Biomonitoring of Multiple Mycotoxins in Urine by GC-MS/MS: A Pilot Study on Patients with Esophageal Cancer in Golestan Province, Northeastern Iran.

A pilot study to investigate the occurrence of 10 mycotoxins (deoxynivalenol, DON; 3-acetyldeoxynivalenol, 3-ADON; 15-acetyldeoxynivalenol, 15-ADON; fusarenon-X, FUS-X; diacetoxyscirpenol, DAS; nivalenol, NIV; neosolaniol, NEO; zearalenone, ZON; zearalanone, ZAN; T-2 toxin, T-2; and HT-2 toxin, HT-2) in esophageal cancer patients was performed with the urinary biomarkers approach in Golestan, Iran. Urine multimycotoxin analysis was performed by dispersive liquid-liquid microextraction and gas chromatography-tandem mass spectrometry (GC-MS/MS) analysis, and values were normalized with urinary creatinine (µg/g). Four mycotoxins, namely NEO (40%), HT-2 (17.6%), DON (10%), and HT-2 (5.8%), were detected in the analyzed urine samples. DON was only detected in the control group (5.09 µg/g creatinine), while T-2 (44.70 µg/g creatinine) was only present in the esophageal cancer group. NEO and HT-2 were quantified in both control and case groups, showing average of positive samples of 9.09 and 10.45 µg/g creatinine for NEO and 16.81 and 29.09 µg/g creatinine for HT-2, respectively. Mycotoxin co-occurrence was observed in three samples as binary (NEO/HT-2 and T-2/HT-2) and ternary (DON/NEO/HT-2) combinations, reaching total concentrations of 44.58, 79.13, and 30.04 µg/g creatinine, respectively. Further investigations are needed to explore a causal association between mycotoxin contamination and esophageal cancer. For this pilot study in Golestan, the low sample size was a very limiting factor.

Urinary microRNA biomarkers for detecting the presence of esophageal cancer.

Esophageal cancer (EC) including esophageal squamous cell carcinoma (ESCC) and adenocarcinoma (EAC) generally exhibits poor prognosis; hence, a noninvasive biomarker enabling early detection is necessary. Age- and sex-matched 150 healthy controls (HCs) and 43 patients with ESCC were randomly divided into two groups: 9 individuals in the discovery cohort for microarray analysis and 184 individuals in the training/test cohort with cross-validation for qRT-PCR analysis. Using 152 urine samples (144 HCs and 8 EACs), we validated the urinary miRNA biomarkers for EAC diagnosis. Among eight miRNAs selected in the discovery cohort, urinary levels of five miRNAs (miR-1273f, miR-619-5p, miR-150-3p, miR-4327, and miR-3135b) were significantly higher in the ESCC group than in the HC group, in the training/test cohort. Consistently, these five urinary miRNAs were significantly different between HC and ESCC in both training and test sets. Especially, urinary miR-1273f and miR-619-5p showed excellent values of area under the curve (AUC) ≥ 0.80 for diagnosing stage I ESCC. Similarly, the EAC group had significantly higher urinary levels of these five miRNAs than the HC group, with AUC values of approximately 0.80. The present study established novel urinary miRNA biomarkers that can early detect ESCC and EAC.

Renal metastases from esophageal cancer and retroperitoneal lymphoma detected via chromosome duplications identified by fluorescence in situ hybridization in urine exfoliated cells: First 2 case reports.

