Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Short Communication
  • Published:

Photoreceptor-targeted gene delivery using intravitreally administered AAV vectors in dogs

Gene Therapy volume 23, pages 223–230 (2016)Cite this article

Subjects

A Corrigendum to this article was published on 07 April 2016

Abstract

Delivery of therapeutic transgenes to retinal photoreceptors using adeno-associated virus (AAV) vectors has traditionally required subretinal injection. Recently, photoreceptor transduction efficiency following intravitreal injection (IVT) has improved in rodent models through use of capsid-mutant AAV vectors; but remains limited in large animal models. Thickness of the inner limiting membrane (ILM) in large animals is thought to impair retinal penetration by AAV. Our study compared two newly developed AAV vectors containing multiple capsid amino acid substitutions following IVT in dogs. The ability of two promoter constructs to restrict reporter transgene expression to photoreceptors was also evaluated. AAV vectors containing the interphotoreceptor-binding protein (IRBP) promoter drove expression exclusively in rod and cone photoreceptors, with transduction efficiencies of ~4% of cones and 2% of rods. Notably, in the central region containing the cone-rich visual streak, 15.6% of cones were transduced. Significant regional variation existed, with lower transduction efficiencies in the temporal regions of all eyes. This variation did not correlate with ILM thickness. Vectors carrying a cone-specific promoter failed to transduce a quantifiable percentage of cone photoreceptors. The newly developed AAV vectors containing the IRBP promoter were capable of producing photoreceptor-specific transgene expression following IVT in the dog.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1
Figure 2
Figure 3
Figure 4

Similar content being viewed by others

References

  1. Nonnenmacher M, Weber T . Intracellular transport of recombinant adeno-associated virus vectors. Gene Ther 2012; 19: 649–658.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Asokan A, Schaffer DV, Samulski RJ . The AAV vector toolkit: poised at the clinical crossroads. Mol Ther 2012; 20: 699–708.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Dalkara D, Sahel JA . Gene therapy for inherited retinal degenerations. C R Biol 2014; 337: 185–192.

    Article  PubMed  Google Scholar 

  4. Jacobson SG, Cideciyan AV, Ratnakaram R, Heon E, Schwartz SB, Roman AJ 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.

    Article  CAS  PubMed  Google Scholar 

  5. Nork TM, Murphy CJ, Kim CB, Ver Hoeve JN, Rasmussen CA, Miller PE et al. Functional and anatomic consequences of subretinal dosing in the cynomolgus macaque. Arch Ophthalmol 2012; 130: 65–75.

    Article  CAS  PubMed  Google Scholar 

  6. Li Q, Miller R, Han PY, Pang J, Dinculescu A, Chiodo V et al. Intraocular route of AAV2 vector administration defines humoral immune response and therapeutic potential. Mol Vis 2008; 14: 1760–1769.

    CAS  PubMed  PubMed Central  Google Scholar 

  7. Dudus L, Anand V, Acland GM, Chen SJ, Wilson JM, Fisher KJ et al. Persistent transgene product in retina, optic nerve and brain after intraocular injection of rAAV. Vision Res 1999; 39: 2545–2553.

    Article  CAS  PubMed  Google Scholar 

  8. Beltran WA, Cideciyan AV, Lewin AS, Iwabe S, Khanna H, Sumaroka A et al. Gene therapy rescues photoreceptor blindness in dogs and paves the way for treating human X-linked retinitis pigmentosa. Proc Natl Acad Sci USA 2012; 109: 2132–2137.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Dalkara D, Byrne LC, Lee T, Hoffmann NV, Schaffer DV, Flannery JG . Enhanced gene delivery to the neonatal retina through systemic administration of tyrosine-mutated AAV9. Gene Ther 2012; 19: 176–181.

    Article  CAS  PubMed  Google Scholar 

  10. Dalkara D, Byrne LC, Klimczak RR, Visel M, Yin L, Merigan WH et al. In vivo-directed evolution of a new adeno-associated virus for therapeutic outer retinal gene delivery from the vitreous. Sci Transl Med 2013; 5: 189ra76.

