polqi1 Search Results


polqi1  (MedChemExpress)
N/A
PolQi1 is a selective inhibitor targeting the Polθ domain of DNA polymerase. PolQi1 inhibits the Polθ-mediated microhomology end joining (TMEJ/alt-EJ) pathway, reducing insertion/deletion (Indels) and imprecise editing events during DNA repair. PolQi1 can enhance the
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polqi1  (TargetMol)
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polqi2 (wo2020243459  (AstraZeneca ltd)
90
AstraZeneca ltd polqi2 (wo2020243459
( A-C ) Editing efficiencies of different repair events at gMej target site with indicated donor DNA for HEK293T wild-type (wt) cells and three knockout pools of enzymes involved in MMEJ repair. TIDE analysis estimated knockout efficiencies of cell pools were PARP1 = 94%, POLQ = 92% and LIG3 = 88%. Experiments were performed with 1 μM AZD7648 or DMSO control treatment 3 hours before plasmid transfections. Horizontal lines illustrate the mean knock-in efficiency in DMSO or AZD7648 treated wt cells. Bar graphs show mean values ± SD (n=3, technical replicates). ( D ) Knock-in efficiencies of ssDNA donor integration at selected target sites with 1 μM AZD7648 or DMSO for HEK293T wt and POLQ -KO cell pools. Bar graphs represent mean values ± SD (n=3, technical replicates). Statistical differences were evaluated using Student’s paired t test (two-tailed) * P < 0.05, ** P < 0.01, *** P < 0.001. ( E ) Frequencies of different repair events at gMej target site in plasmid and ssDNA transfected HEK293T cells. Cells were treated 1-3 hours before transfections with DMSO,1 μM AZD7648, and 1 μM AZD7648 in combination with 3 μM Pol⊖ inhibitor PolQi1 or <t>PolQi2.</t> Bar graphs represent mean values ± SD (n=3, biological replicates). ( F ) Percentage of different repair events in Sp Cas9-inducible hiPSC transfected with sgRNA gMej in presence of 1 μM DNA-PK inhibitor AZD7648, or 1 μM AZD7648 in combination with 3 μM PolQi1 or PolQi2. Bar graphs depict mean editing efficiencies ± SD (n=5, biological replicates).
Polqi2 (Wo2020243459, supplied by AstraZeneca ltd, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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( A-C ) Editing efficiencies of different repair events at gMej target site with indicated donor DNA for HEK293T wild-type (wt) cells and three knockout pools of enzymes involved in MMEJ repair. TIDE analysis estimated knockout efficiencies of cell pools were PARP1 = 94%, POLQ = 92% and LIG3 = 88%. Experiments were performed with 1 μM AZD7648 or DMSO control treatment 3 hours before plasmid transfections. Horizontal lines illustrate the mean knock-in efficiency in DMSO or AZD7648 treated wt cells. Bar graphs show mean values ± SD (n=3, technical replicates). ( D ) Knock-in efficiencies of ssDNA donor integration at selected target sites with 1 μM AZD7648 or DMSO for HEK293T wt and POLQ -KO cell pools. Bar graphs represent mean values ± SD (n=3, technical replicates). Statistical differences were evaluated using Student’s paired t test (two-tailed) * P < 0.05, ** P < 0.01, *** P < 0.001. ( E ) Frequencies of different repair events at gMej target site in plasmid and ssDNA transfected HEK293T cells. Cells were treated 1-3 hours before transfections with DMSO,1 μM AZD7648, and 1 μM AZD7648 in combination with 3 μM Pol⊖ inhibitor PolQi1 or PolQi2. Bar graphs represent mean values ± SD (n=3, biological replicates). ( F ) Percentage of different repair events in Sp Cas9-inducible hiPSC transfected with sgRNA gMej in presence of 1 μM DNA-PK inhibitor AZD7648, or 1 μM AZD7648 in combination with 3 μM PolQi1 or PolQi2. Bar graphs depict mean editing efficiencies ± SD (n=5, biological replicates).

