Extracellular vesicles as a source of prostate cancer biomarkers in liquid biopsies: a decade of research

AbstractProstate cancer is a global cancer burden and considerable effort has been made through the years to identify biomarkers for the disease. Approximately a decade ago, the potential of analysing extracellular vesicles in liquid biopsies started to be envisaged. This was the beginning of a new exciting area of research investigating the rich molecular treasure found in extracellular vesicles to identify biomarkers for a variety of diseases. Vesicles released from prostate cancer cells and cells of the tumour microenvironment carry molecular information about the disease that can be analysed in several biological fluids. Numerous studies document the interest of researchers in this field of research. However, methodological issues such as the isolation of vesicles have been challenging. Remarkably, novel technologies, including those based on nanotechnology, show promise for the further development and clinical use of extracellular vesicles as liquid biomarkers. Development of biomarkers is a long and complicated process, and there are still not many biomarkers based on extracellular vesicles in clinical use. However, the knowledge acquired during the last decade constitutes a solid basis for the future development of liquid biopsy tests for prostate cancer. These are urgently needed to bring prostate cancer treatment to the next level in precision medicine.

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Immunomagnetic sequential ultrafiltration (iSUF) platform for enrichment and purification of extracellular vesicles from biofluids

Scientific Reports ◽

10.1038/s41598-021-86910-y ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Jingjing Zhang ◽

Luong T. H. Nguyen ◽

Richard Hickey ◽

Nicole Walters ◽

Xinyu Wang ◽

...

Keyword(s):

Extracellular Vesicles ◽

Culture Media ◽

Metastatic Breast ◽

Clinical Samples ◽

Clinical Use ◽

Immunomagnetic Beads ◽

Liquid Biopsies ◽

Cell Culture Media ◽

Non Invasive ◽

Healthy Donors

AbstractExtracellular vesicles (EVs) derived from tumor cells have the potential to provide a much-needed source of non-invasive molecular biomarkers for liquid biopsies. However, current methods for EV isolation have limited specificity towards tumor-derived EVs that limit their clinical use. Here, we present an approach called immunomagnetic sequential ultrafiltration (iSUF) that consists of sequential stages of purification and enrichment of EVs in approximately 2 h. In iSUF, EVs present in different volumes of biofluids (0.5–100 mL) can be significantly enriched (up to 1000 times), with up to 99% removal of contaminating proteins (e.g., albumin). The EV recovery rate by iSUF for cell culture media (CCM), serum, and urine corresponded to 98.0% ± 3.6%, 96.0% ± 2.0% and 94.0% ± 1.9%, respectively (p > 0.05). The final step of iSUF enables the separation of tumor-specific EVs by incorporating immunomagnetic beads to target EV subpopulations. Serum from a cohort of clinical samples from metastatic breast cancer (BC) patients and healthy donors were processed by the iSUF platform and the isolated EVs from patients showed significantly higher expression levels of BC biomarkers (i.e., HER2, CD24, and miR21).

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Diagnostic and prognostic potential of the proteomic profiling of serum-derived extracellular vesicles in prostate cancer

Cell Death and Disease ◽

10.1038/s41419-021-03909-z ◽

2021 ◽

Vol 12 (7) ◽

Author(s):

Michele Signore ◽

Romina Alfonsi ◽

Giulia Federici ◽

Simona Nanni ◽

Antonio Addario ◽

...

Keyword(s):

