Low cleaved caspase-7 levels indicate unfavourable outcome across all breast cancers.

Elevated levels of the anti-apoptotic BCL2 protein associate with favourable outcome in breast cancer. We investigated whether executioner caspase activation downstream of mitochondrial apoptosis was associated with, or independent, of BCL2's prognostic signature in breast cancer. Levels of pro- and anti-apoptotic BCL2 family proteins were quantified in triple negative breast cancer (TNBC) samples and utilised to calculate BCL2 profiles of 845 breast cancer patients. Biomarkers including single apoptosis proteins and network-enriched apoptosis system signatures were evaluated using uni- and multi-variate Cox-models. In both TNBC and non-TNBC breast cancer, the anti-apoptotic BCL2 protein was particularly abundant when compared to other solid tumours. High BCL2 protein levels were prognostic of favourable outcome across all breast cancers (HR 0.4, 95% CI 0.2-0.6, Wald p  KEY MESSAGE: BCL2 levels are elevated in breast cancer where they are marker of good prognosis. BCL2 and active caspase levels correlate negatively; yet, active caspases indicate good outcome. Low BCL2 and low caspase-7 are highly prognostic of unfavourable outcome across all breast cancers. BCL2 levels indicate molecular subtype and tumour proliferation status in breast cancer.</p

Systems analysis of protein signatures predicting Cetuximab responses in KRAS, NRAS, BRAF and PIK3CA wild-type patient-derived xenograft models of metastatic colorectal cancer

Antibodies targeting the human epidermal growth factor receptor (EGFR) are used for the treatment of RAS wild-type metastatic colorectal cancer. A significant proportion of patients remains unresponsive to this therapy. Here, we performed a reverse-phase protein array-based (phospho)protein analysis of 63 KRAS, NRAS, BRAF and PIK3CA wild-type metastatic CRC tumours. Responses of tumours to anti-EGFR therapy with cetuximab were recorded in patient-derived xenograft (PDX) models. Unsupervised hierarchical clustering of pretreatment tumour tissue identified three clusters, of which Cluster C3 was exclusively composed of responders. Clusters C1 and C2 exhibited mixed responses. None of the three protein clusters exhibited a significant correlation with transcriptome-based subtypes. Analysis of protein signatures across all PDXs identified 14 markers that discriminated cetuximab-sensitive and cetuximab-resistant tumours: PDK1 (S241), caspase-8, Shc (Y317), Stat3 (Y705), p27, GSK-3β (S9), HER3, PKC-α (S657), EGFR (Y1068), Akt (S473), S6 ribosomal protein (S240/244), HER3 (Y1289), NF-κB-p65 (S536) and Gab-1 (Y627). Least absolute shrinkage and selection operator and binominal logistic regression analysis delivered refined protein signatures for predicting response to cetuximab. (Phospo-)protein analysis of matched pretreated and posttreated models furthermore showed significant reduction of Gab-1 (Y627) and GSK-3β (S9) exclusively in responding models, suggesting novel targets for treatment

Systems biology analysis identifies molecular determinants of chemotherapy-induced diarrhoea

Chemotherapy-induced diarrhoea (CID) is a common dose-limiting adverse event in patients with cancer. Here, we hypothesise that chemotherapy evokes apoptosis in normal gut epithelium, contributes to CID and that patients with increased risk of CID can be identified using a systems model of BCL-2 protein interactions (DR_MOMP) that calculates the sensitivity of cells to undergo apoptosis. Normal adjacent gut epithelium tissue was collected during resection surgery from a cohort of 35 patients with stage II–III colorectal cancer (CRC) who were subsequently treated with capecitabine, XELOX or FOLFOX. Clinical follow-up, type and grade of adverse events during adjuvant chemotherapy were recorded. The level of five BCL-2 proteins required for the calculation of the DR_MOMP score was quantified together with 62 additional signalling proteins related to apoptotic pathways. Odds ratios for the occurrence of diarrhoea were determined using multinomial logistic regression (MLR). Patients treated with capecitabine who had a DR_MOMP score equal or higher than the mean had a significantly lower frequency of diarrhoea significantly compared to patients below the mean. High DR_MOMP scores indicate high apoptosis resistance. No statistical difference was observed in patients treated with XELOX or FOLFOX. Using MLR, we found that levels of apoptosis-related proteins caspase-8, p53 and XIAP statistically interacted with the DR_MOMP stress dose. Markers of MAPK signalling were prognostic for diarrhoea independently of DR_MOMP. In conclusion, apoptosis sensitivity and MAPK signalling status of the adjacent normal gut epithelium of chemotherapy-naïve patients represent promising biomarkers to identify patients with CRC with increased risk of CID

Molecular subtyping combined with biological pathway analyses to study regorafenib response in clinically relevant mouse models of colorectal cancer.

Purpose: Regorafenib (REG) is approved for the treatment of metastatic colorectal cancer, but has modest survival benefit and associated toxicities. Robust predictive/early response biomarkers to aid patient stratification are outstanding. We have exploited biological pathway analyses in a patient-derived xenograft (PDX) trial to study REG response mechanisms and elucidate putative biomarkers.Experimental design: Molecularly subtyped PDXs were annotated for REG response. Subtyping was based on gene expression (CMS, consensus molecular subtype) and copy-number alteration (CNA). Baseline tumor vascularization, apoptosis, and proliferation signatures were studied to identify predictive biomarkers within subtypes. Phospho-proteomic analysis was used to identify novel classifiers. Supervised RNA sequencing analysis was performed on PDXs that progressed, or did not progress, following REG treatment.Results:Improved REG response was observed in CMS4, although intra-subtype response was variable. Tumor vascularity did not correlate with outcome. In CMS4 tumors, reduced proliferation and higher sensitivity to apoptosis at baseline correlated with response. Reverse phase protein array (RPPA) analysis revealed 4 phospho-proteomic clusters, one of which was enriched with non-progressor models. A classification decision tree trained on RPPA- and CMS-based assignments discriminated non-progressors from progressors with 92% overall accuracy (97% sensitivity, 67% specificity). Supervised RNA sequencing revealed that higher basal EPHA2 expression is associated with REG resistance.Conclusions:Subtype classification systems represent canonical "termini a quo" (starting points) to support REG biomarker identification, and provide a platform to identify resistance mechanisms and novel contexts of vulnerability. Incorporating functional characterization of biological systems may optimize the biomarker identification process for multitargeted kinase inhibitors.</p