mesothelioma (MPM).We investigated the associations between physical function, symptom burden, and QoL in order to identify key factors that may be targeted for supportive care interventions.Methods: This was a prospective, cohort study of MPM patients enrolled consecutively at a single center.Baseline metrics were obtained at the first visit and repeated every 3 months for up to 5 visits.Symptom and QoL metrics were assessed via the EORTC QLQ-C30 questionnaire.Lower extremity function was measured with the Short Physical Performance Battery (SPPB), which includes balance, gait, and chair stand tests.Spearman rank correlation was used to examine baseline relationships between physical function and symptom and QoL measures.Univariate and multivariate Cox proportional hazards models were used to calculate hazard ratios for all-cause mortality.Results: Seventy-six patients were enrolled between May 2019 and March 2020.Median age was 70 (range 41-89); 70% were male; 92% were White (non-Hispanic); 76.3% had epithelioid histology.The prevalence of baseline symptoms were: fatigue (80.3%), dyspnea (72.4%), insomnia (63.2%), pain (58.7%), constipation (39.5%), appetite loss (36.8%), nausea (23.7%), and diarrhea (17.1%).The mean EORTC global health status/QoL score was 67.2 (SD 19.7), mean EORTC summary score was 78.3 (SD 15.4), and mean SPPB score was 10.7 (SD 1.5).Total SPPB score was significantly correlated with global health status/QoL, summary health, fatigue, dyspnea, and appetite loss (rj.35j, p<.01), but not with pain or insomnia.These correlations were preserved with chair stand performance, but QoL and symptom associations with balance and gait speed were weaker and generally not statistically significant.At time of data analysis, 33 patients (43.4%) were deceased.Decreased survival was associated with baseline severity of fatigue (HR 1.30 per 10-point increase in symptom score, 95% CI 1.12-1.51),dyspnea (HR 1.28, 95% CI 1.11-1.47),and appetite loss (HR 1.27, 95% CI 1.12-1.45).A higher total SPPB score was associated with better survival (HR 0.76, 95% CI 0.61-0.95),while among SPPB components, only chair stand performance was significantly associated with survival (HR 0.67, 95% CI 0.49-0.91).Conclusions: This study confirms the high symptom burden of patients with MPM, with fatigue, dyspnea, insomnia and pain affecting more than half of patients.Our findings additionally show that lower extremity function, specifically measured by ability to perform chair stands, is significantly associated with symptom burden and QoL and may have prognostic significance for MPM patients.Lower extremity function may be a modifiable factor that supportive care interventions can target to improve QoL for patients.

The fibroblast growth factor (FGF) signaling axis is an important driver of malignant pleural mesothelioma (MPM) growth but the specific contribution of most FGFs has not been explored in detail. In the current study, we focus on FGF5 in MPM and demonstrate its link to poor patient survival, increased tumor cell growth and migration, resistance to cisplatin and increased sensitivity to FGFR inhibitors.

Background: ETS factors are an evolutionary conserved family of transcription factors with a wide range of cell molecular functions including embryonic development and immune cell function. YK-4-279 is a novel ETS-factor inhibitor originally designed to inhibit the transcriptional activity of the EWS-Fli1 fusion protein in Ewing sarcoma. Analysis of several other human cancer entities revealed promising activity also in human melanoma cells, often overexpressing ETS factors. Drug resistance development, the main obstacle for sustained clinical response, involves changes both in tumor cells as well as the tumor environment including tumor-associated macrophages.

Solitary fibrous tumors of the pleura (SFTP) are rare benign or malignant cancers of mesenchymal origin. SFTPs are clinically classified by the expression of immunohistochemical markers CD34, CD99 and Bcl-2. Furthermore, NAB2-STAT6 gene fusion is a distinct hallmark associated with this very rare disease. The clinical course for SFTP patients still lacks consistent treatment guidelines and follow-up strategies. SFTP therapy mostly comprises of total resection only. Hence, the role of chemo-, radio-, and targeted therapy remains elusive. Thus, in-depth molecular analyses are inevitably needed.