Prostate Cancer ◽

Extracellular Vesicles ◽

Biological Fluids ◽

Prognostic Significance ◽

Diagnostic Procedures ◽

Protein Microarrays ◽

Specific Protein ◽

Clinical Settings ◽

Proteomic Profiling ◽

Molecular Tests

AbstractExtracellular vesicles (EVs) and their cargo represent an intriguing source of cancer biomarkers for developing robust and sensitive molecular tests by liquid biopsy. Prostate cancer (PCa) is still one of the most frequent and deadly tumor in men and analysis of EVs from biological fluids of PCa patients has proven the feasibility and the unprecedented potential of such an approach. Here, we exploited an antibody-based proteomic technology, i.e. the Reverse-Phase Protein microArrays (RPPA), to measure key antigens and activated signaling in EVs isolated from sera of PCa patients. Notably, we found tumor-specific protein profiles associated with clinical settings as well as candidate markers for EV-based tumor diagnosis. Among others, PD-L1, ERG, Integrin-β5, Survivin, TGF-β, phosphorylated-TSC2 as well as partners of the MAP-kinase and mTOR pathways emerged as differentially expressed endpoints in tumor-derived EVs. In addition, the retrospective analysis of EVs from a 15-year follow-up cohort generated a protein signature with prognostic significance. Our results confirm that serum-derived EV cargo may be exploited to improve the current diagnostic procedures while providing potential prognostic and predictive information. The approach proposed here has been already applied to tumor entities other than PCa, thus proving its value in translational medicine and paving the way to innovative, clinically meaningful tools.

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The itinerary of circulating miRNAs – implications for cancer progression and diagnosis

Journal of Medical Science ◽

10.20883/medical.e516 ◽

2021 ◽

Vol 90 (2) ◽

pp. e516

Author(s):

Przemysław Szałata ◽

Anna-Maria Guner ◽

Michalina Raczkowska ◽

Julia Smyrek ◽

Dominika Szaj ◽

...

Keyword(s):

Nucleic Acids ◽

Extracellular Vesicles ◽

Cancer Progression ◽

Target Cell ◽

Biological Fluids ◽

Tumour Microenvironment ◽

High Density Lipoproteins ◽

Cancer Type ◽

Circulating Mirna ◽

Extracellular Mirna

microRNAs (miRNAs) are non-coding RNAs that regulate gene expression and protect cells from foreign nucleic acids. miRNA is produced in the nucleus and processed in the cytoplasm. These small nucleic acid molecules are released from cells to the extracellular matrix (extracellular miRNA, ex-miRNA) and reach blood plasma (circulating miRNA). Circulating miRNA can also be detected in other biological fluids, such as saliva, cerebrospinal fluid or urine, and it is usually carried by proteins or extracellular vesicles. Argonaute-miRNA, or miRNA-lipoprotein complex, protect miRNA from being degraded. The entrance of extracellular miRNA into a target cell is mediated by endocytosis and membrane fusion of extracellular vesicles. Additionally, miRNA can also be delivered in high-density lipoproteins by means of interactions with scavenger receptors. miRNAs absorbed into a cell can act as tumour promoters (oncomirs), or suppressors by inhibiting the translation process of the target mRNAs, thus, affecting cells in the tumour microenvironment. miRNA can impact other cells by supporting tumour growth, promoting angiogenesis and modulating the immune system. Molecular high-throughput methods are employed to detect circulating miRNA, and a potentially helpful diagnostic test has been designed to characterise the cancer type. In this review, we aim to summarise the itinerary of miRNAs from a source cell to a target cell, as well as to show how this class of small nucleic acids participates in intercellular communication. Finally, we highlight examples of miRNAs usage as potential molecular markers and discuss treatment approaches in clinical trials.

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Potential of miRNAs in urinary extracellular vesicles for management of active surveillance in prostate cancer patients

British Journal of Cancer ◽

10.1038/s41416-021-01598-1 ◽

2021 ◽

Author(s):

Manuel Ramirez-Garrastacho ◽

Viktor Berge ◽

Aija Linē ◽

Alicia Llorente

Keyword(s):

Prostate Cancer ◽

Cancer Patients ◽

Extracellular Vesicles ◽

Active Surveillance ◽

Liquid Biopsies ◽

Qpcr Analysis ◽

Risk Prostate Cancer ◽

Low Risk Prostate Cancer ◽

Grade 3 ◽

Independent Cohort

Abstract Background Active surveillance is an alternative to radical treatment for patients with low-risk prostate cancer, which could also benefit some patients with intermediate risk. We have investigated the use of miRNA in urinary extracellular vesicles to stratify these patients. Methods NGS was performed to profile the miRNAs from small urinary extracellular vesicles in a cohort of 70 patients with prostate cancer ISUP Grade 1, 2 or 3. The most promising candidates were then analysed by RT-qPCR in a new cohort of 60 patients. Results NGS analysis identified nine miRNAs differentially expressed in at least one of the comparisons. The largest differences were found with miR-1290 (Grade 3 vs. 1), miR-320a-3p (Grade 3 vs. 2) and miR-155-5p (Grade 2 vs. 1). Combinations of 2–3 miRNAs were able to differentiate between two ISUP grades with an AUC 0.79–0.88. RT-qPCR analysis showed a similar trend for miR-186-5p and miR-30e-5p to separate Grade 3 from 2, and miR-320a-3p to separate Grade 2 from 1. Conclusions Using NGS, we have identified several miRNAs that discriminate between prostate cancer patients with ISUP Grades 1, 2 and 3. Moreover, miR-186-5p, miR-320a-3p and miR-30e-5p showed a similar behaviour in an independent cohort using an alternative analytical method. Our results show that miRNAs from urinary vesicles can be potentially useful as liquid biopsies for active surveillance.

