopthalmia, which have been reported in a majority of previously described cases. One subject had exophthalmia and strabismus, while another had bilateral Peters anomaly and sclerocornea, thus expanding the phenotype associated with BMP4 loss-of-function variants.
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http://dx.doi.org/10.1038/s41431-019-0423-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6777538PMC
September 2019

Prospective evaluation of anesthetic protocols during pediatric ophthalmic surgery.

Eur J Ophthalmol 2019 Nov 3;29(6):606-614. Epub 2018 Oct 3.

Department of Ophthalmology, University Hospital of Nantes, Nantes, France.

Purpose: To date, no protocol of anesthesia for pediatric ophthalmic surgery is unanimously recognized. The primary anesthetic risks are associated with strabismus surgery, including oculocardiac reflex, postoperative nausea and vomiting, and postoperative pain.

Methods: This was a prospective, monocentric, observational study conducted in a tertiary pediatric ophthalmic unit. Our anesthetic protocol for strabismus surgery included postoperative nausea and vomiting prevention using dexamethasone and ondansetron. No drug-based prevention of oculocardiac reflex or local/locoregional anesthesia was employed.

Results: A total of 106 pediatric ophthalmic surgeries completed between November 2015 and May 2016 were analyzed. The mean patient age was 4.4 (range: 0.2-7.3, standard deviation: 2.4) years. Ambulatory rate was 90%. Oculocardiac reflex incidence was 65% during strabismus surgery (34/52), 50% during congenital cataract surgery (4/8), 33% during intramuscular injection of botulinum toxin (1/3), and 0% during other procedures. No asystole occurred. Postoperative nausea and vomiting incidence was 9.6% after strabismus surgery (5/52) and 0% following the other procedures. One child was hospitalized for one night because of persistent postoperative nausea and vomiting. Postoperative pain generally occurred early on in the recovery room and was quickly controlled. Its incidence was higher in patients who underwent strabismus surgery (27%) than in those who underwent other procedures (9%).

Conclusion: Morbidity associated with ophthalmic pediatric surgery is low and predominantly associated with strabismus surgery. The benefit-risk ratio and cost-effectiveness of oculocardiac reflex prevention should be questioned. Our postoperative nausea and vomiting rate is low, thanks to the use of a well-managed multimodal strategy. Early postoperative pain is usually well-treated but could probably be more effectively prevented.
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http://dx.doi.org/10.1177/1120672118804798DOI Listing
November 2019

Expanding the phenotype of the X-linked BCOR microphthalmia syndromes.

Hum Genet 2019 Sep 4;138(8-9):1051-1069. Epub 2018 Jul 4.

UDEAR, UMR 1056 Inserm, Université de Toulouse, Toulouse, France.

Two distinct syndromes arise from pathogenic variants in the X-linked gene BCOR (BCL-6 corepressor): oculofaciocardiodental (OFCD) syndrome, which affects females, and a severe microphthalmia ('Lenz'-type) syndrome affecting males. OFCD is an X-linked dominant syndrome caused by a variety of BCOR null mutations. As it manifests only in females, it is presumed to be lethal in males. The severe male X-linked recessive microphthalmia syndrome ('Lenz') usually includes developmental delay in addition to the eye findings and is caused by hypomorphic BCOR variants, mainly by a specific missense variant c.254C > T, p.(Pro85Leu). Here, we detail 16 new cases (11 females with 4 additional, genetically confirmed, affected female relatives; 5 male cases each with unaffected carrier mothers). We describe new variants and broaden the phenotypic description for OFCD to include neuropathy, muscle hypotonia, pituitary underdevelopment, brain atrophy, lipoma and the first description of childhood lymphoma in an OFCD case. Our male X-linked recessive cases show significant new phenotypes: developmental delay (without eye anomalies) in two affected half-brothers with a novel BCOR variant, and one male with high myopia, megalophthalmos, posterior embryotoxon, developmental delay, and heart and bony anomalies with a previously undescribed BCOR splice site variant. Our female OFCD cases and their affected female relatives showed variable features, but consistently had early onset cataracts. We show that a mosaic carrier mother manifested early cataract and dental anomalies. All female carriers of the male X-linked recessive cases for whom genetic confirmation was available showed skewed X-inactivation and were unaffected. In view of the extended phenotype, we suggest a new term of X-linked BCOR-related syndrome.
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http://dx.doi.org/10.1007/s00439-018-1896-xDOI Listing
September 2019

Safety and Long-Term Efficacy of AAV4 Gene Therapy in Patients with RPE65 Leber Congenital Amaurosis.

Mol Ther 2018 01 19;26(1):256-268. Epub 2017 Sep 19.

Ophthalmology Department, University Hospital Centre (CHU) de Nantes, Nantes, France; INSERM UMR 1089, University of Nantes, CHU de Nantes, Nantes France.

