Prognostic correlation between specialized capillary endothelial cells and lung adenocarcinoma.

Background: In-depth analysis of the functional changes occurring in endothelial cells (ECs) involved in capillary formation can help to elucidate the mechanism of tumour vascular growth. Methods: Appropriate datasets were retrieved from the GEO database to obtain single-cell data on LUAD samples and adjacent normal tissue samples. ECs were selected by an automatic annotation program in R and further subdivided based on reported EC marker genes. Functional changes in different types of capillary ECs were then visualized, and the concrete expression was classified by genetic data in the TCGA. Finally, a prognostic model was constructed to predict immunoinfiltration status, survival and drug therapy effects. Results: The LUAD data contained in the GSE183219 dataset were suitable for our analysis. After dimensionality reduction analysis and cell annotation, EC general capillary and EC aerocyte subsets as capillary specialized phenotypes showed a series of functional changes in tumour samples, with a total of 108 genes found to undergo functional changes. Use of CellPhoneDB revealed a close interaction of activity between ECs. After integration of TCGA, GSE68465 and GSE11969 datasets, the genes obtained were analysed by cluster analysis and risk model construction, identifying 8 genes. Drug sensitivity, immune cell and molecular differences can be accurately predicted. Conclusions: EC general capillary and EC aerocyte subsets are recognized capillary ECs in the tumour microenvironment, and the functional changes between them are relevant to the prognosis and treatment of LUAD patients and have the potential to be used in target therapy.

Predicting Differences in Treatment Response and Survival Time of Lung Adenocarcinoma Patients Based on a Prognostic Risk Model of Glycolysis-Related Genes.

Background: Multiple factors influence the survival of patients with lung adenocarcinoma (LUAD). Specifically, the therapeutic outcomes of treatments and the probability of recurrence of the disease differ among patients with the same stage of LUAD. Therefore, effective prognostic predictors need to be identified. Methods: Based on the tumor mutation burden (TMB) data obtained from The Cancer Genome Atlas (TCGA) database, LUAD patients were divided into high and low TMB groups, and differentially expressed glycolysis-related genes between the two groups were screened. The least absolute shrinkage and selection operator (LASSO) and Cox regression were used to obtain a prognostic model. A receiver operating characteristic (ROC) curve and a calibration curve were generated to evaluate the nomogram that was constructed based on clinicopathological characteristics and the risk score. Two data sets (GSE68465 and GSE11969) from the Gene Expression Omnibus (GEO) were used to verify the prognostic performance of the gene. Furthermore, differences in immune cell distribution, immune-related molecules, and drug susceptibility were assessed for their relationship with the risk score. Results: We constructed a 5-gene signature (FKBP4, HMMR, B4GALT1, SLC2A1, STC1) capable of dividing patients into two risk groups. There was a significant difference in overall survival (OS) times between the high-risk group and the low-risk group (p < 0.001), with the low-risk group having a better survival outcome. Through multivariate Cox analysis, the risk score was confirmed to be an independent prognostic factor (HR = 2.709, 95% CI = 1.981-3.705, p < 0.001), and the ROC curve and nomogram exhibited accurate prediction performance. Validation of the data obtained in the GEO database yielded similar results. Furthermore, there were significant differences in sensitivity to immunotherapy, cisplatin, paclitaxel, gemcitabine, docetaxel, gefitinib, and erlotinib between the low-risk and high-risk groups. Conclusion: Our results reveal that glycolysis-related genes are feasible predictors of survival and the treatment response of patients with LUAD.

Novel lncRNA LINC01614 Facilitates Bladder Cancer Proliferation, Migration and Invasion Through the miR-217/RUNX2/Wnt/ß-Catenin Axis.

