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Background: The recurrence rate of thyroid cancer can be as high as 30%. The purpose of this study was to examine changes of urine exosomal peptide levels after thyroidectomy in patients with thyroid cancer to determine if levels can predict the risk of recurrence. Methods: Patients >20 years old as newly diagnosed with papillary thyroid cancer who had received a thyroidectomy were recruited. Urine samples were collected at 12 months after enrollment to the study, and 1 year later. Urine exosomes containing different peptides were identified and compared. Results: A total of 70 patients were enrolled in the study, and were classified by the interval between surgery and enrollment: 42 patients with < 5 years between surgery and enrollment, 14 patients between 5-10 years, and 14 patients longer than 10 years. No recurrence was observed in any patient during the 2 years after enrollment. No significant differences were found in the levels of serum proteins or urine exosomal peptides between groups, or between intervals. Known risk factors for high-risk thyroid cancer had only a mild correlation with serum protein levels and urine exosomal peptides. Conclusion: Our study revealed the long-term basal fluctuation ranges of serum proteins and urine exosomal peptides in patients with thyroid cancer who underwent thyroidectomy. For high-risk patients after thyroidectomy, concentrations of serum proteins or urine exosomal peptides within the ranges may indicate there is a lower risk of thyroid cancer recurrence during long-term follow-up. Trial Registration: ClinicalTrials.gov: NCT03488134.

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OBJECTIVE: To detect the expression level of urinary exosomal lncRNA SNHG16 in patients with bladder cancer and healthy individuals and explore its clinical application value in the diagnosis of bladder cancer. METHODS: Urine samples were collected from 42 patients with bladder cancer and 42 healthy volunteers who visited Lu'an Hospital of Anhui Medical University and the Second Hospital of Tianjin Medical University from January 2020 to December 2022. The expression levels of lncRNA SNHG16 in urinary exosomes of the two groups were detected by RT‒qPCR, and their correlation with clinical pathological parameters of bladder cancer patients was analysed. An Receiver Operating Characteristic(ROC) curve was drawn to analyse the diagnostic value of urinary exosomal lncRNA SNHG16 for bladder cancer and compared with urinary cytology. RESULTS: The expression of urinary exosomal lncRNA SNHG16 in patients with bladder cancer was significantly higher (P < 0.05), and the expression level had no correlation with the age, sex, pathological T stage, pathological grade, or tumour size of bladder cancer patients (P > 0.05). The Area Under Curve(AUC) of urinary exosomal lncRNA SNHG16 in diagnosing bladder cancer was 0.791, which was superior to that of urinary cytology (AUC = 0.597). CONCLUSION: Urinary exosomal lncRNA SNHG16 with high expression can serve as a potential diagnostic biological marker for bladder cancer.

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BACKGROUND: Urine exosomal bkv-miR-B1-5p is associated with BK virus (BKV) nephropathy (BKVN); however, its posttransplantation changes and predictability for BKVN have not been determined in kidney transplant recipients (KTRs). METHODS: Urine exosomal bkv-miR-B1-5p and urine and plasma BKV DNA were measured at 2 weeks and 3, 6, and 12 months posttransplant in 83 KTRs stratified into biopsy-proven or presumptive BKVN, BKV viruria, and no evidence of BKV reactivation. Joint model, multivariable Cox model and receiver operating characteristic curve (ROC) were used to investigate the association of each assay with the following events: a composite of biopsy-proven or presumptive BKVN, and biopsy-proven BKVN. RESULTS: Urine exosomal bkv-miR-B1-5p and urine and plasma BKV DNA showed similar posttransplant time-course changes. Joint models incorporating serial values demonstrated significant associations of all assays with the events, and Cox analyses using single time point values at 2 weeks posttransplant showed that only urine exosomal bkv-miR-B1-5p was significantly associated with the events, although it did not outperform urine BKV DNA in ROC analyses. CONCLUSIONS: Urine exosomal bkv-miR-B1-5p was associated with BKVN as were urine and plasma BKV DNA loads on serial follow-up, and might have potential as a predictive marker for BKVN during the early posttransplant period. CLINICAL TRIALS REGISTRATION: Clinical Research Information Service (https://cris.nih.go.kr/cris/), KCT0001010.

