Taotao Liu

Metabolomic profiling of human urine in hepatocellular carcinoma patients using gas chromatography/mass spectrometry

With the technique of metabolomics, gas chromatography/mass spectrometry (GC/MS), urine or serum metabolites can be assayed to explore disease biomarkers. In this work, we present a metabolomic method to investigate the urinary metabolic difference between hepatocellular carcinoma (HCC, n - 20) male patients and normal male subjects (n - 20). The urinary endogenous metabolome was assayed using chemical derivatization followed by GC/MS. After GC/MS analysis, 103 metabolites were detected, of which 66 were annotated as known compounds. By a two sample t-test statistics with p < 0.05, 18 metabolites were shown to be significantly different between the HCC and control groups. A diagnostic model was constructed with a combination of 18 marker metabolites or together with alphafetoprotein, using principal component analysis and receiver-operator characteristic curves. The multivariate statistics of the diagnostic model yielded a separation between the two groups with an area under the curve value of 0.9275. This non-invasive technique of identifying HCC biomarkers from urine may have clinical utility.

Investigation of volatile biomarkers in liver cancer blood using solid-phase microextraction and gas chromatography/mass spectrometry.

Solid-phase microextraction (SPME) followed by gas chromatography/mass spectrometry (GC/MS) was used for the detection of liver cancer volatile biomarkers. Headspace SPME conditions (fiber coating, extraction temperature and extraction time) and desorption conditions were optimized and applied to the determination of volatiles in human blood. Between the liver cancer group (n = 19) and the normal group (n = 18), positive rates of 19 volatile compounds among the total of 47 detected were found to be different with statistical significance (p < 0.05, chi-squared test). We suggested hexanal, 1-octen-3-ol and octane, of the 19 compounds, as biomarkers of liver cancer with clinical diagnostic value. The sensitivity and specificity of 94.7% and 100% for hexanal, 84.2% and 100% for 1-octen-3-ol, and 89.5% and 100% for octane were obtained, respectively, after the cutoff values had been properly established. These results show that SPME-GC/MS is a simple, rapid and sensitive method for the investigation of volatile disease markers in human blood.

Metabolomic investigation of gastric cancer tissue using gas chromatography/mass spectrometry

Gastric cancer screening or diagnosis is mainly based on endoscopy and biopsy. The aim of this study was to identify the difference of metabolomic profile between normal and malignant gastric tissue, and to further explore tumor biomarkers. Chemical derivatization together with gas chromatography/mass spectrometry (GC/MS) was utilized to obtain the metabolomic information of the malignant and non-malignant tissues of gastric mucosae in 18 gastric cancer patients. Acquired metabolomic data was analyzed using the Wilcoxon rank sum test to find the tissue metabolic biomarkers for gastric cancer. A diagnostic model for gastric cancer was constructed using principal component analysis (PCA), and was assessed with receiver-operating characteristic (ROC) curves. Results showed that 18 metabolites were detected differently between the malignant tissues and the adjacent non-malignant tissues of gastric mucosa. Five metabolites were also detected differently between the non-invasive tumors and the invasive tumors. The diagnostic model could discriminate tumors from normal mucosae with an area under the curve (AUC) value of 0.9629, and another diagnostic model constructed for clinical staging was assessed with an AUC value of 0.969. We conclude that the metabolomic profile of malignant gastric tissue was different from normal, and that the selected tissue metabolites could probably be applied for clinical diagnosis or staging for gastric cancer.

Metabolomic study for diagnostic model of oesophageal cancer using gas chromatography/mass spectrometry.

