A noninvasive multianalytical approach establishment for risk assessment and gastric cancer screening.

Effective screening and early detection are critical to improve the prognosis of gastric cancer (GC). Our study aims to explore noninvasive multianalytical biomarkers and construct integrative models for preliminary risk assessment and GC detection. Whole genomewide methylation marker discovery was conducted with CpG tandems target amplification (CTTA) in cfDNA from large asymptomatic screening participants in a high-risk area of GC. The methylation and mutation candidates were validated simultaneously using one plasma from patients at various gastric lesion stages by multiplex profiling with Mutation Capsule Plus (MCP). Helicobacter pylori specific antibodies were detected with a recomLine assay. Integrated models were constructed and validated by the combination of multianalytical biomarkers. A total of 146 and 120 novel methylation markers were found in CpG islands and promoter regions across the genome with CTTA. The methylation markers together with the candidate mutations were validated with MCP and used to establish a 133-methylation-marker panel for risk assessment of suspicious precancerous lesions and GC cases and a 49-methylation-marker panel as well as a 144-amplicon-mutation panel for GC detection. An integrated model comprising both methylation and specific antibody panels performed better for risk assessment than a traditional model (AUC, 0.83 and 0.63, P < .001). A second model for GC detection integrating methylation and mutation panels also outperformed the traditional model (AUC, 0.82 and 0.68, P = .005). Our study established methylation, mutation and H. pylori-specific antibody panels and constructed two integrated models for risk assessment and GC screening. Our findings provide new insights for a more precise GC screening strategy in the future.

Beneficial effects of endoscopic screening on gastric cancer and optimal screening interval: a population-based study.

BACKGROUND : The effectiveness of endoscopic screening on gastric cancer has not been widely investigated in China and the screening interval of repeated screening has not been determined. METHODS : In a population-based prospective study, we included 375,800 individuals, 14,670 of whom underwent endoscopic screening (2012-2018). We assessed the associations between endoscopic screening and risk of incident gastric cancer and gastric cancer-specific mortality, and examined changes in overall survival and disease-specific survival following screening. The optimal screening interval for repeated endoscopy for early detection of gastric cancer was explored. RESULTS : Ever receiving endoscopic screening significantly decreased the risk of invasive gastric cancer (age- and sex-adjusted relative risk [RR] 0.69, 95 % confidence interval [CI] 0.52-0.92) and gastric cancer-specific mortality (RR 0.33, 95 %CI 0.20-0.56), particularly for noncardia gastric cancer. Repeated screening strengthened the beneficial effect on invasive gastric cancer-specific mortality of one-time screening. Among invasive gastric cancers, screening-detected individuals had significantly better overall survival (RR 0.18, 95 %CI 0.13-0.25) and disease-specific survival (RR 0.18, 95 %CI 0.13-0.25) than unscreened individuals, particularly for those receiving repeated endoscopy. For individuals with intestinal metaplasia or low grade intraepithelial neoplasia, repeated endoscopy at an interval of < 2 years, particularly within 1 year, significantly enhanced the detection of early gastric cancer, compared with repeated screening after 2 years (P-trend = 0.02). CONCLUSION : Endoscopic screening prevented gastric cancer occurrence and death, and improved its prognosis in a population-based study. Repeated endoscopy enhanced the effectiveness. Screening interval should be based on gastric lesion severity.

DNA methylation signatures associated with prognosis of gastric cancer.

BACKGROUND: Few studies have examined prognostic outcomes-associated molecular signatures other than overall survival (OS) for gastric cancer (GC). We aimed to identify DNA methylation biomarkers associated with multiple prognostic outcomes of GC in an epigenome-wide association study. METHODS: Based on the Cancer Genome Atlas (TCGA), DNA methylation loci associated with OS (n = 381), disease-specific survival (DSS, n = 372), and progression-free interval (PFI, n = 383) were discovered in training set subjects (false discovery rates < 0.05) randomly selected for each prognostic outcome and were then validated in remaining subjects (P-values < 0.05). Key CpGs simultaneously validated for OS, DSS, and PFI were further assessed for disease-free interval (DFI, n = 247). Gene set enrichment analyses were conducted to explore the Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathways simultaneously enriched for multiple GC prognostic outcomes. Methylation correlated blocks (MCBs) were identified for co-methylation patterns associated with GC prognosis. Based on key CpGs, risk score models were established to predict four prognostic outcomes. Spearman correlation analyses were performed between key CpG sites and their host gene mRNA expression. RESULTS: We newly identified DNA methylation of seven CpGs significantly associated with OS, DSS, and PFI of GC, including cg10399824 (GRK5), cg05275153 (RGS12), cg24406668 (MMP9), cg14719951(DSC3), and cg25117092 (MED12L), and two in intergenic regions (cg11348188 and cg11671115). Except cg10399824 and cg24406668, five of them were also significantly associated with DFI of GC. Neuroactive ligand-receptor interaction pathway was suggested to play a key role in the effect of DNA methylation on GC prognosis. Consistent with individual CpG-level association, three MCBs involving cg11671115, cg14719951, and cg24406668 were significantly associated with multiple prognostic outcomes of GC. Integrating key CpG loci, two risk score models performed well in predicting GC prognosis. Gene body DNA methylation of cg14719951, cg10399824, and cg25117092 was associated with their host gene expression, whereas no significant associations between their host gene expression and four clinical prognostic outcomes of GC were observed. CONCLUSIONS: We newly identified seven CpGs associated with OS, DSS, and PFI of GC, with five of them also associated with DFI, which might inform patient stratification in clinical practices.

