31 results on '"Suja Hiriyanna"'
Search Results
2. A CEP290 C-Terminal Domain Complements the Mutant CEP290 of Rd16 Mice In Trans and Rescues Retinal Degeneration
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Suddhasil Mookherjee, Holly Yu Chen, Kevin Isgrig, Wenhan Yu, Suja Hiriyanna, Rivka Levron, Tiansen Li, Peter Colosi, Wade Chien, Anand Swaroop, and Zhijian Wu
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Biology (General) ,QH301-705.5 - Abstract
Summary: Mutations in CEP290 cause ciliogenesis defects, leading to diverse clinical phenotypes, including Leber congenital amaurosis (LCA). Gene therapy for CEP290-associated diseases is hindered by the 7.4 kb CEP290 coding sequence, which is difficult to deliver in vivo. The multi-domain structure of the CEP290 protein suggests that a specific CEP290 domain may complement disease phenotypes. Thus, we constructed AAV vectors with overlapping CEP290 regions and evaluated their impact on photoreceptor degeneration in Cep290rd16/rd16 and Cep290rd16/rd16;Nrl−/− mice, two models of CEP290-LCA. One CEP290 fragment (the C-terminal 989 residues, including the domain deleted in mutant mice) reconstituted CEP290 function and resulted in cone preservation and delayed rod death. The CEP290 C-terminal domain also improved cilia phenotypes in mouse embryonic fibroblasts and iPSC-derived retinal organoids carrying the Cep290rd16 mutation. Our study strongly argues for in trans complementation of CEP290 mutations by a cognate fragment and suggests therapeutic avenues. : CEP290 mutations are the leading cause of Leber congenital amaurosis, a devastating inherited blindness. Mookherjee et al. show that the in-frame deletion of Cep290 in rd16 mice can be complemented by expressing a cognate protein fragment in trans, suggesting a new avenue for therapy development of CEP290 mutations. Keywords: CEP290, AAV, gene therapy, photoreceptors, retinal degeneration, LCA, ciliopathy
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- 2018
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3. A visual circuit uses complementary mechanisms to support transient and sustained pupil constriction
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William Thomas Keenan, Alan C Rupp, Rachel A Ross, Preethi Somasundaram, Suja Hiriyanna, Zhijian Wu, Tudor C Badea, Phyllis R Robinson, Bradford B Lowell, and Samer S Hattar
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retinal circuitry ,neurotransmitters ,ipRGC ,melanopsin ,vision ,neuropeptides ,Medicine ,Science ,Biology (General) ,QH301-705.5 - Abstract
Rapid and stable control of pupil size in response to light is critical for vision, but the neural coding mechanisms remain unclear. Here, we investigated the neural basis of pupil control by monitoring pupil size across time while manipulating each photoreceptor input or neurotransmitter output of intrinsically photosensitive retinal ganglion cells (ipRGCs), a critical relay in the control of pupil size. We show that transient and sustained pupil responses are mediated by distinct photoreceptors and neurotransmitters. Transient responses utilize input from rod photoreceptors and output by the classical neurotransmitter glutamate, but adapt within minutes. In contrast, sustained responses are dominated by non-conventional signaling mechanisms: melanopsin phototransduction in ipRGCs and output by the neuropeptide PACAP, which provide stable pupil maintenance across the day. These results highlight a temporal switch in the coding mechanisms of a neural circuit to support proper behavioral dynamics.
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- 2016
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4. Ocular and systemic safety of a recombinant AAV8 vector for X-linked retinoschisis gene therapy: GLP studies in rabbits and Rs1-KO mice
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Dario Marangoni, Ronald A Bush, Yong Zeng, Lisa L Wei, Lucia Ziccardi, Camasamudram Vijayasarathy, Joshua T Bartoe, Kiran Palyada, Maria Santos, Suja Hiriyanna, Zhijian Wu, Peter Colosi, and Paul A Sieving
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Genetics ,QH426-470 ,Cytology ,QH573-671 - Abstract
X-linked retinoschisis (XLRS) is a retinal disease caused by mutations in the gene encoding the protein retinoschisin (RS1) and is one of the most common causes of macular degeneration in young men. Our therapeutic approach for XLRS is based on the administration of AAV8-scRS/IRBPhRS, an adeno-associated viral vector coding the human RS1 protein, via the intravitreal (IVT) route. Two Good Laboratory Practice studies, a 9-month study in New Zealand White rabbits (n = 124) injected with AAV8-scRS/IRBPhRS at doses of 2E9, 2E10, 2E11, and 1.5E12 vector genomes/eye (vg/eye), and a 6-month study in Rs1-KO mice (n = 162) dosed with 2E9 and 2E10 vg/eye of the same vector were conducted to assess ocular and systemic safety. A self-resolving, dose-dependent vitreal inflammation was the main ocular finding, and except for a single rabbit dosed with 1.5E12 vg/eye, which showed a retinal detachment, no other ocular adverse event was reported. Systemic toxicity was not identified in either species. Biodistribution analysis in Rs1-KO mice detected spread of vector genome in extraocular tissues, but no evidence of organ or tissues damage was found. These studies indicate that IVT administration of AAV8-scRS/IRBPhRS is safe and well tolerated and support its advancement into a phase 1/2a clinical trial for XLRS.
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- 2016
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5. Rd9 is a naturally occurring mouse model of a common form of retinitis pigmentosa caused by mutations in RPGR-ORF15.
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Debra A Thompson, Naheed W Khan, Mohammad I Othman, Bo Chang, Lin Jia, Garrett Grahek, Zhijian Wu, Suja Hiriyanna, Jacob Nellissery, Tiansen Li, Hemant Khanna, Peter Colosi, Anand Swaroop, and John R Heckenlively
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Medicine ,Science - Abstract
Animal models of human disease are an invaluable component of studies aimed at understanding disease pathogenesis and therapeutic possibilities. Mutations in the gene encoding retinitis pigmentosa GTPase regulator (RPGR) are the most common cause of X-linked retinitis pigmentosa (XLRP) and are estimated to cause 20% of all retinal dystrophy cases. A majority of RPGR mutations are present in ORF15, the purine-rich terminal exon of the predominant splice-variant expressed in retina. Here we describe the genetic and phenotypic characterization of the retinal degeneration 9 (Rd9) strain of mice, a naturally occurring animal model of XLRP. Rd9 mice were found to carry a 32-base-pair duplication within ORF15 that causes a shift in the reading frame that introduces a premature-stop codon. Rpgr ORF15 transcripts, but not protein, were detected in retinas from Rd9/Y male mice that exhibited retinal pathology, including pigment loss and slowly progressing decrease in outer nuclear layer thickness. The levels of rhodopsin and transducin in rod outer segments were also decreased, and M-cone opsin appeared mislocalized within cone photoreceptors. In addition, electroretinogram (ERG) a- and b-wave amplitudes of both Rd9/Y male and Rd9/Rd9 female mice showed moderate gradual reduction that continued to 24 months of age. The presence of multiple retinal features that correlate with findings in individuals with XLRP identifies Rd9 as a valuable model for use in gaining insight into ORF15-associated disease progression and pathogenesis, as well as accelerating the development and testing of therapeutic strategies for this common form of retinal dystrophy.
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- 2012
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6. In vitro modeling and rescue of ciliopathy associated with IQCB1/NPHP5 mutations using patient-derived cells
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Kamil, Kruczek, Zepeng, Qu, Emily, Welby, Hiroko, Shimada, Suja, Hiriyanna, Milton A, English, Wadih M, Zein, Brian P, Brooks, and Anand, Swaroop
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Cytoskeletal Proteins ,Calmodulin ,Antigens, Neoplasm ,Mutation ,Genetics ,Humans ,Calmodulin-Binding Proteins ,Cell Cycle Proteins ,Cell Biology ,Retinal Pigments ,Biochemistry ,Ciliopathies ,Developmental Biology - Abstract
Mutations in the IQ calmodulin-binding motif containing B1 (IQCB1)/NPHP5 gene encoding the ciliary protein nephrocystin 5 cause early-onset blinding disease Leber congenital amaurosis (LCA), together with kidney dysfunction in Senior-Løken syndrome. For in vitro disease modeling, we obtained dermal fibroblasts from patients with NPHP5-LCA that were reprogrammed into induced pluripotent stem cells (iPSCs) and differentiated into retinal pigment epithelium (RPE) and retinal organoids. Patient fibroblasts and RPE demonstrated aberrantly elongated ciliary axonemes. Organoids revealed impaired development of outer segment structures, which are modified primary cilia, and mislocalization of visual pigments to photoreceptor cell soma. All patient-derived cells showed reduced levels of CEP290 protein, a critical cilia transition zone component interacting with NPHP5, providing a plausible mechanism for aberrant ciliary gating and cargo transport. Disease phenotype in NPHP5-LCA retinal organoids could be rescued by adeno-associated virus (AAV)-mediated IQCB1/NPHP5 gene augmentation therapy. Our studies thus establish a human disease model and a path for treatment of NPHP5-LCA.
