William A. Beltran, Ph.D. - US grants

A multi-investigator, multi-center research plan is proposed to develop and test gene-based retinal therapy in animal models (mouse and dog) for translation to patients with autosomal dominant RP caused by mutations in the rhodopsin gene (RHO). RHO mutations constitute one of the most common molecularly-identified causes of human RP, and more than 100 of them account for > 12 % of RP. The proposal has been divided into 4 aims that will: (Aim#1) develop viral vectors, promoters, knockdown constructs and replacement cDNAs, and compare the efficacy of a RHO cDNA augmentation approach, to that of an allele-independent knockdown and replacement strategy in two mouse models; (Aim #2) evaluate in a large animal model (dog) which of these strategies provides optimal rescue of rods, (Aim #3) develop outcome measures for clinical trials of gene therapy in RHO-ADRP patients, and (Aim #4) evaluate the optimal strategy and vector construct (based on results of Aims #1 and 2) in pre-clinical safety studies. Six coordinated modules (M) are described, each with a specific set of aims that contributes in a unique but complementary way to the translational studies. M1 (Vector Development) will provide AAVs carrying knockdown (siRNA, ribozymes) reagents, and resistant (hardened) RHO cDNAs. M2 (Small Animal-mouse- Therapy Studies) will test the 2 gene therapy approaches in two mouse models. M3 (Large Animal Experiemntal Support) will produce the dogs, and provide infrastructure resources for this work). M4 (Large anima I- dog - Therapy Studies) will test the 2 approaches in a naturally -occurring canine model of RHO-ADRP. M5 (Human RHO-ADRP) will identify retinal regions that can be targeted for focal retinal therapy in patients. M6 (Vector safety studies in Animals) will conduct GLP-based preclinical toxicology and biodistribution studies in small and large animals to test the safety of the optimal (lead) therapeutic vector as the essential first step fro FDA consideration of an IND for a future Phase I Clinical Trial. The research studies described in this proposal represent a continuation of a longstanding collaboration between the module scientists that already has brought retinal gene therapy for RPE65-LCA patients to a Phase I clinical trial.

DESCRIPTION (provided by applicant): A multi-investigator, multi-center research plan is proposed to develop and test gene-based retinal therapy in animal models (mouse and dog) for translation to patients with autosomal dominant RP caused by mutations in the rhodopsin gene (RHO). RHO mutations constitute one of the most common molecularly-identified causes of human RP, and more than 100 of them account for > 12 % of RP. The proposal has been divided into 4 aims that will: (Aim#1) develop viral vectors, promoters, knockdown constructs and replacement cDNAs, and compare the efficacy of a RHO cDNA augmentation approach, to that of an allele-independent knockdown and replacement strategy in two mouse models; (Aim #2) evaluate in a large animal model (dog) which of these strategies provides optimal rescue of rods, (Aim #3) develop outcome measures for clinical trials of gene therapy in RHO-ADRP patients, and (Aim #4) evaluate the optimal strategy and vector construct (based on results of Aims #1 and 2) in pre-clinical safety studies. Six coordinated modules (M) are described, each with a specific set of aims that contributes in a unique but complementary way to the translational studies. M1 (Vector Development) will provide AAVs carrying knockdown (siRNA, ribozymes) reagents, and resistant (hardened) RHO cDNAs. M2 (Small Animal-mouse- Therapy Studies) will test the 2 gene therapy approaches in two mouse models. M3 (Large Animal Experiemntal Support) will produce the dogs, and provide infrastructure resources for this work). M4 (Large anima I- dog - Therapy Studies) will test the 2 approaches in a naturally -occurring canine model of RHO-ADRP. M5 (Human RHO-ADRP) will identify retinal regions that can be targeted for focal retinal therapy in patients. M6 (Vector safety studies in Animals) will conduct GLP-based preclinical toxicology and biodistribution studies in small and large animals to test the safety of the optimal (lead) therapeutic vector as the essential first step ro FDA consideration of an IND for a future Phase I Clinical Trial. The research studies described in this proposal represent a continuation of a longstanding collaboration between the module scientists that already has brought retinal gene therapy for RPE65-LCA patients to a Phase I clinical trial.