TERT promoter mutations in meningiomas were recently found to be strongly prognostic and associated with malignant progression and risk of recurrence. However, the cellular factors underlying meningioma aggressiveness are comparably unexplored. Due to the limited meningioma cell lines available worldwide, it was aim of our study to analyse an extended cohort of WHO grade I-III meningiomas regarding the impact of TERT promoter mutations, telomerase activity and TERT mRNA expression on cell propagation in vitro. Tumor tissue from 128 meningiomas was analysed for TERT promoter mutations by direct sequencing. Telomerase activity (TA), TERT mRNA expression and telomere lengths were investigated by TRAP assay, RT-PCR, and qPCR, respectively. Correspondingly, tumor-derived primo-cell cultures (121/128) were established. The impact of the ETS-transcription factor inhibitor YK-4–279 on cell viability was analysed in selected primo-cell cultures by MTT assays. To elucidate whether ETS- transcription factor inhibition affects TERT promoter activity, luciferase-based reporter assays were performed. TERT promoter mutations were found in 6 % of all samples analysed and were associated with a significantly upregulated telomerase activity and TERT mRNA expression (p<0.0001 both). Meningioma derived primo-cell cultures were able to be propagated with comparable success rates from TA positive and- negative (55% versus 53%) tumors. In contrast, within the meningioma cohort harbouring TERT promoter mutations, 75% (6/8) viable primo-cell cultures could be established. Out of these, 2 primo-cell cultures (2/6; 33%) developed into stable immortalized cell lines. This strongly indicates that TERT promoter mutations but not telomerase activity or TERT mRNA expression were predicting indefinite cell growth in vitro. TERT promoter mutated meningioma cells were hypersensitive against the ETS transcription factor inhibitor YK-4–279 inducing a significant downregulation of TERT mRNA expression. Accordingly, as shown by the luciferase-reporter assay, the enhanced activity of the mutated TERT promoter was completely blocked by presence of YK-4–279. Patients with TERT promoter mutated meningiomas exhibited a significantly shorter overall survival (p=0.0006; 53.8 vs 115.6 months). Aggressiveness of meningiomas reflected by cell immortalization in vitro is dependent on TERT promoter mutations. This fits well with the data of worse prognosis for meningioma patients harbouring a TERT promoter-mutated tumor. Additionally, TERT promoter mutations might open a promising novel therapeutic opportunity in progressive meningiomas.

Malignant pleural mesothelioma (MPM) is characterized by aggressive growth, limited therapeutic options and rapid recurrence following treatment. A better understanding of biological factors underlying MPM aggressiveness offers the chance to improve therapeutic strategies. Growth factors of the TGF-beta superfamily including TGF-beta itself, activins and BMPs have been repeatedly linked to MPM growth. In the current study, we focus on the role of Smad7, a key intracellular antagonist of TGF-beta and activin signaling, in MPM.

Background: Colorectal carcinoma (CRC) is the third most common cancer worldwide. Platinum-based anticancer compounds still constitute one mainstay of systemic CRC treatment despite limitations due to adverse effects and resistance development. Trabectedin has shown promising antitumor effects in CRC, however, again resistance development may occur. In this study, we aimed to develop strategies to circumvent or even exploit acquired trabectedin resistance in novel CRC treatment regimens.Methods: Human HCT116 CRC cells were selected for acquired trabectedin resistance in vitro and characterised by cell biological as well as bioinformatic approaches. In vivo xenograft experiments were conducted.Results: Selection of HCT116 cells for trabectedin resistance resulted in p53-independent hypersensitivity of the selected subline against cisplatin. Bioinformatic analyses of mRNA microarray data suggested deregulation of nucleotide excision repair and particularly loss of the ubiquitin ligase CUL4A in trabectedin-selected cells. Indeed, transient knockdown of CUL4A sensitised parental HCT116 cells towards cisplatin. Trabectedin selected but not parental HCT116 xenografts were significantly responsive towards cisplatin treatment.Conclusions: Trabectedin selection-mediated CUL4A loss generates an Achilles heel in CRC cancer cells enabling effective cisplatin treatment. Hence, inclusion of trabectedin in cisplatin-containing cancer treatment regimens might cause profound synergism based on reciprocal resistance prevention.

