Journal: EMBO Molecular Medicine

Article Title: Targeting cIAP2 in a novel senolytic strategy prevents glioblastoma recurrence after radiotherapy

doi: 10.1038/s44321-025-00201-x

Figure Lengend Snippet: ( A ) List of BCL-2 and IAP family members showing fold changes (Log 2 FC) in gene expression in irradiated (IR) LN229 or A172 cells relative to mock-irradiated (mock) cells along with mRNA transcripts levels in counts per million (CPM), as assessed by RNA sequencing 10 days after irradiation with 10 Gy of X-rays ( n = 3). Genes with Log 2 fold change greater than 3 are highlighted in red. ( B ) MA plot showing abundance of transcripts represented by Log 2 CPM ( y axis) vs . Log 2 fold change in gene expression ( x axis) in irradiated cells relative to mock-irradiated cells ( n = 3 biological replicates per condition for each cell line). BCL-2 and IAP family gene members are denoted with numbers. Genes with log 2 fold change (Log 2 FC) cutoff of −1 and 1, and Log 2 CPM higher than −1 were considered differentially expressed, as denoted by dashed lines. ( C ) Kaplan–Meier curve showing correlation of higher BIRC3 expression levels with poor prognosis in GBMLGG patients in TCGA ( n = 606) and CGGA ( n = 657) cohorts as evidenced by Hazard Ratio (logrank) of 3.441 and 1.811, respectively. P < 0.0001 for both plots. ( D ) Plot shows mean relative expression of BIRC3 + /− SD in mock-irradiated vs. irradiated (IR) GBM cells 10 days after exposure to 10 Gy of X-rays, as assessed by qRT-PCR ( n = 3 biological replicates comprising three technical replicates). A two-tailed Student’s t test was performed; exact P values from left to right: 0.00001, 0.00048, 0.00397, 0.00012. ( E ) Whole cell extracts from mock-irradiated or irradiated GBM cell lines were western blotted with anti-cIAP2 antibody. Actin serves as loading control. ( F ) Senescent LN229 cells (10 days after exposure to 10 Gy) were treated with the IKK inhibitor BMS-345541 (BMS) or DMSO as control for 72 h ( n = 3 biological replicates comprising three technical replicates), and mean relative expression of BIRC3 + /− SD was assessed by qRT-PCR (a two-tailed Student’s t test was performed; P = 0.00000002, 0.00000003, respectively) or ( G ) western blotting for cIAP2. ( H ) Naive LN229 cells were exposed to conditioned media from senescent (CM-SEN) or non-senescent (CM-NS) cells for the indicated times, and expression of cIAP2 assessed by western blotting. ( I ) Naive LN229 cells were treated with BMS-345541 or DMSO as control for 2 h before exposure to CM-SEN ( n = 3 biological replicates comprising 3 technical replicates) and mean relative expression of BIRC3 + /− SD was assessed by qRT-PCR (a two-tailed Student’s t test was performed; P = 0.00001, 0.00002, respectively) or ( J ) western blotting for cIAP2. .

Article Snippet: Anti-c-IAP1 (D5G9) Rabbit mAb , Cell Signaling Technology , 7065.

Techniques: Gene Expression, Irradiation, RNA Sequencing, Expressing, Quantitative RT-PCR, Two Tailed Test, Western Blot, Control

Journal: EMBO Molecular Medicine

Article Title: Targeting cIAP2 in a novel senolytic strategy prevents glioblastoma recurrence after radiotherapy

