Correcting a pathogenic mitochondrial DNA mutation by base editing in mice

Science Translational Medicine, Journal Year: 2025, Volume and Issue: 17(783)

Published: Jan. 29, 2025

Primary mitochondrial disorders are most often caused by deleterious mutations in the DNA (mtDNA). Here, we used a DddA-derived cytosine base editor (DdCBE) to introduce compensatory edit mouse model that carries pathological mutation transfer RNA (tRNA) alanine (mt-tRNAAla) gene. Because original m.5024C→T (G→A mt-tRNAAla) destabilizes mt-tRNAAla aminoacyl stem, designed m.5081G→A (C→T could restore secondary structure of tRNAAla stem. For this, DdCBE gene construct was initially tested an mutant cell line. The reduced amounts these cells were increased after editing up 78% mtDNA. Then, packaged recombinant adeno-associated virus 9 (AAV9) and intravenously administered retro-orbital injections into mice. Expression transduced observed heart skeletal muscle. Total restored muscle dose-dependent manner. Lactate amounts, which heart, also decreased treated However, highest dose AAV9-DdCBE induced severe adverse effects vivo because extensive mtDNA off-target it generated. These results show although is promising therapy tool for disorders, doses therapeutic constructs must be carefully monitored avoid editing.

Language: Английский

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Engineering mtDNA deletions by reconstituting end joining in human mitochondria

Cell, Journal Year: 2025, Volume and Issue: unknown

Published: March 1, 2025

Language: Английский

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Multiple roles of mitochondria in tumorigenesis and treatment: from mechanistic insights to emerging therapeutic strategies

ONCOLOGIE, Journal Year: 2025, Volume and Issue: unknown

Published: March 17, 2025

Language: Английский

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Engineering mtDNA Deletions by Reconstituting End-Joining in Human Mitochondria

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2024, Volume and Issue: unknown

Published: Oct. 17, 2024

Recent breakthroughs in the genetic manipulation of mitochondrial DNA (mtDNA) have enabled precise introduction base substitutions and effective removal genomes carrying harmful mutations. However, reconstitution mtDNA deletions responsible for severe myopathies age-related diseases has not yet been achieved human cells. Here, we developed a method to engineer specific cells by co-expressing end-joining (EJ) machinery targeted endonucleases. As proof-of-concept, used mito-EJ mito-ScaI generate panel clonal cell lines harboring ∼3.5 kb deletion with full spectrum heteroplasmy. Investigating these isogenic revealed critical threshold ∼75% deleted genomes, beyond which exhibited depletion OXPHOS proteins, metabolic disruption, impaired growth galactose-containing media. Single-cell multiomic analysis two distinct patterns nuclear gene deregulation response accumulation; one triggered at another progressively responding increasing In summary, co-expression programable nucleases provides powerful tool model disease-associated different types. Establishing large-scale varying levels heteroplasmy is valuable resource understanding impact on guiding development potential therapeutic strategies.

Language: Английский

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ENZYMATIC TOOLS FOR MITOCHONDRIAL GENOME MANIPULATION

Biochimie, Journal Year: 2024, Volume and Issue: unknown

Published: Oct. 1, 2024

Language: Английский

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