Gene Editing for Therapy and Reverse Genetics of Blood Diseases

A special issue of Genes (ISSN 2073-4425). This special issue belongs to the section "Human Genomics and Genetic Diseases".

Deadline for manuscript submissions: closed (20 March 2023) | Viewed by 12354

Special Issue Editor


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Guest Editor
Molecular Genetics Thalassaemia Department, The Cyprus Institute of Neurology & Genetics, Nicosia 2371, Cyprus
Interests: advanced therapies; genetic modifiers; regulation of globin expression; rare anemias
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Special Issue Information

Dear Colleagues,

Gene editing is increasingly taking center stage for both basic research and translational studies. The key driver of this development is the rapidly growing adoption of editing technology and, in particular, fast-evolving RNA-guided CRISPR/Cas tools, which, with their versatility and ease of use, have facilitated the development of double-strand-break-independent editors and the exploration of new fields of application. In particular blood biology and disorders are a favorite focus of gene editing, motivated by a relatively high prevalence of monogenic, infectious and complex diseases affecting blood cells, and helped by the accessibility of hematopoietic stem and progenitor cells for manipulation.

This Special Issue aims to showcase the application of gene editing technology in therapy development and research for blood diseases. In this context, we welcome Articles, Communications and Reviews providing new insights into (i) developmental and disease mechanisms; (ii) the establishment or study of disease models; (iii) the creation of new editing platforms and molecules for diagnosis, functional study or therapy; and (vi) the refinement of corresponding delivery, targeting and culture procedures.

The topics of interest in the context of gene editing and blood biology include but are not limited to:

  • Editing primary disease-causing mutations;
  • Editing disease modifiers, developmental regulators and pathologically relevant pathways;
  • The treatment or development of cellular and animal models by editing;
  • Improvements in post-editing cell survival, stemness and engraftment behavior;
  • Improvements in the in vivo or in vitro efficiency and specificity of delivery for editing tools;
  • The improvement of or insights into in vivo and ex vivo selection mechanisms for editing events;
  • Comparisons of editing platforms, of gene therapy strategies and of vectors for the delivery of editors and/or templates;
  • New or improved editing technology for RNA, the nuclear or mitochondrial genome and the epigenome;
  • The application of editing technology for the diagnosis and for the detection and verification of disease biomarkers;
  • The modification of the balance between homology-directed repair, non-homologous end joining and other editing events;
  • Safety assessment and optimization for on- and off-target activity.
Dr. Carsten W. Lederer
Guest Editor

Manuscript Submission Information

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Keywords

  • CRISPR/Cas (clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein (Cas))
  • TALEN (Tal effector nuclease)
  • ZFN (zinc finger nuclease)
  • Triplex-forming nucleic acid analog
  • Meganuclease/homing endonuclease
  • Nanoparticles
  • Viral vector
  • Gene therapy
  • Hematopoiesis
  • Erythropoiesis

Published Papers (4 papers)

