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Self-complementary AAV-mediated gene therapy restores cone function and prevents cone degeneration in two models of Rpe65 deficiency

Abstract

To test whether fast-acting, self-complimentary (sc), adeno-associated virus-mediated RPE65 expression prevents cone degeneration and/or restores cone function, we studied two mouse lines: the Rpe65-deficient rd12 mouse and the Rpe65-deficient, rhodopsin null (‘that is, cone function-only’) Rpe65−/−::Rho−/− mouse. scAAV5 expressing RPE65 was injected subretinally into one eye of rd12 and Rpe65−/−::Rho−/− mice at postnatal day 14 (P14). Contralateral rd12 eyes were injected later, at P35. Rd12 behavioral testing revealed that rod vision loss was prevented with either P14 or P35 treatment, whereas cone vision was only detected after P14 treatment. Consistent with this observation, P35 treatment only restored rod electroretinogram (ERG) signals, a result likely due to reduced cone densities at this time point. For Rpe65−/−::Rho−/− mice in which there is no confounding rod contribution to the ERG signal, cone cells and cone-mediated ERGs were also maintained with treatment at P14. This work establishes that a self-complimentary AAV5 vector can restore substantial visual function in two genetically distinct models of Rpe65 deficiency within 4 days of treatment. In addition, this therapy prevents cone degeneration but only if administered before extensive cone degeneration, thus supporting continuation of current Leber's congenital amaurosis-2 clinical trials with an added emphasis on cone subtype analysis and early intervention.

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References

  1. Marlhens F, Bareil C, Griffoin JM, Zrenner E, Amalric P, Eliaou C et al. Mutations in RPE65 cause Leber's congenital amaurosis. Nat Genet 1997; 17: 139–141.

    Article  CAS  PubMed  Google Scholar 

  2. Rohrer B, Lohr HR, Humphries P, Redmond TM, Seeliger MW, Crouch RK . Cone opsin mislocalization in Rpe65−/− mice: a defect that can be corrected by 11-cis retinal. Invest Ophthalmol Vis Sci 2005; 46: 3876–3882.

    Article  PubMed  Google Scholar 

  3. Zhang H, Fan J, Li S, Karan S, Rohrer B, Palczewski K et al. Trafficking of membrane-associated proteins to cone photoreceptor outer segments requires the chromophore 11-cis-retinal. J Neurosci 2008; 28: 4008–4014.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Feathers KL, Lyubarsky AL, Khan NW, Teofilo K, Swaroop A, Williams DS et al. Nrl-knockout mice deficient in Rpe65 fail to synthesize 11-cis retinal and cone outer segments. Invest Ophthalmol Vis Sci 2008; 49: 1126–1135.

    Article  PubMed  Google Scholar 

  5. Jacobson SG, Aleman TS, Cideciyan AV, Heon E, Golczak M, Beltran WA et al. Human cone photoreceptor dependence on RPE65 isomerase. Proc Natl Acad Sci USA 2007; 104: 15123–15128.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Jacobson SG, Cideciyan AV, Aleman TS, Sumaroka A, Windsor EA, Schwartz SB et al. Photoreceptor layer topography in children with Leber congenital amaurosis caused by RPE65 mutations. Invest Ophthalmol Vis Sci 2008; 49: 4573–4577.

    Article  PubMed  Google Scholar 

  7. Mata NL, Ruiz A, Radu RA, Bui TV, Travis GH . Chicken retinas contain a retinoid isomerase activity that catalyzes the direct conversion of all-trans-retinol to 11-cis-retinol. Biochemistry 2005; 44: 11715–11721.

    Article  CAS  PubMed  Google Scholar 

  8. Muniz A, Villazana-Espinoza ET, Hatch AL, Trevino SG, Allen DM, Tsin AT . A novel cone visual cycle in the cone-dominated retina. Exp Eye Res 2007; 85: 175–184.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Schonthaler HB, Lampert JM, Isken A, Rinner O, Mader A, Gesemann M et al. Evidence for RPE65-independent vision in the cone-dominated zebrafish retina. Eur J Neurosci 2007; 26: 1940–1949.