Suppression of Helicobacter pylori infection by daily cranberry intake: A double-blind, randomized, placebo-controlled trial.

BACKGROUND AND AIM: Dietary strategies that contribute to reducing incidence of Helicobacter pylori infection without negative side effects are highly desirable owing to worldwide bacterial prevalence and carcinogenesis potential. The aim of this study was to determine dosage effect of daily cranberry consumption on H. pylori suppression over time in infected adults to assess the potential of this complementary management strategy in a region with high gastric cancer risk and high prevalence of H. pylori infection. METHODS: This double-blind, randomized, placebo-controlled trial on 522 H. pylori-positive adults evaluated dose-response effects of proanthocyanidin-standardized cranberry juice, cranberry powder, or their placebos on suppression of H. pylori at 2 and 8 weeks by 13 C-urea breath testing and eradication at 45 days post-intervention. RESULTS: H. pylori-negative rates in placebo, low-proanthocyanidin, medium-proanthocyanidin, and high-proanthocyanidin cranberry juice groups at week 2 were 13.24%, 7.58%, 1.49%, and 13.85% and at week 8 were 7.35%, 7.58%, 4.48%, and 20.00%, respectively. Consumption of high-proanthocyanidin juice twice daily (44 mg proanthocyanidin/240-mL serving) for 8 weeks resulted in decreased H. pylori infection rate by 20% as compared with other dosages and placebo (P < 0.05). Percentage of H. pylori-negative participants increased from 2 to 8 weeks in subjects who consumed 44 mg proanthocyanidin/day juice once or twice daily, showing a statistically significant positive trend over time. Encapsulated cranberry powder doses were not significantly effective at either time point. Overall trial compliance was 94.25%. Cranberry juice and powder were well-tolerated. CONCLUSIONS: Twice-daily consumption of proanthocyanidin-standardized cranberry juice may help potentiate suppression of H. pylori infection. TRIAL REGISTRATION: ChiCTR1800017522, per WHO ICTRP.

Effect of Helicobacter pylori on gastrointestinal microbiota: a population-based study in Linqu, a high-risk area of gastric cancer.

OBJECTIVE: Gastrointestinal microbiota may be involved in Helicobacter pylori-associated gastric cancer development. The aim of this study was to explore the possible microbial mechanisms in gastric carcinogenesis and potential dysbiosis arising from H. pylori infection. DESIGN: Deep sequencing of the microbial 16S ribosomal RNA gene was used to investigate alterations in paired gastric biopsies and stool samples in 58 subjects with successful and 57 subjects with failed anti-H. pylori treatment, relative to 49 H. pylori negative subjects. RESULTS: In H. pylori positive subjects, richness and Shannon indexes increased significantly (both p<0.001) after successful eradication and showed no difference to those of negative subjects (p=0.493 for richness and p=0.420 for Shannon index). Differential taxa analysis identified 18 significantly altered gastric genera after eradication. The combination of these genera into a Microbial Dysbiosis Index revealed that the dysbiotic microbiota in H. pylori positive mucosa was associated with advanced gastric lesions (chronic atrophic gastritis and intestinal metaplasia/dysplasia) and could be reversed by eradication. Strong coexcluding interactions between Helicobacter and Fusobacterium, Neisseria, Prevotella, Veillonella, Rothia were found only in advanced gastric lesion patients, and were absent in normal/superficial gastritis group. Changes in faecal microbiota included increased Bifidobacterium after successful H. pylori eradication and more upregulated drug-resistant functional orthologs after failed treatment. CONCLUSION: H. pylori infection contributes significantly to gastric microbial dysbiosis that may be involved in carcinogenesis. Successful H. pylori eradication potentially restores gastric microbiota to a similar status as found in uninfected individuals, and shows beneficial effects on gut microbiota.

Association of genetic polymorphisms of interleukins with gastric cancer and precancerous gastric lesions in a high-risk Chinese population.