The aim of this study was the evaluation of the safety and efficacy of unilateral subretinal injection of the adeno-associated vector (AAV) serotypes 2 and 4 (AAV2/4) RPE65-RPE65 vector in patients with Leber congenital amaurosis (LCA) associated with RPE65 gene deficiency. We evaluated ocular and general tolerance and visual function up to 1 year after vector administration in the most severely affected eye in nine patients with retinal degeneration associated with mutations in the RPE65 gene. Patients received either low (1.22 × 10 to 2 × 10 vector genomes [vg]) or high (between 3.27 × 10 and 4.8 × 10 vg) vector doses. An ancillary study, in which six of the original nine patients participated, extended the follow-up period to 2-3.5 years. All patients showed good ophthalmological and general tolerance to the rAAV2/4-RPE65-RPE65 vector. We observed a trend toward improved visual acuity in patients with nystagmus, stabilization and improvement of the visual field, and cortical activation along visual pathways during fMRI analysis. OCT analysis after vector administration revealed no retinal thinning, except in cases of macular detachment. Our findings show that the rAAV2/4.RPE65.RPE65 vector was well tolerated in nine patients with RPE65-associated LCA. Efficacy parameters varied between patients during follow-up.
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http://dx.doi.org/10.1016/j.ymthe.2017.09.014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5763029PMC
January 2018

Vitrectomy Before Intravitreal Injection of AAV2/2 Vector Promotes Efficient Transduction of Retinal Ganglion Cells in Dogs and Nonhuman Primates.

Hum Gene Ther Methods 2016 06;27(3):122-34

1 Atlantic Gene Therapies, INSERM UMR 1089, Université de Nantes, CHU de Nantes, Nantes, France.

Recombinant adeno-associated virus (AAV) has emerged as a promising vector for retinal gene delivery to restore visual function in certain forms of inherited retinal dystrophies. Several studies in rodent models have shown that intravitreal injection of the AAV2/2 vector is the optimal route for efficient retinal ganglion cell (RGC) transduction. However, translation of these findings to larger species, including humans, is complicated by anatomical differences in the eye, a key difference being the comparatively smaller volume of the vitreous chamber in rodents. Here, we address the role of the vitreous body as a potential barrier to AAV2/2 diffusion and transduction in the RGCs of dogs and macaques, two of the most relevant preclinical models. We intravitreally administered the AAV2/2 vector carrying the CMV-eGFP reporter cassette in dog and macaque eyes, either directly into the vitreous chamber or after complete vitrectomy, a surgical procedure that removes the vitreous body. Our findings suggest that the vitreous body appears to trap the injected vector, thus impairing the diffusion and transduction of AAV2/2 to inner retinal neurons. We show that vitrectomy before intravitreal vector injection is an effective means of overcoming this physical barrier, improving the transduction of RGCs in dog and macaque retinas. These findings support the use of vitrectomy in clinical trials of intravitreal gene transfer techniques targeting inner retinal neurons.
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http://dx.doi.org/10.1089/hgtb.2016.034DOI Listing
June 2016

AAV-mediated Gene Therapy Halts Retinal Degeneration in PDE6β-deficient Dogs.

Mol Ther 2016 05 9;24(5):867-76. Epub 2016 Feb 9.

Atlantic Gene Therapies, INSERM UMR 1089, Université de Nantes, CHU de Nantes, Nantes, France.

We previously reported that subretinal injection of AAV2/5 RK.cpde6β allowed long-term preservation of photoreceptor function and vision in the rod-cone dysplasia type 1 (rcd1) dog, a large animal model of naturally occurring PDE6β deficiency. The present study builds on these earlier findings to provide a detailed assessment of the long-term effects of gene therapy on the spatiotemporal pattern of retinal degeneration in rcd1 dogs treated at 20 days of age. We analyzed the density distribution of the retinal layers and of particular photoreceptor cells in 3.5-year-old treated and untreated rcd1 dogs. Whereas no rods were observed outside the bleb or in untreated eyes, gene transfer halted rod degeneration in all vector-exposed regions. Moreover, while gene therapy resulted in the preservation of cones, glial cells and both the inner nuclear and ganglion cell layers, no cells remained in vector-unexposed retinas, except in the visual streak. Finally, the retinal structure of treated 3.5-year-old rcd1 dogs was identical to that of unaffected 4-month-old rcd1 dogs, indicating near complete preservation. Our findings indicate that gene therapy arrests the degenerative process even if intervention is initiated after the onset of photoreceptor degeneration, and point to significant potential of this therapeutic approach in future clinical trials.
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http://dx.doi.org/10.1038/mt.2016.37DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4881772PMC
May 2016

High prevalence of PRPH2 in autosomal dominant retinitis pigmentosa in france and characterization of biochemical and clinical features.

