Dynamic genomic changes in methotrexate-resistant human cancer cell lines beyond DHFR amplification suggest potential new targets for preventing drug resistance.

BACKGROUND: Although DHFR gene amplification has long been known as a major mechanism for methotrexate (MTX) resistance in cancer, the early changes and detailed development of the resistance are not yet fully understood. METHODS: We performed genomic, transcriptional and proteomic analyses of human colon cancer cells with sequentially increasing levels of MTX-resistance. RESULTS: The genomic amplification evolved in three phases (pre-amplification, homogenously staining region (HSR) and extrachromosomal DNA (ecDNA)). We confirm that genomic amplification and increased expression of DHFR, with formation of HSRs and especially ecDNAs, is the major driver of resistance. However, DHFR did not play a detectable role in the early phase. In the late phase (ecDNA), increase in FAM151B protein level may also have an important role by decreasing sensitivity to MTX. In addition, although MSH3 and ZFYVE16 may be subject to different posttranscriptional regulations and therefore protein expressions are decreased in ecDNA stages compared to HSR stages, they still play important roles in MTX resistance. CONCLUSION: The study provides a detailed evolutionary trajectory of MTX-resistance and identifies new targets, especially ecDNAs, which could help to prevent drug resistance. It also presents a proof-of-principal approach which could be applied to other cancer drug resistance studies.

TOB1 modulates neutrophil phenotypes to influence gastric cancer progression and immunotherapy efficacy.

Introduction: The ErbB-2.1(TOB1) signaling transducer protein is a tumor-suppressive protein that actively suppresses the malignant phenotype of gastric cancer cells. Yet, TOB1 negatively regulates the activation and growth of different immune cells. Understanding the expression and role of TOB1 in the gastric cancer immune environment is crucial to maximize its potential in targeted immunotherapy. Methods: This study employed multiplex immunofluorescence analysis to precisely delineate and quantify the expression of TOB1 in immune cells within gastric cancer tissue microarrays. Univariate and multivariate Cox analyses were performed to assess the influence of clinical-pathological parameters, immune cells, TOB1, and double-positive cells on the prognosis of gastric cancer patients. Subsequent experiments included co-culture assays of si-TOB1-transfected neutrophils with AGS or HGC-27 cells, along with EdU, invasion, migration assays, and bioinformatics analyses, aimed at elucidating the mechanisms through which TOB1 in neutrophils impacts the prognosis of gastric cancer patients. Results: We remarkably revealed that TOB1 exhibits varying expression levels in both the nucleus (nTOB1) and cytoplasm (cTOB1) of diverse immune cell populations, including CD8+ T cells, CD66b+ neutrophils, FOXP3+ Tregs, CD20+ B cells, CD4+ T cells, and CD68+ macrophages within gastric cancer and paracancerous tissues. Significantly, TOB1 was notably concentrated in CD66b+ neutrophils. Survival analysis showed that a higher density of cTOB1/nTOB1+CD66b+ neutrophils was linked to a better prognosis. Subsequent experiments revealed that, following stimulation with the supernatant of tumor tissue culture, the levels of TOB1 protein and mRNA in neutrophils decreased, accompanied by enhanced apoptosis. HL-60 cells were successfully induced to neutrophil-like cells by DMSO. Neutrophils-like cells with attenuated TOB1 gene expression by si-TOB1 demonstrated heightened apoptosis, consequently fostering a malignant phenotype in AGS and HCG-27 cells upon co-cultivation. The subsequent analysis of the datasets from TCGA and TIMER2 revealed that patients with high levels of TOB1 combined neutrophils showed better immunotherapy response. Discussion: This study significantly advances our comprehension of TOB1's role within the immune microenvironment of gastric cancer, offering promising therapeutic targets for immunotherapy in this context.

Contribution of PGAP3 co-amplified and co-overexpressed with ERBB2 at 17q12 involved poor prognosis in gastric cancer.