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- 2022
7. Gene Therapy of Dominant CRX-Leber Congenital Amaurosis using Patient Stem Cell-Derived Retinal Organoids
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Ananya Samanta, James Gentry, Kamil Kruczek, Zepeng Qu, Benjamin R. Fadl, Brian P. Brooks, Colin J Chu, Zhijian Wu, Zachary Batz, Linn Gieser, Laura Campello, Suja Hiriyanna, Anand Swaroop, and Mugdha Samant
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0301 basic medicine ,Retinal degeneration ,Opsin ,genetic structures ,Genetic enhancement ,Leber Congenital Amaurosis ,medicine.disease_cause ,Biochemistry ,chemistry.chemical_compound ,0302 clinical medicine ,disease modeling ,Child ,Induced pluripotent stem cell ,transcription factor ,Mutation ,iPSC ,Cell Differentiation ,AAV ,Dependovirus ,Organoids ,Phenotype ,Child, Preschool ,Female ,Single-Cell Analysis ,pluripotent stem cells ,Stem cell ,Adult ,Induced Pluripotent Stem Cells ,3-D organoids ,Biology ,Models, Biological ,Retina ,Article ,03 medical and health sciences ,scRNA-seq ,Genetics ,medicine ,Humans ,Photoreceptor Cells ,Homeodomain Proteins ,therapy ,Opsins ,Sequence Analysis, RNA ,Retinal ,Genetic Therapy ,Cell Biology ,medicine.disease ,eye diseases ,030104 developmental biology ,chemistry ,Trans-Activators ,retinal degeneration ,Cancer research ,Homeobox ,sense organs ,Transcriptome ,transcriptome ,030217 neurology & neurosurgery ,Developmental Biology - Abstract
Summary Mutations in the photoreceptor transcription factor gene cone-rod homeobox (CRX) lead to distinct retinopathy phenotypes, including early-onset vision impairment in dominant Leber congenital amaurosis (LCA). Using induced pluripotent stem cells (iPSCs) from a patient with CRX-I138fs48 mutation, we established an in vitro model of CRX-LCA in retinal organoids that showed defective photoreceptor maturation by histology and gene profiling, with diminished expression of visual opsins. Adeno-associated virus (AAV)-mediated CRX gene augmentation therapy partially restored photoreceptor phenotype and expression of phototransduction-related genes as determined by single-cell RNA-sequencing. Retinal organoids derived from iPSCs of a second dominant CRX-LCA patient carrying K88N mutation revealed the loss of opsin expression as a common phenotype, which was alleviated by AAV-mediated augmentation of CRX. Our studies provide a proof-of-concept for developing gene therapy of dominant CRX-LCA and other CRX retinopathies., Graphical Abstract, Highlights • Leber congenital amaurosis caused by CRX mutations is modeled in retinal organoids • Patient iPSCs-derived organoids show impaired expression of visual opsins • AAV-mediated CRX delivery partially restores expression of phototransduction genes • Gene therapy is applicable to mutations in DNA-binding and transactivation domains, In this article, Swaroop and colleagues show impaired expression of opsin visual pigments in retinal organoids derived from iPSC lines of patients with Leber congenital amaurosis, caused by dominant mutations in the photoreceptor transcription factor gene CRX. AAV-mediated CRX gene augmentation partially restores the expression of phototransduction genes, suggesting a potential therapeutic strategy.
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- 2021
8. A Common Outer Retinal Change in Retinal Degeneration by Optical Coherence Tomography Can Be Used to Assess Outcomes of Gene Therapy
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Zhijian Wu, Wenbo Li, Myung Kuk Joe, Wenhan Yu, Suja Hiriyanna, Shreya A Shah, Haohua Qian, and Mones Abu-Asab
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Retinal degeneration ,Pathology ,medicine.medical_specialty ,Genetic enhancement ,Mutant ,H&E stain ,Retina ,Mice ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Optical coherence tomography ,Genetics ,medicine ,Animals ,Humans ,Molecular Biology ,Pathological ,Research Articles ,030304 developmental biology ,0303 health sciences ,medicine.diagnostic_test ,business.industry ,Retinal Degeneration ,Retinal ,Genetic Therapy ,medicine.disease ,Disease Models, Animal ,medicine.anatomical_structure ,chemistry ,030220 oncology & carcinogenesis ,Molecular Medicine ,sense organs ,business ,Tomography, Optical Coherence - Abstract
Identifying early disease hallmarks in animal models with slow disease progression may expedite disease detection and assessment of treatment outcomes. Using optical coherence tomography, a widely applied noninvasive method for monitoring retinal structure changes, we analyzed retinal optical sections from six mouse lines with retinal degeneration caused by mutations in different disease-causing genes. While images from wild-type mice revealed four well-separated hyper-reflective bands in the outer retina (designated as outer retina reflective bands, ORRBs) at all ages, the second band (ORRB2) and the third band (ORRB3) were merged in retinas of five mutant mouse lines at early ages, suggesting the pathological nature of this alteration. This ORRB change appeared to be degenerating photoreceptor related, and occurred before obvious morphological changes that can be identified on both hematoxylin and eosin-stained sections and electron microscopic sections. Importantly, the merging of ORRB2 and ORRB3 was reversed by treatment with adeno-associated viral vector-mediated gene replacement therapies, and this restoration occurred much earlier than measurable functional or structural improvement. Our data suggest that the ORRB change could be a common hallmark of early retinal degeneration and its restoration could be used for rapid and noninvasive assessment of therapeutic effects following gene therapy or other treatment interventions.
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- 2019
9. Retinal Structure and Gene Therapy Outcome in Retinoschisin-Deficient Mice Assessed by Spectral-Domain Optical Coherence Tomography
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Ya Xian Wang, Zhijian Wu, Suja Hiriyanna, Ronald S. Petralia, Ronald A. Bush, Yong Zeng, Camasamudram Vijayasarathy, Paul A. Sieving, and Hongman Song
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0301 basic medicine ,Pathology ,genetic structures ,Retinoschisis ,Ophthalmology & Optometry ,Medical and Health Sciences ,chemistry.chemical_compound ,Mice ,0302 clinical medicine ,Microscopy, Immunoelectron ,Tomography ,Mice, Knockout ,Microscopy ,medicine.diagnostic_test ,Articles ,Biological Sciences ,medicine.anatomical_structure ,Intravitreal Injections ,outer retina reflective band ,Photoreceptor inner segment ,X-linked retinoschisis ,AAV gene therapy ,Tomography, Optical Coherence ,medicine.medical_specialty ,Knockout ,Genetic Vectors ,Retina ,03 medical and health sciences ,Optical coherence tomography ,outer nuclear layer ,medicine ,Electroretinography ,Outer Limiting Membrane ,Animals ,Outer nuclear layer ,Eye Proteins ,Immunoelectron ,optical coherence tomography ,business.industry ,Animal ,Retinal ,Genetic Therapy ,medicine.disease ,eye diseases ,Disease Models, Animal ,030104 developmental biology ,chemistry ,Optical Coherence ,Disease Models ,030221 ophthalmology & optometry ,sense organs ,business ,Cell Adhesion Molecules - Abstract
Author(s): Zeng, Yong; Petralia, Ronald S; Vijayasarathy, Camasamudram; Wu, Zhijian; Hiriyanna, Suja; Song, Hongman; Wang, Ya-Xian; Sieving, Paul A; Bush, Ronald A | Abstract: PurposeSpectral-domain optical coherence tomography (SD-OCT) was used to characterize the retinal phenotype, natural history, and treatment responses in a mouse model of X-linked retinoschisis (Rs1-KO) and to identify new structural markers of AAV8-mediated gene therapy outcome.MethodsOptical coherence tomography scans were performed on wild-type and Rs1-KO mouse retinas between 1 and 12 months of age and on Rs1-KO mice after intravitreal injection of AAV8-scRS/IRBPhRS (AAV8-RS1). Cavities and photoreceptor outer nuclear layer (ONL) thickness were measured, and outer retina reflective band (ORRB) morphology was examined with age and after AAV8-RS1 treatment. Outer retina reflective band morphology was compared to immunohistochemical staining of the outer limiting membrane (OLM) and photoreceptor inner segment (IS) mitochondria and to electron microscopy (EM) images of IS.ResultsRetinal cavity size in Rs1-KO mice increased between 1 and 4 months and decreased thereafter, while ONL thickness declined steadily, comparable to previous histologic studies. Wild-type retina had four ORRBs. In Rs1-KO, ORRB1was fragmented from 1 month, but was normal after 8 months; ORRB2 and ORRB3 were merged at all ages. Outer retina reflective band morphology returned to normal after AAV-RS1 therapy, paralleling the recovery of the OLM and IS mitochondria as indicated by anti-β-catenin and anti-COX4 labeling, respectively, and EM.ConclusionsSpectral-domain OCT is a sensitive, noninvasive tool to monitor subtle changes in retinal morphology, disease progression, and effects of therapies in mouse models. The ORRBs may be useful to assess the outcome of gene therapy in the treatment of X-linked retinoschisis patients.
