Skip to main content

Advertisement

Log in

Superior human hepatocyte transduction with adeno-associated virus vector serotype 7

  • Article
  • Published:
Gene Therapy Submit manuscript

Abstract

Although therapeutic outcomes have been achieved in hemophilia patients after delivery of clotting factor genes to the liver using adeno-associated virus (AAV) vectors, it is well known that the preclinical results generated from hemophilia animal models have not been directly predictive of successful translation in humans. To address this discrepancy humanized mouse models have recently been used to predict AAV transduction efficiency for human hepatocytes. In this study we evaluated AAV vector transduction from several serotypes in human liver hepatocytes xenografted into chimeric mice. After systemic administration of AAV vectors encoding a GFP transgene in humanized mice, the liver was harvested for either immunohistochemistry staining or flow cytometry assay for AAV human hepatocyte transduction analysis. We observed that AAV7 consistently transduced human hepatocytes more efficiently than other serotypes in both immunohistochemistry assay and flow cytometry analysis. To better assess the future application of AAV7 for systemic administration in the treatment of hemophilia or other liver diseases, we analyzed the prevalence of neutralizing antibodies (NAbs) to AAV7 in sera from healthy subjects and patients with hemophilia. In the general population, the prevalence of NAbs to AAV7 was lower than that of AAV2 or AAV3B. However, a higher prevalence of AAV7 NAbs was found in patients with hemophilia. In summary, results from this study suggest that AAV7 vectors should be considered as an effective vehicle for human liver targeting in future clinical trials.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Subscribe and save

Springer+ Basic
EUR 32.99 /Month
  • Get 10 units per month
  • Download Article/Chapter or Ebook
  • 1 Unit = 1 Article or 1 Chapter
  • Cancel anytime
Subscribe now

Buy Now

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

References

  1. George LA, Sullivan SK, Giermasz A, Rasko JEJ, Samelson-Jones BJ, Ducore J. et al. Hemophilia B gene therapy with a high-specific-activity factor IX variant. N Engl J Med. 2017;377:2215–27. https://doi.org/10.1056/NEJMoa1708538.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Nathwani AC, Reiss UM, Tuddenham EG, Rosales C, Chowdary P, McIntosh J. et al. Long-term safety and efficacy of factor IX gene therapy in hemophilia B. N Engl J Med. 2014;371:1994–2004. https://doi.org/10.1056/NEJMoa1407309.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Nathwani AC, Tuddenham EG, Rangarajan S, Rosales C, McIntosh J, Linch DC. et al. Adenovirus-associated virus vector-mediated gene transfer in hemophilia B. N Engl J Med. 2011;365:2357–65. https://doi.org/10.1056/NEJMoa1407309.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Baruteau J, Waddington SN, Alexander IE, Gissen P. Gene therapy for monogenic liver diseases: clinical successes, current challenges and future prospects. J Inherit Metab Dis. 2017;40:497–517. https://doi.org/10.1007/s10545-017-0053-3.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. George LA. Hemophilia gene therapy comes of age. Blood Adv. 2017;1:2591–9. https://doi.org/10.1182/bloodadvances.2017009878.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Hocquemiller M, Giersch L, Audrain M, Parker S, Cartier N. Adeno-associated virus-based gene therapy for CNS diseases. Hum Gene Ther. 2016;27:478–96. https://doi.org/10.1089/hum.2016.087.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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

    Article  CAS  Google Scholar 

  8. Balakrishnan B, Jayandharan GR. Basic biology of adeno-associated virus (AAV) vectors used in gene therapy. Curr Gene Ther. 2014;14:86–100.

    Article  CAS  Google Scholar 

  9. Zincarelli C, Soltys S, Rengo G, Rabinowitz JE. Analysis of AAV serotypes 1-9 mediated gene expression and tropism in mice after systemic injection. Mol Ther. 2008;16:1073–80. https://doi.org/10.1038/mt.2008.76.

