Non-invasive Biomarkers For Early Diagnosis Research Articles

Abstract B020: DNA methylation and pancreatic cancer in select gene promoter regions: A repeated measures case-control study of US military service members

Abstract Introduction: Pancreatic cancer (PC) is a devastating disease, its etiology is not fully understood, and there is a paucity of non-invasive biomarkers for early diagnosis. While DNA methylation (DNAm) holds promise, associations between DNAm and PC risk are poorly understood, especially regarding temporal changes in associations. Therefore, we aimed to evaluate potential associations between DNAm in pre-diagnostic serum on 9 genes (BNIP3, CDKN1C, CLDN4, LCN2, p16, RASSF1, SFN, SPARC, and TFPI-2) and 2 repetitive elements (Alu and LINE-1) and PC risk in a US Department of Defense cohort. Methods : The study population included 82 PC cases, diagnosed 1991 - 2007, and 229 controls individually- matched on age (within 5 years), sex, race, and serum draw time period. Cases had up to 3 pre- diagnostic serum samples. Controls had 1 serum sample collected 0-10 years before their selection date (date of case diagnosis). DNA was extracted from serum and the percentage of cytosines at a given CpG site that were methylated was quantified via Pyrosequencing. For serum samples within a time window (<2, 2-<5, and 5-10 years pre-diagnosis), we computed odds ratios (ORs) and 95% confidence intervals (95% CIs) for the association of each CpG site with PC risk (66 sites in total across all genes and repetitive elements) with conditional logistic regression models. DNAm was evaluated as both a binary variable and a continuous variable. Results : In samples collected 5-10 years before diagnosis, those with no DNAm (0% DNAm) had increased odds of PC compared to those with DNAm (>0% DNAm) in the promoter region of BNIP3 (Chr10:131982293 OR, 95% CI: 5.63, 1.16 – 27.28) and those with lower DNAm (<median) had increased odds of PC compared to those with higher DNAm (≥median) in the promoter regions of CDKN1C (Chr11:2886398 OR, 95% CI: 4.07, 1.16 – 14.31; Chr11:2886388 OR, 95% CI: 3.51, 1.24 – 9.96) and RASSF1 (Chr3:50340841 OR, 95% CI: 3.98, 1.15 – 17.45). Similarly, in samples collected 2 - <5 years before diagnosis, lower DNAm (<median) was associated with increased odds of PC compared to those with higher DNAm (≥median) in the promoter region of TFPI-2 (Chr7:93890717 OR, 95% CI: 3.82, 1.20 – 12.15). In models with DNAm as a continuous measure and in samples collected <2 years before diagnosis, a 1% increase in DNAm in the promoter region of SFN was associated with increased odds of PC (Chr1:26863188 OR, 95% CI: 1.23, 1.01 – 1.51). There was no evidence of associations between DNAm on Alu, CLDN4, LCN2, LINE-1, p16, or SPARC and PC risk. Conclusions : Our findings of increased risk of PC associated with lower DNAm on BNIP3, CDKN1C and TFPI-2 and higher DNAm on SFN genes suggest there may be molecular patterns occurring prior to PC diagnosis that could be considered further for developing non-invasive biomarkers for early diagnosis of this aggressive carcinoma. The opinions and assertions expressed herein do not reflect the official policy or position of the Uniformed Services University, the Department of Defense, or the National Cancer Institute. Citation Format: Jordan McAdam, Ruth M Pfeiffer, Alexander P Keil, Ann Meyer, Anwar E Ahmed, Dechang Chen, Jennifer Rusiecki. DNA methylation and pancreatic cancer in select gene promoter regions: A repeated measures case-control study of US military service members [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr B020.

Insight into the Role of the miR-584 Family in Human Cancers.