The locus at 17q12 erb-b2 receptor tyrosine kinase 2 (ERBB2) has been heavily amplificated and overexpressed in gastric cancer (GC), but it remains to be elucidated about the clinical significance of the co-amplification and co-overexpression of PGAP3 gene located around ERBB2 in GC. The profile of PGAP3 and ERBB2 in four GC cell lines and tissue microarrays containing 418 primary GC tissues was assessed to investigate the co-overexpression and clinical significance of the co-amplified genes, and to evaluate the impact of the co-amplified genes on the malignancy of GC. Co-amplification of PGAP3 and ERBB2 accompanied with co-overexpression was observed in a haploid chromosome 17 of NCI-N87 cells with double minutes (DMs). PGAP3 and ERBB2 were overexpressed and positively correlated in 418 GC patients. Co-overexpression of the PGAP3 and ERBB2 was correlated with T stage, TNM stage, tumour size, intestinal histological type and poor survival proportion in 141 GC patients. In vitro, knockdown of the endogenous PGAP3 or ERBB2 decreased cell proliferation and invasion, increased G1 phase accumulation and induced apoptosis in NCI-N87 cells. Furthermore, combined silencing of PGAP3 and ERBB2 showed an additive effect on resisting proliferation of NCI-N87 cells compared with targeting ERBB2 or PGAP3 alone. Taken together, the co-overexpression of PGAP3 and ERBB2 may be crucial due to its significant correlation with clinicopathological factors of GC. Haploid gain of PGAP3 co-amplified with ERBB2 is sufficient to facilitate the malignancy and progression of GC cells in a synergistic way.

Multidimensional difference analysis in gastric cancer patients between high and low latitude.

Genetic variation has been shown to affect tumor growth and progression, and the temperature at different latitudes may promote the evolution of genetic variation. Geographical data with latitudinal information is of importance to understand the interplay between genetic variants and environmental influence, such as the temperature, in gastric cancer (GC). In this study, we classified the GC samples from The Cancer Genome Atlas database into two groups based on the latitudinal information of patients and found that GC samples with low-latitude had better clinical outcomes. Further analyses revealed significant differences in other clinical factors such as disease stage and grade between high and low latitudes GC samples. Then, we analyzed the genomic and transcriptomic differences between the two groups. Furthermore, we evaluated the activity score of metabolic pathways and infiltrating immune cells in GC samples with different latitudes using the single-sample gene set enrichment analysis algorithm. These results showed that GC samples at low-latitude had lower tumor mutation burden and subclones as well as higher DNA repair activities. Meanwhile, we found that most immune cells were associated with the prognosis of low-latitude GC patients. At last, we constructed and validated an immune-related prognostic model to evaluate the prognosis of GC samples at different latitudes. This study has provided a further understanding of the geographical contribution to GC at the multiomic level and may benefit the individualized treatment of GC patients at different latitudes.

Genetic Diversity Analysis of the Chinese Daur Ethnic Group in Heilongjiang Province by Complete Mitochondrial Genome Sequencing.

Mitochondrial DNA (mtDNA) has the characteristics of maternal inheritance, high mutation rate, high copy number, and no recombination. As the most powerful tool for studying the origin and evolution of modern humans, mtDNA has great significance in the research of population genetics and evolutionary genetics. Here, we provide new insights into the maternal genetic history of the Daur ethnic group by generating complete mitochondrial genomes from a total of 146 Daur individuals in China. We also collected the published complete mitochondrial genome sequences of 5,094 individuals from 56 worldwide populations as reference data to further explore the matrilineal genetic landscape of the Daur ethnic group. First, the haplotype diversity was 0.9943 ± 0.0019 and nucleotide diversity was 0.0428 ± 0.0210. The neutrality tests of the Daur group showed significant negative values and the mismatch distribution curve was obviously distributed in a unimodal pattern. The results showed that the Daur ethnic group has high genetic diversity and may have experienced recent population expansion. In addition, the main haplogroups of the Daur population were haplogroup D (31.51%), M* (20.55%), C (10.28%), F (7.53%), and B (6.85%), all of which were prevalent in northern China. It probably implies the northern Chinese origin of the Daur population. The PCA, F ST, and phylogenetic analysis results indicated that the Daur group formed a cluster with East Asian populations, and had few genetic differences with the populations in northern China. More importantly, we found that disease-related mutation sites of the mitochondrial genome may be related to ethnic groups, which may have important implications for the prevention and occurrence of specific diseases. Overall, this study revealed the complexity and diversity of the matrilineal genetic background of the Daur ethnic group. Meanwhile, it provided meaningful data for the research on the diversity of the human genome.