    Article  CAS  PubMed  Google Scholar 

  10. Srivastava A. In vivo tissue-tropism of adeno-associated viral vectors. Curr Opin Virol. 2016;21:75–80. https://doi.org/10.1016/j.coviro.2016.08.003.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Nathwani AC, Gray JT, Ng CY, Zhou J, Spence Y, Waddington SN. et al. Self-complementary adeno-associated virus vectors containing a novel liver-specific human factor IX expression cassette enable highly efficient transduction of murine and nonhuman primate liver. Blood. 2006;107:2653–61. https://doi.org/10.1182/blood-2005-10-4035.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Nathwani AC, Rosales C, McIntosh J, Rastegarlari G, Nathwani D, Raj D. et al. Long-term safety and efficacy following systemic administration of a self-complementary AAV vector encoding human FIX pseudotyped with serotype 5 and 8 capsid proteins. Mol Ther. 2011;19:876–85. https://doi.org/10.1038/mt.2010.274.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Nathwani AC, Gray JT, McIntosh J, Ng CY, Zhou J, Spence Y. et al. Safe and efficient transduction of the liver after peripheral vein infusion of self-complementary AAV vector results in stable therapeutic expression of human FIX in nonhuman primates. Blood. 2007;109:1414–21. https://doi.org/10.1182/blood-2006-03-010181.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Wang L, Calcedo R, Nichols TC, Bellinger DA, Dillow A, Verma IM. et al. Sustained correction of disease in naive and AAV2-pretreated hemophilia B dogs: AAV2/8-mediated, liver-directed gene therapy. Blood. 2005;105:3079–86. https://doi.org/10.1182/blood-2004-10-3867.

    Article  CAS  PubMed  Google Scholar 

  15. Jiang H, Couto LB, Patarroyo-White S, Liu T, Nagy D, Vargas JA. et al. Effects of transient immunosuppression on adenoassociated, virus-mediated, liver-directed gene transfer in rhesus macaques and implications for human gene therapy. Blood. 2006;108:3321–8. https://doi.org/10.1182/blood-2006-04-017913.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Manno CS, Pierce GF, Arruda VR, Glader B, Ragni M, Rasko JJ. et al. Successful transduction of liver in hemophilia by AAV-Factor IX and limitations imposed by the host immune response. Nat Med. 2006;12:342–7. https://doi.org/10.1038/nm1358.

    Article  CAS  PubMed  Google Scholar 

  17. Nietupski JB, Hurlbut GD, Ziegler RJ, Chu Q, Hodges BL, Ashe KM. et al. Systemic administration of AAV8-alpha-galactosidase A induces humoral tolerance in nonhuman primates despite low hepatic expression. Mol Ther. 2011;19:1999–2011. https://doi.org/10.1038/mt.2011.119.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Davidoff AM, Gray JT, Ng CY, Zhang Y, Zhou J, Spence Y. et al. Comparison of the ability of adeno-associated viral vectors pseudotyped with serotype 2, 5, and 8 capsid proteins to mediate efficient transduction of the liver in murine and nonhuman primate models. Mol Ther. 2005;11:875–88. https://doi.org/10.1016/j.ymthe.2004.12.022.

    Article  CAS  PubMed  Google Scholar 

  19. Markusic DM, Nichols TC, Merricks EP, Palaschak B, Zolotukhin I, Marsic D. et al. Evaluation of engineered AAV capsids for hepatic factor IX gene transfer in murine and canine models. J Transl Med. 2017;15:94. https://doi.org/10.1186/s12967-017-1200-1.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Bissig-Choisat B, Wang L, Legras X, Saha PK, Chen L, Bell P. et al. Development and rescue of human familial hypercholesterolaemia in a xenograft mouse model. Nat Commun. 2015;6:7339 https://doi.org/10.1038/ncomms8339.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Azuma H, Paulk N, Ranade A, Dorrell C, Al-Dhalimy M, Ellis E. et al. Robust expansion of human hepatocytes in Fah−/−/Rag2−/−/Il2rg−/− mice. Nat Biotechnol. 2007;25:903–10. https://doi.org/10.1038/nbt1326.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Boutin S, Monteilhet V, Veron P, Leborgne C, Benveniste O, Montus MF. et al. Prevalence of serum IgG and neutralizing factors against adeno-associated virus (AAV) types 1, 2, 5, 6, 8, and 9 in the healthy population: implications for gene therapy using AAV vectors. Hum Gene Ther. 2010;21:704–12. https://doi.org/10.1089/hum.2009.182.