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- 2016
10. A CEP290 C-Terminal Domain Complements the Mutant CEP290 of Rd16 Mice In Trans and Rescues Retinal Degeneration
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Holly Y. Chen, Zhijian Wu, Kevin Isgrig, Suja Hiriyanna, Peter Colosi, Wenhan Yu, Rivka Levron, Suddhasil Mookherjee, Tiansen Li, Anand Swaroop, and Wade W. Chien
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0301 basic medicine ,Retinal degeneration ,Male ,genetic structures ,Mutant ,Leber Congenital Amaurosis ,Cell Cycle Proteins ,030105 genetics & heredity ,Biology ,medicine.disease_cause ,General Biochemistry, Genetics and Molecular Biology ,Retina ,Article ,03 medical and health sciences ,Mice ,Antigens, Neoplasm ,Ciliogenesis ,medicine ,Animals ,Humans ,Cilia ,Eye Proteins ,lcsh:QH301-705.5 ,Mice, Knockout ,Mutation ,Cilium ,Retinal Degeneration ,Genetic Therapy ,Dependovirus ,medicine.disease ,Phenotype ,Cell biology ,Complementation ,Ciliopathy ,Cytoskeletal Proteins ,Disease Models, Animal ,030104 developmental biology ,Basic-Leucine Zipper Transcription Factors ,lcsh:Biology (General) ,Retinal Cone Photoreceptor Cells ,Female ,sense organs - Abstract
Summary: Mutations in CEP290 cause ciliogenesis defects, leading to diverse clinical phenotypes, including Leber congenital amaurosis (LCA). Gene therapy for CEP290-associated diseases is hindered by the 7.4 kb CEP290 coding sequence, which is difficult to deliver in vivo. The multi-domain structure of the CEP290 protein suggests that a specific CEP290 domain may complement disease phenotypes. Thus, we constructed AAV vectors with overlapping CEP290 regions and evaluated their impact on photoreceptor degeneration in Cep290rd16/rd16 and Cep290rd16/rd16;Nrl−/− mice, two models of CEP290-LCA. One CEP290 fragment (the C-terminal 989 residues, including the domain deleted in mutant mice) reconstituted CEP290 function and resulted in cone preservation and delayed rod death. The CEP290 C-terminal domain also improved cilia phenotypes in mouse embryonic fibroblasts and iPSC-derived retinal organoids carrying the Cep290rd16 mutation. Our study strongly argues for in trans complementation of CEP290 mutations by a cognate fragment and suggests therapeutic avenues. : CEP290 mutations are the leading cause of Leber congenital amaurosis, a devastating inherited blindness. Mookherjee et al. show that the in-frame deletion of Cep290 in rd16 mice can be complemented by expressing a cognate protein fragment in trans, suggesting a new avenue for therapy development of CEP290 mutations. Keywords: CEP290, AAV, gene therapy, photoreceptors, retinal degeneration, LCA, ciliopathy
- Published
- 2018
11. Retinal AAV8-RS1 Gene Therapy for X-Linked Retinoschisis: Initial Findings from a Phase I/IIa Trial by Intravitreal Delivery
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Sten Kjellstrom, Paul A. Sieving, Lisa L. Wei, Camasamudram Vijayasarathy, Yong Zeng, H. Nida Sen, Zhijian Wu, Henry E. Wiley, Brett G. Jeffrey, Peter Colosi, J. Fraser Wright, Suja Hiriyanna, Ronald A. Bush, Catherine A Cukras, Amy Turriff, Tae Kwon Park, Dario Marangoni, Lucia Ziccardi, Cukras, C, Wiley, He, Jeffrey, Bg, Sen, Hn, Turriff, A, Zeng, Y, Vijayasarathy, C, Marangoni, D, Ziccardi, L, Kjellstrom, S, Park, Tk, Hiriyanna, S, Wright, Jf, Colosi, P, Wu, Z, Bush, Ra, Wei, Ll, and Sieving, Pa.
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Male ,0301 basic medicine ,Technology ,genetic structures ,Genetic enhancement ,Retinoschisin Protein ,Retinoschisis ,Eye ,Medical and Health Sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Drug Discovery ,Retinal detachment ,clinical trial ,Middle Aged ,Biological Sciences ,gene therapy ,AAV vector ,medicine.anatomical_structure ,Tolerability ,6.1 Pharmaceuticals ,Intravitreal Injections ,Molecular Medicine ,Female ,X-linked retinoschisis ,Biotechnology ,Adult ,medicine.medical_specialty ,Clinical Trials and Supportive Activities ,X-linked retinoschisi ,Retina ,ocular disease ,retinal disease ,Young Adult ,03 medical and health sciences ,Clinical Research ,Ophthalmology ,Genetics ,medicine ,Humans ,Eye Proteins ,Eye Disease and Disorders of Vision ,Molecular Biology ,Aged ,Pharmacology ,business.industry ,Neurosciences ,Evaluation of treatments and therapeutic interventions ,Retinal ,Genetic Therapy ,medicine.disease ,eye diseases ,Clinical trial ,030104 developmental biology ,chemistry ,Mutation ,030221 ophthalmology & optometry ,business - Abstract
This study evaluated the safety and tolerability of ocular RS1 adeno-associated virus (AAV8-RS1) gene augmentation therapy to the retina of participants with X-linked retinoschisis (XLRS). XLRS is a monogenic trait affecting only males, caused by mutations in the RS1 gene. Retinoschisin protein is secreted principally in the outer retina, and its absence results in retinal cavities, synaptic dysfunction, reduced visual acuity, and susceptibility to retinal detachment. This phase I/IIa single-center, prospective, open-label, three-dose-escalation clinical trial administered vector to nine participants with pathogenic RS1 mutations. The eye of each participant with worse acuity (≤63 letters; Snellen 20/63) received the AAV8-RS1 gene vector by intravitreal injection. Three participants were assigned to each of three dosage groups: 1e9 vector genomes (vg)/eye, 1e10 vg/eye, and 1e11 vg/eye. The investigational product was generally well tolerated in all but one individual. Ocular events included dose-related inflammation that resolved with topical and oral corticosteroids. Systemic antibodies against AAV8 increased in a dose-related fashion, but no antibodies against RS1 were observed. Retinal cavities closed transiently in one participant. Additional doses and immunosuppressive regimens are being explored to pursue evidence of safety and efficacy (ClinicalTrials.gov: NCT02317887).
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- 2018
12. Long-term rescue of cone photoreceptor degeneration in retinitis pigmentosa 2 (RP2)-knockout mice by gene replacement therapy
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Yichao Li, Peter Colosi, Zhijian Wu, Anand Swaroop, Hemant Khanna, Tiansen Li, Kayleigh Kaneshiro, Wei Li, Suja Hiriyanna, Haohua Qian, Linjing Li, and Suddhasil Mookherjee
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Retinal degeneration ,Opsin ,genetic structures ,Genetic Vectors ,Biology ,Retinal Cone Photoreceptor Cells ,Mice ,GTP-Binding Proteins ,Retinitis pigmentosa ,Electroretinography ,Genetics ,medicine ,Animals ,Humans ,Pyrophosphatases ,Eye Proteins ,Molecular Biology ,Genetics (clinical) ,Retinal regeneration ,Mice, Knockout ,Gene therapy of the human retina ,medicine.diagnostic_test ,Retinal Degeneration ,Intracellular Signaling Peptides and Proteins ,Membrane Proteins ,Genetic Diseases, X-Linked ,Genetic Therapy ,Articles ,General Medicine ,medicine.disease ,eye diseases ,HEK293 Cells ,Mutation ,Cancer research ,sense organs ,Retinitis Pigmentosa ,Photopic vision - Abstract
Retinal neurodegenerative diseases are especially attractive targets for gene replacement therapy, which appears to be clinically effective for several monogenic diseases. X-linked forms of retinitis pigmentosa (XLRP) are relatively severe blinding disorders, resulting from progressive photoreceptor dysfunction primarily caused by mutations in RPGR or RP2 gene. With a goal to develop gene therapy for the XLRP-RP2 disease, we first performed detailed characterization of the Rp2-knockout (Rp2-KO) mice and observed early-onset cone dysfunction, which was followed by progressive cone degeneration, mimicking cone vision impairment in XLRP patients. The mice also exhibited distinct and significantly delayed falling phase of photopic b-wave of electroretinogram (ERG). Concurrently, we generated a self-complementary adeno-associated viral (AAV) vector carrying human RP2-coding sequence and demonstrated its ability to mediate stable RP2 protein expression in mouse photoreceptors. A long-term efficacy study was then conducted in Rp2-KO mice following AAV-RP2 vector administration. Preservation of cone function was achieved with a wide dose range over 18-month duration, as evidenced by photopic ERG and optomotor tests. The slower b-wave kinetics was also completely restored. Morphologically, the treatment preserved cone viability, corrected mis-trafficking of M-cone opsin and restored cone PDE6 expression. The therapeutic effect was achieved even in mice that received treatment at an advanced disease stage. The highest AAV-RP2 dose group demonstrated retinal toxicity, highlighting the importance of careful vector dosing in designing future human trials. The wide range of effective dose, a broad treatment window and long-lasting therapeutic effects should make the RP2 gene therapy attractive for clinical development.
