This article systematically reviews gene regulation therapies (CRT) for genetic disorders caused by abnormal gene dosage, with a focus on non-editing gene regulation technologies based on dCas9, zinc finger proteins, and TALEs, highlighting their therapeutic potential in various disease models and offering new approaches for diseases that are difficult to address with conventional gene therapy.
Literature Overview
The article 'Modulating gene regulation to treat genetic disorders,' published in Nature Reviews. Drug Discovery, reviews and summarizes an emerging strategy—cis-regulatory therapy (CRT)—for treating genetic disorders caused by abnormal gene dosage through modulation of gene expression levels by targeting cis-regulatory elements (CREs). The article details CRT technical platforms, including zinc finger proteins, TALEs, and the CRISPR-dCas9 system, and their applications in gene upregulation, downregulation, epigenetic modifications, and three-dimensional genome looping. The authors also discuss advances in animal models of CRT for haploinsufficiency, repeat expansion diseases, and epigenetically silenced disorders, along with challenges in delivery systems and clinical translation. The entire passage is coherent and logical, ending with a Chinese period.Background Knowledge
Abnormal levels of gene expression—either too high or too low—can lead to human diseases. Conventional gene therapies, such as gene replacement, are limited by vector capacity and struggle with delivery of large genes. Although gene editing technologies can repair mutations, they carry risks of off-target effects and permanent DNA breaks. Cis-regulatory therapy (CRT) uses nuclease-deficient gene regulation systems to target promoters, enhancers, silencers, and other cis-regulatory elements, enabling precise control of endogenous gene expression without genome cutting, thus offering enhanced safety. CRT can be used to upregulate haploinsufficient genes (e.g., SIM1, SCN1A), activate paralogous genes (e.g., LAMA1, utrophin) to compensate for lost function, or downregulate pathogenic alleles (e.g., mutant HTT, DUX4). Additionally, CRT can regulate gene expression through epigenetic mechanisms such as DNA methylation, histone modification, or chromatin looping. Although CRT has shown therapeutic potential in cellular and animal models, challenges including delivery efficiency, tissue specificity, long-term stability, and immunogenicity remain key barriers to clinical translation. This study systematically outlines the technical framework and application prospects of CRT, providing theoretical and practical guidance for developing novel therapies for dosage-sensitive genetic disorders.
Research Methods and Experiments
The research team conducted a systematic review of multiple CRT platforms, including zinc finger proteins, TALEs, and the CRISPR-dCas9 system, which can be fused with transcriptional activation or repression domains, epigenetic modifying enzymes, or chromatin looping factors to target specific cis-regulatory elements and modulate gene expression. In models of haploinsufficiency disorders, targeting the promoters or enhancers of SIM1 or SCN1A with dCas9-VP64 or dCas9-V160 systems effectively upregulated gene expression and improved obesity and seizure phenotypes in mice. For Duchenne muscular dystrophy (DMD), activation of paralogous genes such as utrophin or klotho using a CRISPRa system improved muscle pathology. In Huntington’s disease models, targeting the mutant HTT allele with TALE-KRAB or ZF-KRAB systems enabled allele-specific suppression. Additionally, demethylation of the FMR1 gene promoter using a dCas9-Tet1 system reactivated its expression, offering a therapeutic strategy for Fragile X syndrome. Chromatin looping technologies such as the CLOuD9 system can force interactions between the fetal γ-globin gene promoter and enhancer, activating its expression for the treatment of β-thalassemia and sickle cell disease.Key Conclusions and Perspectives
Research Significance and Prospects
CRT represents a novel class of gene therapy strategies whose core advantage lies in restoring physiological function by modulating endogenous gene expression levels without altering the genomic sequence, thereby avoiding the risks of off-target effects and insertional mutagenesis associated with conventional gene editing. This approach is particularly suitable for diseases involving large genes (e.g., DMD, LAMA2-related muscular dystrophy) and haploinsufficiency disorders, addressing the limitations of rAAV-based gene replacement therapies.
Future research should focus on developing more efficient, tissue-specific delivery systems (e.g., novel AAV serotypes, lipid nanoparticles), optimizing targeting efficiency and specificity of regulatory elements, and exploring reversible or regulatable CRT systems for fine-tuned dosage control. Additionally, long-term safety, immunogenicity, and stability of epigenetic memory need further validation in large animal models. As the technology matures, CRT is expected to become an important tool for treating a wide range of genetic disorders.
Conclusion
This article systematically summarizes recent advances in cis-regulatory therapy (CRT) for treating genetic disorders. CRT uses nuclease-deficient gene regulation systems to target cis-regulatory elements, enabling precise modulation of endogenous gene expression and offering new strategies for long-gene disorders and haploinsufficiency diseases that are difficult to treat with conventional gene therapy. The study demonstrates CRT's therapeutic potential in various animal models, including upregulation of genes such as SIM1, SCN1A, and utrophin to improve obesity, epilepsy, and DMD phenotypes, suppression of mutant HTT or DUX4 to alleviate Huntington’s disease and facioscapulohumeral muscular dystrophy, and reactivation of the FMR1 gene through epigenetic editing. Furthermore, chromatin looping technologies can activate fetal γ-globin, offering a potential functional cure for β-hemoglobinopathies. Although CRT holds great promise, delivery efficiency, long-term stability, immunogenicity, and off-target effects remain key barriers to clinical translation. Future efforts should focus on optimizing delivery systems, enhancing targeting specificity, and validating long-term safety in large animal models. With technological advancements, CRT is poised to become a vital approach for treating numerous genetic disorders, advancing the field of precision medicine.