    Article  CAS  PubMed  Google Scholar 

  23. Calcedo R, Morizono H, Wang L, McCarter R, He J, Jones D. et al. Adeno-associated virus antibody profiles in newborns, children, and adolescents. Clin Vaccine Immunol. 2011;18:1586–8. https://doi.org/10.1128/CVI.05107-11.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Calcedo R, Vandenberghe LH, Gao G, Lin J, Wilson JM. Worldwide epidemiology of neutralizing antibodies to adeno-associated viruses. J Infect Dis. 2009;199:381–90. https://doi.org/10.1086/595830.

    Article  PubMed  Google Scholar 

  25. Mingozzi F, Chen Y, Edmonson SC, Zhou S, Thurlings RM, Tak PP. et al. Prevalence and pharmacological modulation of humoral immunity to AAV vectors in gene transfer to synovial tissue. Gene Ther. 2013;20:417–24. https://doi.org/10.1038/gt.2012.55.

    Article  CAS  PubMed  Google Scholar 

  26. Halbert CL, Miller AD, McNamara S, Emerson J, Gibson RL, Ramsey B. et al. Prevalence of neutralizing antibodies against adeno-associated virus (AAV) types 2, 5, and 6 in cystic fibrosis and normal populations: implications for gene therapy using AAV vectors. Hum Gene Ther. 2006;17:440–7. https://doi.org/10.1089/hum.2006.17.440.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Arruda VR, Stedman HH, Haurigot V, Buchlis G, Baila S, Favaro P. et al. Peripheral transvenular delivery of adeno-associated viral vectors to skeletal muscle as a novel therapy for hemophilia B. Blood. 2010;115:4678–88. https://doi.org/10.1182/blood-2009-12-261156.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Haurigot V, Mingozzi F, Buchlis G, Hui DJ, Chen Y, Basner-Tschakarjan E. et al. Safety of AAV factor IX peripheral transvenular gene delivery to muscle in hemophilia B dogs. Mol Ther. 2010;18:1318–29. https://doi.org/10.1038/mt.2010.73.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Scallan CD, Jiang H, Liu T, Patarroyo-White S, Sommer JM, Zhou S. et al. Human immunoglobulin inhibits liver transduction by AAV vectors at low AAV2 neutralizing titers in SCID mice. Blood. 2006;107:1810–7. https://doi.org/10.1182/blood-2005-08-3229.

    Article  CAS  PubMed  Google Scholar 

  30. Xiao X, Li J, Samulski RJ. Production of high-titer recombinant adeno-associated virus vectors in the absence of helper adenovirus. J Virol. 1998;72:2224–32.

    CAS  PubMed  PubMed Central  Google Scholar 

  31. Lisowski L, Dane AP, Chu K, Zhang Y, Cunningham SC, Wilson EM. et al. Selection and evaluation of clinically relevant AAV variants in a xenograft liver model. Nature. 2014;506:382–6. https://doi.org/10.1038/nature12875.

    Article  CAS  PubMed  Google Scholar 

  32. Wang L, Bell P, Somanathan S, Wang Q, He Z, Yu H. et al. Comparative study of liver gene transfer with AAV vectors based on natural and engineered AAV capsids. Mol Ther. 2015;23:1877–87. https://doi.org/10.1038/mt.2015.179.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Li S, Ling C, Zhong L, Li M, Su Q, He R. et al. Efficient and targeted transduction of nonhuman primate liver with systemically delivered optimized AAV3B vectors. Mol Ther. 2015;23:1867–76. https://doi.org/10.1038/mt.2015.174.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Vercauteren K, Hoffman BE, Zolotukhin I, Keeler GD, Xiao JW, Basner-Tschakarjan E. et al. Superior in vivo transduction of human hepatocytes using engineered AAV3 capsid. Mol Ther. 2016;24:1042–9. https://doi.org/10.1038/mt.2016.61.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Gao GP, Alvira MR, Wang L, Calcedo R, Johnston J, Wilson JM. Novel adeno-associated viruses from rhesus monkeys as vectors for human gene therapy. Proc Natl Acad Sci USA. 2002;99:11854–9. https://doi.org/10.1073/pnas.182412299.

