Small extracellular vesicles (sEVs) are nanosized membrane-enclosed sacs secreted by living cells into the surrounding extracellular space. They do not contain functional nuclei and are incapable of replication [1]. The sEVs are commonly categorized into two primary subtypes based on their origin and features: ectosomes or microvesicles (MVEs) and exosomes. Ectosomes typically range from 150 to 2000 nm in size and are formed through the outward budding of the plasma membrane [2]. Exosomes are typically smaller, ranging from 30 to 150 nm in diameter. They are originated as intraluminal vesicles (ILVs) within multivesicular bodies (MVBs) during endosome maturation and are released into extracellular space when MVBs fuse with the plasma membrane [[3], [4], [5]]. Exosomes were discovered over three decades ago, and were first studied by Rose Johnstone in 1980s in the reticulocytes of rats and sheep [4,5]. However, according to the 2018 guidelines from the International Society for Extracellular Vesicles (MISEV2018), exosomes should be included under the umbrella term of “sEVs”, accompanied by specific descriptors such as size, density, cell source, and experimental conditions, to avoid overrated claims on the specific vesicle types having overlapping size ranges that are practically impossible to isolate without overlap [2,6]. sEVs are naturally derived carriers that offer several advantages over synthetic delivery systems. Due to their capacity to transport DNA, RNA, proteins, small molecules, and other therapeutic agents, sEVs are being largely explored as promising drug delivery vehicles for the treatment of cancer and various other diseases [7]. Notably, cancer cells produce and release a higher amount of sEVs compared to normal cells, distributing them into both the tumor microenvironment (TME) and the bloodstream [8]. Oncogenic factors transported by sEVs can promote cell proliferation, immune system evasion, and angiogenesis, all of which contribute to tumor growth and progression [9]. Early research on sEVs in cancer primarily focused on their involvement in altering the tumor microenvironment (TME), advancing tumor development, and facilitating metastasis. More recently, their potential as anti-cancer agents have gained attention due to their ability to modulate immune responses [8].

High-throughput proteomics provides a quick platform of identifying the protein-based biomarkers from various biological sample sources. Likewise, several proteomics studies on EV proteome have been reported throughout the last decade. Among them, the quantitative proteomics studies provide information on the quantitative abundance for the list of identified EV proteomes.

Current review aims to deliver a comprehensive overview of the quantitative proteomics investigations that have been carried out on the sEV protein contents. The write up systematically describes sEVs and the workflow that one should follow while working with sEV proteome for a quantitative analysis using mass-spectrometry-based proteomics tools. This review focuses particularly on the studies that utilized clinical cancer samples for sEV isolation followed by characterization of the proteome contents. Proteins belonging to the functional classes of adhesion molecules, inflammatory molecules and immunoglobulins were highly enriched in serum-derived sEVs of various cancer patients. Interestingly some of these proteins including Apolipoprotein A 4 (APOA4), Serum amyloid A-4 protein (SAA4), Hemoglobin subunit Delta (HBD) and Hemoglobin subunit Beta (HBB) were reported in multiple studies involving diverse cancers and sample types. These candidates thus in particular should further be studied for their corresponding translational use in diagnosis and treatment of cancers.