    Article  PubMed  Google Scholar 

  11. Kay CN, Ryals RC, Aslanidi GV, Min SH, Ruan Q, Sun J et al. Targeting photoreceptors via intravitreal delivery using novel, capsid-mutated AAV vectors. PLoS One 2013; 8: e62097.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Zhong L, Li B, Jayandharan G, Mah CS, Govindasamy L, Agbandje-McKenna M et al. Tyrosine-phosphorylation of AAV2 vectors and its consequences on viral intracellular trafficking and transgene expression. Virology 2008; 381: 194–202.

    Article  CAS  PubMed  Google Scholar 

  13. Boye S, Bennett A, VanVliet K, Dinculescu A, White M, Peterson J et al. Heparan sulfate affinity dictates transduction of photoreceptors from the vitreous by capsid mutated AAV2 variants. Invest Ophthalmol Vis Sci 2014; 55: (abstract 3337).

  14. Gabriel N, Hareendran S, Sen D, Gadkari RA, Sudha G, Selot R et al. Bioengineering of AAV2 capsid at specific serine, threonine, or lysine residues improves its transduction efficiency in vitro and in vivo. Hum Gene Ther Methods 2013; 24: 80–93.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Yin L, Greenberg K, Hunter JJ, Dalkara D, Kolstad KD, Masella BD et al. Intravitreal injection of AAV2 transduces macaque inner retina. Invest Ophthalmol Vis Sci 2011; 52: 2775–2783.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Mowat FM, Gornik KR, Dinculescu A, Boye SL, Hauswirth WW, Petersen-Jones SM et al. Tyrosine capsid-mutant AAV vectors for gene delivery to the canine retina from a subretinal or intravitreal approach. Gene Ther 2014; 21: 96–105.

    Article  CAS  PubMed  Google Scholar 

  17. Petersen-Jones SM, Komaromy AM . Dog models for blinding inherited retinal dystrophies. Hum Gene Ther Clin Dev 2015; 26: 15–26.

    Article  CAS  PubMed  Google Scholar 

  18. Beltran WA, Cideciyan AV, Guziewicz KE, Iwabe S, Swider M, Scott EM et al. Canine retina has a primate fovea-like bouquet of cone photoreceptors which is affected by inherited macular degenerations. PLoS One 2014; 9: e90390.

    Article  PubMed  PubMed Central  Google Scholar 

  19. Acland GM, Aguirre GD, Ray J, Zhang Q, Aleman TS, Cideciyan AV et al. Gene therapy restores vision in a canine model of childhood blindness. Nat Genet 2001; 28: 92–95.

    CAS  PubMed  Google Scholar 

  20. Komaromy AM, Alexander JJ, Rowlan JS, Garcia MM, Chiodo VA, Kaya A et al. Gene therapy rescues cone function in congenital achromatopsia. Hum Mol Genet 2010; 19: 2581–2593.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Annear MJ, Bartoe JT, Barker SE, Smith AJ, Curran PG, Bainbridge JW et al. Gene therapy in the second eye of RPE65-deficient dogs improves retinal function. Gene Ther 2011; 18: 53–61.

    Article  CAS  PubMed  Google Scholar 

  22. Mowat FM, Bartoe JT, Bruewer A, Dinculescu A, Boye SL, Hauswirth WW et al. Evaluation of rod photoreceptor function and preservation following retinal gene therapy in the PDE6A mutant dog. Invest Ophthalmol Vis Sci 2012; 53: (Abstract 1928).

  23. Yeh C, Iwabe S, Boye S, McDaid K, Harman C, Wen R et al. Optimization of cone-directed AAV-mediated gene augmentation therapy for CNGB3-achromatopsia by use of the IRBP/GNAT2-promoter and intravitreal CNTF administration. Invest Ophthalmol Vis Sci 2013; 54: (Abstract 1937).