Am J Ophthalmol 2015 Feb 5;159(2):302-14. Epub 2014 Nov 5.

Institut National de la Santé et de la Recherche Médicale, U1051, Institute for Neurosciences of Montpellier, Montpellier, France; University of Montpellier 1, Montpellier, France; University of Montpellier 2, Montpellier, France; CHRU, Genetics of Sensory Diseases, Montpellier, France.

Purpose: To assess the prevalence of PRPH2 in autosomal dominant retinitis pigmentosa (adRP), to report 6 novel mutations, to characterize the biochemical features of a recurrent novel mutation, and to study the clinical features of adRP patients.

Design: Retrospective clinical and molecular genetic study.

Methods: Clinical investigations included visual field testing, fundus examination, high-resolution spectral-domain optical coherence tomography (OCT), fundus autofluorescence imaging, and electroretinogram (ERG) recording. PRPH2 was screened by Sanger sequencing in a cohort of 310 French families with adRP. Peripherin-2 protein was produced in yeast and analyzed by Western blot.

Results: We identified 15 mutations, including 6 novel and 9 previously reported changes in 32 families, accounting for a prevalence of 10.3% in this adRP population. We showed that a new recurrent p.Leu254Gln mutation leads to protein aggregation, suggesting abnormal folding. The clinical severity of the disease in examined patients was moderate with 78% of the eyes having 1-0.5 of visual acuity and 52% of the eyes retaining more than 50% of the visual field. Some patients characteristically showed vitelliform deposits or macular involvement. In some families, pericentral RP or macular dystrophy were found in family members while widespread RP was present in other members of the same families.

Conclusions: The mutations in PRPH2 account for 10.3% of adRP in the French population, which is higher than previously reported (0%-8%) This makes PRPH2 the second most frequent adRP gene after RHO in our series. PRPH2 mutations cause highly variable phenotypes and moderate forms of adRP, including mild cases, which could be underdiagnosed.
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http://dx.doi.org/10.1016/j.ajo.2014.10.033DOI Listing
February 2015

Frequency and clinical pattern of vitelliform macular dystrophy caused by mutations of interphotoreceptor matrix IMPG1 and IMPG2 genes.

Ophthalmology 2014 Dec 29;121(12):2406-14. Epub 2014 Jul 29.

Centre de Référence Maladies Sensorielles Génétiques, Hôpital Gui de Chauliac, Montpellier, France; Montpellier University, Montpellier, France; Institute for Neurosciences, INSERM, Montpellier, France.

Purpose: To assess the frequency of and to characterize the clinical spectrum and optical coherence tomography findings of vitelliform macular dystrophy linked to IMPG1 and IMPG2, 2 new causal genes expressed in the interphotoreceptor matrix.

Design: Retrospective epidemiologic, clinical, electrophysiologic, and molecular genetic study.

Participants: The database of a national referral center specialized in genetic sensory diseases was screened for patients with a macular vitelliform dystrophy without identified mutation or small deletion or large rearrangement in BEST1 and PRPH2 genes. Forty-nine families were included.

Methods: Clinical, imaging, and electro-oculogram findings were reviewed. Mutation screening of IMPG1 and IMPG2 genes were performed systematically.

Main Outcome Measures: Frequency, inheritance, and clinical pattern of vitelliform dystrophy associated with IMPG1 and IMPG2 mutations were characterized.

Results: IMPG1 was the causal gene in 3 families (IMPG1 1-3, 11 patients) and IMPG2 in a fourth family (2 patients). With an autosomal dominant transmission, families 1 and 2 had the c.713T→G (p.Leu238Arg) mutation in IMPG1 and family 4 had the c.3230G→T (p.Cys1077Phe) mutation in IMPG2. Patients with IMPG1 or IMPG2 mutations had a late onset and moderate visual impairment (mean visual acuity, 20/40; mean age of onset, 42 years), even in the sporadic case of family 3 with a presumed recessive transmission (age at onset, 38 years; mean visual acuity, 20/50). Drusen-like lesions adjacent to the vitelliform deposits were observed in 9 of 13 patients. The vitelliform material was above the retinal pigment epithelium (RPE) at any stage of the macular dystrophy, and this epithelium was well preserved and maintained its classical reflectivity on spectral-domain optical coherence tomography (SD-OCT). Electro-oculogram results were normal or borderline in 9 cases.

