Human MiniPromoters for ocular-rAAV expression in ON bipolar, cone, corneal, endothelial, Müller glial, and PAX6 cells

• Published in: Gene therapy Vol. 28; no. 6; pp. 351 - 372
• Main Authors: Korecki, Andrea J., Cueva-Vargas, Jorge L., Fornes, Oriol, Agostinone, Jessica, Farkas, Rachelle A., Hickmott, Jack W., Lam, Siu Ling, Mathelier, Anthony, Zhou, Michelle, Wasserman, Wyeth W., Di Polo, Adriana, Simpson, Elizabeth M.
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 01-06-2021; Nature Publishing Group; Stockton Press
• Subjects: 14/35; 14/63; 42/44; 631/1647/2017; 631/61/201; 64/60; 692/699/375; Animals; Biomedical and Life Sciences; Biomedicine; Cell Biology; Clinical trials; Cornea; Deoxyribonucleic acid; Dependoviruses; DNA; Endothelial Cells; Gene Expression; Gene Therapy; Genetic aspects; Genetic engineering; Genetic research; Human Genetics; Humans; Intravenous administration; Methods; Mice; Mueller cells; Nanotechnology; Neonates; Neuroglia; Pax6 protein; PAX6 Transcription Factor - genetics; Promoter Regions, Genetic; Promoters; Promoters (Genetics); Retina; Retinal Cone Photoreceptor Cells; Stroma; Canada
• ISSN: 0969-7128; 1476-5462
• DOI: 10.1038/s41434-021-00227-z

Small and cell-type restricted promoters are important tools for basic and preclinical research, and clinical delivery of gene therapies. In clinical gene therapy, ophthalmic trials have been leading the field, with over 50% of ocular clinical trials using promoters that restrict expression based on cell type. Here, 19 human DNA MiniPromoters were bioinformatically designed for rAAV, tested by neonatal intravenous delivery in mouse, and successful MiniPromoters went on to be tested by intravitreal, subretinal, intrastromal, and/or intravenous delivery in adult mouse. We present promoter development as an overview for each cell type, but only show results in detail for the recommended MiniPromoters: Ple265 and Ple341 (
PCP2
) ON bipolar, Ple349 (
PDE6H
) cone, Ple253 (
PITX3
) corneal stroma, Ple32 (
CLDN5
) endothelial cells of the blood–retina barrier, Ple316 (
NR2E1
) Müller glia, and Ple331 (
PAX6
) PAX6 positive. Overall, we present a resource of new, redesigned, and improved MiniPromoters for ocular gene therapy that range in size from 784 to 2484 bp, and from weaker, equal, or stronger in strength relative to the ubiquitous control promoter smCBA. All MiniPromoters will be useful for therapies involving small regulatory RNA and DNA, and proteins ranging from 517 to 1084 amino acids, representing 62.9–90.2% of human proteins.