RATIONALE: Renal-occupying lesions positive for urine fluorescence in situ hybridization (FISH) are usually considered urothelial carcinomas. Here, we describe 2 cases of renal metastases with chromosome duplications in urine exfoliated cells. PATIENT SYMPTOMS: Patient 1, a 56-year-old male with a history of esophageal cancer, was admitted to our hospital on May 2017 after presenting with right back pain with microscopic hematuria for 1 month. Magnetic resonance imaging (MRI) showed right renal space-occupying lesions (5.4 cm × 4.6 cm) and multiple enlarged lymph nodes in the right renal hilum and retroperitoneum. The cystoscopy results were negative, and FISH analysis of urine exfoliated cells was positive, indicative of chromosome 3, 7, and 17 amplifications. Patient 2 was a 50-year-old male who was admitted to our hospital on May 2019 with no obvious cause of abdominal pain and abdominal distension (lasting for 7 days), with a serum creatinine level of 844 µmol/L. Patient 2 had no hematuria or fever, and MRI showed left renal inferior and medial space-occupying lesions, and multiple mesenteric nodules at the junction of the left adrenal gland, retroperitoneum, abdomen, and pelvis, which were partially fused. The tumor lesions were approximately 3.1 cm × 2.3 cm in size. The urine FISH results were positive, indicating chromosome 3, 7, and 17 amplifications. DIAGNOSES: Both patients were diagnosed with renal tumors with unknown pathology. INTERVENTIONS: Patient 1 underwent laparoscopic resection of the kidney and ureter, and sleeve cystectomy. The postoperative pathological diagnosis was metastatic keratinized squamous cell carcinoma, with squamous cell carcinoma in the right hilar lymph node. Histological FISH of the primary esophageal cancer and renal metastases were consistent with the urine FISH test results. Patient 2 underwent a biopsy of the left renal inferior and retroperitoneal areas, and was diagnosed with diffuse large B-cell lymphoma. OUTCOMES: Patient 1 survived 6 months after urological surgery. After treating patient 2 with the R-CHOP regimen and kinase inhibitors, his renal function recovered significantly and the mass become undetectable. LESSONS: Our results imply that FISH-positive renal occupying lesions should be considered as potential renal metastases with chromosome aberrations when making a differential diagnosis.

Attenuated total reflection Fourier-transform infrared spectral discrimination in human bodily fluids of oesophageal transformation to adenocarcinoma.

Diagnostic tools for the detection of early-stage oesophageal adenocarcinoma (OAC) are urgently needed. Our aim was to develop an accurate and inexpensive method using biofluids (plasma, serum, saliva or urine) for detecting oesophageal stages through to OAC (squamous; inflammatory; Barrett's; low-grade dysplasia; high-grade dysplasia; OAC) using attenuated total reflection Fourier-transform infrared (ATR-FTIR) spectroscopy. ATR-FTIR spectroscopy coupled with variable selection methods, with successive projections or genetic algorithms (GA) combined with quadratic discriminant analysis (QDA) were employed to identify spectral biomarkers in biofluids for accurate diagnosis in a hospital setting of different stages through to OAC. Quality metrics (Accuracy, Sensitivity, Specificity and F-score) and biomarkers of disease were computed for each model. For plasma, GA-QDA models using 15 wavenumbers achieved 100% classification for four classes. For saliva, PCA-QDA models achieved 100% for the inflammatory stage and high-quality metrics for other classes. For serum, GA-QDA models achieved 100% performance for the OAC stage using 13 wavenumbers. For urine, PCA-QDA models achieved 100% performance for all classes. Selected wavenumbers using a Student's t-test (95% confidence interval) identified a differentiation of the stages on each biofluid: plasma (929 cm-1 to 1431 cm-1, associated with DNA/RNA and proteins); saliva (1000 cm-1 to 1150 cm-1, associated with DNA/RNA region); serum (1435 cm-1 to 1573 cm-1, associated with methyl groups of proteins and Amide II absorption); and, urine (1681 cm-1 to 1777 cm-1, associated with a high frequency vibration of an antiparallel ß-sheet of Amide I and stretching vibration of lipids). Our methods have demonstrated excellent efficacy for a rapid, cost-effective method of diagnosis for specific stages to OAC. These findings suggest a potential diagnostic tool for oesophageal cancer and could be translated into clinical practice.

Nuclear magnetic resonance-based metabolomics and metabolic pathway networks from patient-matched esophageal carcinoma, adjacent noncancerous tissues and urine.