  24. Ivanova E, Hwang GS, Pan ZH, Troilo D . Evaluation of AAV-mediated expression of Chop2-GFP in the marmoset retina. Invest Ophthalmol Vis Sci 2010; 51: 5288–5296.

    Article  PubMed  PubMed Central  Google Scholar 

  25. Jayandharan GR, Zhong L, Li B, Kachniarz B, Srivastava A . Strategies for improving the transduction efficiency of single-stranded adeno-associated virus vectors in vitro and in vivo. Gene Ther 2008; 15: 1287–1293.

    Article  CAS  PubMed  Google Scholar 

  26. Byrne LC, Ozturk BE, Lee T, Fortuny C, Visel M, Dalkara D et al. Retinoschisin gene therapy in photoreceptors, Muller glia or all retinal cells in the Rs1h-/- mouse. Gene Ther 2014; 21: 585–592.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Schultz BR, Chamberlain JS . Recombinant adeno-associated virus transduction and integration. Mol Ther 2008; 16: 1189–1199.

    Article  CAS  PubMed  Google Scholar 

  28. Donsante A, Miller DG, Li Y, Vogler C, Brunt EM, Russell DW et al. AAV vector integration sites in mouse hepatocellular carcinoma. Science 2007; 317: 477.

    Article  CAS  PubMed  Google Scholar 

  29. Tenenbaum L, Lehtonen E, Monahan PE . Evaluation of risks related to the use of adeno-associated virus-based vectors. Curr Gene Ther 2003; 3: 545–565.

    Article  CAS  PubMed  Google Scholar 

  30. Dyka FM, Boye SL, Ryals RC, Chiodo VA, Boye SE, Hauswirth WW . Cone specific promoter for use in gene therapy of retinal degenerative diseases. Adv Exp Med Biol 2014; 801: 695–701.

    Article  PubMed  PubMed Central  Google Scholar 

  31. Dalkara D, Kolstad KD, Caporale N, Visel M, Klimczak RR, Schaffer DV et al. Inner limiting membrane barriers to AAV-mediated retinal transduction from the vitreous. Mol Ther 2009; 17: 2096–2102.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Cehajic-Kapetanovic J, Le Goff MM, Allen A, Lucas RJ, Bishop PN . Glycosidic enzymes enhance retinal transduction following intravitreal delivery of AAV2. Mol Vis 2011; 17: 1771–1783.

    CAS  PubMed  PubMed Central  Google Scholar 

  33. Kolstad KD, Dalkara D, Guerin K, Visel M, Hoffmann N, Schaffer DV et al. Changes in adeno-associated virus-mediated gene delivery in retinal degeneration. Hum Gene Ther 2010; 21: 571–578.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Johnson TV, Bull ND, Martin KR . Transplantation prospects for the inner retina. Eye 2009; 23: 1980–1984.

    Article  CAS  PubMed  Google Scholar 

  35. Kim H, Robinson S, Csaky K . Investigating the movement of intravitreal human serum albumin nanoparticles in the vitreous and retina. Pharm Res 2009; 26: 329–337.

    Article  CAS  PubMed  Google Scholar 

  36. Koo H, Moon H, Han H, Na JH, Huh MS, Park JH et al. The movement of self-assembled amphiphilic polymeric nanoparticles in the vitreous and retina after intravitreal injection. Biomaterials 2012; 33: 3485–3493.

    Article  CAS  PubMed  Google Scholar 

  37. Aartsen WM, van Cleef KW, Pellissier LP, Hoek RM, Vos RM, Blits B et al. GFAP-driven GFP expression in activated mouse Muller glial cells aligning retinal blood vessels following intravitreal injection of AAV2/6 vectors. PLoS One 2010; 5: e12387.

    Article  PubMed  PubMed Central  Google Scholar 

  38. Matsumoto B, Blanks JC, Ryan SJ . Topographic variations in the rabbit and primate internal limiting membrane. Invest Ophthalmol Vis Sci 1984; 25: 71–82.