Conclusions: IMPG1 and IMPG2 are new causal genes in 8% of families negative for BEST1 and PRPH2 mutations. These genes should be screened in adult-onset vitelliform dystrophy with (1) moderate visual impairment, (2) drusen-like lesions, (3) normal reflectivity of the RPE line on SD-OCT, and (4) vitelliform deposits located between ellipsoid and interdigitation lines on SD-OCT. These clinical characteristics are not observed in the classical forms of BEST1 or PRPH2 vitelliform dystrophies.
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http://dx.doi.org/10.1016/j.ophtha.2014.06.028DOI Listing
December 2014

Successful gene therapy in the RPGRIP1-deficient dog: a large model of cone-rod dystrophy.

Mol Ther 2014 Feb 4;22(2):265-277. Epub 2013 Oct 4.

INSERM UMR 1089, Institut de Recherche Thérapeutique 1, Université de Nantes, Nantes, France. Electronic address:

For the development of new therapies, proof-of-concept studies in large animal models that share clinical features with their human counterparts represent a pivotal step. For inherited retinal dystrophies primarily involving photoreceptor cells, the efficacy of gene therapy has been demonstrated in canine models of stationary cone dystrophies and progressive rod-cone dystrophies but not in large models of progressive cone-rod dystrophies, another important cause of blindness. To address the last issue, we evaluated gene therapy in the retinitis pigmentosa GTPase regulator interacting protein 1 (RPGRIP1)-deficient dog, a model exhibiting a severe cone-rod dystrophy similar to that seen in humans. Subretinal injection of AAV5 (n = 5) or AAV8 (n = 2) encoding the canine Rpgrip1 improved photoreceptor survival in transduced areas of treated retinas. Cone function was significantly and stably rescued in all treated eyes (18-72% of those recorded in normal eyes) up to 24 months postinjection. Rod function was also preserved (22-29% of baseline function) in four of the five treated dogs up to 24 months postinjection. No detectable rod function remained in untreated contralateral eyes. More importantly, treatment preserved bright- and dim-light vision. Efficacy of gene therapy in this large animal model of cone-rod dystrophy provides great promise for human treatment.
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http://dx.doi.org/10.1038/mt.2013.232DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3918913PMC
February 2014

Mutations in IMPG1 cause vitelliform macular dystrophies.

Am J Hum Genet 2013 Sep 29;93(3):571-8. Epub 2013 Aug 29.

INSERM U1051, Institute for Neurosciences of Montpellier, Université Montpellier 1, Montpellier, France.

Vitelliform macular dystrophies (VMD) are inherited retinal dystrophies characterized by yellow, round deposits visible upon fundus examination and encountered in individuals with juvenile Best macular dystrophy (BMD) or adult-onset vitelliform macular dystrophy (AVMD). Although many BMD and some AVMD cases harbor mutations in BEST1 or PRPH2, the underlying genetic cause remains unknown for many affected individuals. In a large family with autosomal-dominant VMD, gene mapping and whole-exome sequencing led to the identification of a c.713T>G (p.Leu238Arg) IMPG1 mutation, which was subsequently found in two other families with autosomal-dominant VMD and the same phenotype. IMPG1 encodes the SPACR protein, a component of the rod and cone photoreceptor extracellular matrix domains. Structural modeling indicates that the p.Leu238Arg substitution destabilizes the conserved SEA1 domain of SPACR. Screening of 144 probands who had various forms of macular dystrophy revealed three other IMPG1 mutations. Two individuals from one family affected by autosomal-recessive VMD were homozygous for the splice-site mutation c.807+1G>T, and two from another family were compound heterozygous for the mutations c.461T>C (p.Leu154Pro) and c.1519C>T (p.Arg507(∗)). Most cases had a normal or moderately decreased electrooculogram Arden ratio. We conclude that IMPG1 mutations cause both autosomal-dominant and -recessive forms of VMD, thus indicating that impairment of the interphotoreceptor matrix might be a general cause of VMD.
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http://dx.doi.org/10.1016/j.ajhg.2013.07.018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3769927PMC
September 2013

Exudative retinopathy, cerebral calcifications, duodenal atresia, preaxial polydactyly, micropenis, microcephaly and short stature: a new syndrome?

Am J Med Genet A 2013 Aug 3;161A(8):1829-32. Epub 2013 Jul 3.

CHU Nantes, Service de Génétique Médicale, Nantes, France; INSERM, UMR-S 957, 1 Rue Gaston Veil, Nantes, France.

The association of Coats disease with intrauterine growth retardation, intracranial calcification, leukodystrophy, brain cysts, osteopenia, and gastrointestinal bleeding defines Coats plus syndrome caused by mutations in the CTC1 gene, encoding conserved telomere maintenance component 1. Here, we report on a child with exudative retinopathy, cerebral calcifications, duodenal atresia, preaxial polydactyly, micropenis, microcephaly, and short stature, in whom no mutations in CTC1 were found. Our patient shares some features seen in other diseases associated with telomere shortening including Hoyeraal-Hreidarsson and Revesz syndromes. We therefore measured telomere length by Flow-Fish which was normal. The association of duodenal atresia and microcephaly also suggested a diagnosis of Feingold syndrome. However, direct sequencing of MYCN was normal, and we did not detect any hemizygous deletion of the miR-17∼92 polycistronic miRNA cluster. To our knowledge, the phenotype we report on has not been described previously, leading us to speculate that this condition may represent a new syndrome.
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http://dx.doi.org/10.1002/ajmg.a.36021DOI Listing
August 2013

Maculopathy and spinocerebellar ataxia type 1: a new association?

