Abstract
LbCpf1, derived from Lachnospiraceae bacterium ND2006, is a CRISPR RNA-guided endonuclease and holds promise for therapeutic applications. Here we show that LbCpf1 can be used for therapeutic gene editing in a mouse model of age-related macular degeneration (AMD). The intravitreal delivery of LbCpf1, targeted to two angiogenesis-associated genes encoding vascular endothelial growth factor A (Vegfa) and hypoxia inducing factor 1a (Hif1a), using adeno-associated virus, led to efficient gene disruption with no apparent off-target effects in the retina and retinal pigment epithelium (RPE) cells. Importantly, LbCpf1 targeted to Vegfa or Hif1a in RPE cells reduced the area of laser-induced choroidal neovascularization as efficiently as aflibercept, an anti-VEGF drug currently used in the clinic, without inducing cone dysfunction. Unlike aflibercept, LbCpf1 targeted to Vegfa or Hif1a achieved a long-term therapeutic effect on CNV, potentially avoiding repetitive injections. Taken together, these results indicate that LbCpf1-mediated in vivo genome editing to ablate pathologic angiogenesis provides an effective strategy for the treatment of AMD and other neovascularization-associated diseases.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Bacterial Proteins / administration & dosage
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Bacterial Proteins / genetics
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Cell Line
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Choroidal Neovascularization / etiology
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Choroidal Neovascularization / genetics
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Choroidal Neovascularization / pathology
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Choroidal Neovascularization / therapy*
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Clostridiales / genetics
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Clustered Regularly Interspaced Short Palindromic Repeats / genetics
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Dependovirus / genetics
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Disease Models, Animal
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Endonucleases / administration & dosage
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Endonucleases / genetics
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Gene Editing / methods*
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Genetic Therapy / methods*
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Genetic Vectors / administration & dosage*
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Genetic Vectors / genetics
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Humans
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Hypoxia-Inducible Factor 1, alpha Subunit / genetics
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Intravitreal Injections
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Lasers / adverse effects
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Macular Degeneration / etiology
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Macular Degeneration / genetics
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Macular Degeneration / pathology
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Macular Degeneration / therapy*
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Male
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Mice
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Mice, Inbred C57BL
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RNA, Guide, CRISPR-Cas Systems / genetics
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Retinal Pigment Epithelium / blood supply
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Retinal Pigment Epithelium / pathology
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Retinal Pigment Epithelium / radiation effects
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Specific Pathogen-Free Organisms
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Treatment Outcome
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Vascular Endothelial Growth Factor A / genetics
Substances
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Bacterial Proteins
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Hif1a protein, mouse
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Hypoxia-Inducible Factor 1, alpha Subunit
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RNA, Guide, CRISPR-Cas Systems
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Vascular Endothelial Growth Factor A
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vascular endothelial growth factor A, mouse
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Endonucleases