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

2021 ◽  
Author(s):  
Emilia A. Zin ◽  
Daisy Han ◽  
Jennifer Tran ◽  
Nikolas Morisson-Welch ◽  
Meike Visel ◽  
...  
Keyword(s):  
Gene Therapy ◽  
Quantitative Methods ◽  
Vision Loss ◽  
Neuronal Ceroid Lipofuscinosis ◽  
Cerebellar Atrophy ◽  
Gene Replacement ◽  
Therapeutic Benefit ◽  
Lysosomal Function ◽  
Knockout Mouse Model

AbstractNeuronal 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.

In vivo potency testing of subretinal rAAV5.hCNGB1 gene therapy in the Cngb1 knockout mouse model of retinitis pigmentosa

2021 ◽  
Author(s):  
Johanna E Wagner ◽  
Lena Zobel ◽  
Maximilian Joachim Gerhardt ◽  
Catherine R O'Riordan ◽  
Amy Frederick ◽  
...  
Keyword(s):  
Gene Therapy ◽  
Mouse Model ◽  
Retinitis Pigmentosa ◽  
Knockout Mouse ◽  
Knockout Mouse Model

scholarly journals Progress in Gene Editing Tools and Their Potential for Correcting Mutations Underlying Hearing and Vision Loss

2021 ◽  
Vol 3 ◽  
Author(s):  
Catherine Botto ◽  
Deniz Dalkara ◽  
Aziz El-Amraoui
Keyword(s):  
Vision Loss ◽  
Genetic Diseases ◽  
Point Mutations ◽  
Photoreceptor Cells ◽  
Gene Replacement ◽  
Easy Access ◽  
Irreversible Damage ◽  
Partial Restoration ◽  
Dna And Rna

Blindness and deafness are the most frequent sensory disorders in humans. Whatever their cause — genetic, environmental, or due to toxic agents, or aging — the deterioration of these senses is often linked to irreversible damage to the light-sensing photoreceptor cells (blindness) and/or the mechanosensitive hair cells (deafness). Efforts are increasingly focused on preventing disease progression by correcting or replacing the blindness and deafness-causal pathogenic alleles. In recent years, gene replacement therapies for rare monogenic disorders of the retina have given positive results, leading to the marketing of the first gene therapy product for a form of childhood hereditary blindness. Promising results, with a partial restoration of auditory function, have also been reported in preclinical models of human deafness. Silencing approaches, including antisense oligonucleotides, adeno-associated virus (AAV)–mediated microRNA delivery, and genome-editing approaches have also been applied to various genetic forms of blindness and deafness The discovery of new DNA- and RNA-based CRISPR/Cas nucleases, and the new generations of base, prime, and RNA editors offers new possibilities for directly repairing point mutations and therapeutically restoring gene function. Thanks to easy access and immune-privilege status of self-contained compartments, the eye and the ear continue to be at the forefront of developing therapies for genetic diseases. Here, we review the ongoing applications and achievements of this new class of emerging therapeutics in the sensory organs of vision and hearing, highlighting the challenges ahead and the solutions to be overcome for their successful therapeutic application in vivo.

scholarly journals Aggregation chimeras provide evidence of in vivo intercellular correction in ovine CLN6 neuronal ceroid lipofuscinosis (Batten disease)

2021 ◽  
Author(s):  
Lucy A. Barry ◽  
Graham W. Kay ◽  
Nadia L. Mitchell ◽  
Samantha J. Murray ◽  
Nigel P. Jay ◽  
...  
Keyword(s):  
Vision Loss ◽  
Inflammatory Responses ◽  
Neuronal Ceroid Lipofuscinosis ◽  
Batten Disease ◽  
Brain Regions ◽  
Glial Activation ◽  
Neuronal Ceroid Lipofuscinoses ◽  
Lysosomal Storage ◽  
Storage Body