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- 2015
13. Preclinical Safety Evaluation of a Recombinant AAV8 Vector for X-Linked Retinoschisis After Intravitreal Administration in Rabbits
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Lisa L. Wei, Dario Marangoni, Ronald A. Bush, Peter Colosi, Zhijian Wu, Paul A. Sieving, Suja Hiriyanna, Yong Zeng, Henry E. Wiley, Caroline J. Zeiss, and Camasamudram Vijayasarathy
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medicine.medical_specialty ,Retinoschisis ,Genetic enhancement ,Genetic Vectors ,DNA, Recombinant ,Eye ,Viral vector ,chemistry.chemical_compound ,Ophthalmology ,Genetics ,medicine ,Animals ,Eye Proteins ,Eye Disease and Disorders of Vision ,Research Articles ,Genetics (clinical) ,Recombinant ,business.industry ,Retinal ,DNA ,Genetic Therapy ,Gene Therapy ,Intravitreal administration ,Dependovirus ,Macular degeneration ,medicine.disease ,eye diseases ,Surgery ,Clinical trial ,chemistry ,Intravitreal Injections ,Rabbits ,RETINOSCHISIN ,business - Abstract
X-linked retinoschisis (XLRS) is a retinal disease caused by mutations in the gene encoding the protein retinoschisin (RS1) and one of the most common causes of macular degeneration in young men. Currently, no FDA-approved treatments are available for XLRS and a replacement gene therapy could provide a promising strategy. We have developed a novel gene therapy approach for XLRS, based on the administration of AAV8-scRS/IRBPhRS, an adeno-associated viral vector coding the human RS1 protein, via the intravitreal route. On the basis of our prior study in an Rs1-KO mouse, this construct transduces efficiently all the retinal layers, resulting in an RS1 expression similar to that observed in the wild-type and improving retinal structure and function. In support of a clinical trial, we carried out a study to evaluate the ocular safety of intravitreal administration of AAV8-scRS/IRBPhRS into 39 New Zealand White rabbits. Two dose levels of vector, 2e(10) and 2e(11) vector genomes per eye (vg/eye), were tested and ocular inflammation was monitored over a 12-week period by serial ophthalmological and histopathological analysis. A mild ocular inflammatory reaction, consisting mainly of vitreous infiltrates, was observed within 4 weeks from injection, in both 2e(10) and 2e(11) vg/eye groups and was likely driven by the AAV8 capsid. At 12-week follow-up, ophthalmological examination revealed no clinical signs of vitreitis in either of the dose groups. However, while vitreous inflammatory infiltrate was significantly reduced in the 2e(10) vg/eye group at 12 weeks, some rabbits in the higher dose group still showed persistence of inflammatory cells, histologically. In conclusion, intravitreal administration of AAV8-scRS/IRBPhRS into the rabbit eye produces a mild and transient intraocular inflammation that resolves, at a 2e(10) vg/eye dose, within 3 months, and does not cause irreversible tissue damages. These data support the initiation of a clinical trial of intravitreal administration of AAV8-scRS/IRBPhRS in XLRS patients.
- Published
- 2014
14. Molecular codes for cell type specification in Brn3 retinal ganglion cells
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Octavian Popescu, Katherine Chuang, Tudor C. Badea, Miruna Georgiana Ghinia, Friedrich Kretschmer, Matthew Brooks, Szilard Sajgo, Zhijian Wu, and Suja Hiriyanna
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Male ,Retinal Ganglion Cells ,0301 basic medicine ,Cell type ,genetic structures ,Upstream and downstream (transduction) ,Synaptogenesis ,Dendrite ,Cell Communication ,Biology ,Retinal ganglion ,Transcriptome ,Mice ,03 medical and health sciences ,medicine ,Animals ,Transcription factor ,Multidisciplinary ,Gene Expression Profiling ,Brain ,High-Throughput Nucleotide Sequencing ,Membrane Proteins ,eye diseases ,Axon Guidance ,Transcription Factor Brn-3 ,030104 developmental biology ,medicine.anatomical_structure ,PNAS Plus ,Female ,Axon guidance ,sense organs ,Neuroscience - Abstract
Visual information is conveyed from the eye to the brain by distinct types of retinal ganglion cells (RGCs). It is largely unknown how RGCs acquire their defining morphological and physiological features and connect to upstream and downstream synaptic partners. The three Brn3/Pou4f transcription factors (TFs) participate in a combinatorial code for RGC type specification, but their exact molecular roles are still unclear. We use deep sequencing to define (i) transcriptomes of Brn3a- and/or Brn3b-positive RGCs, (ii) Brn3a- and/or Brn3b-dependent RGC transcripts, and (iii) transcriptomes of retinorecipient areas of the brain at developmental stages relevant for axon guidance, dendrite formation, and synaptogenesis. We reveal a combinatorial code of TFs, cell surface molecules, and determinants of neuronal morphology that is differentially expressed in specific RGC populations and selectively regulated by Brn3a and/or Brn3b. This comprehensive molecular code provides a basis for understanding neuronal cell type specification in RGCs.
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- 2017
15. Nrl knockdown by AAV-delivered CRISPR/Cas9 prevents retinal degeneration in mice
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Xun Sun, Suddhasil Mookherjee, Matthew Brooks, Yasaman Ataeijannati, Jung-Woong Kim, Zhijian Wu, Vijender Chaitankar, Wenhan Yu, Tiansen Li, Lijin Dong, Suja Hiriyanna, and Anand Swaroop
- Subjects
0301 basic medicine ,Retinal degeneration ,Leucine zipper ,genetic structures ,Cell Survival ,Science ,General Physics and Astronomy ,Degeneration (medical) ,Biology ,Article ,General Biochemistry, Genetics and Molecular Biology ,Mice ,03 medical and health sciences ,chemistry.chemical_compound ,Retinal Rod Photoreceptor Cells ,Retinitis pigmentosa ,medicine ,Animals ,CRISPR ,Eye Proteins ,Gene Editing ,Genetics ,Gene knockdown ,Multidisciplinary ,Cas9 ,Retinal Degeneration ,Retinal ,General Chemistry ,Dependovirus ,medicine.disease ,eye diseases ,Cell biology ,Basic-Leucine Zipper Transcription Factors ,030104 developmental biology ,chemistry ,Gene Knockdown Techniques ,Retinal Cone Photoreceptor Cells ,sense organs ,CRISPR-Cas Systems ,Retinitis Pigmentosa - Abstract
In retinitis pigmentosa, loss of cone photoreceptors leads to blindness, and preservation of cone function is a major therapeutic goal. However, cone loss is thought to occur as a secondary event resulting from degeneration of rod photoreceptors. Here we report a genome editing approach in which adeno-associated virus (AAV)-mediated CRISPR/Cas9 delivery to postmitotic photoreceptors is used to target the Nrl gene, encoding for Neural retina-specific leucine zipper protein, a rod fate determinant during photoreceptor development. Following Nrl disruption, rods gain partial features of cones and present with improved survival in the presence of mutations in rod-specific genes, consequently preventing secondary cone degeneration. In three different mouse models of retinal degeneration, the treatment substantially improves rod survival and preserves cone function. Our data suggest that CRISPR/Cas9-mediated NRL disruption in rods may be a promising treatment option for patients with retinitis pigmentosa., Retinitis pigmentosa is mainly caused by mutations that initially affect survival of rod photoreceptors, leading to secondary loss of cones. Here the authors use gene editing to prevent rod degeneration, leading to survival of cones and improved vision in mice.
- Published
- 2017
16. Author response: A visual circuit uses complementary mechanisms to support transient and sustained pupil constriction
- Author
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Preethi Somasundaram, Tudor C. Badea, William T Keenan, Rachel A. Ross, Zhijian Wu, Samer Hattar, Suja Hiriyanna, Alan C. Rupp, Bradford B. Lowell, and Phyllis R. Robinson
- Subjects
0301 basic medicine ,Pupil constriction ,03 medical and health sciences ,030104 developmental biology ,Computer science ,Transient (oscillation) ,Neuroscience - Published
- 2016
17. Preclinical Dose-Escalation Study of Intravitreal AAV-RS1 Gene Therapy in a Mouse Model of X-linked Retinoschisis: Dose-Dependent Expression and Improved Retinal Structure and Function
- Author
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Maria José Santos, Sten Kjellstrom, Peter Colosi, Zhijian Wu, Yong Zeng, Ronald A. Bush, Paul A. Sieving, Camasamudram Vijayasarathy, Jinbo Li, and Suja Hiriyanna
- Subjects
0301 basic medicine ,Male ,Pathology ,Time Factors ,Genetic enhancement ,Medical Biotechnology ,Retinoschisis ,Gene Expression ,Eye ,chemistry.chemical_compound ,Mice ,Genes, X-Linked ,Transduction, Genetic ,Tomography ,Research Articles ,Mice, Knockout ,medicine.diagnostic_test ,Gene Therapy ,Dependovirus ,Immunohistochemistry ,medicine.anatomical_structure ,Intravitreal Injections ,Molecular Medicine ,Development of treatments and therapeutic interventions ,Erg ,Tomography, Optical Coherence ,Biotechnology ,medicine.medical_specialty ,Knockout ,Clinical Sciences ,Genetic Vectors ,Biology ,Retina ,03 medical and health sciences ,Transduction ,Rare Diseases ,Genetic ,Clinical Research ,medicine ,Genetics ,Electroretinography ,Animals ,Outer nuclear layer ,Eye Proteins ,Molecular Biology ,Eye Disease and Disorders of Vision ,5.2 Cellular and gene therapies ,Animal ,Neurosciences ,Retinal ,Genetic Therapy ,X-Linked ,medicine.disease ,Disease Models, Animal ,030104 developmental biology ,chemistry ,Genes ,Optical Coherence ,Disease Models ,RETINOSCHISIN ,Cell Adhesion Molecules - Abstract
Gene therapy for inherited retinal diseases has been shown to ameliorate functional and structural defects in both animal models and in human clinical trials. X-linked retinoschisis (XLRS) is an early-age onset macular dystrophy resulting from loss of an extracellular matrix protein (RS1). In preparation for a human clinical gene therapy trial, we conducted a dose-range efficacy study of the clinical vector, a self-complementary AAV delivering a human retinoschisin (RS1) gene under control of the RS1 promoter and an interphotoreceptor binding protein enhancer (AAV8-scRS/IRBPhRS), in the retinoschisin knockout (Rs1-KO) mouse. The therapeutic vector at 1 × 10(6) to 2.5 × 10(9) (1E6-2.5E9) vector genomes (vg)/eye or vehicle was administered to one eye of 229 male Rs1-KO mice by intravitreal injection at 22 ± 3 days postnatal age (PN). Analysis of retinal function (dark-adapted electroretinogram, ERG), structure (cavities and outer nuclear layer thickness) by in vivo retinal imaging using optical coherence tomography, and retinal immunohistochemistry (IHC) for RS1 was done 3-4 months and/or 6-9 months postinjection (PI). RS1 IHC staining was dose dependent across doses ≥1E7 vg/eye, and the threshold for significant improvement in all measures of retinal structure and function was 1E8 vg/eye. Higher doses, however, did not produce additional improvement. At all doses showing efficacy, RS1 staining in Rs1-KO mouse was less than that in wild-type mice. Improvement in the ERG and RS1 staining was unchanged or greater at 6-9 months than at 3-4 months PI. This study demonstrates that vitreal administration of AAV8 scRS/IRBPhRS produces significant improvement in retinal structure and function in the mouse model of XLRS over a vector dose range that can be extended to a human trial. It indicates that a fully normal level of RS1 expression is not necessary for a therapeutic effect.
