Fig 2: TCGA-SKCM-based analysis validates the prognostic Bax, Bak, Smac signature.Independent validation of the prognostic signature at transcriptome level in the SKCM-TCGA metastatic sub-cohort. a Survival analysis in the SKCM-TCGA sub-cohort (n = 79 patients diagnosed with stage III or IV metastatic melanoma before 2010). An optimised chi-square-based cutoff was determined to divide patients with high (red) and low (blue) normalised BAX, BAK1 and DIABLO (Smac) expression (log2(FPKM-UQ + 1)). Kaplan–Meier curves (follow-up from sample procurement) were compared by log-rank test. b mRNA amounts for BAX, BAK1 and DIABLO (Smac) (blue: mRNA level below cutoff, red: mRNA level above cutoff). c Survival analysis in the metastatic TCGA-SKCM sub-cohort based on the expression profiles in Fig. 4b. Log-rank test for trend was used to compare three Kaplan–Meier curves representing patients with combined low BAX, BAK1 and DIABLO (Smac) expression (blue), combined high expression (red) or with mixed expression (black).

Fig 3: Pattern recognition allows predicting patient prognosis.a A principal component analysis was performed on the H scores of nine apoptotic proteins. To provide a visualisation of the spatial clustering, patients samples were positioned in a 3D scatter plot defined by the first three principal components and colour-coded according to their PFS time (red < 12 months, n = 33; blue > 12 months, n = 17). Linear discriminant analysis (LDA) correctly segmented 72% of the patients. Leave One Out Cross Validation (LOOCV) combined with LDA predicted the correct class for 74% of the patients. b 3D scatter plot showing the spatial clustering of patients with short and long PFS based on the H scores for Bax, Bak and Smac. LDA correctly segmented 80% of the patients and LOOCV-LDA achieved 78% prediction accuracy. c Comparison of the performance of the two classifiers shown in a and b. The receiver operating characteristic curves (ROCs) and respective areas under the curve (AUC) are shown.

Fig 4: Combination of ZD55-TIS and SNS-032 enhances suppression of tumor cell proliferation. Pancreatic cancer cells (A) PANC-1 and (B) BxPC-3 were treated with ZD55-TIS (1, 2, 4, 8 and 16 MOI), SNS-032 (5, 10, 20, 40 and 80 ng/ml), or ZD55-TIS plus SNS-032 for 48 h. The image represents three independent experiments. Cell viability was evaluated by MTT assay and the synergistic effect of ZD55-TIS combined with SNS-032 on PANC-1 and BxPC-3 was quantified by CI analysis and expressed as log (CI) vs. fractional effect. Where calculable, 95% confidence intervals are shown. **P<0.05. ZD55-TIS, ZD55-TRAIL-IETD-Smac; TRAIL, tumor necrosis factor-related apoptosis-inducing ligand; IETD, isoleucine-aspartate-threonine-glutamate; Smac, second mitochondria-derived activator of caspase; CI, combination index.

Fig 5: Manual scoring confirms association of low Bak, Bax and Smac protein expression with improved PFS.a Correlation between automated and manual H scores. Manual scores were obtained from two independent pathologists blinded to patient follow-up. Correlation was analysed using Spearman’s rank correlation coefficient. b Median H scores were used as cutoff to separate the patients with high (red line) and low (blue line) expression of each protein. Log-rank test was used to compare the Kaplan–Meier curves for progression-free survival from the date of sample procurement.