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Emerging Role of Extracellular Vesicles in Prostate Cancer

Endocrinology ◽

10.1210/endocr/bqab139 ◽

2021 ◽

Author(s):

Megan Ludwig ◽

Rhea Rajvansh ◽

Justin M Drake

Keyword(s):

Prostate Cancer ◽

Extracellular Vesicles ◽

Prostate Gland ◽

The United States ◽

Liquid Biopsies ◽

Cellular Processes ◽

Non Invasive ◽

Common Cancer ◽

Meaningful Communication

Abstract Prostate cancer (PCa) is the second most common cancer amongst men in the United States. While the use of PSA has improved the ability to screen and ultimately diagnose PCa, there still remains false positives due to non-cancerous conditions in the prostate gland itself and other prognostic biomarkers for PCa are needed. Contents within extracellular vesicles (EVs) have emerged as promising biomarkers that can give valuable information about disease state, and have the additional benefit of being acquired through non-invasive liquid biopsies. Meaningful communication between cancer cells and the microenvironment are carried by EVs, which impact important cellular processes in prostate cancer such as metastasis, immune regulation, and drug resistance.

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Molecular Profile Study of Extracellular Vesicles for the Identification of Useful Small “Hit” in Cancer Diagnosis

Applied Sciences ◽

10.3390/app112210787 ◽

2021 ◽

Vol 11 (22) ◽

pp. 10787

Author(s):

Giusi Alberti ◽

Christian M. Sánchez-López ◽

Alexia Andres ◽

Radha Santonocito ◽

Claudia Campanella ◽

...

Keyword(s):

Extracellular Vesicles ◽

Cancer Progression ◽

Cancer Metastasis ◽

Biological Fluids ◽

Cell Communication ◽

Response To Treatment ◽

Molecular Profile ◽

Cell Of Origin ◽

Liquid Biopsies ◽

Precision Therapy

Tumor-secreted extracellular vesicles (EVs) are the main mediators of cell-cell communication, permitting cells to exchange proteins, lipids, and metabolites in varying physiological and pathological conditions. They contain signature tumor-derived molecules that reflect the intracellular status of their cell of origin. Recent studies have shown that tumor cell-derived EVs can aid in cancer metastasis through the modulation of the tumor microenvironment, suppression of the immune system, pre-metastatic niche formation, and subsequent metastasis. EVs can easily be isolated from a variety of biological fluids, and their content makes them useful biomarkers for the diagnosis, prognosis, monitorization of cancer progression, and response to treatment. This review aims to explore the biomarkers of cancer cell-derived EVs obtained from liquid biopsies, in order to understand cancer progression and metastatic evolution for early diagnosis and precision therapy.

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Role of exosomes in diagnosis and therapy of prostate cancer

I P Pavlov Russian Medical Biological Herald ◽

10.23888/pavlovj2020283399-405 ◽

2020 ◽

Vol 28 (3) ◽

pp. 399-405

Author(s):

Fabrizio Fontana ◽

Olga A. Babenko

Keyword(s):

Prostate Cancer ◽

Cancer Cells ◽

Tumor Cells ◽

Biological Fluids ◽

Diagnosis And Treatment ◽

Diagnosis And Therapy ◽

The Future ◽

Important Direction ◽

Potential Biomarkers

Aim of this letter is to attract the attention of journal readers to the study of exosomes as an important direction in the development of Oncology, in particular, in the diagnosis and treatment of prostate cancer. Exosomes are produced by tumor cells and regulate proliferation, metastasis, and the development of chemoresistance. Their extraction from biological fluids allows further use of these vesicles as potential biomarkers of prostate cancer. In the future, exosomes can be successfully used in the delivery of drugs and other anti-tumor substances to cancer cells.

