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.

  • Article
  • Published:

IgG-cleavage protein allows therapeutic AAV gene delivery in passively immunized MPS IIIA mice

Gene Therapy volume 30, pages 377–385 (2023)Cite this article

Subjects

Abstract

The widespread pre-existing αAAV-Abs in humans pose a critical challenge in translation of AAV gene therapy. The IgG degrading enzyme of Streptococci (IdeS) is demonstrated to specifically cleave IgG of humans and other species (not mouse). This study developed a modified new modified IdeS protein product (IdeSop). When incubated in vitro, IdeSop was shown to completely cleave human and rabbit IgGs within 6 h. To test IdeSop in a disease setting, we established a rabbitized αAAV9-Ab+ mouse by an IV infusion of purified acute αAAV9-Ab+ rabbit IgG into MPS IIIA mice, resulting in serum αAAV9-IgG at 1:6,400 and αAAV9-nAbs at 1:800. IdeSop-Ab-cleavage was shown to be dose-dependent. An IV IdeSop infusion at the effective doses resulted in rapid IgG depletion and clearance of pre-existing αAAV9-IgG and αAAV9-nAbs in rabbitized αAAV9-Abs+ MPS IIIA mice. Importantly, an IV injection of a high dose AAV9-hSGSHop vector (5 × 1013vg/kg) at 24 h post IdeSop treatment led to transduction as effective in αAAV9-Abs+ MPS IIIA mice, as in αAAV9-Abs-negative controls. We believe that transient IdeSop administration may offer a great tool to address the pre-existing-αAAV-Abs for the translation of rAAV gene therapy to treat diseases in humans, making effective rAAV gene therapy available to all patients in need.

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

Fig. 1: Expression of functional recombinant IdeS.
Fig. 2: Dose response IgG cleavage of IdeSop.
Fig. 3: Depletion of αAAV9-IgG by IdeSop in vivo in rabbitized mice wt C57BL/6 mice were given an IV injection of 100 μl of purified αAAV9-Ab-positive Rb IgG via tail vein.
Fig. 4: IdeSop αAAV9-IgG depletion retained the transduction efficiency of IV-delivered scAAV9-hSGSH vector in rabbitized αAAV9-IgG+ MPS IIIA mice MPS IIIA mice (n = 8) were given an IV injection of Rb-IgG, of which half (n = 4) were then given an IV injection of IdeSop (0.45 mg/kg).
Fig. 5: IdeSop αAAV9-IgG depletion allow efficient rSGSH expression and clearance of GAG contents in the CNS and periphery tissues in rabbitized αAAV9-IgG+ MPS IIIA mice following an IV AAV9-hSGSH vector delivery MPS IIIA mice (n = 4/group) were given an IV injection of Rb-IgG, of which four were then given an IV injection of IdeSop (0.45 mg/kg).

Similar content being viewed by others

References

  1. 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.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. 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.

    Article  CAS  PubMed  Google Scholar 

  3. 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.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Foust KD, Wang X, McGovern VL, Braun L, Bevan AK, Haidet AM, et al. Rescue of the spinal muscular atrophy phenotype in a mouse model by early postnatal delivery of SMN. Nat Biotechnol. 2010;28:271–4.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Fu H, Dirosario J, Killedar S, Zaraspe K, McCarty DM. Correction of neurological disease of mucopolysaccharidosis IIIB in adult mice by rAAV9 trans-blood-brain barrier gene delivery. Mol Ther. 2011;19:1025–33.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Ruzo A, Marco S, Garcia M, Villacampa P, Ribera A, Ayuso E, et al. Correction of Pathological Accumulation of Glycosaminoglycans in Central Nervous System and Peripheral Tissues of MPSIIIA Mice Through Systemic AAV9 Gene Transfer. Hum Gene Ther. 2012;23:1237–46.

    Article  CAS  PubMed  Google Scholar 

  7. Foust KD, Nurre E, Montgomery CL, Hernandez A, Chan CM, Kaspar BK. Intravascular AAV9 preferentially targets neonatal neurons and adult astrocytes. Nat Biotechnol. 2009;27:59–65.

    Article  CAS  PubMed  Google Scholar 

  8. Duque S, Joussemet B, Riviere C, Marais T, Dubreil L, Douar AM, et al. Intravenous administration of self-complementary AAV9 enables transgene delivery to adult motor neurons. Mol Ther. 2009;17:1187–96.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Fu H, Cataldi MP, Ware TA, Zaraspe K, Meadows AS, Murrey DA, et al. Functional correction of neurological and somatic disorders at later stages of disease in MPS IIIA mice by systemic scAAV9-hSGSH gene delivery. Mol Ther Methods Clin Dev. 2016;3:16036.