AbstractThe neuronal ceroid lipofuscinoses (NCLs; Batten disease) are fatal, mainly childhood, inherited neurodegenerative lysosomal storage diseases. Sheep affected with a CLN6 form display progressive regionally defined glial activation and subsequent neurodegeneration, indicating that neuroinflammation may be causative of pathogenesis. In this study, aggregation chimeras were generated from homozygous unaffected normal and CLN6 affected sheep embryos, resulting in seven chimeric animals with varied proportions of normal to affected cells. These sheep were classified as affected-like, recovering-like or normal-like, based on their cell-genotype ratios and their clinical and neuropathological profiles.Neuropathological examination of the affected-like animals revealed intense glial activation, prominent storage body accumulation and severe neurodegeneration within all cortical brain regions, along with vision loss and decreasing intracranial volumes and cortical thicknesses consistent with ovine CLN6 disease. In contrast, intercellular communication affecting pathology was evident at both the gross and histological level in the normal-like and recovering-like chimeras, resulting in a lack of glial activation and rare storage body accumulation in only a few cells. Initial intracranial volumes of the recovering-like chimeras were below normal but progressively recovered to about normal by two years of age. All had normal cortical thicknesses, and none went blind. Extended neurogenesis was evident in the brains of all the chimeras.This study indicates that although CLN6 is a membrane bound protein, the consequent defect is not cell intrinsic. The lack of glial activation and inflammatory responses in the normal-like and recovering-like chimeras indicate that newly generated cells are borne into a microenvironment conducive to maturation and survival.

Liver-Directed AAV Gene Therapy for Gaucher Disease

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 3354-3354 ◽  
Author(s):  
Carlos J Miranda ◽  
Miriam Canavese ◽  
Elisa Chisari ◽  
Jalpa Pandya ◽  
Clement Cocita ◽  
...  
Keyword(s):  
Gene Therapy ◽  
Gaucher Disease ◽  
Therapeutic Benefit ◽  
Wild Type ◽  
Employment Equity ◽  
Target Organs ◽  
Equity Ownership

Introduction: Gaucher disease (GD), one of the most common lysosomal storage disorders, is an autosomal recessive condition resulting from mutations in the GBA gene that codes for the b-glucocerebrosidase (GCase) enzyme. Over 90% of patients have type 1 GD, which is characterised by lipid engorged macrophages (known as Gaucher cells) in multiple organs, including spleen, liver and bone marrow, with no overt involvement of the central nervous system (CNS). The current standard of care for type 1 GD patients includes enzyme replacement therapy (ERT), which provides good overall therapeutic benefit. However, ERT is administered intravenously every other week, resulting in a high cumulative cost and a significant treatment burden. Furthermore, disease manifestations, such as pulmonary and skeletal disease, remain unresolved with ERT. Gene therapy is emerging as a very promising avenue of treatment for various monogenic disorders and has the potential to provide sustained levels of GCase enzyme expression after a single treatment. Here we have evaluated liver-directed gene therapy in vitro and in vivo for the treatment of GD. Methods: Adeno-associated virus (AAV) constructs were optimised to express full-length wild-type GCase protein (GBA AAV) and packaged in AAV8 capsids for in vivo mouse studies, or our novel AAVS3 capsid for in vitro studies in a human cell line. GCase activity was determined fluorometrically with 4-Methylumbelliferyl-β-D-glucopyranoside and activity was based on a 4-methylumbelliferone standard curve. Levels of GCase in plasma, and uptake in GD target organs were compared between our GBA AAV optimized construct and ERT treatment with velaglucerase alfa (VPRIV®) in C57BL/6 wild type mice. Doses used ranged from 2x109 to 2x1012 vg/kg for GBA AAV constructs and 60 U/kg for ERT. Results: Our initial proof of concept studies for liver-directed AAV gene therapy of GD used an AAV construct encoding the native full-length human GBA cDNA (RC-04-01). After a single intravenous injection into mice, RC-04-01 led to a dose-dependent expression of GCase in liver and robust levels of enzymatically active GCase in plasma. Based on these preliminary data, 37 GBA AAV constructs with optimisations to the coding sequence and changes to the promoter, signal peptide, and polyA sequences were designed and evaluated in vitro for GCase production. Among them, 6 constructs outperformed RC-04-01 and were further tested in mice. Construct RC-04-26 showed the highest GCase activity levels both in vivo and in vitro. At a dose of 2x1012 vg/kg, RC-04-26 showed plasma active GCase levels up to 9.4-fold higher than RC-04-01. Upon GBA AAV infusion, plasma GCase levels were steady and sustained for the duration of the entire study period (9 months). In addition, RC-04-26 infusion into mice resulted in robust uptake of GCase by cells in spleen, bone marrow and lung, demonstrating that liver-produced GCase is taken up by macrophages present in the GD target organs. Lastly, GCase bioavailability was evaluated after a single administration of RC-04-26 or ERT. With velaglucerase alfa, GCase was rapidly cleared from the bloodstream and tissues. However, RC-04-26 resulted in sustained and steady levels of GCase, with an overall bioavailability of GCase uptake over 170-fold higher than with ERT. Conclusions: Our current data support the hypothesis that a single administration of an optimised liver directed GBA AAV vector results in sustained elevation of GCase in the bloodstream and higher level of GCase bioavailability for uptake into macrophages than velaglucerase alfa. This observation supports further development of AAV gene therapy for Gaucher disease with the potential for enhanced therapeutic benefit from a one-off administration. Disclosures Miranda: Freeline Therapeutics: Employment, Equity Ownership. Canavese:Freeline Therapeutics: Employment, Equity Ownership. Chisari:Freeline Therapeutics: Employment, Equity Ownership. Pandya:Freeline: Employment, Equity Ownership. Cocita:Freeline Therapeutics: Employment, Equity Ownership. Portillo:Freeline: Employment, Equity Ownership. McIntosh:Freeline Therapeutics: Consultancy, Equity Ownership. Kia:Freeline Therapeutics: Employment, Equity Ownership. Foley:Freeline: Employment, Equity Ownership. Dane:Freeline: Employment, Equity Ownership. Jeyakumar:Freeline Therapeutics: Employment, Equity Ownership. Sheridan:Freeline Therapeutics: Employment, Equity Ownership. Corbau:Freeline: Employment, Equity Ownership. Nathwani:Freeline: Consultancy, Equity Ownership, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties.

