Genersatztherapie bei genetisch bedingter Zapfenblindheit

Zusammenfassung

Achromatopsie ist eine autosomal-rezessiv vererbte Netzhauterkrankung, die durch den angeborenen kompletten Funktionsverlust der Zapfenphotorezeptoren charakterisiert ist. Etwa 80 % der Patienten weisen Mutationen in der α- oder β-Untereinheit (A3 oder B3) des cGMP-gesteuerten Kationenkanals CNG (cyclic nucleotide-gated channel) der Zapfenphotorezeptoren auf. Homolog zum humanen Krankheitsbild zeigen CNGA3-defiziente Mäuse einen zapfenspezifischen Funktionsverlust, der zur Degeneration der betroffenen Zapfenphotorezeptorzelle führt. Am Forschungsinstitut für Augenheilkunde in Tübingen gelang es nun erstmalig im Tiermodell für Achromatopsie ACHM2, eine Netzhauterkrankung umfassend zu therapieren. Im vorliegenden Artikel werden der auf subretinaler Injektion von rekombinanten Adeno-assoziierten Viren (rAAV) basierende Ansatz im Detail dargelegt und die eingesetzten nicht-invasiven Diagnostikmethoden ERG, SLO und OCT zur Erfolgs- und Qualitätskontrolle beschrieben. Der nachweisbare Therapieerfolg zeigt sich in einem Funktionsgewinn des Zapfensystems und in einer weiterführenden neuronalen Verarbeitung des retinalen Signals bis hin zu einem zapfendominierten Verhalten der behandelten Tiere. Die herausragenden Ergebnisse sind Ausgangspunkt für die erste humane Translation einer Gentherapie für Achromatopsie in Deutschland.

Abstract

Achromatopsia is an autosomal recessive inherited retinal disease caused by a complete loss of cone photoreceptor function. About 80 % of achromatopsia patients show mutations in the alpha or beta subunit (A3 and B3) of the cGMP controlled cation channel CNG (cyclic nucleotide-gated channel) of cone photoreceptors. Homologous to the human disease, CNGA3 deficient mice reveal a loss of cone specific functionality leading to degeneration of affected cone photoreceptors. The Institute for Ophthalmic Research in Tübingen has now succeeded in curing achromatopsia ACHM2 in an animal model. In this article, we explain the recombinant adeno-associated virus-based approach in detail. Furthermore, applied non-invasive diagnostic techniques for quality and success control, ERG, SLO and OCT, are described. The success of the therapy is indicated by a restored cone photoreceptor function as well as the neuronal processing of retinal signals resulting in a specific, cone-mediated behaviour. The outstanding results derived from the animal model are the starting point for the first human translation of a gene therapy for achromatopsia in Germany.

Schlüsselwörter

rAAV-basierte Genersatztherapie - nicht invasive Diagnostikmethoden - Achromatopsie - CNGA3-Kanalprotein - erbliche Netzhauterkrankungen

Key words

AAV-mediated gene therapy - non-invasive diagnostic methods - achromatopsia - CNGA3 channel protein - hereditary retinal diseases

