Potential Non-invasive Diagnostic Biomarker Research Articles

Association of rs6983267 at 8q24, HULC rs7763881 polymorphisms and serum lncRNAs CCAT2 and HULC with colorectal cancer in Egyptian patients

The impact of HULC rs7763881 on colorectal cancer (CRC) susceptibility is not yet known. Also, the biological function of the cancer-related rs6983267 remains unclear. We investigated the association of these SNPs with the risk of CRC and adenomatous polyps (AP), their correlation with CCAT2 and HULC expression, and the potential of serum CCAT2 and HULC as biomarkers for CRC. 120 CRC patients, 30 AP patients, and 96 healthy controls were included. Genotyping and serum lncRNAs were assayed by qPCR. Studied SNPs were not associated with AP susceptibility. rs6983267 GG was associated with increased CRC risk, whereas rs7763881 AC was protective. rs7763881 and rs6983267 CT haplotype was protective. Serum CCAT2 and HULC were upregulated in CRC and AP patients versus controls and discriminated these groups by ROC analysis. rs6983267 GG and rs7763881 AA patients demonstrated higher serum CCAT2 and HULC compared with GT/TT and AC, respectively. rs6983267 and serum HULC predicted CRC diagnosis among non-CRC groups (AP + controls) by multivariate analysis. Studied SNPs or serum long noncoding RNAs weren’t correlated with nodal or distant metastasis. In conclusion, rs6983267 and rs7763881 are potential genetic markers of CRC predisposition and correlate with serum CCAT2 and HULC, two novel potential non-invasive diagnostic biomarkers for CRC.

Bile acid profiles in neonatal intrahepatic cholestasis caused by citrin deficiency

BackgroundNeonatal intrahepatic cholestasis caused by citrin deficiency (NICCD) is characterized by conjugated hyperbilirubinemia and increased plasma bile acid concentrations. However, the underlying mechanisms remain unclear. We established a liquid chromatography tandem mass spectrometry (LC-MS/MS) method for simultaneously quantifying plasma bile acids and examined bile acid profiles in NICCD infants. MethodsWe measured 15 bile acids within 15min and found a wide linear range for individual bile acids. ResultsThe within-run and run-to-run CV of all bile acids was 1.2–10.9% and 3.1–10.8%, respectively, with a mean recovery of 90.5–112.6%. Compared to infants with citrullinemia without mutations in SLC25A13 (non-NICCD), NICCD infants showed increased plasma total bile acid concentrations (mean: 201 vs. 42μM, p<0.001), with a distinct bile acid profile characterized by increased conjugated primary bile acid concentrations. The calculated ratios, including primary/secondary bile acid (714 vs. 235, p<0.05) and conjugated/free bile acid (371 vs. 125, p<0.05) ratios, were higher in NICCD infants. The area under receiver operating characteristic curve for conjugated/free bile acid ratio to identify infants with NICCD was 0.871 (95% confidence interval, 0.713–1.0). ConclusionsTogether, our findings indicated plasma bile acid profile as a potential noninvasive diagnostic biomarker for NICCD.

Circulating miR-155, miR-145 and let-7c as diagnostic biomarkers of the coronary artery disease

Aim: Specific expression profiles of circulating microRNA (miRNAs) have been associated with several diseases such as cancer and cardiovascular injury, therefore miRNA have emerged as potential noninvasive diagnostic biomarkers of diseases. The aim of the present study was to investigate circulating miRNAs differentially regulated between patients with coronary artery disease (CAD) and healthy control subjects, and,determine their potential diagnostic value for CAD.

The overexpression of salivary cytokeratins as potential diagnostic biomarkers in head and neck squamous cell carcinomas.

BackgroundCytokeratin (CK) intermediate filaments are demonstrated to have enormous potential in regulating cellular motility and cancer progression. There are more than 20 divergent CKs that have been identified, of which CK 8, 17, 18 and 19 are reported to be elevated in the tumour biopsies of head and neck cancer squamous cell carcinoma (HNSCC) patients. However, CK expression profiles in the saliva of HNSCC patients have not been investigated. We aim to investigate the mRNA expression profiles of CKs in saliva collected from healthy controls, HPV-negative and -positive HNSCC patients.MethodsOral rinse samples were collected from 42 cancer-free healthy controls (age-matched) and patients who have been diagnosed with HPV-negative (n = 20) and -positive (n = 48) HNSCC.ResultsHere, we report that the mRNA expression profiles of CKs differed in saliva collected from healthy controls and HNSCC patients. The mRNA expression levels of CK 8 and 18 were significantly elevated in saliva collected from HPV-negative HNSCC patients; whilst, CK 17 and 19 were expressed at a higher mRNA level in saliva collected from HPV-positive HNSCC patients compared to healthy controls. Importantly, receiver operating characteristic (ROC) analysis showed salivary CK 8 and 18 to have superior sensitivity and specificity in discriminating the HPV-negative HNSCC patients from healthy controls (80% and 86%) as well as between HPV-negative and -positive HNSCC patients (75% and 81%).ConclusionIn summary, we have demonstrated that an aberrant expression of salivary CKs may serve as a potential non-invasive diagnostic biomarker in HNSCC.

