Abstract
Inherited retinal dystrophies and optic neuropathies cause chronic disabling loss of visual function. The development of recombinant adeno-associated viral vectors (rAAV) gene therapies in all disease fields have been promising, but the translation to the clinic has been slow. The safety and efficacy profiles of rAAV are linked to the dose of applied vectors. DNA changes in the rAAV gene cassette affect potency, the expression pattern (cell-specificity), and the production yield. Here, we present a library of rAAV vectors and elements that provide a workflow to design novel vectors. We first performed a meta-analysis on recombinant rAAV elements in clinical trials (2007–2020) for ocular gene therapies. We analyzed 33 unique rAAV gene cassettes used in 57 ocular clinical trials. The rAAV gene therapy vectors used six unique capsid variants, 16 different promoters, and six unique polyadenylation sequences. Further, we compiled a list of promoters, enhancers, and other sequences used in current rAAV gene cassettes in preclinical studies. Then, we give an update on pro-viral plasmid backbones used to produce the gene therapy vectors, inverted terminal repeats, production yield, and rAAV safety considerations. Finally, we assess rAAV transgene and bioactivity assays applied to cells or organoids in vitro, explants ex vivo, and clinical studies.
Highlights
Many reviews have been written on recombinant adeno-associated virus vector tropism in ocular tissue, rAAV host cell infection, and potential rAAV-treatable inherited retinal diseases [1,2,3,4,5,6,7,8,9]
The second gene is transcribed without a promoter Synthetic polyA signal/transcriptional pause site frp, pGL4.25 Block-iT PolII miR vector system based on miR-155 expressing artificial miRNAs engineered to a target sequence resulting in target cleavage 7-to-45 fold AAV production increase in physical titer mini-intronic plasmid (MIP) backbones for AAV production increased transgene expression by 40–100 fold in vivo
The removal of a Woodchuck posttranscriptional regulatory element (WPRE) sequence reduced the expression by 80%, whereas the use of a synthetic polyadenylation sequence (49 bp) in combination with two SV40 late upstream elements (50 bp) increased the GFP expression compared to the use of a robust bGHpolyA sequence
Summary
Many reviews have been written on recombinant adeno-associated virus vector (rAAV) tropism in ocular tissue, rAAV host cell infection, and potential rAAV-treatable inherited retinal diseases [1,2,3,4,5,6,7,8,9]. We will review the ocular gene therapies developed over the past 20 years focusing on the diversity of elements incorporated in rAAV vectors. We will provide a library of validated elements that allows other researchers to streamline the modification of their vectors. Novel medical therapies, such as gene therapies, need to be carefully optimized to demonstrate their efficacy and safety before going into clinical human trials [10]. We will explore the developments in preclinical models, for example, cellular models, human induced pluripotent stem cell (hiPSC) derived retinal organoid disease models, retinal explant models, and tropism studies in non-human primates
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
