首页|Multiplex gene editing reduces oxalate production in primary hyperoxaluria type 1
Multiplex gene editing reduces oxalate production in primary hyperoxaluria type 1
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国家科技期刊平台
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Targeting key enzymes that generate oxalate precursors or substrates is an alternative strategy to eliminate primary hyperoxaluria type Ⅰ(PH1),the most common and life-threatening type of primary hyperoxaluria.The compact Clustered Regularly Interspaced Short Palindromic Repeats(CRISPR)from the Prevotella and Francisella 1(Cpf1)protein simplifies multiplex gene editing and allows for all-in-one adeno-associated virus(AAV)delivery.We hypothesized that the multiplex capabilities of the Cpf1 system could help minimize oxalate formation in PH1 by simultaneously targeting the hepatic hydroxyacid oxidase 1(Hao1)and lactate dehydrogenase A(Ldha)genes.Study cohorts included treated PH1 rats(AgxtQ84X rats injected with AAV-AsCpf1 at 7 days of age),phosphate-buffered saline(PBS)-injected PH1 rats,untreated PH1 rats,and age-matched wild-type(WT)rats.The most efficient and specific CRISPR RNA(crRNA)pairs targeting the rat Hao1 and Ldha genes were initially screened ex vivo.In vivo experiments demonstrated efficient genome editing of the Hao1 and Ldha genes,primarily resulting in small deletions.This resulted in decreased transcription and translational expression of Hao1 and Ldha.Treatment significantly reduced urine oxalate levels,reduced kidney damage,and alleviated nephrocalcinosis in rats with PH1.No liver toxicity,ex-liver genome editing,or obvious off-target effects were detected.We demonstrated the AAV-AsCpf1 system can target multiple genes and rescue the pathogenic phenotype in PH1,serving as a proof-of-concept for the development of multiplex genome editing-based gene therapy.
Department of Pediatric Urology,Xinhua Hospital Affiliated to Shanghai Jiao Tong University School of Medicine,Shanghai 200092,China
Shanghai Key Laboratory of Regulatory Biology,Institute of Biomedical Sciences and School of Life Sciences,East China Normal University,Shanghai 200241,China
Division of Pediatric Urology,Children's Hospital of Fudan University,Shanghai 201102,China
Science and Technology Commission of Shanghai MunicipalityScience and Technology Commission of Shanghai MunicipalityNational Natural Science Foundation of China
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