Among the non-coding RNAs, the aberrant expression of microRNAs (miRNAs) is well described in the oncology field. It is clear that the altered expression of miRNAs is crucial for a variety of processes such as proliferation, apoptosis, motility, angiogenesis and metastasis insurgence. Considering these aspects, RNA-based therapies and the use of miRNAs as non-invasive biomarkers for early diagnosis are underlined as promising opportunities against cancer death. In the era of precision medicine, significant progress in next-generation sequencing (NGS) techniques has broadened knowledge regarding the miRNAs expression profile in cancer tissues and in the blood of cancer patients. In this scenario, pre-clinical and clinical studies suggested that the members of the miR-584 family, i.e., miR-584-5p and -3p, are prominent players in cancer development and progression. Under some conditions, these miRNAs are under-expressed in cancer tissues acting as tumor suppressors, while in other conditions, they are overexpressed, acting as oncogenes increasing the aggressive behavior of cancer cells. The aim of this review is to provide a comprehensive and up-to-date overview on the expression, upstream genes, molecular targets and signaling pathways influenced by the miR-584 family (i.e., miR-584-3p and -5p) in various human solid and hematological cancers. To achieve this goal, 64 articles on this topic are discussed. Among these articles, 55 are focused on miR-584-5p, and it is outlined how this miRNA could be used in future applications as a potential new therapeutic strategy and diagnostic tool.

The Sera miRNA Pattern in Patients Inflammatory Bowel Disease

Inflammatory bowel disease comprising Crohn's disease and ulcerative colitis presents with periods of flares and remission. Many reports have identified different dysregulated miRNAs in patients with IBD. Finding new biomarkers in IBD patients can help to launch a novel non-invasive approach for diagnosis and prognosis for patients with UC and CD. This study aimed to evaluate the plasma expression pattern of the miRNAs panel in IBD patients compared to healthy individuals. 73 plasma samples were included; 58 patients with IBD (33 individuals in flare and 25 in remission phase) and 15 healthy controls were enrolled in the study. The miRNA expression was measured by qRT-PCR using miScript SYBR Green PCR Kit (QIAGEN). Our results showed the expression level of miR-16-5P was significantly increased in the active phase compared to the inactive phase (P=0.0138) and in CD patients compared to UC patients (P=0.0216). There was a significant difference in the expression of miR-29a in Crohn's patients compared to healthy subjects (P=0.04). Measuring the expression of mir-106a; a significant increase was observed compared to healthy individuals (P=0.03) and patients with CD (P=0.0143) in proportion of UC patients’ group. The miR-126 expression significantly increased in patients with active disease compared to patients in the inactive phase (P=0.0413) and healthy controls (P=0.0492). This study showed evidence for differential expression levels of plasma panel of miR-16, miR-29a, miR-106a, and miR-126 in IBD patients compare to healthy individuals. We illustrate that miRNAs in plasma correlate with disease activity and can be used as a practical and non-invasive biomarker for early diagnosis and monitoring of the treatment protocol.

Intestinal metabolomics in premature infants with late-onset sepsis

We aimed to investigate the characteristics of intestinal metabolomics and non-invasive biomarkers for early diagnosis of late-onset sepsis (LOS) by analyzing gut metabolites in preterm infants with LOS. We collected stool samples from septic and healthy preterm infants for analysis by liquid chromatography–mass spectrometry. 123 different metabolites were identified and 13 pathways were mainly involved. Glycine, serine, and threonine metabolism; glyoxylate and dicarboxylic acid metabolism; glutathione metabolism; primary bile acid biosynthesis; steroid synthesis; pentose and glucuronic acid interconversion may be involved in the pathogenesis of LOS in preterm infants. The significant changes of N-Methyldopamine, cellulose, glycine, gamma-Glutamyltryptophan, N-Ribosylnicotinamide and 1alpha, 25-dihydroxycholecalciferol showed specific diagnostic values and as non-invasive biomarkers for LOS.

A new methodology to reveal potential nucleic acid modifications associated with the risk of endometrial cancer through dispersive solid-phase extraction coupled with UHPLC-QE-Orbitrap-MS/MS and HPLC-UV.

