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Manufacturing Considerations and Challenges for AAV Ocular Gene Therapy

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Ophthalmic Product Development

Part of the book series: AAPS Advances in the Pharmaceutical Sciences Series ((AAPS,volume 37))

Abstract

Gene therapy has the potential to revolutionize the treatment of debilitating ocular diseases, and adeno-associated viral vectors (AAV) are particularly attractive. However, technical hurdles that remain in their manufacture and product formulation may hamper the development of new treatments. Such issues arise from the structural properties of AAV and differ between serotypes due to their varying capsid structure. Whereas the manufacture and quality control of AAV vectors generally exploit many techniques, materials and procedures that have been developed for other biotherapeutics, the unique characteristics of AAV capsid structure, and the AAV genome encapsidation process introduce complexities not commonly encountered hitherto. In this chapter we examine these issues and review progress with overcoming the technical challenges that arise. In particular, the development of AAVs using directed evolution or rational design techniques will result in novel vectors that enable more efficient transduction of diseased tissues and which can be manufactured and stored more reliably and safely.

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References

  • 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(6):685–7.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Allocca M, Mussolino C, Garcia-Hoyos M, Sanges D, Iodice C, Petrillo M, et al. Novel adeno-associated virus serotypes efficiently transduce murine photoreceptors. J Virol. 2007;81(20):11372–80.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Anzo K, Harada M, Okada T. Enhanced kinetics of pseudo first-order hydrolysis in liquid phase coexistent with ice. J Phys Chem A. 2013;117(41):10619–25.

    Article  CAS  PubMed  Google Scholar 

  • Auricchio A. Pseudotyped AAV vectors for constitutive and regulated gene expression in the eye. Vision Res. 2003;43(8):913–8.

    Article  CAS  PubMed  Google Scholar 

  • Auricchio A, Kobinger G, Anand V, Hildinger M, O’Connor E, Maguire AM, et al. Exchange of surface proteins impacts on viral vector cellular specificity and transduction characteristics: the retina as a model. Hum Mol Genet. 2001;10(26):3075–81.

    Article  CAS  PubMed  Google Scholar 

  • Ayuso E, Mingozzi F, Bosch F. Production, purification and characterization of adeno-associated vectors. Curr Gene Ther. 2010;10(6):423–36.

    Article  CAS  PubMed  Google Scholar 

  • 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 U S A. 2012;109(6):2132–7.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Bennett A, et al. Thermal stability as a determinant of serotype identity. Mol Ther Methods Clin Dev. 2017;6:171–82. Accessed 15 Sep 2017. https://doi.org/10.1016/j.omtm.2017.07.003.eCollection.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Bennicelli J, Wright JF, Komaromy A, Jacobs JB, Hauck B, Zelenaia O, et al. Reversal of blindness in animal models of Leber congenital amaurosis using optimized AAV2-mediated gene transfer. Mol Ther. 2008;16(3):458–65.

    Article  CAS  PubMed  Google Scholar 

  • Bhatnagar BS, Bogner RH, Pikal MJ. Protein stability during freezing: separation of stresses and mechanisms of protein stabilization. Pharm Dev Technol. 2007;12(5):505–23.

    Article  CAS  PubMed  Google Scholar 

  • Bhatnagar BS, Pikal MJ, Bogner RH. Study of the individual contributions of ice formation and freeze-concentration on isothermal stability of lactate dehydrogenase during freezing. J Pharm Sci. 2008;97(2):798–814.

    Article  CAS  PubMed  Google Scholar 

  • Blanco MA, Sahin E, Robinson AS, Roberts CJ. J Phys Chem B. 2013;117:16013–28.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Boye SE, Boye SL, Lewin AS, Hauswirth WW. A comprehensive review of retinal gene therapy. Mol Ther. 2013;21(3):509–19.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Carvalho LS, Xu J, Pearson RA, Smith AJ, Bainbridge JW, Morris LM, 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(16):3161–75.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Clément N, Grieger JC. Manufacturing of recombinant adeno-associated viral vectors for clinical trials. Mol Ther Methods Clin Dev. 2016;3:16002.