    Article  PubMed  PubMed Central  Google Scholar 

  10. Bailey RM, Armao D, Nagabhushan Kalburgi S, Gray SJ. Development of Intrathecal AAV9 Gene Therapy for Giant Axonal Neuropathy. Mol Ther Methods Clin Dev. 2018;9:160–71.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. 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.

    Article  CAS  PubMed  Google Scholar 

  12. 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.

    Article  PubMed  Google Scholar 

  13. Fu H, Meadows AS, Pineda RJ, Kunkler KL, Truxal KV, McBride KL, et al. Differential Prevalence of Antibodies Against Adeno-Associated Virus in Healthy Children and Patients with Mucopolysaccharidosis III: Perspective for AAV-Mediated Gene Therapy. Human Gene Ther Clin Dev. 2017;28:187–96.

    Article  CAS  Google Scholar 

  14. 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.

    Article  CAS  PubMed  Google Scholar 

  15. Wang L, Calcedo R, Bell P, Lin J, Grant RL, Siegel DL, et al. Impact of pre-existing immunity on gene transfer to nonhuman primate liver with adeno-associated virus 8 vectors. Human Gene Ther. 2011;22:1389–401.

    Article  CAS  Google Scholar 

  16. McIntosh JH, Cochrane M, Cobbold S, Waldmann H, Nathwani SA, Davidoff AM, et al. Successful attenuation of humoral immunity to viral capsid and transgenic protein following AAV-mediated gene transfer with a non-depleting CD4 antibody and cyclosporine. Gene Ther. 2012;19:78–85.

    Article  CAS  PubMed  Google Scholar 

  17. Mingozzi F, Chen Y, Murphy SL, Edmonson SC, Tai A, Price SD, et al. Pharmacological modulation of humoral immunity in a nonhuman primate model of AAV gene transfer for hemophilia B. Mol Ther. 2012;20:1410–6.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Corti M, Elder M, Falk D, Lawson L, Smith B, Nayak S, et al. B-Cell Depletion is Protective Against Anti-AAV Capsid Immune Response: A Human Subject Case Study. Mol Ther Method Clin Dev. 2014;1:14033.

    Article  Google Scholar 

  19. Chicoine LG, Montgomery CL, Bremer WG, Shontz KM, Griffin DA, Heller KN, et al. Plasmapheresis eliminates the negative impact of AAV antibodies on microdystrophin gene expression following vascular delivery. Mol Ther. 2014;22:338–47.

    Article  CAS  PubMed  Google Scholar 

  20. Chicoine LG, Rodino-Klapac LR, Shao G, Xu R, Bremer WG, Camboni M, et al. Vascular delivery of rAAVrh74.MCK.GALGT2 to the gastrocnemius muscle of the rhesus macaque stimulates the expression of dystrophin and laminin alpha2 surrogates. Mol Ther. 2014;22:713–24.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Velazquez VM, Meadows AS, Pineda RJ, Camboni M, McCarty DM, Fu H. Effective Depletion of Pre-existing Anti-AAV Antibodies Requires Broad Immune Targeting. Mol Ther Methods Clin Dev. 2017;4:159–68.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Mingozzi F, Anguela XM, Pavani G, Chen Y, Davidson RJ, Hui DJ, et al. Overcoming preexisting humoral immunity to AAV using capsid decoys. Sci Transl Med. 2013;5:194ra92.

    Article  PubMed  PubMed Central  Google Scholar 

  23. Maersch S, Huber A, Buning H, Hallek M, Perabo L. Optimization of stealth adeno-associated virus vectors by randomization of immunogenic epitopes. Virology. 2010;397:167–75.

    Article  CAS  PubMed  Google Scholar 

  24. Asokan A, Samulski RJ. AAV does the shuffle. Nat Biotechnol. 2006;24:158–60.

    Article  CAS  PubMed  Google Scholar 

  25. Lee GK, Maheshri N, Kaspar B, Schaffer DV. PEG conjugation moderately protects adeno-associated viral vectors against antibody neutralization. Biotechnol Bioeng. 2005;92:24–34.

    Article  CAS  PubMed  Google Scholar 

  26. Monteilhet V, Saheb S, Boutin S, Leborgne C, Veron P, Montus MF, et al. A 10 patient case report on the impact of plasmapheresis upon neutralizing factors against adeno-associated virus (AAV) types 1, 2, 6, and 8. Mol Ther. 2011;19:2084–91.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Unzu C, Hervas-Stubbs S, Sampedro A, Mauleon I, Mancheno U, Alfaro C, et al. Transient and intensive pharmacological immunosuppression fails to improve AAV-based liver gene transfer in non-human primates. J Transl Med. 2012;10:122.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Leborgne C, Barbon E, Alexander JM, Hanby H, Delignat S, Cohen DM, et al. IgG-cleaving endopeptidase enables in vivo gene therapy in the presence of anti-AAV neutralizing antibodies. Nat Med. 2020;26:1096–101.