Differential Infiltration of Immune Cells Driven by Tumor Heterogeneity Reveals Two Immune Subtypes in Lung Adenocarcinoma.

Intra-tumoral heterogeneity (ITH) is a critical factor leading to aggressive progression and response to immunotherapy in lung adenocarcinoma (LUAD). However, the relationship between ITH and immune cells in the tumor microenvironment (TME) has not been systematically elucidated. In the present study, we evaluated the ITH status of LUAD samples based on the mutational data obtained from The Cancer Genome Atlas database. First, we identified five key immune pathways with a significantly continuous downtrend among normal, low-heterogeneous, and high-heterogeneous samples and further excavated nine key immune cells related to the key immune pathways and tumor heterogeneity. Then, two immune subtypes were defined by a consensus clustering algorithm based on the infiltration of these immune cells. Differences between these two immune subtypes were remarkable, including alterations of tumor mutation burden and DNA copy number variation at the genomic level, various metabolic pathways, and the different clinical outcome, which was also validated in two independent Gene Expression Omnibus datasets. The results revealed that ITH was significantly associated with prognosis and infiltrating immune cells in the TME. Our study provides novel insights in understanding the relationship between ITH and immune cells and contributes to the immunotherapy of LUAD patients.

Analysis of DNA Repair-Related Prognostic Function and Mechanism in Gastric Cancer.

DNA repair mechanisms have been proven to be essential for cells, and abnormalities in DNA repair could cause various diseases, such as cancer. However, the diversity and complexity of DNA repair mechanisms obscure the functions of DNA repair in cancers. In addition, the relationships between DNA repair, the tumor mutational burden (TMB), and immune infiltration are still ambiguous. In the present study, we evaluated the prognostic values of various types of DNA repair mechanisms and found that double-strand break repair through single-strand annealing (SSA) and nonhomologous end-joining (NHEJ) was the most prognostic DNA repair processes in gastric cancer (GC) patients. Based on the activity of these two approaches and expression profiles, we constructed a HR-LR model, which could accurately divide patients into high-risk and low-risk groups with different probabilities of survival and recurrence. Similarly, we also constructed a cancer-normal model to estimate whether an individual had GC or normal health status. The prognostic value of the HR-LR model and the accuracy of the cancer-normal model were validated in several independent datasets. Notably, low-risk samples, which had higher SSA and NHEJ activities, had more somatic mutations and less immune infiltration. Furthermore, the analysis found that low-risk samples had higher and lower methylation levels in CpG islands (CGIs) and open sea regions respectively, and had higher expression levels of programmed death-ligand 1 (PD-L1) and lower methylation levels in the promoter of the gene encoding PD-L1. Moreover, low-risk samples were characterized primarily by higher levels of CD4+ memory T cells, CD8+ naive T cells, and CD8+ TEM cells than those in high-risk samples. Finally, we proposed a decision tree and nomogram to help predict the clinical outcome of an individual. These results provide an improved understanding of the complexity of DNA repair, the TMB, and immune infiltration in GC, and present an accurate prognostic model for use in GC patients.

The Overexpression of TOB1 Induces Autophagy in Gastric Cancer Cells by Secreting Exosomes.