    Article  PubMed  PubMed Central  Google Scholar 

  • Conchilli-Solé O, de Groot NS, Avilés FX, Vendrell J, Daura X, Ventura S. AGGRESCAN: a server for the prediction and evaluation of “hot-spots” of aggregation in polypeptides. BMC Bioinform. 2007;8:65.

    Article  CAS  Google Scholar 

  • Connolly BD, Le L, Patapoff TW, Cromwell MEME, Moore JMR. Protein aggregation in frozen trehalose formulations: effects of composition, cooling rate, and storage temperature. J Pharm Sci. 2015;104:4170–84.

    Article  CAS  PubMed  Google Scholar 

  • Croyle MA, Cheng X, Wilson JM. Development of formulations that enhance physical stability of viral vectors for gene therapy. Gene Ther. 2001;8(17):1281–90.

    Article  CAS  PubMed  Google Scholar 

  • Di Mattia MA, et al. Structural insight into the unique properties of adeno-associated virus serotype 9. J Virol. 2012;86(12):6947–58. https://doi.org/10.1128/JVI.07232-11.

    Article  CAS  Google Scholar 

  • Flannery JG, Zolotukhin S, Vaquero MI, LaVail MM, Muzyczka N, Hauswirth WW. Efficient photoreceptor-targeted gene expression in vivo by recombinant adeno-associated virus. Proc Natl Acad Sci U S A. 1997;94(13):6916–21.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Franks F, Hatley RHM. Stability of proteins at subzero temperatures: thermodynamics and some ecological consequences. Pure Appl Chem. 1991;63(10):1367–80.

    Article  CAS  Google Scholar 

  • Gao G, Qu G, Burnham MS, Huang J, Chirmule N, Joshi B, Yu QC, Marsh JA, Conceicao CM, Wilson JM. Hum Gene Ther. 2000;11:2079–91.

    Article  CAS  PubMed  Google Scholar 

  • Gao G, Vandenberghe LH, Wilson JM. New recombinant serotypes of AAV vectors. Curr Gene Ther. 2005;5(3):285–97.

    Article  CAS  PubMed  Google Scholar 

  • Gu JH, Beekman A, Wu T, Piedmonte DM, Baker P, Eschenberg M, Hale M, Goldenbergh M. Beyond glass transitions: studying the highly viscous and elastic behavior of frozen protein formulations using low temperature rheology and its potential implications on protein stability. Pharm Res. 2013;30:387–401.

    Article  CAS  PubMed  Google Scholar 

  • Howard DB, Harvey BK. Assaying the stability and inactivation of AAV serotype 1 vectors. Hum Gene Ther Methods. 2017;28(1):39–48.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Israelachvili J. Intermolecular and surface forces. San Diego, CA: Academic Press; 1985.

    Google Scholar 

  • Komáromy AM, Alexander JJ, Cooper AE, Chiodo VA, Glushakova LG, Acland GM, et al. Targeting gene expression to cones with human cone opsin promoters in recombinant AAV. Gene Ther. 2008;15(14):1049–55.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • 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(13):2581–93.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Kotterman MA, Schaffer DV. Engineering adeno-associated viruses for clinical gene therapy. Nat Rev Genet. 2014;15(7):445–51.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Kwon I, Schaffer DV. Designer gene delivery vectors: molecular engineering and evolution of adeno-associated viral vectors for enhanced gene transfer. Pharm Res. 2008;25(3):489–99.

    Article  CAS  PubMed  Google Scholar 

  • Lebherz C, Maguire A, Tang W, Bennett J, Wilson JM. Novel AAV serotypes for improved ocular gene transfer. J Gene Med. 2008;10(4):375–82.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Levine H, Shalaev E, Franks F. Solid-liquid state diagrams in pharmaceutical lyophilisation: crystallization of solutes. In: Levine H, editor. Amorphous food and pharmaceutical systems. London: The Royal Society of Chemistry; 2002. p. 200–15.

