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A drug-tunable Flt23k gene therapy for controlled intervention in retinal neovascularization

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Chen, J, Lin, F-L, Leung, JYK ORCID: 0000-0001-8798-3932, Tu, L, Wang, J-H, Chuang, Y-F, Li, F, Shen, H-H, Dusting, GJ, Wong, VHY, Lisowski, L, Hewitt, AW ORCID: 0000-0002-5123-5999, Bui, BV, Zhong, J and Liu, G-S ORCID: 0000-0003-3379-724X 2020 , 'A drug-tunable Flt23k gene therapy for controlled intervention in retinal neovascularization' , Angiogenesis, no. Septem , pp. 1-14 , doi: 10.1007/s10456-020-09745-7.

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Abstract

Gene therapies that chronically suppress vascular endothelial growth factor (VEGF) represent a new approach for managing retinal vascular leakage and neovascularization. However, constitutive suppression of VEGF in the eye may have deleterious side effects. Here, we developed a novel strategy to introduce Flt23k, a decoy receptor that binds intracellular VEGF, fused to the destabilizing domain (DD) of Escherichia coli dihydrofolate reductase (DHFR) into the retina. The expressed DHFR(DD)-Flt23k fusion protein is degraded unless "switched on" by administering a stabilizer; in this case, the antibiotic trimethoprim (TMP). Cells transfected with the DHFR(DD)-Flt23k construct expressed the fusion protein at levels correlated with the TMP dose. Stabilization of the DHFR(DD)-Flt23k fusion protein by TMP was able to inhibit intracellular VEGF in hypoxic cells. Intravitreal injection of self-complementary adeno-associated viral vector (scAAV)-DHFR(DD)-Flt23k and subsequent administration of TMP resulted in tunable suppression of ischemia-induced retinal neovascularization in a rat model of oxygen-induced retinopathy (OIR). Hence, our study suggests a promising novel approach for the treatment of retinal neovascularization. Schematic diagram of the tunable system utilizing the DHFR(DD)-Flt23k approach to reduce VEGF secretion. a The schematic shows normal VEGF secretion. b Without the ligand TMP, the DHFR(DD)-Flt23k protein is destabilized and degraded by the proteasome. c In the presence of the ligand TMP, DHFR(DD)-Flt23k is stabilized and sequestered in the ER, thereby conditionally inhibiting VEGF. Green lines indicate the intracellular and extracellular distributions of VEGF. Blue lines indicate proteasomal degradation of the DHFR(DD)-Flt23k protein. Orange lines indicate the uptake of cell-permeable TMP. TMP, trimethoprim; VEGF, vascular endothelial growth factor; ER, endoplasmic reticulum.

Item Type: Article
Authors/Creators:Chen, J and Lin, F-L and Leung, JYK and Tu, L and Wang, J-H and Chuang, Y-F and Li, F and Shen, H-H and Dusting, GJ and Wong, VHY and Lisowski, L and Hewitt, AW and Bui, BV and Zhong, J and Liu, G-S
Keywords: gene therapy, diabetic retinopathy, angiogenesis, VEGF, AAV, destabilizing domain, Flt23k, retinal neovascularization, trimethoprim
Journal or Publication Title: Angiogenesis
Publisher: Springer Netherlands
ISSN: 0969-6970
DOI / ID Number: 10.1007/s10456-020-09745-7
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Copyright 2020 Springer Nature B.V.

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