We previously confirmed that transducer of ERBB2, 1 (TOB1) gene, can induce autophagy in gastric cancer cells. Studies have shown the biogenesis of exosomes overlaps with different autophagy processes, which helps to maintain the self-renewal and homeostasis of body cells. This study is aimed at verifying whether overexpressing TOB1 induces autophagy by secreting exosomes in gastric cancer cells and its underlying mechanisms. Differential ultracentrifugation was used to extracted the exosomes from the culture medium of gastric cancer cell line AGS-TOB1 ectopically overexpressing TOB1 (exo-AGS-TOB1, experimental group) and AGS-empty-vector cell line with low expression of endogenous TOB1 (exo-AGS-Vector, control group). Exosomal markers CD9 and TSG101 were determined in both the cell supernatants of exo-AGS-TOB1 and exo-AGS-Vector by Western blot. Under the transmission electron microscope (TEM), the exosomes were round and saucer-like vesicles with double-layer membrane structure, and the vesicles showed different translucency due to different contents. The peak size of exosomes detected by nanoparticle tracking analysis (NTA) was about 100 nm. When the exosomes of exo-AGS-TOB1 and exo-AGS-Vector were cocultured with TOB1 knockdown gastric cancer cell line HGC-27-TOB1-6E12 for 48 hours, the conversion of autophagy-related protein LC3-I to LC3-II in HGC-27-TOB1-6E12 gastric cancer cells cocultured with exo-AGS-TOB1 was significantly higher than that in the control group, and the ratio of LC3-II/LC3-I was statistically different (P < 0.05). More autophagosomes in HGC-27-TOB1-6E12 cells cocultured with exo-AGS-TOB1 for 48 hours were observed under TEM, while fewer autophagosomes were found in the control group. Lastly, miRNAs were differentially expressed by cell supernatant-exosomal whole transcriptome sequencing. Thus, our results provide new insights into TOB1-induced autophagy in gastric cancer.

The promoter methylation drives down-regulation mode of HIC1 in gastric cancer, its molecular characteristics and downstream functional pathways.

Gastric cancer is a common malignant tumor of the gastrointestinal tract with a high incidence and mortality rate. Previous results have suggested that the HIC1 gene might be a tumor suppressor candidate in gastric cancer. However, several critical points need to be elucidated: (1) The correlation of HIC1 promoter methylation with its specific expression level in gastric cancer; (2) The molecular characterization of HIC1 promoter methylation; (3) The possible mechanism by which HIC1 performs its inhibitory role in gastric cancer. To address these questions, we retrieved data from TCGA database to analyze HIC1 promoter methylation levels and transcript expression data, and performed targeted region bisulfite sequencing on three stable HIC1 down-regulated cell lines and normal control cell lines, and performed whole transcriptome and metabolite assays in HIC1 knockout cell lines by CRISPR-Cas9 technique. Results demonstrated that HIC1 promoter hypermethylation might be a crucial driving force leading to its down-regulation in HIC1 expression in gastric cancer. This implicated that promoter CG methylation of HIC1 might play a major role in the development of gastric carcinogenesis. Besides, HIC1 may suppress gastric cancer progression by maintaining the normal cellular metabolism, and inhibiting the mTOR signaling pathway activity.

Involvement of TOB1 on autophagy in gastric cancer AGS cells via decreasing the activation of AKT/mTOR signaling pathway.

