Directed Evolution of AAV Serotype 5 for Increased Hepatocyte Transduction and Retained Low Humoral Seroreactivity
| Autor: | Randolph Qian, Juan Li, Bin Xiao, Xiao Xiao |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2020 |
| Předmět: |
0301 basic medicine
lcsh:QH426-470 Genetic enhancement Transgene viruses Population Virus 03 medical and health sciences Transduction (genetics) 0302 clinical medicine Genetics medicine lcsh:QH573-671 education Molecular Biology education.field_of_study biology lcsh:Cytology Virology lcsh:Genetics 030104 developmental biology medicine.anatomical_structure 030220 oncology & carcinogenesis Hepatocyte Humoral immunity biology.protein Molecular Medicine Original Article Antibody |
| Zdroj: | Molecular Therapy. Methods & Clinical Development Molecular Therapy: Methods & Clinical Development, Vol 20, Iss, Pp 122-132 (2021) |
| ISSN: | 2329-0501 |
| Popis: | Most recombinant adeno-associated virus (AAV) capsids utilized in liver gene therapy have significant levels of pre-existing neutralizing antibodies in the human population. These neutralizing factors limit the patient pools eligible for receiving AAV-mediated therapies. AAV serotype 5 (AAV5) does not face the same barrier of humoral immunity as most AAV serotypes due to its low seroprevalence. However, AAV5 can only facilitate a low level of transgene expression in the liver, constraining its application to a small number of liver diseases. To improve the liver transduction of AAV5 while retaining its low seroprevalence, we constructed a library of AAV5 mutants via random mutagenesis and screened in Huh7 cells. Two molecularly evolved AAV5 variants, MV50 and MV53, demonstrated significantly increased transduction efficiency in Huh7 cells (∼12×) and primary human hepatocytes (∼10×). All variants had retained low seroreactivity toward pooled human immunoglobulin G (IgG) when compared to AAV5, which was significantly less seroreactive than AAV9. Functional characterization of the mutants also revealed insights into the functions of various domains, especially the VR-I, in the AAV5 capsid. The result is AAV5 variant capsids with much enhanced human hepatocyte transduction, potentially useful for liver-directed gene therapy. Graphical Abstract AAV5 has a distinct advantage in evading pre-existing neutralizing antibodies but is hampered by poor transduction efficiency. To improve AAV5-based liver gene therapy, we evolved and screened AAV5 mutants on human liver cell lines. The result is that more efficient human liver tropic AAV5 capsids retained low humoral seroreactivity. |
| Databáze: | OpenAIRE |
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