    Google Scholar 

  • Lotery AJ, Yang GS, Mullins RF, Russell SR, Schmidt M, Stone EM, et al. Adeno-associated virus type 5: transduction efficiency and cell-type specificity in the primate retina. Hum Gene Ther. 2003;14(17):1663–71.

    Article  CAS  PubMed  Google Scholar 

  • Lund DB, Fennema OR, Powrie WD. Enzymic and acid hydrolysis of sucrose as influenced by freezing. J Food Sci. 1969;34:378–82.

    Article  CAS  Google Scholar 

  • Madigan VJ, Asokan A. Engineering AAV receptor footprints for gene therapy. Curr Opin Virol. 2016;18:89–96.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Mancuso K, Hauswirth WW, Li Q, Connor TB, Kuchenbecker JA, Mauck MC, et al. Gene therapy for red-green colour blindness in adult primates. Nature. 2009;461(7265):784–7.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Manfredi A, Marrocco E, Puppo A, Cesi G, Sommella A, Della Corte M, et al. Combined rod and cone transduction by adeno-associated virus 2/8. Hum Gene Ther. 2013;24(12):982–92.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Martino AT, Basner-Tschakarjan E, Markusic DM, Finn JD, Hinderer C, Zhou S, et al. Engineered AAV vector minimizes in vivo targeting of transduced hepatocytes by capsid-specific CD8+ T cells. Blood. 2013;121(12):2224–33.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Michalakis S, Mühlfriedel R, Tanimoto N, Krishnamoorthy V, Koch S, Fischer MD, et al. Restoration of cone vision in the CNGA3−/− mouse model of congenital complete lack of cone photoreceptor function. Mol Ther. 2010;18(12):2057–63.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Mussolino C, Della Corte M, Rossi S, Viola F, Di Vicino U, Marrocco E, et al. AAV-mediated photoreceptor transduction of the pig cone-enriched retina. Gene Ther. 2011;18(7):637–45.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Nass SA, Mattingly MA, Woodcock DA, Burnham BL, Ardinger JA, Osmond SE, et al. Universal method for the purification of recombinant AAV vectors of differing serotypes. Mol Ther Methods Clin Dev. 2018;9:33–46.

    Article  CAS  PubMed  Google Scholar 

  • Nestolaa P, Peixotoa C, Villaine L, Alvesa PM, Carrondoa MJT, Mota JPB. Rational development of two flowthrough purification strategies for adenovirus type 5 and retro virus-like particles. J Chromatogr A. 2015;1426:91–101.

    Article  CAS  Google Scholar 

  • Petrs-Silva H, Dinculescu A, Li Q, Min SH, Chiodo V, Pang JJ, et al. High-efficiency transduction of the mouse retina by tyrosine-mutant AAV serotype vectors. Mol Ther. 2009;17(3):463–71.

    Article  CAS  PubMed  Google Scholar 

  • Piedmonte DM, Sumers C, McAuley A, Karamuji L, Ratnaswamy G. Sorbitol crystallization can Lead to protein aggregation in frozen protein formulations. Pharm Res. 2007;24:136–46.

    Article  CAS  PubMed  Google Scholar 

  • Pincock RE. Reactions in frozen systems. Acc Chem Res. 1969;2:97–103.

    Article  CAS  Google Scholar 

  • Pincock RE, Kiovsky TE. Kinetics of reactions in frozen solutions. J Chem Educ. 1966;43:358–60.

    Article  CAS  Google Scholar 

  • Qu G, Bahr-Davidson J, Prado J, Tai A, Cataniag F, McDonnell J, et al. Separation of adeno-associated virus type 2 empty particles from genome containing vectors by anion-exchange column chromatography. J Virol Methods. 2007;140(1–2):183–92.