    Article  CAS  PubMed  Google Scholar 

  36. Daya S, Berns KI. Gene therapy using adeno-associated virus vectors. Clin Microbiol Rev. 2008;21:583–93. https://doi.org/10.1128/CMR.00008-08.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Gao GP, Lu Y, Sun X, Johnston J, Calcedo R, Grant R, et al. High-level transgene expression in nonhuman primate liver with novel adeno-associated virus serotypes containing self-complementary genomes. J Virol. 2006;80:6192–4. https://doi.org/10.1128/JVI.00526-06.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Gao G, Lu Y, Calcedo R, Grant RL, Bell P, Wang L. et al. Biology of AAV serotype vectors in liver-directed gene transfer to nonhuman primates. Mol Ther. 2006;13:77–87. https://doi.org/10.1016/j.ymthe.2005.08.017.

    Article  CAS  PubMed  Google Scholar 

  39. Wang L, Louboutin JP, Bell P, Greig JA, Li Y, Wu D, et al. Muscle-directed gene therapy for hemophilia B with more efficient and less immunogenic AAV vectors. J Thromb Haemost. 2011;9:2009–19. https://doi.org/10.1111/j.1538-7836.2011.04491.x.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Bish LT, Morine K, Sleeper MM, Sanmiguel J, Wu D, Gao G. et al. Adeno-associated virus (AAV) serotype 9 provides global cardiac gene transfer superior to AAV1, AAV6, AAV7, and AAV8 in the mouse and rat. Human gene therapy. 2008;19:1359–68. https://doi.org/10.1089/hum.2008.123.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Lebherz C, Maguire A, Tang W, Bennett J, Wilson JM, Novel AAV. serotypes for improved ocular gene transfer. J Gene Med. 2008;10:375–82. https://doi.org/10.1002/jgm.1126.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Strobel B, Miller FD, Rist W, Lamla T. Comparative analysis of cesium chloride- and iodixanol-based purification of recombinant adeno-associated viral vectors for preclinical applications. Human Gene Therapy Methods. 2015;26:147–57. https://doi.org/10.1089/hgtb.2015.051

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Gao K, Li M, Zhong L, Su Q, Li J, Li S, et al. Empty virions in AAV8 vector preparations reduce transduction efficiency and may cause total viral particle dose-limiting side-effects. Mol Ther Methods Clin Dev. 2014;1:20139. https://doi.org/10.1038/mtm.2013.9.

    Article  CAS  PubMed  Google Scholar 

  44. Erles K, Sebökovà P, Schlehofer JR. Update on the prevalence of serum antibodies (IgG and IgM) to adeno-associated virus (AAV). J Med Virol. 1999;59:406–11.

    Article  CAS  Google Scholar 

  45. Li C, Narkbunnam N, Samulski RJ, Asokan A, Hu G, Jacobson LJ. et al. Neutralizing antibodies against adeno-associated virus examined prospectively in pediatric patients with hemophilia. Gene Ther. 2012;19:288–94. https://doi.org/10.1038/gt.2011.90.

    Article  CAS  PubMed  Google Scholar 

  46. Murphy SL, Li H, Mingozzi F, Sabatino DE, Hui DJ, Edmonson SA, et al. Diverse IgG subclass responses to adeno-associated virus infection and vector administration. J Med Virol. 2009;81:65–74. https://doi.org/10.1002/jmv.21360.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

We thank UNC Vector Core for AAV vector production. The authors acknowledge the UNC Histological Research Core and Flow Cytometry Core Facilities for their assistance in liver immunohistochemistry and flow cytometry analysis. This work was supported by National Institutes of Health Grants R01HL125749 and R01HL144661 (to TCN and CL), and R01AI117408 and P01HL112761 (to RJS and CL).

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Chengwen Li.

Ethics declarations

Conflict of interest

RJS is the founder and a shareholder at Asklepios BioPharmaceutical and Bamboo Therapeutics, Inc. He holds patents that have been licensed by UNC to Asklepios Biopharmaceutical, for which he receives royalties. He has consulted for Baxter Healthcare and has received payment for speaking. CL is a cofounder of Bedrock Therapeutics, Inc. He has licensed patents by UNC and has received royalties from Bedrock Therapeutics and Asklepios Biopharmaceutical. The other authors declare that they have no conflict of interest.

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Shao, W., Pei, X., Cui, C. et al. Superior human hepatocyte transduction with adeno-associated virus vector serotype 7. Gene Ther 26, 504–514 (2019). https://doi.org/10.1038/s41434-019-0104-5

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/s41434-019-0104-5

  • Springer Nature Limited

This article is cited by

Navigation