Journal: Cancers

Article Title: WEE1 Inhibition Enhances Anti-Apoptotic Dependency as a Result of Premature Mitotic Entry and DNA Damage

doi: 10.3390/cancers11111743

Figure Lengend Snippet: Apoptosis induced by AZD1775 in diffuse large B-cell lymphoma (DLBCL). ( A ) Cell viability analysis was measured with propidium iodide flow cytometry in DLBCL cell lines treated with AZD1775 for 72 h. Data were plotted as the mean ± SD ( n = 3). ( B ) Representative flow cytometry of apoptosis (Annexin V/Propidium iodide) induced by treatment of AZD1775 for 18 h in representative cell lines SUDHL-10 and SC-1. ( C ) Mitochondrial response of DLBCL cell lines treated with AZD1775 for 18 h, plotted as the delta mitochondrial outer membrane permeabilization (ΔMOMP%). ΔMOMP% was calculated by subtracting the percentage treated MOMP from percentage untreated MOMP. Cell line SUDHL-10 was treated with 0.25 µM AZD1775, cell lines OCI-LY3, U-2932, SUDHL-2, and SUDHL-5 were treated with 0.5 µM AZD1775 and cell lines SUDHL-4, SUDHL-6, and SC-1 were treated with 1 µM AZD1775. Data were plotted as the mean ± SD ( n = 3). Statistical analysis was performed using a one-sample t -test as compared to untreated control cells (* p ≤ 0.05).

Article Snippet: Cells were incubated at 0.5 × 10 6 cells/mL for 18 h with AZD1775 (WEE1 inhibitor, Selleckchem No.S1525, Houston, TX, USA) , palbociclib (CDK4/6 inhibitor, Selleckchem No.S1116), RO3306 (CDK1 inhibitor, Selleckchem No.S7747), roscovitine (CDK2 inhibitor, Selleckchem No. S1153), nocodazole (M1404, Sigma Aldrich, St. Louis, MO, USA), or ultraviolet (UV) radiation (IBL 637, CisBioInternational, Gif-sur-Yvette, France).

Techniques: Flow Cytometry, Membrane, Control

Journal: Cancers

Article Title: WEE1 Inhibition Enhances Anti-Apoptotic Dependency as a Result of Premature Mitotic Entry and DNA Damage

doi: 10.3390/cancers11111743

Figure Lengend Snippet: Cellular effect of AZD1775 treatment in DLBCL. ( A ) Representative examples of flow cytometry cell cycle and γH2AX analysis of AZD1775 in SUDHL-10 and SC-1 treated for 18 h. ( B ) Correlation of the mitochondrial response (ΔMOMP%) for 10 µM NOXA in SUDHL-10 or 0.1 µM BAD in SC-1 versus the percentage of G1 phase cells treated with AZD1775. ( C ) Correlation of the mitochondrial response (ΔMOMP%) for 10 µM NOXA in SUDHL-10 or 0.1 µM BAD in SC-1 versus the percentage of S/G2/M phase cells treated with AZD1775. ( D ) Correlation of the mitochondrial response (ΔMOMP%) of 10 µM NOXA in SUDHL-10 or 0.1 µM BAD in SC-1 versus the percentage of γH2AX cells treated with AZD1775. Cells were treated with 0, 0.1, 0.2, 0.3, 0.4, 0.5, 0.75, or 1.0 µM AZD1775 for 18 h. ( E ) Representative examples of cell cycle flow cytometry and γH2AX analysis in SC-1 treated with 1 µM AZD1775, 50 ng/mL nocodazole, and 20 J/m 2 UV radiation. ( F ) Dynamic BH3 profile for 0.3 µM BIM, 10 µM NOXA, 10 µM HRK, and 0.1 µM BAD of SC-1 cells treated with AZD1775, nocodazole, and (ultra violet) UV radiation. Data were plotted as the mean ± SD ( n = 3). Statistical analysis was performed using a one-sample t -test as compared to untreated control cells (* p ≤ 0.05).

Article Snippet: Cells were incubated at 0.5 × 10 6 cells/mL for 18 h with AZD1775 (WEE1 inhibitor, Selleckchem No.S1525, Houston, TX, USA) , palbociclib (CDK4/6 inhibitor, Selleckchem No.S1116), RO3306 (CDK1 inhibitor, Selleckchem No.S7747), roscovitine (CDK2 inhibitor, Selleckchem No. S1153), nocodazole (M1404, Sigma Aldrich, St. Louis, MO, USA), or ultraviolet (UV) radiation (IBL 637, CisBioInternational, Gif-sur-Yvette, France).

