Identification of miRNAs Enriched in Extracellular Vesicles Derived from Serum Samples of Breast Cancer Patients

Abstract

MicroRNAs derived from extracellular vesicles (EV-miRNAs) are circulating miRNAs considered as potential new diagnostic markers for cancer that can be easily detected in liquid biopsies. In this study, we performed RNA sequencing analysis as a screening strategy to identify EV-miRNAs derived from serum of clinically well-annotated breast cancer (BC) patients from the south of Brazil. EVs from three groups of samples (healthy controls (CT), luminal A (LA), and triple-negative (TNBC)) were isolated from serum using a precipitation method and analyzed by RNA-seq (screening phase). Subsequently, four EV-miRNAs (miR-142-5p, miR-150-5p, miR-320a, and miR-4433b-5p) were selected to be quantified by quantitative real-time PCR (RT-qPCR) in individual samples (test phase). A panel composed of miR-142-5p, miR-320a, and miR-4433b-5p distinguished BC patients from CT with an area under the curve (AUC) of 0.8387 (93.33% sensitivity, 68.75% specificity). The combination of miR-142-5p and miR-320a distinguished LA patients from CT with an AUC of 0.9410 (100% sensitivity, 93.80% specificity). Interestingly, decreased expression of miR-142-5p and miR-150-5p were significantly associated with more advanced tumor grades (grade III), while the decreased expression of miR-142-5p and miR-320a was associated with a larger tumor size. These results provide insights into the potential application of EVs-miRNAs from serum as novel specific markers for early diagnosis of BC.

MicroRNAs from Liquid Biopsy Derived Extracellular Vesicles: Recent Advances in Detection and Characterization Methods

“…However, this technique is time-consuming, labor-intense, and requires big sample volumes [108][109][110]. A method already employed on a more regular basis is EV precipitation [72,89,93]. Several comparison studies of these two methods exist.…”

“…Therefore, we will focus on the EV miRNA signature identified in other biofluid specimens as alternative biomarker sources. Current studies focus on the extracellular vesicles mainly isolated from human blood, or, respectively, serum or plasma [ 71 , 72 , 73 , 74 ]. However, extracellular vesicles can be isolated from a broad variety of human body fluids such as urine [ 75 , 76 ], saliva [ 77 , 78 ], bronchoalveolar liquid [ 79 ], pleural lavage [ 80 ], and cerebrospinal fluid [ 81 ] and even more uncommon ones like tears [ 82 ], semen [ 83 ], menstrual blood [ 84 ], peritoneal lavage [ 85 , 86 ], bile [ 87 ], and pancreatic juice [ 88 ].…”

Circulating MicroRNAs as Prognostic and Therapeutic Biomarkers in Breast Cancer Molecular Subtypes

“…Finally, Ozawa et al [69] analyzed miRNAs extracted from extracellular vesicles of 16 Luminal A and 15 TNBC patients compared to the controls. They selected four miRNAs (miR-142-5p, -150-5p, -320a, -4433b-5p) and demonstrated that miR-142-5p, miR-320a, and miR-4433b-5p were able to distinguish BC patients from healthy individuals with 93% sensitivity and 68% specificity.…”

Modulatory Role of microRNAs in Triple Negative Breast Cancer with Basal-Like Phenotype

“…EMT; cancer stem cell-like property; cytoskeleton reorganization [148,154] Cell proliferation, migration, and invasion [162] miR-155 TSPAN5 Promotion of stem cell proliferation and cellular proliferation [18,163] miR-150 [18] miR-142-5p miR-142-3p PTEN Cell proliferation and apoptosis [18,75,164,165] miR-135b…”