doi: 10.1038/s44321-025-00201-x

Figure Lengend Snippet: ( A ) Kaplan–Meier curve showing lack of correlation of Bcl2l10 expression levels with prognosis in GBMLGG patients in TCGA ( n = 606) and CGGA ( n = 657) cohorts as evidenced by Hazard Ratio (logrank) of 0.9692 and 0.8629 and P values of 0.1943 and 0.2640, respectively. ( B ) Plot shows mean relative expression of BIRC2 + /− SD in mock-irradiated vs. irradiated (IR) GBM cells 10 days after exposure to 10 Gy of X-rays, as assessed by qRT-PCR ( n = 3 biological replicates comprising 3 technical replicates each). A two-tailed Student’s t test was performed; exact P values from left to right: 0.0000001, 0.0010112, 0.0044335, 0.9295618. ( C ) Whole cell extracts from mock-irradiated or irradiated GBM cell lines were western blotted with anti-cIAP1 antibody. Actin serves as loading control. ( D ) Senescent GBM cells (10 days after exposure to 10 Gy) were treated with the IKK inhibitor BMS-345541 (BMS) or DMSO as control for 72 h ( n = 3 biological replicates comprising 3 technical replicates each), and mean relative expression of BIRC3 + /− SD was assessed by qRT-PCR (a two-tailed Student’s t test was performed; A172 p = 0.00000003, 0.00000232; U118 p = 0.00000220, 0.00026887; U87 P = 0.00000818, 0.00118914, respectively) or ( E ) western blotting for cIAP2. ( F ) Naive GBM cells were exposed to conditioned media from senescent (CM-SEN) or non-senescent (CM-NS) cells for the indicated times, and expression of cIAP2 assessed by western blotting. ( G ) Naive GBM cells were treated with BMS-345541 or DMSO as control for 2 h before exposure to CM-SEN ( n = 3 biological replicates comprising 3 technical replicates each), and mean relative expression of BIRC3 + /− SD was assessed by qRT-PCR (a two-tailed Student’s t test was performed; A172 P = 0.000627, 0.000622; U118 P = 0.000001, 0.000001; U87 P = 0.000593, 0.000936, respectively) or ( H ) western blotting for cIAP2. ( I ) GBM cells were exposed to conditioned media from senescent or non-senescent cells ( n = 3) and radiation sensitivity measured by the colony survival assay. The mean percentage of surviving colonies +/− SD ( y axis) is plotted against the corresponding radiation dose ( x axis). .

Article Snippet: Anti-c-IAP1 (D5G9) Rabbit mAb , Cell Signaling Technology , 7065.

Techniques: Expressing, Irradiation, Quantitative RT-PCR, Two Tailed Test, Western Blot, Control, Clonogenic Cell Survival Assay

Journal: EMBO Molecular Medicine

Article Title: Targeting cIAP2 in a novel senolytic strategy prevents glioblastoma recurrence after radiotherapy

doi: 10.1038/s44321-025-00201-x

Figure Lengend Snippet: ( A ) GBM12 cells were irradiated with 10 Gy of X-rays and treated with the cIAP2 inhibitor birinapant (or DMSO as control) after 10 days for the indicated times, and cIAP2 levels and cleavage of caspase-8 were assessed by western blotting. Actin serves as loading control. ( B ) Whole cell extracts from mock-irradiated or irradiated GBM PDX cultures treated with DMSO or birinapant for 2 h were western blotted with anti-CIAP2 antibody. ( C ) Plots show mean relative expression of BIRC3 + /− SD in mock-irradiated (Mock) vs. irradiated (IR) PDX cultures 10 days after exposure to 10 Gy of X-rays, as assessed by qRT-PCR ( n = 3 biological replicates comprising three technical replicates). A two-tailed Student’s t test was performed; GBM6 P = 0.00004, GBM12 P = 0.00001, GBM43 P = 0.00015, GBM123 P = 0.00053, GBM148 P = 0.00026, GBM245 P = 0.00016. ( D ) Naive PDX cultures were exposed to conditioned media from senescent (CM-SEN) or non-senescent (CM-NS) cultures for the indicated times, and expression of cIAP2 assessed by western blotting. ( E ) Mock-irradiated or irradiated PDX cultures were treated with birinapant or DMSO as control ( n = 3–5 replicates per PDX) and viability was quantified by the MTT assay (normalized to that of DMSO-treated cells). The drug was replaced every 72 h. Plots show mean viability +/− SD. .

Article Snippet: Anti-c-IAP1 (D5G9) Rabbit mAb , Cell Signaling Technology , 7065.

Techniques: Irradiation, Control, Western Blot, Expressing, Quantitative RT-PCR, Two Tailed Test, MTT Assay

Journal: EMBO Molecular Medicine

Article Title: Targeting cIAP2 in a novel senolytic strategy prevents glioblastoma recurrence after radiotherapy

doi: 10.1038/s44321-025-00201-x

Figure Lengend Snippet: Reagents and tools table

Article Snippet: Anti-c-IAP1 (D5G9) Rabbit mAb , Cell Signaling Technology , 7065.