BACKGROUND: We previously identified the tumor suppressor gene TOB1 as related to gastric cancer. The purpose of this study was to explore whether TOB1 induces autophagy through the AKT/mTOR signaling pathway in gastric cancer. METHODS: Western blotting was used to detect the protein levels of TOB1, LC3, AKT, mTOR, phosphorylated (p) AKT, and p-mTOR. A double fluorescent GFP-RFP-LC3 fusion protein was used to trace autophagy by laser confocal microscopy. Autophagosomes were observed by transmission electron microscopy. RESULTS: The conversion of LC3-I to LC3-II and the LC3-II/LC3-I ratio were significantly increased in AGS cells overexpressing TOB1 compared with control cells. Fluorescence imaging showed LC3 puncta at 48 h, and these puncta increased significantly at 72 h after TOB1 transfection compared with control tumor cells. The presence of autophagosomes in AGS cells was observed at 72 h after TOB1 transfection by transmission electron microscopy, and no autophagosomes were found in the control cells. Moreover, the levels of p-AKT and p -mTOR were lower in AGS cells than in control cancer cells. CONCLUSION: Our results provide novel insight that TOB1 might suppress gastric cancer by inducing autophagy, possibly through decreasing phosphorylation and the subsequent activation of the AKT/mTOR signaling pathway.

The functional variant in promoter of OVCA1 was associated with the risk of gastric cancer in the northeast Chinese Han population.

We previously found allelic deletions on chromosomes 17 in primary gastric cancers (GC) using microsatellite markers for loss of heterozygosity (LOH). OVCA1 lies in one of these regions (17q21.33). The association between single nucleotide polymorphism (SNP) of OVCA1 gene and risk of gastric cancer is not yet clear. In this study, the peripheral blood of 505 gastric cancer patients and 544 healthy controls were genotyped for six SNPs (rs2273981, rs1131600, rs3752963, rs3803806, rs2236375, and rs1051322) of OVCA1, to evaluate the association of these SNPs with the risk of gastric cancer in the Han population in northeast China. The effect of rs2273981 located in the promoter region of OVCA1 on the transcription activity was determined using dual luciferase reporter assay. We found that the association between the AA + AG genotype of rs2273981 and the risk of gastric cancer was significant in smokers (AA + AG vs. GG, OR = 2.47, 95% CI = 1.04 - 5.87, P < 0.05). Stratified analysis of the clinicopathological parameters revealed that rs1131600 AG + GG genotype were significantly associated with increased gastric tumor volume (AG + GG vs. AA, OR = 1.81, 95% CI = 1.00 - 3.29, P < 0.05). The rs2236375 CT + TT genotype was also significantly associated with increased gastric tumor volume (CT + TT vs. CC, OR = 2.65, 95% CI = 1.38 - 5.10, P < 0.05). Additionally, by interacting with the transcription factor AP2A, the GG genotype the rs2273981 increased the transcription activity of OVCA1 compared with AA genotype, thus involved in gastric cancer development.

Genetic Polymorphism of Drug Metabolic Gene CYPs, VKORC1, NAT2, DPYD and CHST3 of Five Ethnic Minorities in Heilongjiang Province, Northeast China.

INTRODUCTION: Genetic variability in genes encoding drug-metabolizing enzymes may contribute to the heterogeneity of drug responses in different populations. Extensive research in pharmacogenomics in major populations around the world provides us with a great deal of information about drug-related genetic polymorphisms. OBJECTIVE: The purpose of this study was to detect the genetic variation of drug-metabolism-related genes in the five ethnic minorities Daur, Hezhen, Ewenki, Mongolian and Manchu in China, and to analyze the distribution differences among ethnic groups. METHODS: We genotyped 32 SNPs of drug metabolism genes in 882 healthy Chinese volunteers from five ethnic groups. The genotype frequency and allele frequency of the five ethnic groups were calculated, and the different variants among the five ethnic groups were compared by chi-square test. Genetic parameters were analyzed using Popgene software. The genetic structure of five ethnic minorities was analyzed by principal component analysis, and compared with 26 populations. RESULTS: We found that SNPs of genes related to drug metabolism existed diversity in different populations. Among them, rs8192766 and rs9419082 in CYP2E1 showed statistical differences between Daur and Manchu, and NAT2 rs1801280 showed statistical differences between Hezhen and Mongolian. In addition, the five populations we studied had the smallest differences with EAS populations. There was haplotype diversity in CHST3, VKORC1, CYP1A2 and CYP2E1 genes in the five ethnic minorities, and these haplotype polymorphisms were related to the use of corresponding drug doses. Cluster analysis shows that the five ethnic minorities in Heilongjiang Province are clustered together with the EAS populations. CONCLUSION: These results suggest that understanding the diversity of drug-related genetic markers is critical for individualized drug gene therapy programs in ethnic minorities in China as well as in populations highly mixed with these ethnic groups.