BACKGROUND: Several studies have demonstrated a correlation between esophageal cancer (EC) and perturbed urinary metabolomic profiles, but none has described the correlation between urine metabolite profiles and those of the tumor and adjacent esophageal mucosa in the same patient. AIM: To investigate how urinary metabolic phenotypes were linked to the changes in the biochemical landscape of esophageal tumors. METHODS: Nuclear magnetic resonance-based metabolomics were applied to esophageal tumor tissues and adjacent normal mucosal tissues alongside patient-matched urine samples. RESULTS: Analysis revealed that specific metabolite changes overlapped across both metrics, including glucose, glutamate, citrate, glycine, creatinine and taurine, indicating that the networks for metabolic pathway perturbations in EC, potentially involved in but not limited to disruption of fatty acid metabolism, glucose and glycolytic metabolism, tricarboxylic acid cycle and glutaminolysis. Additionally, changes in most urinary biomarkers correlated with changes in biomarker candidates in EC tissues, implying enhanced energy production for rapid cell proliferation. CONCLUSION: Overall, these associations provide evidence for distinct metabolic signatures and pathway disturbances between the tumor tissues and urine of EC patients, and changes in urinary metabolic signature could reflect reprogramming of the aforementioned metabolic pathways in EC tissues. Further investigation is needed to validate these initial findings using larger samples and to establish the underlying mechanism of EC progression.

Development of a metabolic pathway-based pseudo-targeted metabolomics method using liquid chromatography coupled with mass spectrometry.

The pseudo-targeted metabolomics approach was developed recently which combined the advantages of untargeted and targeted analysis. However, the current pseudo-targeted analysis method has limitations due to the technical characteristics. In this study, a novel metabolic pathway-based pseudo-targeted approach was proposed for urine metabolomics analysis using an ultra-high-performance liquid chromatography (UPLC)-MS/MS system operated in the multiple reaction monitoring (MRM) mode. MRM ion pairs were acquired from urine samples through untargeted analysis using UPLC-HRMS, as well as by searching for metabolites in related pathways in relevant databases and from previous relevant research, including amino acids, fatty acids, nucleosides, carnitines, glycolysis metabolites, and steroids. This improved pseudo-targeted method exhibited good repeatability and precision, and no complicated peak alignment was required. As a proof of concept, the developed novel method was applied to the discovery of urine biomarkers for patients with esophageal squamous cell carcinoma (ESCC). The results showed that ESCC patients had altered acylcarnitines, amino acids, nucleosides, and steroid derivative levels et al. compared to those of healthy controls. The novelty of this study lies in the fact that it provides an approach for acquiring MRM ion pairs not only from untargeted MS analysis but also from targeted searching for metabolites in related metabolic pathways. By improving the detection limit of low-abundance metabolites, it enlarges the range for the discovery of potential biomarkers. Our work provides a foundation for achieving pseudo-targeted metabolomics analysis on the widely used LC-MS/MS MRM platform.

A noninvasive cancer detection strategy based on gold nanoparticle surface-enhanced raman spectroscopy of urinary modified nucleosides isolated by affinity chromatography.

The search for tumor biomarkers in the urine for cancer diagnosis is currently a hot topic in clinical oncology, with potential for cancer screening and diagnosis. Modified nucleosides excreted through the urine are considered to be a general tumor marker for various cancer types. Herein, we explore a new method that utilizes surface-enhanced Raman scattering (SERS) spectroscopy to obtain a complete biochemical profile of urinary modified nucleosides. In our method, modified nucleosides are first isolated from urine sample utilizing the excellent separation ability of affinity chromatography; then supplemented with gold (Au) nanoparticles as substrate for SERS spectroscopy analysis. The obtained SERS spectra present rich diagnostic and fingerprinting type signatures of urinary modified nucleosides. The utility of this new method in cancer detection was evaluated by analyzing urine samples from three groups of subjects: nasopharyngeal cancer patients (n=62), esophageal cancer patients (n=55), and healthy volunteers (n=52). Partial least squares and linear discriminant analysis (PLS-DA) were used to analyze and classify the SERS spectra of urinary modified nucleosides from nasopharyngeal cancer, esophageal cancer, and the normal group, achieving diagnostic sensitivities of 95.2%, 90.9% and 98.1% and specificities of 97.2%, 98.2% and 95.7%, respectively. These results demonstrated great potential of this novel method for non-invasive and label-free cancer detection and screening.