• Literatur

• 1 Sohocki MM, Daiger SP, Bowne SJ et al. Prevalence of mutations causing retinitis pigmentosa and other inherited retinopathies. Hum Mutat 2001; 17: 42-51
• 2 Rattner A, Sun H, Nathans J. Molecular genetics of human retinal disease. Annu Rev Genet 1999; 33: 89-131
• 3 Berger W, Kloeckener-Gruissem B, Neidhardt J. The molecular basis of human retinal and vitreoretinal diseases. Prog Retin Eye Res 2010; 29: 335-375
• 4 Surace EM, Auricchio A. Versatility of AAV vectors for retinal gene transfer. Vision Res 2008; 48: 353-359
• 5 Thiadens AA, Phan TM, Zekveld-Vroon RC et al. Clinical course, genetic etiology, and visual outcome in cone and cone-rod dystrophy. Ophthalmology 2012; 119: 819-826
• 6 Francois J. Heredity in Ophthalmology. CV Mosby: St Louis; 1961
• 7 Sharpe LT, Nordby K. Total Colour Blindness: An Introduction. In: Hess RF, Sharpe LT, Nordby K, eds. Night Vision: basic, clinical and applied Aspects. Cambridge, UK: Cambridge University Press; 1990: 253-289
• 8 Sharpe LT, Stockman A, Jägle H et al. Opsin Genes, Cone Photopigments and Colour Blindness. In: Gegenfurtner KR, Sharpe LT, eds. Color Vision: From Genes to Perception. Cambridge, UK: Cambridge University Press; 1999
• 9 Park WL, Sunness JS. Red contact lenses for alleviation of photophobia in patients with cone disorders. Am J Ophthalmol 2004; 137: 774-775
• 10 Schornack M. Prescription and management of contact lenses in patients with monocular visual impairment. Optometry 2007; 78: 652-656
• 11 Young RS, Krefman RA, Fishman GA. Visual improvements with red-tinted glasses in a patient with cone dystrophy. Arch Ophthalmol 1982; 100: 268-271
• 12 Kohl S, Baumann B, Rosenberg T et al. Mutations in the cone photoreceptor G-protein alpha-subunit gene GNAT2 in patients with achromatopsia. Am J Hum Genet 2002; 71: 422-425
• 13 Aligianis IA, Forshew T, Johnson S et al. Mapping of a novel locus for achromatopsia (ACHM4) to 1p and identification of a germline mutation in the alpha subunit of cone transducin (GNAT2). J Med Genet 2002; 39: 656-660
• 14 Rosenberg T, Baumann B, Kohl S et al. Variant phenotypes of incomplete achromatopsia in two cousins with GNAT2 gene mutations. Invest Ophthalmol Vis Sci 2004; 45: 4256-4262
• 15 Ouechtati F, Merdassi A, Bouyacoub Y et al. Clinical and genetic investigation of a large Tunisian family with complete achromatopsia: identification of a new nonsense mutation in GNAT2 gene. J Hum Genet 2011; 56: 22-28
• 16 Chang B, Grau T, Dangel S et al. A homologous genetic basis of the murine cpfl1 mutant and human achromatopsia linked to mutations in the PDE6C gene. Proc Natl Acad Sci U S A 2009; 106: 19581-19586
• 17 Thiadens AA, den Hollander AI, Roosing S et al. Homozygosity mapping reveals PDE6C mutations in patients with early-onset cone photoreceptor disorders. Am J Hum Genet 2009; 85: 240-247
• 18 Kohl S, Coppieters F, Meire F et al. A nonsense mutation in PDE6H causes autosomal-recessive incomplete achromatopsia. Am J Hum Genet 2012; 91: 527-532
• 19 Wissinger B, Gamer D, Jägle H et al. CNGA3 mutations in hereditary cone photoreceptor disorders. Am J Hum Genet 2001; 69: 722-737
• 20 Kohl S, Marx T, Giddings I et al. Total colourblindness is caused by mutations in the gene encoding the alpha-subunit of the cone photoreceptor cGMP-gated cation channel. Nat Genet 1998; 19: 257-259
• 21 Sundin OH, Yang JM, Li Y et al. Genetic basis of total colourblindness among the Pingelapese islanders. Nat Genet 2000; 25: 289-293
• 22 Winick JD, Blundell ML, Galke BL et al. Homozygosity mapping of the Achromatopsia locus in the Pingelapese. Am J Hum Genet 1999; 64: 1679-1685
• 23 Milunsky A, Huang XL, Milunsky J et al. A locus for autosomal recessive achromatopsia on human chromosome 8q. Clin Genet 1999; 56: 82-85
