June 2023
Volume 64, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2023
A base editing strategy for Best vitelliform macular dystrophy utilizing retinal pigment epithelium from patient-derived iPSCs
Sushma Kalmodia; Jennifer Aparicio; Assaf Beck; Andrew Salas; Narine Harutyunyan; Ryan Schmidt; David Cobrinik; Aaron Nagiel
Author Affiliations & Notes
  • Sushma Kalmodia
    The Vision Center and Saban Research Institute, Children's Hospital Los Angeles, Children's Hospital Los Angeles, Los Angeles, California, United States
  • Jennifer Aparicio
    The Vision Center and Saban Research Institute, Children's Hospital Los Angeles, Children's Hospital Los Angeles, Los Angeles, California, United States
  • Assaf Beck
    The Vision Center and Saban Research Institute, Children's Hospital Los Angeles, Children's Hospital Los Angeles, Los Angeles, California, United States
  • Andrew Salas
    The Vision Center and Saban Research Institute, Children's Hospital Los Angeles, Children's Hospital Los Angeles, Los Angeles, California, United States
  • Narine Harutyunyan
    The Vision Center and Saban Research Institute, Children's Hospital Los Angeles, Children's Hospital Los Angeles, Los Angeles, California, United States
  • Ryan Schmidt
    Department of Pathology and Laboratory Medicine, Children’s Hospital Los Angeles, and Keck School of Medicine, University of Southern California, University of Southern California Keck School of Medicine, Los Angeles, California, United States
  • David Cobrinik
    The Vision Center and Saban Research Institute, Children's Hospital Los Angeles, Children's Hospital Los Angeles, Los Angeles, California, United States
    USC Roski Eye Institute, University of Southern California Keck School of Medicine, Los Angeles, California, United States
  • Aaron Nagiel
    The Vision Center and Saban Research Institute, Children's Hospital Los Angeles, Children's Hospital Los Angeles, Los Angeles, California, United States
    USC Roski Eye Institute, University of Southern California Keck School of Medicine, Los Angeles, California, United States
  • Footnotes
    Commercial Relationships   Sushma Kalmodia None; Jennifer Aparicio None; Assaf Beck None; Andrew Salas None; Narine Harutyunyan None; Ryan Schmidt None; David Cobrinik None; Aaron Nagiel Lexitas, Eyebiotech, Biogen, Novartis, Atsena, Janssen., Code C (Consultant/Contractor)
  • Footnotes
    Support  California Institute for Regenerative Medicine- Project No 000015187
Investigative Ophthalmology & Visual Science June 2023, Vol.64, 2731. doi:
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      Sushma Kalmodia, Jennifer Aparicio, Assaf Beck, Andrew Salas, Narine Harutyunyan, Ryan Schmidt, David Cobrinik, Aaron Nagiel; A base editing strategy for Best vitelliform macular dystrophy utilizing retinal pigment epithelium from patient-derived iPSCs. Invest. Ophthalmol. Vis. Sci. 2023;64(8):2731.

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Abstract

Purpose : Best vitelliform macular dystrophy (BVMD) is an autosomal dominant (AD) inherited retinal dystrophy (IRD) caused by variants in BEST1 and leading to defects in retinal pigment epithelium (RPE) cells. In this study, we created patient-derived induced pluripotent stem cells (iPSCs) and differentiated them into RPE in order to characterize the pathogenic phenotype and test a base editing strategy to correct or abolish the expression of the pathogenic BEST1 allele.

Methods : Patients diagnosed with BVMD at Children’s Hospital Los Angeles were identified with the two pathogenic variants 1) BEST1 c.851A>G (Tyr284Cys) and 2) BEST1 c.240C>A (Phe80Leu). IPSCs were generated from blood and differentiated to RPE cells. RPE monolayers were characterized for marker expression, transepithelial resistance (TER), and ATP -stimulated fluid transport. Plasmids expressing cytosine base editor (CBE) and gRNA were delivered by Neon transfection to the patient iPSCs (c.851A>G) to correct the expression of the pathogenic allele

Results : BVMD patient-derived iPSC were differentiated into RPE cells and showed typical RPE morphology with polygonal-shaped epithelial monolayers and pigmented granules. These cells expressed RPE markers such as ZO1 (tight junctions), bestrophin and MCT3 (basolateral polarity), and Na, K-ATPase, (apical polarity). TER was higher in the BEST1 c.240C>A variant and lower for the BEST1 c.851A>G variant compared to the control. ATP-stimulated apical fluid transport was impaired in the BEST1 c.240C>A compared to the control. Western blotting showed two different molecular weights for these BEST1 variants. Finally, CBE and gRNA transfection resulted in 47% correction efficiency in BEST1 c.851A>G iPSCs lines.

Conclusions : This study demonstrates a strategy for obtaining BEST1 patient-derived iPSCs, differentiating them into RPE cells, characterizing the RPE phenotype, and assessing base editing gene correction strategies.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.

This work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.
Attribution-NonCommercial-NoDerivatives
Copyright © 2024 Association for Research in Vision and Ophthalmology.
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