- Published
- 2016
18. Ocular and Systemic Safety of a Recombinant AAV8 Vector for X-linked Retinoschisis Gene Therapy: GLP studies in rabbits and Rs1-KO mice
- Author
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Joshua T. Bartoe, Lucia Ziccardi, Zhijian Wu, Camasamudram Vijayasarathy, Paul A. Sieving, Kiran Palyada, Lisa L. Wei, Dario Marangoni, Ronald A. Bush, Maria José Santos, Suja Hiriyanna, Peter Colosi, Yong Zeng, Marangoni, D, Bush, Ra, Zeng, Y, Wei, Ll, Ziccardi, L, Vijayasarathy, C, Bartoe, Jt, Palyada, K, Santos, M, Hiriyanna, S, Wu, Z, Colosi, P, and Sieving, Pa.
- Subjects
0301 basic medicine ,Pathology ,medicine.medical_specialty ,lcsh:QH426-470 ,Genetic enhancement ,Retinoschisis ,Inflammation ,Eye ,Article ,Viral vector ,03 medical and health sciences ,chemistry.chemical_compound ,Genetics ,medicine ,lcsh:QH573-671 ,Eye Disease and Disorders of Vision ,Molecular Biology ,lcsh:Cytology ,business.industry ,Neurosciences ,Retinal detachment ,Retinal ,Gene Therapy ,Macular degeneration ,medicine.disease ,eye diseases ,lcsh:Genetics ,030104 developmental biology ,chemistry ,Molecular Medicine ,sense organs ,medicine.symptom ,business ,RETINOSCHISIN ,Biotechnology - Abstract
X-linked retinoschisis (XLRS) is a retinal disease caused by mutations in the gene encoding the protein retinoschisin (RS1) and is one of the most common causes of macular degeneration in young men. Our therapeutic approach for XLRS is based on the administration of AAV8-scRS/IRBPhRS, an adeno-associated viral vector coding the human RS1 protein, via the intravitreal (IVT) route. Two Good Laboratory Practice studies, a 9-month study in New Zealand White rabbits (n = 124) injected with AAV8-scRS/IRBPhRS at doses of 2E9, 2E10, 2E11, and 1.5E12 vector genomes/eye (vg/eye), and a 6-month study in Rs1-KO mice (n = 162) dosed with 2E9 and 2E10 vg/eye of the same vector were conducted to assess ocular and systemic safety. A self-resolving, dose-dependent vitreal inflammation was the main ocular finding, and except for a single rabbit dosed with 1.5E12 vg/eye, which showed a retinal detachment, no other ocular adverse event was reported. Systemic toxicity was not identified in either species. Biodistribution analysis in Rs1-KO mice detected spread of vector genome in extraocular tissues, but no evidence of organ or tissues damage was found. These studies indicate that IVT administration of AAV8-scRS/IRBPhRS is safe and well tolerated and support its advancement into a phase 1/2a clinical trial for XLRS.
- Published
- 2016
19. A long-term efficacy study of gene replacement therapy for RPGR-associated retinal degeneration
- Author
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Zhijian Wu, Tiansen Li, Peter Colosi, Maria M Campos, Chun Gao, Suja Hiriyanna, Robert N. Fariss, Suddhasil Mookherjee, Haohua Qian, Paul A. Sieving, and Anand Swaroop
- Subjects
Retinal degeneration ,Male ,Opsin ,Genetic enhancement ,Neurodegenerative ,Inbred C57BL ,Eye ,Medical and Health Sciences ,Mice ,Genetics (clinical) ,Genetics ,Mice, Knockout ,Genetics & Heredity ,Gene therapy of the human retina ,medicine.diagnostic_test ,General Medicine ,Retinitis pigmentosa GTPase regulator ,Articles ,Exons ,Gene Therapy ,Dependovirus ,Biological Sciences ,medicine.anatomical_structure ,Development of treatments and therapeutic interventions ,Retinitis Pigmentosa ,Biotechnology ,Knockout ,Genetic Vectors ,Biology ,Retina ,Open Reading Frames ,Rare Diseases ,Retinitis pigmentosa ,medicine ,Electroretinography ,Animals ,Humans ,Eye Proteins ,Molecular Biology ,Eye Disease and Disorders of Vision ,5.2 Cellular and gene therapies ,Animal ,Neurosciences ,Genetic Therapy ,medicine.disease ,eye diseases ,Mice, Inbred C57BL ,Disease Models, Animal ,Disease Models ,Mutation ,Cancer research ,Carrier Proteins - Abstract
Mutations in the retinitis pigmentosa GTPase regulator (RPGR) gene account for >70% of X-linked retinitis pigmentosa (XLRP) and 15-20% of all inherited retinal degeneration. Gene replacement therapy for RPGR-XLRP was hampered by the relatively slow disease progression in mouse models and by difficulties in cloning the full-length RPGR-ORF15 cDNA that includes a purine-rich 3'-coding region; however, its effectiveness has recently been demonstrated in four dogs with RPGR mutations. To advance the therapy to clinical stage, we generated new stable vectors in AAV8 or AAV9 carrying mouse and human full-length RPGR-ORF15-coding sequence and conducted a comprehensive long-term dose-efficacy study in Rpgr-knockout mice. After validating their ability to produce full-length proteins that localize to photoreceptor connecting cilia, we evaluated various vector doses in mice during a 2-year study. We demonstrate that eyes treated with a single injection of mouse or human RPGR-ORF15 vector at an optimal dose maintained the expression of RPGR-ORF15 throughout the study duration and exhibited higher electroretinogram amplitude, thicker photoreceptor layer and better targeting of opsins to outer segments compared with sham-treated eyes. Furthermore, mice that received treatment at an advanced age also showed remarkable preservation of retinal structure and function. Retinal toxicity was observed at high vector doses, highlighting the importance of careful dose optimization in future clinical experiments. Our long-term dose-efficacy study should facilitate the design of human trials with human RPGR-ORF15 vector as a clinical candidate.