• 24 Grau T, Artemyev NO, Rosenberg T et al. Decreased catalytic activity and altered activation properties of PDE6C mutants associated with autosomal recessive achromatopsia. Hum Mol Genet 2011; 20: 719-730
• 25 Kohl S, Varsanyi B, Antunes GA et al. CNGB3 mutations account for 50 % of all cases with autosomal recessive achromatopsia. Eur J Hum Genet 2005; 13: 302-308
• 26 Michaelides M, Aligianis IA, Ainsworth JR et al. Progressive cone dystrophy associated with mutation in CNGB3. Invest Ophthalmol Vis Sci 2004; 45: 1975-1982
• 27 Thiadens AA, Slingerland NW, Roosing S et al. Genetic etiology and clinical consequences of complete and incomplete achromatopsia. Ophthalmology 2009; 116: 1984-1989
• 28 Kohl S, Kitiratschky V, Papke M et al. Genes and mutations in autosomal dominant cone and cone-rod dystrophy. Adv Exp Med Biol 2012; 723: 337-343
• 29 Johnson S, Michaelides M, Aligianis IA et al. Achromatopsia caused by novel mutations in both CNGA3 and CNGB3. J Med Genet 2004; 41: e20
• 30 Kaupp UB, Seifert R. Cyclic nucleotide-gated ion channels. Physiol Rev 2002; 82: 769-824
• 31 Zhong H, Molday LL, Molday RS et al. The heteromeric cyclic nucleotide-gated channel adopts a 3A:1B stoichiometry. Nature 2002; 420: 193-198
• 32 Zagotta WN, Siegelbaum SA. Structure and function of cyclic nucleotide-gated channels. Annu Rev Neurosci 1996; 19: 235-263
• 33 Hofmann F, Biel M, Kaupp UB et al. International Union of Pharmacology. XLII. Compendium of voltage-gated ion channels: cyclic nucleotide-modulated channels. Pharmacol Rev 2003; 55: 587-589
• 34 Hofmann F, Biel M, Kaupp UB. International Union of Pharmacology. LI. Nomenclature and structure-function relationships of cyclic nucleotide-regulated channels. Pharmacol Rev 2005; 57: 455-462
• 35 Ali RR, Reichel MB, Thrasher AJ et al. Gene transfer into the mouse retina mediated by an adeno-associated viral vector. Hum Mol Genet 1996; 5: 591-594
• 36 Allocca M, Mussolino C, Garcia-Hoyos M et al. Novel adeno-associated virus serotypes efficiently transduce murine photoreceptors. J Virol 2007; 81: 11372-11380
• 37 Bainbridge JW, Mistry A, Schlichtenbrede FC et al. Stable rAAV-mediated transduction of rod and cone photoreceptors in the canine retina. Gene Ther 2003; 10: 1336-1344
• 38 Buch PK, Bainbridge JW, Ali RR. AAV-mediated gene therapy for retinal disorders: from mouse to man. Gene Ther 2008; 15: 849-857
• 39 Pang JJ, Lauramore A, Deng WT et al. Comparative analysis of in vivo and in vitro AAV vector transduction in the neonatal mouse retina: effects of serotype and site of administration. Vision Res 2008; 48: 377-385
• 40 Chen J, Tucker CL, Woodford B et al. The human blue opsin promoter directs transgene expression in short-wave cones and bipolar cells in the mouse retina. Proc Natl Acad Sci U S A 1994; 91: 2611-2615
• 41 Yang GS, Schmidt M, Yan Z et al. Virus-mediated transduction of murine retina with adeno-associated virus: effects of viral capsid and genome size. J Virol 2002; 76: 7651-7660
• 42 Glushakova LG, Timmers AM, Pang J et al. Human blue-opsin promoter preferentially targets reporter gene expression to rat s-cone photoreceptors. Invest Ophthalmol Vis Sci 2006; 47: 3505-3513
• 43 Komáromy AM, Alexander JJ, Cooper AE et al. Targeting gene expression to cones with human cone opsin promoters in recombinant AAV. Gene Ther 2008; 15: 1049-1055 Erratum in: Gene Ther 2008; 15: 1073; Glushakova LG [added]. Gene Ther 2011; 18: 1179
• 44 Lebherz C, Maguire A, Tang W et al. Novel AAV serotypes for improved ocular gene transfer. J Gene Med 2008; 10: 375-382
• 45 Smith RH. Adeno-associated virus integration: virus versus vector. Gene Therapy 2008; 15: 817-822
