Correction of metabolic abnormalities in a mouse model of glycogen storage disease type Ia by CRISPR/Cas9-based gene editing

Arnaoutova, Irina; Zhang, Lisa; Chen, Hung-Dar; Mansfield, Brian C.; Chou, Janice Y.

doi:10.1016/j.ymthe.2020.12.027

Cited by 21 publications

(50 citation statements)

References 25 publications

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“…We have previously shown that GSD-Ia mice expressing ≥3% of normal hepatic G6Pase-α activity maintain glucose homeostasis. 9,10,19 We now show that the dsODN insertional strategy restored hepatic G6Pase-α activity to $4% of normal hepatic activity in the G6pc-R83C mice and increased the 0% survival rate to age 6 weeks for the untreated G6pc-R83C mice to a 100% survival rate to age 16 weeks for the dsODN-treated mice. Furthermore, the LNP-dsODN-treated mice could maintain glucose homeostasis and sustain long hours of fasting.…”

Section: Introductionmentioning

confidence: 60%

“…19 Consequently, we used the G6pc-R83 (wild-type) and G6pc-R83/R83C (heterozygote) mice with indistinguishable phenotype as controls. 19 Mice were maintained on a standard NIH-31 Open formula mouse/ rat sterilizable diet from Envigo. To sustain the survival of the untreated G6pc-R83C mice beyond age 1 week, a glucose therapy consisting of daily subcutaneous injection of 100-150 μL of 15% glucose per mouse starting at age 8 days.…”

Section: Animalsmentioning

confidence: 99%

“…Liver microsome isolation and microsomal phosphohydrolase assays were performed as described previously. 9,19 In phosphohydrolase assays, reaction mixtures (50 μL) containing 50 mM sodium cacodylate buffer, pH 6.5, 2 mM EDTA, 10 mM G6P, and appropriate amounts of microsomal preparations were incubated at 30 C for 10 min. Disrupted microsomal membranes were prepared by incubating intact membranes in 0.2% deoxycholate for 20 min at 4 C. Nonspecific phosphatase activity was estimated by pre-incubating disrupted microsomal preparations at pH 5 for 10 min at 37 C to inactivate the acid-labile G6Pase-α.…”

Section: Phosphohydrolase Assaysmentioning

confidence: 99%

“…We therefore explored alternative genetic technologies for GSD-Ia therapy, such as CRISPR/Cas9-based gene editing. [13][14][15][16][17][18] We previously generated a G6pc-R83C mouse strain 19 carrying the prevalent pathogenic G6PC-p.R83C variant 20,21 and showed that the G6pc-R83C mice exhibit the pathophysiology of impaired glucose homeostasis mimicking human GSD-Ia. 19 In an initial exploration of CRISP/Cas-9-based editing using AAV to deliver the CRISPR reagents, we showed that a homology-directed repair (HDR) strategy could correct the abnormal metabolic phenotype of neonatal G6pc-R83C mice.…”

Section: Introductionmentioning

confidence: 99%

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CRISPR/Cas9‐based double‐strand oligonucleotide insertion strategy corrects metabolic abnormalities in murine glycogen storage disease type‐Ia

Samanta

George

Arnaoutova

et al. 2023

J of Inher Metab Disea

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Glycogen storage disease type‐Ia (GSD‐Ia), characterized by impaired blood glucose homeostasis, is caused by a deficiency in glucose‐6‐phosphatase‐α (G6Pase‐α or G6PC). Using the G6pc‐R83C mouse model of GSD‐Ia, we explored a CRISPR/Cas9‐based double‐strand DNA oligonucleotide (dsODN) insertional strategy that uses the non‐homologous end joining repair mechanism to correct the pathogenic p.R83C variant in G6pc exon‐2. The strategy is based on the insertion of a short dsODN into G6pc exon‐2 to disrupt the native exon, and to introduce an additional splice acceptor site and the correcting sequence. When transcribed and spliced the edited gene would generate a wild‐type mRNA encoding the native G6Pase‐α protein. The editing reagents formulated in lipid nanoparticles (LNP) were delivered to the liver. Mice were treated either with one dose of LNP‐dsODN at age 4 weeks or with 2 doses of LNP‐dsODN at age 2 and 4 weeks. The G6pc‐R83C mice receiving successful editing expressed ~4% of normal hepatic G6Pase‐α activity, maintained glucose homeostasis, lacked hypoglycemic seizures, and displayed normalized blood metabolite profile. The outcomes are consistent with preclinical studies supporting previous gene augmentation therapy which is currently in clinical trials. This editing strategy may offer the basis for a therapeutic approach with an earlier clinical intervention than gene augmentation, with the additional benefit of a potentially permanent correction of the GSD‐Ia phenotype.This article is protected by copyright. All rights reserved.