- Published
- 2015
20. 266. In Vivo Rod Photoreceptor Reprogramming Using AAV-Delivered CRISPR/Cas9 Rescues Retinal Degeneration
- Author
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Suddhasil Mookherjee, Suja Hiriyanna, Zhijian Wu, Xun Sun, Yasaman Ataeijannati, Wenhan Yu, Tiansen Li, Jung-Woong Kim, Lijin Dong, and Anand Swaroop
- Subjects
Retinal degeneration ,genetic structures ,Biology ,Cell fate determination ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Drug Discovery ,Retinitis pigmentosa ,Genetics ,medicine ,Molecular Biology ,Gene knockout ,Pharmacology ,Gene knockdown ,medicine.diagnostic_test ,Retinal ,medicine.disease ,Cone cell ,Cell biology ,medicine.anatomical_structure ,chemistry ,030221 ophthalmology & optometry ,Molecular Medicine ,sense organs ,030217 neurology & neurosurgery ,Electroretinography - Abstract
Retinitis pigmentosa (RP) is the most common form of inherited retinal dystrophy and the leading cause of inherited blindness, due to mutations in any of the over 60 genes/loci identified so far. The disease is characterized by an initial loss of rod photoreceptors and secondary cone cell death. Since cone photoreceptors are responsible for day time vision and visual acuity, preserving cone functions in RP patients is a priority when developing treatment strategies. NRL is a transcription factor that determines the rod photoreceptor cell fate during retinal development. Acute gene knockout of Nrl in mice was shown to reprogram adult rods into cone-like cells, rendering them resistant to effects of mutations in rod-specific genes and consequently preventing secondary cone loss (Montana CL, et al. PNAS, 2013; 110: 1732-7). With a goal to develop this approach for treatment of RP, we used adeno-associated virus (AAV)-delivered CRISPR/Cas9 for Nrl-knockdown in rod photoreceptors. AAV vectors were constructed to carry a photoreceptor-specific Cas9 nuclease expression cassette or a single-guided RNA (sgRNA) targeting Nrl or eGFP gene. The Cas9 and the sgRNA vectors were co-delivered into mice by subretinal administration. Potency of the AAV-CRISPR/Cas9 system was validated by EGFP knockdown in a mouse line with eGFP-labeled rods. Nrl knockdown was conducted in wild-type C57/Bl6 or Crxp-Nrl, a mouse line with rod-only photoreceptors. Molecular, histological and functional alterations were examined by next generation sequencing, immunoblot analysis, immunofluorescence, electron microscopy, and electroretinography (ERG). Our results showed that eGFP and Nrl were efficiently knocked down following AAV-CRISPR/Cas9 treatment. For Nrl knockdown, almost all insertions and deletions were detected in the targeted Nrl locus, and very few mutations were identified in ten potential off-target loci. A majority of the transduced rods acquired characteristics of cone photoreceptors following Nrl-CRISPR/Cas9 vector treatment, as demonstrated by reduced expression of rod-specific genes and enhanced expression of cone-specific genes, loss of the unique rod chromatin pattern, and diminished rod ERG response. Rescue of retinal degeneration was assessed in three mouse models harboring either recessive or dominant rod-specific mutations. In all three models, the Nrl-CRISPR/Cas9 vector treated eyes maintained significantly better photoreceptor viability and cone function than control eyes, as revealed by remarkably thicker photoreceptor layer, higher cone cell number, greater cone ERG amplitude and better optomotor behavior. In conclusion, AAV-CRISPR-mediated Nrl gene knockdown can efficiently reprogram rods into cone-like photoreceptors and prevent secondary cone death in retinal degeneration, which could be developed into a viable treatment for RP in humans.
- Published
- 2016
21. Phenotype correction of hemophilia A mice by spliceosome-mediated RNA trans-splicing
- Author
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Mariano A. Garcia-Blanco, Christopher E. Walsh, Suja Hiriyanna, Lloyd G. Mitchell, Hengjun Chao, Robert C Bartel, and S. Gary Mansfield
- Subjects
congenital, hereditary, and neonatal diseases and abnormalities ,Messenger RNA ,Spliceosome ,Genetic enhancement ,Mutant ,RNA ,General Medicine ,RNA repair ,Biology ,Molecular biology ,General Biochemistry, Genetics and Molecular Biology ,hemic and lymphatic diseases ,Knockout mouse ,Gene - Abstract
Conventional gene therapy of hemophilia A relies on the transfer of factor VIII (FVIII; encoded by the F8 gene) cDNA. We carried out spliceosome-mediated RNA trans-splicing (SMaRT) to repair mutant FVIII mRNA. A pre-trans-splicing molecule (PTM) corrected endogenous FVIII mRNA in F8 knockout mice with the hemophilia A phenotype, producing sufficient functional FVIII to correct the hemophilia A phenotype. This is the first description of phenotypic correction of a genetic defect by RNA repair in a knockout animal model. Our results indicate the feasibility of using SMaRT to repair RNA for the treatment of genetic diseases.
- Published
- 2003
22. Retinal Histopathology of an XLRP Carrier with a Mutation in the RPGR Exon ORF15
- Author
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Sinoj K John, Anand Swaroop, Julie E. Smith, Ann H. Milam, Debra K. Breuer, Beverly M. Yashar, Gustavo D. Aguirre, and Suja Hiriyanna
- Subjects
Retinal degeneration ,Genetics ,Pathology ,medicine.medical_specialty ,Retina ,Retinal pigment epithelium ,genetic structures ,Retinal ,Biology ,medicine.disease ,eye diseases ,Sensory Systems ,Cellular and Molecular Neuroscience ,Ophthalmology ,Exon ,chemistry.chemical_compound ,Atrophy ,medicine.anatomical_structure ,chemistry ,Retinitis pigmentosa ,medicine ,sense organs ,X chromosome - Abstract
X-linked retinitis pigmentosa comprises the severe forms of RP, with early onset of night blindness, rapid constriction of visual fields and eventual loss of central acuity. Of the five distinct XLRP loci identified on the X chromosome, mutations have been found only in the RP2 and RPGR genes. Of these, mutations in RPGR are more common, particularly in a mutational hot spot that was identified in the newly discovered exon ORF15. We report on an extended family with a microdeletion in RPGR exon ORF15 and the retinal histopathology of a female carrier of this mutation. We found a 1bp deletion at position 632 in exon ORF15 in affected members of family XLRP-319. This mutation alters the reading frame of the predicted RPGR protein, resulting in a premature stop codon. The mutation segregated with disease in three generations of the family and was associated with severe early onset retinal disease in affected men. The retina from a 75 year old carrier female donor had slight photoreceptor loss in the less diseased areas. More severe atrophy with retinal pigment epithelium (RPE) migration was present in areas of the mid- and far periphery. By immunocytochemistry, loss of rhodopsin labelling in rods was found in the areas of focal atrophy and loss of uniform cone spacing was apparent even in well preserved regions. Small multifocal areas of outer retinal degeneration were present in the better preserved regions of the eye. In these foci, rod and cone loss did not coincide. The dissociation of rod and cone degeneration in areas of focal disease is consistent with random X-inactivation early in embryonic development and the occurrence of distinct patterns of radial (rod) and tangential (cone) dispersion during clonal expansion early in photoreceptor differentiation.
- Published
- 2002
23. A Comprehensive Mutation Analysis of RP2 and RPGR in a North American Cohort of Families with X-Linked Retinitis Pigmentosa
- Author
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Bersabell Asaye, E. Filippova, Robert H. Lyons, John R. Heckenlively, Alan J. Mears, Maria A. Musarella, Paul A. Sieving, Debra K. Breuer, Alessandro Iannaccone, Raf Vervoort, Samuel G. Jacobson, Ceren Acar, Patricia G. Wheeler, Dennis R. Hoffman, Gerald A. Fishman, Alan F. Wright, David G. Birch, Anand Swaroop, Suja Hiriyanna, Beverly M. Yashar, and Ian R. MacDonald
- Subjects
Male ,Proband ,X Chromosome ,Genotype ,Genetic Linkage ,DNA Mutational Analysis ,Biology ,medicine.disease_cause ,Cohort Studies ,Open Reading Frames ,Exon ,GTP-Binding Proteins ,Genetic linkage ,Retinitis pigmentosa ,Genetics ,medicine ,Humans ,Genetics(clinical) ,Eye Proteins ,Genetics (clinical) ,X chromosome ,Mutation ,Polymorphism, Genetic ,Genetic heterogeneity ,Intracellular Signaling Peptides and Proteins ,Genetic Variation ,Membrane Proteins ,Proteins ,Exons ,Articles ,Retinitis pigmentosa GTPase regulator ,medicine.disease ,eye diseases ,Phenotype ,North America ,Carrier Proteins ,Retinitis Pigmentosa - Abstract
X-linked retinitis pigmentosa (XLRP) is a clinically and genetically heterogeneous degenerative disease of the retina. At least five loci have been mapped for XLRP; of these, RP2 and RP3 account for 10%-20% and 70%-90% of genetically identifiable disease, respectively. However, mutations in the respective genes, RP2 and RPGR, were detected in only 10% and 20% of families with XLRP. Mutations in an alternatively spliced RPGR exon, ORF15, have recently been shown to account for 60% of XLRP in a European cohort of 47 families. We have performed, in a North American cohort of 234 families with RP, a comprehensive screen of the RP2 and RPGR (including ORF15) genes and their 5' upstream regions. Of these families, 91 (39%) show definitive X-linked inheritance, an additional 88 (38%) reveal a pattern consistent with X-linked disease, and the remaining 55 (23%) are simplex male patients with RP who had an early onset and/or severe disease. In agreement with the previous studies, we show that mutations in the RP2 gene and in the original 19 RPGR exons are detected in10% and approximately 20% of XLRP probands, respectively. Our studies have revealed RPGR-ORF15 mutations in an additional 30% of 91 well-documented families with X-linked recessive inheritance and in 22% of the total 234 probands analyzed. We suggest that mutations in an as-yet-uncharacterized RPGR exon(s), intronic changes, or another gene in the region might be responsible for the disease in the remainder of this North American cohort. We also discuss the implications of our studies for genetic diagnosis, genotype-phenotype correlations, and gene-based therapy.