Global metabolomics reveals potential urinary biomarkers of esophageal squamous cell carcinoma for diagnosis and staging.

We performed a metabolomics study using liquid chromatography-mass spectrometry (LC-MS) combined with multivariate data analysis (MVDA) to discriminate global urine profiles in urine samples from esophageal squamous cell carcinoma (ESCC) patients and healthy controls (NC). Our work evaluated the feasibility of employing urine metabolomics for the diagnosis and staging of ESCC. The satisfactory classification between the healthy controls and ESCC patients was obtained using the MVDA model, and obvious classification of early-stage and advanced-stage patients was also observed. The results suggest that the combination of LC-MS analysis and MVDA may have potential applications for ESCC diagnosis and staging. We then conducted LC-MS/MS experiments to identify the potential biomarkers with large contributions to the discrimination. A total of 83 potential diagnostic biomarkers for ESCC were screened out, and 19 potential biomarkers were identified; the variations between the differences in staging using these potential biomarkers were further analyzed. These biomarkers may not be unique to ESCCs, but instead result from any malignant disease. To further elucidate the pathophysiology of ESCC, we studied related metabolic pathways and found that ESCC is associated with perturbations of fatty acid ß-oxidation and the metabolism of amino acids, purines, and pyrimidines.

Esophageal squamous cell cancer in a highly endemic region.

AIM: To identify risk factors associated with esophageal cancer in Zambia and association between dietary intake and urinary 8-iso prostaglandin F2α (8-isoPGF2α). METHODS: We conducted a prospective, case control study at the University Teaching Hospital. Subjects included both individuals admitted to the hospital and those presenting for an outpatient upper endoscopy. Esophageal cancer cases were compared to age and sex-matched controls. Cases were defined as patients with biopsy proven esophageal cancer; controls were defined as subjects without endoscopic evidence of esophageal cancer. Clinical and dietary data were collected using a standard questionnaire, developed a priori. Blood was collected for human immunodeficiency virus (HIV) serology. Urine was collected, and 8-isoPGF2α was measured primarily by enzyme-linked immunosorbent assay and expressed as a ratio to creatinine. RESULTS: Forty five controls (mean age 54.2 ± 15.3, 31 male) and 27 cases (mean age 54.6 ± 16.4, 17 males) were studied. Body mass index was lower in cases (median 16.8) than controls (median 23.2), P = 0.01. Histopathologically, 25/27 (93%) were squamous cell carcinoma and 2/27 (7%) adenocarcinoma. More cases smoked cigarettes (OR = 11.24, 95%CI: 1.37-92.4, P = 0.02) but alcohol consumption and HIV seropositivity did not differ significantly (P = 0.14 for both). Fruit, vegetables and fish consumption did not differ significantly between groups (P = 0.11, 0.12, and 0.10, respectively). Mean isoprostane level was significantly higher in cases (0.03 ng/mg creatinine) than controls (0.01 ng/mg creatinine) (OR = 2.35, 95%CI: 1.19-4.65, P = 0.014). CONCLUSION: Smoking and isoprostane levels were significantly associated with esophageal cancer in Zambians, but diet, HIV status, and alcohol consumption were not.

Programmed cell death 6 interacting protein (PDCD6IP) and Rabenosyn-5 (ZFYVE20) are potential urinary biomarkers for upper gastrointestinal cancer.

PURPOSE: Cancer of the upper digestive tract (uGI) is a major contributor to cancer-related death worldwide. Due to a rise in occurrence, together with poor survival rates and a lack of diagnostic or prognostic clinical assays, there is a clear need to establish molecular biomarkers. EXPERIMENTAL DESIGN: Initial assessment was performed on urine samples from 60 control and 60 uGI cancer patients using MS to establish a peak pattern or fingerprint model, which was validated by a further set of 59 samples. RESULTS: We detected 86 cluster peaks by MS above frequency and detection thresholds. Statistical testing and model building resulted in a peak profiling model of five relevant peaks with 88% overall sensitivity and 91% specificity, and overall correctness of 90%. High-resolution MS of 40 samples in the 2-10 kDa range resulted in 646 identified proteins, and pattern matching identified four of the five model peaks within significant parameters, namely programmed cell death 6 interacting protein (PDCD6IP/Alix/AIP1), Rabenosyn-5 (ZFYVE20), protein S100A8, and protein S100A9, of which the first two were validated by Western blotting. CONCLUSIONS AND CLINICAL RELEVANCE: We demonstrate that MS analysis of human urine can identify lead biomarker candidates in uGI cancers, which makes this technique potentially useful in defining and consolidating biomarker patterns for uGI cancer screening.