scholarly journals Hyperfunctional coagulation factor IX improves the efficacy of gene therapy in hemophilic mice

Blood ◽  
2012 ◽  
Vol 120 (23) ◽  
pp. 4517-4520 ◽  
Author(s):  
Alessio Cantore ◽  
Nisha Nair ◽  
Patrizia Della Valle ◽  
Mario Di Matteo ◽  
Janka Màtrai ◽  
...  
Keyword(s):  
Gene Therapy ◽  
Coagulation Factor ◽  
Therapeutic Index ◽  
Therapeutic Benefit ◽  
Factor Ix ◽  
Promising Strategy ◽  
Transfer Vectors ◽  
B Mice ◽  
In Vivo Administration

Abstract Gene therapy may provide a cure for hemophilia and overcome the limitations of protein replacement therapy. Increasing the potency of gene transfer vectors may allow improvement of their therapeutic index, as lower doses can be administered to achieve therapeutic benefit, reducing toxicity of in vivo administration. Here we generated codon-usage optimized and hyperfunctional factor IX (FIX) transgenes carrying an R338L amino acid substitution (FIX Padua), previously associated with clotting hyperactivity and thrombophilia. We delivered these transgenes to hemophilia B mice by hepatocyte-targeted integration-competent and -defective lentiviral vectors. The hyperfunctional FIX transgenes increased FIX activity reconstituted in the plasma without detectable adverse effects, allowing correction of the disease phenotype at lower vector doses and resulting in improved hemostasis in vivo. The combined effect of codon optimization with the hyperactivating FIX-R338L mutation resulted in a robust 15-fold gain in potency and therefore provides a promising strategy to improve the efficacy, feasibility, and safety of hemophilia gene therapy.

scholarly journals Gene Therapy in Movement Disorders: A Systematic Review of Ongoing and Completed Clinical Trials

2021 ◽  
Vol 12 ◽  
Author(s):  
Aristide Merola ◽  
Noelle Kobayashi ◽  
Alberto Romagnolo ◽  
Brenton A. Wright ◽  
Carlo Alberto Artusi ◽  
...  
Keyword(s):  
Gene Therapy ◽  
Clinical Trials ◽  
Phase I ◽  
Movement Disorders ◽  
Viral Vector ◽  
Gene Replacement ◽  
Therapeutic Benefit ◽  
Included Study ◽  
Acid Decarboxylase ◽  
Searching Strategy

