Recombinant AAV vectors for enhanced expression of authentic IgG

Sebastian P. Fuchs, José M. Martinez-Navio, Guangping Gao, Ronald Charles Desrosiers

Research output: Contribution to journalArticle

9 Citations (Scopus)

Abstract

Adeno-associated virus (AAV) has become a vector of choice for the treatment of a variety of genetic diseases that require safe and long-term delivery of a missing protein. Muscledirected gene transfer for delivery of protective antibodies against AIDS viruses and other pathogens has been used experimentally in mice and monkeys. Here we examined a number of variations to AAV vector design for the ability to produce authentic immunoglobulin G (IgG) molecules. Expression of rhesus IgG from a single single-stranded AAV (ssAAV) vector (one vector approach) was compared to expression from two self-complementary AAV (scAAV) vectors, one for heavy chain and one for light chain (two vector approach). Both the one vector and the two vector approaches yielded considerable levels of expressed fulllength IgG. A number of modifications to the ssAAV expression system were then examined for their ability to increase the efficiency of IgG expression. Inclusion of a furin cleavage sequence with a linker peptide just upstream of the 2A self-cleaving sequence from footand-mouth disease virus (F2A) increased IgG expression approximately 2 fold. Inclusion of these sequences also helped to ensure a proper sequence at the C-terminal end of the heavy chain. Inclusion of the post-transcriptional regulatory element from woodchuck hepatitis virus (WPRE) further increased IgG expression 1.5-2.0 fold. IgG1 versions of the two rhesus IgGs that were examined consistently expressed better than the IgG2 forms. In contrast to what has been reported for AAV2-mediated expression of other proteins, introduction of capsid mutations Y445F and Y731F did not increase ssAAV1-mediated expression of IgG as determined by transduction experiments in cell culture. Our findings provide a rational basis for AAV vector design for expression of authentic IgG.

Original languageEnglish (US)
Article numbere0158009
JournalPLoS One
Volume11
Issue number6
DOIs
StatePublished - Jun 1 2016

Fingerprint

Dependovirus
immunoglobulin G
Viruses
Immunoglobulin G
Woodchuck hepatitis virus
mouth diseases
Mouth Diseases
viruses
Woodchuck Hepatitis B Virus
Transcriptional Regulatory Elements
capsid
Furin
genetic disorders
Gene transfer
Inborn Genetic Diseases
gene transfer
monkeys
Capsid Proteins
Pathogens
cell culture

ASJC Scopus subject areas

  • Medicine(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)

Cite this

Recombinant AAV vectors for enhanced expression of authentic IgG. / Fuchs, Sebastian P.; Martinez-Navio, José M.; Gao, Guangping; Desrosiers, Ronald Charles.

In: PLoS One, Vol. 11, No. 6, e0158009, 01.06.2016.

Research output: Contribution to journalArticle

Fuchs, Sebastian P. ; Martinez-Navio, José M. ; Gao, Guangping ; Desrosiers, Ronald Charles. / Recombinant AAV vectors for enhanced expression of authentic IgG. In: PLoS One. 2016 ; Vol. 11, No. 6.
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