Helicobacter pylori (H. pylori) infection and cytokine-mediated inflammatory responses play important roles in gastric cancer (GC) pathogenesis. To investigate an association between genetic polymorphisms in interleukin (IL)-1ß, IL-4R, IL-8, IL-10, IL-16, IL-18RAP, IL-22, and IL-32 and risks of GC and its precursors, a population-based study was conducted in Linqu County. Genotypes were determined by Sequenom MassARRAY platform in 132 GC cases and 1198 subjects with gastric lesions. The H. pylori status was determined by (13)C-urea breath test ((13)C-UBT) or enzyme-linked immunosorbent assay (ELISA). Among 11 candidate single nucleotide polymorphisms (SNPs), subjects carrying IL-18RAP rs917997 AA genotype were associated with risk of GC [adjusted odds ratio (OR) = 1.83, 95 % confidence interval (CI) 1.14-2.92] or chronic atrophic gastritis (CAG; OR = 1.55, 95 % CI 1.07-2.24). The risk of GC was also increased in subjects carrying IL-32 rs2015620 A allele (AA + AT; OR = 1.92, 95 % CI 1.09-3.39). Moreover, elevated risks of CAG (OR = 2.64, 95 % CI 1.89-3.69), intestinal metaplasia (IM; OR = 5.58, 95 % CI 3.86-8.05), and dysplasia (DYS; OR = 1.64, 95 % CI 1.18-2.26) were observed in subjects with IL-22 rs1179251 CC genotype. Stratified analysis indicated that risks of GC and its precursors were elevated in subjects with IL-32 rs2015620 A allele (AA + AT) or IL-22 rs1179251 CC genotype and H. pylori infection, and significant interactions between these two SNPs and H. pylori infection were found. These findings suggested that IL-18RAP rs917997, IL-32 rs2015620, IL-22 rs1179251, and interactions between these polymorphisms and H. pylori infection were associated with risks of gastric lesions. Genetic polymorphisms of interleukins may play crucial roles in H. pylori-induced gastric carcinogenesis.

Helicobacter pylori antibody responses and evolution of precancerous gastric lesions in a Chinese population.

Helicobacter pylori-specific proteins are involved in gastric carcinogenesis. To investigate the seroprevalence of six H. pylori-specific antibodies in patients with different gastric histology, and the impact of seropositivities on the evolution of precancerous gastric lesions, a follow-up study was conducted in Linqu County, China. The seropositivities for CagA, VacA, GroEL, UreA, HcpC and gGT were assessed by recomLine analysis in 573 H. pylori-positive subjects and correlated with evolution of precancerous gastric lesions. We found that the score of H. pylori recomLine test was significantly increased in subjects with chronic atrophic gastritis (CAG, p < 0.0001) or intestinal metaplasia (IM, p = 0.0125), and CagA was an independent predictor of advanced gastric lesions, adjusted odds ratios (ORs) were 2.54 (95% CI = 1.42-4.55) for IM and 2.38 (95% CI = 1.05-5.37) for dysplasia (DYS). Moreover, seropositivities for CagA and GroEL were identified as independent predictors for progression of gastric lesions in a longitudinal study, and ORs were 2.89 (95% CI = 1.27-6.59) and 2.20 (95% CI = 1.33-3.64), respectively. Furthermore, the risk of progression was more pronounced in subjects with more than three positive antigens (p(for) trend = 0.0003). This population-based study revealed that seropositivities for CagA and GroEL might be potential markers to identify patients infected with high-risk H. pylori strains, which are related to the development of GC in a Chinese high-risk population, and recomLine test might serve as a tool for risk stratification.

Magnetic resonance imaging-based lymph node radiomics for predicting the metastasis of evaluable lymph nodes in rectal cancer.

BACKGROUND: Lymph node (LN) staging in rectal cancer (RC) affects treatment decisions and patient prognosis. For radiologists, the traditional preoperative assessment of LN metastasis (LNM) using magnetic resonance imaging (MRI) poses a challenge. AIM: To explore the value of a nomogram model that combines Conventional MRI and radiomics features from the LNs of RC in assessing the preoperative metastasis of evaluable LNs. METHODS: In this retrospective study, 270 LNs (158 nonmetastatic, 112 metastatic) were randomly split into training (n = 189) and validation sets (n = 81). LNs were classified based on pathology-MRI matching. Conventional MRI features [size, shape, margin, T2-weighted imaging (T2WI) appearance, and CE-T1-weighted imaging (T1WI) enhancement] were evaluated. Three radiomics models used 3D features from T1WI and T2WI images. Additionally, a nomogram model combining conventional MRI and radiomics features was developed. The model used univariate analysis and multivariable logistic regression. Evaluation employed the receiver operating characteristic curve, with DeLong test for comparing diagnostic performance. Nomogram performance was assessed using calibration and decision curve analysis. RESULTS: The nomogram model outperformed conventional MRI and single radiomics models in evaluating LNM. In the training set, the nomogram model achieved an area under the curve (AUC) of 0.92, which was significantly higher than the AUCs of 0.82 (P < 0.001) and 0.89 (P < 0.001) of the conventional MRI and radiomics models, respectively. In the validation set, the nomogram model achieved an AUC of 0.91, significantly surpassing 0.80 (P < 0.001) and 0.86 (P < 0.001), respectively. CONCLUSION: The nomogram model showed the best performance in predicting metastasis of evaluable LNs.

