BackgroundCirculating cell-free miRNAs have emerged as promising minimally-invasive biomarkers for early detection, prognosis and monitoring of cancer. They can exist in the bloodstream incorporated into extracellular vesicles (EVs) and ribonucleoprotein complexes. However, it is still debated if EVs contain biologically meaningful amounts of miRNAs and may provide a better source of miRNA biomarkers than whole plasma. The aim of this study was to systematically compare the diagnostic potential of prostate cancer-associated miRNAs in whole plasma and in plasma EVs.MethodsRNA was isolated from whole plasma and plasma EV samples from a well characterised cohort of 50 patient with prostate cancer (PC) and 22 patients with benign prostatic hyperplasia (BPH). Nine miRNAs known to have a diagnostic potential for PC in cell-free blood were quantified by RT-qPCR and the relative quantities were compared between patients with PC and BPH and between PC patients with Gleason score ≥ 8 and ≤6.ResultsOnly a small fraction of the total cell-free miRNA was recovered from the plasma EVs, however the EV-incorporated and whole plasma cell-free miRNA profiles were clearly different. Four of the miRNAs analysed showed a diagnostic potential in our patient cohort. MiR-375 could differentiate between PC and BPH patients when analysed in the whole plasma, while miR-200c-3p and miR-21-5p performed better when analysed in plasma EVs. EV-incorporated but not whole plasma Let-7a-5p level could distinguish PC patients with Gleason score ≥ 8 vs ≤6.ConclusionsThis study demonstrates that for some miRNA biomarkers EVs provide a more consistent source of RNA than whole plasma, while other miRNAs show better diagnostic performance when tested in the whole plasma.

Adult stem cells (SCs) participate in tissue repair and homeostasis regulation. The relative ease of SC handling and their therapeutic effect has made of these cell popular candidates for cellular therapy. However, several problems interfere with their clinical application in cancer treatment, like safety issues, unpredictable pro-tumour effects, and tissue entrapment. Therefore cell-free therapies that exhibit SC properties are being investigated. It is now well known that adult SCs exhibit their therapeutic effect via paracrine mechanisms. In addition to secretory proteins, SCs also release extracellular vesicles (EV) that deliver their contents to the target cells. Cancer treatment is one of the most promising applications of SC-EVs. Moreover, SC-EVs could be modified to improve targeted drug delivery. The aim of the review is to summarise current knowledge of adult SC-EV application in cancer treatment and to emphasise future opportunities and challenges in cancer treatment.

Introduction: Extracellular vesicles (EVs) have emerged as a very attractive source of cancer- derived RNA biomarkers for the early detection, prognosis and monitoring of various cancers, including prostate cancer (PC). However, biofluids contain a mixture of EVs released from a variety of tissues and the fraction of total EVs that are derived from PC tissue is not known. Moreover, the optimal biofluid—plasma or urine—that is more suitable for the detection of EV- enclosed RNA biomarkers is not yet clear.Methodology: In the current study, we performed RNA sequencing analysis of plasma and urinary EVs collected before and after radical prostatectomy, and matched tumor and normal prostate tissues of 10 patients with prostate cancer.Results and Discussion: The most abundant RNA biotypes in EVs were miRNA, piRNA, tRNA, lncRNA, rRNA and mRNA. To identify putative cancer-derived RNA biomarkers, we searched for RNAs that were overexpressed in tumor as compared to normal tissues, present in the pre-operation EVs and decreased in the post-operation EVs in each RNA biotype. The levels of 63 mRNAs, 3 lncRNAs, 2 miRNAs and 1 piRNA were significantly increased in the tumors and decreased in the post-operation urinary EVs, thus suggesting that these RNAs mainly originate from PC tissue. No such RNA biomarkers were identified in plasma EVs. This suggests that the fraction of PC-derived EVs in urine is larger than in plasma and allows the detection and tracking of PC-derived RNAs.

adjacent normal tissues. The upregulated NUB1 transcripts were observed in the breast cancer. METABRIC cohort highlighted that patients with low NUB1 transcripts had a poorer survival in the ER-negative subgroup of breast cancer patients [hazard ratio (HR)=0.66, 95% confidence interval (CI)=0. 5-0.87, p=0.003] and triple negative subgroup (HR=0.67, 95% CI=0. 47-0.96, p=0.028). NUB1 knockdown inhibits in vitro cell growth in MDA-MB-231. AIPL1 protein forms multimers in cancer cells. NUB1 protein moved into the nucleus in hypoxia (0.1% O 2 48 hours). p21 and p27 proteins accumulated in NUB1-knockdown MDA-MB-231 cells. The prolonged G 0 /G 1 cell cycle arrests resulted in cell death through the neddylation-dependent CUL1-p27-p21 and CUL2-VHL axis. We also demonstrated that HIF1a protein could be neddylated upon reoxygenation. In a retrospective study of Oxford breast cancer cohort, the lower cytoplasmic expression (n=57) prognosed worse overall survival than higher cytoplasmic expression (n=57) (HR=1.779, 95% CI=1. 006-3.346, p=0.048 Introduction Sarcomas often show a limited response to current treatments. A hypothesis to explain this resistance relies on the existence of subpopulations of drug-resistant cancer stem cells (CSCs) responsible for relapses and metastasis. Therefore, there is a need for patient-close disease models amenable for testing the effect of anti-cancer therapies on CSCs. As a proof-of principle of their possible utility in future personalise medicine strategies, we have established a panel of patient-derived sarcoma primary cell lines, characterised their CSC subpopulations and tested the anti-tumour activity of the mithramycin analogue EC-8042 on these cell lines. Material and methods Fresh surgical samples of sarcomas were disaggregated and put in culture. In those cell lines adapted to grow in vitro, their proliferation ability and their potential to growth tumour xenografts were evaluated. CSCs subpopulations were characterised according to their ability to grow as tumorspheres, and the presence of cells presenting ALDH1 activity (Aldefluor © activity) or transcriptional activity due to SOX2/OCT4 in cells expressing the reporter system Sore6 (Sore6 activity). Cell survival, apoptotic induction and modulation of Sore6 activity was analysed after drug treatments. Finally, the expression/phosphorylation of signalling proteins was analysed by Western blotting. Results and discussions We have established a panel of patientderived primary cell lines able to reproduce in vivo the histopathological features of their tumours of origin. Most of these cell lines were able to grow as serially passaged tumorspheres and showed between 1% and 15% of Aldefluor © positive cells. In addition, we detected Sore6 activity in 25% of cells in a cell line expressing the Sore6 reporter. Finally, these cell lines showed a distinct sensitivity to EC8042, with IC 50 values ranging from 0.1 to 1 mM. EC-8042 was able to inhibit the expression of CSC-related factors such as SOX2, NOTCH1 and C-MYC in a dose-dependent fa...