    Article  PubMed  PubMed Central  Google Scholar 

  10. Wang JS, Estevez ME, Cornwall MC, Kefalov VJ . Intra-retinal visual cycle required for rapid and complete cone dark adaptation. Nat Neurosci 2009; 12: 295–302.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Bainbridge JW, Smith AJ, Barker SS, Robbie S, Henderson R, Balaggan K et al. Effect of gene therapy on visual function in Leber's congenital amaurosis. N Engl J Med 2008; 358: 2231–2239.

    Article  CAS  PubMed  Google Scholar 

  12. Maguire AM, Simonelli F, Pierce EA, Pugh Jr EN, Mingozzi F, Bennicelli J et al. Safety and efficacy of gene transfer for Leber's congenital amaurosis. N Engl J Med 2008; 358: 2240–2248.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Hauswirth W, Aleman TS, Kaushal S, Cideciyan AV, Schwartz SB, Wang L 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.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Cideciyan AV, Aleman TS, Boye SL, Schwartz SB, Kaushal S, Roman AJ et al. Human gene therapy for RPE65 isomerase deficiency activates the retinoid cycle of vision but with slow rod kinetics. Proc Natl Acad Sci USA 2008; 105: 15112–15117.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Cideciyan AV, Hauswirth WW, Aleman TS, Kaushal S, Schwartz SB, Boye SL et al. Vision one year after gene therapy for Leber congenital amaurosis. N Engl J Med 2009; 361: 725–727.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Cronin T, Leveillard T, Sahel JA . Retinal degenerations: from cell signaling to cell therapy: preclinical and clinical issues. Curr Gene Ther 2007; 7: 121–129.

    Article  CAS  PubMed  Google Scholar 

  17. Samardzija M, Tanimoto N, Kostic C, Beck S, Oberhauser V, Joly S et al. In conditions of limited chromophore supply rods entrap 11-cis-retinal leading to loss of cone function and death. Hum Mol Genet 2009; 18: 1266–1275.

    Article  CAS  PubMed  Google Scholar 

  18. Carroll J, Choi SS, Williams DR . In vivo imaging of the photoreceptor mosaic of a rod monochromat. Vision Res 2008; 48: 2564–2568.

    Article  PubMed  PubMed Central  Google Scholar 

  19. Seeliger MW, Grimm C, Stahlberg F, Friedburg C, Jaissle G, Zrenner E et al. New views on RPE65 deficiency: the rod system is the source of vision in a mouse model of Leber congenital amaurosis. Nat Genet 2001; 29: 70–74.

    Article  CAS  PubMed  Google Scholar 

  20. Hauswirth WW, Petrs-Silva H, Min S-H, Boye SE, Liu JM, Mani S et al. Self-complementary AAV vectors promote fast and efficient transduction of mouse retina. Invest Ophthalmol Vis Sci 2006; 47 ARVO E-Abstract 839.

  21. Yokoi K, Kachi S, Zhang HS, Gregory PD, Spratt SK, Samulski RJ et al. Ocular gene transfer with self-complementary AAV vectors. Invest Ophthalmol Vis Sci 2007; 48: 3324–3328.

    Article  PubMed  Google Scholar 

  22. Natkunarajah M, Trittibach P, McIntosh J, Duran Y, Barker SE, Smith AJ et al. Assessment of ocular transduction using single-stranded and self-complementary recombinant adeno-associated virus serotype 2/8. Gene Therapy 2008; 15: 463–467.

    Article  CAS  PubMed  Google Scholar 

  23. Li Q, Kong F, Li X, Dai X, Liu X, Zheng Q et al. Gene therapy following subretinal AAV5 vector delivery is not affected by a previous intravitreal AAV5 vector administration in the partner eye. Mol Vis 2009; 15: 267–275.

    CAS  PubMed  PubMed Central  Google Scholar 

  24. Leveillard T, Mohand-Said S, Lorentz O, Hicks D, Fintz AC, Clerin E et al. Identification and characterization of rod-derived cone viability factor. Nat Genet 2004; 36: 755–759.

    Article  CAS  PubMed  Google Scholar 

  25. Sahel JA . Saving cone cells in hereditary rod diseases: a possible role for rod-derived cone viability factor (RdCVF) therapy. Retina 2005; 25: S38–S39.