A multiserological line assay to potentially discriminate current from past Helicobacter pylori infection.

OBJECTIVES: Early diagnosis is important in controlling Helicobacter pylori-induced gastritis and progression to gastric malignancy. Serological testing is an efficient non-invasive diagnostic method, but currently does not allow differentiation between active and past infections. To fill this diagnostic gap we investigated the diagnostic value of a panel of ten H. pylori-specific antibodies in individuals with different H. pylori infection status within a German population. METHODS: We used the recomLine Helicobacter IgG 2.0 immunoblotting assay to analyse ten H. pylori-specific antibodies in serum samples collected from 1108 volunteers. From these, 788 samples were used to build exposure and infection status models and 320 samples for model validation. H. pylori infection status was verified by histological examination. We applied logistic regression to select antibodies correlated to infection status and developed, with independent validation, discriminating models and risk scores. Receiving operating characteristic analysis was performed to assess the accuracy of the discriminating models. RESULTS: Antibody reactivity against cytotoxin-associated gene A (CagA), H. pylori chaperone (GroEL), and hook-associated protein 2 homologue (FliD) was independently associated with the risk of H. pylori exposure with ORs and 95% CIs of 99.24 (46.50-211.80), 46.17 (17.45-122.17), and 22.16 (8.46-55.04), respectively. A risk score comprising these three selected antibodies differentiated currently H. pylori infected or eradicated participants from negatives with an area under the curve of 0.976 (95% CI: 0.965-0.987) (Model 1). Seropositivity for vacuolating cytotoxin A (VacA), GroEL, FliD, H. pylori adhesin A (HpaA), and γ-glutamyl transpeptidase (gGT) was associated with a current infection with an area under the curve of 0.870 (95% CI: 0.837-0.903), which may help discriminate currently infected patients from eradicated ones (Model 2). DISCUSSION: The recomLine assay is sensitive and specific in determining H. pylori infection and eradication status and thus represents a valuable tool in the management of H. pylori infection.

[Serological assessment of Helicobacter pylori-specific antibodies and their association with gastric lesions in a high-risk population].

OBJECTIVE: To determine the distributions of six Helicobacter pylori (Hp)-specific antibodies in a high-risk population of gastric cancer (GC) and explore the relationship between Hp virulence factors and precancerous gastric lesions. METHODS: Based on the two intervention trials conducted in Linqu County, the seropositivities for CagA, VacA, GroEL, UreA, HcpC and GGT were assessed by recombinant immunoassay (recomLine) in 623 participants with H. pylori infection determined by (13)C-urea breath test ((13)C-UBT) and/or enzyme linked immunosorbent assay (ELISA). RESULTS: In a total of 623 participants were detected by recomLine analysis, of which 594 were Hp-positive. The seropositivities rates of CagA, VacA, GroEL, UreA, HcpC and GGT were 84.0%, 38.2%, 66.7%, 17.7%, 58.8% and 42.8%, respectively. A total of 523 participants were determined as type I infection of Hp, accounting for 88.1%. Compared with superficial gastritis (SG), the infection rate of Hp type I was higher in the chronic atrophic gastritis (CAG) (P = 0.001). CONCLUSIONS: The results of this population-based study suggest that the virulence factors of Hp may be related to the development of GC in a Chinese high-risk population. The recomLine analysis may serve as a tool for identification of Hp strains and prediction of high-risk population of GC.

Allium vegetable intake associated with the risk of incident gastric cancer: a continuous follow-up study of a randomized intervention trial.