Nucleic acid modifications have attracted increasing attention in recent years since they have been found to be related to a number of diseases including cancer. Previous studies have shown that the early development of endometrial cancer (EC) is often accompanied by changes in methylation levels of related genes, and the expression of related proteins that regulate reactive oxygen species (ROS) shows significant differences in EC cells and tissues. However, it has not been reported whether nucleic acid modifications related to methylation or ROS can serve as biomarkers for EC. Accurate quantification of these nucleic acid modifications still has challenges because their amounts in urine are very low and the interferences in urine are complicated. In this study, a novel dispersive solid-phase extraction (DSPE) method based on chitosan-carbon nanotube-Al2O3 (CS-CNT-Al2O3) has been established for the analysis of 5-hydroxymethyluracil (5mU), 5-methyl-2'-deoxycytidine (5-mdC), 5-hydroxymethyl-2'-deoxycytidine (5-hmdC), 5-formyl-2'-deoxycytidine (5-fdC), and 8-hydroxy-2'-deoxyguanosine (8-OHdG) in EC patient urine samples coupled with UHPLC-QE-Orbitrap-MS/MS and HPLC-UV. Firstly, the synthesis of the CS-CNT-Al2O3 nanocomposite was conducted by a sono-coprecipitation method and was characterized by scanning electron microscope (SEM), energy dispersive spectrometer (EDS), and Fourier transform infrared (FTIR). Under the optimal extraction conditions of DSPE, we successfully quantified 5mU, 5-mdC, 5-hmdC, 5-fdC, and 8-OHdG in urine samples from 37 EC patients and 39 healthy controls. The results showed that there were significant differences in the levels of 5-mdC, 5-hmdC, 5-fdC, and 8-OHdG in EC patients compared to the healthy control group. The receiver operator characteristic (ROC) curve analysis was carried out to evaluate the potential of 5-mdC, 5-hmdC, 5-fdC, and 8-OHdG to distinguish EC patients from healthy volunteers. The area under the curve (AUC) for 5-mdC, 5-hmdC, 5-fdC, and 8-OHdG was 0.7412, 0.667, 0.8438, and 0.7981, respectively. It indicated that 5-mdC, 5-hmdC, 5-fdC, and 8-OHdG had certain potential in distinguishing between EC patients and healthy volunteers and they could act as potential non-invasive biomarkers for early diagnosis of EC. Moreover, the present study would stimulate investigations of the effects of nucleic acid modifications on the initiation and progression of EC.

Metabolomic machine learning predictor for diagnosis and prognosis of gastric cancer

Gastric cancer (GC) represents a significant burden of cancer-related mortality worldwide, underscoring an urgent need for the development of early detection strategies and precise postoperative interventions. However, the identification of non-invasive biomarkers for early diagnosis and patient risk stratification remains underexplored. Here, we conduct a targeted metabolomics analysis of 702 plasma samples from multi-center participants to elucidate the GC metabolic reprogramming. Our machine learning analysis reveals a 10-metabolite GC diagnostic model, which is validated in an external test set with a sensitivity of 0.905, outperforming conventional methods leveraging cancer protein markers (sensitivity < 0.40). Additionally, our machine learning-derived prognostic model demonstrates superior performance to traditional models utilizing clinical parameters and effectively stratifies patients into different risk groups to guide precision interventions. Collectively, our findings reveal the metabolic landscape of GC and identify two distinct biomarker panels that enable early detection and prognosis prediction respectively, thus facilitating precision medicine in GC.

Non-invasive biomarkers for early diagnosis of pancreatic cancer risk: metabolite genomewide association study based on the KCPS-II cohort

BackgroundPancreatic cancer is a lethal disease with a high mortality rate. The difficulty of early diagnosis is one of its primary causes. Therefore, we aimed to discover non-invasive biomarkers that facilitate the early diagnosis of pancreatic cancer risk.MethodsThe study subjects were randomly selected from the Korean Cancer Prevention Study-II and matched by age, sex, and blood collection point [pancreatic cancer incidence (n = 128) vs. control (n = 256)]. The baseline serum samples were analyzed by non-targeted metabolomics, and XGBoost was used to select significant metabolites related to pancreatic cancer incidence. Genomewide association study for the selected metabolites discovered valuable single nucleotide polymorphisms (SNPs). Moderation and mediation analysis were conducted to explore the variables related to pancreatic cancer risk.ResultsEleven discriminant metabolites were selected by applying a cut-off of 4.0 in XGBoost. Five SNP presented significance in metabolite-GWAS (p ≤ 5 × 10–6) and logistic regression analysis. Among them, the pair metabolite of rs2370981, rs55870181, and rs72805402 displayed a different network pattern with clinical/biochemical indicators on comparison with allelic carrier and non-carrier. In addition, we demonstrated the indirect effect of rs59519100 on pancreatic cancer risk mediated by γ-glutamyl tyrosine, which affects the smoking status. The predictive ability for pancreatic cancer on the model using five SNPs and four pair metabolites with the conventional risk factors was the highest (AUC: 0.738 [0.661–0.815]).ConclusionsSignatures involving metabolites and SNPs discovered in the present research may be closely associated with the pathogenesis of pancreatic cancer and for use as predictive biomarkers allowing early pancreatic cancer diagnosis and therapy.