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Separation of Extracellular Vesicles by DNA-Directed Immunocapturing Followed by Enzymatic Release

10.26434/chemrxiv.12234683 ◽

2020 ◽

Author(s):

Dario Brambilla ◽

Laura Sola ◽

Elisa Chiodi ◽

Natasa Zarovni ◽

Diogo Fortunato ◽

...

Keyword(s):

Cell Culture ◽

Extracellular Vesicles ◽

Culture Media ◽

Biological Fluids ◽

Imaging Techniques ◽

Cell Communication ◽

Disease Diagnosis ◽

Cell Culture Supernatant ◽

Transmission Electron ◽

Downstream Analysis

Extracellular vesicles (EVs) have attracted great interest among researchers due to their role in cell-cell communication, disease diagnosis, and drug delivery. In spite of their potential in the medical field, there is no consensus on the best method for separating microvesicles from cell culture supernatant and complex biological fluids. Obtaining a good recovery yield and preserving physical characteristics is critical for the diagnostic and therapeutic use of EVs. The separation is made complex by the fact that blood and cell culture media, contain a large number of nanoparticles in the same size range. Methods that exploit immunoaffinity capture provide high purity samples and overcome the issues of currently used separation methods. However, the release of captured nanovesicles requires harsh conditions that hinder their use in certain types of downstream analysis. Herein, a novel capture and release approach for small extracellular vesicles (sEVs), based on DNAdirected immobilization of antiCD63 antibody is presented. The flexible DNAlinker increases the capture efficiency and allows releasing of EVs by exploiting the endonucleasic activity of DNAse I. This separation protocol works under mild conditions, enabling the release of intact vesicles that can be successfully analyzed by imaging techniques. In this article sEVs recovered from plasma were characterized by established techniques for EVs analysis including nanoparticle tracking and transmission electron microscopy.<br>

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miRNA-expression in prostate cancer-associated fibroblasts and their extracellular vesicles

European Urology ◽

10.1016/s0302-2838(21)00800-9 ◽

2021 ◽

Vol 79 ◽

pp. S584

Author(s):

J. Linxweiler ◽

H. Ayoubian ◽

D. Himbert ◽

M. Saar ◽

M. Stöckle ◽

...

Keyword(s):

Prostate Cancer ◽

Extracellular Vesicles ◽

Mirna Expression ◽

Cancer Associated Fibroblasts

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Molecular and Circulating Biomarkers of Brain Tumors

International Journal of Molecular Sciences ◽

10.3390/ijms22137039 ◽

2021 ◽

Vol 22 (13) ◽

pp. 7039

Author(s):

Wojciech Jelski ◽

Barbara Mroczko

Keyword(s):

Brain Tumors ◽

Tumor Cells ◽

Circulating Tumor Cells ◽

Extracellular Vesicles ◽

Dna Methyltransferase ◽

Growth Factor Receptor ◽

Response To Treatment ◽

Liquid Biopsies ◽

Methylguanine Dna Methyltransferase ◽

The Central Nervous System

Brain tumors are the most common malignant primary intracranial tumors of the central nervous system. They are often recognized too late for successful therapy. Minimally invasive methods are needed to establish a diagnosis or monitor the response to treatment of CNS tumors. Brain tumors release molecular information into the circulation. Liquid biopsies collect and analyze tumor components in body fluids, and there is an increasing interest in the investigation of liquid biopsies as a substitute for tumor tissue. Tumor-derived biomarkers include nucleic acids, proteins, and tumor-derived extracellular vesicles that accumulate in blood or cerebrospinal fluid. In recent years, circulating tumor cells have also been identified in the blood of glioblastoma patients. In this review of the literature, the authors highlight the significance, regulation, and prevalence of molecular biomarkers such as O6-methylguanine-DNA methyltransferase, epidermal growth factor receptor, and isocitrate dehydrogenase. Herein, we critically review the available literature on plasma circulating tumor cells (CTCs), cell-free tumors (ctDNAs), circulating cell-free microRNAs (cfmiRNAs), and circulating extracellular vesicles (EVs) for the diagnosis and monitoring of brain tumor. Currently available markers have significant limitations.While much research has been conductedon these markers, there is still a significant amount that we do not yet understand, which may account for some conflicting reports in the literature.

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