Differential expression of circulating miR-21, miR-142-3p and miR-155 in renal transplant recipients with impaired graft function.

The discovery of circulating microRNAs (miRNAs), as potential noninvasive diagnostic biomarkers, has opened new avenues of research for identifying patients with chronic failure in renal transplantation. The present study aimed to investigate the expression levels of four immune-related miRNAs (miR-21, miR-31, miR-142-3p and miR-155) in plasma samples of renal recipients. The plasma expression levels of the miRNAs were evaluated by quantitative real-time PCR (qPCR) in 53 renal recipients with long-term stable allograft function, SGF (N=27), and with biopsy-proven interstitial fibrosis and tubular atrophy (IFTA) (N=26) and also healthy controls (N=15). The possible correlation between clinical parameters and the circulating miRNAs and the receiver-operating characteristic (ROC) analysis were performed. Our results showed that expression of miR-21 (p=0.023), miR-142-3p (p=0.048) and miR-155 (p=0.005) was significantly upregulated in plasma samples of recipients with IFTA in comparison with SGF and healthy control groups. Concentration of miR-21 (∆Ct value) in plasma was negatively correlated with creatinine (r=-0.432, p=0.028) and positively correlated with eGFR (r=0.423, p=0.031). The multivariate ROC curve analysis indicated that miR-21, miR-142-3p and miR-155 in plasma samples could discriminate almost most of the IFTA patients (area under curve=0.802, sensitivity=81%, specificity=92%). Our data suggested that altered expression of miR-21, miR-142-3p and miR-155 in plasma samples may be associated with renal dysfunction and can be used for graft monitoring.

Circulating miR-155, miR-145 and let-7c as diagnostic biomarkers of the coronary artery disease

Coronary artery disease (CAD) is the most prevalent cause of mortality and morbidity worldwide and the number of individuals at risk is increasing. To better manage cardiovascular diseases, improved tools for risk prediction including the identification of novel accurate biomarkers are needed. MicroRNA (miRNA) are essential post-transcriptional modulators of gene expression leading to mRNA suppression or translational repression. Specific expression profiles of circulating miRNA have emerged as potential noninvasive diagnostic biomarkers of diseases. The aim of this study was to identify the potential diagnostic value of circulating miRNA with CAD. Circulating miR-145, miR-155, miR-92a and let-7c were selected and validated by quantitative PCR in 69 patients with CAD and 30 control subjects from the cross-sectional study GENES. The expression of miR-145, miR-155 and let-7c showed significantly reduced expression in patients with CAD compared to controls. Multivariate logistic regression analysis revealed that low levels of circulating let-7c, miR-145 and miR-155 were associated with CAD. Receiver operating curves analysis showed that let-7c, miR-145 or miR-155 were powerful markers for detecting CAD. Furthermore, we demonstrated that the combination of the three circulating miRNA managed to deliver a specific signature for diagnosing CAD.

Evaluation of the Role of Circulating Long Non-Coding RNA H19 as a Promising Novel Biomarker in Plasma of Patients with Gastric Cancer.

H19 is one of the long non-coding RNAs (LncRNA) that is related to the progression of many diseases including cancers. This work was carried out to study the level of the long non-coding RNA; H19, in plasma of patients with gastric cancer (GC) and to assess its significance in their clinical management. Sixty-two participants were enrolled in the present study. The first group included 32 GC patients. The second group was formed of 30 age and sex matched healthy volunteers serving as a control group. Plasma samples were used to assess H19 gene expression using real-time quantitative PCR technique. H19 expression was up-regulated and closely related to TNM cancer stages in GC patients. Using Receiver Operating Characteristic (ROC) curve analysis, a cutoff level of 0.5 was set for H19 expression to diagnose GC cases achieving a sensitivity of 68.75%, specificity of 56.67%, positive predictive value (PPV) 62.86% and negative predictive value (NPV) 62.96% with an area under the curve (AUC) of 72.4%. Combined use of Carcinoembryonic Antigen (CEA) and H19 level in GC diagnosis was evaluated using ROC curve revealing improvement in performance with an area under the curve of 80.4%. Up-regulation of H19 is closely associated with gastric cancer displaying progressive up-regulation in advanced stages of the disease implementing its role as a potential non-invasive diagnostic biomarker in gastric cancer and as a novel tool in gastric cancer management with better performance achieved on using both CEA and H19 simultaneously.

