Ocular Localization and Transduction by Adenoviral Vectors Are Serotype-Dependent and Can Be Modified by Inclusion of RGD Fiber Modifications

Ueyama, Kazuhiro; Mori, Keisuke; Shoji, T.; Omata, H.; Gehlbach, Peter; Brough, Douglas E.; Wei, Lisa L.; Yoneya, Shin

doi:10.1371/journal.pone.0108071

Cited by 20 publications

(21 citation statements)

References 32 publications

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“…By quantifying the number of corrected TM cells in vivo, we found an estimated transduction efficiency of at least ∼60-70% using Ad5-cas9 and/or Ad5-crMYOC. Of note, we also found Cas9 expression in the cornea endothelium and parts of the iris and ciliary body, similar to previous studies with adenovirus transduction (36)(37)(38). Western blot analysis data also showed reductions in myocilin, BiP, Calnexin, PDI, ATF4, and CHOP levels, confirming our ICC data indicating gRNA targeting in Tg-MYOC Y437H mice (Fig.…”

Section: Crispr-cas9 Lowers Iop and Prevents Further Glaucomatous Damagesupporting

confidence: 91%

CRISPR-Cas9–based treatment of myocilin-associated glaucoma

Jain

Zode

Kasetti

et al. 2017

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

155

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Primary open-angle glaucoma (POAG) is a leading cause of irreversible vision loss worldwide, with elevated intraocular pressure (IOP) a major risk factor. Myocilin (MYOC) dominant gain-offunction mutations have been reported in ∼4% of POAG cases. MYOC mutations result in protein misfolding, leading to endoplasmic reticulum (ER) stress in the trabecular meshwork (TM), the tissue that regulates IOP. We use CRISPR-Cas9-mediated genome editing in cultured human TM cells and in a MYOC mouse model of POAG to knock down expression of mutant MYOC, resulting in relief of ER stress. In vivo genome editing results in lower IOP and prevents further glaucomatous damage. Importantly, using an ex vivo human organ culture system, we demonstrate the feasibility of human genome editing in the eye for this important disease.myocilin | CRISPR | glaucoma | trabecular meshwork | genome editing

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Section: Crispr-cas9 Lowers Iop and Prevents Further Glaucomatous Damagesupporting

confidence: 91%

CRISPR-Cas9–based treatment of myocilin-associated glaucoma

Jain

Zode

Kasetti

et al. 2017

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

155

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“…AdV requires handling at biosafety level (BSL) 2. The nonspecific transduction can be made more specific by transcriptional targeting (Gonzalez et al, 2004) or choosing select sub-serotypes (Ueyama et al, 2014). …”

Section: Established Methods Of Ocular Gene Therapymentioning

confidence: 99%

“…A TM preferential promoter is the matrix Gla protein promoter (Gonzalez et al, 2004) or chitinase 3-like 1 promoter (Liton et al, 2005). Ueyama et al found that an unmodified Ad35 serotype preferentially transduces TM cells (Ueyama et al, 2014). …”