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Research

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15 pages, 1537 KiB  
Article
Allele-Specific Disruption of a Common STAT3 Autosomal Dominant Allele Is Not Sufficient to Restore Downstream Signaling in Patient-Derived T Cells
by Saskia König, Manfred Fliegauf, Manuel Rhiel, Bodo Grimbacher, Tatjana I. Cornu, Toni Cathomen and Claudio Mussolino
Genes 2022, 13(10), 1912; https://doi.org/10.3390/genes13101912 - 20 Oct 2022
Cited by 1 | Viewed by 1646
Abstract
Dominant negative mutations in the STAT3 gene account for autosomal dominant hyper-IgE syndrome (AD-HIES). Patients typically present high IgE serum levels, recurrent infections, and soft tissue abnormalities. While current therapies focus on alleviating the symptoms, hematopoietic stem cell transplantation (HSCT) has recently been [...] Read more.
Dominant negative mutations in the STAT3 gene account for autosomal dominant hyper-IgE syndrome (AD-HIES). Patients typically present high IgE serum levels, recurrent infections, and soft tissue abnormalities. While current therapies focus on alleviating the symptoms, hematopoietic stem cell transplantation (HSCT) has recently been proposed as a strategy to treat the immunological defect and stabilize the disease, especially in cases with severe lung infections. However, because of the potentially severe side effects associated with allogeneic HSCT, this has been considered only for a few patients. Autologous HSCT represents a safer alternative but it requires the removal of the dominant negative mutation in the patients’ cells prior to transplantation. Here, we developed allele-specific CRISPR-Cas9 nucleases to selectively disrupt five of the most common STAT3 dominant negative alleles. When tested ex vivo in patient-derived hematopoietic cells, allele-specific disruption frequencies varied in an allele-dependent fashion and reached up to 62% of alleles harboring the V637M mutation without detectable alterations in the healthy STAT3 allele. However, assessment of the gene expression profiles of the STAT3 downstream target genes revealed that, upon activation of those edited patient cells, mono-allelic STAT3 expression (functional haploinsufficiency) is not able to sufficiently restore STAT3-dependent signaling in edited T cells cultured in vitro. Moreover, the stochastic mutagenesis induced by the repair of the nuclease-induced DNA break could further contribute to dominant negative effects. In summary, our results advocate for precise genome editing strategies rather than allele-specific gene disruption to correct the underlying mutations in AD-HIES. Full article
(This article belongs to the Special Issue Gene Editing for Therapy and Reverse Genetics of Blood Diseases)
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18 pages, 1849 KiB  
Article
Co-Treatment of Erythroid Cells from β-Thalassemia Patients with CRISPR-Cas9-Based β039-Globin Gene Editing and Induction of Fetal Hemoglobin
by Lucia Carmela Cosenza, Cristina Zuccato, Matteo Zurlo, Roberto Gambari and Alessia Finotti
Genes 2022, 13(10), 1727; https://doi.org/10.3390/genes13101727 - 26 Sep 2022
Cited by 4 | Viewed by 2389
Abstract
Gene editing (GE) is an efficient strategy for correcting genetic mutations in monogenic hereditary diseases, including β-thalassemia. We have elsewhere reported that CRISPR-Cas9-based gene editing can be employed for the efficient correction of the β039-thalassemia mutation. On the other hand, robust [...] Read more.
Gene editing (GE) is an efficient strategy for correcting genetic mutations in monogenic hereditary diseases, including β-thalassemia. We have elsewhere reported that CRISPR-Cas9-based gene editing can be employed for the efficient correction of the β039-thalassemia mutation. On the other hand, robust evidence demonstrates that the increased production of fetal hemoglobin (HbF) can be beneficial for patients with β-thalassemia. The aim of our study was to verify whether the de novo production of adult hemoglobin (HbA) using CRISPR-Cas9 gene editing can be combined with HbF induction protocols. The gene editing of the β039-globin mutation was obtained using a CRISPR-Cas9-based experimental strategy; the correction of the gene sequence and the transcription of the corrected gene were analyzed by allele-specific droplet digital PCR and RT-qPCR, respectively; the relative content of HbA and HbF was studied by high-performance liquid chromatography (HPLC) and Western blotting. For HbF induction, the repurposed drug rapamycin was used. The data obtained conclusively demonstrate that the maximal production of HbA and HbF is obtained in GE-corrected, rapamycin-induced erythroid progenitors isolated from β039-thalassemia patients. In conclusion, GE and HbF induction might be used in combination in order to achieve the de novo production of HbA together with an increase in induced HbF. Full article
(This article belongs to the Special Issue Gene Editing for Therapy and Reverse Genetics of Blood Diseases)
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Review

