D - Synthetic/Molecular Conjugates and Physical Methods for Delivery of Gene Therapeutics -> Synthetic/Molecular Conjugates and Physical Methods for Delivery of Gene Therapeutics (including exosomes)
29: Shuttle Peptides-Mediated Local RNP Delivery of Editing Complex in the Sensory Organs of the Inner Ear and Retina In Vivo in Adult Mice
Type: Oral Abstract Session
Presentation Details
Session Title: Synthetic/Molecular Conjugates and Physical Methods for Delivery of Gene Therapeutics I
Location: Petree Hall C
Start Time: 5/17/2023 16:45
End Time: 5/17/2023 17:00
Genetic hearing loss and blindness are prevalent with limited treatment options. Despite the tremendous progress in the development of editing therapy for hearing loss and blindness, the ability to deliver the editing complex into the inner ear and retina remains the major challenge to realizing the potential. For viral vectors, issues like immunogenicity, genome integration and long-term effect remain a safety concern. In addition, with the more complex editing strategies including base editing and prime editing, multiple components need to be packaged into multiple AAVs, further limiting their efficiency. We have developed an amphiphilic S10 shuttle peptide and demonstrated delivery efficiency of CRISPR-associated nuclease ribonucleoprotein (RNP) to the respiratory tract of mice. In this study, we screened additional peptides of different charges and lengths to evaluate the application of shuttle peptides to deliver editing RNP complex into the sensory organs of the inner ear and retina in adult mice by local injection in vivo. The shuttle peptide-RNP formulation was delivered locally into the inner ear through the round window membrane (RWM) and retina by subretinal injection in adult Ai14 mice with tdTomato (tdT) reporter. We identified shuttle peptides capable of RNP delivery into multiple inner ear sensory epithelial cell types and retinal cells with efficient editing. In the inner ear, the shuttle peptides mediated efficient delivery and editing in the supporting cell subtypes throughout the entire cochlear turn. Damage to outer hair cells was detected as a result of the delivery. In the retina, the shuttle peptides mediated efficient delivery and editing in the retinal pigment epithelial (RPE) cells and Muller cells, including those at a distance from the injection site. Muller cells at the injection site were also edited after injection. Using a shuttle peptide mixed with fluorescent cargo Cy5-NLS for the delivery into the inner ear, we detected Cy5 labeling in the majority of inner ear sensory epithelial cell types in adult mice, suggesting further modifications may target more inner ear cell types. The identification of shuttle peptides capable of efficient in vivo RNP delivery of editing agents in the mature mammalian inner ear and retina opens a new avenue to develop editing therapy for human hearing loss and blindness.
Wan Du1, Xue Cheng2, Qin Liu3, Wei Wei1, Rossano Butcher3, David Guay2, Zheng-Yi Chen1
1Department of Otolaryngology-Head and Neck Surgery, Harvard Medical School/Massachusetts Eye and Ear, Boston, MA,2Feldan Therapeutics, Quebec, QC, Canada,3Department of Ophthalmology, Harvard Medical School/Massachusetts Eye and Ear, Boston, MA
W. Du: None.
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