Introduction: We sought to provide an overview of the published and currently ongoing movement disorders clinical trials employing gene therapy, defined as a technology aiming to modulate the expression of one or more genes to achieve a therapeutic benefit.Methods: We systematically reviewed movement disorders gene therapy clinical trials from PubMed and ClinicalTrials.gov using a searching strategy that included Parkinson disease (PD), Huntington disease (HD), amino acid decarboxylase (AADC) deficiency, multiple system atrophy (MSA), progressive supranuclear palsy (PSP), dystonia, tremor, ataxia, and other movement disorders. Data extracted included study characteristics, investigational product, route of administration, safety/tolerability, motor endpoints, and secondary outcomes (i.e., neuroimaging, biomarkers).Results: We identified a total of 46 studies focusing on PD (21 published and nine ongoing), HD (2 published and 5 ongoing), AADC deficiency (4 published and 2 ongoing), MSA (2 ongoing), and PSP (1 ongoing). In PD, intraparenchymal infusion of viral vector-mediated gene therapies demonstrated to be safe and showed promising preliminary data in trials aiming at restoring the synthesis of dopamine, enhancing the production of neurotrophic factors, or modifying the functional interaction between different nodes of the basal ganglia. In HD, monthly intrathecal delivery of an antisense oligonucleotide (ASO) targeting the huntingtin protein (HTT) mRNA proved to be safe and tolerable, and demonstrated a dose-dependent reduction of the cerebrospinal fluid levels of mutated HTT, while a small phase-I study testing implantable capsules of cells engineered to synthesize ciliary neurotrophic factor failed to show consistent drug delivery. In AADC deficiency, gene replacement studies demonstrated to be relatively safe in restoring catecholamine and serotonin synthesis, with promising outcomes. Ongoing movement disorders clinical trials are focusing on a variety of gene therapy approaches including alternative viral vector serotypes, novel recombinant genes, novel delivery techniques, and ASOs for the treatment of HD, MSA, and distinct subtypes of PD (LRRK2 mutation or GBA1 mutation carriers).Conclusion: Initial phase-I and -II studies tested the safety and feasibility of gene therapy in PD, HD, and AADC deficiency. The ongoing generation of clinical trials aims to test the efficacy of these approaches and explore additional applications for gene therapy in movement disorders.

SiRNA technology, the gene therapy of the future?

2008 ◽  
Vol 149 (4) ◽  
pp. 153-159 ◽  
Author(s):  
Zsuzsanna Rácz ◽  
Péter Hamar
Keyword(s):  
Gene Therapy ◽  
Short Interfering Rna ◽  
The Future ◽  
Interfering Rna

A genetikában új korszak kezdődött 17 éve, amikor a petúniában felfedezték a koszuppressziót. Később a koszuppressziót azonosították a növényekben és alacsonyabb rendű eukariótákban megfigyelt RNS-interferenciával (RNSi). Bár a növényekben ez ősi vírusellenes gazdaszervezeti védekezőmechanizmus, emlősökben az RNSi élettani szerepe még nincs teljesen tisztázva. Az RNSi-t rövid kettős szálú interferáló RNS-ek (short interfering RNA, siRNS) irányítják. A jelen cikkben összefoglaljuk az RNSi történetét és mechanizmusát, az siRNS-ek szerkezete és hatékonysága közötti összefüggéseket, a célsejtbe való bejuttatás virális és nem virális módjait. Az siRNS-ek klinikai alkalmazásának legfontosabb akadálya az in vivo alkalmazás. Bár a hidrodinamikus kezelés állatokban hatékony, embereknél nem alkalmazható. Lehetőséget jelent viszont a szervspecifikus katéterezés. A szintetizált siRNS-ek ismert mellékhatásait szintén tárgyaljuk. Bár a génterápia ezen új területén számos problémával kell szembenézni, a sikeres in vitro és in vivo kísérletek reményt jelentenek emberi betegségek siRNS-sel történő kezelésére.