Journal: bioRxiv

Article Title: Simultaneous inhibition of DNA-PK and Polϴ improves integration efficiency and precision of genome editing

doi: 10.1101/2022.12.15.520396

Figure Lengend Snippet: ( A-C ) Editing efficiencies of different repair events at gMej target site with indicated donor DNA for HEK293T wild-type (wt) cells and three knockout pools of enzymes involved in MMEJ repair. TIDE analysis estimated knockout efficiencies of cell pools were PARP1 = 94%, POLQ = 92% and LIG3 = 88%. Experiments were performed with 1 μM AZD7648 or DMSO control treatment 3 hours before plasmid transfections. Horizontal lines illustrate the mean knock-in efficiency in DMSO or AZD7648 treated wt cells. Bar graphs show mean values ± SD (n=3, technical replicates). ( D ) Knock-in efficiencies of ssDNA donor integration at selected target sites with 1 μM AZD7648 or DMSO for HEK293T wt and POLQ -KO cell pools. Bar graphs represent mean values ± SD (n=3, technical replicates). Statistical differences were evaluated using Student’s paired t test (two-tailed) * P < 0.05, ** P < 0.01, *** P < 0.001. ( E ) Frequencies of different repair events at gMej target site in plasmid and ssDNA transfected HEK293T cells. Cells were treated 1-3 hours before transfections with DMSO,1 μM AZD7648, and 1 μM AZD7648 in combination with 3 μM Pol⊖ inhibitor PolQi1 or PolQi2. Bar graphs represent mean values ± SD (n=3, biological replicates). ( F ) Percentage of different repair events in Sp Cas9-inducible hiPSC transfected with sgRNA gMej in presence of 1 μM DNA-PK inhibitor AZD7648, or 1 μM AZD7648 in combination with 3 μM PolQi1 or PolQi2. Bar graphs depict mean editing efficiencies ± SD (n=5, biological replicates).

Article Snippet: AZ31, AZ20, AZD2281, AZD2811, AZD7648, TLR1 (WO2014183850), M9831/VX-984, FEN-IN-1, PolQi1 (WO2021028643), PolQi2 (WO2020243459) were provided by AstraZeneca (Gothenburg, SE).

Techniques: Knock-Out, Plasmid Preparation, Transfection, Knock-In, Two Tailed Test

( A ) Heatmaps show mean editing efficiencies (InDel frequencies) for HEK3 and HEK4 on- and off-target sites (n=3, technical replicates). Cells were treated with DMSO, 1 μM AZD7648 and 1 μM AZD7648 with 3 μM PolQi1 or PolQi2. Ctrl refers to non-targeting control guide. Divergent bases are indicated on the right (red small letters). ( B ) Quantification of large deletions from long read sequencing data of HBEGF -edited HEK293T cells treated with indicated compounds in following concentrations 1 μM AZD7648, 3 μM PolQi1, 3 μM PolQi2. Bar graphs present percentage of deletion index (n=1). ( C ) Simultaneous genome editing in HEK293T cells at HBEGF and PCSK9 loci for translocation analysis in presence of inhibitors (1 μM AZD7648, 3 μM PolQi1 and 3 μM PolQi2) or DMSO control. Dot-plots depict mean translocation frequencies analysed with ddPCR ± SD (n=3, technical replicates).

Journal: bioRxiv

Article Title: Simultaneous inhibition of DNA-PK and Polϴ improves integration efficiency and precision of genome editing

doi: 10.1101/2022.12.15.520396

Figure Lengend Snippet: ( A ) Heatmaps show mean editing efficiencies (InDel frequencies) for HEK3 and HEK4 on- and off-target sites (n=3, technical replicates). Cells were treated with DMSO, 1 μM AZD7648 and 1 μM AZD7648 with 3 μM PolQi1 or PolQi2. Ctrl refers to non-targeting control guide. Divergent bases are indicated on the right (red small letters). ( B ) Quantification of large deletions from long read sequencing data of HBEGF -edited HEK293T cells treated with indicated compounds in following concentrations 1 μM AZD7648, 3 μM PolQi1, 3 μM PolQi2. Bar graphs present percentage of deletion index (n=1). ( C ) Simultaneous genome editing in HEK293T cells at HBEGF and PCSK9 loci for translocation analysis in presence of inhibitors (1 μM AZD7648, 3 μM PolQi1 and 3 μM PolQi2) or DMSO control. Dot-plots depict mean translocation frequencies analysed with ddPCR ± SD (n=3, technical replicates).