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BACKGROUND: Approximately 70%-80% of kidney cancers are clear cell renal cell carcinomas (CCRCCs). Patient management is based on imaging (abdominal ultrasound and computerized tomography), surgical excision of the tumor, and pathological analysis. A tissue biopsy is therefore necessary to confirm the diagnosis and avoid unnecessary nephrectomy. For metastatic cancers, a tissue biopsy is essential for establishing the targeted therapy. This biopsy of tumor material is invasive and painful. Other techniques such as liquid biopsy would help reduce the need for tissue biopsy. The development of a simple biological test for diagnosis is essential. CA9 is a powerful marker for the diagnosis of CCRCC. Exosomes have become a major source of liquid biopsy because they carry tumor proteins, RNA, and lipids. Urine is the most convenient biological liquid for exosome sampling. OBJECTIVE: The aim of this study (PEP-C study) is mainly to determine whether it is possible to detect urinary exosomal CA9 for the molecular diagnosis of CCRCC. METHODS: This study will include 60 patients with CCRCC and 40 noncancer patients. Exosomes will be isolated from urine samples and exosomal CA9 will be detected by transmission electron microscopy, flow cytometry, and reverse transcription-quantitative polymerase chain reaction. RESULTS: This study is currently underway with funding support from the CHU Saint-Etienne of France. CONCLUSIONS: We expect to demonstrate that urinary tumor exosomes could be a novel liquid biopsy to diagnose CCRCC and to guide clinicians in treatment decision-making. TRIAL REGISTRATION: ClinicalTrials.gov NCT04053855; https://clinicaltrials.gov/ct2/show/NCT04053855. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/24423.

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BACKGROUND: Developing a noninvasive clinical test to accurately diagnose kidney allograft rejection is critical to improve allograft outcomes. Urinary exosomes, tiny vesicles released into the urine that carry parent cells' proteins and nucleic acids, reflect the biologic function of the parent cells within the kidney, including immune cells. Their stability in urine makes them a potentially powerful tool for liquid biopsy and a noninvasive diagnostic biomarker for kidney-transplant rejection. METHODS: Using 192 of 220 urine samples with matched biopsy samples from 175 patients who underwent a clinically indicated kidney-transplant biopsy, we isolated urinary exosomal mRNAs and developed rejection signatures on the basis of differential gene expression. We used crossvalidation to assess the performance of the signatures on multiple data subsets. RESULTS: An exosomal mRNA signature discriminated between biopsy samples from patients with all-cause rejection and those with no rejection, yielding an area under the curve (AUC) of 0.93 (95% CI, 0.87 to 0.98), which is significantly better than the current standard of care (increase in eGFR AUC of 0.57; 95% CI, 0.49 to 0.65). The exosome-based signature's negative predictive value was 93.3% and its positive predictive value was 86.2%. Using the same approach, we identified an additional gene signature that discriminated patients with T cell-mediated rejection from those with antibody-mediated rejection (with an AUC of 0.87; 95% CI, 0.76 to 0.97). This signature's negative predictive value was 90.6% and its positive predictive value was 77.8%. CONCLUSIONS: Our findings show that mRNA signatures derived from urinary exosomes represent a powerful and noninvasive tool to screen for kidney allograft rejection. This finding has the potential to assist clinicians in therapeutic decision making.

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A magnetic porous carbon-dependent platform is established to separate and determine N-glycans from urine exosomes of healthy people and patients with gastric cancer. The results of the comparison reveal that 6 N-glycans shared by the two groups are downregulated, most of which present core fucose or bisecting N-acetylglucosamine (GlcNAc) type. In addition, five shared N-glycans including two of sialic acid type are upregulated. These obvious differences indicate the close relationship between glycans and gastric cancer thus permitting early diagnosis. A magnetic porous carbon material (FeMPC) from MIL-101(Fe) was employed to separate and analyze N-glycans from urine exosomes of healthy people and patients with gastric cancer.

