High-efficiency Transduction of the Mouse Retina by Tyrosine-mutant AAV Serotype Vectors

Petrs‐Silva, Hilda; Dinculescu, Astra; Li, Qiuhong; Min, Seok-Hong; Chiodo, Vince A.; Pang, Jijing; Zhong, Li; Zolotukhin, Sergei; Srivastava, Arun; Lewin, Alfred S.; Hauswirth, William W.

doi:10.1038/mt.2008.269

Cited by 354 publications

(360 citation statements)

References 47 publications

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“…In response to axonal injury, 50%-75% of RGCs die by 2 wks (31). Mice carrying a floxed allele of Dlk (Dlk fl/fl ) were injected intravitreally with a self-complementary, capsid-modified adeno-associated virus 2 (AAV2) (32,33) expressing Cre. Injection of the AAV2-Cre resulted in Cre expression in nearly 100% of RGCs ( Fig.…”

Section: Dlk Inhibition Promotes Rgc Survival In Vivo After Optic Nervementioning

confidence: 99%

Functional genomic screening identifies dual leucine zipper kinase as a key mediator of retinal ganglion cell death

Welsbie

Yang

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

237

307

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Glaucoma, a major cause of blindness worldwide, is a neurodegenerative optic neuropathy in which vision loss is caused by loss of retinal ganglion cells (RGCs). To better define the pathways mediating RGC death and identify targets for the development of neuroprotective drugs, we developed a high-throughput RNA interference screen with primary RGCs and used it to screen the full mouse kinome. The screen identified dual leucine zipper kinase (DLK) as a key neuroprotective target in RGCs. In cultured RGCs, DLK signaling is both necessary and sufficient for cell death. DLK undergoes robust posttranscriptional up-regulation in response to axonal injury in vitro and in vivo. Using a conditional knockout approach, we confirmed that DLK is required for RGC JNK activation and cell death in a rodent model of optic neuropathy. In addition, tozasertib, a small molecule protein kinase inhibitor with activity against DLK, protects RGCs from cell death in rodent glaucoma and traumatic optic neuropathy models. Together, our results establish a previously undescribed drug/drug target combination in glaucoma, identify an early marker of RGC injury, and provide a starting point for the development of more specific neuroprotective DLK inhibitors for the treatment of glaucoma, nonglaucomatous forms of optic neuropathy, and perhaps other CNS neurodegenerations.G laucoma is the leading cause of irreversible blindness worldwide (1). It is a neurodegenerative disease in which vision loss is caused by the axonal injury and death of retinal ganglion cells (RGCs) (2), the projection neurons that process and transmit vision from the retina to the brain. Current therapies (i.e., surgery, laser, and eye drops) all act by lowering intraocular pressure (IOP). However, pressure reduction can be difficult to achieve, and even with significant pressure lowering, RGC loss can continue. Efforts have therefore been made to develop neuroprotective agents that would complement IOP-lowering therapies by directly inhibiting the RGC cell death process (3, 4). However, no neuroprotective agent has yet been approved for clinical use.Protein kinases provide attractive targets for the development of neuroprotective agents. A number of kinases, including cyclindependent kinases, death-associated protein kinases, JNK1-3, MAPKs, and glycogen synthase kinase-3β, are involved in neuronal cell death (5-12). An additional attraction is that protein kinases are readily druggable. The pharmacology and medicinal chemistry of kinase inhibitors are well-developed, with kinases now being the most important class of drug targets after G protein-coupled receptors (13). Although the primary clinical use of kinase inhibitors continues to be as antineoplastic agents, increasing attention is being paid to their use in other areas (14,15).To identify, in a comprehensive and unbiased manner, kinases that could serve as targets for neuroprotective glaucoma therapy, we screened the entire mouse kinome for kinases whose inhibition promotes RGC survival. For this screen, we develope...

