Fig. 5: Entinostat enhances apoptosis in response to IAP antagonists via Ku70, RIPK1 and caspase-8. | Cell Death & Disease

Fig. 5: Entinostat enhances apoptosis in response to IAP antagonists via Ku70, RIPK1 and caspase-8.

From: Cytoplasmic FLIP(S) and nuclear FLIP(L) mediate resistance of castrate-resistant prostate cancer to apoptosis induced by IAP antagonists

Fig. 5

a Western blot of Ku70 in PC3, DU145 and VCaP cells following immunoprecipitation with anti-acetylated Lysine (AcK) or IgG isotype control antibody after 24 h treatment with 2.5 µM Entinostat(Ent) (Left). Right: Western blot of FLIP, Ku70, Hyperacetylated Histone4 (Ac-H4) in PC3, DU145 and VCaP cells treated for 24 h with 2.5 µM Entinostat (Ent). b Western blot of FLIP, Ku70 and HSP90 and PARP in nuclear and cytoplasmic fractions from PC3 and DU145 cells treated for 24 h with 2.5 μM Entinostat(ENT) or transfected with 20 nM scrambled control(SC) or Ku70 siRNA. c Annexin-V/PI flow cytometry analysis of PC3 cells transfected for 24 h with 20 nM scrambled control (SC) or RIPK1 siRNA followed by 24 h of 2.5 µM Entinostat(Ent) and a further 24 h with 1 µM TL32711 and 10 ng/mL TNFα combination. Western blot analysis of RIPK1 and β-actin in PC3 cells treated with 10 nM SC or RIPK1 siRNA. d Annexin-V/PI flow cytometry (Left)and cell viability assay (Right) in PC3 cells pre-treated for 1 h with 20 µM Necrostatin-1 (RIPK1 kinase inhibitor), GSK’840 (RIPK3 kinase inhibitor Necrosulfonamide (MLKL inhibitor) followed by pre-treatment for 24 h with 2.5 µM Entinostat(Ent) and a further 24 h with 1 µM TL32711 and 10 ng/mL TNFα combination. e Annexin-V/PI flow cytometry (Left) and cell viability assay (Right) in PC3 cells pre-treated for 1 h with 20 µM z-VAD-fmk followed by pre-treatment for 24 h with 2.5 µM Entinostat(Ent) and a further 24 h with 1 µM TL32711 and 10 ng/mL TNFα combination. f Annexin-V/PI flow cytometry analysis of PC3 empty vector(EV) and procaspase-8(C8) CRISPR cells following pre-treatment for 24 h with 2.5 µM Entinostat(Ent) and a further 24 h with 1 µM TL32711 and 10 ng/mL TNFα combination. *p ≤ 0.05, **p ≤ 0.01 and ***p ≤ 0.001

Back to article page