BACKGROUND: Allium vegetable components have antibacterial, antioxidative, and immune modulation properties, thus potentially exhibiting antitumor effects. Despite evidence from case-control studies, prospective studies linking allium vegetables with gastric cancer (GC) have been sparse. OBJECTIVE: In a prospective study, we examined whether allium vegetable intake would change the risk of GC occurrence and whether the associations would be modified by vitamin supplementation, garlic supplementation, and Helicobacter pylori (H. pylori) treatment. METHODS: The study was conducted on the basis of the Shandong Intervention Trial, a randomized, placebo-controlled, factorial-designed trial (1995-2003) in a well-recognized high-risk area for GC in China. Participants were continuously followed up to December 2017 for 22.3 y (1995-2017). A total of 3229 subjects were included, with information on the intake of allium vegetables (garlic vegetables and scallions), collected by structured questionnaires in 1994. The associations of total and individual allium vegetable intake with the risk of GC were examined, respectively. RESULTS: During the follow-up, 144 incident cases of GC were identified. Garlic vegetable intake was associated with a decreased risk of incident GC (P-trend = 0.02; OR: 0.83; 95% CI: 0.70, 0.98, per 1 kg/y increment), whereas scallion intake showed no association (P-trend = 0.80). An inverse association of the risk of GC with total allium vegetable and garlic vegetable intake was particularly stronger among those receiving the placebo for vitamin supplementation or garlic supplementation, indicating potential effect modifications by nutritional supplementation on allium vegetable intake and the risk of developing GC. Similar findings were found for analyses of the combined prevalence of dysplasia or GC. CONCLUSIONS: We found a significant reduction in the risk of developing GC with increasing dietary intake of allium vegetables, particularly garlic vegetables. The findings add to the literature on the potential inverse association of garlic vegetable intake with the risk of GC, therefore holding public health implications for dietary recommendations. This trial was registered at clinicaltrials.gov as NCT00339768.

Metabolite Alterations and Interactions with Microbiota in Helicobacter pylori-Associated Gastric Lesions.

Metabolites and their interactions with microbiota may be involved in Helicobacter pylori-associated gastric lesion development. This study aimed to explore metabolite alterations upon H. pylori eradication and possible roles of microbiota-metabolite interactions in progression of precancerous lesions. Targeted metabolomics assays and 16S rRNA gene sequencing were conducted to investigate metabolic and microbial alterations of paired gastric biopsy specimens in 58 subjects with successful and 57 subjects with failed anti-H. pylori treatment. Integrative analyses were performed by combining the metabolomics and microbiome profiles from the same intervention participants. A total of 81 metabolites were significantly altered after successful eradication compared to failed treatment, including acylcarnitines, ceramides, triacylglycerol, cholesterol esters, fatty acid, sphingolipids, glycerophospholipids, and glycosylceramides, with P values of <0.05 for all. The differential metabolites showed significant correlations with microbiota in baseline biopsy specimens, such as negative correlations between Helicobacter and glycerophospholipids, glycosylceramide, and triacylglycerol (P < 0.05 for all), which were altered by eradication. The characteristic negative correlations between glycosylceramides and Fusobacterium, Streptococcus, and Gemella in H. pylori-positive baseline biopsy specimens were further noticed in active gastritis and intestinal metaplasia (P < 0.05 for all). A panel including differential metabolites, genera, and their interactions may help to discriminate high-risk subjects who progressed from mild to advanced precancerous lesions in short-term and long-term follow-up periods with areas under the curve (AUC) of 0.914 and 0.801, respectively. Therefore, our findings provide new insights into the metabolites and microbiota interactions in H. pylori-associated gastric lesion progression. IMPORTANCE In this study, a panel was established including differential metabolites, genera, and their interactions, which may help to discriminate high-risk subjects for progression from mild lesions to advanced precancerous lesions in short-term and long-term follow-up.

Plasma lipids signify the progression of precancerous gastric lesions to gastric cancer: a prospective targeted lipidomics study.

Rationale: Gastric cancer (GC) is preceded by a stepwise progression of precancerous gastric lesions. Distinguishing individuals with precancerous gastric lesions that have progression potential to GC is an important need. Perturbated lipid metabolism, particularly the dysregulation of de novo lipogenesis, is involved in gastric carcinogenesis. We conducted the first prospective lipidomics study exploring lipidomic signatures for the risk of gastric lesion progression and early GC. Methods: Our two-stage study of targeted lipidomics enrolled 400 subjects from the National Upper Gastrointestinal Cancer Early Detection Program in China, including 200 subjects of GC and different gastric lesions in the discovery and validation stages. Of validation stage, 152 cases with gastric lesions were prospectively followed for the progression of gastric lesions for a median follow-up of 580 days (interquartile range 390-806 days). We examined the lipidomic signatures associated with the risk of advanced gastric lesions and their progression to GC. Our published tissue proteomic data were referred to further investigate highlighted lipids with their biologically related protein expression in gastric mucosa. Results: We identified 11 plasma lipids significantly inversely associated with the risk of gastric lesion progression and GC occurrence. These lipids were integrated as latent profiles to identify 5 clusters of lipid expression that had distinct risk of gastric lesion progression. The latent profiles significantly improved the ability to predict the progression potential of gastric lesions (AUC: 0.82 vs 0.68, Delong's P = 4.6×10-4) and risk of early GC (AUC: 0.81 vs 0.55, P = 6.3×10-5). Significant associations were found between highlighted lipids, their biologically correlated proteins and the risk of GC, supporting the role of the pathways involving monocarboxylic acid metabolism and lipid transport and catabolic process in GC. Conclusions: Our study revealed the lipidomic signatures associated with the risk of gastric lesion progression and GC occurrence, exhibiting translational implications for GC prevention.