Retinal Changes in Parkinson's Disease: A Non-invasive Biomarker for Early Diagnosis.

Parkinson's disease (PD) is caused due to degeneration of dopaminergic neurons in the substantia nigra pars compacta (SNpc) which leads to the depletion of dopamine in the body. The lack of dopamine is mainly due to aggregation of misfolded α-synuclein which causes motor impairment in PD. Dopamine is also required for normal retinal function and the light-dark vision cycle. Misfolded α-synuclein present in inner retinal layers causes vision-associated problems in PD patients. Hence, individuals with PD also experience structural and functional changes in the retina. Mutation in LRRK2, PARK2, PARK7, PINK1, or SNCA genes and mitochondria dysfunction also play a role in the pathophysiology of PD. In this review, we discussed the different etiologies which lead to PD and future prospects of employing non-invasive techniques and retinal changes to diagnose the onset of PD earlier.

Targeted demethylation of the CDO1 promoter based on CRISPR system inhibits the malignant potential of breast cancer cells.

Cysteine dioxygenase 1 (CDO1) is frequently methylated, and its expression is decreased in many human cancers including breast cancer (BC). However, the functional and mechanistic aspects of CDO1 inactivation in BC are poorly understood, and the diagnostic significance of serum CDO1 methylation remains unclear. We performed bioinformatics analysis of publicly available databases and employed MassARRAY EpiTYPER methylation sequencing technology to identify differentially methylated sites in the CDO1 promoter of BC tissues compared to normal adjacent tissues (NATs). Subsequently, we developed a MethyLight assay using specific primers and probes for these CpG sites to detect the percentage of methylated reference (PMR) of the CDO1 promoter. Furthermore, both LentiCRISPR/dCas9-Tet1CD-based CDO1-targeted demethylation system and CDO1 overexpression strategy were utilized to detect the function and underlying mechanism of CDO1 in BC. Finally, the early diagnostic value of CDO1 as a methylation biomarker in BC serum was evaluated. CDO1 promoter was hypermethylated in BC tissues, which was related to poor prognosis (p<.05). The CRISPR/dCas9-based targeted demethylation system significantly reduced the PMR of CDO1 promotor and increased CDO1 expression in BC cells. Consequently, this leads to suppression of cell proliferation, migration and invasion. Additionally, we found that CDO1 exerted a tumour suppressor effect by inhibiting the cell cycle, promoting cell apoptosis and ferroptosis. Furthermore, we employed the MethyLight to detect CDO1 PMR in BC serum, and we discovered that serum CDO1 methylation was an effective non-invasive biomarker for early diagnosis of BC. CDO1 is hypermethylated and acts as a tumour suppressor gene in BC. Epigenetic editing of abnormal CDO1 methylation could have a crucial role in the clinical treatment and prognosis of BC. Additionally, serum CDO1 methylation holds promise as a valuable biomarker for the early diagnosis and management of BC.