    Article  CAS  PubMed  Google Scholar 

  • Qua W, Wanga M, Wua Y, Lva Y, Wanga Q, Xua R. Calcium-ion-modulated ceramic hydroxyapatite resin for the scalable purification of recombinant adeno-associated virus serotype 9. J Chromatogr B. 2015;990:15–22.

    Article  CAS  Google Scholar 

  • Rayaprolu V, et al. Comparative analysis of adeno-associated virus capsid stability and dynamics. J Virol. 2013;87(24):13150–60. https://doi.org/10.1128/JVI.01415-13.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Schnodt M, Büning H. Improving the quality of adeno-associated viral vector preparations: the challenge of product-related impurities. Hum Gene Ther Methods. 2017;28(3):101–8.

    Article  PubMed  CAS  Google Scholar 

  • Schön C, Biel M, Michalakis S. Retinal gene delivery by adeno-associated virus (AAV) vectors: strategies and applications. Eur J Pharm Biopharm. 2015;95(Pt B):343–52.

    Article  PubMed  CAS  Google Scholar 

  • Schwegman JJ, Carpenter JF, Nail SL. Evidence of partial unfolding of proteins at the ice/freeze-concentrate interface by infrared microscopy. J Pharm Sci. 2009;98(9):3239–46.

    Article  CAS  PubMed  Google Scholar 

  • Singh SK, Kolhe P, Mehta AP, Chico SC, Lary AL, Huang M. Frozen state storage instability of a monoclonal antibody: aggregation as a consequence of trehalose crystallization and protein unfolding. Pharm Res. 2011;28(4):873–85.

    Article  CAS  PubMed  Google Scholar 

  • Sommer J, Sanftner L, Feng L, Suzuki B, Zhen Z, Powell S, et al. Optimal recovery of AAV-AADC vector from Teflon catheters using Polysorbate 80 or Pluronic F68 (abstract 768). Mol Ther. 2002;5(Suppl):S251.

    Google Scholar 

  • Summerford C, Samulski RJ. Viral receptors and vector purification: new approaches for generating clinical-grade reagents. Nat Med. 1999;5(5):587–8.

    Article  CAS  PubMed  Google Scholar 

  • Trapani I, Puppo A, Auricchio A. Vector platforms for gene therapy of inherited retinopathies. Prog Retin Eye Res. 2014;43:108–28.

    Article  CAS  PubMed  Google Scholar 

  • Vandenberghe LH, Bell P, Maguire AM, Cearley CN, Xiao R, Calcedo R, et al. Dosage thresholds for AAV2 and AAV8 photoreceptor gene therapy in monkey. Sci Transl Med. 2011;3(88):88ra54.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Venkatakrishnan B, Yarbrough J, Domsic J, Bennett A, Bothner B, Kozyreva OG, et al. Structure and dynamics of adeno-associated virus serotype 1 VP1-unique N-terminal domain and its role in capsid trafficking. J Virol. 2013;87(9):4974–84.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Wang Q, Lock M, Prongay AJ, Alvira MR, Petkov B, Wilson JM. Identification of an adeno-associated virus binding epitope for AVB sepharose affinity resin. Mol Ther Methods Clin Dev. 2015;2:15040.

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Weber M, Rabinowitz J, Provost N, Conrath H, Folliot S, Briot D, et al. Recombinant adeno-associated virus serotype 4 mediates unique and exclusive long-term transduction of retinal pigmented epithelium in rat, dog, and nonhuman primate after subretinal delivery. Mol Ther. 2003;7(6):774–81.

    Article  CAS  PubMed  Google Scholar 

  • Wright JF. Product-related impurities in clinical-grade recombinant AAV vectors: characterization and risk assessment. Biomedicine. 2014;2(1):80–97.

    Google Scholar 

  • Wright JF, Qu G, Tang C, Sommer JM. Recombinant adeno-associated virus: formulation challenges and strategies for a gene therapy vector. Curr Opin Drug Discov Devel. 2003;6(2):174–8.