    Article  PubMed  Google Scholar 

  26. Redmond TM, Yu S, Lee E, Bok D, Hamasaki D, Chen N et al. Rpe65 is necessary for production of 11-cis-vitamin A in the retinal visual cycle. Nat Genet 1998; 20: 344–351.

    Article  CAS  PubMed  Google Scholar 

  27. Znoiko SL, Rohrer B, Lu K, Lohr HR, Crouch RK, Ma JX . Downregulation of cone-specific gene expression and degeneration of cone photoreceptors in the Rpe65−/ mouse at early ages. Invest Ophthalmol Vis Sci 2005; 46: 1473–1479.

    Article  PubMed  Google Scholar 

  28. Bemelmans AP, Kostic C, Crippa SV, Hauswirth WW, Lem J, Munier FL et al. Lentiviral gene transfer of RPE65 rescues survival and function of cones in a mouse model of Leber congenital amaurosis. PLoS Med 2006; 3: e347.

    Article  PubMed  PubMed Central  Google Scholar 

  29. Chen Y, Moiseyev G, Takahashi Y, Ma JX . RPE65 gene delivery restores isomerohydrolase activity and prevents early cone loss in Rpe65−/ mice. Invest Ophthalmol Vis Sci 2006; 47: 1177–1184.

    Article  PubMed  Google Scholar 

  30. Cramer W, Stowell RE . Carcinogenesis in the mouse's skin by the infrequent application at long intervals of methylcholanthrene. Cancer Res 1941; 1: 849–852.

    CAS  Google Scholar 

  31. Humphries MM, Rancourt D, Farrar GJ, Kenna P, Hazel M, Bush RA et al. Retinopathy induced in mice by targeted disruption of the rhodopsin gene. Nat Genet 1997; 15: 216–219.

    Article  CAS  PubMed  Google Scholar 

  32. Lem J, Krasnoperova NV, Calvert PD, Kosaras B, Cameron DA, Nicolo M et al. Morphological, physiological, and biochemical changes in rhodopsin knockout mice. Proc Natl Acad Sci USA 1999; 96: 736–741.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Hewitt AT, Lindsey JD, Carbott D, Adler R . Photoreceptor survival-promoting activity in interphotoreceptor matrix preparations: characterization and partial purification. Exp Eye Res 1990; 50: 79–88.

    Article  CAS  PubMed  Google Scholar 

  34. Bird AC . Investigation of disease mechanisms in retinitis pigmentosa. Ophthalmic Paediatr Genet 1992; 13: 57–66.

    Article  CAS  PubMed  Google Scholar 

  35. Petters RM, Alexander CA, Wells KD, Collins EB, Sommer JR, Blanton MR et al. Genetically engineered large animal model for studying cone photoreceptor survival and degeneration in retinitis pigmentosa. Nat Biotechnol 1997; 15: 965–970.

    Article  CAS  PubMed  Google Scholar 

  36. Ostrer H, Pullarkat RK, Kazmi MA . Glycosylation and palmitoylation are not required for the formation of the X-linked cone opsin visual pigments. Mol Vis 1998; 4: 28.

    CAS  PubMed  Google Scholar 

  37. Jaissle GB, May CA, Reinhard J, Kohler K, Fauser S, Lutjen-Drecoll E et al. Evaluation of the rhodopsin knockout mouse as a model of pure cone function. Invest Ophthalmol Vis Sci 2001; 42: 506–513.

    CAS  PubMed  Google Scholar 

  38. Pang J, Chang B, Hawes NL, Hurd RE, Davisson MT, Li J et al. Retinal degeneration 12 (rd12): a new, spontaneously arising mouse model for human Leber congenital amaurosis (LCA). Mol Vis 2005; 11: 152–162.

    CAS  PubMed  Google Scholar 

  39. Haire SE, Pang J, Boye SL, Sokal I, Craft CM, Palczewski K et al. Light-driven cone arrestin translocation in cones of postnatal guanylate cyclase-1 knockout mouse retina treated with AAV-GC1. Invest Ophthalmol Vis Sci 2006; 47: 3745–3753.