Techniques: Flow Cytometry, Control

Journal: Cancers

Article Title: WEE1 Inhibition Enhances Anti-Apoptotic Dependency as a Result of Premature Mitotic Entry and DNA Damage

doi: 10.3390/cancers11111743

Figure Lengend Snippet: Rescue of AZD1775-induced cellular effects by CDK inhibitors RO3306, palbociclib and roscovitine. ( A ) Representative examples of apoptotic cells, cell cycle, and γH2AX flow cytometry analysis of SC-1 treated with 10 µM RO3306, 1 µM palbociclib, 15 µM roscovitine, and 1 µM AZD1775 after 18 h of incubation. ( B ) Dynamic BH3 profile for 0.3 µM BIM, 10 µM NOXA, 10 µM HRK, and 0.1 µM BAD of SC-1 cells treated with RO3306, palbociclib, roscovitine, and AZD1775 after 18 h of incubation. Delta mitochondrial outer membrane permeabilization (ΔMOMP%) was calculated by subtracting the percentage treated MOMP from percentage untreated MOMP. Data were plotted as the mean ± SD ( n = 3). Statistical analysis was performed using a one-sample t -test as compared to untreated control cells (* p ≤ 0.05).

Article Snippet: Cells were incubated at 0.5 × 10 6 cells/mL for 18 h with AZD1775 (WEE1 inhibitor, Selleckchem No.S1525, Houston, TX, USA) , palbociclib (CDK4/6 inhibitor, Selleckchem No.S1116), RO3306 (CDK1 inhibitor, Selleckchem No.S7747), roscovitine (CDK2 inhibitor, Selleckchem No. S1153), nocodazole (M1404, Sigma Aldrich, St. Louis, MO, USA), or ultraviolet (UV) radiation (IBL 637, CisBioInternational, Gif-sur-Yvette, France).

Techniques: Flow Cytometry, Incubation, Membrane, Control

Journal: Cancers

Article Title: WEE1 Inhibition Enhances Anti-Apoptotic Dependency as a Result of Premature Mitotic Entry and DNA Damage

doi: 10.3390/cancers11111743

Figure Lengend Snippet: Combination of AZD1775 and BH3 mimetic drugs in DLBCL cell lines. ( A ) Cell viability flow cytometry data for the cell lines SC-1, SUDHL-5, and SUDHL-10 pre-treated with 1 µM AZD1775 and incubated with increasing concentrations of venetoclax. Data were normalized to the control and plotted as the mean ± SD ( n = 4). ( B ) Venetoclax IC50 values as calculated from cell viability curves. Data were plotted as the mean ± SD ( n = 4). ( C ) Cell viability flow cytometry data for SC-1, SUDHL-5, and SUDHL-10 pre-treated with 1 µM AZD1775 and incubated with increasing concentrations of S63845. Data were normalized to the control and plotted as the mean ± SD ( n = 4). ( D ) S63845 IC50 values as calculated from cell viability curves. Data plotted as the mean ± SD ( n = 4). Statistical analysis was performed using a two-tailed paired t -test (* p ≤ 0.05) (** p ≤ 0.01).

Article Snippet: Cells were incubated at 0.5 × 10 6 cells/mL for 18 h with AZD1775 (WEE1 inhibitor, Selleckchem No.S1525, Houston, TX, USA) , palbociclib (CDK4/6 inhibitor, Selleckchem No.S1116), RO3306 (CDK1 inhibitor, Selleckchem No.S7747), roscovitine (CDK2 inhibitor, Selleckchem No. S1153), nocodazole (M1404, Sigma Aldrich, St. Louis, MO, USA), or ultraviolet (UV) radiation (IBL 637, CisBioInternational, Gif-sur-Yvette, France).

Techniques: Flow Cytometry, Incubation, Control, Two Tailed Test

Journal: Cancers

Article Title: WEE1 Inhibition Enhances Anti-Apoptotic Dependency as a Result of Premature Mitotic Entry and DNA Damage

doi: 10.3390/cancers11111743

Figure Lengend Snippet: Dynamic BH3 profile and validation experiments in a DLBCL patient treated with AZD1775. ( A ) Static BH3 profile and ( B ) dynamic BH3 profile of DLBCL patient cells treated with 0.5 µM AZD1775 for 18 h. Alamethicin and PUMA2A were used as positive and negative controls, respectively. Delta mitochondrial outer membrane permeabilization (ΔMOMP%) was calculated by subtracting the percentage treated MOMP from percentage untreated MOMP. ( C ) Cell viability flow cytometry data for DLBCL patient cells pre-treated with AZD1775 for 18 h and incubated with increasing concentrations of venetoclax and ( D ) navitoclax for 24 h. Data were normalized to the control.

Article Snippet: Cells were incubated at 0.5 × 10 6 cells/mL for 18 h with AZD1775 (WEE1 inhibitor, Selleckchem No.S1525, Houston, TX, USA) , palbociclib (CDK4/6 inhibitor, Selleckchem No.S1116), RO3306 (CDK1 inhibitor, Selleckchem No.S7747), roscovitine (CDK2 inhibitor, Selleckchem No. S1153), nocodazole (M1404, Sigma Aldrich, St. Louis, MO, USA), or ultraviolet (UV) radiation (IBL 637, CisBioInternational, Gif-sur-Yvette, France).

Techniques: Biomarker Discovery, Membrane, Flow Cytometry, Incubation, Control