XIAP Therapy Increases Survival of Transplanted Rod Precursors in a Degenerating Host Retina

Department of Ophthalmology and Visual Sciences, University of Michigan, Ann Arbor, Michigan, USA.
Investigative ophthalmology & visual science (Impact Factor: 3.4). 10/2010; 52(3):1567-72. DOI: 10.1167/iovs.10-5998
Source: PubMed

ABSTRACT To assess the survival of rod precursor cells transplanted into the Rd9 mouse, a model of X-linked retinal degeneration, and the effect of antiapoptotic therapy with X-linked inhibitor of apoptosis (XIAP) on preventing cell loss.
Dissociated retinal cells from P4 Nrlp-GFP mice were transplanted into the subretinal space of 2-, 5-, and 8-month-old Rd9 mice. Histology, immunohistochemistry, and quantification of integrated cells were performed every month for up to 3 months after transplantation. XIAP delivery to donor cells was accomplished by transfection with adenoassociated virus (AAV-XIAP). Intraretinal activation of immune modulators was assessed using a quantitative real-time polymerase chain reaction-based immune response array.
GFP-positive rod precursors were able to integrate into the outer nuclear layer (ONL) of the Rd9 retina. Transplanted cells underwent morphologic differentiation with the formation of inner and outer segments and synaptic projections to bipolar cells. Integration of donor cells into the ONL increased as a function of host age at the time of transplantation. The number of integrated cells was maximal at 1 month after transplantation and then decreased with time. Survival of integrated cells was significantly increased when donor cells were pretreated with AAV-XIAP. We did not detect any donor cell-specific activation of inflammation within the host retina.
Survival of integrated cells decreases with time after transplantation but can be significantly increased with XIAP antiapoptotic therapy. Preventing programmed cell death through XIAP therapy may be an important component of future therapeutic retinal cell transplantation strategies.