    CAS  PubMed  Google Scholar 

  • Wright JF, Le T, Prado J, Bahr-Davidson J, Smith PH, Zhen Z, et al. Identification of factors that contribute to recombinant AAV2 particle aggregation and methods to prevent its occurrence during vector purification and formulation. Mol Ther. 2005;12(1):171–8.

    Article  CAS  PubMed  Google Scholar 

  • Wu C, Shamblin S, Varshney D, Shalaev E. Advance understanding of buffer behavior during lyophilization. In: Varshney D, Singh M, editors. Lyophilized biologics and vaccines: modality-based approaches. New York: Springer; 2015. p. 25–42.

    Chapter  Google Scholar 

  • Xie Q, Hare J, Turnigan J, Chapman MS. Large-scale production, purification and crystallization of wild-type adeno-associated virus-2. J Virol Methods. 2004;122(1):17–27.

    Article  CAS  PubMed  Google Scholar 

  • Yang GS, Schmidt M, Yan Z, Lindbloom JD, Harding TC, Donahue BA, et al. Virus-mediated transduction of murine retina with adeno-associated virus: effects of viral capsid and genome size. J Virol. 2002;76(15):7651–60.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Young JE, Vogt T, Gross KW, Khani SC. A short, highly active photoreceptor-specific enhancer/promoter region upstream of the human rhodopsin kinase gene. Invest Ophthalmol Vis Sci. 2003;44(9):4076–85.

    Article  PubMed  Google Scholar 

  • Zhong L, Li B, Mah CS, Govindasamy L, Agbandje-McKenna M, Cooper M, et al. Next generation of adeno-associated virus 2 vectors: point mutations in tyrosines lead to high-efficiency transduction at lower doses. Proc Natl Acad Sci U S A. 2008;105(22):7827–32.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Zipper DB, Goldschmidt L, Teng PK, Riek R, Eisenberg D. Identifying the amylome, proteins capable of forming amyloid-like fibrils. Proc Natl Acad Sci U S A. 2010;107:3487–92.

    Article  Google Scholar 

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Acknowledgments

Authors thank Drs. Michael Sztucki and Theyencheri Narayanan for their expert assistance with the SAXS experiment.

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Authors and Affiliations

  1. Eye Care Research, Abbvie, Irvine, CA, USA

    Gerard A. Rodrigues

  2. Pharmaceutical Sciences, R&D, Abbvie, Irvine, CA, USA

    Evgenyi Shalaev, Shaoxin Feng & Dinen Shah

  3. Pharmaceutical R&D, Allergan plc, Irvine, CA, USA

    Thomas K. Karami

  4. Analytical QC, Glaukos Corporation, San Clemente, CA, USA

    Thomas K. Karami

  5. Combination Product Development, AbbVie, Irvine, CA, USA

    James Cunningham

  6. Allergan Aesthetics R&D, AbbVie, Irvine, CA, USA

    Hongwen M. Rivers

  7. Department of Chemical Engineering and Biotechnology, University of Cambridge, Cambridge, UK

    Nigel K. H. Slater

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  1. Gerard A. Rodrigues
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  2. Evgenyi Shalaev
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  3. Thomas K. Karami
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  4. James Cunningham
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  5. Hongwen M. Rivers
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  6. Shaoxin Feng
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  8. Nigel K. H. Slater
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Corresponding author

Correspondence to Nigel K. H. Slater .

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Editors and Affiliations

  1. Chief Operating Officer, Tarsus Pharmaceuticals Inc, Irvine, CA, USA

    Seshadri Neervannan

  2. Professor of Pharmaceutical Sciences, University of Colorado Anschutz Medical Campus, Aurora, CO, USA

    Uday B. Kompella

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Rodrigues, G.A. et al. (2021). Manufacturing Considerations and Challenges for AAV Ocular Gene Therapy. In: Neervannan, S., Kompella, U.B. (eds) Ophthalmic Product Development. AAPS Advances in the Pharmaceutical Sciences Series, vol 37. Springer, Cham. https://doi.org/10.1007/978-3-030-76367-1_22

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