The prognosis for oesophageal cancer is poor. Attempts have been made for the identification of biomarkers for early diagnosis. Metabolomic panel has been evaluated as potential candidate biomarkers. With gas chromatography/mass spectrometry (GC/MS) as a sensitive modality for metabolomics, various tissue metabolites can be detected and identified. We hypothesized that tissue metabolomic biomarkers may be identifiable and diagnostically useful for oesophageal cancer. We present a metabolomic method of chemical derivatization followed by GC/MS to analyze the metabolic difference in biopsied specimens between oesophageal cancer and corresponding normal mucosae obtained from 20 oesophageal cancer patients. The GC/MS data was analyzed using a two sample t-test to explore the potential metabolic biomarkers for oesophageal cancer. A diagnostic model was constructed to discriminate normal from malignant samples, using principal component analysis (PCA) and receiver-operating characteristic (ROC) curves. t-Test showed a total of 20 marker metabolites detected were found to be different with statistical significance (P<0.05). The multivariate logistic analysis yielded a complete distinction between the two groups. The diagnostic model could discriminate tumors from normal mucosae with an area under the curve (AUC) value of 1. Our findings suggest that this assay may potentially provide a new metabolomic biomarker for oesophageal cancer.

A serum metabolomic investigation on hepatocellular carcinoma patients by chemical derivatization followed by gas chromatography/mass spectrometry

The purpose of this study was to investigate the serum metabolic difference between hepatocellular carcinoma (HCC, n = 20) male patients and normal male subjects (n = 20). Serum metabolome was detected through chemical derivatization followed by gas chromatography/mass spectrometry (GC/MS). The acquired GC/MS data was analyzed by stepwise discriminant analysis (SDA) and support vector machine (SVM). The metabolites including butanoic acid, ethanimidic acid, glycerol, L-isoleucine, L-valine, aminomalonic acid, D-erythrose, hexadecanoic acid, octadecanoic acid, and 9,12-octadecadienoic acid in combination with each other gave the strongest segregation between the two groups. By applying these variables, our method provided a diagnostic model that could well discriminate between HCC patients and normal subjects. More importantly, the error count estimate for each group was 0%. The total classifying accuracy of the discriminant function tested by SVM 20-fold cross validation was 75%. This technique is different from traditional ones and appears to be a useful tool in the area of HCC diagnosis.

GC/MS-based metabolomic approach to validate the role of urinary sarcosine and target biomarkers for human prostate cancer by microwave-assisted derivatization

A recent study showed that sarcosine may be potentially useful for the diagnosis and prognosis of prostate cancer (PCa). The aim of this study was to validate diagnostic value of sarcosine for PCa, to evaluate urine metabolomic profiles in patients with PCa in comparison of non-cancerous control, and to further explore the other potential metabolic biomarkers for PCa. Isotope dilution gas chromatography/mass spectrometry (ID GC/MS) metabolomic approach was applied to evaluate sarcosine using [methyl-D3]-sarcosine as an internal standard. Microwave-assisted derivatization (MAD) together with GC/MS was utilized to obtain the urinary metabolomic information in 20 PCa patients compared with eight patients with benign prostate hypertrophy and 20 healthy men. Acquired metabolomic data were analyzed using a two-sample t test. Diagnostic models for PCa were constructed using principal component analysis and were assessed with receiver–operating characteristic curves. Results showed that the urinary sarcosine level has no statistical difference between the PCa group and the control group. In addition, nine metabolomic markers between the PCa group and the healthy male group were selected, which constructed a diagnostic model with a high area under the curve value of 0.9425. We conclude that although urinary sarcosine value has limited potential in the diagnostic algorithm of PCa, urinary metabolomic panel based on GC/MS assay following MAD may potentially become a diagnostic tool for PCa.

