June 2022
Volume 63, Issue 7
Open Access
ARVO Annual Meeting Abstract  |   June 2022
Delivery of genome editors to the mouse eye using engineered virus-like particles
Author Affiliations & Notes
  • Samuel Wang Du
    Ophthalmology, University of California Irvine School of Medicine, Irvine, California, United States
    Physiology & Biophysics, University of California Irvine School of Medicine, Irvine, California, United States
  • Samagya Banskota
    Broad Institute, Cambridge, Massachusetts, United States
    Chemistry & Chemical Biology, Harvard University, Cambridge, Massachusetts, United States
  • Aditya Raguram
    Broad Institute, Cambridge, Massachusetts, United States
    Chemistry & Chemical Biology, Harvard University, Cambridge, Massachusetts, United States
  • Susie Suh
    Ophthalmology, University of California Irvine School of Medicine, Irvine, California, United States
    Pharmacology, Case Western Reserve University, Cleveland, Ohio, United States
  • Elliot H Choi
    Ophthalmology, University of California Irvine School of Medicine, Irvine, California, United States
    Pharmacology, Case Western Reserve University, Cleveland, Ohio, United States
  • Grazyna Palczewska
    Ophthalmology, University of California Irvine School of Medicine, Irvine, California, United States
  • David R Liu
    Broad Institute, Cambridge, Massachusetts, United States
    Pharmacology, Case Western Reserve University, Cleveland, Ohio, United States
  • Krzysztof Palczewski
    Ophthalmology, University of California Irvine School of Medicine, Irvine, California, United States
    Physiology & Biophysics, University of California Irvine School of Medicine, Irvine, California, United States
  • Footnotes
    Commercial Relationships   Samuel Du None; Samagya Banskota Broad Institute, Code P (Patent); Aditya Raguram Broad Institute, Code P (Patent); Susie Suh None; Elliot Choi None; Grazyna Palczewska Polgenix, Inc, Code E (Employment); David Liu Prime Medicine, Beam Therapeutics, Pairwise Plants, Chroma Medicine, Code C (Consultant/Contractor), Prime Medicine, Beam Therapeutics, Pairwise Plants, Chroma Medicine, Code O (Owner), Broad Institute, Code P (Patent); Krzysztof Palczewski Polgenix, Inc, Code C (Consultant/Contractor)
  • Footnotes
    Support  NIH UG3AI150551, NIH U01AI142756, NIH R35GM118062, NIH RM1HG009490, NIH R01EY009339, NIH T32GM095450, NIH F30EY029136, NIH T32GM007250, NIH T32GM007250, NIH T32GM008620, the Bill and Melinda Gates Foundation, an NSF graduate fellowship, and HHMI
Investigative Ophthalmology & Visual Science June 2022, Vol.63, 2823 – A0339. doi:
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    • Get Citation

      Samuel Wang Du, Samagya Banskota, Aditya Raguram, Susie Suh, Elliot H Choi, Grazyna Palczewska, David R Liu, Krzysztof Palczewski; Delivery of genome editors to the mouse eye using engineered virus-like particles. Invest. Ophthalmol. Vis. Sci. 2022;63(7):2823 – A0339.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Precision genome editing could advance greatly the treatment of inherited retinal diseases (IRDs), as shown by the reversion of IRD-causing mutations by CRISPR/Cas9 and restoration of visual function in animal models. However, before broader human application, it is imperative to improve the safety profile of vectors for genome editor delivery to minimize deleterious off-target and bystander effects, which can occur when they are expressed over prolonged periods of time. As genomic DNA edits are permanent, a one-time treatment with transiently delivered genome editors should be sufficient to correct an IRD mutation. One such promising method is the administration of genome editors as preformed proteins and ribonucleoproteins encapsulated within virus-like particles (VLPs).

Methods : All animal procedures were approved by the IACUC at UC Irvine and conformed to the recommendations of the American Veterinary Medical Association Panel on Euthanasia and ARVO.
We developed engineered VLPs (eVLPs), an improved VLP platform for packaging and delivering genome editor proteins and RNPs. eVLP possess many of the same advantages as viral vectors, including cell-type tropism and large cargo capacity, but offer significant safety advantages over lentiviral and AAV vectors, such as decreased off-target and bystander editing, and no risk of genomic integration. We exploited the property of Moloney murine leukemia virus which accommodates C-terminal fusion of proteins to its gag structural protein to ensure proper and controlled packaging of genome editors within the eVLPs.

Results : We produced eVLP constructs carrying Cre recombinase or adenine base editors (ABEs). eVLPs efficiently delivered their cargoes to cell lines in vitro. When injected subretinally, eVLPs effected genomic changes in the RPE, such as loxP recombination and A to G nucleotide changes, as assessed by imaging and ERG. Importantly, these eVLPs correct mutations with similar on-target and substantially lower bystander and off-target editing compared to virally delivered ABEs.

Conclusions : By iteratively modifying the surface glycoprotein of eVLPs, we will screen for eVLPs which facilitate the delivery of genome editors to photoreceptors, which historically have been difficult to target with enveloped viruses. We hope the adaptation of this technology will enable the broader application of genome editors to the mouse and human eye.

This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.

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