Biology subjects

Visel, M.

Publications and source records attributed to Visel, M..

6 recordsLinked to original sources

bioRxiv2024

Truncated Complement Factor H Y402 Gene Therapy Cures C3 Glomerulonephritis

Patients with both age-related macular degeneration (AMD) and C3 glomerulonephritis (C3G) are challenged by the absence of effective therapies to reverse and eliminate their disease burden. Capitalizing on complement dysregulation as both a significant risk factor for AMD and the known pathophysiology of C3G, we investigated the potential for adeno-associated virus (AAV) delivery of complement factor H (CFH) to rescue C3G in a Cfh-/- mouse model of C3G. While past efforts to treat C3G using exogenous human CFH resulted in limited success before immune rejection led to a foreign protein response, our findings demonstrate the capacity for long-term AAV-mediated delivery of truncated CFH (tCFH) to restore inhibition of the alternative pathway of complement and ultimately reverse C3G without immune rejection. Comparing results from the administration of several tCFH vectors also revealed significant differences in their relative efficiency and efficacy. These discoveries pave the way for subsequent development of AAV-mediated tCFH replacement therapy for patients with C3G, while simultaneously demonstrating proof of concept for a parallel AAV-mediated tCFH gene augmentation therapy for patients with AMD.

pathology↗

bioRxiv2022

Restoration of high-sensitivity patterned vision in motion with an engineered light-gated G protein-coupled receptor

Inherited retinal degenerations (IRDs) result in blindness due to apoptotic cell death of rods and cones, but spare other retinal neurons, providing a potential that delivery of a light-activated signaling protein to surviving neurons may restore vision. We previously demonstrated that aspects of vision could be restored by introduction into surviving cells of a G protein-coupled receptor for glutamate (mGluR) bearing a tethered photoswitchable agonist. However, this system, containing one photoswitchable agonist per glutamate binding site, yielded low sensitivity, responding only to visual stimuli at the intensity of bright outdoor light, similar to channelrhodopsins. To increase sensitivity, we designed a multi-branched photoswitch, bearing four light-activatable glutamates for each glutamate binding site. When tethered to a modified mGluR2 expressed in retinal ganglion cells via intravitreal AAV gene delivery, this photoswitch boosted sensitivity by ~100-fold compared to the unbranched (single photo-ligand) photoswitch. This improvement in sensitivity enabled an IRD mouse model (rd1) to perform visually-guided object recognition under incidental room light and pattern recognition using standard LCD computer displays. The restored line pattern differentiation approached the acuity reported for normal mouse vision. Pattern recognition functioned as well as wildtype vision with line patterns moving at speeds of up to 36{degrees}/s. In summary, this two-component chemical-optogenetic approach combines high sensitivity and high acuity with superior motion vision, and, unlike optogenetic gene therapy, can be adjusted for dose, upgraded, as new photoswitches are developed, and discontinued at will.

neuroscience↗

bioRxiv2021

Targeting ON-bipolar cells by AAV gene therapy stably reverses LRIT3-congenital stationary night blindness

AAV gene therapies aimed at curing inherited retinal diseases to date have typically focused on photoreceptors and retinal pigmented epithelia within the relatively accessible outer retina. However, therapeutic targeting in diseases such as congenital stationary night blindness (CSNB) that involve defects in ON-bipolar cells (ON-BCs) within the mid-retina has been challenged by the relative inaccessibility of the target cell in intact retinas, the limited transduction efficiency of these cells by existing AAV serotypes, poor availability of established ON-BC-specific promoters, and absence of appropriate patient-relevant large animal models. Here, we demonstrate safe and effective ON-BC targeting by AAV gene therapy in a recently characterized naturally-occurring canine model of CSNB, LRIT3-CSNB. To effectively target ON-BCs, new AAV capsid variants with ON-BC tropism and ON-BC specific modified GRM6 promoters were adopted to ensure cell-specific transgene expression. Notably, subretinal injection of one vector, AAVK9#4-shGRM6-cLRIT3-WPRE, significantly recovered rod-derived b-wave in all treated eyes (6/6) of adult dogs injected at 1-3 years of age. The robust therapeutic effect was evident 7 weeks post-injection and was sustained for at least 1 year in all treated eyes. Scotopic vision was significantly improved in treated eyes based on visually-guided obstacle course navigation. Restoration of LRIT3 signals was confirmed by immunohistochemistry. Thus, we report on the first ON-BC functional rescue in a large animal model using a novel AAV capsid variant and modified promoter construct optimized for ON-BC specificity, thereby establishing both proof-of-concept and a novel translational platform for treatment of CSNB in patients with defects in photoreceptor-to-bipolar signaling. SignificanceCanine models of inherited retinal diseases have informed and advanced AAV gene therapies targeting specific cells, including photoreceptors in the outer retina, for treatment of blinding diseases in human patients. However, therapeutic targeting of diseases such as congenital stationary night blindness (CSNB) that exhibit defects in ON-bipolar cells (ON-BCs) of the mid-retina remains under-developed. Using a new LRIT3 mutant canine model of CSNB exhibiting ON-BC dysfunction, we tested the ability of a cell-specific AAV capsid and promotor to specifically target ON-BCs for gene delivery. Notably, subretinal injection of AAV-LRIT3 vector demonstrated safety and efficacy with robust and stable rescue of ERG signals and night vision up to 1 year, paving the way for clinical trials in CNSB patients.