Article Snippet: AZ31, AZ20, AZD2281, AZD2811, AZD7648, TLR1 (WO2014183850), M9831/VX-984, FEN-IN-1, PolQi1 (WO2021028643), PolQi2 (WO2020243459) were provided by AstraZeneca (Gothenburg, SE).

Techniques: Sequencing, Translocation Assay

( A-C ) Quantification of luminescence signal via HiBiT lytic assay to estimate protein tagging efficiencies with HiBiT and Halo-HiBiT tags at indicated genomic loci and cell lines. Experiments were performed with 1 μM AZD7648, and 1 μM AZD7648 in combination with 3 μM PolQi1 or PolQi2. Data represent mean luminescence signal after background subtraction ± SD (Hela HiBiT, n=4; Jurkat HiBiT, n=3; Jurkat HaloTag-HiBiT, n=3; all technical replicates). ( D ) Bar graphs represent percentage of Jurkat cell clones containing HaloTag-HiBiT knock-in assessed with the Nano-Glo HiBiT lytic detection system for the indicated target genes. Cells were treated with DMSO or 1 μM AZD7648 and 3 μM PolQi2. Numbers show positive / analysed clones ( E ) ddPCR analysis of HaloTag-HiBiT positive Jurkat cell clones. Bar graphs show percentage of homozygous HaloTag-HiBiT integration at different loci using DMSO or 1 μM AZD7648 and 3 μM PolQi2. Numbers show homozygous / heterozygous clones. ( F ) Flow cytometry analysis to measure GFP integration into the TRAC locus in primary human CD3+ T cells derived from three individual human donors in presence of 1 μM AZD7648, and 1 μM AZD7648 in combination with 3 μM PolQi1 or PolQi2. KI efficiencies were evaluated seven days post electroporation. Data represent percentage of GFP positive cells in the viable cell population.

Journal: bioRxiv

Article Title: Simultaneous inhibition of DNA-PK and Polϴ improves integration efficiency and precision of genome editing

doi: 10.1101/2022.12.15.520396

Figure Lengend Snippet: ( A-C ) Quantification of luminescence signal via HiBiT lytic assay to estimate protein tagging efficiencies with HiBiT and Halo-HiBiT tags at indicated genomic loci and cell lines. Experiments were performed with 1 μM AZD7648, and 1 μM AZD7648 in combination with 3 μM PolQi1 or PolQi2. Data represent mean luminescence signal after background subtraction ± SD (Hela HiBiT, n=4; Jurkat HiBiT, n=3; Jurkat HaloTag-HiBiT, n=3; all technical replicates). ( D ) Bar graphs represent percentage of Jurkat cell clones containing HaloTag-HiBiT knock-in assessed with the Nano-Glo HiBiT lytic detection system for the indicated target genes. Cells were treated with DMSO or 1 μM AZD7648 and 3 μM PolQi2. Numbers show positive / analysed clones ( E ) ddPCR analysis of HaloTag-HiBiT positive Jurkat cell clones. Bar graphs show percentage of homozygous HaloTag-HiBiT integration at different loci using DMSO or 1 μM AZD7648 and 3 μM PolQi2. Numbers show homozygous / heterozygous clones. ( F ) Flow cytometry analysis to measure GFP integration into the TRAC locus in primary human CD3+ T cells derived from three individual human donors in presence of 1 μM AZD7648, and 1 μM AZD7648 in combination with 3 μM PolQi1 or PolQi2. KI efficiencies were evaluated seven days post electroporation. Data represent percentage of GFP positive cells in the viable cell population.

Article Snippet: AZ31, AZ20, AZD2281, AZD2811, AZD7648, TLR1 (WO2014183850), M9831/VX-984, FEN-IN-1, PolQi1 (WO2021028643), PolQi2 (WO2020243459) were provided by AstraZeneca (Gothenburg, SE).

Techniques: Clone Assay, Knock-In, Flow Cytometry, Derivative Assay, Electroporation