BACKGROUND: LncRNA plays a vital role in tumorigenesis and development. This study aimed to explore the novel lncRNA affecting bladder cancer progression. METHODS: The open-access data of bladder cancer patients, including transcriptome profiles and corresponding clinical information were all obtained from The Cancer Genome Atlas database. All the statistical analysis were performed using R software, SPSS and GraphPad Prism 8. CCK8, colony formation, apoptosis detection and tumorigenicity assay were used to assess cell proliferation ability. Transwell assay and wound-healing assay were used to evaluate cell metastasis potential. RESULTS: Our result showed that the lncRNA LINC01614 was highly expressed in bladder cancer tissue and cell lines. Meanwhile, patients with high LINC01614 expression level tend to have poor clinical features and shorter survival time. Further experiments demonstrated that the inhibition of LINC01614 could significantly hamper the proliferation and invasion of bladder cancer cells. Then, we found that the LINC01614 could regulate RUNX2 expression through miR-137. GSEA analysis indicated that the Wnt/ß-catenin signaling pathway might be the downstream pathway of LINC01614. Further experiments showed that the LINC01614 act as an oncogene in bladder cancer partly depending on the RUNX2/Wnt/ß-catenin axis, making it an underlying therapeutic target. CONCLUSION: In all, LINC01614 facilitates bladder cancer cells proliferation, migration and invasion through the miR-217/RUNX2/Wnt/ß-catenin axis.

Integrin-associated CD151 is a suppressor of prostate cancer progression.

Owing to the complexity of interacting molecular networks on the cell surface, integrin-associated tetraspanin CD151 remains controversial regarding its clinical importance and functional impact in prostate cancer. The current study evaluated dynamics and clinical importance of CD151 expression and its function in prostate cancer by IHC analysis of two independent patient cohorts (n=80, 181), bioinformatic interrogation of the TCGA database, and evaluation of gene knockdown effect at the cellular level. Our data showed that aside from high mRNA expression, CD151 was primarily localized to intercellular junctions at the plasma membrane in normal prostate glands or benign tissues, regardless of nature of antibodies used. By contrast, in primary tumors from patients with metastatic disease, CD151 was largely localized in the cytosol. Furthermore, the level of the cell-cell junction-linked CD151 was inversely associated with Gleason grade and tumor stage (P<0.001 for both). The portion of primary tumors expressing junctional CD151 was also three-fold less in the metastatic patient population than its counterpart (P<0.001). In line with these observations, CD151 and its associated α3ß1 or α6ß4 integrin inversely correlated with androgen receptor (AR) at the mRNA level (Spearman coefficient: -0.44, -0.48 and -0.42) in the TCGA cohort. Expression of these adhesion molecules also correlated with DNA methylation in their promoters (Spearman coefficient: -0.37, -0.71 and -0.82). Combined, these data suggest that CD151 and associated integrins are linked to tumor metastasis through AR and the epigenetic program. Meanwhile, CD151 knockdown in E-cadherin-positive tumor cells led to increased cell proliferation and induction of the epithelial-mesenchymal transition (EMT)-like phenotype. Given the strong RGD-binding integrin dependence of EMT-featured tumor cells, we examined focal adhesion kinase (FAK), their key signaling effector, in the above patient cohorts. In contrast to CD151, FAK exhibited positive correlation with tumor grade and stage as well as AR and p53 inactivation at either mRNA, protein or genomic level. Taken together, our results suggest that CD151 represses prostate cancer by antagonizing cell proliferation, EMT and the signaling of RGD-binding integrins. Since this anti-tumorigenic role is prone to the AR-mediated transcriptional and epigenetic regulation, CD151 and possibly α3ß1 and α6ß4 integrins are of potential biomarkers for metastatic prostate cancer.

Deletion of tetraspanin CD151 alters the Wnt oncogene-induced mammary tumorigenesis: A cell type-linked function and signaling.