Selected ion flow tube mass spectrometry analysis of volatile metabolites in urine headspace for the profiling of gastro-esophageal cancer.

Urine is considered an ideal biofluid for clinical investigation because it is obtained noninvasively and relatively large volumes are easily acquired. In this study, selected ion flow tube mass spectrometry (SIFT-MS) has been applied for the quantification of volatile organic compounds (VOCs) in the headspace vapor of urine samples, which were retrieved from three groups of patients with gastro-esophageal cancer, noncancer diseases of the upper gastro-intestinal tract, and a healthy cohort. Eleven VOCs have been investigated in this study. The concentrations of seven VOCs-acetaldehyde, acetone, acetic acid, hexanoic acid, hydrogen sulfide, methanol, and phenol-were found to be significantly different between cancer, positive control, and healthy groups using the Kruskal-Wallis test. The concentrations of acetaldehyde, acetone, acetic acid, hexanoic acid, hydrogen sulfide, and methanol were increased in the cancer cohort compared with healthy controls while the concentration of phenol decreased. The differences in the concentrations of ethanol, propanol, methyl phenol, and ethyl phenol were not significant between cancer and control groups. Receiver operating characteristics (ROC) analysis was applied for a combination of six VOCs (acetaldehyde, acetone, acetic acid, hexanoic acid, hydrogen sulfide, and methanol) to discriminate cancer patients from noncancer controls. The integrated area under ROC curve is 0.904. This result indicates that VOC profiling may be suitable in identifying those at high risk of gastro-esophageal cancer. Therefore, further investigations should be undertaken to assess the potential for VOC profiling as a new screening test in gastro-esophageal cancer.

Tobacco smoke biomarkers and cancer risk among male smokers in the Shanghai cohort study.

Metabolites of tobacco smoke constituents can be quantified in urine and other body fluids providing a realistic measure of carcinogen and toxicant dose in a smoker. Many previous studies have demonstrated that these metabolites - referred to as biomarkers in this paper - are related to tobacco smoke exposure. The studies reviewed here were designed to answer another question: are these substances also biomarkers of cancer risk? Using a prospective study design comparing biomarker levels in cancer cases and controls, all of whom were smokers, the results demonstrate that several of these biomarkers - total cotinine, total 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), r-1-,t-2,3,c-4-tetrahydroxy-1,2,3,4-tetrahydrophenanthrene (PheT), and total N'-nitrosonornicotine (NNN) - are biomarkers of cancer risk. Therefore, these biomarkers have the potential to become part of a cancer risk prediction algorithm for smokers.

Urinary metabolomic signature of esophageal cancer and Barrett's esophagus.