Immobilization-free dual-aptamer-based photoelectrochemical platform for ultrasensitive exosome assay

Exosomes, involved in cancer-specific biological processes, are promising noninvasive biomarkers for early diagnosis of cancer. Herein, an immobilization-free dual-aptamer-based photoelectrochemical (PEC) biosensor was proposed for the enrichment and quantification of cancer exosome based on photoactive bismuch oxyiodide/gold/cadmium sulfide (BiOI/Au/CdS) composites, nucleic acid-based recognition and signal amplification. In this biosensor, the recognition of exosome by two aptamers would trigger the deoxyribonucleotidyl transferase (TdT) enzyme-aided polymerization, leading to the enrichment of alkaline phosphatase (ALP) on Fe3O4 surface. After magnetic separation, ALP could catalyze the generation of ascorbic acid (AA) as electron donor and initiate the following redox cycle reaction for further signal amplification. Furthermore, all the above processes were performed in solution, the recognition and signal amplification efficiency would be superior than the heterogeneous strategy owing to the avoidance of steric hindrance effect. As a result, the proposed PEC biosensor was capable of enriching and detecting of cancer exosomes with high sensitivity and selectivity. The linear range of the biosensor was from 1.0 × 102 particles·μL−1 to 1.0 × 106 particles·μL−1 and the detection limit was estimated to be 21 particles·μL−1. Therefore, the proposed PEC biosensor holds great promise in quantifying tumor exosome for nondestructive early clinical cancer diagnosis and various other bioassay applications.

Transfer RNA-derived fragments as novel biomarkers of the onset and progression of gastric cancer.

Gastric cancer (GC) is a particularly malignant disease; thus, early diagnosis and treatment are especially important. Transfer RNA-derived small RNAs (tsRNAs) have been implicated in the onset and progression of various cancers. Therefore, the aim of this study was to explore the role of tRF-18-79MP9P04 (previously named tRF-5026a) in the onset and progression of GC. Expression levels of tRF-18-79MP9P04 were quantified in gastric mucosa specimens of healthy controls and plasma samples of patients with different stages of GC. The results showed that plasma levels of tRF-18-79MP9P04 were significantly decreased in the early and advanced stages of GC. The results of the nucleocytoplasmic separation assay found that tRF-18-79MP9P04 was localized in the nuclei of GC cells. High-throughput transcriptome sequencing identified genes regulated by tRF-18-79MP9P04 in GC cells, and the function of tRF-18-79MP9P04 was predicted by bioinformatics. Collectively, the findings of this study suggest that tRF-18-79MP9P04 would be useful as non-invasive biomarker for early diagnosis of GC and is related to cornification, the type I interferon signaling pathway, RNA polymerase II activities, and DNA binding.

Hypomethylation of Tumor necrosis factor-like cytokine 1A(TL1A) and its decoy receptor 3 expressive level increase has diagnostic value in HBV-associated cirrhosis

For patients with cirrhosis, early diagnosis is the key to delaying the development of liver fibrosis and improving prognosis. This study aimed to investigate the clinical significance of TL1A, which is a susceptibility gene for hepatic fibrosis, and DR3 in the development of cirrhosis and fibrosis. We analyzed the expression of TL1A, DR3, and other inflammatory cytokines associated with liver fibrosis in serum and PBMCs in 200 patients.TL1A methylation level was lower in patients with HBV-associated LC than in the other groups. In addition, the mRNA level and serum of TL1A and DR3 expression levels were found to increase in the LC. Hypomethylation of the TL1A promoter is present in HBV-associated LC, and TL1A and DR3 are highly expressed in HBV-associated cirrhosis. These results indicate that TL1A and DR3 may play an important role in the pathogenesis of LC and TL1A methylation levels may serve as a noninvasive biomarker for early diagnosis and progression of LC.

Diagnostic accuracy of circular RNA for diabetes Mellitus: a systematic review and diagnostic Meta-analysis

