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Mol Ther. 2018 Jan 3;26(1):289-303. doi: 10.1016/j.ymthe.2017.09.021. Epub 2017 Sep 25.

Bioengineered AAV Capsids with Combined High Human Liver Transduction In Vivo and Unique Humoral Seroreactivity.

Author information

1
Departments of Pediatrics and Genetics, Stanford University, Stanford, CA 94305, USA.
2
Translational Vectorology Group, Children's Medical Research Institute, University of Sydney, Sydney, NSW, Australia.
3
Oregon Stem Cell Center, Oregon Health & Science University, Portland, OR 97239, USA.
4
Genethon and INSERM U951, Evry, France.
5
Department of Haematology, UCL Cancer Institute, London, UK.
6
Department of Surgery, St. Jude Children's Research Hospital, Memphis, TN 38105, USA.
7
Cytogenetic Shared Resource, St. Jude Children's Research Hospital, Memphis, TN 38105, USA.
8
Department of Haematology, UCL Cancer Institute, London, UK; Department of Haematology and Katharine Dormandy Haemophilia Centre & Thrombosis Unit, Royal Free London NHS Foundation Trust Hospital, London, UK; National Health Services Blood and Transplant, Watford, UK.
9
Genethon and INSERM U951, Evry, France; University Pierre and Marie Curie, Paris, France.
10
Translational Vectorology Group, Children's Medical Research Institute, University of Sydney, Sydney, NSW, Australia; Military Institute of Hygiene and Epidemiology (MIHE), Puławy, Poland.
11
Departments of Pediatrics and Genetics, Stanford University, Stanford, CA 94305, USA. Electronic address: markay@stanford.edu.

Abstract

Existing recombinant adeno-associated virus (rAAV) serotypes for delivering in vivo gene therapy treatments for human liver diseases have not yielded combined high-level human hepatocyte transduction and favorable humoral neutralization properties in diverse patient groups. Yet, these combined properties are important for therapeutic efficacy. To bioengineer capsids that exhibit both unique seroreactivity profiles and functionally transduce human hepatocytes at therapeutically relevant levels, we performed multiplexed sequential directed evolution screens using diverse capsid libraries in both primary human hepatocytes in vivo and with pooled human sera from thousands of patients. AAV libraries were subjected to five rounds of in vivo selection in xenografted mice with human livers to isolate an enriched human-hepatotropic library that was then used as input for a sequential on-bead screen against pooled human immunoglobulins. Evolved variants were vectorized and validated against existing hepatotropic serotypes. Two of the evolved AAV serotypes, NP40 and NP59, exhibited dramatically improved functional human hepatocyte transduction in vivo in xenografted mice with human livers, along with favorable human seroreactivity profiles, compared with existing serotypes. These novel capsids represent enhanced vector delivery systems for future human liver gene therapy applications.

KEYWORDS:

AAV; evolution; hepatocyte; human; library; liver; neutralization; screen; transduction

PMID:
29055620
PMCID:
PMC5763027
DOI:
10.1016/j.ymthe.2017.09.021
[Indexed for MEDLINE]
Free PMC Article

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