The rs9953490 polymorphism of DAL-1 gene is associated with gastric cancer risk in the Han population in Northeast China.

BACKGROUND: DAL-1 gene was reported to inhibit proliferation, migration, invasion, and epithelial to mesenchymal transition (EMT) of gastric cancer (GC) cells in our previous study. The association between the genomic variants in DAL-1 gene with risk of GC is still unclear. METHODS: In this study, 505 GC cases and 544 healthy controls (HCs) were collected to evaluate the association between six single nucleotide polymorphisms (SNPs) (rs7240736, rs73937194, rs3817466, rs8082898, rs73381527, rs9953490) of DAL-1 gene and GC risk in the Han population in Northeast China. RESULTS: The TA + AA genotypes of rs9953490 were significantly associated with an increased risk in N3 compared with N0 subgroup (adjusted OR = 4.56, 95% CI = 1.49-13.98, P = 0.008), and also showed evident association with an increased risk in TNM stage III compared with stage I-II (adjusted OR = 2.33, 95% CI = 1.16-4.67, P = 0.017). CONCLUSION: The rs9953490 of DAL-1 gene may play an important role in the occurrence and development of GC in the Han population in Northeast China.

Molecular structure and evolution mechanism of two populations of double minutes in human colorectal cancer cells.

Gene amplification chiefly manifests as homogeneously stained regions (HSRs) or double minutes (DMs) in cytogenetically and extrachromosomal DNA (ecDNA) in molecular genetics. Evidence suggests that gene amplification is becoming a hotspot for cancer research, which may be a new treatment strategy for cancer. DMs usually carry oncogenes or chemoresistant genes that are associated with cancer progression, occurrence and prognosis. Defining the molecular structure of DMs will facilitate understanding of the molecular mechanism of tumorigenesis. In this study, we re-identified the origin and integral sequence of DMs in human colorectal adenocarcinoma cell line NCI-H716 by genetic mapping and sequencing strategy, employing high-resolution array-based comparative genomic hybridization, high-throughput sequencing, multiplex-fluorescence in situ hybridization and chromosome walking techniques. We identified two distinct populations of DMs in NCI-H716, confirming their heterogeneity in cancer cells, and managed to construct their molecular structure, which were not investigated before. Research evidence of amplicons distribution in two different populations of DMs suggested that a multi-step evolutionary model could fit the module of DM genesis better in NCI-H716 cell line. In conclusion, our data implicated that DMs play a very important role in cancer progression and further investigation is necessary to uncover the role of the DMs.

Association between HIC1 promoter methylation and solid tumor: A meta-analysis.

The epigenetic silencing of tumor suppressor genes by promoter methylation plays an increasingly important role in cancer research. A number of studies have reported the contribution of HIC1 promoter methylation towards the occurrence and development of solid tumors, even though HIC1 promoter methylation has also been found in normal and benign tissue samples. We sought to perform a more accurate and comprehensive meta-analysis to assess the association between HIC1 promoter methylation and cancer risk. We searched and retrieved all published studies on HIC1 promoter methylation in PubMed, Google Scholar, Embase, Cochrane Library, and Web of Science databases. After two reviewers checked the studies and extracted the necessary data independently, the meta-analysis was performed using STATA 12.0 software. A total of 14 case-control studies (949 cancer patients, 282 benign, and 371 normal controls) were included in our study. We report a significantly elevated HIC1 promoter methylation in tumor samples compared to normal (OR = 7.02, 95 % CI 3.12-15.78, P < 0.001) and benign controls (OR = 2.69, 95 % CI 1.13-6.42, P = 0.025). Subgroup analysis stratified by ethnicity showed a significantly reduced heterogeneity among North American (I 2 = 0.0 %, P = 0.502) and European (I 2 = 33.7 %, P = 0.183) samples. In addition, heterogeneity was significantly reduced among MSP based detection method (I 2 = 36.4 %, P = 0.139) when samples were stratified based on the methylation detection methods. The overall outcome demonstrated that HIC1 promoter methylation may be involved in the occurrence and development of solid tumors and has the potential to serve as an epigenetic maker in various specific tumors.