J Neuroophthalmol 2013 Sep;33(3):225-31

Department of Ophthalmology, Nantes University Hospital, Nantes, France.

Background: Autosomal dominant cerebellar ataxia is a rare heterogeneous group of diseases characterized by cerebellar symptoms, often associated with other multisystemic signs. Mild optic neuropathy has been associated with spinocerebellar ataxia type 1 (SCA1), but macular dysfunction has been reported in only 2 cases. We report the first family with SCA1 with several members affected by visual loss related to maculopathy.

Methods: Cross-sectional clinical and electrophysiological study of a family with genetically confirmed SCA1. Patients with unexplained visual loss were included.

Results: Four patients from the same family, carrying the same genetic mutation, were examined. Testing revealed an increased CAG trinucleotide repeat number within the SCA1 gene. Genetic testing results for SCA7 were negative. Visual acuities ranged between 20/20 and 20/200. Visual fields revealed central scotomas in most of the eyes, and funduscopy was unremarkable in most patients. Central retinal thinning of the retina or disorganized photoreceptor layers were found with optical coherence tomography (OCT). In one patient, multifocal electroretinography (mfERG) revealed central retinal dysfunction.

Conclusions: A clinically subtle or even occult maculopathy can occur in association with SCA1. Macular OCT and mfERG can be abnormal even in asymptomatic patients. Unexplained visual loss in SCA1 patients should prompt evaluation of macular function, even if clinical signs of maculopathy are absent.
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http://dx.doi.org/10.1097/WNO.0b013e31828d4addDOI Listing
September 2013

Determinants of ocular deviation in esotropic subjects under general anesthesia.

J Pediatr Ophthalmol Strabismus 2013 May-Jun;50(3):155-60. Epub 2013 Mar 5.

Department of Ophthalmology, Hôpital Gui de Chauliac, Montpellier, France.

Purpose: The authors attempted to identify the determinants of ocular deviation in a population of patients with esotropia under general anesthesia.

Methods: Forty-one patients with esotropia were included. Horizontal ocular deviation was evaluated by the photographic Hirschberg test both in the awakened state and under general anesthesia before surgery. Changes in ocular deviation were measured and a multivariate analysis was used to assess its clinical determinants.

Results: The mean age (± standard deviation [SD]) of study subjects was 13 ± 11 years and 51% were females. The mean spherical equivalent refraction of the right eye was 2.44 ± 2.50 diopters (D), with no significant difference between eyes (P = .26). The mean ocular deviation changed significantly, from 33.5 ± 12.5 prism diopters (PD) at preoperative examination to 8.8 ± 11.4 PD under general anesthesia (P = .0001). The changes in ocular deviation positively correlated with the pre-operative ocular deviation (correlation coefficient r = 0.59, P = .0001) and negatively correlated with patient age (correlation coefficient r = -0.53, P = .0001). These two determinants remained significant after multivariate adjustment of the following variables: preoperative ocular deviation; age; gender; spherical equivalent refraction; and number of previous strabismus surgeries (model r(2) = 0.49, P = .0001).

Conclusions: The ocular position under general anesthesia was reported as a key factor in the surgical treatment of subjects with esotropia; therefore, its clinical determinants were assessed. The authors observed that preoperative ocular deviation and patient age were the main factors that influenced the ocular position under general anesthesia.
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http://dx.doi.org/10.3928/01913913-20130226-01DOI Listing
March 2014

Restoration of vision in the pde6β-deficient dog, a large animal model of rod-cone dystrophy.

Mol Ther 2012 Nov 24;20(11):2019-30. Epub 2012 Jul 24.

Translational Gene Therapy for Retinal and Neuromuscular Diseases, INSERM UMR 1089, Institut de Recherche Thérapeutique 1, Université de Nantes, Nantes, France.