BackgroundThis study aimed to investigate the pooled diagnostic ability of circular RNA (circRNA) molecules for diabetes mellitus.MethodsWe searched PubMed, Scopus, and Web of Science for relevant studies. A total of 2070 participants, including 775 diabetic patients and 1295 healthy individuals, from five studies were included in this meta-analysis. True positive, true negative, false positive, and false negative data were extracted to calculate pooled sensitivity, specificity, positive and negative likelihood ratios, diagnostic odds ratio, and area under the receiver operating characteristics curve. The Deeks’ funnel plot was applied for publication bias assessment, Cochran’s Q test and I2 index were applied for inter-study heterogeneity assessment. Besides, a subgroup analysis was performed for determining the source of heterogeneity between studies. P value < 0.05 was considered significance. All analysis were done by STATA version 14.ResultsCircRNA presented a sensitivity of 76% (95% confidence interval [95%CI]: 66-84%), specificity of 77% (95%CI: 58-89%), positive LR of 3.25 (95%CI: 1.69–6.23), negative LR of 0.31 (95%CI: 0.21–0.46), DOR of 10.41 (95%CI: 4.26–25.41), and AUC of 0.82 (95%CI: 0.79–0.85) for diabetes mellitus detection. More specifically, hsa_circ_0054633 showed a sensitivity of 67% (95%CI: 53-81%) and a specificity of 82% (95%CI: 63-100%).ConclusionCircRNAs show highly accurate diagnostic capability for type 2 diabetes mellitus and gestational diabetes mellitus. High sensitivity of circRNAs introduces them as potential noninvasive biomarkers for early diagnosis of diabetes mellitus and their high specificity introduces them as potential therapeutic targets by regulation of their expression.

Biomarkers for Early Detection, Prognosis, and Therapeutics of Esophageal Cancers.

Esophageal cancer (EC) is the deadliest cancer worldwide, with a 92% annual mortality rate per incidence. Esophageal squamous cell carcinoma (ESCC) and esophageal adenocarcinoma (EAC) are the two major types of ECs, with EAC having one of the worst prognoses in oncology. Limited screening techniques and a lack of molecular analysis of diseased tissues have led to late-stage presentation and very low survival durations. The five-year survival rate of EC is less than 20%. Thus, early diagnosis of EC may prolong survival and improve clinical outcomes. Cellular and molecular biomarkers are used for diagnosis. At present, esophageal biopsy during upper endoscopy and histopathological analysis is the standard screening modality for both ESCC and EAC. However, this is an invasive method that fails to yield a molecular profile of the diseased compartment. To decrease the invasiveness of the procedures for diagnosis, researchers are proposing non-invasive biomarkers for early diagnosis and point-of-care screening options. Liquid biopsy involves the collection of body fluids (blood, urine, and saliva) non-invasively or with minimal invasiveness. In this review, we have critically discussed various biomarkers and specimen retrieval techniques for ESCC and EAC.

The Emerging Role of Circular RNA Homeodomain Interacting Protein Kinase 3 and Circular RNA 0046367 through Wnt/Beta-Catenin Pathway on the Pathogenesis of Nonalcoholic Steatohepatitis in Egyptian Patients.

Non-alcoholic fatty liver disease is a major problem worldwide that needs non-invasive biomarkers for early diagnosis and treatment response assessment. We aimed to assess the correlation between circRNA-HIPK3 and miRNA-29a expression and its role as miRNA-29a sponge, as well as the correlation between circRNA-0046367 and miRNA-34a expression and its role as miRNA-34a sponge and their effect on regulation of the Wnt/β catenin pathway, which may provide a new target for treatment of non-alcoholic steatohepatitis. the research was performed on 110 participants: group (I): fifty-five healthy donors served as controls and group (II): fifty-five patients with fatty liver pattern on abdominal ultrasound. Lipid profile and liver functions were assessed. RT-PCR was performed to assess the RNAs: circRNA-HIPK3, circRNA-0046367, miRNA-29a, miRNA-34a and Wnt mRNA gene expression. ELISA was performed to determine β-catenin protein levels. miRNA-34a and circRNA-HIPK3 expression were significantly greater, while miRNA-29a and circRNA-0046367 expression were significantly less, in patients than in controls. Wnt/β-catenin regulated by miRNA-29a and miRNA-34a showed a significant decrease that leads to its abnormal effect on lipid metabolism. our results imply that miRNA-29a can be investigated as a target for circRNA-HIPK3, while miRNA-34a can be investigated as a target for circRNA-0046367, and that circRNA-HIPK3 and circRNA-0046367 may have emerging roles that can affect the pathogenesis of nonalcoholic steatohepatitis through the Wnt/β-catenin pathway and thus be used as therapeutic targets for the disease.