BACKGROUND: Esophageal adenocarcinoma (EAC) often presents at a late, incurable stage, and mortality has increased substantially, due to an increase in incidence of EAC arising out of Barrett's esophagus. When diagnosed early, however, the combination of surgery and adjuvant therapies is associated with high cure rates. Metabolomics provides a means for non- invasive screening of early tumor-associated perturbations in cellular metabolism. METHODS: Urine samples from patients with esophageal carcinoma (n = 44), Barrett's esophagus (n = 31), and healthy controls (n = 75) were examined using (1)H-NMR spectroscopy. Targeted profiling of spectra using Chenomx software permitted quantification of 66 distinct metabolites. Unsupervised (principal component analysis) and supervised (orthogonal partial least-squares discriminant analysis OPLS-DA) multivariate pattern recognition techniques were applied to discriminate between samples using SIMCA-P(+) software. Model specificity was also confirmed through comparison with a pancreatic cancer cohort (n = 32). RESULTS: Clear distinctions between esophageal cancer, Barrett's esophagus and healthy controls were noted when OPLS-DA was applied. Model validity was confirmed using two established methods of internal validation, cross-validation and response permutation. Sensitivity and specificity of the multivariate OPLS-DA models were summarized using a receiver operating characteristic curve analysis and revealed excellent predictive power (area under the curve = 0.9810 and 0.9627 for esophageal cancer and Barrett's esophagus, respectively). The metabolite expression profiles of esophageal cancer and pancreatic cancer were also clearly distinguishable with an area under the receiver operating characteristics curve (AUROC) = 0.8954. CONCLUSIONS: Urinary metabolomics identified discrete metabolic signatures that clearly distinguished both Barrett's esophagus and esophageal cancer from controls. The metabolite expression profile of esophageal cancer was also discrete from its precursor lesion, Barrett's esophagus. The cancer-specific nature of this profile was confirmed through comparison with pancreatic cancer. These preliminary results suggest that urinary metabolomics may have a future potential role in non-invasive screening in these conditions.

The involvement of serum serotonin levels producing radiation-induced bystander effects for an in vivo assay with fractionated high dose-rate (HDR) brachytherapy.

PURPOSE: The primary goal of this investigation was to observe whether measurable levels of bystander factor(s) can be detected in esophageal carcinoma patients' urine samples taken after undergoing high dose rate (HDR) intraluminal brachytherapy (ILBT). However, a small pilot study was developed to evaluate whether serotonin [5-Hydroxytryptamine (5-HT)] serum levels play an active role in the mechanisms of radiation-induced bystander effects (RIBE) at high doses. MATERIALS AND METHODS: In the present study, a colony-forming in vivo assay was developed and used for the detection of non-targeted effects. Samples of urine were collected from five esophageal carcinoma patients undergoing fractionated HDR-ILBT. To observe whether 5-HT modulates the bystander effect at higher doses, different batches of foetal bovine serum (FBS) and 5-HT were tested on the same urine samples before and after brachytherapy. RESULTS: Some of our data suggests statistically significant evidence for serotonin playing an active role as a signalling molecule at higher doses when patients underwent HDR-ILBT. CONCLUSION: However, a more thorough investigation, with a larger sample size, is warranted before serotonin can be known to play a role in bystander effects at this particular dose range and treatment regime.

Revealing the metabonomic variation of EC using ¹H-NMR spectroscopy and its association with the clinicopathological characteristics.

In this study, (1)H NMR-based metabonomics has been applied to investigate esophageal cancer metabolic signatures in plasma and urine, purpose of assessing the diagnostic potential of this approach and gaining novel insights into esophageal cancer metabolism and systemic effects. Plasma and urine samples from esophageal cancer patients (n = 108) and a control healthy group (n = 40) were analyzed by Nuclear Magnetic Resonance (NMR) spectroscopy (600 MHz), and their spectral profiles subjected to Orthogonal Projections to Latent Structures (OPLS-DA) for multivariate statistics. Potential metabolic biomarkers were identified using data base comparisons used for examining the significance of metabolites. Compared to healthy controls, esophageal cancer plasma had higher levels of dimethylamine, α-glucose, ß-glucose, citric acid, together with lower levels of Leucine, alanine, isoleucine, valine, glycoprotein, lactate, acetone, acetate, choline, isobutyrate, unsaturated lipid, VLDL, LDL, 1-methylhistidine; Compared to healthy controls, esophageal cancer urine had higher levels of Mannitol, glutamate, γ-propalanine, phenylalanine, acetate, allantoin, pyruvate, tyrosine, ß-glucose and guinolinate, together with lower levels of N-acetylcysteine, valine, dihydrothymine, hippurate, methylguanidine, 1-methylnicotin- amide and Citric acid; Very good discrimination between cancer and control groups was achieved by multivariate modeling of plasma and urinary profiles. (1)H NMR-based metabolite profiling analysis was shown to be an effective approach to differentiating between patients with EC and healthy subjects. Good sensitivity and selectivity were shown by using the metabolite markers discovered to predict the classification of samples from the healthy control group and the patients with the disease. Plasma and urine metabolic profiling may have potential for early diagnosis of EC and may enhance our understanding of its mechanisms.