Urine proteomic signatures predicting the progression from premalignancy to malignant gastric cancer.

BACKGROUND: Early detection of gastric cancer (GC) remains challenging. We aimed to examine urine proteomic signatures and identify protein biomarkers that predict the progression of gastric lesions and risk of GC. METHODS: A case-control study was initially designed, covering subjects with GC and gastric lesions of different stages. Subjects were aged 40-69 years, without prior diagnosis of renal or urological diseases. We enrolled a total of 255 subjects, with 123 in the discovery stage from Linqu, China, a high-risk area for GC and 132 in the validation stage from Linqu and Beijing. A prospective study was further designed for a subset of 60 subjects with gastric lesions, which were followed for 297-857 days. FINDINGS: We identified 43 differentially expressed urine proteins in subjects with GC vs. mild or advanced gastric lesions. Baseline urinary levels of ANXA11, CDC42, NAPA and SLC25A4 were further positively associated with risk of gastric lesion progression. Three of them, except for SLC25A4, also had higher expression in GC than non-GC tissues. Integrating these four proteins showed outstanding performance in predicting the progression of gastric lesions (AUC (95% CI): 0.92 (0.83-1.00)) and risk of GC (AUC (95% CI): 0.81 (0.73-0.89) and 0.84 (0.77-0.92) for GC vs. mild or advanced gastric lesions respectively). INTERPRETATION: This study revealed distinct urine proteomic profiles and a panel of proteins that may predict the progression of gastric lesions and risk of GC. These biomarkers in a non-invasive approach may have translational significance for defining high-risk populations of GC and its early detection. FUNDING: Funders are listed in the Acknowledgement.

Identification and Validation of Plasma Metabolomic Signatures in Precancerous Gastric Lesions That Progress to Cancer.

Importance: Metabolic deregulation plays an important role in gastric cancer (GC) development. To date, no studies have comprehensively explored the metabolomic profiles along the cascade of gastric lesions toward GC. Objective: To draw a metabolic landscape and define metabolomic signatures associated with the progression of gastric lesions and risk of early GC. Design, Setting, and Participants: A 2-stage, population-based cohort study was initiated in 2017 in Linqu County, Shandong Province, China, a high-risk area for GC. Prospective follow-up was conducted during the validation stage (June 20, 2017, to May 27, 2020). A total of 400 individuals were included based on the National Upper Gastrointestinal Cancer Early Detection Program in China. The discovery stage involved 200 individuals with different gastric lesions or GC (high-grade intraepithelial neoplasia or invasive GC). The validation stage prospectively enrolled 152 individuals with gastric lesions who were followed up for 118 to 1063 days and 48 individuals with GC. Exposures: Metabolomic profiles and metabolite signatures were examined based on untargeted plasma metabolomics assay. Main Outcomes and Measures: The risk of GC overall and early GC (high-grade intraepithelial neoplasia), and progression of gastric lesions. Results: Of the 400 participants, 124 of 200 (62.0%) in the discovery set were men; mean (SD) age was 56.8 (7.5) years. In the validation set, 136 of 200 (68.0%) were men; mean (SD) age was 57.5 (8.1) years. Distinct metabolomic profiles were noted for gastric lesions and GC. Six metabolites, including α-linolenic acid, linoleic acid, palmitic acid, arachidonic acid, sn-1 lysophosphatidylcholine (LysoPC)(18:3), and sn-2 LysoPC(20:3) were significantly inversely associated with risk of GC overall and early GC (high-grade intraepithelial neoplasia). Among these metabolites, the first 3 were significantly inversely associated with gastric lesion progression, especially for the progression of intestinal metaplasia (α-linolenic acid: OR, 0.42; 95% CI, 0.18-0.98; linoleic acid: OR, 0.43; 95% CI, 0.19-1.00; and palmitic acid: OR, 0.32; 95% CI, 0.13-0.78). Compared with models including only age, sex, Helicobacter pylori infection, and gastric histopathologic findings, integrating these metabolites significantly improved the performance for predicting the progression of gastric lesions (area under the curve [AUC], 0.86; 95% CI, 0.70-1.00 vs AUC, 0.69; 95% CI, 0.50-0.88; P = .02) and risk of early GC (AUC, 0.83; 95% CI, 0.58-1.00 vs AUC, 0.61; 95% CI, 0.31-0.91; P = .03). Conclusions and Relevance: This study defined metabolite signatures that might serve as meaningful biomarkers for assessing high-risk populations and early diagnosis of GC, possibly advancing targeted GC prevention and control.

