Recombinant adeno-associated virus as a delivery platform for ocular gene therapy: A comprehensive review

Abstract

Adeno-associated virus (AAV) has emerged as a leading platform for in vivo gene therapy, particularly in ocular diseases. AAV-based therapies are characterized by low pathogenicity and broad tissue tropism and have demonstrated clinical success, as exemplified by voretigene neparvovec-rzyl (Luxturna) being the first gene therapy to be approved by the U.S. Food and Drug Administration to treat RPE65-associated Leber congenital amaurosis (LCA). However, several challenges remain in the development of AAV-based gene therapies, including immune responses, limited cargo capacity, and the need for enhanced transduction efficiency, especially for intravitreal delivery to photoreceptors and retinal pigment epithelium cells. This review explores the biology of AAVs in the context of gene therapy, innovations in capsid engineering, and clinical advancements in AAV-based ocular gene therapy. We highlight ongoing clinical trials targeting inherited retinal diseases and acquired conditions, discuss immune-related limitations, and examine novel strategies for enhancing AAV vector performance to address current barriers.

Keywords: AAV; adeno-associated virus; capsid engineering; immune response; inherited retinal diseases; ocular gene therapy.

Graphical abstract

Figure 1

Characterization of AAV capsid structures, genomes, and tissue tropisms (A) AAV capsid surface model demonstrates the icosahedral 2-, 3- and 5-fold axes. (B) Genome structure of wtAAV and rAAV. Within the wtAAV genome, the transcription of Rep78 and Rep52 genes is regulated by promoters p5 and p19, respectively. These transcripts can undergo alternative splicing, producing two shorter transcripts, Rep68 and Rep40. Additionally, the transcription of the Cap gene is regulated by promoter p40. The Cap transcripts undergo alternative splicing, resulting in three viral structural proteins: VP1, VP2, and VP3. The rAAV genome is created by replacing viral genes with an expression cassette containing the transgene of interest flanked by two ITRs, the sole cis element necessary for DNA replication and packaging. rAAV production is accomplished by providing Rep, Cap and, Ad helper gene functions in trans. (C) Preferential tissue tropisms of natural AAV serotypes.

Figure 2

Analysis of rAAV-based gene therapy clinical trials for ocular disorders A total of 78 clinical trials (both ongoing and completed) related to rAAV-based ocular gene therapy were identified (ClinicalTrials.gov; Table S1). The diagram specifically encompasses distinct investigational drugs belonging to the same phase within identical disease categories (i.e., if a particular investigational drug is involved in multiple trials within the same phase, it is considered only once). As such, it is important to note that this diagram provides a selective view and does not account for the total number of clinical trials of rAAV-based ocular gene therapy.

Figure 3

Challenges in rAAV-based ocular gene therapy First, pre-existing antibodies against rAAVs can prevent cell/tissue transduction following intravitreal administration. Second, ocularly delivered rAAVs can induce both innate and adaptive immune responses in the eye, resulting in severe inflammation and significantly affecting therapeutic efficacy and safety. Third, rAAV, specifically rAAV2, molecules delivered through intravitreal injection are not able to penetrate the outer retina where photoreceptors and RPE are located, both of which are degenerated in most IRDs. This limitation is primarily due to binding between rAAV2 and HSPG. HSPG is the primary receptor that rAAV2 binds to and is enriched in both ILM and outer limiting membrane (OLM), as well as other barriers, such as the extracellular matrix in the retina.

Figure 4

Schematic representation of subretinal, intravitreal, and suprachoroidal injection of rAAV (A) Subretinal injection is a major administration route used in most current clinical trials for rAAV-based ocular gene therapy. rAAVs delivered through this route mainly transduce RPE cells and photoreceptors. (B) Intravitreal injection is a promising route of administration because it is easier to deliver and less invasive compared to subretinal injection. However, this administration route limits the rAAV transduction mainly to the inner retinal cells due to multiple barrier layers in the retina. (C) A microinjector-based suprachoroidal injection, which can be conducted in an office setting, facilitates the delivery of vectors to the suprachoroidal space, which is situated between the choroid and sclera. Once administered into the suprachoroidal space, rAAVs spread posteriorly and circumferentially.