Journal: bioRxiv

Article Title: Unlocking DNA Damage Sensitivity of Cancer Cells: The Potential of Splicing Inhibitors

doi: 10.1101/2023.10.08.561421

Figure Lengend Snippet: A – Dose-response curves from MTT assays of SKOV3 cells treated with a fixed concentration of pladienolide B (1.56 nM) and various concentrations of carboplatin, cisplatin, doxorubicin, etoposide, or gemcitabine simultaneously for 48 hours. DMSO was used as a control instead of pladienolide B. B – Dose-response curves from MTT assays of SKOV3 cells pretreated with 1.56 nM pladienolide B or DMSO (control) for different durations (0, 6, 9, 12, 24, 48, 72, or 96 hours) followed by treatment with different concentrations of cisplatin for 48 hours. C – CellEvent Caspase-3/7 Green Flow Cytometry Assay of SKOV3 cells after different types of treatment. “Pl-B + CP” - pretreatment with pladienolide B (1.56 nM, 48 hours) followed by treatment with cisplatin (10 µM, 24 hours); “CP” - pretreatment with DMSO (48 hours) followed by treatment with cisplatin (10 µM, 24 hours); “Pl-B” - pretreatment with pladienolide B (1.56 nM, 48 hours) followed by fresh medium without pladienolide B (24 hours); “DMSO” - pretreatment with DMSO (48 hours) followed by fresh medium without DMSO (24 hours). D – Dose-response curves from MTT assays of various cell lines (MESOV, OVCAR3, TOV112D, TOV21G, A2789, Hep G2, HEY, HT29, A549, ID8, MDA-MB-231) treated with a fixed concentration of pladienolide B (1.56 nM) and different concentrations of cisplatin simultaneously for 48 hours. DMSO served as the control instead of pladienolide B. Each data point in A, B, D represents mean values ± SD (n = 3). IC50 values were determined by fitting a normalized model to data with nonlinear regression using GraphPad Prism software. Significance assessed via a paired, two-tailed Student’s t-test.

Article Snippet: Human ovarian cancer cell line OVCAR3 (ATCC, HTB-161) and murine colon carcinoma cell line CT26 (ATCC, CRL-2638) were grown in RPMI medium supplemented with 10% FBS, 2 mM L-glutamine, and 1% penicillin/streptomycin. hTERT-immortalized cells that were isolated from the fallopian tube of a female donor FT282 (ATCC, CRL-3449) was grown as adherent monolayers in DMEM medium supplemented with 10% FBS, 2 mM L-glutamine, and 1% penicillin/streptomycin.

Techniques: Concentration Assay, Control, Flow Cytometry, Software, Two Tailed Test

Journal: bioRxiv

Article Title: Unlocking DNA Damage Sensitivity of Cancer Cells: The Potential of Splicing Inhibitors

doi: 10.1101/2023.10.08.561421

Figure Lengend Snippet: A – Dose-response curves from MTT assays of various cell lines (MESOV, OVCAR3, TOV112D, TOV21G, A2780, Hep G2, HEY, HT29, A549, ID8, MDA-MB-231) pretreated with 1.56 nM pladienolide B (48 hours) followed by treatment with different concentrations of cisplatin for 48 hours. DMSO was used as the control instead of pladienolide B. B – Dose-response curves from MTT assays of human primary fibroblasts pretreated with 1.56 nM pladienolide B (48 hours) followed by treatment with different concentrations of cisplatin for 48 hours. C – Dose-response curves from MTT assays of FT282 cells pretreated with 1.56 nM pladienolide B (48 hours) followed by treatment with different concentrations of cisplatin for 48 hours. D-E – Dose-response curves from MTT assays of pladienolide B-resistant SKOV3 cells treated with different concentrations of pladienolide B (D) or cisplatin (E) for 48 hours. The method for generating resistant SKOV3 cells is described in the “Methods” section. F – Dose-response curves from MTT assays of SKOV3 cells treated with different concentrations of H3B-8800 (another splicing inhibitor) for 48 hours. G – Dose-response curves from MTT assays of SKOV3 (red line) and cisplatin-resistant SKOV3 (light red line) cells treated with different concentrations of cisplatin for 48 hours. H – Dose-response curves from MTT assays of cisplatin-resistant SKOV3 cells pretreated with 1.56 nM pladienolide B (48 hours) followed by treatment with different concentrations of cisplatin for 48 hours. I – Dose-response curves from MTT assays of SKOV3 cells pretreated with 50 nM H3B-8800 (48 hours) followed by treatment with different concentrations of cisplatin for 48 hours. J-K – Synergy landscapes for SKOV3 (J) and A549 (K) cells after simultaneous or sequential treatment with different doses of DMSO (y-axis) and cisplatin (x-axis). Zero-Interaction Potency (ZIP) synergy scores were calculated for the combination of DMSO in different volume range (according to Pl-B volume for 0 nM–6.24 nM concentration range) and cisplatin (0 µM–80 µM) for SKOV3 and A549 cells. For simultaneous regimen, сells were treated with DMSO and cisplatin simultaneously (48 hours). For sequential regimen, cells were pretreated with different volumes of DMSO (48 hours) followed by treatment with different concentrations of cisplatin for 48 hours. Cell survival was calculated in comparison to cisplatin untreated cells (DMSO only). ZIP values below 0 indicate antagonism (blue), 0 - 10 indicate additivity (from white to light red), and above 10 (corresponding to a deviation from the reference model above 10%) indicate synergy (dark red). L-M – Synergy landscapes for HepG2 (L) and HT29 (M) cells. Each data point represents mean values ± SD (n = 3). IC50 values were determined by fitting a normalized model to data with nonlinear regression using GraphPad Prism software.