Phosphorylation of TOB1 at T172 and S320 is critical for gastric cancer proliferation and progression.

We previously revealed that increased phosphorylation of TOB1, a tumor suppressor protein, may promote the progression of gastric cancer. However, the phosphorylated sites on TOB1 and their functional implication in gastric cancer remain to be clarified. Here, we addressed these questions using the gastric mucosal epithelial cell line GES-1 and three gastric cancer cell lines (HGC-27, AGS, and MKN-1). Compared with the control GES-1 cells, the gastric cancer cells showed decreased levels of TOB1 protein and increased levels of phosphorylated TOB1 (p-TOB1) by Western blotting. Then, TOB1 protein was enriched and purified by immunoprecipitation. Two novel phosphorylation sites at threonine 172 (T172) and serine 320 (S320) in TOB1 were identified in gastric cancer MKN-1 cells using LC-MS/MS. Furthermore, treatment with the serine/threonine kinase inhibitor staurosporine (STS; 2 nmol/L, 8 h) significantly decreased the levels of p-TOB1. As a result, the proliferation, migration, and invasion of gastric cancer cells were diminished, accompanied by an increased proportion of cells in G1 phase and a decreased proportion of cells in G2 phase. Taken together, these findings indicate for the first time that TOB1 is phosphorylated at T172 and S320 in gastric cancer cells, which are sensitive to STS. Downregulation of p-TOB1 levels by STS treatment can weaken the malignant phenotype of gastric cancer cells and block their progression through the cell cycle. Moreover, STS may exert its antiproliferative activity in gastric cancers by restoring TOB1 protein activity.

Decreased expression levels of DAL-1 and TOB1 are associated with clinicopathological features and poor prognosis in gastric cancer.

PURPOSE: We previously demonstrated that the functional inactivation of DAL-1 and TOB1 promotes an aggressive phenotype in gastric cancer cells, but the links between both genes and the survival of patients with gastric cancer are unknown. Here, we investigated the correlations of the expression levels of DAL-1 and TOB1 with the progression of gastric cancer. METHODS: A total of 270 patients who underwent resectable gastrectomy were included. The expression of DAL-1 and TOB1 was detected by immunohistochemistry. RESULTS: Low expression of DAL-1 in cancer tissue was significantly associated with tumor site (p < 0.05), histological grade (p < 0.01), depth of invasion (p < 0.05), lymph node metastasis status (p < 0.05), Lauren classification (p < 0.001), and clinical stage (p < 0.01). A lower level of TOB1 was observed in gastric cancer patients with diffuse type disease compared to patients with either intestinal or mixed type disease (p < 0.001). Additionally, Spearman's correlation analysis revealed that decreased expression of DAL-1 was positively correlated with low TOB1 expression (r=0.304, p < 0.001). The survival analysis showed that low levels of DAL-1 and TOB1 were significantly associated with poor survival of gastric cancer patients (p <0.001 and p < 0.05, respectively). CONCLUSION: The downregulation of DAL-1 and TOB1 expression is associated with shorter survival of gastric cancer patients. Hence, DAL-1 and TOB1 may be considered potential novel markers for predicting the outcomes of patients with gastric cancer.

Prevalence of chromosomal abnormalities identified by copy number variation sequencing in high-risk pregnancies, spontaneous abortions, and suspected genetic disorders.

