An open debate in antiangiogenic therapies is about their consequence on tumor invasiveness and metastasis, which is undoubtedly relevant for patients currently treated with antiangiogenics, such as renal cell carcinoma patients. To address, this we developed an extensive series of 27 patient biopsy-derived orthotopic xenograft models (Ren-PDOX) that represent inter-patient heterogeneity. In specific tumors, antiangiogenics produced increased invasiveness and metastatic dissemination, while in others aggressiveness remained unchanged. Mechanistically, species-discriminative RNA sequencing identified a tumor cell-specific differential expression profile associated with tumor progression and aggressivity in TCGA RCC patients. Gene filtering using an invasion-annotated patient series pinpointed two candidate genes, of which ALDH1A3 differentiated the pro-invasive subtype of Ren-PDOXs. Validation in an independent series of 15 antiangiogenic-treated patients confirmed that pre-treatment ALDH1A3 can significantly discriminate patients with pro-aggressive response upon treatment. Overall, results confirm that effects of antiangiogenic drugs on tumor invasion and metastasis are heterogeneous and may profoundly affect the natural progression of tumors and promote malignancy. Furthermore, we identify a specific molecular biomarker that could be used to select patients that better benefit from treatment.

Micro-RNAs (miRNAs) play a crucial role in controlling intestinal epithelial barrier function partly by modulating the expression of tight junction (TJ) proteins. We have previously shown differential messenger RNA (mRNA) expression correlated with ultrastructural abnormalities of the epithelial barrier in patients with diarrhoea-predominant IBS (IBS-D). However, the participation of miRNAs in these differential mRNA-associated findings remains to be established. Our aims were (1) to identify miRNAs differentially expressed in the small bowel mucosa of patients with IBS-D and (2) to explore putative target genes specifically involved in epithelial barrier function that are controlled by specific dysregulated IBS-D miRNAs.

PTOV1 is a transcription and translation regulator and a promoter of cancer progression. Its overexpression in prostate cancer induces transcription of drug resistance and self-renewal genes, and docetaxel resistance. Here we studied PTOV1 ability to directly activate the transcription of ALDH1A1 and CCNG2 by binding to specific promoter sequences. Chromatin immunoprecipitation and electrophoretic mobility shift assays identified a DNA-binding motif inside the PTOV-A domain with similarities to known AT-hooks that specifically interacts with ALDH1A1 and CCNG2 promoters. Mutation of this AT-hook-like sequence significantly decreased the expression of ALDH1A1 and CCNG2 promoted by PTOV1. Immunohistochemistry revealed the association of PTOV1 with mitotic chromosomes in high grade prostate, colon, bladder, and breast carcinomas. Overexpression of PTOV1, ALDH1A1, and CCNG2 significantly correlated with poor prognosis in prostate carcinomas and with shorter relapse-free survival in colon carcinoma. The previously described interaction with translation complexes and its direct binding to ALDH1A1 and CCNG2 promoters found here reveal the PTOV1 capacity to modulate the expression of critical genes at multiple levels in aggressive cancers. Remarkably, the AT-hook motifs in PTOV1 open possibilities for selective targeting its nuclear and/or cytoplasmic activities.

PTOV1 is an adaptor protein with functions in diverse processes, including gene transcription and protein translation, whose overexpression is associated with a higher proliferation index and tumor grade in prostate cancer (PC) and other neoplasms. Here we report its interaction with the Notch pathway and its involvement in PC progression.

Corticotropin-releasing factor (CRF) has been identified in intestinal mucosal eosinophils and associated with psychological stress and gut dysfunction. Irritable bowel syndrome (IBS) is commonly characterized by altered intestinal motility, immune activation, and increased gut barrier permeability along with heightened susceptibility to psychosocial stress. Despite intensive research, the role of mucosal eosinophils in stress-associated gut dysfunction remains uncertain. In this study, we evaluated eosinophil activation profile and CRF content in the jejunal mucosa of diarrhea-predominant IBS (IBS-D) and healthy controls (HC) by gene/protein expression and transmission electron microscopy. We also explored the association between intestinal eosinophil CRF and chronic stress, and the potential mechanisms underlying the stress response by assessing eosinophil response to neuropeptides. We found that mucosal eosinophils displayed higher degranulation profile in IBS-D as compared to HC, with increased content of CRF in the cytoplasmic granules, which significantly correlated with IBS clinical severity, life stress background and depression. Eosinophils responded to substance P and carbachol by increasing secretory activity and CRF synthesis and release, without promoting pro-inflammatory activity, a profile similar to that found in mucosal eosinophils from IBS-D. Collectively, our results suggest that intestinal mucosal eosinophils are potential contributors to stress-mediated gut dysfunction through CRF production and release.