Circulating microRNAs as a Fingerprint for Liver Cirrhosis

Background Sensitive and specific detection of liver cirrhosis is an urgent need for optimal individualized management of disease activity. Substantial studies have identified circulation miRNAs as biomarkers for diverse diseases including chronic liver diseases. In this study, we investigated the plasma miRNA signature to serve as a potential diagnostic biomarker for silent liver cirrhosis. Methods A genome-wide miRNA microarray was first performed in 80 plasma specimens. Six candidate miRNAs were selected and then trained in CHB-related cirrhosis and controls by qPCR. A classifier, miR-106b and miR-181b, was validated finally in two independent cohorts including CHB-related silent cirrhosis and controls, as well as non−CHB-related cirrhosis and controls as validation sets, respectively. Results A profile of 2 miRNAs (miR-106b and miR-181b) was identified as liver cirrhosis biomarkers irrespective of etiology. The classifier constructed by the two miRNAs provided a high diagnostic accuracy for cirrhosis (AUC = 0.882 for CHB-related cirrhosis in the training set, 0.774 for CHB-related silent cirrhosis in one validation set, and 0.915 for non−CHB-related cirrhosis in another validation set). Conclusion Our study demonstrated that the combined detection of miR-106b and miR-181b has a considerable clinical value to diagnose patients with liver cirrhosis, especially those at early stage.

Intratumor Hypoxia Promotes Immune Tolerance by Inducing Regulatory T Cells via TGF-β1 in Gastric Cancer

Regulatory T cell (Treg)-mediated immunosuppression represents one of the crucial tumor immune evasion mechanisms and is a main obstacle for successful tumor immunotherapy. Hypoxia, a common feature of solid tumors, has been associated with potentiated immunosuppression, decreased therapeutic response, malignant progression and local invasion. Unfortunately, the link between hypoxia and Treg-mediated immune tolerance in gastric cancer remains poorly understood. In our study, Tregs and hypoxia inducible factor-1α were found to be positively correlated with each other and were increased with the tumor progression. A subsequent in vitro study indicated that supernatants derived from gastric cancer cells under hypoxic condition, could induce the expression of Foxp3 via TGF-β1. These findings confirmed the crucial role of Tregs as a therapeutic target in gastric cancer therapy and provided helpful thoughts for the design of immunotherapy for gastric cancer in the future.

Ubiquitin C-terminal Hydrolase 37, a novel predictor for hepatocellular carcinoma recurrence, promotes cell migration and invasion via interacting and deubiquitinating PRP19

Ubiquitin C-terminal hydrolase 37 (UCH37) plays a crucial role in numerous biological processes and is also involved in oncogenesis. In this study, clinicopathologic data showed that UCH37 was over-expressed in hepatocellular carcinoma (HCC) cancerous tissues and was a significant predictor for time to recurrence (TTR). In vitro, we discovered that UCH37 could promote cell migration and invasion. Subsequently, we utilized Isobaric Tags for Relative and Absolute Quantitation (iTRAQ) to identify differentially expressed proteins in UCH37 over-expressing cells compared with the control cells, and found that PRP19, an essential RNA splicing factor, was up-regulated. The relationship between UCH37, PRP19 and the capability of cell migration and invasion was further confirmed. Collectively, this study demonstrated that UCH37 could promote cell migration and invasion in HCC cell lines through interacting and deubiquitinating PRP19, and suggested that UCH37 could be a novel predictor for HCC recurrence after curative resection.

Gas chromatography/mass spectrometry screening of serum metabolomic biomarkers in hepatitis B virus infected cirrhosis patients.

Abstract Background: Metabolomic studies have been applied to disease biomarkers selection. With the metabolomic technique, gas chromatography/mass spectrometry (GC/MS), human serum metabolites can be detected and identified. The purpose of this study was to investigate the serum metabolic profile of hepatitis B virus (HBV) infected cirrhosis patients and to detect disease biomarkers. Methods: HBV infected non-cirrhosis male subjects (n=20) and HBV infected cirrhosis male patients (n=20) participated in this experiment. Serum metabolome was detected through chemical derivatization followed by GC/MS. The high-flux metabolomic data were analyzed by stepwise discriminant analysis. Results: Out of the 41 metabolites detected in serum, we selected metabolites, including acetic acid, sorbitol, D-lactic acid, hexanoic acid, 1-naphthalenamine, butanoic acid, phosphoric acid, D-glucitol, and glucose, which in combination with each other could segregate the two groups. The error count was 0% for the non-cirrhosis group and 25% for the cirrhosis group. Conclusions: This technique can be used to select biomarkers for hepatic cirrhosis. Clin Chem Lab Med 2009;47:305–10.