In primary melanoma, the amount of vascular endothelial growth factor C (VEGF-C) expression and lymphangiogenesis predicts the probability of metastasis to sentinel nodes, but conditions boosting VEGF-C expression in melanoma are poorly characterized. By comparative mRNA expression analysis of a set of 22 human melanoma cell lines, we found a striking negative correlation between VEGF-C and microphthalmia-associated transcription factor (MITF) expression, which was confirmed by data mining in GEO databases of human melanoma Affymetrix arrays. Moreover, in human patients, high VEGF-C and low MITF levels in primary melanoma significantly correlated with the chance of metastasis. Pathway analysis disclosed the respective c-Jun N-terminal kinase and p38/mitogen-activated protein kinase activities as being responsible for the inverse regulation of VEGF-C and MITF. Predominant c-Jun N-terminal kinase signaling results in a VEGF-C-low/MITFhigh phenotype; these melanoma cells are highly proliferative, show low mobility, and are poorly lymphangiogenic. Predominant p38 signaling results in a VEGF-C-high/MITFlow phenotype, corresponding to a slowly cycling, highly mobile, lymphangiogenic, and metastatic melanoma. In conclusion, the relative c-Jun N-terminal kinase and p38 activities determine the biological behavior of melanoma. VEGF-C and MITF levels serve as surrogate markers for the respective c-Jun N-terminal kinase and p38 activities and may be used to predict the risk of metastasis in primary melanoma.

Glioblastoma multiforme (GBM) is a heterogeneous, highly aggressive primary brain tumor with strongly variable patient survival. Because reliable prognostic biomarkers are lacking, we investigated the relation between telomerase-associated parameters and the disease course.Telomerase-associated parameters were determined in 100 GBM tissues and associated with clinical characteristics and overall survival. Expressions of telomere length, telomerase activity (TA), and human telomerase reverse transcriptase (hTERT) were analyzed by quantitative PCR, telomeric repeat amplification protocol assay, and reverse transcriptasePCR, respectively. Mutation status of isocitrate dehydrogenase (IDH)1 was determined by direct sequencing, and O-6-methylguanine DNA methyltransferase (MGMT) promoter methylation by methylation-specific PCR.Of 100 GBM tissues, 61 were positive for both hTERT mRNA and TA, with a highly significant correlation between both parameters (linear regression, P .0001). Telomere length determination revealed a significant difference between the hTERT/TA-positive and -negative subgroups, with markedly longer telomeres in the hTERT/TA-negative cohort (unpaired Students t-test, P .0001). Accordingly, significantly shorter telomeres were detected in GBM tissues derived from older patients (60 y at diagnosis, P .0001). While no association of telomere parameters with MGMT promoter status was found, all tumors with IDH1 mutation (6/100) were negative for both hTERT expression and TA and harbored significantly longer telomeres. Patients with tumors lacking hTERT expression/TA showed a significant survival benefit (KaplanMeier test, both P .01), which, however, was based exclusively on the younger patient subgroup (60 y, both P .005; 60 y, both ns).Telomerase activation is not an independent prognostic parameter in GBM but predicts aggressive tumor behavior solely in a younger patient cohort.

Material and Methods: Based on literature data and miRanda database we have focused on analysis of 16 miRNAs (miR-21, miR-126, miR-205, miR-23a, miR-23b, miR-335*, miR-369-3p, miR-3163, miR-491-3p, miR-548x, miR-548p, miR-576-5p, miR-590-5p, miR-655, miR-656, miR-944) that target genes involved in cell proliferation, differentiation, apoptosis and resistance.Our patient cohort consisted of 52 formalin-fixed paraffin-embedded (FFPE) samples from operable non-small cell lung cancer (NSCLC) patients treated in the University Hospital in Olomouc in years 1996-2000.Total RNA was isolated and after reverse transcription with miRNA-specific looped primers and preamplification PCR, real-time PCR was used for miRNAs quantification.Statistical methods were used to correlate the expression levels of miRNAs and different variables.For survival analysis the Kaplan-Meier method was used. Results:The results of our pilot study suggested that expression of miR-205 positively correlated with overall survival of NSCLC patients (p < 0.095) in stage II patients and higher level of miR-944 expression (p Conclusion: The quantification of miR-205, miR-590-5p and miR-944 levels seems to be the useful diagnostic tool for better stratification of NSCLC patients and they might be used as diagnostic and prognostic markers.