Defects in the β subunit of rod cGMP phosphodiesterase 6 (PDE6β) are associated with autosomal recessive retinitis pigmentosa (RP), a childhood blinding disease with early retinal degeneration and vision loss. To date, there is no treatment for this pathology. The aim of this preclinical study was to test recombinant adeno-associated virus (AAV)-mediated gene addition therapy in the rod-cone dysplasia type 1 (rcd1) dog, a large animal model of naturally occurring PDE6β deficiency that strongly resembles the human pathology. A total of eight rcd1 dogs were injected subretinally with AAV2/5RK.cpde6β (n = 4) or AAV2/8RK.cpde6β (n = 4). In vivo and post-mortem morphological analysis showed a significant preservation of the retinal structure in transduced areas of both AAV2/5RK.cpde6β- and AAV2/8RK.cpde6β-treated retinas. Moreover, substantial rod-derived electroretinography (ERG) signals were recorded as soon as 1 month postinjection (35% of normal eyes) and remained stable for at least 18 months (the duration of the study) in treated eyes. Rod-responses were undetectable in untreated contralateral eyes. Most importantly, dim-light vision was restored in all treated rcd1 dogs. These results demonstrate for the first time that gene therapy effectively restores long-term retinal function and vision in a large animal model of autosomal recessive rod-cone dystrophy, and provide great promise for human treatment.
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http://dx.doi.org/10.1038/mt.2012.134DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3498794PMC
November 2012

Whole-exome sequencing identifies mutations in GPR179 leading to autosomal-recessive complete congenital stationary night blindness.

Am J Hum Genet 2012 Feb;90(2):321-30

Institut National de la Santé et de la Recherche Médicale, Paris, France.

Congenital stationary night blindness (CSNB) is a heterogeneous retinal disorder characterized by visual impairment under low light conditions. This disorder is due to a signal transmission defect from rod photoreceptors to adjacent bipolar cells in the retina. Two forms can be distinguished clinically, complete CSNB (cCSNB) or incomplete CSNB; the two forms are distinguished on the basis of the affected signaling pathway. Mutations in NYX, GRM6, and TRPM1, expressed in the outer plexiform layer (OPL) lead to disruption of the ON-bipolar cell response and have been seen in patients with cCSNB. Whole-exome sequencing in cCSNB patients lacking mutations in the known genes led to the identification of a homozygous missense mutation (c.1807C>T [p.His603Tyr]) in one consanguineous autosomal-recessive cCSNB family and a homozygous frameshift mutation in GPR179 (c.278delC [p.Pro93Glnfs(∗)57]) in a simplex male cCSNB patient. Additional screening with Sanger sequencing of 40 patients identified three other cCSNB patients harboring additional allelic mutations in GPR179. Although, immunhistological studies revealed Gpr179 in the OPL in wild-type mouse retina, Gpr179 did not colocalize with specific ON-bipolar markers. Interestingly, Gpr179 was highly concentrated in horizontal cells and Müller cell endfeet. The involvement of these cells in cCSNB and the specific function of GPR179 remain to be elucidated.
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http://dx.doi.org/10.1016/j.ajhg.2011.12.007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3276675PMC
February 2012

Regulation of retinal function but nonrescue of vision in RPE65-deficient dogs treated with doxycycline-regulatable AAV vectors.

Mol Ther 2010 Jun 30;18(6):1085-93. Epub 2010 Mar 30.

Laboratoire de Thérapie Génique, INSERM UMR U649, Institut de Recherche Thérapeutique 1, Université de Nantes, Nantes, France.

In previous studies, we demonstrated that recombinant adeno-associated virus (rAAV)-mediated gene transfer of the doxycycline (Dox)-regulatable system allows for the regulation of erythropoietin (EPO) expression in the retina of nonhuman primates after intravenous or oral administration of Dox. In addition, it was shown that administrating different amounts of Dox resulted in a dose-response dynamic of transgene expression. Adeno-associated viral gene therapy has raised hope for the treatment of patients with Leber congenital amaurosis, caused by mutations in the retinal pigment epithelium (RPE)-specific gene RPE65. The preliminary results of three clinical trials suggest some improvement in visual function. However, further improvements might be necessary to optimize vision recovery and this means developing vectors able to generate transgene expression at physiological levels. The purpose of this study was to investigate the ability of the Dox-regulatable system to regulate retinal function in RPE65(-/-) Briard dogs. rAAV vectors expressing RPE65 under the control of either the TetOff and TetOn Dox-regulated promoters or the cytomegalovirus (CMV) constitutive promoter were generated and administered subretinally to seven RPE65-deficient dogs. We demonstrate that the induction and deinduction of retinal function, as assessed by electroretinography (ERG), can be achieved using a Dox-regulatable system, but do not lead to any recovery of vision.
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http://dx.doi.org/10.1038/mt.2010.46DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2889751PMC
June 2010

Detection of intact rAAV particles up to 6 years after successful gene transfer in the retina of dogs and primates.

Mol Ther 2009 Mar 23;17(3):516-23. Epub 2008 Dec 23.

INSERM U649, CHU-Hotel Dieu, Nantes, France.

