BACKGROUND Extracellular vesicles (EVs) contain a rich cargo of different RNA species with specialized functions and clinical applications. However, the landscape and characteristics of extracellular vesicle long RNA (exLR) in human blood remain largely unknown. METHODS We presented an optimized strategy for exLR sequencing (exLR-seq) of human plasma. The sample cohort included 159 healthy individuals, 150 patients with cancer (5 cancer types), and 43 patients with other diseases. Bioinformatics approaches were used to analyze the distribution and features of exLRs. Support vector machine algorithm was performed to construct the diagnosis classifier, and diagnostic efficiency was evaluated by ROC analysis. RESULTS More than 10000 exLRs, including mRNA, circRNA, and lncRNA, were reliably detected in each exLR-seq sample from 1–2 mL of plasma. We observed that blood EVs contain a substantial fraction of intact mRNAs and a large number of assembling spliced junctions; circRNA was also enriched in blood EVs. Interestingly, blood exLRs reflected their tissue origins and the relative fractions of different immune cell types. Additionally, the exLR profile could distinguish patients with cancer from healthy individuals. We further showed that 8 exLRs can serve as biomarkers for hepatocellular carcinoma (HCC) diagnosis with high diagnostic efficiency in training [area under the curve (AUC) = 0.9527; 95% CI, 0.9170–0.9883], validation cohort (AUC = 0.9825; 95% CI, 0.9606–1), and testing cohort (AUC = 0.9627; 95% CI, 0.9263–0.9991). CONCLUSIONS In summary, this study revealed abundant exLRs in human plasma and identified diverse specific markers potentially useful for cancer diagnosis.

Detection of breast cancer at an early stage is the key for successful treatment and improvement of outcome. However the limitations of mammography are well recognized, especially for those women with premenopausal breast cancer. Novel approaches to breast cancer screening are necessary, especially in the developing world where mammography is not feasible. In this study, we examined the promoter methylation of six genes (SFN, P16, hMLH1, HOXD13, PCDHGB7 and RASSF1a) in circulating free DNA (cfDNA) extracted from serum. We used a high-throughput DNA methylation assay (MethyLight) to examine serum from 749 cases including breast cancer patients, patients with benign breast diseases and healthy women. The six-gene methylation panel test achieved 79.6% and 82.4% sensitivity with a specificity of 72.4% and 78.1% in diagnosis of breast cancer when compared with healthy and benign disease controls, respectively. Moreover, the methylation panel positive group showed significant differences in the following independent variables: (a) involvement of family history of tumors; (b) a low proliferative index, ki-67; (c) high ratios in luminal subtypes. Additionally the panel also complemented some breast cancer cases which were neglected by mammography or ultrasound. These data suggest that epigenetic markers in serum have potential for diagnosis of breast cancer.

BackgroundChemoresistance has become a major obstacle for cancer therapy in clinic. Long noncoding RNAs (lncRNAs) have been reported to play critical roles in the development of chemoresistance in various tumors, including gastric cancer (GC). However, the role of HOXA transcript at the distal tip (HOTTIP) within extracellular vesicles (exosomes) in cisplatin-resistant GC cells remains largely unknown.Materials and methodsCell proliferation, migration and invasion were detected using Cell Counting Kit-8 (CCK-8) and transwell assays, respectively. Western blot assay was employed to analyze the protein levels of E-cadherin, N-cadherin, Vimentin, CD63, CD83, GRP78, HMGA1, and high-mobility group A1 (HMGA1). The expression levels of HOTTIP, microRNA-218 (miR-218) and HMGA1were measured by quantitative real-time polymerase chain reaction (qRT-PCR). The interaction between miR-218 and HOTTIP or HMGA1 was predicted by bioinformatics software and confirmed by the dual-luciferase reporter and RNA immunoprecipitation (RIP) assays.ResultsCell proliferation, migration, invasion, and epithelial-mesenchymal transition (EMT) were promoted in cisplatin-resistant GC cells. HOTTIP level was upregulated in cisplatin-resistant GC cells and its downregulation enhanced cisplatin sensitivity. Moreover, extracellular HOTTIP could be incorporated into exosomes and transmitted to sensitive cells, thus disseminating cisplatin resistance. Additionally, exosomal HOTTIP promoted cisplatin resistance via activating HMGA1 in GC cells. Interestingly, HMGA1 was a target of miR-218 and miR-218 could directly bind to HOTTIP. Clinically, high expression of exosomal HOTTIP in serum was associated with poor response to cisplatin treatment in GC patients.ConclusionExosomal HOTTIP contributed to cisplatin resistance in GC cells by regulating miR-218/HMGA1 axis, providing a novel avenue for the treatment of GC.

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