Figure S1 . (B) DrugZ scores comparing relative enrichment/depletion sgRNAs targeting given genes in OVCAR8 cells treated with PARPi olaparib compared to DMSO across genome-wide screen. (C) DrugZ scores from (B) restricted to only druggable genes. (D–F) Depletion of UBA1 induces sensitivity to PARP inhibition. Cells were transfected with control siRNA, UBA1 siRNA1.1, and UBA1 siRNA1.2, treated with indicated doses of olaparib for 10 days, and then stained with crystal violet. Representative image of OVCAR8 cells shown (D). Quantification of relative cell viability by extraction of crystal violet for OVCAR8 (E) and HCC1806 (F) cells. Graphs represent mean +/− standard deviation. Two-way ANOVA with Holm-Sidak post hoc test. ∗∗∗∗ p < 0.0001. n = 3. See
Figure S2 . (G) Relative cell viability following treatment with varying concentrations of olaparib or TAK243 alone or the combination for 96 h in various ovarian cancer cell lines (OVCAR8, SKOV3, FUOV1, HOC1, and HOC8) and breast cancer cell lines (MDA-MB-468, HCC1806, BT549, and MCF7). The combination index (CI) was calculated with the Chou-Talalay equation. Experiments were performed in triplicates with two independent biological repeats ( n = 2). Graphs represent the mean with standard deviation. " width="100%" height="100%">
Journal: Cell Reports Medicine
Article Title: UBA1 inhibition sensitizes cancer cells to PARP inhibitors
doi: 10.1016/j.xcrm.2024.101834
Figure Lengend Snippet: Genome-wide CRISPR KO screen identifies UBA1 as a druggable target to enhance PARP inhibitor efficacy (A) Experimental outline of genome-wide CRISPR KO screen in OVCAR8 ovarian cancer cells (created with biorender.com ). See Figure S1 . (B) DrugZ scores comparing relative enrichment/depletion sgRNAs targeting given genes in OVCAR8 cells treated with PARPi olaparib compared to DMSO across genome-wide screen. (C) DrugZ scores from (B) restricted to only druggable genes. (D–F) Depletion of UBA1 induces sensitivity to PARP inhibition. Cells were transfected with control siRNA, UBA1 siRNA1.1, and UBA1 siRNA1.2, treated with indicated doses of olaparib for 10 days, and then stained with crystal violet. Representative image of OVCAR8 cells shown (D). Quantification of relative cell viability by extraction of crystal violet for OVCAR8 (E) and HCC1806 (F) cells. Graphs represent mean +/− standard deviation. Two-way ANOVA with Holm-Sidak post hoc test. ∗∗∗∗ p < 0.0001. n = 3. See Figure S2 . (G) Relative cell viability following treatment with varying concentrations of olaparib or TAK243 alone or the combination for 96 h in various ovarian cancer cell lines (OVCAR8, SKOV3, FUOV1, HOC1, and HOC8) and breast cancer cell lines (MDA-MB-468, HCC1806, BT549, and MCF7). The combination index (CI) was calculated with the Chou-Talalay equation. Experiments were performed in triplicates with two independent biological repeats ( n = 2). Graphs represent the mean with standard deviation.
Article Snippet: FUOV1 , DSMZ , ACC-444.
Techniques: Genome Wide, CRISPR, Inhibition, Transfection, Control, Staining, Extraction, Standard Deviation
Figures S3 A and S3B), triple-negative breast cancer cell lines (BT549 and HCC1806), and non-malignant mammary epithelial cells (MCF-10A) treated with olaparib, TAK243, or the combination for 10 days. Concentrations given in
Table S2 . Graphs represent mean +/− standard deviation. ANOVA with Holm-Sidak post hoc test. ∗∗∗∗ p < 0.0001. n = 3. See also
Figure S4 . (C and D) Representative image (C) and quantification (D) of patient-derived ovarian cancer organoids treated either with 1 μM olaparib, 10 nM TAK243, combination thereof, or DMSO vehicle control for 9 days. Scale bar, 1 mm. Representative time-lapsed quantification shown in
Figure S3 C. ANOVA with Holm-Sidak post hoc test. ∗∗ p < 0.01, ∗ p < 0.05. n = 3. " width="100%" height="100%">
Journal: Cell Reports Medicine
Article Title: UBA1 inhibition sensitizes cancer cells to PARP inhibitors
doi: 10.1016/j.xcrm.2024.101834
Figure Lengend Snippet: Validation of TAK243 and PARP inhibition in extended 2D and 3D assays (A and B) Representative images (A) and quantification (B) of ovarian cancer cell lines (OVCAR8, SKOV3, FUOV1, HOC8, and COV362 PARPi-R, see Figures S3 A and S3B), triple-negative breast cancer cell lines (BT549 and HCC1806), and non-malignant mammary epithelial cells (MCF-10A) treated with olaparib, TAK243, or the combination for 10 days. Concentrations given in Table S2 . Graphs represent mean +/− standard deviation. ANOVA with Holm-Sidak post hoc test. ∗∗∗∗ p < 0.0001. n = 3. See also Figure S4 . (C and D) Representative image (C) and quantification (D) of patient-derived ovarian cancer organoids treated either with 1 μM olaparib, 10 nM TAK243, combination thereof, or DMSO vehicle control for 9 days. Scale bar, 1 mm. Representative time-lapsed quantification shown in Figure S3 C. ANOVA with Holm-Sidak post hoc test. ∗∗ p < 0.01, ∗ p < 0.05. n = 3.
Article Snippet: FUOV1 , DSMZ , ACC-444.
Techniques: Biomarker Discovery, Inhibition, Standard Deviation, Derivative Assay, Control
Table S3 . See
Table S4 . (C) Western blot validation of RPPA results in HCC1806, OVCAR8, and FUOV1 cell lines, and in OVCAR8 and HCC1806 following treatment with indicated doses of TAK243. " width="100%" height="100%">
Journal: Cell Reports Medicine
Article Title: UBA1 inhibition sensitizes cancer cells to PARP inhibitors
doi: 10.1016/j.xcrm.2024.101834
Figure Lengend Snippet: TAK243 treatment downregulates DNA damage repair pathway-related proteins (A) Experimental outline of reverse-phase protein array (RPPA) experiment (created with biorender.com ). (B) Heatmap showing changes in protein expression determined by RPPA following 48 h treatment with TAK243 in 10 different ovarian, breast, and colorectal cell lines. Red color indicates upregulated proteins and blue color indicates downregulated proteins. One-sample t test using average change in protein expression for each cell line ( n = 10), with Benjamini-Hochberg procedure to correct for multiple comparisons. TAK243 concentrations given in Table S3 . See Table S4 . (C) Western blot validation of RPPA results in HCC1806, OVCAR8, and FUOV1 cell lines, and in OVCAR8 and HCC1806 following treatment with indicated doses of TAK243.
Article Snippet: FUOV1 , DSMZ , ACC-444.
Techniques: Protein Array, Expressing, Western Blot, Biomarker Discovery
Journal: Cell Reports Medicine
Article Title: UBA1 inhibition sensitizes cancer cells to PARP inhibitors
doi: 10.1016/j.xcrm.2024.101834
Figure Lengend Snippet:
Article Snippet: FUOV1 , DSMZ , ACC-444.
Techniques: Recombinant, CRISPR, Negative Control, Genome Wide, Software