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Section: Introductionmentioning

confidence: 60%

Section: Animalsmentioning

confidence: 99%

Section: Phosphohydrolase Assaysmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

CRISPR/Cas9‐based double‐strand oligonucleotide insertion strategy corrects metabolic abnormalities in murine glycogen storage disease type‐Ia

Samanta

George

Arnaoutova

et al. 2023

J of Inher Metab Disea

Self Cite

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“…Glycogen storage disease Ia (GSD1A, OMIM #232200), also known as von Gierke disease, is caused by pathogenic mutations in the glucose-6-phosphatase alpha subunit ( G6PC ) gene that result in the accumulation of glycogen throughout the body. Recently, the highly prevalent G6PC p.R83C variant was subjected to in vivo CRISPR-based genome editing in mouse models using two AAVs, one encoding SaCas9 and the other encoding sgRNA, 68 resulting in normalization of G6Pase activity, reductions in serum insulin levels, and long-term survival.…”

Section: Clinical Applications Of Crispr-based Genome Editingmentioning

confidence: 99%

Basic Principles and Clinical Applications of CRISPR-Based Genome Editing

Lim

Kim

2022

Yonsei Med J

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Advances in sequencing technologies have facilitated the discovery of previously unknown genetic variants in both inherited and acquired disorders, and tools to correct these pathogenic variants are rapidly evolving. Since the first introduction of CRISPR-Cas9 in 2012, the field of CRISPR-based genome editing has progressed immensely, giving hope to many patients suffering from genetic disorders that lack effective treatment. In this review, we will examine the basic principles of CRISPR-based genome editing, explain the mechanisms of new genome editors, including base editors and prime editors, and evaluate the therapeutic possibilities of CRISPR-based genome editing by focusing on recently published clinical trials and animal studies. Although efficacy and safety issues remain a large concern, we cannot deny that CRISPR-based genome editing will soon be prevalent in clinical practice.

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Gene therapy for glycogen storage diseases

Koeberl

Koch

Lim

et al. 2023

J of Inher Metab Disea

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Glycogen storage disorders (GSDs) are inherited disorders of metabolism resulting from the deficiency of individual enzymes involved in the synthesis, transport, and degradation of glycogen. This literature review summarizes the development of gene therapy for the GSDs. The abnormal accumulation of glycogen and deficiency of glucose production in GSDs lead to unique symptoms based upon the enzyme step and tissues involved, such as liver and kidney involvement associated with severe hypoglycemia during fasting and the risk of long‐term complications including hepatic adenoma/carcinoma and end stage kidney disease in GSD Ia from glucose‐6‐phosphatase deficiency, and cardiac/skeletal/smooth muscle involvement associated with myopathy +/− cardiomyopathy and the risk for cardiorespiratory failure in Pompe disease. These symptoms are present to a variable degree in animal models for the GSDs, which have been utilized to evaluate new therapies including gene therapy and genome editing. Gene therapy for Pompe disease and GSD Ia has progressed to Phase I and Phase III clinical trials, respectively, and are evaluating the safety and bioactivity of adeno‐associated virus vectors. Clinical research to understand the natural history and progression of the GSDs provides invaluable outcome measures that serve as endpoints to evaluate benefits in clinical trials. While promising, gene therapy and genome editing face challenges with regard to clinical implementation, including immune responses and toxicities that have been revealed during clinical trials of gene therapy that are underway. Gene therapy for the glycogen storage diseases is under development, addressing an unmet need for specific, stable therapy for these conditions.

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Correction of metabolic abnormalities in a mouse model of glycogen storage disease type Ia by CRISPR/Cas9-based gene editing

Cited by 21 publications

References 25 publications

CRISPR/Cas9‐based double‐strand oligonucleotide insertion strategy corrects metabolic abnormalities in murine glycogen storage disease type‐Ia

CRISPR/Cas9‐based double‐strand oligonucleotide insertion strategy corrects metabolic abnormalities in murine glycogen storage disease type‐Ia

Basic Principles and Clinical Applications of CRISPR-Based Genome Editing

Gene therapy for glycogen storage diseases

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