neuroscience↗

bioRxiv2021

Outcomes of Progranulin Gene Therapy in the Retina are Dependent on Time of Delivery

Neuronal ceroid lipofuscinosis (NCL) is a family of neurodegenerative diseases caused by mutations to genes related to lysosomal function. One variant, CNL11, is caused by mutations to the gene encoding the protein progranulin. Primarily secreted by microglia, progranulin regulates neuronal lysosomal function once endocytosed. Absence of progranulin causes cerebellar atrophy, seizures, ataxia, dementia and vision loss. As progranulin gene therapies targeting the brain are developed, it is also advantageous to focus on the retina, as its characteristics are beneficial for gene therapy development: the retina is easily visible through direct imaging, can be assessed through quantitative methods in vivo, requires smaller amounts of AAV and AAV can be administered via a less invasive surgery. In this study we characterize the retinal degeneration in a progranulin knockout mouse model of CLN11 and study the effects of gene replacement at different time points. All mice heterologously expressing progranulin showed reduction in lipofuscin deposits and microglia infiltration. While mice that receive systemic AAV9.2YF-scCAG-PGRN at post-natal day 3 or 4 show a reduction in retina thinning, mice injected intravitreally at months 1 and 6 with 7m8-scCAG-PGRN show no improvement, and mice injected at 12 months of age show increased retinal thinning in comparison to their controls. Thus, delivery of progranulin proves to be time-sensitive, requiring early administration for optimal therapeutic benefit.

neuroscience↗

bioRxiv2020

scAAVengr: Single-cell transcriptome-based quantification of engineered AAVs in non-human primate retina

Adeno-associated virus (AAV)-mediated gene therapies are rapidly advancing to the clinic, and AAV engineering has resulted in vectors with increased ability to deliver therapeutic genes. Although the choice of vector is critical, quantitative comparison of AAVs, especially in large animals, remains challenging. Here, we developed an efficient single-cell AAV engineering pipeline (scAAVengr) to quantify efficiency of AAV-mediated gene expression across all cell types. scAAVengr allows for definitive, head-to-head comparison of vectors in the same animal. To demonstrate proof-of-concept for the scAAVengr workflow, we quantified - with cell-type resolution - the abilities of naturally occurring and newly engineered AAVs to mediate gene expression in primate retina following intravitreal injection. A top performing variant, K912, was used to deliver SaCas9 and edit the rhodopsin gene in macaque retina, resulting in editing efficiency similar to infection rates detected by the scAAVengr workflow. These results validate scAAVengr as a powerful method for development of AAV vectors.

bioengineering↗

bioRxiv2019

In vivo directed evolution of AAV in the primate retina

Efficient AAV-mediated gene delivery remains a significant obstacle to effective retinal gene therapies. Here, we apply directed evolution - guided by deep sequencing and followed by direct in vivo secondary selection of high-performing vectors with a GFP-barcoded library - to create AAV viral capsids with new capabilities to deliver genes to the outer retina in primates. A replication incompetent library, produced via providing rep in trans, was created to mitigate risk of AAV propagation. Six rounds of in vivo selection with this library in primates - involving intravitreal library administration, recovery of genomes from outer retina, and extensive next generation sequencing of each round - resulted in vectors with redirected tropism to the outer retina and increased gene delivery efficiency to retinal cells. These new viral vectors expand the toolbox of vectors available for primate retina, and may enable less invasive delivery of therapeutic genes to patients, potentially offering retina-wide infection at a similar dosage to vectors currently in clinical use.

neuroscience↗