Suchergebnisse

Background: Histone deacetylase inhibitors (HDACis) like vorinostat are promising radiosensitisers in prostate cancer, but their effect under hypoxia is not known. We investigated gene expression associated with radiosensitisation of normoxic and hypoxic prostate cancer cells by vorinostat. Methods: Cells were exposed to vorinostat under normoxia or hypoxia and subjected to gene expression profiling before irradiation and clonogenic survival analysis. Results: Pretreatment with vorinostat led to radiosensitisation of the intrinsically radioresistant DU 145 cells, but not the radiosensitive PC-3 and 22Rv1 cells, and was independent of hypoxia status. Knockdown experiments showed that the sensitisation was not caused by repression of hypoxia-inducible factor HIF1 or tumour protein TP53. Global deregulation of DNA repair and chromatin organisation genes was associated with radiosensitisation under both normoxia and hypoxia. A radiosensitisation signature with expression changes of 56 genes was generated and valid for both conditions. For eight signature genes, baseline expression also correlated with sensitisation, showing potential as pretreatment biomarker. The hypoxia independence of the signature was confirmed in a clinical data set. Conclusions: Pretreatment with HDACi may overcome radioresistance of hypoxic prostate tumours by similar mechanisms as under normoxia. We propose a gene signature to predict radiosensitising effects independent of hypoxia status. ; Financial support was provided by the European Union 7th Framework Program (Grant No. 222741-METOXIA), the Norwegian Cancer Society (Grant No. 107438 - PR-2007-0179) and The South-Eastern Norway Regional Health Authority (Grant No. 2015020).

The tumor-suppressor protein p16 is paradoxically overexpressed in cervical cancer (CC). Despite its potential as a biomarker, its clinical value and the reasons for its failure in tumor suppression remain unclear. Our purpose was to determine p16 clinical and biological significance in CC. p16 expression pattern was examined by immunohistochemistry in 78 CC cases (high-grade squamous intraepithelial lesions (HSILs) and squamous cell carcinomas of the cervix –SCCCs). CC cell proliferation and invasion were monitored by real-time cell analysis and Transwell® invasion assay, respectively. Cytoplasmic p16 interactors were identified from immunoprecipitated extracts by liquid chromatography-tandem mass spectrometry, and colocalization was confirmed by double-immunofluorescence. We observed that SCCCs showed significantly more cytoplasmic than nuclear p16 expression than HSILs. Importantly, nuclear p16 absence significantly predicted poor outcome in SCCC patients irrespective of other clinical parameters. Moreover, we demonstrated that cytoplasmic p16 interacted with CDK4 and other unreported proteins, such as BANF1, AKAP8 and AGTRAP, which could sequester p16 to avoid nuclear translocation, and then, impair its anti-tumor function. Our results suggest that the absence of nuclear p16 could be a diagnostic biomarker between HSIL and SCCC, and an independent prognostic biomarker in SCCC; and explain why p16 overexpression fails to stop CC growth. ; This research was funded by the Health Department of the Navarre Government (Grant number 21/11), the Navarre Breast Cancer Patients' Association (SARAY) and the "Proof of concept project on proteomic research" from Proteored (Grant number ProteoRed-0000029). The Proteomics Unit of Navarrabiomed is a member of Proteored, PRB3-ISCIII, and is supported by Grant PT17/0019/009, of the PE I + D + I 2013–2016 funded by ISCIII and ERDF. S.M. was a recipient of a predoctoral grant from the Public University of Navarre. E.M.-S. was a recipient of a fellowship from the Spanish Ministry of Science, Innovation and Universities (PTA2015-11895-I).

No therapeutic targets and molecular biomarkers are available in cervical cancer (CC) management. In other cancer types, micro-RNA-877-3p (miR-877-3p) has been associated with events relevant for CC development. Thus, we aimed to determine miR-877-3p role in CC. miR-877-3p levels were examined by quantitative-PCR in 117 cervical lesions and tumors. Effects on CC cell proliferation, migration, and invasion were evaluated upon anti-miR-877-3p transfection. miR-877-3p dependent molecular mechanism was comprehensively explored by proteomics, dual-luciferase reporter assay, western blot, and immunohistochemistry. Cervical tumors expressed higher miR-877-3p levels than benign lesions. miR-877-3p promoted CC cell migration and invasion, at least partly by modulating cytoskeletal protein folding through the chaperonin-containing T-complex protein 1 complex. Notably, miR-877-3p silencing synergized with paclitaxel. Interestingly, miR-877-3p downregulated the levels of an in silico-predicted target, ZNF177, whose expression and subcellular location significantly distinguished high-grade squamous intraepithelial lesions (HSILs) and squamous cell carcinomas of the cervix (SCCCs). Cytoplasmic ZNF177 was significantly associated with worse progression-free survival in SCCC. Our results suggest that: (i) miR-877-3p is a potential therapeutic target whose inhibition improves paclitaxel effects; (ii) the expression and location of its target ZNF177 could be diagnostic biomarkers between HSIL and SCCC; and (iii) cytoplasmic ZNF177 is a poor-prognosis biomarker in SCCC. ; This research was funded by the Health Department of the Navarre Government (Grant number 21/11), the Navarre Breast Cancer Patients' Association (SARAY), and the "Proof of concept project on proteomic research" from Proteored (Grant number ProteoRed-0000029). The Proteomics Unit of Navarrabiomed is a member of Proteored, PRB3-ISCIII, and is supported by grant PT17/0019/009, of the PE I + D + I 2013-2016 funded by ISCIII and ERDF. S.M. was a recipient of a predoctoral grant from the Public University of Navarre. I.A. was supported by Miguel Servet contract CP15/00176 from the Instituto de Salud Carlos III-FEDER. E.M.-S. was a recipient of a fellowship from the Spanish Ministry of Science, Innovation, and Universities (PTA2015-11895-I)