Gene transfer to the retina using recombinant adeno-associated viral (rAAV) vectors has proven to be an effective option for the treatment of retinal degenerative diseases in several animal models and has recently advanced into clinical trials in humans. To date, intracellular trafficking of AAV vectors and subsequent capsid degradation has been studied only in vitro, but the fate of AAV particles in transduced cells following subretinal injection has yet to be elucidated. Using electron microscopy and western blot, we analyzed retinas of one primate and four dogs that had been subretinally injected with AAV2/4, -2/5, or -2/2 serotypes and that displayed efficient gene transfer over several years. We show that intact AAV particles are still present in retinal cells, for up to 6 years after successful gene transfer in these large animals. The persistence of intact vector particles in the target organ, several years postadministration, is totally unexpected and, therefore, represents a new and unanticipated safety issue to consider at a time when gene therapy clinical trials raise new immunological concerns.
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http://dx.doi.org/10.1038/mt.2008.283DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2835079PMC
March 2009

Subretinal delivery of recombinant AAV serotype 8 vector in dogs results in gene transfer to neurons in the brain.

Mol Ther 2008 May 11;16(5):916-23. Epub 2008 Mar 11.

INSERM U649, CHU Hotel-Dieu, Nantes, France.

Recombinant adeno-associated virus (rAAV) vectors are among the most efficient gene delivery vehicles for gene transfer to the retina. This study evaluates the behavior of the rAAV8 serotype vector with regard to intraocular delivery in rats and dogs. Subretinal delivery of an AAV2/8.gfp vector results in efficient gene transfer in the retinal pigment epithelium (RPE), the photoreceptors and, surprisingly, in the cells of the inner nuclear layer as well as in ganglion cells. Most importantly, in dogs, gene transfer also occurred distal to the injection site in neurons of the lateral geniculate nucleus of the brain. Because green fluorescent protein (GFP) was detected along the visual pathway within the brain, we analyzed total DNA extracted from various brain slices using PCR. Vector sequences were detected in many parts of the brain, but chiefly in the contralateral hemisphere.
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http://dx.doi.org/10.1038/mt.2008.41DOI Listing
May 2008

Gene therapeutic prospects in early onset of severe retinal dystrophy: restoration of vision in RPE65 Briard dogs using an AAV serotype 4 vector that specifically targets the retinal pigmented epithelium.

Bull Mem Acad R Med Belg 2006 ;161(10-12):497-508; discussion 508-9

Université de Nantes.

Previous studies have tested gene replacement therapy in RPE65 deficient dogs using recombinant adeno-associated virus 2/2 (rAAV2/2), -2/1 or -2/5 mediated delivery of the RPE65 gene. They all documented restoration of dark- and light-adapted ERG responses and improved psychophysical outcomes. Use of a specific RPE65 promoter and a rAAV vector that targets transgene expression specifically to the retinal pigmented epithelium (RPE) may, however, provide a safer setting for the long-term therapeutic expression of RPE65. Subretinal injection of rAAV2 pseudotyped with serotype 4 (rAAV2/4) specifically targets the RPE. The purpose of our study was to evaluate a rAAV2/4 vector carrying a human RPE65cDNA driven by a human RPE65 promoter, for the ability to restore vision in RPE65-/- purebred Briard dogs. Recombinant rAAV2/4 and rAAV2/2 vectors containing similar human RPE65 promoter and cDNA cassettes were generated and administered subretinally in 8 affected dogs, ages 8 to 30 months (n = 6 with rAAV2/4, n = 2 with rAAV2/2). Although fluorescein angiography and OCT examinations displayed retinal abnormalities in treated retinas, electrophysiological analysis demonstrated that restoration of rod and cone photoreceptor function started as soon as 15 days post-injection, reaching maximal function at 3 months post-injection, and remaining stable thereafter in all animals treated at 8 to 11 months of age. As assessed by the ability of these animals to avoid obstacles in both dim and normal light, functional vision was restored in the treated eye, while the untreated contralateral eye served as an internal control. The dog treated at a later age (30 months) did not recover retinal function or vision, suggesting that there might be a therapeutic window for the successful treatment of RPE65 -/- dogs by gene replacement therapy.
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June 2007

Long-term doxycycline-regulated transgene expression in the retina of nonhuman primates following subretinal injection of recombinant AAV vectors.

Mol Ther 2006 May 25;13(5):967-75. Epub 2006 Jan 25.

INSERM UMR U649, CHU-Hotel Dieu, Bât. J. Monnet, 30 Avenue J. Monnet, 44035 Nantes Cedex 01, France.

