2024-03-29T08:05:10Zhttps://eprints.lib.hokudai.ac.jp/dspace-oai/requestoai:eprints.lib.hokudai.ac.jp:2115/733342022-11-17T02:08:08Zhdl_2115_20044hdl_2115_124Mitochondrial transgene expression via an artificial mitochondrial DNA vector in cells from a patient with a mitochondrial diseaseIshikawa, TakuyaSomiya, KanaMunechika, ReinaHarashima, HideyoshiYamada, YumaMitochondriaTransgene expressionIndependent clinical studyMitochondrial disease's patientMitochondrial deliveryMITO-Porter460To achieve mitochondrial gene therapy, developing a mitochondrial transgene expression system that produces therapeutic proteins in mitochondria of disease cells is essential. We previously reported on the design of pCMV-mtLuc (CGG) containing a CMV promotor and a NanoLuc (Nluc) luciferase gene that records adjustments to the mitochondrial codon system, and showed that the mitochondrial transfection of pCMV-mtLuc (CGG) resulted in the efficient production of the Nluc luciferase protein in human HeLa cells. This mitochondrial transfection was achieved using a MITO-Porter, a liposome-based carrier for delivering a cargo to mitochondria via membrane fusion. We report herein that mitochondrial transfection using the MITO-Porter results in mitochondrial transgene expression in G625A fibroblasts obtained from a patient with a mitochondrial disease. We investigated the effect of promoters and the basic structure of pCMV-mtLuc (CGG) on gene expression efficiency, and were able to construct a high performance mitochondrial DNA vector, pCMV-mtLuc (CGG) [hND4] that contains a human mitochondrial endogenous gene. We also constructed an RP/KALA-MITO-Porter composed of the KALA peptide (cell-penetrating peptide) with a mitochondrial RNA aptamer to enhance cellular uptake and mitochondrial targeting. Finally, the mitochondrial transfection of pCMV-mtLuc (CGG) [hND4] in G625A fibroblasts using the RP/KALA-MITO-Porter resulted in strong mitochondrial transgene expression.ElsevierJournal Articleapplication/pdfhttp://hdl.handle.net/2115/73334https://eprints.lib.hokudai.ac.jp/dspace/bitstream/2115/73334/1/WoS_84356_Yamada.pdf0168-3659Journal of controlled release2741091172018-03-28enginfo:doi/10.1016/j.jconrel.2018.02.005©2018. This manuscript version is made available under the CC-BY-NC-ND 4.0 license http://creativecommons.org/licenses/by-nc-nd/4.0/author