A multi-investigator, multi-center research plan is proposed to develop and test gene-based retinal therapy in animal models (mouse and dog) for translation to patients with autosomal dominant RP caused by mutations in the rhodopsin gene (RHO). RHO mutations constitute one of the most common molecularly-identified causes of human RP, and more than 100 of them account for > 12 % of RP. The proposal has been divided into 4 aims that will: (Aim#1) develop viral vectors, promoters, knockdown constructs and replacement cDNAs, and compare the efficacy of a RHO cDNA augmentation approach, to that of an allele-independent knockdown and replacement strategy in two mouse models; (Aim #2) evaluate in a large animal model (dog) which of these strategies provides optimal rescue of rods, (Aim #3) develop outcome measures for clinical trials of gene therapy in RHO-ADRP patients, and (Aim #4) evaluate the optimal strategy and vector construct (based on results of Aims #1 and 2) in pre-clinical safety studies. Six coordinated modules (M) are described, each with a specific set of aims that contributes in a unique but complementary way to the translational studies. M1 (Vector Development) will provide AAVs carrying knockdown (siRNA, ribozymes) reagents, and resistant (hardened) RHO cDNAs. M2 (Small Animal-mouse- Therapy Studies) will test the 2 gene therapy approaches in two mouse models. M3 (Large Animal Experiemntal Support) will produce the dogs, and provide infrastructure resources for this work). M4 (Large anima I- dog - Therapy Studies) will test the 2 approaches in a naturally -occurring canine model of RHO-ADRP. M5 (Human RHO-ADRP) will identify retinal regions that can be targeted for focal retinal therapy in patients. M6 (Vector safety studies in Animals) will conduct GLP-based preclinical toxicology and biodistribution studies in small and large animals to test the safety of the optimal (lead) therapeutic vector as the essential first step fro FDA consideration of an IND for a future Phase I Clinical Trial. The research studies described in this proposal represent a continuation of a longstanding collaboration between the module scientists that already has brought retinal gene therapy for RPE65-LCA patients to a Phase I clinical trial.

Project summary . A multi-investigator, multi-center plan is proposed to develop gene-based retinal therapies for LCA- ciliopathies using the dog NPHP5 model. A subgroup of these human ciliopathies show early onset and profound congenital retinal and visual malfunction that results from abnormally developed photoreceptors (PR) that subsequently degenerate. The proposal builds on success achieved in the current grant period in moving RPGR-XLRP to a clinical trial, and recent studies in the NPHP5 model showing that a scAAV2/8- based viral vector, together with the human GRK1 promoter and human full-length NPHP5 cDNA, rescues ERG rod and cone function, and vision, for at least 1 year, but that PRs continue to degenerate, albeit at a much slower rate. This vector now serves as the benchmark test vector to assess treatment paradigms to optimize PR targeting, infectivity and therapeutic transgene expression that will result in permanent disease correction. The proposal will evaluate gene therapy in dogs having this aggressive and severe LCA- ciliopathy, and is divided into three aims that will: 1- establish the benchmark dose and disease stage treatment efficacy of the test vector; 2- select the lead vector pseudotype and promoter with optimized transduction efficiency, and efficacy in targeting different PR disease stages; 3- facilitate translational studies by defining the natural history of the disease in dogs and patients, the effect of treatment in dogs, and determining the degree of PR/retinal disease still amenable to treatment. While the test platform is the NPHP5-LCA dog model, the therapeutic questions addressed apply broadly to other LCA-ciliopathies. Four coordinated groups [a.k.a. modules (M)], are described that take advantage of the special expertise of each group to create a complementary and focused approach to the proposed translational studies. M1 (Large Animal Experimental) will produce the dog models, and provide infrastructure resources for the work; M2 (Large Animal Therapy) will carry out therapy studies and develop measures for outcome assessment; M3 (Non-invasive Patient and Dog Studies) will establish functional and structural disease features in the patients and model, and evaluate success of therapies in dogs using non-invasive outcome measures that correlate with ex vivo morphologic studies; M4 (Molecular Therapeutic Development) will provide therapeutic vectors. The research studies described in this proposal represents a continuation of a longstanding collaboration between the module scientists that already has brought retinal gene therapy for the treatment of patients with RPE65-LCA (Phase I clinical trial), and CNGB3-achromatopsia and RPGR-XLRP (both in final preparations for clinical trials). The University of Pennsylvania leads this collaboration with the University of Florida.

Instrumentation - Project Summary The VRC Instrumentation Core Module supports the design and construction of specialized laboratory instrumentation. It supports a machine shop, electronics shop, and their associated personnel. The module provides design consultation, construction of precision mechanical and electronic apparatus, modifications to commercial instruments to meet specialized vision research needs, and instrument repair and maintenance. The module has been used heavily by VRC Participating Investigators over the past 5 years, and we expect that this will continue.