• 46 Schnepp BC, Jensen RL, Chen CL et al. Characterization of adeno-associated virus genomes isolated from human tissues. J Virol 2005; 79: 14793-14803
• 47 Grieger JC, Samulski RJ. Adeno-associated virus vectorology, manufacturing, and clinical applications. Methods Enzymol 2012; 507: 229-254
• 48 Surace EM, Auricchio A. Versatility of AAV vectors for retinal gene transfer. Vision Res 2008; 48: 353-359
• 49 Petrs-Silva H, Dinculescu A, Li Q et al. High-efficiency transduction of the mouse retina by tyrosine-mutant AAV serotype vectors. Mol Ther 2009; 17: 463-471
• 50 Allocca M, Doria M, Petrillo M et al. Serotype-dependent packaging of large genes in adeno-associated viral vectors results in effective gene delivery in mice. J Clin Invest 2008; 118: 1955-1964
• 51 Acland GM, Aguirre GD, Ray J et al. Gene therapy restores vision in a canine model of childhood blindness. Nat Genet 2001; 28: 92-95
• 52 Ali RR, Sarra GM, Stephens C et al. Restoration of photoreceptor ultrastructure and function in retinal degeneration slow mice by gene therapy. Nat Genet 2000; 25: 306-310
• 53 Bainbridge JW, Smith AJ, Barker SS et al. Effect of gene therapy on visual function in Leberʼs congenital amaurosis. N Engl J Med 2008; 358: 2231-2239
• 54 Cideciyan AV, Aleman TS, Boye SL et al. Human gene therapy for RPE65 isomerase deficiency activates the retinoid cycle of vision but with slow rod kinetics. Proc Natl Acad Sci U S A 2008; 105: 15112-15117
• 55 Hauswirth WW, Aleman TS, Kaushal S et al. Treatment of leber congenital amaurosis due to RPE65 mutations by ocular subretinal injection of adeno-associated virus gene vector: short-term results of a phase I trial. Hum Gene Ther 2008; 19: 979-990
• 56 Timmers AM, Zhang H, Squitieri A et al. Subretinal injections in rodent eyes: effects on electrophysiology and histology of rat retina. Mol Vis 2001; 7: 131-137
• 57 Johnson CJ, Berglin L, Chrenek MA et al. Technical brief: subretinal injection and electroporation into adult mouse eyes. Mol Vis 2008; 14: 2211-2226
• 58 Busskamp V, Duebel J, Balya D et al. Genetic reactivation of cone photoreceptors restores visual responses in retinitis pigmentosa. Science 2010; 329: 413-417
• 59 Price J, Turner D, Cepko C. Lineage analysis in the vertebrate nervous system by retrovirus-mediated gene transfer. Proc Natl Acad Sci U S A 1987; 84: 156-160
• 60 Schlichtenbrede FC, da Cruz L, Stephens C et al. Long-term evaluation of retinal function in Prph2Rd2/Rd2 mice following AAV-mediated gene replacement therapy. J Gene Med 2003; 5: 757-764
• 61 Mühlfriedel R, Michalakis S, Garrido MG et al. Optimized technique for subretinal injections in mice. Methods Mol Biol 2013; 935: 343-349
• 62 Liang FQ, Anand V, Maguire AM et al. Intraocular delivery of recombinant virus. Methods Mol Med 2001; 47: 125-139
• 63 Seeliger MW, Beck SC, Pereyra-Muñoz N et al. In vivo confocal imaging of the retina in animal models using scanning laser ophthalmoscopy. Vision Res 2005; 45: 3512-3519
• 64 Huber G, Beck SC, Grimm C et al. Spectral domain optical coherence tomography in mouse models of retinal degeneration. Invest Ophthalmol Vis Sci 2009; 50: 5888-5895
• 65 Fischer MD, Huber G, Beck SC et al. Noninvasive, in vivo assessment of mouse retinal structure using optical coherence tomography. PLoS One 2009; 4: e7507
• 66 Marmor MF, Fulton AB, Holder GE et al. International Society for Clinical Electrophysiology of Vision. MISCEV Standard for full-field clinical electroretinography (2008 update). Doc Ophthalmol 2009; 118: 69-77
• 67 Scholl HP, Zrenner E. Electrophysiology in the investigation of acquired retinal disorders. Surv Ophthalmol 2000; 45: 29-47
• 68 Dalke C, Löster J, Fuchs H et al. Electroretinography as a screening method for mutations causing retinal dysfunction in mice. Invest Ophthalmol Vis Sci 2004; 45: 601-609