Cyclin D1 (Ccnd1) is a proto-oncogen amplified in many different cancers and nuclear accumulation of Ccnd1 is a characteristic of tumor cells. Ccnd1 activates the transcription of a large set of genes involved in cell cycle progress and proliferation. However, Ccnd1 also targets cytoplasmic proteins involved in the regulation of cell migration and invasion. In this work, we have analyzed by immunohistochemistry the localization of Ccnd1 in endometrial, breast, prostate and colon carcinomas with different types of invasion. The number of cells displaying membranous or cytoplasmic Ccnd1 was significantly higher in peripheral cells than in inner cells in both collective and pushing invasion patterns of endometrial carcinoma, and in collective invasion pattern of colon carcinoma. Also, the cytoplasmic localization of Ccnd1 was higher when tumors infiltrated as single cells, budding or small clusters of cells. To evaluate cytoplasmic function of cyclin D1, we have built a variant (Ccnd1-CAAX) that remains attached to the cell membrane therefore sequestering this cyclin in the cytoplasm. Tumor cells harboring Ccnd1-CAAX showed high levels of invasiveness and metastatic potential compared to those containing the wild type allele of Ccnd1. However, Ccnd1- CAAX expression did not alter proliferative rates of tumor cells. We hypothesize that the role of Ccnd1 in the cytoplasm is mainly associated with the invasive capability of tumor cells. Moreover, we propose that subcellular localization of Ccnd1 is an interesting guideline to measure cancer outcome. ; This work was funded by Spanish Ministry of Education and Science (BFU2010-20293/ BMC and BFU2013-42895-P), Catalan Government (SGR-559), and Fondo de Investigaciones Sanitarias (PI10/00604 and PI13/00263). X.M-G was supported by grants 2014SGR138, RD12/0036/0013, and Fundación Asociación Española contra el Cancer. Tumour samples were obtained with the support of Xarxa Catalana de Bancs de Tumors, and Plataforma de Biobancos ISCIII (PT13/0010/0014). NP. Fusté was ...

Background: Colorectal (CRC) and endometrial cancer (EC) are the most common types of cancer in Lynch syndrome (LS). Risk reducing surgeries (RRS) might impact cancer incidence and mortality. Our objectives were to evaluate cumulative incidences of CRC, gynecological cancer and all-cause mortality after RRS in LS individuals. Methods: Retrospective analysis of 976 LS carriers from a single-institution registry. Primary endpoints were cumulative incidence at 75 years of cancer (metachronous CRC in 425 individuals; EC and ovarian cancer (OC) in 531 individuals) and all-cause mortality cumulative incidence, comparing extended (ES) vs. segmental surgery (SS) in the CRC cohort and risk reducing gynecological surgery (RRGS) vs. surveillance in the gynecological cohort. Results: Cumulative incidence at 75 years of metachronous CRC was 12.5% vs. 44.7% (p = 0.04) and all-cause mortality cumulative incidence was 38.6% vs. 55.3% (p = 0.31), for ES and SS, respectively. Cumulative, incidence at 75 years was 11.2% vs. 46.3% for EC (p = 0.001) and 0% vs. 12.7% for OC (p N/A) and all-cause mortality cumulative incidence was 0% vs. 52.7% (p N/A), for RRGS vs. surveillance, respectively. Conclusions: RRS in LS reduces the incidence of metachronous CRC and gynecological neoplasms, also indicating a reduction in all-cause mortality cumulative incidence in females undergoing RRGS. ; This research has been funded by the Instituto de Salud Carlos III and co-funded by European Social Fund—ESF investing in your future—(grant CM19/00099), the Catalan-Balearic Society of Oncology (2018 grant of the Catalan-Balearic Society of Oncology), the Spanish Ministry of Economy and Competitiveness and co-funded by FEDER funds—A way to build Europe—(grant SAF2015-68016-R), CIBERONC and the Government of Catalonia (grants 2017SGR1282 and PERIS SLT002/16/0037). We thank the CERCA Program/Generalitat de Catalunya for institutional support.

