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2022 | Book

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Sequence-Specific DNA Binders for the Therapy of Mitochondrial Diseases

Author: Dr. Takuya Hidaka

Publisher: Springer Nature Singapore

Book Series : Springer Theses

Part of: Springer Professional "Wirtschaft+Technik" , Springer Professional "Technik"

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About this book

This book describes the author’s work on the development of sequence-specific DNA binders for the therapy of mitochondrial diseases. In the first chapter, the author provides a detailed background of pyrrole–imidazole polyamides (PIPs) and mitochondrial disease research followed by chapters presenting the author's own research and discoveries. Firstly, the developed compounds called MITO-PIPs, which recognize specific sequences in mitochondrial DNA, are presented. The following chapter demonstrates how, by introducing a DNA alkylating reagent into a MITO-PIP that recognizes the adjacent sequence to a target mutation, the copy number of mutated mitochondrial DNA was successfully reduced in live cells. Furthermore, because nuclear DNA is another important target for treating mitochondrial diseases, the author demonstrated that tri-arginine vectors can enhance nuclear uptake of PIPs and improve their biological activity in cells.
This work will attract readers’ interest because it paves the way for a transgene-free chemical gene therapy for mitochondrial diseases. The book includes a detailed description of experimental procedures, especially compound synthesis. This description helps readers to have a clear image of the author’s studies and to perform similar and extended studies themselves.

Table of Contents

Frontmatter
Chapter 1. Introduction
Abstract
Mitochondria are ubiquitous cellular organelles and generate most of the energy required for cellular function. Not only as the powerhouse of cells, but mitochondria also act as hubs for biosynthesis and metabolic waste management and as balancers to maintain cellular homeostasis. Due to the central role in cells, defects in mitochondrial function cause a critical impact on our body, making mitochondrion an attractive target for therapeutic purposes. This chapter describes the basic mitochondrial biology and diseases caused by impaired mitochondrial function—mitochondrial diseases—including current therapeutic approaches. In addition, pyrrole-imidazole polyamides, a class of DNA-binding ligands with sequence programmability, are introduced as promising drug candidates to modulate DNA mutation and abnormal RNA transcription.
Takuya Hidaka
Chapter 2. Creation of a Synthetic Ligand for Mitochondrial DNA Sequence Recognition and Promoter-Specific Transcription Suppression
Abstract
Synthetic ligands capable of DNA sequence recognition in human mitochondria are in increasing demand because more and more studies have revealed the relation between mitochondrial genome and diseases. In this chapter, a new type of synthetic DNA-binding ligands, termed MITO-PIPs, was developed by conjugating a mitochondria-penetrating peptide with pyrrole-imidazole polyamides (PIPs). A MITO-PIP that inhibits the binding of mitochondrial transcription factor A to the light-strand promoter (LSP) triggered targeted transcriptional suppression of a downstream gene. A melting temperature analysis revealed sequence-specific DNA binding of the MITO-PIP, and mitochondrial accumulation in HeLa cells was also observed. The tunability of PIPs suggests the potential of the MITO-PIPs as potent modulators of mitochondrial gene transcription and mitochondrial DNA mutations.
Takuya Hidaka
Chapter 3. Allele-Specific Replication Inhibition of Mitochondrial DNA by MITO-PIP Conjugated with Alkylation Reagent
Abstract
Pathogenic mutations in mitochondrial DNA cause genetic disorders named mitochondrial diseases. Although mutated mitochondrial DNA should be eliminated from cells, no chemical-based drugs in clinical trials have the potential to modulate mtDNA mutation and cure mitochondrial diseases permanently. To achieve selective elimination of mitochondrial DNA with mutant adenine base, I develop a conjugate of MITO-PIP and chlorambucil (Chb), which can alkylate an adenine base in a sequence-specific manner. The in vitro alkylation assay shows that a conjugate targeting the point mutation in HeLa S3 cells (m.8950G > A) alkylates the adenine at the mutation site. The conjugate also reduces the proportion of mutant mtDNA in cultured HeLa S3 cells. MITO-PIP–Chb conjugates would be drug candidates to cure mitochondrial diseases caused by pathogenic mutations in mtDNA and paves the way to the gene therapy of mitochondrial diseases by a chemical approach.
Takuya Hidaka
Chapter 4. Enhanced Nuclear Accumulation of Pyrrole-Imidazole Polyamides by Incorporation of the Tri-Arginine Vector
Abstract
Mutations in mitochondrial genes encoded in nuclear DNA cause mitochondrial diseases and are important targets to treat the diseases. Although approaches utilizing pyrrole-imidazole polyamides (PIPs) have potential to modulate nuclear DNA transcription and replication based on DNA sequence information, their application in living cells has achieved limited success due to the moderate or poor nuclear accumulation of PIPs. In this chapter, I show that the tri-arginine moiety enhances nuclear accumulation of 12-ring PIPs without compromising their sequence-selective DNA binding. The tri-arginine vector improves biological activity of PIPs and PIP–tri-arginine conjugates achieve efficient transcription inhibition of SOX2-downstream genes in induced pluripotent stem (iPS) cells and HER2 oncogene in human breast cancer cells. This simple vector expands the application of long PIPs by overcoming the compound delivery problems.
Takuya Hidaka
Backmatter
Metadata
Title
Sequence-Specific DNA Binders for the Therapy of Mitochondrial Diseases
Author
Dr. Takuya Hidaka
Copyright Year
2022
Publisher
Springer Nature Singapore
Electronic ISBN
978-981-16-8436-4
Print ISBN
978-981-16-8435-7
DOI
https://doi.org/10.1007/978-981-16-8436-4

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