    CAS  PubMed  Google Scholar 

  39. Sebag J . Anatomy and pathology of the vitreo-retinal interface. Eye 1992; 6: 541–552.

    Article  PubMed  Google Scholar 

  40. Wolter JR . Pores in the internal limiting membrane of the human retina. Acta Ophthalmol (Copenh) 1964; 42: 971–974.

    Article  CAS  Google Scholar 

  41. Mutlu F, Leopold IH . The structure of human retinal vascular system. Arch Ophthalmol 1964; 71: 93–101.

    Article  CAS  PubMed  Google Scholar 

  42. Zolotukhin S, Potter M, Zolotukhin I, Sakai Y, Loiler S, Fraites TJ Jr et al. Production and purification of serotype 1, 2, and 5 recombinant adeno-associated viral vectors. Methods 2002; 28: 158–167.

    Article  CAS  PubMed  Google Scholar 

  43. Ying S, Fong SL, Fong WB, Kao CW, Converse RL, Kao WW . A CAT reporter construct containing 277 bp GNAT2 promoter and 214 bp IRBP enhancer is specifically expressed by cone photoreceptor cells in transgenic mice. Curr Eye Res 1998; 17: 777–782.

    Article  CAS  PubMed  Google Scholar 

  44. Gearhart PM, Gearhart C, Thompson DA, Petersen-Jones SM . Improvement of visual performance with intravitreal administration of 9-cis-retinal in Rpe65-mutant dogs. Arch Ophthalmol 2010; 128: 1442–1448.

    Article  CAS  PubMed  Google Scholar 

  45. Mowat FM, Breuwer AR, Bartoe JT, Annear MJ, Zhang Z, Smith AJ et al. RPE65 gene therapy slows cone loss in Rpe65-deficient dogs. Gene Ther 2012; 20: 545–555.

    Article  PubMed  Google Scholar 

Download references

Acknowledgements

We thank Janice Querbin and Kristen Koehl for animal assistance, Kristen Gervais and Laurence Occelli for imaging assistance and Tzu-Fen Chang and Joe Hauptman for statistical assistance. We also thank Cheryl Craft for hCAR antibody production, as well as Vince Chiodo and the Retinal Gene Therapy Vector lab for AAV purification. JTB and RFB acknowledge funding from Michigan State University College of Veterinary Medicine Endowed Research Fund. SLB, SEB, WWH, and AMK acknowledge funding from Foundation Fighting Blindness. SEB acknowledges funding from NIH Grant R01 EY024280. AMK acknowledges funding from NIH Grant R01 EY019304. SPJ acknowledges funding from Myers-Dunlap Endowment for Canine Health.

Author information

Authors and Affiliations

  1. Small Animal Clinical Sciences, Michigan State University, East Lansing, MI, USA

    R F Boyd, A M Komáromy, S M Petersen-Jones & J T Bartoe

  2. Diagnostic Center for Population and Animal Health, Michigan State University, East Lansing, MI, USA

    D G Sledge

  3. Department of Ophthalmology, University of Florida College of Medicine, Gainesville, FL, USA

    S L Boye, S E Boye & W W Hauswirth

Authors
  1. R F Boyd
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. D G Sledge
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. S L Boye
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. S E Boye
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. W W Hauswirth
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. A M Komáromy
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. S M Petersen-Jones
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. J T Bartoe
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to S M Petersen-Jones.

Ethics declarations

Competing interests

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. The remaining authors declare no conflict of interest.

Additional information

Supplementary Information accompanies this paper on Gene Therapy website

Supplementary information

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Boyd, R., Sledge, D., Boye, S. et al. Photoreceptor-targeted gene delivery using intravitreally administered AAV vectors in dogs. Gene Ther 23, 223–230 (2016). https://doi.org/10.1038/gt.2015.96

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/gt.2015.96

This article is cited by

Search

Advanced search

Quick links