Triple Vectors Expand AAV Transfer Capacity in the Retina

Mol Ther. 2018 Feb 7;26(2):524-541. doi: 10.1016/j.ymthe.2017.11.019. Epub 2017 Dec 5.

Abstract

Retinal gene transfer with adeno-associated viral (AAV) vectors holds great promise for the treatment of inherited retinal degenerations (IRDs). One limit of AAV is its transfer capacity of about 5 kb, which can be expanded to about 9 kb, using dual AAV vectors. This strategy would still not suffice for treatment of IRDs such as Usher syndrome type 1D or Alström syndrome type I (ALMS) due to mutations in CDH23 or ALMS1, respectively. To overcome this limitation, we generated triple AAV vectors, with a maximal transfer capacity of about 14 kb. Transcriptomic analysis following triple AAV transduction showed the expected full-length products along a number of aberrant transcripts. However, only the full-length transcripts are efficiently translated in vivo. We additionally showed that approximately 4% of mouse photoreceptors are transduced by triple AAV vectors and showed correct localization of recombinant ALMS1. The low-photoreceptor transduction levels might justify the modest and transient improvement we observe in the retina of a mouse model of ALMS. However, the levels of transduction mediated by triple AAV vectors in pig retina reached 40% of those observed with single vectors, and this bodes well for further improving the efficiency of triple AAV vectors in the retina.

Keywords: AAV; gene therapy; large gene; retina; triple AAV.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cadherins / genetics
  • Cadherins / metabolism
  • Dependovirus / genetics*
  • Gene Expression
  • Gene Expression Regulation, Viral
  • Gene Transfer Techniques
  • Genes, Reporter
  • Genetic Therapy
  • Genetic Vectors / administration & dosage
  • Genetic Vectors / genetics*
  • HEK293 Cells
  • Humans
  • Mice
  • Mice, Knockout
  • Recombination, Genetic*
  • Retina / metabolism*
  • Swine
  • Transcription, Genetic
  • Transduction, Genetic*
  • Transgenes

Substances

  • Cadherins
  • Cdh23 protein, mouse