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Protein N-glycosylation plays crucial roles in many biological processes and has close association with the occurrence and development of various cancers. Therefore, it is necessary to analyze the abnormal changes of N-glycopeptides in complex biological samples for biomarker discovery. However, due to their low abundance and poor ionization, N-glycopeptides identification in complex samples by mass spectrometry (MS) is still a challenging task. In this work, a novel magnetic hydrophilic material was prepared by serial functionalization of ultra-thin two-dimensional molybdenum disulfide with Fe3O4 nanoparticles, gold nanowire and glutathione (MoS2-Fe3O4-Au/NWs-GSH) for efficient N-glycopeptides enrichment. The advantage of using the new nanocomposite is threefold. First, the introduction of magnetic Fe3O4 nanoparticles efficiently simplifies the enrichment process. Second, the gold nanowire modification enlarges the surface area of the nanocomposites to facilitate interaction with N-glycopeptides. Third, the employment of highly hydrophilic glutathione leads to specific HILIC-based retention of N-glycopeptides. Low femtomolar detection sensitivity and 1:1000 enrichment selectivity can be achieved using MoS2-Fe3O4-Au/NWs-GSH enrichment and bio-mass spectrometry analysis. Successful applications in human urine exosome and serum proteins were demonstrated by the enrichment and identification of 1250 and 489 N-glycopeptides, respectively. This remarkable data set of N-glycoproteome indicates the application potential of the novel nanocomposites for N-glycopeptides enrichment in complex biological samples and for related glycoproteome studies.

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BACKGROUND: An early sepsis-induced acute kidney injury (sepsis-AKI) biomarker is currently in needed. Urinary neutrophil gelatinase-associated lipocalin (uNGAL) is a candidate of sepsis-AKI biomarker but with different cut-point values. Urinary exosomal activating transcriptional factor 3 (uATF3) has been mentioned as an interesting biomarker. METHODS: We conducted experiments in mice and a prospective, multicenter study in patients as a proof of concept that urine exosome is an interesting biomarker. An early expression of ATF3 in kidney of CD-1 mice at 6 h after cecal ligation and puncture implied the possibility of uATF3 as an early sepsis-AKI biomarker. Increase serum creatinine (Scr) ≥0.3 mg/dL from the baseline was used as an AKI diagnosis and urine was analyzed for uATF3 and uNGAL. Patients with baseline Scr at admission ≥1.5 mg/dL were excluded. RESULTS: The analysis showed higher Scr, uNGAL and uATF3 in patients with sepsis-AKI in comparison with patients with sepsis-non-AKI and healthy volunteers. A fair correlation, r2 = 0.47, between uATF3 and uNGAL was showed in sepsis-AKI group with Scr ≥2 mg/dL. To see if uATF3 could be an early sepsis-AKI biomarker, urine sample was collected daily during the first week of the admission. In sepsis-AKI and sepsis-non-AKI groups, uNGAL were 367 ± 43 ng/mL and 183 ± 23 ng/mL, respectively; and uATF3 were 19 ± 4 ng/mL and 1.4 ± 0.8 ng/mL, respectively. With the mean value of uNGAL and uATF3 in sepsis AKI as a cut-off level, AUROC of uNGAL and uATF3 were 64% (95% CI 0.54 to 0.74) and 84% (95% CI 0.77 to 0.91), respectively. CONCLUSIONS: Urine exosome is an interesting source of urine biomarker and uATF3 is an interesting sepsis-AKI biomarker.

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PURPOSE: Using prostatic fluids rich in glycoproteins like prostate-specific antigen and prostatic acid phosphatase (PAP), the goal of this study was to identify the structural types and relative abundance of glycans associated with prostate cancer status for subsequent use in emerging MS-based glycopeptide analysis platforms. EXPERIMENTAL DESIGN: A series of pooled samples of expressed prostatic secretions (EPS) and exosomes reflecting different stages of prostate cancer disease were used for N-linked glycan profiling by three complementary methods, MALDI-TOF profiling, normal-phase HPLC separation, and triple quadropole MS analysis of PAP glycopeptides. RESULTS: Glycan profiling of N-linked glycans from different EPS fluids indicated a global decrease in larger branched tri- and tetra-antennary glycans. Differential exoglycosidase treatments indicated a substantial increase in bisecting N-acetylglucosamines correlated with disease severity. A triple quadrupole MS analysis of the N-linked glycopeptides sites from PAP in aggressive prostate cancer pools was done to cross-reference with the glycan profiling data. CONCLUSION AND CLINICAL RELEVANCE: Changes in glycosylation as detected in EPS fluids reflect the clinical status of prostate cancer. Defining these molecular signatures at the glycopeptide level in individual samples could improve current approaches of diagnosis and prognosis.

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