Sporadic synchronous endometrial (ECs) and ovarian cancers (OCs), although clinically considered to be independent primaries, have been shown to be clonally related and likely constitute metastases from each other. We sought to define whether synchronous ECs/OCs in patients with DNA mismatch repair (MMR)-deficiency syndromes would be clonally-related. We subjected synchronous ECs/OCs from four patients (LS3-LS6) with clinically confirmed Lynch syndrome (LS) and one patient with constitutional mismatch repair deficiency syndrome (CMMRD) to massively parallel sequencing targeting 468 cancer-related genes. Somatic mutations, copy number alterations (CNAs), clonal relatedness and clonal decomposition analyses were performed using previously described bioinformatics methods. All synchronous ECs/OCs analyzed were considered independent primaries based on clinicopathologic criteria. Sequencing analysis revealed that the ECs/OCs of three cases (LS2-CMMRD, L3, L4) harbored similar repertoires of somatic mutations and CNAs and were clonally-related. In these three cases, a subset of subclonal mutations in the EC became clonal in the OC, suggesting that the EC was likely the substrate from which the OC developed. LS5's EC/OC had distinct mutational profiles but shared specific CNAs. In contrast, LS6's EC/OC harbored distinct somatic mutation and lacked CNAs, consistent with each tumor constituting an independent primary lesion. In LS5 and LS6, PTEN mutations and PTEN loss of protein expression were found to be restricted to the EC. Finally, re-analysis of sequencing data of sporadic synchronous ECs/OCs supported the observations made in the current study that the directionality of progression is likely from the endometrium to the ovary. In conclusion, contrary to sporadic synchronous ECs/OCs, which are almost invariably clonally-related, ECs/OCs simultaneously involving the uterus and ovary in LS patients may represent distinct primary tumors. A subset of MMR-deficiency syndrome-related synchronous ECs/OCs, however, may originate from a single primary tumor at variance with their clinical diagnosis, with the endometrium being the likeliest site of origin. ; JSR-F was funded in part by a Breast Cancer Research Foundation grant, BW in part by Breast Cancer Research Foundation, Cycle for Survival and Stand Up To Cancer grants, F.P. partially by a National Institutes of Health/ National Cancer Institute K12 CA184746 grant, XM-G by AECC (grupos estables) and GC and MP by the Spanish Ministry of Economy and Competitiveness and co-funded by FEDER funds -a way to build Europe- (grant SAF2015-68016-R), CIBERONC and the Government of Catalonia (grant 2017SGR1282). XM-G, GC and MP thank CERCA Programme for institutional support. Research reported in this publication was supported in part by a Cancer Center Support Grant of the NIH/NCI (Grant No. P30CA008748).

The causal mechanism for cancer predisposition in Lynch-like syndrome (LLS) remains unknown. Our aim was to elucidate the constitutional basis of mismatch repair (MMR) deficiency in LLS patients throughout a comprehensive (epi)genetic analysis. One hundred and fifteen LLS patients harboring MMR-deficient tumors and no germline MMR mutations were included. Mutational analysis of 26 colorectal cancer (CRC)-associated genes was performed. Pathogenicity of MMR variants was assessed by splicing and multifactorial likelihood analyses. Genome-wide methylome analysis was performed by the Infinium Human Methylation 450K Bead Chip. The multigene panel analysis revealed the presence of two MMR gene truncating mutations not previously found. Of a total of 15 additional MMR variants identified, five -present in 6 unrelated individuals- were reclassified as pathogenic. In addition, 13 predicted deleterious variants in other CRC-predisposing genes were found in 12 probands. Methylome analysis detected one constitutionalMLH1epimutation, but no additional differentially methylated regions were identified in LLS compared to LS patients or cancer-free individuals. In conclusion, the use of an ad-hoc designed gene panel combined with pathogenicity assessment of variants allowed the identification of deleterious MMR mutations as well as new LLS candidate causal genes. Constitutional epimutations in non-LS-associated genes are not responsible for LLS. ; This work was funded by the Spanish Ministry of Economy and Competitiveness and cofunded by FEDER funds -a way to build Europe-(grants SAF2012-33636, SAF2015-68016-R and SAF2016-80888-R), CIBERONC, RTICC Network (RD12/0036/0031 and RD12/0036/0008), the Spanish Association Against Cancer (AECC) (080253), the Government of Catalonia (grant 2014SGR338, 2017SGR1282 and PERIS SLT002/16/0037), Fundacion Mutua Madrilena (grant AP114252013). We thank CERCA Programme for institutional support. ED was supported by a grant from the Spanish Ministry of Economy and Competitiveness. The AECC fellowship to MG-A. AF was supported by a grant from the Catalonian Health Department (SLT002/16/00409). FM was supported by CIBERONC. The Mexican National Council for Science and Technology (CONACyT) fellowship to GV.

