Coding and non-coding co-expression network analysis identifies key modules and driver genes associated with precursor lesions of gastric cancer.

BACKGROUND: Helicobacter pylori infection is the most important risk factor for gastric cancer (GC). Human gastric adenocarcinoma develops after long-term H. pylori infection via the Correa cascade. This carcinogenic pathway describes the progression from gastritis to atrophy, intestinal metaplasia (IM), dysplasia and GC. Patients with atrophy and intestinal metaplasia are considered to have precancerous lesions of GC (PLGC). H. pylori eradication and endoscopy surveillance are currently the only interventions for preventing GC. Better knowledge of the biology of human PLGC may help find stratification markers and contribute to better understanding of biological mechanisms. One way to achieve this is by using co-expression network analysis. Weighted gene co-expression network analysis (WGCNA) is often used to identify modules from co-expression networks and relate them to clinical traits. It also allows identification of driver genes that may be critical for PLGC. AIM: The purpose of this study was to identify co-expression modules and differential gene expression in dyspeptic patients at different stages of the Correa pathway. METHODS: We studied 96 gastric biopsies from 78 patients that were clinically classified as: non-active (n = 10) and chronic-active gastritis (n = 20), atrophy (n = 12), and IM (n = 36). Gene expression of coding RNAs was determined by microarrays and non-coding RNAs by RNA-seq. The WGCNA package was used for network construction, module detection, module preservation and hub and driver gene selection. RESULTS: WGCNA identified 20 modules for coding RNAs and 4 for each miRNA and small RNA class. Modules were associated with antrum and corpus gastric locations, chronic gastritis and IM. Notably, coding RNA modules correlated with the Correa cascade. One was associated with the presence of H. pylori. In three modules, the module eigengene (ME) gradually increased in the stages toward IM, while in three others the inverse relationship was found. One miRNA module was negatively correlated to IM and was used for a mRNA-miRNA integration analysis. WGCNA also uncovered driver genes. Driver genes show both high connectivity within a module and are significantly associated with clinical traits. Some of those genes have been previously involved in H. pylori carcinogenesis, but others are new. Lastly, using similar external transcriptomic data, we confirmed that the discovered mRNA modules were highly preserved. CONCLUSION: Our analysis captured co-expression modules that provide valuable information to understand the pathogenesis of the progression of PLGC.

Cross-sectional study of human coding- and non-coding RNAs in progressive stages of Helicobacter pylori infection.

Helicobacter pylori infects 4.4 billion individuals worldwide and is considered the most important etiologic agent for peptic ulcers and gastric cancer. Individual response to H. pylori infection is complex and depends on complex interactions between host and environmental factors. The pathway towards gastric cancer is a sequence of events known as Correa's model of gastric carcinogenesis, a stepwise inflammatory process from normal mucosa to chronic-active gastritis, atrophy, metaplasia and gastric adenocarcinoma. This study examines gastric clinical specimens representing different steps of the Correa pathway with the aim of identifying the expression profiles of coding- and non-coding RNAs that may have a role in Correa's model of gastric carcinogenesis. We screened for differentially expressed genes in gastric biopsies by employing RNAseq, microarrays and qRT-PCR. Here we provide a detailed description of the experiments, methods and results generated. The datasets may help other scientists and clinicians to find new clues to the pathogenesis of H. pylori and the mechanisms of progression of the infection to more severe gastric diseases. Data is available via ArrayExpress.

Expression profile of circulating microRNAs in the Correa pathway of progression to gastric cancer.

BACKGROUND: Helicobacter pylori infection causes long-term chronic active gastritis, a risk factor for the intestinal and diffuse forms of gastric cancer. Most gastric cancers develop in a stepwise progression from chronic active gastritis to precursor lesions of gastric cancer. The early detection of gastric cancer improves survival. Studies with recent evidence have proposed circulating-microRNAs as biomarkers of cancer. OBJECTIVE: The purpose of this study was to explore the circulating-microRNA profile from H. pylori infection to gastric adenocarcinoma. METHODS: One hundred and twenty-three patients were enrolled and assigned to the discovery or the validation sets. In the discovery phase, circulating-microRNAs were measured by dye-based quantitative polymerase chain reaction and a selection of circulating-microRNAs was validated by probe-based quantitative polymerase chain reaction. A quality control protocol was used. RESULTS: One hundred and sixty-seven circulating-microRNAs were detected. Precursor lesions of gastric cancer and gastric cancer patients showed the downregulation of eight and five circulating-microRNAs, respectively. We further validated the deregulation of miR-196a-5p in precursor lesions of gastric cancer and the deregulation of miR-134-5p, miR-144-3p and miR-451a in gastric cancer. However, circulating-microRNAs exhibited moderate diagnostic performance due to the overlap of circulating-microRNA expression between non-cancer and cancer patients. miR-144-3p/miR-451a expression levels were correlated. Interestingly, these microRNAs are in 17q11.2, a site of rearrangements associated with gastric cancer. CONCLUSION: Circulating-microRNAs are deregulated in precancerous and gastric cancer patients but efforts are needed to improve their diagnostic accuracy.

Occult H. pylori infection partially explains 'false-positive' results of (13)C-urea breath test.

BACKGROUND: In a previous study, UBiT-100 mg, (Otsuka, Spain), a commercial (13)C-urea breath test omitting citric acid pre-treatment, had a high rate of false-positive results; however, it is possible that UBiT detected low-density 'occult' infection missed by other routine reference tests. We aimed to validate previous results in a new cohort and to rule out the possibility that false-positive UBiT were due to an 'occult' infection missed by reference tests. METHODS: Dyspeptic patients (n = 272) were prospectively enrolled and UBiT was performed, according to the manufacturer's recommendations. Helicobacter pylori infection was determined by combining culture, histology and rapid urease test results. We calculated UBiT sensitivity, specificity, positive and negative predictive values (with 95% CI). In addition, we evaluated 'occult' H. pylori infection using two previously-validated polymerase chain reaction (PCR) methods for urease A (UreA) and 16 S sequences in gastric biopsies. We included 44 patients with a false-positive UBiT, and two control groups of 25 patients each, that were positive and negative for all H. pylori tests. RESULTS: UBiT showed a false-positive rate of 17%, with a specificity of 83%. All the positive controls and 12 of 44 patients (27%) with false-positive UBiT were positive for all two PCR tests; by contrast, none of our negative controls had two positive PCR tests. CONCLUSIONS: UBiT suffers from a high rate of false-positive results and sub-optimal specificity, and the protocol skipping citric acid pre-treatment should be revised; however, low-density 'occult' H. pylori infection that was undetectable by conventional tests accounted for around 25% of the 'false-positive' results.