Rapid and reliable diagnosis of prostate cancer (PCa) is highly desirable as current used methods lack specificity. In addition, identification of PCa biomarkers that can classify patients into high- and low-risk groups for disease progression at early stage will improve treatment decision-making. Here, we describe a set of protein-combination panels in urinary extracellular vesicles (EVs), defined by targeted proteomics and immunoblotting techniques that improve early non-invasive detection and stratification of PCa patients.We report a two-protein combination in urinary EVs that classifies benign and PCa patients (ADSV-TGM4), and a combination of five proteins able to significantly distinguish between high- and low-grade PCa patients (CD63-GLPK5-SPHM-PSA-PAPP). Proteins composing the panels were validated by immunohistochemistry assays in tissue microarrays (TMAs) confirming a strong link between the urinary EVs proteome and alterations in PCa tissues. Moreover, ADSV and TGM4 abundance yielded a high diagnostic potential in tissue and promising TGM4 prognostic power. These results suggest that the proteins identified in urinary EVs distinguishing high- and low grade PCa are a reflection of histological changes that may be a consequence of their functional involvement in PCa development. In conclusion, our study resulted in the identification of protein-combination panels present in urinary EVs that exhibit high sensitivity and specificity for PCa detection and patient stratification. Moreover, our study highlights the potential of targeted proteomic approaches-such as selected reaction monitoring (SRM)-as diagnostic assay for liquid biopsies via urinary EVs to improve diagnosis and prognosis of suspected PCa patients.

The human sex hormone-binding globulin (SHBG) gene comprises at least 6 different transcription units (TU-1, -1A, -1B, -1C, -1D and -1E), and is regulated by no less than 6 different promoters. The best characterized are TU-1 and TU-1A: TU-1 is responsible for producing plasma SHBG, while TU-1A is transcribed and translated in the testis. Transcription of the recently described TU-1B, -1C, and -1D has been demonstrated in human prostate tissue and prostate cancer cell lines, as well as in other human cell lines such as HeLa, HepG2, HeK 293, CW 9019 and imr 32. However, there are no reported data demonstrating their translation. In the present study, we aimed to determine whether TU-1A and TU-1B are indeed translated in the human prostate and whether 5' UTR exons 1A and 1B differently regulate SHBG translation.

The human Sex Hormone Binding Globulin (SHBG) gene, located at 17p13.1, comprises, at least, two different transcription units regulated by two different promoters. The first transcription unit begins with the exon 1 sequence and is responsible for the production of plasma SHBG by the hepatocytes, while the second begins with an alternative exon 1 sequence, which replaces the exon 1 present in liver transcripts. Alternative exon 1 transcription and translation has only been demonstrated in the testis of transgenic mice containing an 11-kb human SHBG transgene and in the human testis. Our goal has been to further characterize the 5' end of the SHBG gene and analyze the presence of the SHBG alternative transcripts in human prostate tissue and derived cell lines.

Clear cell renal cell carcinoma (ccRCC) is the most common and aggressive subtype of renal cancer. STAT3 pathway is altered in these tumors and p-STAT3 Ser727 is an independent prognostic factor for ccRCC. Protein kinase CK2 is altered in different types of tumors and overexpression of CK2α is considered predictive of bad prognosis and metastatic risk. CK2 subunits analyses in ccRCC samples showed increased CK2α/α' nuclear content in all cases, but decreased cytosolic CK2β (CK2βcyt) levels in the more advanced tumors. Stable downregulation of CK2β in renal proximal tubular (HK-2) and clear cell adenocarcinoma (786-O) cells triggered changes in E-cadherin, vimentin and Snail1 protein levels indicative of epithelial-to-mesenchymal transition (EMT), and increased HIF-α. Moreover, CK2β was required in order to observe STAT3 Ser727 phosphorylation in HK-2 but not in 786-O cells. We also observed that CK2β improved the prognostic value of p-STAT3 Ser727, as CK2βcyt>41 (median value) discriminates patients free of disease for a period of 10 years upon surgery, from those with CK2βcyt<41, when p-STAT3 Ser727levels are low. We conclude that CK2β down-regulation might represent a mechanism to support EMT and angiogenesis and that CK2βcyt levels are instrumental to refine prognosis of ccRCC patients with low p-STAT3 Ser727 levels.

To analyze how Proclarix is valuable to appropriately select candidates for multiparametric magnetic resonance imaging (mpMRI) and derived biopsies, among men with suspected prostate cancer (PCa). Proclarix is a new marker computing the clinically significant PCa (csPCa) risk, based on serum thosmbospondin-1, cathepsin D, prostate-specific antigen (PSA) and percent free PSA, in addition to age, that has been developed in men with serum PSA 2 to 10 ng/mL, prostate volume ≥35 mL, and normal digital rectal examination (DRE).