rAAV-compatible MiniPromoters for restricted expression in the brain and eye.

Journal Article (Journal Article)

BACKGROUND: Small promoters that recapitulate endogenous gene expression patterns are important for basic, preclinical, and now clinical research. Recently, there has been a promising revival of gene therapy for diseases with unmet therapeutic needs. To date, most gene therapies have used viral-based ubiquitous promoters-however, promoters that restrict expression to target cells will minimize off-target side effects, broaden the palette of deliverable therapeutics, and thereby improve safety and efficacy. Here, we take steps towards filling the need for such promoters by developing a high-throughput pipeline that goes from genome-based bioinformatic design to rapid testing in vivo. METHODS: For much of this work, therapeutically interesting Pleiades MiniPromoters (MiniPs; ~4 kb human DNA regulatory elements), previously tested in knock-in mice, were "cut down" to ~2.5 kb and tested in recombinant adeno-associated virus (rAAV), the virus of choice for gene therapy of the central nervous system. To evaluate our methods, we generated 29 experimental rAAV2/9 viruses carrying 19 different MiniPs, which were injected intravenously into neonatal mice to allow broad unbiased distribution, and characterized in neural tissues by X-gal immunohistochemistry for icre, or immunofluorescent detection of GFP. RESULTS: The data showed that 16 of the 19 (84 %) MiniPs recapitulated the expression pattern of their design source. This included expression of: Ple67 in brain raphe nuclei; Ple155 in Purkinje cells of the cerebellum, and retinal bipolar ON cells; Ple261 in endothelial cells of brain blood vessels; and Ple264 in retinal Müller glia. CONCLUSIONS: Overall, the methodology and MiniPs presented here represent important advances for basic and preclinical research, and may enable a paradigm shift in gene therapy.

Full Text

Duke Authors

Cited Authors

  • de Leeuw, CN; Korecki, AJ; Berry, GE; Hickmott, JW; Lam, SL; Lengyell, TC; Bonaguro, RJ; Borretta, LJ; Chopra, V; Chou, AY; D'Souza, CA; Kaspieva, O; Laprise, S; McInerny, SC; Portales-Casamar, E; Swanson-Newman, MI; Wong, K; Yang, GS; Zhou, M; Jones, SJM; Holt, RA; Asokan, A; Goldowitz, D; Wasserman, WW; Simpson, EM

Published Date

  • May 10, 2016

Published In

Volume / Issue

  • 9 / 1

Start / End Page

  • 52 -

PubMed ID

  • 27164903

Pubmed Central ID

  • PMC4862195

Electronic International Standard Serial Number (EISSN)

  • 1756-6606

Digital Object Identifier (DOI)

  • 10.1186/s13041-016-0232-4


  • eng

Conference Location

  • England