Jul 2026· International Journal of Molecular Sciences· Vol 27, pp. 6467· 0 citations· 137 references
Medicine
TL;DR
Overall, delivery systems for reversible genetic medicines should be judged by tissue selectivity, functional editing, duration of action, reversibility after treatment withdrawal, safety, manufacturability, and clinical controllability.
Abstract
Reversible genetic medicines are emerging as controllable alternatives to permanent genome editing by enabling programmable modulation of RNA sequence, transcript abundance, chromatin state, and gene expression without irreversible genomic alteration. However, reversibility is not a single binary property: transient editor exposure, decay of the molecular effect, recovery of cellular function, and clinical capacity to stop, redose, or counteract treatment may diverge. This review therefore distinguishes mechanistic, functional, and clinical reversibility while examining targeted delivery systems for RNA-editing and epigenome-editing therapeutics. Key payloads include ADAR-recruiting oligonucleotides, CRISPR-Cas13 RNA editors, guide RNAs, chemically modified RNAs, editor-encoding mRNAs, dCas9 transcriptional regulators, DNA methylation editors, histone-modifying systems, and CRISPRoff-like platforms. We evaluate extracellular and intracellular delivery barriers, including nuclease degradation, immune recognition, renal clearance, liver uptake, cellular entry, endosomal escape, cytoplasmic release, nuclear localization, chromatin access, editing-window duration, off-target activity, immunogenicity, repeat-dosing feasibility, manufacturing, quality control, potency assays, and regulatory translation. Overall, delivery systems for reversible genetic medicines should be judged by tissue selectivity, functional editing, duration of action, reversibility after treatment withdrawal, safety, manufacturability, and clinical controllability.
Epigenome editing has emerged as a transformative extension of conventional genome engineering, offering precise and programmable control of gene expression without altering the underlying DNA sequence. In contrast to the genetic modifications that introduce permanent sequence changes, epigenome editing exploits reversible and potentially heritable epigenetic mechanisms such as DNA methylation, histone modifications, chromatin remodelling, and RNA-mediated gene silencing to regulate transcriptional states. Advances in next-generation sequencing technologies have enabled high-resolution mapping of epigenetic landscapes, revealing the dynamic and context-dependent nature of chromatin regulation across development and environmental conditions. The integration of programmable DNA-binding platforms, including zinc finger proteins, transcription activator-like effectors, and CRISPR/dCas9-based systems, with epigenetic effector domains has revolutionized locus specific manipulation of chromatin states. These tools facilitate targeted gene activation or repression (CRISPRa/CRISPRi), as well as direct rewriting of epigenetic marks, providing unprecedented opportunities to dissect gene regulatory networks and trait expression. In plants, epigenome editing has shown particular promise for functional genomics, stress adaptation, and crop improvement, enabling fine-tuning of agriculturally important traits such as yield, quality, and stress tolerance. This review comprehensively discusses the molecular mechanisms underlying epigenetic regulation, key epigenome editing tools, their modes of action, and the emerging applications of epigenome editing in plant biology, highlighting it as a potential tool for sustainable agriculture and precision breeding. This review provides a comprehensive and integrated perspective on epigenome editing by linking molecular mechanisms, advanced editing tools, and practical applications in plant systems, thereby offering a foundation for future research and the development of climate-resilient and sustainable crop improvement strategies.
Marifa Gulzar, Gazala Nazir, F. Mushtaq et al.· Saudi Journal of Food Securi...· 0 citations
A comprehensive review of delivery modalities of CRISPR systems solely in vivo that underpin their therapeutic translation and outlines the remaining barriers to durable, tissue-selective, and broadly deployable CRISPR therapeutics is provided.
L. Martin, Jure Bohinc, Alessandra Recchia et al.· International Journal on Bio...· 0 citations
CRISPR editors including nucleases, base editors, and prime editors can efficiently correct disease-causing genetic variants or disrupt target genes. Editing outcomes are commonly evaluated in cultured primary cells, patient-derived cells, or engineered cell lines to study the impact of genetic variation or as a first step before initiating animal studies or clinical translation. Delivery of editors as mRNA together with synthetic guide RNAs into mammalian cells can improve editing efficiency relative to plasmid-based approaches and prevent issues such as DNA integration or off-target editing from sustained expression. This article presents a workflow to prepare genome editor mRNA by in vitro transcription (IVT), including co-transcriptional capping and chemically-modified nucleotides, electroporate editor mRNA and guide RNAs into primary human fibroblasts, induced pluripotent stem cells (iPSCs), or lymphoblastoid cell lines (LCLs), and quantify editing outcomes by targeted amplicon sequencing on an Illumina platform followed by analysis using CRISPResso2. This workflow enables quantitative benchmarking of guide RNAs, electroporation parameters, and editor variants, and supports downstream applications including single-cell cloning, phenotypic assays, preclinical animal studies, and therapeutic development.
Maria Viskadourou, J. Workman, Emma V Burke et al.· Journal of Visualized Experi...· 0 citations
Cell-specific epigenetic editing holds very high therapeutic value for atherosclerosis, cardiomyopathy, and fibrosis, provided that delivery, specificity, and safety challenges are also addressed.
Majed Alsulami, Mahmood Rasool, Ahmed Masoud et al.· The Cardiology· 0 citations
To systematically map cellular factors constraining nonviral genome editing, influencing uptake and intracellular trafficking, we develop a genome-wide CRISPR screening platform linking perturbation of 19,114 genes to editing outcomes in human cells. We identify six negative regulators of delivery whose depletion increases editing efficiency by up to six-fold across diverse payloads, loci, and cell types. We test the top two factors, GJB2 and BET1L, in two distinct human models: correction of a pathogenic adenine base mutation in KCNJ13 and introduction of a cytosine base mutation in the GABAA receptor gene. Depletion of either improves base-editing outcomes by 6-fold, potentially through effects on delivery. In a patient-derived model of retinal channelopathy, knockdown of either gene improves lipid nanoparticle base editing efficiency by over 3.5-fold. This enables functional restoration of Kir7.1 ion channels in a subset of edited cells, highlighting cellular barriers as actionable targets to enhance the potency of genetic therapies. Low editing efficiency of nonviral delivery in post mitotic tissues presents a challenge to the field of gene therapy. Here, authors dissect the genetic regulators of nonviral delivery in post mitotic retinal epithelial cells describe strategies for improved base editor delivery and editing.
Shivani Saxena, Meha Kabra, Amr A. Abdeen et al.· Nature Communications· 0 citations
A novel genome-wide CRISPR screening strategy that will facilitate the systematic engineering of novel nonviral genome editing delivery methods, where the identified novel gene hits can be further used to increase editing efficiency for other therapeutically relevant cell types.
Shivani Saxena, Meha Kabra, Amr A. Abdeen et al.· bioRxiv· 2 citations