Exosomal MicroRNAs Are Diagnostic Biomarkers and Can Mediate Cell–Cell Communication in Renal Cell Carcinoma

BackgroundApart from an invasive biopsy, currently no tools are available to confirm the diagnosis of clear cell renal cell carcinoma (ccRCC); this resulted in approximately 30% of patients being diagnosed with metastatic disease. ObjectiveTo determine whether urinary microRNAs (miRNAs) can serve as biomarkers to confirm the diagnosis of ccRCC. Design, setting, and participantsGlobal miRNA expression was assessed in 28 preoperative urine samples from patients with ccRCC and 18 healthy participants. The independent validation set consisted of 81 ccRCC patients, 24 patients with benign lesions, and 33 healthy participants. We extracted both cell-free and exosomal RNA for miRNA expression analysis using miRNA-specific polymerase chain reaction assays. We also investigated exosomal miRNA secretion in cell line models and performed exosome transfer between RCC and endothelial cell types. Outcome measurements and statistical analysisReceiver operating characteristic analysis was applied to identify the discrimination power of miRNAs. Results and limitationsOverall, miR-126-3p combined with miR-449a or with miR-34b-5p could significantly distinguish ccRCC patients from healthy participants (miR-126-3p–miR-449a: area under the curve [AUC]: 0.84; 95% confidence interval [CI], 0.7620–0.9151; p<0.001; miR-126-3p–miR-34b-5p: AUC: 0.79; 95% CI, 0.7013–0.8815; p<0.001). The combination of miR-126-3p and miR-34b-5p was also able to distinguish small renal masses (pT1a, ≤4cm) from healthy controls (AUC: 0.79; 95% CI, 0.6848–0.8980; p<0.001). Using miR-126-3p and miR-486-5p in combination, we were able to differentiate between benign lesions and ccRCC (AUC: 0.85; 95% CI, 0.7295–0.9615; p<0.01). The expression of a number of miRNAs returned to a level comparable with health after surgery. Kidney cancer cell lines were found to secrete exosomal miR-126-3p, miR-17-5p, miR-21-3p, and miR-25-3p, and these miRNAs were found to be internalized by other cell types. ConclusionsWe identified exosomal miRNAs as potential noninvasive diagnostic urinary biomarkers for ccRCC and provided evidence that miRNAs are secreted by the tumor and can function as a tool for intercellular communication. Patient summaryWe identified urinary microRNAs that can serve as diagnostic biomarkers for clear cell renal cell carcinoma.

Pilot Study of Serum MicroRNA-21 as a Diagnostic and Prognostic Biomarker in Egyptian Breast Cancer Patients.

MicroRNAs are small RNA molecules that bind to complementary sequences of target messenger RNAs and down-regulate their translation to protein or degrade them. MicroRNAs play critical roles in many different cellular processes. Hence, aberrant microRNA expression is common in a variety of disorders, including cancer. In this work, we quantified serum microRNA-21 (miR-21) expression levels in 30 breast cancer patients, 30 cancer-free individuals with risk factors for developing breast cancer, and another 30 controls without risk factors, in order to test the role of miR-21 as a possible diagnostic and prognostic biomarker in breast cancer. Our results indicated that miR-21 expression was elevated in asymptomatic high-risk individuals (2.98-fold) compared with healthy non-risk controls (p < 0.001), and was increased in almost all sera of cancer patients (12.72-fold) compared with healthy controls (p < 0.001). Higher levels of serum miR-21 were also correlated with tumors of higher grades, more nodal involvement, distal metastasis and advanced clinical stages (p < 0.01). Furthermore, over-expression levels declined towards normal after surgical tumor resection (p < 0.001). In conclusion, our findings demonstrate that serum miR-21 expression profile may serve as a potential non-invasive diagnostic and prognostic biomarker for breast cancer.

Circulating miR-765 and miR-149: potential noninvasive diagnostic biomarkers for geriatric coronary artery disease patients.