Adeno-associated viral gene therapy has shown promise for the treatment of inherited and acquired retinal disorders. In most applications, regulation of expression is a critical concern for both safety and efficacy. The purpose of our study was to evaluate the ability of the tetracycline-regulatable system to establish long-term transgene regulation in the retina of nonhuman primates. Three rAAV vectors expressing the tetracycline-dependent transactivator (rtTA) under the control of either the ubiquitous CAG promoter or the specific RPE65 promoter (AAV2/5.CAG.TetOn.epo, AAV2/4.CAG.TetOn.epo, and AAV2/4.RPE65.TetOn.epo) were generated and administered subretinally to seven macaques. We demonstrated that repeated inductions of transgene expression in the nonhuman primate retina can be achieved using a Tet-inducible system via rAAV vector administration over a long period (2.5 years). Maximum erythropoietin (EPO) secretion in the anterior chamber depends upon the rAAV serotype and the nature of the promoter driving rtTA expression. We observed that the EPO isoforms produced in the retina differ from one another based on the transduced cell type of origin within the retina and also differ from both the physiological EPO isoforms and the isoforms produced by AAV-transduced skeletal muscle.
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http://dx.doi.org/10.1016/j.ymthe.2005.12.001DOI Listing
May 2006

Postsurgical assessment and long-term safety of recombinant adeno-associated virus-mediated gene transfer into the retinas of dogs and primates.

Arch Ophthalmol 2005 Apr;123(4):500-6

Institut National de la Santé et de la Récherche Médicale UMR U649, 44035 Nantes, France.

Objective: To evaluate, in dogs and primates, the short-term effects of subretinal injection and the safety of long-term recombinant adeno-associated virus (rAAV)-mediated transgene expression with respect to retinal morphology and function.

Methods: Subretinal delivery of rAAV (serotype 2, 4, or 5) was performed unilaterally in 14 beagles and 9 macaques. Postsurgical condition was evaluated during a 2-month follow-up study. Three dogs and 1 primate were examined for the long-term study. Green fluorescent protein expression was monitored by fluorescent retinal imaging. Retinal anatomy and function were assessed by angiography and electroretinography, respectively.

Results: Transgene expression was observed in 20 of 23 subretinally injected animals (both with and without vitrectomy). We did not detect an inflammatory response in any of the 23 treated subjects. In the long-term study, transgene expression was detected at the latest points evaluated: 36 months for the rAAV-2-injected dog, 24 months for the rAAV-4 and rAAV-5 dogs, and more than 18 months for the rAAV-4-injected primate. Angiography examinations were performed and showed no retinal abnormalities. Functional evaluation showed normal electroretinographic amplitude responses that were similar to those of the noninjected contralateral eyes.

Conclusions: Subretinal injection of the rAAV vector in dogs and primates is a safe procedure with no perioperative complications and a high rate of successful retinal gene transfer. The retinal anatomy and function remained unchanged, despite persistent transgene expression up to 36 months postinjection with rAAV-2, -4, or -5. Additionally, we observed no other adverse effects, such as tumor formation due to possible insertional mutagenesis. These short- and long-term studies on rAAV transgene expression using large animals are encouraging for the prospects of ocular gene therapy applications in humans.

Clinical Relevance: These short- and long-term studies on rAAV transgene expression using large animals are encouraging for the prospects of ocular gene therapy applications in humans.
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http://dx.doi.org/10.1001/archopht.123.4.500DOI Listing
April 2005

Biodistribution of rAAV vectors following intraocular administration: evidence for the presence and persistence of vector DNA in the optic nerve and in the brain.

Mol Ther 2005 Feb;11(2):275-83

INSERM UMR U649, CHU-Hotel DIEU, Bâtiment J. Monnet, 30 Avenue J. Monnet, 44035 Nantes Cedex 01, France.

The purpose of our study was to evaluate the biodistribution of rAAV vectors following subretinal or intravitreal injection. In rats, we performed subretinal or intravitreal injections of rAAV-2/2.CMV.gfp. In large animals, rAAV-2/4.CMV.gfp or rAAV-2/5.CMV.gfp was delivered into the subretinal space while rAAV-2/2.CMV.gfp was delivered either to the subretinal space or to the vitreous. In euthanized animals, we undertook a complete necropsy. In animals maintained alive, we collected blood and tissue samples from the submandibular lymph node, liver, and gonads. We analyzed total DNA, extracted from various tissue samples and peripheral blood mononuclear cells (PBMC), by PCR. Following subretinal or intravitreal injections in rats and in large animals, vector sequences were not detected in the liver or in the gonads but were occasionally found in PBMC. An unexpected result was the detection of rAAV sequences in the optic nerve following subretinal injection. The most striking finding was the detection of vector sequences in the brain, along the visual pathway, in rAAV-2/2 intravitreally injected dogs. These findings raise safety concerns regarding intraocular administration of rAAV vectors and will have an impact on the development of future gene therapy trials for retinal diseases.
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http://dx.doi.org/10.1016/j.ymthe.2004.09.022DOI Listing
February 2005