Background: Vaults are highly conserved ribonucleoprotein particles ubiquitously expressed in eukaryotic organisms.They predominantly consist of the 110 kDa major vault protein (MVP) and have been implicated in the regulation of multiple cellular processes including transport mechanisms, chemoresistance, and intracellular signalling pathways.While in normal brain the expression is low, MVP levels are consistently upregulated in glioblastoma multiforme (GBM).Aim of this study was to investigate whether MVP/vaults have an impact on GBM cell growth and survival, including chemotherapy responsiveness, and to clarify underlying molecular mechanisms.Material and Methods: The MVP protein was stably overexpressed in MVP-low H7 glioma cells.Ectopic and endogenous MVP expression was repressed by MVP mRNA-specific shRNA.Protein expressions were detected by immunofluorescence and Western blot.Consequences of MVP modulation on cell proliferation, survival, chemotherapy response and serum starvation with or without growth factor stimulation were analysed.Additionally, impact of MVP on subcutaneous and orthotopic tumour formation in SCID mice was tested.Results and Discussion: Ectopic MVP expression in H7 glioma cells did not substantially alter sensitivity against diverse chemotherapeutic drugs.However, responsiveness to growth factor stimulation (EGF, serum) was increased paralleled by a significant upregulation of MAPK-and PI3Kpathway indicated by phosphorylation of ERK, AKT and S6.Moreover, MVPtransgenic cells were impressively resistant to apoptotic cell death induced by serum-starvation, an effect reversible by shRNA-mediated MVP-repression.PI3K downstream signalling, namely AKT and S6 phosphorylation, was hyperactivated in MVP-positive as compared to control transfected cells.Accordingly, inhibition of mTOR via temsirolimus or PI3K via LY-294002 induced both complete blockade of S6 and 4EBP phosphorylation and restoration of apoptosis induction by serum starvation of MVP-positive cells.Subcutaneous tumor growth in SCID mice was significantly enhanced, PI3K signalling distinctly increased and the apoptotic cell fraction reduced in orthotopic xenografts from MVP-overexpressing subclones as compared to vector controls.Conclusion: Our data proof a significant contribution of vaults/MVP to the malignant phenotype of human GBM cells by supporting enhanced survival under nutrient starvation based on the deregulation of oncogenic signalling pathways.

resistant to 0.5 mM 17-AAG in cell proliferation assays, whereas IMIM-PC-2 is the only cell line resistant to 0.5 mM radicicol.In general, these inhibitors induce a G 2 /M arrest in sensitive cell lines, except in IMIM-PC-2 cells that are arrested in the G 1 phase of the cell cycle.Especially, HT-29, SW-620 and IMIM-PC-1 cells accumulate in G 2 /M, followed by an increase in subG 1 .Additionally, we have studied the relationship between the presence of the enzyme NAD(P)H: quinone oxidoreductase 1 (NQO1) and 17-AAG activity, because previous studies have shown that this enzyme metabolizes the quinone 17-AAG to hydroquinone (17-AAGH 2 ), which has more affinity for HSP90.However, the levels of NQO1 as a determinant of sensitivity to 17-AAG is still controversial.Conclusions: There are three cell lines especially sensitive to HSP90 inhibitors: HT-29, SW-620 and IMIM-PC-1.Furthermore, our study shows that NVP-AUY922 treatment is the most effective HSP90 inhibitor in pancreatic and colorectal carcinoma cell lines.Therefore, NVP-AUY922 is the best candidate to be used in a clinical setting.