Modulating CRISPR/Cas9 genome-editing activity by small molecules

Chen, Siwei; Deng, Chao; Liu, Bin; Haisma, Hidde J.

doi:10.1016/j.drudis.2021.11.018

Cited by 18 publications

(21 citation statements)

References 123 publications

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“…In addition to Acrs, small molecules are capable of modulating the activity of CRISPR/ Cas9 systems. Small molecules are used to arrest cells at G1/S, G2/M, or S phases, regulate chromatin accessibility, inhibit NHEJ-mediated repair, and promote HDR [332].…”

Section: Crispr/cas9 Activity Modulatorsmentioning

confidence: 99%

CRISPR/Cas9 Landscape: Current State and Future Perspectives

Tyumentseva,

Tyumentsev,

Akimkin

2023

IJMS

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CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 is a unique genome editing tool that can be easily used in a wide range of applications, including functional genomics, transcriptomics, epigenetics, biotechnology, plant engineering, livestock breeding, gene therapy, diagnostics, and so on. This review is focused on the current CRISPR/Cas9 landscape, e.g., on Cas9 variants with improved properties, on Cas9-derived and fusion proteins, on Cas9 delivery methods, on pre-existing immunity against CRISPR/Cas9 proteins, anti-CRISPR proteins, and their possible roles in CRISPR/Cas9 function improvement. Moreover, this review presents a detailed outline of CRISPR/Cas9-based diagnostics and therapeutic approaches. Finally, the review addresses the future expansion of genome editors’ toolbox with Cas9 orthologs and other CRISPR/Cas proteins.

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Section: Crispr/cas9 Activity Modulatorsmentioning

confidence: 99%

CRISPR/Cas9 Landscape: Current State and Future Perspectives

Tyumentseva,

Tyumentsev,

Akimkin

2023

IJMS

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“…Combining CRISPR-Cas9 with the assembled guide RNA library is beneficial for highthroughput screening of vital genes that lead to specific phenotypes and diseases. 7,8 The "phenotype to genotype" approach involves manipulating genomic patterns, selecting cell lines with the phenotype of interest and sequencing the perturbations to identify lethal genes related to cell viability. 9,10 Simultaneously, large-scale functional screenings dependent on cancer type have been carried out in various cell lines with suitable characteristics to evaluate the impact of target gene knockout on cell viability.…”

Section: Introductionmentioning

confidence: 99%

“…The emergence of clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR‐associated protein 9 (Cas9) analysis provides a powerful tool for precision medicine. Combining CRISPR‐Cas9 with the assembled guide RNA library is beneficial for high‐throughput screening of vital genes that lead to specific phenotypes and diseases 7,8 . The “phenotype to genotype” approach involves manipulating genomic patterns, selecting cell lines with the phenotype of interest and sequencing the perturbations to identify lethal genes related to cell viability 9,10 .…”

Section: Introductionmentioning

confidence: 99%

CRISPR‐Cas9 screening identified lethal genes enriched in necroptosis pathway and of prognosis significance in osteosarcoma

Liu

Wang

et al. 2023

The Journal of Gene Medicine

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BackgroundThe present study aimed to identify indispensable genes associated with tumor cell viability according to the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR‐associated protein 9 (Cas9) datasets, which may support new therapeutic targets for patients with osteosarcoma.MethodsThe transcriptome patterns between tumor and normal tissues, which were obtained from the Therapeutically Applicable Research to Generate Effective Treatments dataset, were overlapped with the genomics associated with cell viability screened by CRISPR‐Cas9 technology. Kyoto Encyclopedia of Genes and Genomes and Gene Ontology analyses were employed to determine enrichment pathways related to lethal genes. Least absolute shrinkage and selection operator (LASSO) regression was employed to construct a risk model related to lethal genes for predicting clinical outcomes of osteosarcoma. Univariate and multivariate Cox regressions were conducted to assess the prognostic value of this feature. Weighted gene co‐expression network analysis was performed to identify modules associated with patients with high‐risk score.ResultsIn total, 34 lethal genes were identified in this investigation. These genes were enriched in the necroptosis pathway. The risk model based on LASSO regression algorithm distinguishes patients with high‐risk score from patients with low‐risk score. Compared with low‐risk patients, high‐risk patients showed a shorter overall survival rate in both the training and validation sets. The time‐dependent receiver operating characteristic curves of 1, 3 and 5 years displayed that the risk score has great prediction performance. The necroptosis pathway represents the main difference in biological behavior between the high‐risk group and the low‐risk group. Meanwhile, CDK6 and SMARCB1 may serve as important targets for detecting osteosarcoma progression.ConclusionsThe present study developed a predictive model that outperformed classical clinicopathological parameters for predicting the clinical outcomes of osteosarcoma patients and identified specific lethal genes, including CDK6 and SMARCB1, as well as the necroptosis pathway. These findings may serve as potential targets for future osteosarcoma treatments.

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“…However, it remains elusive as to the location of the underlying pocket(s) in SpCas9 for ligand engagement and how the ligand binding results in the blockade of the DNA-bound state of SpCas9. Addressing these questions has important implications for the potency optimization of these inhibitors and rational design of novel compounds toward the development of safer SpCas9-based therapeutics. ,, …”

Section: Introductionmentioning

confidence: 99%

“…Addressing these questions has important implications for the potency optimization of these inhibitors and rational design of novel compounds toward the development of safer SpCas9based therapeutics. 8,13,14 Molecular dynamics simulations and other computational methods have proved to be powerful in deciphering the structure-dynamics-function relationships of CRISPR-Cas systems (as recently reviewed in ref 15). 16−21 To this end, we set up an integrative computational protocol that encompasses a variety of state-of-the-art approaches of computational biophysics.…”

Section: ■ Introductionmentioning

confidence: 99%

Identification of a Cryptic Binding Site in CRISPR-Cas9 for Targeted Inhibition

Zhang

Zuo

2023

J. Chem. Inf. Model.

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The need for precision control of CRISPR-Cas9 genome editing has created a demand for anti-CRISPR molecules. Recently, the first class of small-molecule Cas9 inhibitors has been identified, verifying the feasibility of regulating CRISPR-Cas9 activity using direct-acting small molecules. However, it remains enigmatic as to the location of the ligand binding site(s) on CRISPR-Cas9 and how the ligand binding leads to Cas9 functional inhibition. Here, we established an integrative computational protocol, including massive binding site mapping, molecular docking, molecular dynamics simulations, and free energy calculations. Ultimately, a Cas9 ligand binding site was discovered from the dynamics trajectories that is hidden within its carboxylterminal domain (CTD), a domain recognizing the protospacer adjacent motif (PAM). Using the top inhibitor BRD0539 as a probe, we demonstrated that the ligand binding induces significant CTD structural rearrangements toward an incompetent conformation for PAM DNA engagement. The revealed molecular mechanism of BRD0539 inhibiting Cas9 is in well agreement with the experimental data. This study provides a structural and mechanistic basis for the potency improvement of existing ligands and the rational discovery of novel small-molecule brakes for developing safer CRISPR-Cas9 technologies.

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Modulating CRISPR/Cas9 genome-editing activity by small molecules

Cited by 18 publications

References 123 publications

CRISPR/Cas9 Landscape: Current State and Future Perspectives

CRISPR/Cas9 Landscape: Current State and Future Perspectives

CRISPR‐Cas9 screening identified lethal genes enriched in necroptosis pathway and of prognosis significance in osteosarcoma

Identification of a Cryptic Binding Site in CRISPR-Cas9 for Targeted Inhibition

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