MicroRNA-210 induces apoptosis in colorectal cancer via induction of reactive oxygen

Additional file 2: Figure S2. Representative flow cytometric histograms of CRC cell lines 72 h post transfection with pre-miR-210 and a control miRNA, respectively. Cells were stained with by PI staining and subjected to FACS analysis

Senescence-associated molecules and tumor-immune-interactions as prognostic biomarkers in colorectal cancer

Background and Aims: The initiation of cellular senescence in response to protumorigenic stimuli counteracts malignant progression in (pre)malignant cells. Besides arresting proliferation, cells entering this terminal differentiation state adopt a characteristic senescence-associated secretory phenotype (SASP) which initiates alterations to their microenvironment and effects immunosurveillance of tumorous lesions. However, some effects mediated by senescent cells contribute to disease progression. Currently, the exploration of senescent cells' impact on the tumor microenvironment and the evaluation of senescence as possible target in colorectal cancer (CRC) therapy demand reliable detection of cellular senescence in vivo. Therefore, specific immunohistochemical biomarkers are required. Our aim is to analyze the clinical implications of senescence detection in colorectal carcinoma and to investigate the interactions of senescent tumor cells and their immune microenvironment in vitro and in vivo. Methods: Senescence was induced in CRC cell lines by low-dose-etoposide treatment and confirmed by Senescence-associated β-galactosidase (SA-β-GAL) staining and fluorescence activated cell sorting (FACS) analysis. Co-cultures of senescent cells and immune cells were established. Multiple cell viability assays, electron microscopy and live cell imaging were conducted. Immunohistochemical (IHC) markers of senescence and immune cell subtypes were studied in a cohort of CRC patients by analyzing a tissue micro array (TMA) and performing digital image analysis. Results were compared to disease-specific survival (DSS) and progression-free survival (PFS). Results: Varying expression of senescence markers in tumor cells was associated with in- or decreased survival of CRC patients. Proximity analysis of p21-positive senescent tumor cells and cytotoxic T cells revealed a significantly better prognosis for patients in which these cell types have the possibility to directly interact. In vitro, NK-92 cells (mimicking natural killer T cells) or TALL-104 cells (mimicking both cytotoxic T cells and natural killer T cells) led to dose-dependent specific cytotoxicity in >75 % of the senescent CRC cells but <20 % of the proliferating control CRC cells. This immune cell-mediated senolysis seems to be facilitated via direct cell-cell contact inducing apoptosis and granule exocytosis. Conclusion: Counteracting tumorigenesis, cellular senescence is of significant relevance in CRC. We show the dual role of senescence bearing both beneficial and malignancy-promoting potential in vivo. Absence as well as exceeding expression of senescence markers are associated with bad prognosis in CRC. The antitumorigenic potential of senescence induction is determined by tumor micromilieu and immune cell-mediated elimination of senescent cells

External validation of molecular subtype classifications of colorectal cancer based on microsatellite instability, CIMP, BRAF and KRAS

Background: Competing molecular classification systems have been proposed to complement the TNM staging system for a better prediction of survival in colorectal cancer (CRC). However, validation studies are so far lacking. The aim of this study was to validate and extend previously published molecular classifications of CRC in a large independent cohort of CRC patients. Methods: CRC patients were recruited into a population-based cohort study (DACHS). Molecular subtypes were categorized based on three previously published classifications. Cox-proportional hazard models, based on the same set of patients and using the same confounders as reported by the original studies, were used to determine overall, cancer-specific, or relapse-free survival for each subtype. Hazard ratios and confidence intervals, as well as Kaplan-Meier plots were compared to those reported by the original studies. Results: We observed similar patterns of worse survival for the microsatellite stable (MSS)/BRAF-mutated and MSS/KRAS-mutated subtypes in our validation analyses, which were included in two of the validated classifications. Of the two MSI subtypes, one defined by additional presence of CIMP-high and BRAF-mutation and the other by tumors negative for CIMP, BRAF and KRAS-mutations, we could not confirm associations with better prognosis as suggested by one of the classifications. For two of the published classifications, we were able to provide results for additional subgroups not included in the original studies (men, other disease stages, other locations). Conclusions: External validation of three previously proposed classifications confirmed findings of worse survival for CRC patients with MSS subtypes and BRAF or KRAS mutations. Regarding MSI subtypes, other patient characteristics such as stage of the tumor, may influence the potential survival benefit. Further integration of methylation, genetic, and immunological information is needed to develop and validate a comprehensive classification that will have relevance for use in clinical practice

A prognostic CpG score derived from epigenome-wide profiling of tumor tissue was independently associated with colorectal cancer survival

