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Effects of Stuffer DNA on the Suppression of Choroidal Neovascularization by a rAAV Expressing a mTOR-Inhibiting shRNA
- Source :
- Molecular Therapy. Methods & Clinical Development
- Publication Year :
- 2019
-
Abstract
- Choroidal neovascularization (CNV) is the defining characteristic of the wet subtype of age-related macular degeneration (AMD), which is a rapidly growing global health problem. Previously, we had demonstrated the therapeutic potential of gene therapy against CNV using short hairpin RNA (shRNA) delivered via recombinant adeno-associated virus (rAAV), which abrogates mammalian-to-mechanistic (mTOR) activity in a novel manner by simultaneously inhibiting both mTOR complexes. Both the target and use of gene therapy represent a novel treatment modality against AMD. Here, the xenogeneic GFP gene used as a reporter in previous studies was removed from the virus vector to further develop the therapeutic for clinical trials. Instead, a stuffer DNA derived from the 3′ UTR of the human UBE3A gene was used to ensure optimal viral genome size for efficient rAAV assembly. The virus vector containing the stuffer DNA, rAAV2-shmTOR-SD, positively compares to one encoding the shRNA and a GFP expression cassette in terms of reducing CNV in a laser-induced mouse model, as determined by fundus fluorescein angiography. These results were confirmed via immunohistochemistry using anti-CD31, while a TUNEL assay showed that rAAV2-shmTOR-SD possesses anti-apoptotic properties as well. The qualities exhibited by rAAV2-shmTOR-SD demonstrate its potential as a human gene therapeutic for the treatment of wet AMD.
- Subjects :
- 0301 basic medicine
genetic structures
Genetic enhancement
Biology
GFP
choroidal neovascularization
Article
law.invention
Viral vector
Small hairpin RNA
03 medical and health sciences
0302 clinical medicine
law
RNA interference
Genetics
medicine
Molecular Biology
PI3K/AKT/mTOR pathway
mammalian target of rapamycin
recombinant adeno-associated virus
stuffer DNA
wet age-related macular degeneration
eye diseases
030104 developmental biology
Choroidal neovascularization
030220 oncology & carcinogenesis
RNAi
Recombinant DNA
Cancer research
Molecular Medicine
Expression cassette
sense organs
medicine.symptom
Subjects
Details
- ISSN :
- 23290501
- Volume :
- 14
- Database :
- OpenAIRE
- Journal :
- Molecular therapy. Methodsclinical development
- Accession number :
- edsair.doi.dedup.....59e0a2e8b3661138fe64841acbb81a5f