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Fig. 2 | Molecular Cancer

Fig. 2

From: Correction to: Transcriptional inhibition by CDK7/9 inhibitor SNS-032 abrogates oncogene addiction and reduces liver metastasis in uveal melanoma

Fig. 2

SNS-032 induces apoptosis in human UM cells. a and b UM cells were incubated with increasing concentrations of SNS-032 for 48 h, apoptosis was determined by flow cytometry after Annexin V-FITC/PI dual staining. a Representative flow dot plots for Omm2.3 are shown; b Quantitative analysis from three independent experiments. Data represent mean ± SEM. *, P < 0.05; **, P < 0.01; ***, P < 0.001, one-way ANOVA, post hoc intergroup comparisons, Tukey’s test. c Dose-dependent cleavage of PARP and activation of caspase-3 were detected by Western blot after exposure with SNS-032 for 48 h in UM cells. d Western blot analysis of levels of cytochrome c in the cytosolic fractionations was performed in UM cells treated with SNS-032 (1.0 μM) for the indicated durations. The cytosolic fractionations were not contaminated as indicated by a mitochondria marker COX II. e SNS-032 induced mitochondrial membrane depolarization in UM cells. 92.1 and Omm2.3 cells incubated with SNS-032 were stained with CMXRos and MTGreen, and then analyzed by flow cytometry. Results for three independent experiments are shown. ***, P < 0.001, one-way ANOVA, post hoc intergroup comparisons, Tukey’s test. f Western blot analysis of apoptosis-related proteins expression was performed in UM cells treated with SNS-032. g SNS-032 dose-dependently decreased mRNA levels of BIRC5 gene (encoding Survivin) in UM cells. *, P < 0.05; **, P < 0.01; ***, P < 0.001, one-way ANOVA, post hoc intergroup comparisons, Tukey’s test. h and i Overexpression of Survivin abrogated whereas knockdown of Survivin potentiated the SNS-032-induced apoptosis. h Omm1 cells were transduced with lentiviral Vector (pTSB) or Survivin (pTSB-Survivin) constructs for 48 h, then treated with SNS-032 for 24 h; i 92.1 cells were transduced with lentiviral Vector (Scramble) or Survivin shRNAs (shSurvivin #1 and #2) constructs for 48 h, then treated with SNS-032 for 24 h. The protein levels of PARP and Survivin were detected by Western blot analysis (top); cell death was examined by trypan blue exclusion assay (bottom). **, P < 0.01; ***, P < 0.001, Student’s t test

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