Urinary levels of the tobacco-specific carcinogen N'-nitrosonornicotine and its glucuronide are strongly associated with esophageal cancer risk in smokers.

N'-Nitrosonornicotine (NNN) and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) are tobacco-specific nitrosamines. NNN and NNK can induce cancers of the esophagus and lung, respectively, in laboratory animals, but data on human esophageal cancer are lacking. The association between levels of NNN and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), an NNK metabolite, in urine samples collected before diagnosis and risk of esophageal cancer was examined in 77 patients with esophageal cancer and 223 individually matched controls, all current smokers, from a cohort of 18244 Chinese men in Shanghai, China, followed from 1986 to 2008. Urinary total NNN (free NNN plus NNN-N-glucuronide) was significantly higher, whereas the percentage of its detoxification product NNN-N-glucuronide was significantly lower in cases than controls. Odds ratios (95% confidence intervals) of esophageal cancer for the second and third tertiles of total NNN were 3.99 (1.25-12.7) and 17.0 (3.99-72.8), respectively, compared with the first tertile after adjustment for urinary total NNAL and total cotinine and smoking intensity and duration (P(trend) < 0.001). The corresponding figures for the percentage of NNN-N-glucuronides were 0.37 (0.17-0.80) and 0.27 (0.11-0.62) (P(trend) = 0.001). Urinary total NNN and the percentage of NNN-N-glucuronides almost completely accounted for the observed association for urinary total NNAL (free NNAL plus its glucuronides), urinary total cotinine and smoking intensity with esophageal cancer risk. These findings along with results of previous studies in laboratory animals support a significant and unique role of NNN in esophageal carcinogenesis in humans.

Fumonisin B1 as a urinary biomarker of exposure in a maize intervention study among South African subsistence farmers.

BACKGROUND: The consumption of maize highly contaminated with carcinogenic fumonisins has been linked to high oesophageal cancer rates. The aim of this study was to validate a urinary fumonisin B1 (UFB1) biomarker as a measure of fumonisin exposure and to investigate the reduction in exposure following a simple and culturally acceptable intervention. METHODS: At baseline home-grown maize, maize-based porridge, and first-void urine samples were collected from female participants (n=22), following their traditional food practices in Centane, South Africa. During intervention the participants were trained to recognize and remove visibly infected kernels, and to wash the remaining kernels. Participants consumed the porridge prepared from the sorted and washed maize on each day of the two-day intervention. Porridge, maize, and urine samples were collected for FB1 analyses. RESULTS: The geometric mean (95% confidence interval) for FB1 exposure based on porridge (dry weight) consumption at baseline and following intervention was 4.84 (2.87-8.14) and 1.87 (1.40-2.51) µg FB1/kg body weight/day, respectively, (62% reduction, P<0.05). UFB1C, UFB1 normalized for creatinine, was reduced from 470 (295-750) at baseline to 279 (202-386) pg/mg creatinine following intervention (41% reduction, P=0.06). The UFB1C biomarker was positively correlated with FB1 intake at the individual level (r=0.4972, P<0.01). Urinary excretion of FB1 was estimated to be 0.075% (0.054%-0.104%) of the FB1 intake. CONCLUSION: UFB1 reflects individual FB1 exposure and thus represents a valuable biomarker for future fumonisin risk assessment. IMPACT: The simple intervention method, hand sorting and washing, could positively impact on food safety and health in communities exposed to fumonisins.

Evaluation of fumonisin biomarkers in a cross-sectional study with two high-risk populations in China.

