Attenuation of Heparan Sulfate Proteoglycan Binding Enhances In Vivo Transduction of Human Primary Hepatocytes with AAV2

Marti Cabanes-Creus; Adrian Westhaus; Renina Gale Navarro; Grober Baltazar; Erhua Zhu; Anais K. Amaya; Sophia H.Y. Liao; Suzanne Scott; Erwan Sallard; Kimberley L. Dilworth; Arkadiusz Rybicki; Matthieu Drouyer; Claus V. Hallwirth; Antonette Bennett; Giorgia Santilli; Adrian J. Thrasher; Mavis Agbandje-McKenna; Ian E. Alexander; Leszek Lisowski

Molecular Therapy: Methods & Clinical Development (Jun 2020)

Attenuation of Heparan Sulfate Proteoglycan Binding Enhances In Vivo Transduction of Human Primary Hepatocytes with AAV2

Marti Cabanes-Creus,
Adrian Westhaus,
Renina Gale Navarro,
Grober Baltazar,
Erhua Zhu,
Anais K. Amaya,
Sophia H.Y. Liao,
Suzanne Scott,
Erwan Sallard,
Kimberley L. Dilworth,
Arkadiusz Rybicki,
Matthieu Drouyer,
Claus V. Hallwirth,
Antonette Bennett,
Giorgia Santilli,
Adrian J. Thrasher,
Mavis Agbandje-McKenna,
Ian E. Alexander,
Leszek Lisowski

Affiliations

Marti Cabanes-Creus: Translational Vectorology Research Unit, Children’s Medical Research Institute, The University of Sydney, Westmead, NSW 2145, Australia
Adrian Westhaus: Translational Vectorology Research Unit, Children’s Medical Research Institute, The University of Sydney, Westmead, NSW 2145, Australia; Great Ormond Street Institute of Child Health, University College London, London WC1N 1EH, UK
Renina Gale Navarro: Translational Vectorology Research Unit, Children’s Medical Research Institute, The University of Sydney, Westmead, NSW 2145, Australia
Grober Baltazar: Translational Vectorology Research Unit, Children’s Medical Research Institute, The University of Sydney, Westmead, NSW 2145, Australia
Erhua Zhu: Translational Vectorology Research Unit, Children’s Medical Research Institute, The University of Sydney, Westmead, NSW 2145, Australia; Gene Therapy Research Unit, Children’s Medical Research Institute & The Children’s Hospital at Westmead, University of Sydney, Westmead, NSW 2145, Australia
Anais K. Amaya: Gene Therapy Research Unit, Children’s Medical Research Institute & The Children’s Hospital at Westmead, University of Sydney, Westmead, NSW 2145, Australia
Sophia H.Y. Liao: Translational Vectorology Research Unit, Children’s Medical Research Institute, The University of Sydney, Westmead, NSW 2145, Australia
Suzanne Scott: Gene Therapy Research Unit, Children’s Medical Research Institute & The Children’s Hospital at Westmead, University of Sydney, Westmead, NSW 2145, Australia; Commonwealth Scientific and Industrial Research Organisation (CSIRO), North Ryde, NSW 2113, Australia
Erwan Sallard: Translational Vectorology Research Unit, Children’s Medical Research Institute, The University of Sydney, Westmead, NSW 2145, Australia
Kimberley L. Dilworth: Translational Vectorology Research Unit, Children’s Medical Research Institute, The University of Sydney, Westmead, NSW 2145, Australia
Arkadiusz Rybicki: Translational Vectorology Research Unit, Children’s Medical Research Institute, The University of Sydney, Westmead, NSW 2145, Australia
Matthieu Drouyer: Translational Vectorology Research Unit, Children’s Medical Research Institute, The University of Sydney, Westmead, NSW 2145, Australia
Claus V. Hallwirth: Gene Therapy Research Unit, Children’s Medical Research Institute & The Children’s Hospital at Westmead, University of Sydney, Westmead, NSW 2145, Australia
Antonette Bennett: Department of Biochemistry and Molecular Biology, Center for Structural Biology, University of Florida, Gainesville, FL 32610, USA
Giorgia Santilli: Great Ormond Street Institute of Child Health, University College London, London WC1N 1EH, UK
Adrian J. Thrasher: Great Ormond Street Institute of Child Health, University College London, London WC1N 1EH, UK
Mavis Agbandje-McKenna: Department of Biochemistry and Molecular Biology, Center for Structural Biology, University of Florida, Gainesville, FL 32610, USA
Ian E. Alexander: Gene Therapy Research Unit, Children’s Medical Research Institute & The Children’s Hospital at Westmead, University of Sydney, Westmead, NSW 2145, Australia; Discipline of Child and Adolescent Health, The University of Sydney, Sydney, NSW 2006, Australia
Leszek Lisowski: Translational Vectorology Research Unit, Children’s Medical Research Institute, The University of Sydney, Westmead, NSW 2145, Australia; Vector and Genome Engineering Facility, Children’s Medical Research Institute, The University of Sydney, Westmead, NSW 2145, Australia; Military Institute of Hygiene and Epidemiology, Biological Threats Identification and Countermeasure Center, 24-100 Puławy, Poland; Corresponding author: Leszek Lisowski, Translational Vectorology Research Unit, Children’s Medical Research Institute, The University of Sydney, Westmead, NSW 2145, Australia.

Journal volume & issue: Vol. 17
pp. 1139 – 1154

Abstract

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Use of the prototypical adeno-associated virus type 2 (AAV2) capsid delivered unexpectedly modest efficacy in an early liver-targeted gene therapy trial for hemophilia B. This result is consistent with subsequent data generated in chimeric mouse-human livers showing that the AAV2 capsid transduces primary human hepatocytes in vivo with low efficiency. In contrast, novel variants generated by directed evolution in the same model, such as AAV-NP59, transduce primary human hepatocytes with high efficiency. While these empirical data have immense translational implications, the mechanisms underpinning this enhanced AAV capsid transduction performance in primary human hepatocytes are yet to be fully elucidated. Remarkably, AAV-NP59 differs from the prototypical AAV2 capsid by only 11 aa and can serve as a tool to study the correlation between capsid sequence/structure and vector function. Using two orthogonal vectorological approaches, we have determined that just 2 of the 11 changes present in AAV-NP59 (T503A and N596D) account for the enhanced transduction performance of this capsid variant in primary human hepatocytes in vivo, an effect that we have associated with attenuation of heparan sulfate proteoglycan (HSPG) binding affinity. In support of this hypothesis, we have identified, using directed evolution, two additional single amino acid substitution AAV2 variants, N496D and N582S, which are highly functional in vivo. Both substitution mutations reduce AAV2’s affinity for HSPG. Finally, we have modulated the ability of AAV8, a highly murine-hepatotropic serotype, to interact with HSPG. The results support our hypothesis that enhanced HSPG binding can negatively affect the in vivo function of otherwise strongly hepatotropic variants and that modulation of the interaction with HSPG is critical to ensure maximum efficiency in vivo. The insights gained through this study can have powerful implications for studies into AAV biology and capsid development for preclinical and clinical applications targeting liver and other organs.

Published in Molecular Therapy: Methods & Clinical Development

ISSN: 2329-0501 (Online)
Publisher: Elsevier
Country of publisher: United States
LCC subjects: Science: Biology (General): Genetics; Science: Biology (General): Cytology
Website: http://www.cell.com/molecular-therapy-family/methods/latest-content

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