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24 pages, 2372 KiB  
Review
Epigenetic Regulation of β-Globin Genes and the Potential to Treat Hemoglobinopathies through Epigenome Editing
by Letizia Fontana, Zoe Alahouzou, Annarita Miccio and Panagiotis Antoniou
Genes 2023, 14(3), 577; https://doi.org/10.3390/genes14030577 - 25 Feb 2023
Cited by 3 | Viewed by 4035
Abstract
Beta-like globin gene expression is developmentally regulated during life by transcription factors, chromatin looping and epigenome modifications of the β-globin locus. Epigenome modifications, such as histone methylation/demethylation and acetylation/deacetylation and DNA methylation, are associated with up- or down-regulation of gene expression. The understanding [...] Read more.
Beta-like globin gene expression is developmentally regulated during life by transcription factors, chromatin looping and epigenome modifications of the β-globin locus. Epigenome modifications, such as histone methylation/demethylation and acetylation/deacetylation and DNA methylation, are associated with up- or down-regulation of gene expression. The understanding of these mechanisms and their outcome in gene expression has paved the way to the development of new therapeutic strategies for treating various diseases, such as β-hemoglobinopathies. Histone deacetylase and DNA methyl-transferase inhibitors are currently being tested in clinical trials for hemoglobinopathies patients. However, these approaches are often uncertain, non-specific and their global effect poses serious safety concerns. Epigenome editing is a recently developed and promising tool that consists of a DNA recognition domain (zinc finger, transcription activator-like effector or dead clustered regularly interspaced short palindromic repeats Cas9) fused to the catalytic domain of a chromatin-modifying enzyme. It offers a more specific targeting of disease-related genes (e.g., the ability to reactivate the fetal γ-globin genes and improve the hemoglobinopathy phenotype) and it facilitates the development of scarless gene therapy approaches. Here, we summarize the mechanisms of epigenome regulation of the β-globin locus, and we discuss the application of epigenome editing for the treatment of hemoglobinopathies. Full article
(This article belongs to the Special Issue Gene Editing for Therapy and Reverse Genetics of Blood Diseases)
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23 pages, 443 KiB  
Review
In Vivo Hematopoietic Stem Cell Genome Editing: Perspectives and Limitations
by Nikoletta Psatha, Kiriaki Paschoudi, Anastasia Papadopoulou and Evangelia Yannaki
Genes 2022, 13(12), 2222; https://doi.org/10.3390/genes13122222 - 27 Nov 2022
Cited by 6 | Viewed by 2990
Abstract
The tremendous evolution of genome-editing tools in the last two decades has provided innovative and effective approaches for gene therapy of congenital and acquired diseases. Zinc-finger nucleases (ZFNs), transcription activator- like effector nucleases (TALENs) and CRISPR-Cas9 have been already applied by ex vivo [...] Read more.
The tremendous evolution of genome-editing tools in the last two decades has provided innovative and effective approaches for gene therapy of congenital and acquired diseases. Zinc-finger nucleases (ZFNs), transcription activator- like effector nucleases (TALENs) and CRISPR-Cas9 have been already applied by ex vivo hematopoietic stem cell (HSC) gene therapy in genetic diseases (i.e., Hemoglobinopathies, Fanconi anemia and hereditary Immunodeficiencies) as well as infectious diseases (i.e., HIV), and the recent development of CRISPR-Cas9-based systems using base and prime editors as well as epigenome editors has provided safer tools for gene therapy. The ex vivo approach for gene addition or editing of HSCs, however, is complex, invasive, technically challenging, costly and not free of toxicity. In vivo gene addition or editing promise to transform gene therapy from a highly sophisticated strategy to a “user-friendly’ approach to eventually become a broadly available, highly accessible and potentially affordable treatment modality. In the present review article, based on the lessons gained by more than 3 decades of ex vivo HSC gene therapy, we discuss the concept, the tools, the progress made and the challenges to clinical translation of in vivo HSC gene editing. Full article
(This article belongs to the Special Issue Gene Editing for Therapy and Reverse Genetics of Blood Diseases)
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