Tetraspanin CD151 is increasingly implicated as a multifaceted mediator of cancer development and progression. Here we investigated the role of CD151 in breast cancer in the context of the Wnt oncogenic activation. Our data showed that removal of one or both of CD151 alleles in the MMTV-Wnt1 model significantly decreased the tumor-free survival of mice from 34 weeks on average to 22 weeks and 18 weeks, respectively. This effect coincided with an accelerated tumor growth and an increased number of Ki-67+ proliferative cells. Mechanistically, the CD151-deficient tumors were largely ER+, and exhibited hyperactivation of the Wnt pathway as reflected by a marked upregulation in ß-catenin and Cyclin D1, and their target genes. In addition, E-cadherin displayed a cytosolic distribution and transcription factor Snail was markedly upregulated. Collectively, this data implies that CD151 suppresses the Wnt1-driven tumorigenesis, at least in part, via counteracting the epithelial-mesenchymal transition (EMT)-like program in luminal epithelial cells. Meanwhile, the proportion of tumor cells expressing CK5 or p63, the biomarkers of myoepithelial/basal cells, markedly decreased in the absence of CD151. This change was accompanied by a decreased invasiveness of tumors and their incompetence to form a long-term cell culture. Consistent with this basal cell-linked role, the CD151 downregulation impairs mammosphere formation in MCF-10A cells and the defect was rescued by re-expression of intact CD151 ORF, but not its integrin binding-defective mutant. Overall, our study suggests that CD151 is a key player in the Wnt oncogene-driven tumorigenesis and impacts breast cancer malignancy in a cell type-dependent manner.

Identification of biomarkers and their functions in dasatinib-resistant pancreatic cancer using bioinformatics analysis.

Dasatinib is a tyrosine kinase inhibitor, which inhibits tumor proliferation by blocking SRC pathways and is considered as a potential treatment of various epithelial neoplasms, including pancreatic cancer. However, dasatinib efficacy is largely limited due to drug resistance. In the present study, bioinformatics strategies were used to investigate the potential mechanisms of dasatinib-resistance in pancreatic cancer. The gene expression profiles of the Panc0403, Panc0504, Panc1005 (dasatinib-sensitive), SU8686, MiaPaCa2 and Panc1 (acquired dasatinib-resistant) cell lines were obtained from the gene expression omnibus database. The differentially expressed genes (DEGs) were then selected using R software. In addition, gene ontology (GO) and pathway enrichment analysis were performed through the Database for Annotation, Visualization and Integrated Discovery. A protein-protein interaction (PPI) network was constructed and analyzed to determine the hub genes using the Search Tool for the Retrieval of Interacting Genes database. A total of 472 DEGs, including vimentin, transmembrane 4 l six family member 18 and S100 calcium binding protein P, were identified. Enrichment analysis by GO function demonstrated that DEGs were associated with extracellular components, signal regulation and binding factors. The analysis of the Kyoto Encyclopedia of Genes and Genomes demonstrated that several adenocarcinoma pathways were enriched, including the phosphoinositide 3-kinases/protein kinase B and mitogen-activated protein kinase signaling pathways. Some hub genes were highlighted following the PPI network construction, including Rac family small GTPase 1, laminin subunit α3, integrin subunit ß4, integrin subunit α2, collagen type VI α1 chain, collagen type I α2 chain, arrestin ß1 and synaptotagmin 1, which may be associated with pancreatic adenocarcinoma prognosis. A total of five out of eight hub genes were highly associated with the overall survival rate (P<0.05). In conclusion, the present study reported novel insights into the mechanisms of dasatinib resistance. Identification of these hub genes may be considered as potential novel treatment targets for dasatinib-resistance in pancreatic cancer.

Establishment and characterization of a novel cell line (cc­006cpm8) of moderately/poorly differentiated colorectal adenocarcinoma derived from a primary tumor of a patient.

