AAV cis-regulatory sequences are correlated with ocular toxicity

Area of Science:

• Ophthalmology
• Gene Therapy
• Molecular Biology

Background:

• Adeno-associated viral vectors (AAVs) are widely used in gene therapy due to their low immunogenicity.
• Despite immune privilege, ocular tissues can mount host-cell responses to AAV vectors.
• Investigating AAV-induced toxicity in ocular cells is critical for safe gene therapy development.

Purpose of the Study:

• To investigate the impact of AAV genome structures and capsid types on ocular cell types.
• To identify factors contributing to AAV-induced toxicity in the eye.
• To inform the design of safer AAV vectors for ocular gene therapy.

Main Methods:

• Subretinal injections of various AAV vectors in mice.
• Comprehensive assays evaluating morphology, inflammation, and physiology of ocular tissues.
• Analysis of host-cell responses, including glial and microglial activation and cytokine production.

Main Results:

• Ocular toxicity was observed in photoreceptors and the retinal pigment epithelium (RPE).
• Müller glia and microglia activation, along with increased TNF-α and IL-1β, indicated inflammation.
• A strong correlation was found between specific cis-regulatory sequences (promoters) and toxicity, with RPE-specific and broadly active promoters being toxic, while photoreceptor-specific promoters were not.

Conclusions:

• Ocular AAV toxicity is primarily linked to specific cis-regulatory sequences and their activity, not transgene, capsid, or preparation method.
• Retinal damage results from host responses, particularly involving the RPE and microglia.
• Designing AAV vectors with appropriate cis-regulatory elements is essential for achieving safe, high-dose ocular gene therapy and enhancing therapeutic efficacy.