Article Snippet: Human ovarian cancer cell line OVCAR3 (ATCC, HTB-161) and murine colon carcinoma cell line CT26 (ATCC, CRL-2638) were grown in RPMI medium supplemented with 10% FBS, 2 mM L-glutamine, and 1% penicillin/streptomycin. hTERT-immortalized cells that were isolated from the fallopian tube of a female donor FT282 (ATCC, CRL-3449) was grown as adherent monolayers in DMEM medium supplemented with 10% FBS, 2 mM L-glutamine, and 1% penicillin/streptomycin.

Techniques: Control, Concentration Assay, Comparison, Software

Journal: Cancer cell

Article Title: A Single Agent Dual Specificity Targeting of FOLR1 and DR5 as an Effective Strategy for Ovarian Cancer

doi: 10.1016/j.ccell.2018.07.005

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Human: OVCAR-3 , ATCC , HTB-161.

Techniques: Virus, Recombinant, Endotoxin Assay, Bioassay, Generated, Software, Plasmid Preparation, SYBR Green Assay, Protease Inhibitor, Transfection

Journal: eLife

Article Title: Identification of PARP-7 substrates reveals a role for MARylation in microtubule control in ovarian cancer cells

doi: 10.7554/eLife.60481

Figure Lengend Snippet: ( A–C ) Western blots showing PARP-7 protein levels after siRNA-mediated knockdown (KD) of PARP7 in HeLa ( A ), OVCAR3 ( B ) and A704 ( C ) cells. Two different siRNAs were used. α-tubulin was used as loading control. ( D–F ) The growth of HeLa ( D ), OVCAR3 ( E ), and A704 ( F ) cells with or without siRNA-mediated knockdown (KD) of PARP7 was assayed by crystal violet staining. Each point represents the mean ± SEM, n = 3. Asterisks indicate significant differences from the corresponding control (Student’s t-test; *p<0.05). ( G–I ) Cell migration assays ( top ) and cell invasion assays ( bottom ) for HeLa ( G ), OVCAR3 ( H ), and A704 ( I ) cells with or without siRNA-mediated knockdown (KD) of PARP7 . ( J–L ) Quantification of cell migration ( left ) and cell invasion ( right ) assays like those shown in ( G–I ). Asterisks indicate significant differences from the control (n = 3, Student’s t-test; **p<0.01).

Article Snippet: Cell line ( Homo sapiens ) , OVCAR3 (female adult ovarian cancer) , ATCC , RRID: CVCL_0465 , .

Techniques: Western Blot, Knockdown, Control, Staining, Migration

Journal: eLife

Article Title: Identification of PARP-7 substrates reveals a role for MARylation in microtubule control in ovarian cancer cells

doi: 10.7554/eLife.60481

Figure Lengend Snippet: Knockdown of PARP7 promotes microtubule stability. Immunofluorescent staining of α-Tubulin in HeLa ( A ), OVCAR3 ( B ), and A704 ( C ) cells after siRNA-mediated knockdown (KD) of PARP7 and treatment with cold or nocodazole. Scale bars = 25 µm. Each experiment was performed three times (n = 3) to ensure reproducibility.

Article Snippet: Cell line ( Homo sapiens ) , OVCAR3 (female adult ovarian cancer) , ATCC , RRID: CVCL_0465 , .

Techniques: Knockdown, Staining

Journal: eLife

Article Title: Identification of PARP-7 substrates reveals a role for MARylation in microtubule control in ovarian cancer cells

doi: 10.7554/eLife.60481

Figure Lengend Snippet:

Article Snippet: Cell line ( Homo sapiens ) , OVCAR3 (female adult ovarian cancer) , ATCC , RRID: CVCL_0465 , .

Techniques: Transfection, Construct, Binding Assay, Recombinant, Mutagenesis, Plasmid Preparation, Expressing, Software