Identification of PDCD1 as a potential biomarker in acute rejection after kidney transplantation via comprehensive bioinformatic analysis.

Acute rejection is a determinant of prognosis following kidney transplantation. It is essential to search for novel noninvasive biomarkers for early diagnosis and prompt treatment. Gene microarray data was downloaded from the Gene Expression Omnibus (GEO) expression profile database and the intersected differentially expressed genes (DEGs) was calculated. We conducted the DEGs with Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis. Distribution of immune cell infiltration was calculated by CIBERSORT. A hub gene marker was identified by intersecting the rejection-related genes from WGCNA and a selected KEGG pathway-T cell receptor signaling pathway (hsa04660), and building a protein-protein interaction network using the STRING database and Cytoscape software. We performed flow-cytometry analysis to validate the hub gene. A total of 1450 integrated DEGs were obtained from five datasets (GSE1563, GSE174020, GSE98320, GSE36059, GSE25902). The GO, KEGG and immune infiltration analysis results showed that AR was mainly associated with T cell activation and various T-cell related pathways. Other immune cells, such as B cells, Macrophage and Dendritic cells were also associated with the progress. After utilizing the WGCNA and PPI network, PDCD1 was identified as the hub gene. The flow-cytometry analysis demonstrated that both in CD4+ and CD8+ T cells, PD1+CD57-, an exhausted T cell phenotype, were downregulated in the acute rejection whole blood samples. Our study illustrated that PDCD1 may be a candidate diagnostic biomarker for acute kidney transplant rejection via integrative bioinformatic analysis.

Photoelectrochemical biosensor based on FTO modified with BiVO4 film and gold nanoparticles for detection of miRNA-25 biomarker and single-base mismatch

MicroRNAs (miRNAs) are of great important as non-invasive biomarkers for early diagnosis of cancer. Based on this, a photoelectrochemical (PEC) biosensor was developed to measure the concentration of microRNA-25 (miR-25), as a lung cancer biomarker. To fabricate the PEC biosensor, a suitable amino modified probe was immobilized on the surface of a florin tin oxide (FTO) electrode modified with bismuth vanadate (BiVO4) film and gold nanoparticles (AuNPs). The fabrication steps of the PEC biosensor were monitored by electrochemical impedance spectroscopy (EIS) and PEC measurements. The results indicate that the combination of BiVO4 film and AuNPs enhances the separation of photogenerated electrons and increase the photocurrent signal. Upon hybridization of the probe immobilized on the biosensor with miR-25, the photocurrent is significantly decreased. This current decrease is attributed to the fact that the hybridization processes blocks the electrode surface by creating a steric hindrance and by decrease the diffusion of AA to the electrode surface, it reduces the AA oxidation and subsequently decreases the capture photogenerated holes. Therefore, the amount of photocurrent decrease is dependent on the miR-25 concentration, and this PEC biosensor can be used to measure miR-25. The developed PEC biosensor shows good sensitivity for the determination of miR-25 in the linear concentration range of 1.0 × 10−14 M to 1.0 × 10−6 M with a detection limit of 3.4 fM, under optimal conditions. Using this biosensor, miR-25 can be detected from a non-complementary and one base mismatch sequence. It is also possible to measure miR-25 in human serum samples with satisfactory results using the PEC biosensor.

Early Diagnosis of Coronary Artery Disease by Inflammatory Biomarkers of Atherosclerosis in Patients with Angina.

