Small molecules enhance CRISPR genome editing in pluripotent stem cells
Open accessAbstract
The bacterial CRISPR-Cas9 system has emerged as an effective tool for sequence-specific gene knockout through non-homologous end joining (NHEJ), but it remains inefficient for precise editing of genome sequences. Here we develop a reporter-based screening approach for high-throughput identification of chemical compounds that can modulate precise genome editing through homology-directed repair (HDR). Using our screening method, we have identified small molecules that can enhance CRISPR-mediated HDR efficiency, 3-fold for large fragment insertions and 9-fold for point mutations. Interestingly, we have also observed that a small molecule that inhibits HDR can enhance frame shift insertion and deletion (indel) mutations mediated by NHEJ. The identified small molecules function robustly in diverse cell types with minimal toxicity. The use of small molecules provides a simple and effective strategy to enhance precise genome engineering applications and facilitates the study of DNA repair mechanisms in mammalian cells.
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An HDR enhancer and NHEJ inhibitor were used to improve homology-directed repair and efficient genome editing, particularly for precise single-nucleotide modifications.
“An HDR enhancer and NHEJ inhibitor were used to improve homology-directed repair and efficient genome editing, particularly for precise single-nucleotide modifications (Figure 4A) [45,46].”
From Precise CRISPR/Cas9 and Cas12 Correction Using Lipoplexes in Retinal Models Derived from Patients with Inherited Retinal Dystrophies (2026) · cites this paper as [46]
In the passage on SNP editing (indexed under Abstract; page unavailable), L755507 increased precise A4V single-nucleotide editing in human iPS cells almost ninefold and reduced indel mutations. The paper does not report using a separate NHEJ inhibitor alongside an HDR enhancer; it describes L755507 as an HDR enhancer that also reduced indels. (LLM, conf=0.87)
Experimental — accuracy not yet measuredThe desired recombination efficacy of HDR often occurs infrequently, which presents enormous challenges for robust applications.
“However, the desired recombination efficacy often occurs infrequently, which presents enormous challenges for robust applications [61, 82–84].”
From Non-viral delivery systems for CRISPR/Cas9-based genome editing: challenges and opportunities (2018) · cites this paper as [83]
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“An HDR enhancer and NHEJ inhibitor were used to improve homology-directed repair and efficient genome editing, particularly for precise single-nucleotide modifications (Figure 4A) [45,46].”
- Authors
- Yu, C., Liu, Y., Ma, T., Liu, K., Xu, S., Zhang, Y., Liu, H., La Russa, M., Xie, M., Ding, S., & Qi, L. S.
- Affiliations
- Stanford University (US)
- Gladstone Institutes (US)
- Corresponding author
- Ding, Sheng
- Qi, Lei S.
- Venue
- Cell stem cell
- Publisher
- Elsevier BV
- Published
- 2015-02
- Volume
- 16
- Issue
- 2
- Pages
- 142-147
- ISSN
- 1875-9777 (ISSN-L)
- 1934-5909
- PMC
- PMC4461869
- OpenAlex
- W2007594606
- Cited by
- 455
- References
- 32
- Citations by year
- 2026: 11 · 2025: 20 · 2024: 10 · 2023: 25 · 2022: 31 · 2021: 38 · 2020: 76 · 2019: 53 · 2018: 43 · 2017: 59 · 2016: 62 · 2015: 26
- FWCI
- 27.51
- Type
- journal article
- Language
- en
- Open access
- bronze
- doi.org
- Licence
- Locations
- Cell stem cell (publishedVersion)
- PubMed (publishedVersion)
- PubMed Central (acceptedVersion)
- eScholarship (California Digital Library) (submittedVersion)
- Funders
- National Institutes of Health (DP5OD017887)
- National Heart, Lung, and Blood Institute (U01HL107436)
- National Institute on Drug Abuse (DA036858, R01DA036858)
- National Institute of General Medical Sciences (P50GM081879, T32GM008568)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › CRISPR and Genetic Engineering
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA regulation and disease
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › DNA Repair Mechanisms
- Keywords
- CRISPR/Cas9, small molecules, homology-directed repair, non-homologous end joining, genome editing efficiency, pluripotent stem cells, indel mutations, DNA repair, high-throughput screening
- MeSH terms
- DNA Repair, Dose-Response Relationship, Drug, Genetic Engineering, HeLa Cells, Humans, Structure-Activity Relationship, Molecular Structure, Genome, Human, Pluripotent Stem Cells, Small Molecule Libraries, High-Throughput Screening Assays, Clustered Regularly Interspaced Short Palindromic Repeats
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