    Article  CAS  PubMed  Google Scholar 

  29. Elmore ZC, Oh DK, Simon KE, Fanous MM, Asokan A Rescuing AAV gene transfer from neutralizing antibodies with an IgG-degrading enzyme. JCI Insight. 2020;5.

  30. von Pawel-Rammingen U, Johansson BP, Bjorck L. IdeS, a novel streptococcal cysteine proteinase with unique specificity for immunoglobulin G. EMBO J. 2002;21:1607–15.

    Article  Google Scholar 

  31. Vincents B, von Pawel-Rammingen U, Bjorck L, Abrahamson M. Enzymatic characterization of the streptococcal endopeptidase, IdeS, reveals that it is a cysteine protease with strict specificity for IgG cleavage due to exosite binding. Biochemistry. 2004;43:15540–9.

    Article  CAS  PubMed  Google Scholar 

  32. Wenig K, Chatwell L, von Pawel-Rammingen U, Bjorck L, Huber R, Sondermann P. Structure of the streptococcal endopeptidase IdeS, a cysteine proteinase with strict specificity for IgG. Proc Natl Acad Sci USA. 2004;101:17371–6.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Johansson BP, Shannon O, Bjorck L. IdeS: a bacterial proteolytic enzyme with therapeutic potential. PLoS ONE. 2008;3:e1692.

    Article  PubMed  PubMed Central  Google Scholar 

  34. Ryan MH, Petrone D, Nemeth JF, Barnathan E, Bjorck L, Jordan RE. Proteolysis of purified IgGs by human and bacterial enzymes in vitro and the detection of specific proteolytic fragments of endogenous IgG in rheumatoid synovial fluid. Mol Immunol. 2008;45:1837–46.

    Article  CAS  PubMed  Google Scholar 

  35. von Pawel-Rammingen U, Johansson BP, Tapper H, Bjorck L. Streptococcus pyogenes and phagocytic killing. Nat Med. 2002;8:1044–5.

    Article  Google Scholar 

  36. Yang R, Otten MA, Hellmark T, Collin M, Bjorck L, Zhao MH, et al. Successful treatment of experimental glomerulonephritis with IdeS and EndoS, IgG-degrading streptococcal enzymes. Nephrol Dial Transplant. 2010;25:2479–86.

    Article  CAS  PubMed  Google Scholar 

  37. Winstedt L, Jarnum S, Nordahl EA, Olsson A, Runstrom A, Bockermann R, et al. Complete Removal of Extracellular IgG Antibodies in a Randomized Dose-Escalation Phase I Study with the Bacterial Enzyme IdeS-A Novel Therapeutic Opportunity. PLoS ONE. 2015;10:e0132011.

    Article  PubMed  PubMed Central  Google Scholar 

  38. Lorant T, Bengtsson M, Eich T, Eriksson BM, Winstedt L, Jarnum S, et al. Safety, immunogenicity, pharmacokinetics, and efficacy of degradation of anti-HLA antibodies by IdeS (imlifidase) in chronic kidney disease patients. Am J Transplant. 2018;11:2752–62.

    Article  Google Scholar 

  39. Lonze BE, Tatapudi VS, Weldon EP, Min ES, Ali NM, Deterville CL, et al. IdeS (Imlifidase): A Novel Agent That Cleaves Human IgG and Permits Successful Kidney Transplantation Across High-strength Donor-specific Antibody. Ann Surg. 2018;268:488–96.

    Article  PubMed  Google Scholar 

  40. Bobo TA, Samowitz PN, Robinson MI, Fu H. Targeting the Root Cause of Mucopolysaccharidosis IIIA with a New scAAV9 Gene Replacement Vector. Mol Ther Methods Clin Dev. 2020;19:474–85.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Grieger JC, Soltys SM, Samulski RJ. Production of Recombinant Adeno-associated Virus Vectors Using Suspension HEK293 Cells and Continuous Harvest of Vector From the Culture Media for GMP FIX and FLT1 Clinical Vector. Mol Ther. 2016;24:287–97.

    Article  CAS  PubMed  Google Scholar 

  42. Bhaumik M, Muller VJ, Rozaklis T, Johnson L, Dobrenis K, Bhattacharyya R, et al. A mouse model for mucopolysaccharidosis type III A (Sanfilippo syndrome). Glycobiology. 1999;9:1389–96.