• 69 Tanimoto N, Muehlfriedel RL, Fischer MD et al. Vision tests in the mouse: Functional phenotyping with electroretinography. Front Biosci (Landmark Ed) 2009; 14: 2730-2737
• 70 Douglas RM, Alam NM, Silver BD et al. Independent visual threshold measurements in the two eyes of freely moving rats and mice using a virtual-reality optokinetic system. Vis Neurosci 2005; 22: 677-684
• 71 Umino Y, Solessio E, Barlow RB. Speed, spatial, and temporal tuning of rod and cone vision in mouse. J Neurosci 2008; 28: 189-198
• 72 Alexander JJ, Umino Y, Everhart D et al. Restoration of cone vision in a mouse model of achromatopsia. Nat Med 2007; 13: 685-687
• 73 Deeb SS. Molecular genetics of color-vision deficiencies. Vis Neurosci 2004; 21: 191-196
• 74 Wiszniewski W, Lewis RA, Lupski JR. Achromatopsia: the CNGB3 p.T383fsX mutation results from a founder effect and is responsible for the visual phenotype in the original report of uniparental disomy 14. Hum Genet 2007; 121: 433-439
• 75 Carvalho LS, Xu J, Pearson RA et al. Long-term and age-dependent restoration of visual function in a mouse model of CNGB3-associated achromatopsia following gene therapy. Hum Mol Genet 2011; 20: 3161-3175
• 76 Aguirre GD, Rubin LF. Pathology of hemeralopia in the Alaskan malamute dog. Invest Ophthalmol 1974; 13: 231-235
• 77 Aguirre GD, Rubin LF. The electroretinogram in dogs with inherited cone degeneration. Invest Ophthalmol 1975; 14: 840-847
• 78 Rubin LF. Clinical features of hemeralopia in the adult Alaskan malamute. J Am Vet Med Assoc 1971; 158: 1696-1698
• 79 Sidjanin DJ, Lowe JK, McElwee JL et al. Canine CNGB3 mutations establish cone degeneration as orthologous to the human achromatopsia locus ACHM3. Hum Mol Genet 2002; 11: 1823-1833
• 80 Komáromy AM, Alexander JJ, Rowlan JS et al. Gene therapy rescues cone function in congenital achromatopsia. Hum Mol Genet 2010; 19: 2581-2593 Erratum in: Hum Mol Genet 2011; 20: 5024
• 81 Michalakis S, Mühlfriedel R, Tanimoto N et al. Restoration of cone vision in the CNGA3-/- mouse model of congenital complete lack of cone photoreceptor function. Mol Ther 2010; 18: 2057-2063
• 82 Biel M, Seeliger M, Pfeifer A et al. Selective loss of cone function in mice lacking the cyclic nucleotide-gated channel CNG3. Proc Natl Acad Sci U S A 1999; 96: 7553-7557
• 83 Michalakis S, Geiger H, Haverkamp S et al. Impaired opsin targeting and cone photoreceptor migration in the retina of mice lacking the cyclic nucleotide-gated channel CNGA3. Invest Ophthalmol Vis Sci 2005; 46: 1516-1524
• 84 Hawes NL, Wang X, Hurd RE. A point mutation in the Cnga3 gene causes cone photoreceptor function loss (Cpfl5) in mice. Paper presented at ARVO, E-Abstract 4579: 2006, Fort Lauderdale. 2006
• 85 Pang JJ, Deng WT, Dai X et al. AAV-mediated cone rescue in a naturally occurring mouse model of CNGA3-achromatopsia. PLoS One 2012; 7: e35250
• 86 Maguire AM, Simonelli F, Pierce EA et al. Safety and efficacy of gene transfer for Leberʼs congenital amaurosis. N Engl J Med 2008; 358: 2240-2248
• 87 Amado D, Mingozzi F, Hui D et al. Safety and efficacy of subretinal readministration of a viral vector in large animals to treat congenital blindness. Sci Transl Med 2010; 2: 21ra16
• 88 Jacobson SG, Cideciyan AV, Ratnakaram R et al. Gene therapy for leber congenital amaurosis caused by RPE65 mutations: safety and efficacy in 15 children and adults followed up to 3 years. Arch Ophthalmol 2012; 130: 9-24
• 89 Zhang S, Wu J, Wu X et al. Enhancement of rAAV2-mediated transgene expression in retina cells in vitro and in vivo by coadministration of low-dose chemotherapeutic drugs. Invest Ophthalmol Vis Sci 2012; 53: 2675-2684