In the present study, the cc­006cpm8 novel colon cell line was established from a sample of right colorectal adenocarcinoma obtained from a woman with liver metastasis. It was possible to culture this cell line for ≥100 passages in vitro with vigorous growth. Morphologically, the cells grew as several layers with tight adhesion to the surface of the culture plate. The morphological, immunological and ultrastructural features of these cells suggested their epithelial origin. The characterization of this cell line indicated a doubling time of 27 h, a colony forming efficiency of 73.2% in semisolid media and a plate efficiency of 66.5% in liquid culture. The modal number of chromosomes was 50. In vivo, the cc­006cpm8 cells underwent tumorigenesis in all nude mice used. Immunohistochemical analysis demonstrated that mutS homolog 2 (MSH2) and MSH6 were expressed; however, mutL homolog 1 and postmeiotic segregation 2 were downregulated in cc­006cpm8 cells. To determine the mutation profile of the cell line analyzed, exome capture DNA sequencing was performed. The results revealed 20 hypermutated exons comprising single nucleotide polymorphisms, and insertion and deletions (InDels), including single nucleotide variants of mucin (MUC)19, MUC16, MUC12, filaggrin and AHNAK nucleoprotein 2, and InDels of ß defensin­126, microRNA­3665, WNK lysine deficient protein kinase 1 and SLAIN motif­containing protein 1. In addition, commonly mutated genes in colorectal cancer and exon mutations of genes in cc­006cpm8 cells were analyzed, including adenomatous polyposis coli, tumor protein p53, Drosophila mothers against decapentaplegic 4, phosphatidylinositol­4,5­bisphosphate 3­kinase catalytic subunit α and Kirsten rat sarcoma, and genes associated with the DNA mismatch repair pathway were investigated.

Clinicopathological and prognostic significance of metastasis-associated in colon cancer-1 in gastric cancer: A meta-analysis.

BACKGROUND: The gene metastasis-associated in colon cancer-1 (MACC1) has been reported to be overexpressed in diverse human malignancies, and an increasing amount of evidence suggests that its overexpression is associated with the development and progression of many human tumors. However, the prognostic and clinicopathological value of MACC1 in gastric cancer remains inconclusive. Therefore, we conducted this meta-analysis to investigate the effect of positive MACC1 expression on clinicopathological features and survival outcomes in gastric cancer. METHODS: Medline, Web of Science, and EMBASE databases were searched for relevant articles published up to 10 April 2018. The correlation of MACC1 expression levels with overall survival and clinicopathological features was analyzed. RESULTS: In this meta-analysis, nine studies with a total of 2103 gastric cancer patients were included. Our results showed that high expression of MACC1 was significantly related to a poor overall survival. Moreover, our meta-analysis showed that MACC1 overexpression was significantly linked to distant metastasis and vascular invasion. There were no significant correlations between positive MACC1 expression and gender, localization, tumor-node-metastasis (TNM) stage, tumor extent (T stage) and lymph node involvement (N stage). CONCLUSIONS: MACC1 expression levels can serve as a novel prognostic factor in gastric cancer patients.

Establishment and characterization of the GC-030-35 cell line derived from gastric hepatoid adenocarcinoma.

PURPOSE: Gastric hepatoid adenocarcinoma is a rare subtype of primary gastric cancer and is a high-grade form of malignancy. However, the pathogenesis and molecular biology of gastric hepatoid adenocarcinoma remain poorly understood. The aim of this study was to establish and characterize a new human gastric hepatoid adenocarcinoma cell line, GC-030-35. MATERIALS AND METHODS: The GC-030-35 cell line was established from tumor cells from a 58-year-old Chinese man with gastric hepatoid adenocarcinoma. The cultured cells underwent immunocytochemistry and flow cytometry to confirm the tumor cell phenotype. RNA sequencing was performed to analyze the differences in gene expression between GC-030-35 cells compared with normal gastric epithelial cells. A zebrafish assay was performed. Gene enrichment analysis and interrogation of the bioinformatics databases, the Gene Ontology (GO) database and the Kyoto Encyclopedia of Genes and Genomes (KEGG) database, were used for pathway analysis. RESULTS: Flow cytometry analysis of the GC-030-35 cells showed a positive expression rate for CD44+ of 10.7%, high cell clonality, an average plating efficiency of 32%, cell-doubling time of 29.2 hours, and cell proliferation for >15 generations in serial culture. The zebrafish assay showed the ability of the GC-030-35 cells to proliferate, promote angiogenesis, and metastasize. RNA sequencing identified the functional clustering of 6,601 differentially expressed genes of GC-030-35, which were significantly different when compared with nonneoplastic gastric epithelial cells. Pathway enrichment analysis and interrogation of the GO and KEGG bioinformatics databases identified genes for microbial metabolism in diverse environments (63 genes), metabolism of xenobiotics by cytochrome P450 (CYP450; 25 genes), and the drug metabolism cytochrome P450 (28 genes). CONCLUSION: A human gastric hepatoid adenocarcinoma cell line, GC-030-35, was developed and characterized by comparison with normal gastric epithelial cells. Bioinformatics and gene analysis data showed that the CYP450 gene was significantly differentially expressed by GC-030-35 cells.

