Benchmarking blood collection tubes and processing intervals for extracellular vesicle performance metrics.

The analysis of extracellular vesicles (EV) in blood samples is under intense investigation and holds the potential to deliver clinically meaningful biomarkers for health and disease. Technical variation must be minimized to confidently assess EV-associated biomarkers, but the impact of pre-analytics on EV characteristics in blood samples remains minimally explored. We present the results from the first large-scale EV Blood Benchmarking (EVBB) study in which we systematically compared 11 blood collection tubes (BCT; six preservation and five non-preservation) and three blood processing intervals (BPI; 1, 8 and 72 h) on defined performance metrics (n = 9).

Integrins are enriched on aberrantly fucosylated tumour-derived urinary extracellular vesicles.

Urinary extracellular vesicles (uEVs) are enriched with glycosylated proteins which have been extensively studied as putative biomarkers of urological cancers. Here, we characterized the glycosylation and integrin profile of EVs derived from urological cancer cell lines. We used fluorescent europium-doped nanoparticles coated with lectins and antibodies to identify a biomarker combination consisting of integrin subunit alpha 3 (ITGA3) and fucose.

Removal of platelets from blood plasma to improve the quality of extracellular vesicle research.

Background: Blood plasma is commonly used for biomarker research of extracellular vesicles (EVs). Removing all cells prior to analysis of EVs is essential.

Objectives: We therefore studied the efficacy of the most commonly used centrifugation protocol to prepare cell-free plasma.

Multicentre, prospective study on local treatment of metastatic prostate cancer (LoMP study).

Objectives: To investigate the role of cytoreductive radical prostatectomy in addition to standard of care for patients with newly diagnosed metastatic prostate cancer.

Materials And Methods: This multicentre, prospective study included asymptomatic patients from 2014 to 2018 (NCT02138721). Cytoreductive radical prostatectomy was offered to all fit patients with resectable tumours, resulting in 40 patients.

Detection of bladder cancer with aberrantly fucosylated ITGA3.

We describe a simple, non-invasive assay to identify fucosylated-glycoisoform of integrin alpha-3 (ITGA3) directly from unprocessed urine. ITGA3 was detected directly from the urine of bladder cancer (BlCa) (n = 13) and benign prostatic hyperplasia (BPH) (n = 9) patients with the use of lectins coated on europium-doped-nanoparticles (Eu-NPs). Lectin Ulex europaeus agglutinin-I (UEA) showed enhanced binding with BlCa-derived ITGA3.

Preparation of Multi-omics Grade Extracellular Vesicles by Density-Based Fractionation of Urine.

The multidimensional cargo of extracellular vesicles (EV) released in urine is a reflection of the pathophysiological processes occurring within their cells and tissues of origin in the urogenital system. Here, we describe a step-by-step protocol for density-based separation of urinary EV with high specificity and repeatability. The implementation of integrative omics allows the study of the molecular complexity of highly purified urinary EV, supporting the identification of EV-specific functions and biomarkers.

Unravelling the proteomic landscape of extracellular vesicles in prostate cancer by density-based fractionation of urine.

Extracellular vesicles (EV) are increasingly being recognized as important vehicles of intercellular communication and promising diagnostic and prognostic biomarkers in cancer. Despite this enormous clinical potential, the plethora of methods to separate EV from biofluids, providing material of highly variable purity, and lacking knowledge regarding methodological repeatability pose a barrier to clinical translation. Urine is considered an ideal proximal fluid for the study of EV in urological cancers due to its direct contact with the urogenital system.

Intradetrusor onabotulinum-a toxin injections in children with therapy-resistant idiopathic detrusor overactivity. A retrospective study.

Introduction And Objective: The use of intravesical onabotulinum-A toxin (BoNT-A) injections in the treatment of idiopathic detrusor overactivity has been widely studied in adults [2-5]. However, in pediatric populations, study groups are small, and results are not yet sufficient to support this treatment as a standard practice. The aim of this study is to determine the effectiveness and safety of this treatment in children with non-neurogenic detrusor overactivity, resistant to conservative therapy.

Performance assessment of total RNA sequencing of human biofluids and extracellular vesicles.

RNA profiling has emerged as a powerful tool to investigate the biomarker potential of human biofluids. However, despite enormous interest in extracellular nucleic acids, RNA sequencing methods to quantify the total RNA content outside cells are rare. Here, we evaluate the performance of the SMARTer Stranded Total RNA-Seq method in human platelet-rich plasma, platelet-free plasma, urine, conditioned medium, and extracellular vesicles (EVs) from these biofluids.

The generation and use of recombinant extracellular vesicles as biological reference material.

Recent years have seen an increase of extracellular vesicle (EV) research geared towards biological understanding, diagnostics and therapy. However, EV data interpretation remains challenging owing to complexity of biofluids and technical variation introduced during sample preparation and analysis. To understand and mitigate these limitations, we generated trackable recombinant EV (rEV) as a biological reference material.

Discovery and validation of a serum microRNA signature to characterize oligo- and polymetastatic prostate cancer: not ready for prime time.

