Journal
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Volume 116, Issue 12, Pages 5785-5794Publisher
NATL ACAD SCIENCES
DOI: 10.1073/pnas.1821000116
Keywords
AAV; gene therapy; toxicity; retina; retinal pigment epithelium
Categories
Funding
- Howard Hughes Medical Institute
- Astellas Pharma Inc.
- City University of Hong Kong Grant [9610345]
- Hong Kong Research Grants Council [21105916]
- National Natural Science Foundation of China [81770937]
- National Institutes of Health/National Eye Institute [K08-EY023993-05]
- Massachusetts Lions Eye Research Fund, Iraty Award
- National Eye Institute Audacious Goals Initiatives Grant [U01 EY025497]
- Howard Hughes Medical Institute Medical Student Fellowship
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Adeno-associated viral vectors (AAVs) have become popular for gene therapy, given their many advantages, including their reduced inflammatory profile compared with that of other viruses. However, even in areas of immune privilege such as the eye, AAV vectors are capable of eliciting host-cell responses. To investigate the effects of such responses on several ocular cell types, we tested multiple AAV genome structures and capsid types using subretinal injections in mice. Assays of morphology, inflammation, and physiology were performed. Pathological effects on photoreceptors and the retinal pigment epithelium (RPE) were observed. Muller glia and microglia were activated, and the proinflammatory cytokines TNF-alpha and IL-1 beta were up-regulated. There was a strong correlation between cis-regulatory sequences and toxicity. AAVs with any one of three broadly active promoters, or an RPE-specific promoter, were toxic, while AAVs with four different photoreceptor-specific promoters were not toxic at the highest doses tested. There was little correlation between toxicity and transgene, capsid type, preparation method, or cellular contaminants within a preparation. The toxic effect was dose-dependent, with the RPE being more sensitive than photoreceptors. Our results suggest that ocular AAV toxicity is associated with certain AAV cis-regulatory sequences and/or their activity and that retinal damage occurs due to responses by the RPE and/or microglia. By applying multiple, sensitive assays of toxicity, AAV vectors can be designed so that they can be used safely at high dose, potentially providing greater therapeutic efficacy.
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