Establishment and Characterization of gc-006-03, a Novel Human Signet Ring Cell Gastric Cancer Cell Line Derived from Metastatic Ascites.

Signet ring cell gastric cancer (SRCGC) is a special type of gastric cancer with rapid progression and poor prognosis. However, few available SRCGC cell lines from Chinese patients can be used for research, the molecular mechanism of its growth and metastasis is still incompletely understood. In this study, we established and characterized a novel SRCGC cell line, gc-006-03.The cells showed a tendency to pile up without contact inhibition. G-band karyotypes of gc-006-03 were revealed hypotriploid with a modal chromosome number of 51. Immunohistochemistry analysis showed that the cells were positive for CEA, CK7, CDX2 and Ki-67(45%), and negative for CK20, TTF1and Li-cadherin. Flow cytometry analysis showed that gc-006-3 had 25% of CD44+ cells. The cells possessed strong clonality and high plating efficiency, and the doubling time was 36h. The cells grew vigorously for more than 100 passages in serial culture. Meanwhile, the cells showed a high rate of tumor formation. Tumors were observed in all of the nude mice (5/5) given injections of the cells. The metastatic capability of the cell line was found in zebrafish injected the cells. The results of whole genome sequencing revealed the unique genomic characteristics of gc-006-03. In summary, this new stable cell line may be useful in basic and clinical research on gastric signet ring cell carcinoma.

Influence of TS (rs34743033) and RUNX1 (rs2014300) gene polymorphisms on survival outcomes of fluorouracil-based chemotherapy in Chinese advanced gastric cancer patients.

BACKGROUND: This study aimed to explore the clinical correlation of single-nucleotide polymorphisms of thymidylate synthase (TS) and runt-related transcription factor 1 (RUNX1) in patients with postoperative stage II and III gastric cancer (GC). PATIENTS AND METHODS: Samples were obtained from 661 patients with postoperative stage II and III GC. TS (rs34743033) and RUNX1 (rs2014300) were genotyped in 261 patients who received postoperative basic platinum and fluorouracil chemotherapy regimens and 400 patients who did not accept chemotherapy. RESULTS: TS (rs34743033) variant genotypes significantly prolonged the median overall survival (OS) time compared to the patients who only received adjuvant chemotherapy (HR 1.604, 95% CI 1.068-2.410, p=0.021). Moreover, 3R/3R variant genotypes were demonstrated to have a positive effect on the OS of patients who received chemotherapy based on cisplatin (HR 1.754, 95% CI 1.041-2.954, p=0.031) compared to oxaliplatin. A stratification analysis indicated that 2R/3R and 2R/2R variant genotypes were associated with inferior survival in GC patients with intestinal-type tumors, tumor less than 5 cm in size, and poorly differentiated tumors (p<0.05). However, RUNX1 (rs2014300) AA genotypes markedly increased the risk of death in GC patients compared with the GG/GA genotypes (p=0.007), but no significant difference was observed between chemotherapy based on platinum. The stratification analysis showed that the GA/AA genotype was significantly associated with inferior survival in well to moderately differentiated tumors (HR 2.001, 95% CI 1.082-3.703, p=0.023). CONCLUSION: These preliminary results indicated that the two polymorphisms had a significant effect on postoperative adjuvant chemotherapy. TS (rs34743033) and RUNX1 (rs2014300) may be used as biomarkers to predict prognosis and select chemotherapy regimens in GC patients.