Purpose: Patients with oligometastatic prostate cancer (PC) may benefit from metastasis-directed therapy (MDT), delaying disease progression and the start of palliative systemic treatment. However, a significant proportion of oligometastatic PC patients progress to polymetastatic PC within a year following MDT, suggesting an underestimation of the metastatic load by current staging modalities. Molecular markers could help to identify true oligometastatic patients eligible for MDT.

Abiraterone and spironolactone in prostate cancer: a combination to avoid.

Disease progression in metastatic castration-resistant prostate cancer (mCRPC) is dependent on androgen signaling. This case describes the complex adaptive androgen signaling mechanisms in mCRPC and illustrates that caution should be exercised when treating these patients with drugs influencing the androgen axis. Single case report and review of the literature.

Importance of metastatic volume in prognostic models to predict survival in newly diagnosed metastatic prostate cancer.

Purpose: To explore the prognostic importance of metastatic volume in a contemporary daily practice cohort of patients with newly diagnosed metastatic hormone-naive prostate cancer (mHNPC) and to develop a pragmatic prognostic model to predict survival for these patients.

Methods: Since 2014, 113 patients with newly diagnosed mHNPC were prospectively registered. Statistical analysis was performed using SPSS 25.

Urinary extracellular vesicle biomarkers in urological cancers: From discovery towards clinical implementation.

Urine contains cellular elements, biochemicals, and proteins derived from glomerular filtration of plasma, renal tubule excretion, and urogenital tract secretions that reflect an individual's metabolic and pathophysiologic state. Despite intensive research into the discovery of urinary biomarkers to facilitate early diagnosis, accurate prognosis and prediction of therapy response in urological cancers, none of these markers has reached widespread use. Their implementation into daily clinical practice is hampered by a substantial degree of heterogeneity in performance characteristics and uncertainty about reliability, clinical utility and cost-effectiveness, in addition to several technical limitations.

Prognostic and Therapeutic Implications of Circulating Androgen Receptor Gene Copy Number in Prostate Cancer Patients Using Droplet Digital Polymerase Chain Reaction.

Background: Resistance mechanisms in the androgen receptor (AR) signaling pathway remain key drivers in the progression to castration-resistant prostate cancer (CRPC) and relapse under antihormonal therapy.

Materials And Methods: We evaluated the circulating AR gene copy number (CN) gain using droplet digital polymerase chain reaction in 21 control and 91 prostate cancer serum samples and its prognostic and therapeutic implications in prostate cancer.

Results: In CRPC, AR CN gain was associated with faster progression to CRPC (P = .

Metastatic burden in newly diagnosed hormone-naive metastatic prostate cancer: Comparing definitions of CHAARTED and LATITUDE trial.

Objectives: No uniformity exists in the definition of metastatic burden in metastatic hormone-naive prostate cancer (mHNPC) across clinical trials making their comparison challenging. We explored definition agreement and prognostic significance of bulky mHNPC according to the CHAARTED and LATITUDE trial.

Materials And Methods: Since 2014, 95 patients with newly diagnosed mHNPC were prospectively registered.

Confounding factors of ultrafiltration and protein analysis in extracellular vesicle research.

Identification and validation of extracellular vesicle (EV)-associated biomarkers requires robust isolation and characterization protocols. We assessed the impact of some commonly implemented pre-analytical, analytical and post-analytical variables in EV research. Centrifugal filters with different membrane types and pore sizes are used to reduce large volume biofluids prior to EV isolation or to concentrate EVs.

EV-TRACK: transparent reporting and centralizing knowledge in extracellular vesicle research.

We argue that the field of extracellular vesicle (EV) biology needs more transparent reporting to facilitate interpretation and replication of experiments. To achieve this, we describe EV-TRACK, a crowdsourcing knowledgebase (http://evtrack.org) that centralizes EV biology and methodology with the goal of stimulating authors, reviewers, editors and funders to put experimental guidelines into practice.

Whole pelvis radiotherapy for pathological node-positive prostate cancer : Oncological outcome and prognostic factors.

Purpose: The goal of this work was to investigate the oncological outcome of whole pelvis radiotherapy (wpRT) in pathologic pelvic lymph node-positive (pN1) prostate cancer (PCa), evaluate the location of relapse, and identify potential prognostic factors.

Patients And Methods: All patients undergoing pelvic lymph node dissection (PLND) since the year 2000 at a single tertiary care center were evaluated. A total of 154 patients with pN1 PCa were treated with wpRT (39 in an adjuvant setting) and 2-3 years of androgen deprivation therapy (ADT).

Function of extracellular vesicle-associated miRNAs in metastasis.

Extracellular RNA (exRNA) is functionally transferrable from donor to recipient cells and is protected from RNAses by electrostatic interactions with proteins or by membrane encapsulation. In addition to bioactive RNA, extracellular vesicles (EVs) contain intraluminal and membrane-associated proteins. The cellular context and fitness affect the composition of EVs and thus the outcome of the communication between the EV-producer and recipient cells.