- Published
- 2002
24. Remapping of the RP15 Locus for X-Linked Cone-Rod Degeneration to Xp11.4-p21.1, and Identification of a De Novo Insertion in the RPGR Exon ORF15
- Author
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Stephen P. Daiger, Suja Hiriyanna, Stacey Fahrner, Beverly M. Yashar, Raf Vervoort, Alan J. Mears, Alan F. Wright, John R. Heckenlively, Anand Swaroop, Linn Gieser, and Paul A. Sieving
- Subjects
Adult ,Male ,X Chromosome ,genetic structures ,Genetic Linkage ,Molecular Sequence Data ,Locus (genetics) ,Biology ,Open Reading Frames ,Exon ,Gene mapping ,Genetic linkage ,Report ,Genotype ,Retinitis pigmentosa ,Genetics ,medicine ,Humans ,Genetics(clinical) ,Amino Acid Sequence ,Genetics (clinical) ,X chromosome ,Recombination, Genetic ,Base Sequence ,Chromosome Mapping ,Exons ,Retinitis pigmentosa GTPase regulator ,medicine.disease ,eye diseases ,Pedigree ,Phenotype ,Haplotypes ,Mutation ,Female ,sense organs ,Retinitis Pigmentosa - Abstract
X-linked forms of retinitis pigmentosa (XLRP) are among the most severe, because of their early onset, often leading to significant vision loss before the 4th decade. Previously, the RP15 locus was assigned to Xp22, by linkage analysis of a single pedigree with “X-linked dominant cone-rod degeneration.” After clinical reevaluation of a female in this pedigree identified her as affected, we remapped the disease to a 19.5-cM interval (DXS1219–DXS993) at Xp11.4-p21.1. This new interval overlapped both RP3 (RPGR) and COD1. Sequencing of the previously published exons of RPGR revealed no mutations, but a de novo insertion was detected in the new RPGR exon, ORF15. The identification of an RPGR mutation in a family with a severe form of cone and rod degeneration suggests that RPGR mutations may encompass a broader phenotypic spectrum than has previously been recognized in “typical” retinitis pigmentosa.
- Published
- 2000
25. Rd9 is a naturally occurring mouse model of a common form of retinitis pigmentosa caused by mutations in RPGR-ORF15
- Author
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Zhijian Wu, John R. Heckenlively, Hemant Khanna, Bo Chang, Anand Swaroop, Debra A. Thompson, Tiansen Li, G. Grahek, Lin Jia, Mohammad Othman, Jacob Nellissery, Suja Hiriyanna, Naheed W. Khan, and Peter Colosi
- Subjects
Male ,Retinal degeneration ,Opsin ,Mouse ,genetic structures ,lcsh:Medicine ,Mice ,chemistry.chemical_compound ,0302 clinical medicine ,lcsh:Science ,Genetics ,0303 health sciences ,Multidisciplinary ,medicine.diagnostic_test ,Reverse Transcriptase Polymerase Chain Reaction ,Retinal Degeneration ,Animal Models ,Exons ,Retinitis pigmentosa GTPase regulator ,Immunohistochemistry ,medicine.anatomical_structure ,Medicine ,Retinal Disorders ,Female ,Transducin ,Retinitis Pigmentosa ,Research Article ,Rhodopsin ,Mice, 129 Strain ,Immunoblotting ,Molecular Sequence Data ,Biology ,Retina ,03 medical and health sciences ,Model Organisms ,Sequence Homology, Nucleic Acid ,Retinitis pigmentosa ,Electroretinography ,medicine ,Animals ,Humans ,Amino Acid Sequence ,Eye Proteins ,Outer nuclear layer ,030304 developmental biology ,Clinical Genetics ,Base Sequence ,Sequence Homology, Amino Acid ,lcsh:R ,Human Genetics ,Retinal ,X-Linked ,medicine.disease ,Molecular biology ,Mice, Mutant Strains ,eye diseases ,Mice, Inbred C57BL ,Ophthalmology ,Disease Models, Animal ,chemistry ,Genetics of Disease ,Mutation ,030221 ophthalmology & optometry ,lcsh:Q ,sense organs ,Carrier Proteins ,Animal Genetics - Abstract
Animal models of human disease are an invaluable component of studies aimed at understanding disease pathogenesis and therapeutic possibilities. Mutations in the gene encoding retinitis pigmentosa GTPase regulator (RPGR) are the most common cause of X-linked retinitis pigmentosa (XLRP) and are estimated to cause 20% of all retinal dystrophy cases. A majority of RPGR mutations are present in ORF15, the purine-rich terminal exon of the predominant splice-variant expressed in retina. Here we describe the genetic and phenotypic characterization of the retinal degeneration 9 (Rd9) strain of mice, a naturally occurring animal model of XLRP. Rd9 mice were found to carry a 32-base-pair duplication within ORF15 that causes a shift in the reading frame that introduces a premature-stop codon. Rpgr ORF15 transcripts, but not protein, were detected in retinas from Rd9/Y male mice that exhibited retinal pathology, including pigment loss and slowly progressing decrease in outer nuclear layer thickness. The levels of rhodopsin and transducin in rod outer segments were also decreased, and M-cone opsin appeared mislocalized within cone photoreceptors. In addition, electroretinogram (ERG) a- and b-wave amplitudes of both Rd9/Y male and Rd9/Rd9 female mice showed moderate gradual reduction that continued to 24 months of age. The presence of multiple retinal features that correlate with findings in individuals with XLRP identifies Rd9 as a valuable model for use in gaining insight into ORF15-associated disease progression and pathogenesis, as well as accelerating the development and testing of therapeutic strategies for this common form of retinal dystrophy.
- Published
- 2012
26. Potent immune responses and in vitro pro-inflammatory cytokine suppression by a novel adenovirus vaccine vector based on rare human serotype 28
- Author
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Megan Fultz, C. Richter King, Suja Hiriyanna, Jessica Bonnell, Jason G. D. Gall, Cassandra Nyberg-Hoffman, Christoph Kahl, and Ping Chen
- Subjects
Serotype ,Adult ,Male ,Adolescent ,T cell ,viruses ,Genetic Vectors ,Biology ,CD8-Positive T-Lymphocytes ,medicine.disease_cause ,Article ,Cell Line ,Membrane Cofactor Protein ,Mice ,Young Adult ,Immune system ,Antigen ,Neutralization Tests ,Seroepidemiologic Studies ,medicine ,Animals ,Humans ,Aged ,Immunity, Cellular ,Mice, Inbred BALB C ,Cytokine Suppression ,General Veterinary ,General Immunology and Microbiology ,Immunogenicity ,Adenoviruses, Human ,Public Health, Environmental and Occupational Health ,Viral Vaccines ,Middle Aged ,Virology ,United States ,Adenovirus vaccine ,Adenoviridae ,Infectious Diseases ,medicine.anatomical_structure ,Immunology ,Antibody Formation ,Leukocytes, Mononuclear ,Molecular Medicine ,Cytokines ,Female ,medicine.drug - Abstract
Adenovirus vaccine vectors derived from rare human serotypes have been shown to be less potent than serotype 5 (Ad5) at inducing immune responses to encoded antigens. To identify highly immunogenic adenovirus vectors, we assessed pro-inflammatory cytokine expression, binding to the CD46 receptor, and immunogenicity. Species D adenoviruses uniquely suppressed pro-inflammatory cytokines and induced high levels of type I interferon. Thus, it was unexpected that a vector derived from a representative serotype, Ad28, induced significantly higher transgene-specific T cell responses than an Ad35 vector. Prime-boost regimens with Ad28, Ad35, Ad14, or Ad5 significantly boosted T cell and antibody responses. The seroprevalence of Ad28 was confirmed to be
- Published
- 2010
27. 714. Gene Therapy Rescues Cone Function and Viability in an Rp2 Knockout Mouse Model for X-Linked Retinitis Pigmentosa Over a Wide Dose Range and a Broad Therapeutic Time Window
- Author
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Peter Colosi, Tiansen Li, Suja Hiriyanna, Kayleigh Kaneshiro, Hemant Khanna, Haohua Qian, Zhijian Wu, Anand Swaroop, and Suddhasil Mookherjee
- Subjects
Pharmacology ,Retinal degeneration ,medicine.medical_specialty ,Visual acuity ,genetic structures ,medicine.diagnostic_test ,Genetic enhancement ,Anatomy ,Biology ,medicine.disease ,eye diseases ,Ophthalmology ,Drug Discovery ,Retinitis pigmentosa ,Knockout mouse ,Genetics ,medicine ,Molecular Medicine ,sense organs ,medicine.symptom ,Molecular Biology ,Erg ,Retinal Dystrophies ,Electroretinography - Abstract
Retinitis pigmentosa (RP) refers to a diverse group of inherited retinal dystrophies characterized by a progressive loss of photoreceptors, resulting in vision impairment or blindness. The prevalence of the disease is 1 in 3,500 to 4,000 worldwide. X-linked RP (XLRP) accounts for ~15% of all RP cases and is one of the most severe forms of retinal degeneration. About 10% of the total XLRP cases are caused by mutations in the RP2 gene. A recently developed Rp2 knockout (Rp2-KO) mouse model exhibits early onset, progressive cone photoreceptor dysfunction and degeneration, mimicking early cone involvement in patients with RP2 mutations. To develop gene therapy for the disease, we generated and characterized a self-complementary AAV8 vector carrying the human RP2 expression cassette (AAV8-scRK-RP2), and examined its efficacy in the Rp2-KO mice. Our initial results demonstrated successful cone function rescue following subretinal administration of the vector to the mice (ASGCT 2014, poster#122). In the present study, we continued to examine the long term safety and efficacy of the vector using a broader dose range. Four to six week-old Rp2-KO mice received unilateral subretinal injections of the vector with doses ranging from 5×107 to 1×109 vector genomes (vg)/eye. The fellow eyes were injected with vehicle as controls. Retinal function was assessed by electroretinography (ERG) and optomotor test up to 18 months of age. Our results revealed significantly higher amplitude and faster recovery of cone ERG in vector-treated eyes than control eyes in almost all dose groups through the duration of monitoring starting from 4 months of age. In addition, remarkably better visual acuity was observed in the vector-treated eyes at 18 months of age. The treatment also reversed M-opsin mis-localization, restored cone PDE expression and maintained cone viability. Interestingly, this rescue was achieved in the Rp2-KO mice even when treated at 10-month of age, suggesting the potential benefit of the treatment to older patients. Retinal toxicity was only observed in mice receiving the highest vector dose (1×109 vg/eye), as reflected by reduced rod ERG amplitude and thinner photoreceptor layer. We conclude that our RP2 AAV vector can rescue cone function and viability over a wide dose range and a broad therapeutic time window, which paves the way for future clinical studies.