Early diagnosis of coronary artery disease (CAD) in patients with chest pain is a challenge. Currently diagnosis of CAD is confirmed by coronary angiography, which is invasive and not easily available in developing countries. Therefore, it is imperative to establish noninvasive biomarker for early diagnosis of CAD in patients with angina and determine the diagnostic accuracy of inflammatory biomarkers of atherosclerosis in comparison to angiography and correlate with severity of CAD in patients with angina. Diagnostic accuracy study was carried out in tertiary care hospitals, Rawalpindi, Pakistan. Total of 58 patients aged 55.24 ± 11.61 years, with chest pain and troponin-I -ve, having coronary artery stenosis ≥50% were included as cases of CAD and 55 subjects having stenosis <50% were included as controls. Nuclear factor kappa-B (NF-κB), tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), and high-sensitivity C-reactive protein (hs-CRP) were analyzed on immunoassay analyzers. The receiver operating characteristic curve analysis revealed significant (P < 0.05) high area under curve (95% confidence interval) with sensitivity and specificity of NF-κB 0.76 (0.65-0.85), 73% and 65%; TNF-α 0.72 (0.61-0.81), 71% and 69%; IL-6 0.62 (0.52-0.71), 64% and 53% and hs-CRP 0.62 (0.52-0.71), and 53% and 56% in CAD patients compared to controls. There was significant positive correlation between NF-κB (r = 0.44), TNF-α (r = 0.37), IL-6 (r = 0.23), and hs-CRP (r = 0.23) with severity of CAD by Gensini score. The inflammatory biomarkers, especially NF-κB and TNF-α, have highest diagnostic accuracy and indicate severity of atherosclerosis in patients with angina. These markers may be used as noninvasive biomarkers to exclude healthy individuals before undergoing angiography.

Exosomal Non-Coding RNAs: New Insights into the Biology of Hepatocellular Carcinoma.

Exosomes, extracellular vesicles with a diameter of 40 to 160 nm, are among the smallest extracellular vesicles released by cells. They deliver different cargoes, including proteins, DNAs, and RNAs, and facilitate communication between cells to coordinate a variety of physiological and pathological functions. Hepatocellular carcinoma (HCC) is the sixth common malignant tumor and the fourth leading cause of cancer-related death worldwide. Its molecular mechanism remains largely unknown, and there is a lack of reliable and noninvasive biomarkers for early diagnosis and prognosis prediction. Mounting evidence has shown that exosomes carry a variety of ncRNAs, such as long non-coding RNAs (lncRNAs), microRNAs (miRNAs), and circular RNAs (circRNAs), which play critical roles in the occurrence and progression of HCC. In this review, we summarize the recent findings of exosomal miRNAs, lncRNAs, and circRNAs in HCC from their impact on the development of HCC to their potential applications in the diagnosis and treatment of HCC.

Possible relation between expression of circulating microRNA and plasma cytokine levels in cases of pulmonary tuberculosis.

Tuberculosis (TB) is a life-threatening infection and early diagnosis is critical for treatment and prevention of transmission. There is evidence of correlation between miRNA expression and cytokine regulation during TB infection. The aim of this study was to determine the relationship between expression levels of miRNAs in plasma and cytokine levels as a potential biomarker for genetic predisposition and/or early diagnosis of TB infection. The expression levels of 86 miRNAs were examined in plasma samples of 44 TB patients and 44 healthy controls by qRT-PCR using BioMarkTM 96.96 Dynamic Array (Fluidigm Corporation, South San Francisco, CA, USA) system. The levels of plasma TNF-α, IFN-γ, IL-1β, IL-4, IL-6, IL-8, IL-10, and IL-12/P40 were examined with ELISA. We identified dysregulation of 18 miRNAs which included upregulation of miR-1, miR-7-5p, miR-9-5p, miR-10a-5p, miR-10b-5p, miR-100-5p, miR-106b-5p, miR-128-3p, miR-133a-3p, miR-143-3p, miR-193a-5p, miR-200b-3p, miR-205-5p, miR-210-3p, and miR-296-5p, and downregulation of miR-15b-5p, miR-16-5p, and miR-25-3p in plasma samples of patients with pulmonary TB (p < 0.05). A significant correlation between the expression levels of miR-1, miR-7-5p, miR-9-5p, miR-10a-5p, miR-10b-5p, miR-15b-5p, miR-100-5p, miR-143-3p, miR-193a-5p, miR-200b-3p, miR-210-3p and cytokine levels of TNF-α, IFN-γ, IL-1β, IL-8 and IL-10 was identified (p < 0.05). We demonstrated that altered expression levels of plasma miRNAs consistent with immunological response have the potential to serve as non-invasive biomarkers for early diagnosis of pulmonary TB. Additional investigations with larger sample sizes will be required to confirm our findings and to determine if miRNAs can be possible targets for TB management strategies.