Delivery Approaches for Therapeutic Genome Editing and Challenges

Ates, Ilayda; Rathbone, Tanner; Stuart, Callie; Bridges, P Hudson; Cottle, Renee N.

doi:10.3390/genes11101113

Cited by 43 publications

(37 citation statements)

References 241 publications

(310 reference statements)

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“…Safe and effective cellular delivery of engineered nucleases, gRNAs, and template sequences constitutes a key step in the process of gene editing. For ex vivo genome editing, approaches for the delivery of the required constituents within target cells can be broadly classified into viral vectors, electroporation, and cell-penetrating peptides [ 69 ]. In primary cells, including HSPCs, nucleases and the associated gRNAs are most effectively delivered by electroporation of mRNA molecules or as an RNP complex between gRNAs and the nuclease (e.g., Cas9) protein.…”

Section: The Process Of Genome Editingmentioning

confidence: 99%

Advances and Obstacles in Homology-Mediated Gene Editing of Hematopoietic Stem Cells

Salisbury-Ruf

Larochelle

2021

JCM

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Homology-directed gene editing of hematopoietic stem and progenitor cells (HSPCs) is a promising strategy for the treatment of inherited blood disorders, obviating many of the limitations associated with viral vector-mediated gene therapies. The use of CRISPR/Cas9 or other programmable nucleases and improved methods of homology template delivery have enabled precise ex vivo gene editing. These transformative advances have also highlighted technical challenges to achieve high-efficiency gene editing in HSPCs for therapeutic applications. In this review, we discuss recent pre-clinical investigations utilizing homology-mediated gene editing in HSPCs and highlight various strategies to improve editing efficiency in these cells.

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Section: The Process Of Genome Editingmentioning

confidence: 99%

Advances and Obstacles in Homology-Mediated Gene Editing of Hematopoietic Stem Cells

Salisbury-Ruf

Larochelle

2021

JCM

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“…The major disadvantage of ZFN technology is the probability of unwanted genes editing at off-target sites [ 41 ]. TALEN is structurally similar to ZFN.…”

Section: Comparison To Zfns and Talensmentioning

confidence: 99%

“…FokI also needs dimerization from two TALENs to introduce DSBs [ 42 , 43 , 44 ]. The major disadvantage of TALEN technology is to induce mutation at off-target sites [ 41 ]. In contrast to them, Cas9 is an RNA-guided nuclease, and has sequence specificity largely due to Watson–Crick base pairing between the target DNA site and its gRNA, apart from direct interaction between Cas9 and PAM [ 14 , 45 ].…”

Section: Comparison To Zfns and Talensmentioning

confidence: 99%

Various Aspects of a Gene Editing System—CRISPR–Cas9

Janik

Niemcewicz

Ceremuga

et al. 2020

IJMS

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The discovery of clustered, regularly interspaced short palindromic repeats (CRISPR) and their cooperation with CRISPR-associated (Cas) genes is one of the greatest advances of the century and has marked their application as a powerful genome engineering tool. The CRISPR–Cas system was discovered as a part of the adaptive immune system in bacteria and archaea to defend from plasmids and phages. CRISPR has been found to be an advanced alternative to zinc-finger nucleases (ZFN) and transcription activator-like effector nucleases (TALEN) for gene editing and regulation, as the CRISPR–Cas9 protein remains the same for various gene targets and just a short guide RNA sequence needs to be altered to redirect the site-specific cleavage. Due to its high efficiency and precision, the Cas9 protein derived from the type II CRISPR system has been found to have applications in many fields of science. Although CRISPR–Cas9 allows easy genome editing and has a number of benefits, we should not ignore the important ethical and biosafety issues. Moreover, any tool that has great potential and offers significant capabilities carries a level of risk of being used for non-legal purposes. In this review, we present a brief history and mechanism of the CRISPR–Cas9 system. We also describe on the applications of this technology in gene regulation and genome editing; the treatment of cancer and other diseases; and limitations and concerns of the use of CRISPR–Cas9.

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“…However, CRISPRi- or Cas13-based therapeutic approaches face several challenges. One of the major limitations is the efficient delivery of a large cargo size (dCas9-KRAB or Cas13 and a guide RNA) to the cells of interest, using viral or nonviral vectors [ 183 ]. Among viral delivery systems, adeno-associated viruses (AAVs) exhibit interesting features, such as high gene transfer efficiency and low immunogenicity.…”

Section: Lncrna-directed Therapeutic Approachesmentioning

confidence: 99%

“…Among viral delivery systems, adeno-associated viruses (AAVs) exhibit interesting features, such as high gene transfer efficiency and low immunogenicity. However, AAVs have small packaging capacity (<5 kb), which limits their attractiveness for CRISPR-based approaches [ 183 , 184 ]. Alternatively, nonviral delivery systems, including lipid- (LNPs) and polymer-based nanoparticles (PNPs), have substantially evolved in the past decades [ 185 ].…”

Section: Lncrna-directed Therapeutic Approachesmentioning

confidence: 99%

Emerging Role and Therapeutic Potential of lncRNAs in Colorectal Cancer

Schwarzmueller

Bril

Vermeulen

et al. 2020

Cancers

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Maintenance of the intestinal epithelium is dependent on the control of stem cell (SC) proliferation and differentiation. The fine regulation of these cellular processes requires a complex dynamic interplay between several signaling pathways, including Wnt, Notch, Hippo, EGF, Ephrin, and BMP/TGF-β. During the initiation and progression of colorectal cancer (CRC), key events, such as oncogenic mutations, influence these signaling pathways, and tilt the homeostatic balance towards proliferation and dedifferentiation. Therapeutic strategies to specifically target these deregulated signaling pathways are of particular interest. However, systemic blocking or activation of these pathways poses major risks for normal stem cell function and tissue homeostasis. Interestingly, long non-coding RNAs (lncRNAs) have recently emerged as potent regulators of key cellular processes often deregulated in cancer. Because of their exceptional tissue and tumor specificity, these regulatory RNAs represent attractive targets for cancer therapy. Here, we discuss how lncRNAs participate in the maintenance of intestinal homeostasis and how they can contribute to the deregulation of each signaling pathway in CRC. Finally, we describe currently available molecular tools to develop lncRNA-targeted cancer therapies.

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Delivery Approaches for Therapeutic Genome Editing and Challenges

Cited by 43 publications

References 241 publications

Advances and Obstacles in Homology-Mediated Gene Editing of Hematopoietic Stem Cells

Advances and Obstacles in Homology-Mediated Gene Editing of Hematopoietic Stem Cells

Various Aspects of a Gene Editing System—CRISPR–Cas9

Emerging Role and Therapeutic Potential of lncRNAs in Colorectal Cancer

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