    Article  CAS  PubMed  Google Scholar 

  43. Murrey DA, Naughton BJ, Duncan FJ, Meadows AS, Ware TA, Campbell KJ, et al. Feasibility and safety of systemic rAAV9-hNAGLU delivery for treating mucopolysaccharidosis IIIB: toxicology, biodistribution, and immunological assessments in primates. Human Gene Ther Clin Dev. 2014;25:72–84.

    Article  CAS  Google Scholar 

  44. Meliani A, Leborgne C, Triffault S, Jeanson-Leh L, Veron P, Mingozzi F. Determination of anti-adeno-associated virus vector neutralizing antibody titer with an in vitro reporter system. Hum Gene Ther Methods. 2015;26:45–53.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Karpova EA, Voznyi YaV, Keulemans JL, Hoogeveen AT, Winchester B, Tsvetkova IV, et al. A fluorimetric enzyme assay for the diagnosis of Sanfilippo disease type A (MPS IIIA). J Inherit Metab Dis. 1996;19:278–85.

    Article  CAS  PubMed  Google Scholar 

  46. van de Lest CH, Versteeg EM, Veerkamp JH, van Kuppevelt TH. - Quantification and characterization of glycosaminoglycans at the nanogram level by a combined azure A-silver staining in agarose gels. Anal Biochem. 1994;221:356–61.

    Article  PubMed  Google Scholar 

  47. Fu H, Kang L, Jennings JS, Moy SS, Perez A, Dirosario J, et al. Significantly increased lifespan and improved behavioral performances by rAAV gene delivery in adult mucopolysaccharidosis IIIB mice. Gene Ther. 2007;14:1065–77.

    Article  CAS  PubMed  Google Scholar 

  48. de Jong JG, Wevers RA, Laarakkers C, Poorthuis BJ. - Dimethylmethylene blue-based spectrophotometry of glycosaminoglycans in untreated urine: a rapid screening procedure for mucopolysaccharidoses. Clin Chem. 1989;35:1472–7.

    Article  PubMed  Google Scholar 

  49. Meadows AS, Pineda RJ, Goodchild L, Bobo TA, Fu H. Threshold for Pre-existing Antibody Levels Limiting Transduction Efficiency of Systemic rAAV9 Gene Delivery: Relevance for Translation. Mol Ther Methods Clin Dev. 2019;13:453–62.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  50. Neufeld EF, Cantz MJ. Corrective factors for inborn errors of mucopolysaccharide metabolism. Ann N Y Acad Sci. 1971;179:580–7.

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We thank the UNC Protein Production & Purification Core for manufacturing of the AbC product, and UNC Vector Core for vector production services. We appreciate Dr. Douglas M. McCarty’s constructive comments and editing assistance. We also thank Dr. Charles Askew for kindly sharing Huh7 cells. This work was supported by a research grant from the National Institute of Health (NIAID R21AI146653) and donations from the Sanfilippo syndrome patient community through Aislinne’s Wish Foundation and the Abby Grace Foundation. Bobo was also supported by a Gene Therapy Fellowship from North Carolina Biotechnology Center (NCBC Grant # 2020-GTF-6903). HF was also supported as site PI by a STTR grant from NIH/NINDS (R41NS115232).

Author information

Author notes

  1. These authors contributed equally: Tierra A. Bobo, Preston N. Samowitz.

Authors and Affiliations

  1. Gene Therapy Center, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA

    Tierra A. Bobo, Preston N. Samowitz, Michael I. Robinson, Laura I. Montes & Haiyan Fu

  2. Division of Genetics and Metabolism, Department of Pediatrics, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA

    Tierra A. Bobo & Haiyan Fu

  3. Protein Production & Purification Core, School of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA

    Lawrence J. Forsberg, Richard Feng & Nathan I. Nicely

Authors
  1. Tierra A. Bobo
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Preston N. Samowitz
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Michael I. Robinson
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Laura I. Montes
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Lawrence J. Forsberg
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Richard Feng
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Nathan I. Nicely
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Haiyan Fu
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

TB designed and performed the experiments, data analysis and wrote the paper; PS performed the experiments, data collection and analysis, and paper preparation; MR performed tissue collection and ELISA assays; LM performed PAGE, ELISA and NAb assays and data analysis; LF, RF, and NN performed protein production, purification and analysis; HF designed the project and wrote the paper.

Corresponding author

Correspondence to Haiyan Fu.

Ethics declarations

Competing interests

HF is the founder and president of NeuroGT, Inc. Other authors declare no competing interests.

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

Springer Nature or its licensor holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Bobo, T.A., Samowitz, P.N., Robinson, M.I. et al. IgG-cleavage protein allows therapeutic AAV gene delivery in passively immunized MPS IIIA mice. Gene Ther 30, 377–385 (2023). https://doi.org/10.1038/s41434-022-00368-9

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/s41434-022-00368-9

Search

Advanced search

Quick links