The purpose of this study was to evaluate the diagnostic value of circulating miR-765 and miR-149 as noninvasive early biomarkers for geriatric coronary artery disease (CAD) patients. A total of 69 angiographically documented CAD patients including 37 stable CAD (72.9 ± 4.2 years) and 32 unstable CAD (72.03 ± 4.3 years) and 20 healthy subjects (71.7 ± 5.2 years), matched for age, sex, smoking habit, hypertension, and diabetes, were enrolled in this study. Compared with healthy subjects, circulating miR-765 levels were increased by 2.9-fold in stable CAD and 5.8-fold in unstable CAD patients, respectively, while circulating miR-149 levels were downregulated by 3.5-fold in stable CAD and 4.2-fold in unstable CAD patients, respectively. Furthermore, plasma levels of miR-765 were found to be positively correlated with ages within control, stable, and unstable groups. The ROC curves of miR-765 and miR-149 represented significant diagnostic values with an area under curve (AUC) of 0.959, 0.972 and 0.938, 0.977 in stable CAD patients and unstable CAD patients as compared with healthy subjects, respectively. Plasma levels of miR-765 and miR-149 might be used as noninvasive biomarkers for the diagnosis of CAD in geriatric people.

Circulating MicroRNA in Digestive Tract Cancers

d t For many decades, cell-free nucleic acids have been known to be present in peripheral blood. Several tudies have identified tumor-specific and/or tumor-assoiated alterations in the circulating nucleic acids of paients with various cancers. In recent years, cell-free miroRNA (miRNA) have been stably detected in the plasma nd serum, like other molecules; their presence in the lood has attracted the attention of researchers due to heir potential use as valuable blood biomarkers.1 MiRNAs are short, noncoding RNAs that play important roles in various physiologic and developmental processes. The mature miRNAs are produced from long primary transcripts through 2 sequential cleavage steps. The long primary miRNA transcript is cleaved by the Drosha complex in the nucleus, generating intermediate precursor miRNA. Precursor miRNA is transported by exportin-5 from the nucleus into the cytoplasm, and then subjected to further cleavage by a Dicer RNAase III enzyme, generating a short double-strand miRNA. One strand (guided strand) of mature miRNA is then incorporated into the RNA-induced silencing complex and subsequently hybridize to the 3=-untranslated region of their target mRNAs to repress translation or degrade these mRNAs. Thus, a single miRNA can influence the expression of hundreds of genes and allow them to function in a coordinated manner. Therefore, miRNAs have been implicated as key molecules in all cellular processes. Numerous studies have shown that alterations in miRNA expression correlate with various diseases, including the development and progression of cancer, and some miRNAs can function as oncogenes or tumor suppressors. These findings have opened up a new and interesting field in the diagnosis of cancer and the treatments of cancer patients. Mitchell et al2 first demonstrated that circulating miRNAs had the potential to be new biomarkers in patients with solid cancers. In recent years, several papers have demonstrated that circulating miRNAs can also be detected in the peripheral blood of patients with digestive tract cancers. Although the origins and physiologic functions of cell-free miRNAs in the blood remain to be fully elucidated, a noninvasive assay for miRNAs should be developed to exploit these molecules as potential diagnostic and prognostic biomarkers. This assay undoubtedly contributes to an improvement in the clinical outcomes of cancer patients. In this article, we review the current state of biological and clinical research regarding circulating miRNAs of digestive tract cancer patients and discuss the future perspectives.

Increased Urinary Neutrophil Gelatinase Associated Lipocalin Levels in a Rat Model of Upper Urinary Tract Infection

Recurrent upper urinary tract infection is a common complication of vesicoureteral reflux that often leads to irreversible renal scarring. In our previous study of a rat model of renal bacterial infection we performed global gene expression profiling of the kidney during the onset of renal scarring. We have further investigated the product of an up-regulated gene product, NGAL, in this animal model to evaluate its potential usefulness as a biomarker of renal scarring. Renal NGAL mRNA and protein levels were examined by real-time polymerase chain reaction, Western blot and immunohistochemistry. Urinary NGAL levels were monitored by direct enzyme-linked immunosorbent assay. Rat renal NGAL mRNA and protein levels were found to be increased soon after bacterial injection. They then decreased rapidly but subsequently persisted at high levels until the 6-week time point after injection. On histological analysis we found that NGAL protein was overproduced in macrophages and renal tubular cells 2 weeks after injection. However, renal tubular cells continued to produce NGAL 6 weeks after injection, whereas this expression was lost in infiltrating cells. Rat urinary NGAL levels were also markedly increased at the early stages of infection and they persisted at high levels throughout the latter stages of the experiment. Urinary NGAL may be a potential noninvasive diagnostic biomarker of renal scarring.