Mitochondrial metabolism and the generation of reactive oxygen species (ROS) contribute to the acquisition of DNA mutations and genomic instability in cancer. How genomic instability influences the metabolic capacity of cancer cells is nevertheless poorly understood. Here, we show that homologous recombination‐defective (HRD) cancers rely on oxidative metabolism to supply NAD+ and ATP for poly(ADP‐ribose) polymerase (PARP)‐dependent DNA repair mechanisms. Studies in breast and ovarian cancer HRD models depict a metabolic shift that includes enhanced expression of the oxidative phosphorylation (OXPHOS) pathway and its key components and a decline in the glycolytic Warburg phenotype. Hence, HRD cells are more sensitive to metformin and NAD+ concentration changes. On the other hand, shifting from an OXPHOS to a highly glycolytic metabolism interferes with the sensitivity to PARP inhibitors (PARPi) in these HRD cells. This feature is associated with a weak response to PARP inhibition in patient‐derived xenografts, emerging as a new mechanism to determine PARPi sensitivity. This study shows a mechanistic link between two major cancer hallmarks, which in turn suggests novel possibilities for specifically treating HRD cancers with OXPHOS inhibitors ; This study has been funded by the Ministerio de Ciencia, Innovación y Universidades, which is part of the Agencia Estatal de Investigación (AEI), through the project SAF2017-85869-R (cofunded by the European Regional Development Fund (ERDF), a way to build Europe) to FV and BFU2015-66030-R to JCP; by the FIS PI15/00854 and FIS PI16/01898 (Instituto Carlos III, cofunded by FEDER funds/European Regional Development Fund (ERDF), a way to build Europe) to MAP and AVillanueva and with the support of the Secretariat for Universities and Research of the Department of Business and Knowledge of the Government of Catalonia (2017SGR449) to FV. We thank the CERCA Program/Generalitat de Catalunya for their institutional support. We particularly wish to acknowledge the collaboration of ...

Background: About 20% of patients diagnosed with endometrial cancer (EC) are considered high-risk with unfavorable prognosis. In the framework of the European Network for Individualized Treatment in EC (ENITEC), we investigated the presence and phenotypic features of Circulating Tumor Cells (CTC) in high-risk EC patients. Methods: CTC isolation was carried out in peripheral blood samples from 34 patients, ranging from Grade 3 Stage IB to Stage IV carcinomas and recurrences, and 27 healthy controls using two methodologies. Samples were subjected to EpCAM-based immunoisolation using the CELLection™ Epithelial Enrich kit (Invitrogen, Dynal) followed by RTqPCR analysis. The phenotypic determinants of endometrial CTC in terms of pathogenesis, hormone receptor pathways, stem cell markers and epithelial to mesenchymal transition (EMT) drivers were asked. Kruskal-Wallis analysis followed by Dunn's post-test was used for comparisons between groups. Statistical significance was set at p < 0.05. Results: EpCAM-based immunoisolation positively detected CTC in high-risk endometrial cancer patients. CTC characterization indicated a remarkable plasticity phenotype defined by the expression of the EMT markers ETV5, NOTCH1, SNAI1, TGFB1, ZEB1 and ZEB2. In addition, the expression of ALDH and CD44 pointed to an association with stemness, while the expression of CTNNB1, STS, GDF15, RELA, RUNX1, BRAF and PIK3CA suggested potential therapeutic targets. We further recapitulated the EMT phenotype found in endometrial CTC through the up-regulation of ETV5 in an EC cell line, and validated in an animal model of systemic dissemination the propensity of these CTC in the accomplishment of metastasis. Conclusions: Our results associate the presence of CTC with high-risk EC. Gene-expression profiling characterized a CTC-plasticity phenotype with stemness and EMT features. We finally recapitulated this CTC-phenotype by over-expressing ETV5 in the EC cell line Hec1A and demonstrated an advantage in the promotion of metastasis in an in vivo mouse model of CTC dissemination and homing. ; ISCIII PI11/00873; Fundación Asociación Española Contra el Cancer (AECC), Grupos Estables 2011; InveNNta (Innovation in Nanomedicine), co-financed by the European Union (EU) through the Operational Programme for Cross-border Cooperation, Spain-Portugal (POCTEP 2007-2013), European Regional Development Fund (ERDF); Helse Vest, Research Council of Norway, Norwegian Cancer Society and Harald Andersens legat (H.B.S.); L. Alonso-Alconada is recipient of fellowship from the Basque Government (Spain).