Genetic Engineering of AAV Capsid Gene for Gene Therapy Application

2020 ◽  
Vol 20 (5) ◽  
pp. 321-332
Author(s):  
Yunbo Liu ◽  
Xu Zhang ◽  
Lin Yang
Keyword(s):  
Gene Therapy ◽  
Neutralizing Antibodies ◽  
Human Subjects ◽  
Genetic Diseases ◽  
Capsid Gene ◽  
Human Populations ◽  
Stable Gene ◽  
Efficient Gene ◽  
Gene Therapy Application

Adeno-associated virus (AAV) is a promising vector for in vivo gene therapy because of its excellent safety profile and ability to mediate stable gene expression in human subjects. However, there are still numerous challenges that need to be resolved before this gene delivery vehicle is used in clinical applications, such as the inability of AAV to effectively target specific tissues, preexisting neutralizing antibodies in human populations, and a limited AAV packaging capacity. Over the past two decades, much genetic modification work has been performed with the AAV capsid gene, resulting in a large number of variants with modified characteristics, rendering AAV a versatile vector for more efficient gene therapy applications for different genetic diseases.

Exosome as a Natural Gene Delivery Vector for Cancer Treatment

2020 ◽  
Vol 20 (11) ◽  
pp. 821-830
Author(s):  
Prasad Pofali ◽  
Adrita Mondal ◽  
Vaishali Londhe
Keyword(s):  
Gene Therapy ◽  
Gene Delivery ◽  
Nucleic Acids ◽  
Cancer Treatment ◽  
Intestinal Barrier ◽  
Gene Carrier ◽  
Gene Delivery Vector ◽  
Delivery Vector

Background: Current gene therapy vectors such as viral, non-viral, and bacterial vectors, which are used for cancer treatment, but there are certain safety concerns and stability issues of these conventional vectors. Exosomes are the vesicles of size 40-100 nm secreted from multivesicular bodies into the extracellular environment by most of the cell types in-vivo and in-vitro. As a natural nanocarrier, exosomes are immunologically inert, biocompatible, and can cross biological barriers like the blood-brain barrier, intestinal barrier, and placental barrier. Objective: This review focusses on the role of exosome as a carrier to efficiently deliver a gene for cancer treatment and diagnosis. The methods for loading of nucleic acids onto the exosomes, advantages of exosomes as a smart intercellular shuttle for gene delivery and therapeutic applications as a gene delivery vector for siRNA, miRNA and Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) and also the limitations of exosomes as a gene carrier are all reviewed in this article. Methods: Mostly, electroporation and chemical transfection are used to prepare gene loaded exosomes. Results: Exosome-mediated delivery is highly promising and advantageous in comparison to the current delivery methods for systemic gene therapy. Targeted exosomes, loaded with therapeutic nucleic acids, can efficiently promote the reduction of tumor proliferation without any adverse effects. Conclusion: In the near future, exosomes can become an efficient gene carrier for delivery and a biomarker for the diagnosis and treatment of cancer.

Non-Viral Vectors for Gene Delivery

2018 ◽  
Vol 9 (1) ◽  
pp. 4-11 ◽  
Author(s):  
Aparna Bansal ◽  
Himanshu
Keyword(s):  
Gene Therapy ◽  
Viral Vectors ◽  
Transfection Efficiency ◽  
Gene Transfection ◽  
Expression System ◽  
Target Cells ◽  
Specific Expression ◽  
Naked Dna

Introduction: Gene therapy has emerged out as a promising therapeutic pave for the treatment of genetic and acquired diseases. Gene transfection into target cells using naked DNA is a simple and safe approach which has been further improved by combining vectors or gene carriers. Both viral and non-viral approaches have achieved a milestone to establish this technique, but non-viral approaches have attained a significant attention because of their favourable properties like less immunotoxicity and biosafety, easy to produce with versatile surface modifications, etc. Literature is rich in evidences which revealed that undoubtedly, non–viral vectors have acquired a unique place in gene therapy but still there are number of challenges which are to be overcome to increase their effectiveness and prove them ideal gene vectors. Conclusion: To date, tissue specific expression, long lasting gene expression system, enhanced gene transfection efficiency has been achieved with improvement in delivery methods using non-viral vectors. This review mainly summarizes the various physical and chemical methods for gene transfer in vitro and in vivo.

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