Background: Results of previous studies on the association of the CpG island methylator phenotype (CIMP) with colorectal cancer (CRC) prognosis were inconsistent and mostly based on different CIMP definitions. The current study aimed to comprehensively investigate the associations between DNA methylation on genes previously used to define CIMP status with CRC survival. Results: Patients with CRC followed up for a median of 5.2 years were divided into a study cohort (n = 568) and a validation cohort (n = 308). DNA methylation was measured in tumor tissue using the Illumina Infinium HumanMethylation450 BeadChip and restricted to 43 genes used to define CIMP status in previous studies. Cox proportional hazard regression models were used to estimate adjusted hazard ratios (HR) and 95% confidence intervals (CI) of survival after CRC, including adjustment for tumor stage, microsatellite instability, and BRAF mutation status. In the study cohort, ten CpG sites were identified to be associated with CRC survival. Seven of these ten CpG sites were also associated with CRC survival in the validation cohort and were used to construct a prognostic score. CRC patients with a prognostic score of the lowest methylation level showed poorer disease-specific survival compared with patients with the highest methylation level in both the study cohort and the validation cohort (HR = 3.11 and 95% CI = 1.97–4.91, and HR = 3.06 and 95% CI = 1.71–5.45, respectively). Conclusions: A CpG panel consisting of seven CpG sites was found to be strongly associated with CRC survival, independent from important clinical factors and mutations associated with CIMP. Further studies are required to confirm these findings

Value of c-MET and Associated Signaling Elements for Predicting Outcomes and Targeted Therapy in Penile Cancer

Whereas the lack of biomarkers in penile cancer (PeCa) impedes the development of efficacious treatment protocols, preliminary evidence suggests that c-MET and associated signaling elements may be dysregulated in this disorder. In the following study, we investigated whether c-MET and associated key molecular elements may have prognostic and therapeutic utility in PeCa. Formalin-fixed, paraffin-embedded tumor tissue from therapy-naïve patients with invasive PeCa was used for tissue microarray (TMA) analysis. Immunohistochemical staining was performed to determine the expression of the proteins c-MET, PPARg, β-catenin, snail, survivin, and n-MYC. In total, 94 PeCa patients with available tumor tissue were included. The median age was 64.9 years. High-grade tumors were present in 23.4%, and high-risk HPV was detected in 25.5%. The median follow-up was 32.5 months. High expression of snail was associated with HPV-positive tumors. Expression of β-catenin was inversely associated with grading. In both univariate COX regression analysis and the log-rank test, an increased expression of PPARg and c-MET was predictive of inferior disease-specific survival (DSS). Moreover, in multivariate analysis, a higher expression of c-MET was independently associated with worse DSS. Blocking c-MET with cabozantinib and tivantinib induced a significant decrease in viability in the primary PeCa cell line UKF-PeC3 isolated from the tumor tissue as well as in cisplatin- and osimertinib-resistant sublines. Strikingly, a higher sensitivity to tivantinib could be detected in the latter, pointing to the promising option of utilizing this agent in the second-line treatment setting

Expression and Functional Characterization of the BNIP3 Protein in Renal Cell Carcinomas

BNIP3 (Bcl-2/adenovirus E1B 19-kDa interacting protein 3) is a BH3-only protein that regulates apoptosis and autophagy. BNIP3 plays also an important role in hypoxia-induced cell response and is regulated by HIF1. Here, we studied a possible association of BNIP3 expression and the prognosis of patients with renal cell carcinomas (RCCs) and examined the functional relevance of BNIP3 in the regulation of cell survival and apoptosis of renal carcinoma cells. BNIP3 expression was determined by immunohistochemistry in RCC tumor tissue samples of 569 patients using a tissue microarray. Functional characterization of BNIP3 in renal carcinoma cells indicates prosurvival effects. In human RCC tumor samples, high cytoplasmic BNIP3 expression was associated with high-grade RCCs and regional lymph node metastasis. BNIP3 expression correlated negatively with disease-specific survival. Multivariate Cox regression analysis retained BNIP3 expression as an independent prognostic factor in patients without distant metastasis. Together, our studies imply that BNIP3 regulates cell survival in RCCs and its expression is an independent prognostic marker in patients with localized RCCs

Decoy Receptor 3 Is a Prognostic Factor in Renal Cell Cancer12

AbstractBACKGROUND: Decoy receptor 3 (DcR3) is a soluble protein that binds to and inactivates the death ligand CD95L. Here, we studied a possible association between DcR3 expression and prognosis in patients with renal cell carcinomas (RCCs). METHODS: A tissue microarray containing RCC tumor tissue samples and corresponding normal tissue samples was generated. Decoy receptor 3 expression in tumors of 560 patients was examined by immunohistochemistry. The effect of DcR3 expression on disease-specific survival and progression-free survival was assessed using univariate analysis and multivariate Cox regression analysis. Decoy receptor 3 serum levels were determined by ELISA. FINDINGS: High DcR3 expression was associated with high-grade (P = .005) and high-stage (P = .048) RCCs. The incidence of distant metastasis (P = .03) and lymph node metastasis (P = .002) was significantly higher in the group with high DcR3 expression. Decoy receptor 3 expression correlated negatively with disease-specific survival (P < .001) and progression-free survival (P < .001) in univariate analyses. A multivariate Cox regression analysis retained DcR3 expression as an independent prognostic factor that outperformed the Karnofsky performance status. In patients with high-stage RCCs expressing DcR3, the 2-year survival probability was 25%, whereas in patients with DcR3-negative tumors, the survival probability was 65% (P < .001). Moreover, DcR3 serum levels were significantly higher in patients with high-stage localized disease (P = .007) and metastatic disease (P = .001). INTERPRETATION: DcR3 expression is an independent prognostic factor of RCC progression and mortality. Therefore, the assessment of DcR3 expression levels offers valuable prognostic information that could be used to select patients for adjuvant therapy studies