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Available from: Kecia L Feathers, Sep 26, 2015
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    • "AAV5 and AAV8-based vectors were used in the aforementioned studies for developing a treatment for LCA1. These vectors have proven utility for photoreceptor-targeted therapy following subretinal injection in multiple mouse and dog models of inherited retinal disease (Min et al., 2005; Alexander et al., 2007; Boye et al., 2010, 2013; Gorbatyuk et al., 2010; Komaromy et al., 2010; Mao et al., 2011; Pang et al., 2011, 2012; Yao et al., 2011; Beltran et al., 2012; Petit et al., 2012; Lheriteau et al., 2014). Because AAV transduction profiles and the activity of promoters can vary across species, the final decision should be dictated by the serotype and promoter combination’s behavior in a species most closely related to man, non-human primate (NHP). "
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    ABSTRACT: Vertebrate species possess two retinal guanylate cyclases (retGC1 and retGC2) and at least two guanylate cyclase activating proteins (GCAPs), GCAP1 and GCAP2. GCAPs function as Ca(2+) sensors that regulate the activity of guanylate cyclases. Together, these proteins regulate cGMP and Ca(2+) levels within the outer segments of rod and cone photoreceptors. Mutations in GUCY2D, the gene that encodes retGC1, are a leading cause of the most severe form of early onset retinal dystrophy, Leber congenital amaurosis (LCA1). These mutations, which reduce or abolish the ability of retGC1 to replenish cGMP in photoreceptors, are thought to lead to the biochemical equivalent of chronic light exposure in these cells. In spite of this, the majority of LCA1 patients retain normal photoreceptor laminar architecture aside from foveal cone outer segment abnormalities, suggesting they may be good candidates for gene replacement therapy. Work began in the 1980s to characterize multiple animal models of retGC1 deficiency. 34 years later, all models have been used in proof of concept gene replacement studies toward the goal of developing a therapy to treat GUCY2D-LCA1. Here we use the results of these studies as well as those of recent clinical studies to address specific questions relating to clinical application of a gene therapy for treatment of LCA1.
    Frontiers in Molecular Neuroscience 05/2014; 7:43. DOI:10.3389/fnmol.2014.00043 · 4.08 Impact Factor
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    • "Most photoreceptor-targeted studies have focused on AAV5 and AAV8-based vectors. AAV5 has proven utility following subretinal injection in mouse and dog models of photoreceptor-mediated disease (Min et al., 2005; Alexander et al., 2007; Boye et al., 2010; Gorbatyuk et al., 2010; Komaromy et al., 2010; Pang et al., 2010; Li et al., 2011; Mao et al., 2011; Yao et al., 2011; Beltran et al., 2012; Pang et al., 2012). In addition , AAV5 is the only serotype tested thus far to confer a gain of visual function to photoreceptors of nonhuman primates (Mancuso et al., 2009). "
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    ABSTRACT: Mutations in GUCY2D are associated with recessive Leber congenital amaurosis-1 (LCA1). GUCY2D encodes photoreceptor-specific, retinal guanylate cyclase-1 (RetGC1). Reports of retinal degeneration in LCA1 are conflicting; some describe no obvious degeneration and others report loss of both rods and cones. Proof of concept studies in models representing the spectrum of phenotypes is warranted. We have previously demonstrated AAV-mediated RetGC1 is therapeutic in GC1ko mice, a model exhibiting loss of cones only. The purpose of this study was to characterize AAV-mediated gene therapy in the RetGC1/RetGC2 double knockout (GCdko) mouse, a model lacking rod and cone function and exhibiting progressive loss of both photoreceptor subclasses. Use of this model also allowed for the evaluation of the functional efficiency of transgenic RetGC1 isozyme. Subretinal delivery of AAV8(Y733F) vector containing the human rhodopsin kinase (hGRK1) promoter driving murine Gucy2e was performed in GCdko mice at various postnatal time points. Treatment resulted in restoration of rod and cone function at all treatment ages and preservation of retinal structure in GCdko mice treated as late as 7 weeks of age. Functional gains and structural preservation were stable for at least 1 year. Treatment also conferred cortical- and subcortical-based visually- guided behavior. Functional efficiency of transgenic RetGC1 was indistinguishable from that of endogenous isozyme in congenic WT mice. This study clearly demonstrates AAV-mediated RetGC1 expression restores function to and preserves structure of rod and cone photoreceptors in a degenerative model of retinal guanylate cyclase deficiency, further supporting development of an AAV-based vector for treatment of LCA1.
    Human gene therapy 12/2012; 24(2). DOI:10.1089/hum.2012.193 · 3.76 Impact Factor
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    • "In our study, we found that AAV5-hGRK1-GFP at a dose of 6.00 · 10 10 vector genomes delivered was well tolerated in the NHP retina, with no apparent photoreceptor toxicity, suggesting the upper limit of toxicity must be above this concentration. The vector dose used in the current study falls within the range proven sufficient for restoration of visual function/preservation of retinal structure following AAV5-mediated treatment of multiple mouse and dog models of photoreceptor-mediated, inherited retinal disease (Alexander et al., 2007; Boye et al., 2010; Gorbatyuk et al., 2010; Komaromy et al., 2010; Li et al., 2011; Mao et al., 2011; Min et al., 2005; Pang et al., 2010; Pang et al., 2008; Pang et al., 2012b; Yao et al., 2011). In at least 11 reports, all of which utilized AAV5-based vectors produced by the same laboratory (University of Florida), vectors were delivered to respective animal models at concentrations between 1 · 10 9 –4 · 10 10 vector genomes delivered. "
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    ABSTRACT: Abstract Adeno-associated virus (AAV) has proven an effective gene delivery vehicle for the treatment of retinal disease. Ongoing clinical trials using a serotype 2 AAV vector to express RPE65 in the retinal pigment epithelium have proven safe and effective. While many proof-of-concept studies in animal models of retinal disease have suggested that gene transfer to the neural retina will also be effective, a photoreceptor-targeting AAV vector has yet to be used in the clinic, principally because a vector that efficiently but exclusively targets all primate photoreceptors has yet to be demonstrated. Here, we evaluate a serotype 5 AAV vector containing the human rhodopsin kinase (hGRK1) promoter for its ability to target transgene expression to rod and cone photoreceptors when delivered subretinally in a nonhuman primate (NHP). In vivo fluorescent fundus imaging confirmed that AAV5-hGRK1-mediated green fluorescent protein (GFP) expression was restricted to the injection blebs of treated eyes. Optical coherence tomography (OCT) revealed a lack of gross pathology after injection. Neutralizing antibodies against AAV5 were undetectable in post-injection serum samples from subjects receiving uncomplicated subretinal injections (i.e., no hemorrhage). Immunohistochemistry of retinal sections confirmed hGRK1 was active in, and specific for, both rods and cones of NHP retina. Biodistribution studies revealed minimal spread of vector genomes to peripheral tissues. These results suggest that AAV5-hGRK1 is a safe and effective AAV serotype/promoter combination for targeting therapeutic transgene expression protein to rods and cones in a clinical setting.
    Human gene therapy 07/2012; 23(10):1101-15. DOI:10.1089/hum.2012.125 · 3.76 Impact Factor
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