John G. Flannery

Professor of Neurobiology

Lab Homepage: http://mcb.berkeley.edu/labs/flannery/

Full Directory Information

Research Interests

Viral Vectors for Gene Therapy of Inherited Retinal Degenerations

Gene therapy has vast potential for treating and potentially curing a number of retinal diseases including glaucoma, age-related macular degeneration, and inherited photoreceptor diseases.  However, gene delivery technologies require significant improvements in cellular targeting, efficiency, and safety before promising findings in animal studies are translated to the clinic.  In particular, for retinal gene therapy it would be highly advantageous to transduce a single cell type that spans the entire retina after an intravitreal injection of a gene delivery vehicle for the subsequent secretion of a general neuroprotective factor throughout the retina.  Unfortunately, there is no vector capable of efficiently infecting the cell type that meets these needs, Muller cells.  Vectors based on adeno-associated virus (AAV) have proven themselves to be highly promising in numerous retinal disease models, but they are also incapable of Muller cell infection.  We have developed novel lentiviral vectors with new properties, including altered receptor binding, which are capable of efficient Muller cell transduction.  In parallel, the basic mechanisms of AAV transduction of Muller cells will be explored in order to develop new AAV pseudotypes capable of Muller cell transduction.  The novel approaches developed in this work will have general impact for the molecular engineering of enhanced viral gene delivery vehicles, and future work will focus on testing these vectors in an animal model of retinal disease.

Current Projects

The retina is a complex tissue in the back of the eye that contains the rod and cone photoreceptor cells. The photoreceptors connect to a network of retinal interneurons. The adjacent retinal pigment epithelium (RPE) supports many of the retina's metabolic functions. The retina is susceptible to a number of blinding diseases, such as age-related macular degeneration, diabetic retinopathy and other inherited retinal degenerations. The inherited retinal degenerations are typified by retinitis pigmentosa (RP), which results in blindness from destruction of photoreceptor cells, and the RPE. This group of conditions affects approximately 100,000 people in the United States. To date, more than 130 genes causing inherited retinopathies in humans have been identified. This makes it possible to identify the cause of RP in approximately 50 percent of patients and the cause of Usher syndrome in 75 percent of patients.

Gene identifications in humans have allowed us to examine the biochemical pathways in these diseases. In addition, gene identification in patients permits us to identify naturally occurring animal models or create new transgenic or knockout animal models with retinal degeneration due to defects in the gene homologs. In particular, we have the examined retinal degeneration in the naturally arising rd mouse strains (defects in the b-subunit of phosphodiesterase). We have also developed transgenic rats, expressing dominant rhodopsin mutations. Most recently we have developed a knockout mouse model of Usher syndrome type 3, which caused progressive blindness and deafness in patients. These animal models are the subject of study to determine the pathophysiological mechanisms whereby these gene defects lead to photoreceptor degeneration and hopefully will lead to pilot studies of novel therapies for retinal degeneration.

Development of effective treatments for retinal diseases.

With increasing insight into the molecular etiologies of several inherited retinal and macular dystrophies, studies from ours and many laboratories have defined several promising therapeutic strategies.

Current projects in our lab involve development of retinal cell specific viral vectors based upon lentivirus and adeno-associated viruses. In previous work, we have demonstrated significant slowing of photoreceptor degeneration in several animal models following gene transfer of neurotrophic agents. Another promising strategy for dominantly inherited retinal diseases involves directly targeting the mutant mRNA product using ribozymes, and siRNA. For recessive null diseases, gene replacement is an option.

We find that gene therapy has vast potential for treating and potentially curing a number of inherited photoreceptor diseases. However, gene delivery technologies require significant improvements in cellular targeting, efficiency, and safety before promising findings in animal studies are translated to the clinic. In particular, for retinal gene therapy it would be highly advantageous to transduce a single cell type that spans the entire retina after an intravitreal injection of a gene delivery vehicle for the subsequent secretion of a general neuroprotective factor throughout the retina. Unfortunately, there is no vector capable of efficiently infecting the cell type that meets these needs, Müller cells. Vectors based on adeno-associated virus (AAV) have proven themselves to be highly promising in numerous retinal disease models, but they are also incapable of Müller cell infection. Recently, we have developed novel lentiviral vectors with new properties, including altered receptor binding, which are capable of efficient Müller cell transduction. In parallel, the basic mechanisms of AAV transduction of Muller cells will be explored in order to develop new AAV pseudotypes capable of Muller cell transduction. The novel approaches developed in this work will have general impact for the molecular engineering of enhanced viral gene delivery vehicles, and future work will focus on testing these vectors in an animal model of retinal disease.

 

Selected Publications

Caporale N, Kolstad KD, Lee T, Tochitsky I, Dalkara D, Trauner D et al. LiGluR Restores Visual Responses in Rodent Models of Inherited Blindness. Mol Ther 2011; 19(7): 1212-9.

Dalkara D, Kolstad KD, Guerin KI, Hoffmann NV, Visel M, Klimczak RR et al. AAV Mediated GDNF Secretion From Retinal Glia Slows Down Retinal Degeneration in a Rat Model of Retinitis Pigmentosa. Mol Ther  2011.

Vastinsalo H, Jalkanen R, Dinculescu A, Isosomppi J, Geller S, Flannery JG et al. Alternative splice variants of the USH3A gene Clarin 1 (CLRN1). Eur J Hum Genet 2011; 19(1): 30-5.

Kolstad KD, Dalkara D, Guerin K, Visel M, Hoffmann N, Schaffer DV et al. Changes in adeno-associated virus-mediated gene delivery in retinal degeneration. Hum Gene Ther 2010; 21(5): 571-8.

Klimczak RR, Koerber JT, Dalkara D, Flannery JG, Schaffer DV. A novel adeno-associated viral variant for efficient and selective intravitreal transduction of rat Muller cells. PLoS One 2009; 4(10): e7467.

Tackenberg MA, Tucker BA, Swift JS, Jiang C, Redenti S, Greenberg KP et al. Muller cell activation, proliferation and migration following laser injury. Mol Vis 2009; 15: 1886-96.

Isosomppi J, Vastinsalo H, Geller SF, Heon E, Flannery JG, Sankila EM. Disease-causing mutations in the CLRN1 gene alter normal CLRN1 protein trafficking to the plasma membrane. Mol Vis 2009; 15: 1806-18.

Geller SF, Guerin KI, Visel M, Pham A, Lee ES, Dror AA et al. CLRN1 is nonessential in the mouse retina but is required for cochlear hair cell development. PLoS Genet 2009; 5(8): e1000607.

Dalkara D, Kolstad KD, Caporale N, Visel M, Klimczak RR, Schaffer DV et al. Inner limiting membrane barriers to AAV-mediated retinal transduction from the vitreous. Mol Ther 2009; 17(12): 2096-102.

Koerber JT, Klimczak R, Jang JH, Dalkara D, Flannery JG, Schaffer DV. Molecular evolution of adeno-associated virus for enhanced glial gene delivery. Mol Ther 2009; 17(12): 2088-95.

Last Updated 2011-08-05