Liquid biopsy refers to the sampling and molecular analysis of the biofluids of circulating tumor cells, extracellular vesicles, nucleic acids, and so forth. Exosomes are small extracellular vesicles with sizes between 30–150 nm. They are secreted by multivesicular bodies through exocytosis in live cells and can participate in intercellular communication due to their contents, including nucleic acids, proteins, and lipids. Herein, we investigate publication frequencies on exosomes over the past 10 years, and review recent clinical studies on liquid biopsy of exosomes in the fields of oncology, pregnancy disorders, cardiovascular diseases, and organ transplantation. We also describe the advantages of exosomes as an effective liquid biopsy tool and the progression of exosome extraction methods. Finally, we depict the commercial development of exosome research and discuss the future role of exosomes in liquid biopsy.

Liver metastases develop in more than half of the patients with colorectal cancer (CRC) and are associated with a poor prognosis. The factors influencing liver metastasis of CRC are poorly characterized, but this information is urgently needed. We have now discovered that small extracellular vesicles (sEVs; exosomes) derived from CRC can be specifically targeted to liver tissue and induce liver macrophage polarization toward an interleukin-6 (IL-6)-secreting proinflammatory phenotype. More importantly, we found that microRNA-21-5p (miR-21) was highly enriched in CRC-derived sEVs and was essential for creating a liver proinflammatory phenotype and liver metastasis of CRC. Silencing either miR-21 in CRC-sEVs or Toll-like receptor 7 (TLR7) in macrophages, to which miR-21 binds, abolished CRC-sEVs’ induction of proinflammatory macrophages. Furthermore, miR-21 expression in plasma-derived sEVs was positively correlated with liver metastasis in CRC patients. Collectively, our data demonstrate a pivotal role of CRC-sEVs in promoting liver metastasis by inducing an inflammatory premetastatic niche through the miR-21–TLR7–IL-6 axis. Thus, sEVs–miR-21 represents a potential prognostic marker and therapeutic target for CRC patients with liver metastasis.

Background : Early screening for colorectal cancer (CRC) is essential to improve its prognosis. Liquid biopsies are increasingly being considered for diagnosing cancer due to low invasiveness and high reproducibility. In addition, circulating extracellular vesicles (crEVs, extracellular vesicles isolated from plasma) expressing tumour‐specific proteins are potential biomarkers for various cancers. Here, we present a data‐independent acquisition (DIA)‐mass spectrometry (MS)‐based diagnostic method for liquid biopsies. Methods : Extracellular vesicles (EVs) were isolated from culture supernatants of human CRC cell lines, and plasma of patients with CRC at different tumour stages, by overnight ultracentrifugation coupled with sucrose density gradient centrifugation. Tumour‐specific EV proteins were prioritized using Tandem Mass Tag (TMT)‐based shotgun proteomics and phosphoproteomics. The results were verified in a second independent cohort and a mouse tumour‐bearing model using Western blotting (WB). The candidate biomarkers were further validated in a third cohort by DIA‐MS. Finally, the DIA‐MS methodology was accelerated to permit high‐throughput detection of EV biomarkers in another independent cohort of patients with CRC and healthy controls. Results : High levels of total and phosphorylated fibronectin 1 (FN1) in crEVs, haptoglobin (HP), S100A9 and fibrinogen α chain (FGA) were significantly associated with cancer progression. FGA was the most dominant biomarker candidate. Analysis of the human CRC cell lines and the mouse model indicated that FGA+ crEVs were likely released by CRC cells. Furthermore, fast DIA‐MS and parallel reaction monitoring (PRM)‐MS both confirmed that FGA+ crEVs could distinguish colon adenoma with an area of curve (AUC) in the receiver operating characteristic (ROC) curve of 0.949 and patients with CRC (AUC of ROC is 1.000) from healthy individuals. The performance outperformed conventional tumour biomarkers. The DIA‐MS quantification of FGA+ crEVs among three groups agreed with that from PRM‐MS. Conclusion : DIA‐MS detection of FGA+ crEVs is a potential rapid and non‐invasive screening tool to identify early stage CRC. Abbreviations: FGA: fibrinogen α chain; CRC: colorectal cancer; crEVs: circulating extracellular vesicles; EV: extracellular vesicles;MS: mass spectrometry; WB: Western blotting; ROC: receiver operating characteristic; PRM: Parallel Reaction Monitoring; GPC1: Glypican‐1; GO: Gene ontology; TEM: transmission electron microscopy; FN1: Fibronectin 1; HP: haptoglobin; TMT: Tandem Mass Tag; LC‐MS/MS: liquid chromatography coupled to tandem mass spectrometry; DIA: data‐independent acquisition; DDA: data‐dependent acquisition; CiRT: Common internal Retention Time standards;AGC: Automatic gain control; AUC: area under curve.

Prospective epidemiological studies have consistently suggested that pancreatic cancer-associated new-onset diabetes mellitus (PC-DM) represents a potential platform for early diagnose of pancreatic cancer (PC). Despite the studies performed, the mechanism behind this phenomenon remains ambiguous. In this study, we explored the effects of two types of exosomes released by murine pancreatic cancer and ductal epithelial cells on murine skeletal muscle cells. The results show that PC-derived exosomes can readily enter C2C12 myotubes, triggering lipidosis and glucose intake inhibition. We also demonstrate that PC-derived exosomes can inhibit insulin and PI3K/Akt signalling, in which insulin-induced FoxO1 nuclear exclusion is preserved and Glut4 trafficking is impaired. Microarray and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses show that exosomal microRNAs (miRNAs) probably play key roles in this process, an assumption that is corroborated by in vitro studies. These results confirm that the insulin resistance (IR) of skeletal muscle cells is governed by PC-derived exosomes through the insulin and PI3K/Akt/FoxO1 signalling pathways, where exosomal miRNAs potentially contribute to this phenomenon. These novel findings pave the way towards a comprehensive understanding of the cancer theories: “metabolic reprogramming” and “metabolic crosstalk”.

To address the increasing need for detecting and validating protein biomarkers in clinical specimens, mass spectrometry (MS)-based targeted proteomic techniques, including the selected reaction monitoring (SRM), parallel reaction monitoring (PRM), and massively parallel data-independent acquisition (DIA), have been developed. For optimal performance, they require the fragment ion spectra of targeted peptides as prior knowledge. In this report, we describe a MS pipeline and spectral resource to support targeted proteomics studies for human tissue samples. To build the spectral resource, we integrated common open-source MS computational tools to assemble a freely accessible computational workflow based on Docker. We then applied the workflow to generate DPHL, a comprehensive DIA pan-human library, from 1096 data-dependent acquisition (DDA) MS raw files for 16 types of cancer samples. This extensive spectral resource was then applied to a proteomic study of 17 prostate cancer (PCa) patients. Thereafter, PRM validation was applied to a larger study of 57 PCa patients and the differential expression of three proteins in prostate tumor was validated. As a second application, the DPHL spectral resource was applied to a study consisting of plasma samples from 19 diffuse large B cell lymphoma (DLBCL) patients and 18 healthy control subjects. Differentially expressed proteins between DLBCL patients and healthy control subjects were detected by DIA-MS and confirmed by PRM. These data demonstrate that the DPHL supports DIA and PRM MS pipelines for robust protein biomarker discovery. DPHL is freely accessible at https://www.iprox.org/page/project.html?id=IPX0001400000.