- Published
- 2015
28. Clinical studies of X-linked retinitis pigmentosa in three Swedish families with newly identified mutations in the RP2 and RPGR-ORF15 genes
- Author
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Suja Hiriyanna, Sten Andréasson, Louise Eksandh, E. Filippova, Beverly M. Yashar, Anand Swaroop, Christina Frennesson, Debra K. Breuer, and Vesna Ponjavic
- Subjects
Retinal degeneration ,Adult ,Male ,DNA Mutational Analysis ,Visual Acuity ,Disease ,Biology ,chemistry.chemical_compound ,Exon ,Open Reading Frames ,GTP-Binding Proteins ,Retinitis pigmentosa ,medicine ,Electroretinography ,Humans ,Eye Proteins ,Gene ,Genetics (clinical) ,Aged ,Genetics ,Sweden ,Chromosomes, Human, X ,Intracellular Signaling Peptides and Proteins ,Membrane Proteins ,Proteins ,Retinal ,Genetic Diseases, X-Linked ,Exons ,Middle Aged ,medicine.disease ,Phenotype ,eye diseases ,Pedigree ,Ophthalmology ,chemistry ,Pediatrics, Perinatology and Child Health ,Mutation ,Female ,X-linked retinitis pigmentosa ,Visual Fields ,Carrier Proteins ,Retinitis Pigmentosa - Abstract
PURPOSE: To describe new disease-causing RP2 and RPGR-ORF15 mutations and their corresponding clinical phenotypes in Swedish families with X-linked retinitis pigmentosa (XLRP) and to establish genotype-phenotype correlations by studying the clinical spectrum of disease in families with a known molecular defect. METHODS: Seventeen unrelated families with RP and an apparent X-linked pattern of disease inheritance were identified from the Swedish RP registry and screened for mutations in the RP2 and RPGR (for the RP3 disease) genes. These families had been previously screened for the RPGR exons 1-19, and disease-causing mutations were identified in four of them. In the remaining 13 families, we sequenced the RP2 gene and the newly discovered RPGR-ORF exon. Detailed clinical evaluations were then obtained from individuals in the three families with identified mutations. RESULTS: Mutations in RP2 and RPGR-ORF15 were identified in three of the 13 families. Clinical evaluations of affected males and carrier females demonstrated varying degrees of retinal dysfunction and visual handicap, with early onset and severe disease in the families with mutations in the ORF15 exon of the RPGR gene. CONCLUSIONS: A total of seven mutations in the RP2 and RPGR genes have been discovered so far in Swedish XLRP families. All affected individuals express a severe form of retinal degeneration with visual handicap early in life, although the degree of retinal dysfunction varies both in hemizygous male patients and in heterozygous carrier females. Retinal disease phenotypes in patients with mutations in the RPGR-ORF15 were more severe than in patients with mutations in RP2 or other regions of the RPGR. (Less)
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- 2003
29. X-Linked Retinitis Pigmentosa: Current Status
- Author
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Alan F. Wright, Anand Swaroop, Maria A. Musarella, Paul A. Sieving, Alan J. Mears, Beverly M. Yashar, Samuel G. Jacobson, Debra K. Breuer, Elena Redolfi, Ileana Zucchi, John R. Heckenlively, Gerald A. Fishman, Sten Andréasson, David G. Birch, Suja Hiriyanna, Maurizio Affer, and Dennis R. Hoffman
- Subjects
Progressive retinal atrophy ,Genetics ,education.field_of_study ,Positional cloning ,Genetic heterogeneity ,Population ,Biology ,medicine.disease ,eye diseases ,Genetic linkage ,Retinitis pigmentosa ,medicine ,Age of onset ,education ,Gene - Abstract
Retinitis pigmentosa (RP) is a clinically and genetically heterogeneous group of retinal degenerative diseases, characterized by nightblindness, progressive restriction of the visual field and pigmentary retinopathy.1 At least 28 different genetic loci have been mapped for autosomal dominant, autosomal recessive, and X-linked forms of RP. [http://www.sph.uth.tmc.edu/Retnet/home.htm] The X-linked RP (XLRP) subtype is the most severe, with an early age of onset and more rapid progression, accounting for 10 to 20% of RP families.2,3 XLRP is also genetically heterogeneous with at least 5 mapped loci: RP2, RP3, RP6, RP23 and RP24, as schematically depicted in Figure 1. By linkage analysis, RP2 is predicted to account for 10–20% of XLRP and RP3 for 70–90%,4–6 depending on the population. Genes for these two major loci have now been cloned. Our laboratory has been involved in the mutational screening and functional analysis of the two identified XLRP genes (RPGR and RP2), as well as the positional cloning of two other XLRP loci (RP6 and RP24). This report summarizes these efforts as well as the current standing of XLRP research.
- Published
- 2001
30. Protein-Truncation Mutations in the RP2 Gene in a North American Cohort of Families with X-Linked Retinitis Pigmentosa
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Cynthia S. Chen, Denise Yan, Alan J. Mears, Samuel G. Jacobson, Ricardo Fujita, Linn Gieser, Stacey Fahrner, Paul A. Sieving, Anand Swaroop, and Suja Hiriyanna
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Male ,Letter ,Genetic Linkage ,Newfoundland and Labrador ,Molecular Sequence Data ,Library science ,Retina ,X chromosome ,03 medical and health sciences ,0302 clinical medicine ,Eye disease ,GTP-Binding Proteins ,Photoreceptor degeneration ,medicine ,Mutation screening ,Genetics ,Humans ,Point Mutation ,Base sequence ,In patient ,Genetics(clinical) ,Sociology ,Amino Acid Sequence ,Eye Proteins ,Frameshift Mutation ,Genetics (clinical) ,Vice president ,030304 developmental biology ,0303 health sciences ,Patient registry ,Blindness ,Base Sequence ,Intracellular Signaling Peptides and Proteins ,Protein Truncation ,Membrane Proteins ,Proteins ,Sequence Analysis, DNA ,medicine.disease ,Founder Effect ,3. Good health ,Genetic diagnosis ,030221 ophthalmology & optometry ,X-linked retinitis pigmentosa ,Gene Deletion ,Retinitis Pigmentosa - Abstract
We thank Drs. Sten Andreasson, David Birch, Nancy Carson, Bernie Chodirker, Mark Evans, Gerald Fishman, John Heckenlively, Dennis Hoffman, Maria Musarella, and Beth Spriggs and Mr. Eric L. Krivchenia for some of the patient samples that were included in the mutation screening. We acknowledge the assistance of Dr. Wolfgang Berger for providing the RP2 primer sequences. We thank Dr. Monika Buraczynska for organization of the patient registry; Dr. Radha Ayyagari for discussions; Dr. Beverly Yashar for counseling; Ms. Cara Coats for assistance in patient collection; Mr. Jason Cook, Ms. Patricia Forsythe, and Ms. Eve Bingham for technical assistance; and Ms. D. Giebel for secretarial assistance. This research was supported by National Institutes of Health (NIH) grants EY05627, EY06094, and EY07961 and by grants from the Foundation Fighting Blindness, the Chatlos Foundation, the Kirby Foundation, the Mackall Trust, and Research to Prevent Blindness. We also acknowledge NIH grants EY07003 (core) and M01-RR00042 (General Clinical Research Center) and a shared equipment grant from the Office of Vice President for Research (University of Michigan). A.S. is recipient of a Lew R. Wasserman Merit Award, and P.A.S., a Senior Scientific Investigator Award, both from Research to Prevent Blindness.
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31. Ocular and systemic safety of a recombinant AAV8 vector for X-linked retinoschisis gene therapy: GLP studies in rabbits and Rs1-KO mice
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Dario Marangoni, Ronald A Bush, Yong Zeng, Lisa L Wei, Lucia Ziccardi, Camasamudram Vijayasarathy, Joshua T Bartoe, Kiran Palyada, Maria Santos, Suja Hiriyanna, Zhijian Wu, Peter Colosi, and Paul A Sieving
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