Levels of serum and urinary sphinganine (Sa) and sphingosine (So), the Sa/So ratio, and urinary-free fumonisin B(1) (FB(1)) were determined in a cross-sectional study consisting of 43 adults in Huaian and 34 adults in Fusui, China. Home-produced corn had 100% contamination with FB(1). There were 93.0% (40/43) of Huaian subjects and 52.9% (18/34) of Fusui subjects with daily FB(1) intakes greater than 2 microg kg(-1) body weight, which showed a significant difference (p < 0.01). Levels of sphinganine and sphingosine and the Sa/So ratio were not correlated with levels of dietary exposure. The median level of the serum Sa/So ratio in Huaian subjects (0.41, range = 0.14-0.85) was significantly lower than that in Fusui subjects (0.78, range = 0.57-1.08) (p < 0.01). The median level of the urinary Sa/So ratio was also significantly lower in Huaian subjects (0.31, range = 0.08-1.33) than in Fusui subjects (0.57, range = 0.03-2.52) (p < 0.01). Urinary-free FB(1) was detected in 83.7% (36/43) of Huaian samples and in 82.4% (28/34) of Fusui urine samples (p > 0.05). However, the median level of urinary-free FB(1) in Huaian subjects, 3.91 (range = 0.06-253.61) ng mg(-1) creatinine, was significantly higher than 0.39 (range = 0.01-3.72) ng mg(-1) creatinine found in Fusui subjects (p < 0.01). These results suggest that urinary-free FB(1) may be a potential biomarker for human fumonisin exposure, while further validation is needed in human epidemiological and intervention studies.

Mass spectrometric detection of candidate protein biomarkers of cancer cachexia in human urine.

Increased membrane permeability and myofibrillar protein breakdown are established features of cancer cachexia. Proteins released from cachectic muscle may be excreted in urine to act as biomarkers of the cachectic process. One-dimensional SDS polyacrylamide gel electrophoresis followed by matrix-assisted laser desorption/ionisation or liquid chromatography tandem mass spectrometry was used to compare the protein content of urine from cachectic (>10% weight loss) (n=8) and weight-stable (n=8) gastro-oesophageal cancer patients and healthy controls (n=8). Plasma creatine kinase concentration was used as a marker of gross muscle breakdown. The number of protein species identified in cachectic samples (median 42; range 28-61; total 199) was greater than that identified in weight-stable cancer (median 15; range 9-28; total 79) and control samples (median 12.5; range 5-18; total 49) (P<0.001). Many of the proteins identified have not been reported previously in the urine of cancer patients. Proteins identified specifically in cachectic samples included muscle (myosin species), cytoskeletal (alpha-spectrin; nischarin) and microtubule-associated proteins (microtubule-actin crosslinking factor; microtubule-associated protein-1B; bullous pemphigoid antigen 1), whereas immunoglobulin kappa-light chain and zinc alpha-2 glycoprotein appeared to represent markers of cancer. The presence of myosin in urine (without an increase in plasma creatine kinase) is consistent with a specific loss of myosin as part of the cachectic process. Urinary proteomics using mass spectrometry can identify muscle-specific and non-muscle-specific candidate biomarkers of cancer cachexia.

Galectin-3 in urine of cancer patients: stage and tissue specificity.

PURPOSE: Galectin-3 has been implicated in advanced stage of cancer disease. In the current study we examined the possibility of urinary galectin-3 levels to stage cancer disease and to follow up therapy. EXPERIMENTAL DESIGN: Urine was collected from all types of cancer patients at different stages including patients undergoing radio/chemotherapy. Galectin-3 level was determined by anti-galectin-3 based ELISA and agglutination assays. Immunoblotting and purification on lactosyl affinity column further confirmed the presence of galectin-3. RESULTS: Cancer samples exhibited stage dependent expression of galectin-3 approx. ranging from 1.0 to 3.3, 4.4 to 5.4, 5.4 to 24.7, 13.1 to 31.9, 13.9 to 32.9 ng/mg C (creatinine) for stage I-V, respectively, at P approximately <0.05 level. Galectin-3 levels were decreased by approx. threefolds after 5th day of therapy. CONCLUSIONS: Sample collection being simple and non-invasive, urinary galectin-3 may be used as a potential diagnostic tool for monitoring or follow up of the stage of cancer disease.