Helicobacter pylori associated aberrant methylation genes in blood leukocyte and gastric mucosa.

Background and Aim: Methylation alterations may be involved in Helicobacter pylori-associated gastric carcinogenesis. This study aims to explore the potential H.pylori-associated methylation biomarkers in blood leukocyte and gastric mucosa. Methods: Five candidate H.pylori-associated aberrant methylation genes were selected from the previous genome-wide profiling panels and validated in blood leukocyte and gastric mucosa in multi-stages (case-control validation between H.pylori positive and negative subjects and self-control validation before and after anti-H.pylori treatment). Results: GNAS methylation level was decreased in blood leukocyte (62.07% v.s. 46.33%, p<0.001) and gastric mucosa (56.30% v.s. 32.42%, p<0.001) of H.pylori positive subjects compared to negative controls. While, MTERF1 methylation level was increased significantly in blood leukocyte (29.57% v.s. 56.02%, p<0.001) and gastric mucosa (31.10% v.s. 47.50%, p<0.001) of positive subjects compared to controls. After successful H.pylori eradication, the methylation levels were increased from 44.87% to 60.88% (p<0.001) for GNAS and decreased from 46.19% to 34.56% (p<0.001) for MTERF1 in blood leukocyte. Similar increasing and decreasing methylation alterations were also found for the two genes after successful eradication in paired gastric mucosa. In TCGA database, an inverse relationship was found between GNAS methylation and mRNA expression (r=-0.12, p=0.027). The GC cases with higher GNAS expression levels showed significantly worse survival (HR, 2.09, 95%CI, 1.22-3.57, p=0.007) compared to lower expression subjects. Conclusions: GNAS and MTERF1 methylation levels may be affected by H.pylori infection in gastric mucosa and blood leukocyte. GNAS may be involved in advanced stage of GC development, although the possible mechanism still needs further study in precancerous lesions.

Microbiota alteration at different stages in gastric lesion progression: a population-based study in Linqu, China.

In addition to Helicobacter pylori (H.pylori), gastric microbiota may be involved in carcinogenesis process. However, the longitudinal study to assess changes in the gastric microbiota associated with the development of gastric carcinogenesis is still limited. The aim of this study is to explore dynamic microbial alterations in gastric cancer (GC) development based on a 4-year endoscopic follow-up cohort in Linqu County, China. Microbial alterations were investigated by deep sequencing of the microbial 16S ribosomal RNA gene in 179 subjects with various gastric lesions, and validated in paired gastric biopsies prospectively collected before and after lesion progression and in non-progression controls. Significant differences were found in microbial diversity and community structure across various gastric lesions, with 62 candidate differential taxa between at least two lesion groups. Further validations identified Helicobacter, Bacillus, Capnocytophaga and Prevotella to be associated with lesion progression-to-dysplasia (DYS)/GC (all P < 0.05), especially for subjects progressing from intestinal metaplasia (IM) to DYS/GC. The combination of the four genera in a microbial dysbiosis index showed a significant difference after lesion progression-to-DYS/GC compared to controls (P = 0.027). The panel including the four genera identified subjects after progression-to-DYS/GC with an area under the receiver-operating curve (AUC) of 0.941. Predictive significance was found before lesion progression-to-DYS/GC with an AUC = 0.776 and an even better AUC (0.927) for subjects progressing from IM to DYS/GC. Microbiota may play different roles at different stages in gastric carcinogenesis. A panel of bacterial genera associated with gastric lesions may help to assess gastric microbial dysbiosis and show potential predictive values for lesion progression. Our findings provide new clues for the microbial mechanism of H.pylori-associated carcinogenesis.

Proteomic profiling identifies signatures associated with progression of precancerous gastric lesions and risk of early gastric cancer.

BACKGROUND: Molecular features underlining the multistage progression of gastric lesions and development of early gastric cancer (GC) are poorly understood, restricting the ability to GC prevention and management. METHODS: We portrayed proteomic landscape and explored proteomic signatures associated with progression of gastric lesions and risk of early GC. Tissue proteomic profiling was conducted for a total of 324 subjects. A case-control study was performed in the discovery stage (n=169) based on populations from Linqu, a known high-risk area for GC in China. We then conducted two-stage validation, including a cohort study from Linqu (n = 56), with prospective follow-up for progression of gastric lesions (280-473 days), and an independent case-control study from Beijing (n = 99). FINDINGS: There was a clear distinction in proteomic features for precancerous gastric lesions and GC. We derived four molecular subtypes of gastric lesions and identified subtype-S4 with the highest progression risk. We found 104 positively-associated and 113 inversely-associated proteins for early GC, with APOA1BP, PGC, HPX and DDT associated with the risk of gastric lesion progression. Integrating these proteomic signatures, the ability to predict progression of gastric lesions was significantly strengthened (areas-under-the-curve=0.88 (95%CI: 0.78-0.99) vs. 0.56 (0.36-0.76), Delong's P = 0.002). Immunohistochemistry assays and examination at mRNA level validated the findings for four proteins. INTERPRETATION: We defined proteomic signatures for progression of gastric lesions and risk of early GC, which may have translational significance for identifying particularly high-risk population and detecting GC at an early stage, improving potential for targeted GC prevention. FUNDING: The funders are listed in the Acknowledgement.