Techniques: Biomarker Discovery, Purification, Sequencing, Negative Control, Plasmid Preparation, Fluorsave, SYBR Green Assay, Western Blot, Stripping, Software

Journal: Scientific Reports

Article Title: Inhibition of AKT enhances chemotherapy efficacy and synergistically interacts with targeting of the Inhibitor of apoptosis proteins in oesophageal adenocarcinoma

doi: 10.1038/s41598-024-83912-4

Figure Lengend Snippet: Characterisation of IAP proteins in OAC cells. ( A ) Western blot analysis of cleaved PARP, cIAP1, cIAP2, XIAP and Survivin following 24 h treatment with ~ IC 30(72 h) doses of chemotherapy in OE33, FLO1 and OE19 cell lines. Western blot analysis of basal IAP protein levels in ( B ) paired sensitive and resistant OAC cell lines and ( C ) an OAC cell line panel. Vinculin was used as a loading control ( D ) Pearson correlation analysis of wild-type XIAP protein levels and ALM301 sensitivity, ~ IC50 (72 h) in the OAC cell line panel (r = 0.79, p < 0.05). XIAP mutant KYAE1 cell line was omitted from the analysis due to predicted functionality effect.

Article Snippet: Western blot analysis was carried out as previously described using antibodies targeting phospho-(S473)-Akt (Cell Signaling Technology Cat# 4060, RRID:AB_2315049), phospho-(T308)-Akt (Cell Signaling Technology Cat# 4056, RRID:AB_331163), pan- Akt (Cell Signaling Technology Cat# 2920, RRID:AB_1147620), PTEN (Cell Signaling Technology Cat# 9559, RRID:AB_390810), PARP (Cell Signaling Technology Cat# 9532, RRID:AB_659884), phospho-γ-H2AX (Cell Signaling Technology Cat# 9718, RRID:AB_2118009), cIAP1 (Cell Signaling Technology Cat# 7065, RRID:AB_10890862), cIAP2 (Cell Signaling Technology Cat# 3130, RRID:AB_10693298), XIAP (Cell Signaling Technology Cat# 2045, RRID:AB_2214866) and Survivin (Cell Signaling Technology Cat# 2808, RRID:AB_2063948).

Techniques: Western Blot, Control, Mutagenesis

Journal: Scientific Reports

Article Title: Inhibition of AKT enhances chemotherapy efficacy and synergistically interacts with targeting of the Inhibitor of apoptosis proteins in oesophageal adenocarcinoma

doi: 10.1038/s41598-024-83912-4

Figure Lengend Snippet: IAP antagonist activity in OAC cells ( A ) Western blot analysis of cleaved PARP, cIAP1, cIAP2, XIAP and Survivin protein levels following 24 h treatment with SMC, BV6. Vinculin was used as a loading control. ( B ) MTT analysis of OAC cell viability following 72 h treatment with BV6.

Article Snippet: Western blot analysis was carried out as previously described using antibodies targeting phospho-(S473)-Akt (Cell Signaling Technology Cat# 4060, RRID:AB_2315049), phospho-(T308)-Akt (Cell Signaling Technology Cat# 4056, RRID:AB_331163), pan- Akt (Cell Signaling Technology Cat# 2920, RRID:AB_1147620), PTEN (Cell Signaling Technology Cat# 9559, RRID:AB_390810), PARP (Cell Signaling Technology Cat# 9532, RRID:AB_659884), phospho-γ-H2AX (Cell Signaling Technology Cat# 9718, RRID:AB_2118009), cIAP1 (Cell Signaling Technology Cat# 7065, RRID:AB_10890862), cIAP2 (Cell Signaling Technology Cat# 3130, RRID:AB_10693298), XIAP (Cell Signaling Technology Cat# 2045, RRID:AB_2214866) and Survivin (Cell Signaling Technology Cat# 2808, RRID:AB_2063948).

Techniques: Activity Assay, Western Blot, Control