Validation of a mitochondrial RNA therapeutic strategy using fibroblasts from a Leigh syndrome patient with a mutation in the mitochondrial ND3 gene

Yuma Yamada,Hitoshi Osaka,Kana Somiya,Hideyoshi Harashima,Akihiko Miyauchi

doi:10.1038/s41598-020-64322-8

Yuma Yamada, Hitoshi Osaka + Show 3 more

Open Access

https://doi.org/10.1038/s41598-020-64322-8

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Journal: Scientific Reports	Publication Date: May 5, 2020
Citations: 29	License type: open-access

Affiliation: Hokkaido University, Jichi Medical University

Abstract

We report on the validation of a mitochondrial gene therapeutic strategy using fibroblasts from a Leigh syndrome patient by the mitochondrial delivery of therapeutic mRNA. The treatment involves delivering normal ND3 protein-encoding mRNA as a therapeutic RNA to mitochondria of the fibroblasts from a patient with a T10158C mutation in the mtDNA coding the ND3 protein, a component of the mitochondrial respiratory chain complex I. The treatment involved the use of a liposome-based carrier (a MITO-Porter) for delivering therapeutic RNA to mitochondria via membrane fusion. The results confirmed that the mitochondrial transfection of therapeutic RNA by the MITO-Porter system resulted in a decrease in the levels of mutant RNA in mitochondria of diseased cells based on reverse transcription quantitative PCR. An evaluation of mitochondrial respiratory activity by respirometry also showed that transfection using the MITO-Porter resulted in an increase in maximal mitochondrial respiratory activity in the diseased cells.

Highlights

There are currently no available treatments for mitochondrial diseases
We examined the use of a non-viral mitochondrial gene delivery system in an attempt to produce mitochondrial transgene expression by delivering a mitochondrial DNA vector to mitochondria
We evaluated a procedure for mitochondrial gene therapy that involves delivering therapeutic RNA to mitochondria using fibroblasts from a patient with a T10158C mutation in the mitochondrial DNA (mtDNA) that codes for the ND3 protein[16]

Summary

Introduction

There are currently no available treatments for mitochondrial diseases. The main cause of a mitochondrial disease is a mutation in the mitochondrial DNA (mtDNA). Since tRNA and rRNA are involved in mitochondrial transcription and the translation of mtDNA, and all of the 13 proteins that are encoded for are subunits of the respiratory chain complex, mutations in mtDNA can result in serious, life-threatening diseases. Some of the previous reports focused on virus vectors resulting in mitochondrial transgene expression first reported by Yu et al.[4] They used an Adeno-associated virus (AAV) as a carrier for achieving mitochondrial gene transfection. We examined the use of a non-viral mitochondrial gene delivery system in an attempt to produce mitochondrial transgene expression by delivering a mitochondrial DNA vector to mitochondria. We evaluated a procedure for mitochondrial gene therapy that involves delivering therapeutic RNA to mitochondria using fibroblasts from a patient with a T10158C mutation in the mtDNA that codes for the ND3 protein[16]. We investigated the therapeutic effect by measuring the mitochondrial respiration of the diseased cells after transfection

Methods

Results

Conclusion