[Determination of serum copper and zinc in different chemical forms by graphite furnace atomic absorption spectrometry with ethanol-EDTA precipitation method].

A simple and reliable method was developed for the determination of serum copper and zinc in different chemical forms by graphite furnace atomic absorption spectrometry (GFAAS) with ethanol-EDTA precipitation. The serum and ethanol were mixed with volume ratio 1 : 2. The mixture was incubated at 70 degrees C and ultra-centrifuged to precipitate proteins. Zinc and copper can be released from albumin after EDTA treating. Thus, macroglobulin-zinc, ceruloplasmin-copper, and albumin-bound zinc or copper could be determined by two proposed precipitation steps. The determination limit of copper (3sigma) was 1.2 microg x L(-1) and the recovery was 92.3%-104%, while the determination limit of zinc (3sigma) was 0.098 microg x L(-1), and the recovery was 90%-107%. This two-step precipitation method can be used to determine serum zinc and copper in various chemical forms in tumor, Wilson patients and heathy human.

Association Between Lifestyle Factors, Vitamin and Garlic Supplementation, and Gastric Cancer Outcomes: A Secondary Analysis of a Randomized Clinical Trial.

Importance: The associations of lifestyle factors with gastric cancer (GC) are still underexplored in populations in China. Long-term nutritional supplementation may prevent GC in high-risk populations, but the possible effect modification by lifestyle factors remains unknown. Objective: To evaluate how lifestyle factors, including smoking, alcohol intake, and diet, may change the risk of GC incidence and mortality and whether the effects of vitamin and garlic supplementation on GC are associated with major lifestyle factors. Design, Setting, and Participants: This is a secondary analysis of the Shandong Intervention Trial, a masked, randomized, placebo-controlled trial that aimed to assess the effect of vitamin and garlic supplementations and Helicobacter pylori treatment on GC in a factorial design with 22.3 years of follow-up. The study took place in Linqu County, Shandong province, China, a high-risk area for GC. Data were collected from Jully 1995 to December 2017. Overall, 3365 participants aged 35 to 64 years identified in 13 randomly selected villages who agreed to undergo gastroscopy were invited to participate in the trial and were included in the analysis. Data analysis was conducted from March to May 2019. Interventions: Participants received vitamin and garlic supplementation for 7.3 years, H pylori treatment for 2 weeks (among participants with H pylori ), or placebo. Main Outcomes and Measures: The primary outcomes were GC incidence and GC mortality (1995-2017). We also examined the progression of gastric lesions (1995-2003) as a secondary outcome. Results: Of the 3365 participants (mean [SD] age, 47.1 [9.2] years; 1639 [48.7%] women), 1677 (49.8%) were randomized to receive active vitamin supplementation, with 1688 (50.2%) receiving placebo, and 1678 (49.9%) receiving active garlic supplementation, with 1687 (50.1%) receiving placebo. Overall, 151 GC cases (4.5%) and 94 GC deaths (2.8%) were identified. Smoking was associated with increased risk of GC incidence (odds ratio, 1.72; 95% CI, 1.003-2.93) and mortality (hazard ratio [HR], 2.01; 95% CI, 1.01-3.98). Smoking was not associated with changes to the effects of vitamin or garlic supplementation. The protective effect on GC mortality associated with garlic supplementation was observed only among those not drinking alcohol (never drank alcohol: HR, 0.33; 95% CI, 0.15-0.75; ever drank alcohol: HR, 0.92; 95% CI, 0.55-1.54; P for interaction = .03), and significant interactions were only seen among participants with H pylori (never drank alcohol: HR, 0.31; 95% CI, 0.12-0.78; ever drank alcohol: HR, 0.91; 95% CI, 0.52-1.60; P for interaction = .04). No significant interactions between vitamin supplementation and lifestyle factors were found. Conclusions and Relevance: In this secondary analysis of a randomized clinical trial, smoking was associated with an increased risk of GC incidence and mortality. Not drinking alcohol was associated with a stronger beneficial effect of garlic supplementation on GC prevention. Our findings provide new insights into lifestyle intervention for GC prevention, suggesting that mass GC prevention strategies may need to be tailored to specific population subgroups to maximize the potential beneficial effect. Trial Registration: ClinicalTrials.gov Identifier: NCT00339768.