    Article  PubMed  Google Scholar 

  40. McCarty DM, Fu H, Monahan PE, Toulson CE, Naik P, Samulski RJ . Adeno-associated virus terminal repeat (TR) mutant generates self-complementary vectors to overcome the rate-limiting step to transduction in vivo. Gene Therapy 2003; 10: 2112–2118.

    Article  CAS  PubMed  Google Scholar 

  41. Hauswirth WW, Lewin AS, Zolotukhin S, Muzyczka N . Production and purification of recombinant adeno-associated virus. Methods Enzymol 2000; 316: 743–761.

    Article  CAS  PubMed  Google Scholar 

  42. Pang J, Chang B, Kumar A, Nusinowitz S, Noorwez SM, Li J et al. Gene therapy restores vision-dependent behavior as well as retinal structure and function in a mouse model of RPE65 Leber congenital amaurosis. Mol Ther 2006; 13: 565–572.

    Article  CAS  PubMed  Google Scholar 

  43. Pang J, Boye SL, Kumar A, Dinculescu A, Deng W, Li J et al. AAV-mediated gene therapy delays retinal degeneration in the rd10 mouse containing a recessive PDEβ mutation. Invest Ophthalmol Vis Sci 2008; 49: 4278–4283.

    Article  PubMed  Google Scholar 

  44. Timmers AM, Zhang H, Squitieri A, Bonzalez-Pola C . Subretinal injections in rodent eyes: effects on electrophysiology and histology of rat retina. Mol Vis 2001; 7: 131–137.

    CAS  PubMed  Google Scholar 

  45. Lei B, Yao G, Zhang K, Hofeldt KJ, Chang B . Study of rod- and cone-driven oscillatory potentials in mice. Invest Ophthalmol Vis Sci 2006; 47: 2732–2738.

    Article  PubMed  Google Scholar 

  46. Lei B, Zhang K, Yue Y, Ghosh A, Duan D . Adeno-associated virus serotype-9 efficiently transduces the photoreceptor terminals. Mol Vis 2009; 17: 1374–1382.

    Google Scholar 

  47. Umino Y, Solessio E, Barlow RB . Speed, spatial, and temporal tuning of rod and cone vision in mouse. J Neurosci 2008; 28: 189–198.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  48. Alexander JJ, Umino Y, Everhart D, Chang B, Min SH, Li Q et al. Restoration of cone vision in a mouse model of achromatopsia. Nat Med 2007; 13: 685–687.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. Douglas RM, Alam NM, Silver BD, McGill TJ, Tschetter WW, Prusky GT . Independent visual threshold measurements in the two eyes of freely moving rats and mice using a virtual-reality optokinetic system. Vis Neuro 2005; 22: 677–684.

    Article  CAS  Google Scholar 

  50. Pang J, Cheng M, Haire SE, Barker E, Planelles V, Blanks JC . Efficiency of lentiviral transduction during development in normal and rd mice. Mol Vis 2006; 12: 756–767.

    CAS  PubMed  Google Scholar 

  51. Pang J, Lauramore A, Deng W, Li Q, Doyle T, Chiodo V et al. Analysis of in vivo and in vitro AAV vector transduction in neonatal mouse retina: effects of serotype and site of administration. Vision Res 2008; 48: 377–385.

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

We thank Vince Chiodo and Thomas Doyle at the University of Florida and Keqing Zhang at the University of Missouri for the technical support. We also acknowledge NIH Grants EY018331, EY13729, EY11123, NS36302, EY08571, EY07758, EY014046, EY06360, EY017246 (to DE), EY00067 (to RB) and grants from the Macular Vision Research Foundation, Foundation Fighting Blindness, Fight for Sight (to DE), Lions of Central NY, NASA (to RB), Juvenile Diabetes Research Foundation and Research to Prevent Blindness Inc. for partial support of this work.

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Correspondence to J Pang.

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WWH and the University of Florida have a financial interest in the use of AAV therapies and own equity in a company (AGTC Inc.) that might, in the future, commercialize some aspects of this work. There is no conflict of interest for other co-authors.

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Pang, J., Boye, S., Lei, B. et al. Self-complementary AAV-mediated gene therapy restores cone function and prevents cone degeneration in two models of Rpe65 deficiency. Gene Ther 17, 815–826 (2010). https://doi.org/10.1038/gt.2010.29

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