Section: Current Challenges and Future Directionsmentioning

confidence: 99%

Gene transfer to the outflow tract

Dang

Loewen

Parikh

et al. 2017

Experimental Eye Research

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Elevated intraocular pressure is the primary cause of open angle glaucoma. Outflow resistance exists within the trabecular meshwork but also at the level of Schlemm's canal and further downstream within the outflow system. Viral vectors allow to take advantage of naturally evolved, highly efficient mechanisms of gene transfer, a process that is termed transduction. They can be produced at biosafety level 2 in the lab using protocols that have evolved considerably over the last 15 to 20 years. Applied by an intracameral bolus, vectors follow conventional as well as uveoscleral outflow pathways. They may affect other structures in the anterior chamber depending on their transduction kinetics which can vary among species when using the same vector. Not all vectors can express long-term, a desirable feature to address the chronicity of glaucoma. Vectors that integrate into the genome of the target cell can achieve transgene function for the life of the transduced cell but are mutagenic by definition. The most prominent long-term expressing vector systems are based on lentiviruses that are derived from HIV, FIV, or EIAV. Safety considerations make non-primate lentiviral vector systems easier to work with as they are not derived from human pathogens. Non-integrating vectors are subject to degradation and attritional dilution during cell division. Lentiviral vectors have to integrate in order to express while adeno-associated viral vectors (AAV) often persist as intracellular concatemers but may also integrate. Adeno- and herpes viral vectors do not integrate and earlier generation systems might be relatively immunogenic. Nonviral methods of gene transfer are termed transfection with few restrictions of transgene size and type but often a much less efficient gene transfer that is also short-lived. Traditional gene transfer delivers exons while some vectors (lentiviral, herpes and adenoviral) allow transfer of entire genes that include introns. Recent insights have highlighted the role of non-coding RNA, most prominently, siRNA, miRNA and lncRNA. SiRNA is highly specific, miRNA is less specific, while lncRNA uses highly complex mechanisms that involve secondary structures and intergenic, intronic, overlapping, antisense, and bidirectional location. Several promising preclinical studies have targeted the RhoA or the prostaglandin pathway or modified the extracellular matrix. TGF-β and glaucoma myocilin mutants have been transduced to elevate the intraocular pressure in glaucoma models. Cell based therapies have started to show first promise. Past approaches have focused on the trabecular meshwork and the inner wall of Schlemm's canal while new strategies are concerned with modification of outflow tract elements that are downstream of the trabecular meshwork.

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“…Several different viral vectors have been used to transduce the TM including: Adenovirus serotypes 5 (Ad5) (Hoffman et al , 1997; Millar et al , 2008), 28 or 35 (Ueyama et al , 2014), self-complementary Adeno-Associated Virus (scAAV) (Buie et al , 2010), Herpes Simplex Virus (HSV) (Spencer et al , 2000), HIV pseudotyped with VEEV-G or VSV-G (Lipinski et al , 2014), and Equine infectious anemia virus (EIAV) pseudotyped with VSV-G (Balaggan et al , 2006). …”

Section: Introductionmentioning

confidence: 99%

Elevation of intraocular pressure in rodents using viral vectors targeting the trabecular meshwork

Pang

Millar

Clark

2015

Experimental Eye Research

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Rodents are increasingly being used as glaucoma models to study ocular hypertension, optic neuropathy, and retinopathy. A number of different techniques are used to elevate intraocular pressure in rodent eyes by artificially obstructing the aqueous outflow pathway. Another successful technique to induce ocular hypertension is to transduce the trabecular meshwork of rodent eyes with viral vectors expressing glaucoma associated transgenes to provide more relevant models of glaucomatous damage to the trabecular meshwork. This technique has been used to validate newly discovered glaucoma pathogenesis pathways as well as to develop rodent models of primary open angle glaucoma. Ocular hypertension has successfully been induced by adenovirus 5 mediated delivery of mutant MYOC, bioactivated TGFβ2, SFRP1, DKK1, GREM1, and CD44. Advantages of this approach are: selective tropism for the trabecular meshwork, the ability to use numerous mouse strains, and the relatively rapid onset of IOP elevation. Disadvantages include mild-to-moderate ocular inflammation induced by the Ad5 vector and sometimes transient transgene expression. Current efforts are focused at discovering less immunogenic viral vectors that have tropism for the trabecular meshwork and drive sufficient transgene expression to induce ocular hypertension. This viral vector approach allows rapid proof of concept studies to study glaucomatous damage to the trabecular meshwork without the expensive and time-consuming generation of transgenic mouse lines.

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Ocular Localization and Transduction by Adenoviral Vectors Are Serotype-Dependent and Can Be Modified by Inclusion of RGD Fiber Modifications

Cited by 20 publications

References 32 publications

CRISPR-Cas9–based treatment of myocilin-associated glaucoma

CRISPR-Cas9–based treatment of myocilin-associated glaucoma

Gene transfer to the outflow tract

Elevation of intraocular pressure in rodents using viral vectors targeting the trabecular meshwork

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