OBJECTIVE: High-throughput sequencing based on copy number variation (CNV-seq) is commonly used to detect chromosomal abnormalities including aneuploidy. This study provides evidence for the prevalence of chromosomal abnormalities in target populations. METHODS: A total of 160 samples, including 83 high-risk pregnancies, 37 spontaneous abortions, and 40 suspected genetic disorders, were analyzed by CNV-seq. Relationships between the incidence of these chromosomal abnormalities and risk factors (e.g. advanced maternal age, abnormal pregnancy history, and family history of congenital disease) were further analyzed by subgroup. RESULTS: A total of 37 (44.6%) high-risk pregnancies, 25 (67.6%) spontaneous abortions, and 22 (55%) suspected genetic disorders had chromosomal abnormalities including aneuploidy and CNVs. There was an increased risk association between the prevalence of aneuploidy and pathogenic-relevant CNV in the fetus or abortive tissue and advanced maternal age. Moreover, a family history of congenital disease was also positively correlated with fetal chromosomal abnormalities in high-risk pregnancies. CONCLUSION: A relatively high prevalence of chromosomal abnormalities was detected in high-risk pregnancies, spontaneous abortions, and suspected genetic disorders, indicating the importance of CNV detection in such populations.

Identification of a pathogenic mutation in a Chinese pedigree with polycystic kidney disease.

Polycystic kidney disease (PKD) is a life­threatening inherited disease with a morbidity of 1:500­1,000 worldwide. Numerous progressively enlarging cysts are observed in the bilateral kidneys of patients with PKD, inducing structural damage and loss of kidney function. The present study analyzed one family with PKD. Whole exome sequencing of the proband was performed to detect the pathogenic gene present in the family. Candidate gene segments for lineal consanguinity in the family were amplified by nest polymerase chain reaction, followed by Sanger sequencing. One novel duplication variant (NM_001009944.2:c.9359dupA:p.Y3120_E3121delinsX) and one missense mutation (c.G9022A:p.V3008M) were detected in PKD1. Additionally, the pathogenic substitutions in PKD1 published from the dataset were analyzed. Following analysis and confirmation, the duplication variant NM_001009944.2:c.9359dupA:p.Y3120_E3121delinsX in PKD1, within the polycystin­1, lipoxygenase, α­toxin domain, was considered to be the pathogenic factor in the examined family with autosomal dominant PKD. Additionally, based on the analysis of 4,805 pathogenic substitutions in PKD1 within various regions, the presence of the missense mutation in the N­terminal domain of polycystin­1 may present high pathogenicity in ADPKD.

Associations between PTPN2 gene polymorphisms and psoriasis in Northeastern China.

Psoriasis is a chronic immune-mediated disease with a complex etiology involving various genetic and immunological factors as well as environmental factors. Psoriasis is thought to be mediated by T-cells polarized to a Th17 fate. PTPN2 encodes the T-cell protein tyrosine phosphatase, which acts as a negative regulator of the JAK/STAT signaling pathways downstream of cytokines and plays a prominent role in T-cell activation, signaling and/or effector function. To evaluate the association between PTPN2 gene polymorphisms and psoriasis in the Northeastern Chinese population. A case-control study was conducted, and 398 patients with psoriasis and 397 healthy controls were genotyped for thirteen genetic polymorphisms in PTPN2. Allele analysis revealed that rs2847297, rs657555 and rs482160 polymorphisms were significantly associated with psoriasis (p = 0.0018, p = 0.0017 and p = 0.0086, respectively). Genotype analysis also revealed that these polymorphisms were significantly associated with psoriasis under different models (codominant, dominant and recessive models) (p < 0.05). In this study, three haplotypes (H1, H7 and H11) were also found to be associated with psoriasis (p = 0.0015, p = 0.0094, and p = 0.0124, respectively). These results indicate that PTPN2 genetic polymorphisms are associated with psoriasis in the Northeastern Chinese population.