Optimization of stealth adeno-associated virus vectors by randomization of immunogenic epitopes

被引:44
作者
Maersch, Stephan [1 ]
Huber, Anke [1 ]
Buening, Hildegard [1 ,2 ]
Hallek, Michael [1 ,2 ]
Perabo, Luca [1 ]
机构
[1] Univ Hosp Cologne, Clin Internal Med 1, D-50937 Cologne, Germany
[2] Univ Cologne, Ctr Mol Med, D-50931 Cologne, Germany
关键词
Adeno-associated virus; Directed evolution; Capsid library; Capsid engineering; Pre-existing immunity; Antibody neutralization; Antibody escaping; Capsid biology; Vector optimization; Tropism; HEPARAN-SULFATE PROTEOGLYCAN; GENE-THERAPY VECTOR; NEUTRALIZING ANTIBODIES; DIRECTED EVOLUTION; IMMUNE-RESPONSES; TYPE-2; CAPSIDS; IN-VITRO; BINDING; IDENTIFICATION; TROPISM;
D O I
10.1016/j.virol.2009.10.021
中图分类号
Q93 [微生物学];
学科分类号
071005 ; 100705 ;
摘要
Therapeutic gene transfer by adeno-associated virus of serotype 2 (AAV-2) vectors is hampered in patients with pre-existing immunity. Molecular engineering was recently used to identify key immunogenic amino acid residues of the viral capsid and generate mutants with decreased antibody recognition. Here we explored the importance of finely tuning amino acid identity at immunogenic sites to optimize vector phenotype. A capsid library was generated by codon randomization at five positions where substitutions were shown to yield antibody evading phenotypes. Screening this library to isolate immune-escaping Mutants allowed an exhaustive scan of combinations of the 20 natural amino acids at each position and yielded variants that remained infectious when incubated with serum or IVIG concentrations that completely neutralize AAV-2. Clones obtained replacing different residues at the same positions displayed strikingly different phenotypes, demonstrating that a precise choice of amino acid substitutions is fundamental to optimize immune-escaping, packaging ability, infectivity and tropism. (C) 2009 Elsevier Inc. All rights reserved.
引用
收藏
页码:167 / 175
页数:9
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