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Section: Dlk Inhibition Promotes Rgc Survival In Vivo After Optic Nervementioning

confidence: 99%

Functional genomic screening identifies dual leucine zipper kinase as a key mediator of retinal ganglion cell death

Welsbie

Yang

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

237

307

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“…Consistent with this hypothesis, they observed that site-directed mutagenesis of surface-exposed tyrosine residues significantly improve the transduction efficiency of AAV vectors (about 10-fold increase) in human epithelial cells in vitro. It has also been demonstrated that tyrosine mutant AAV vectors display enhanced gene delivery efficiencies in vivo when compared to wild-type AAV vectors [40] . Although further investigation is warranted, such enhanced AAV vectors engineered through rational design to avoid cytoplasmic degradation could also potentially enhance the efficiencies of AAV-mediated gene targeting in human stem cells.…”

Section: Molecular Engineering Of Viral Vectorsmentioning

confidence: 99%

Advances in homology directed genetic engineering of human pluripotent and adult stem cells

Ramamoorthi

Curtis²,

Asuri³

2013

WJSC

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The ability to introduce precise genomic modifications in human cells has profound implications for both basic and applied research in stem cells, ranging from identification of genes regulating stem cell self-renewal and multilineage differentiation to therapeutic gene correction and creation of in vitro models of human diseases. However, the overall efficiency of this process is challenged by several factors including inefficient gene delivery into stem cells and low rates of homology directed site-specific targeting. Recent studies report the development of novel techniques to improve gene targeting efficiencies in human stem cells; these methods include molecular engineering of viral vectors to efficiently deliver episomal genetic sequences that can participate in homology directed targeting, as well as the design of synthetic proteins that can introduce double-stranded breaks in DNA to initiate such recombination events. This review focuses on the potential of these new technologies to precisely alter the human stem cell genome and also highlights the possibilities offered by the combination of these complementary strategies.© 2013 Baishideng. All rights reserved. Key words: Human stem cells; Genetic engineering; Engineered viruses; Synthetic restriction endonucleasesCore tip: This manuscript focuses on the development of novel technologies to precisely alter the human stem cell genome and highlights their implications for both basic and applied stem cell research. Specifically, we discuss the development of two main technologies: molecular engineering of viral vectors and design of artificial endonucleases. We also discuss the merits of combining these complementary approaches and suggest other possible strategies that could be explored to further improve genetic engineering of human stem cells.

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“…A further development of AAV as a vector for photoreceptor cell gene targeting has involved mutagenesis of the viral capsid proteins to increase the efficiency of transduction and penetration of the vector across the layers of the neurosensory retina. Mutagenesis of specific capsid tyrosine residues can enable efficient transduction of photoreceptors following intravitreal injection, 28,29 a technique that is safer and technically less challenging than subretinal injection. However, administration of AAV vectors into the vitreous can stimulate more powerful immune responses than are typically evident following administration into the subretinal space, which has the benefit of relative immune privilege.…”

Section: Gene Transfer To Photoreceptor Cellsmentioning

confidence: 99%

Gene supplementation therapy for recessive forms of inherited retinal dystrophies

Smith¹,

Bainbridge²,

Ali³

2011

Gene Ther

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Over the last decade, gene supplementation therapy for inherited retinal degeneration has come of age. Early proof-of-concept studies in animal models of disease showed modest, but genuine improvements in retinal function and/or survival. Further development of the vectors used for gene transfer to the retina has led to better treatment efficacy in a wide variety of animal models, leading in 2008 to the initiation of three clinical trials for Leber congenital amaurosis caused by retinal pigment epithelium 65 deficiency. The results from these trials suggest that the treatment of inherited retinal dystrophy by gene therapy can be safe and effective. Here, we examine the progress of gene supplementation therapy in the retina, and discuss the potential for using gene therapy to treat different forms of inherited retinal degeneration.

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High-efficiency Transduction of the Mouse Retina by Tyrosine-mutant AAV Serotype Vectors

Cited by 354 publications

References 47 publications

Functional genomic screening identifies dual leucine zipper kinase as a key mediator of retinal ganglion cell death

Functional genomic screening identifies dual leucine zipper kinase as a key mediator of retinal ganglion cell death

Advances in homology directed genetic engineering of human pluripotent and adult stem cells

Gene supplementation therapy for recessive forms of inherited retinal dystrophies

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