Background Field cancerization is the process in which a population of normal or pre-malignant cells is affected by oncogenic alterations leading to progressive molecular changes that drive malignant transformation. Aberrant DNA methylation has been implicated in early cancer development in non-small cell lung cancer (NSCLC); however, studies on its role in field cancerization (FC) are limited. This study aims to identify FC-specific methylation patterns that could distinguish between pre-malignant lesions and tumor tissues in NSCLC. Methods We enrolled 52 patients with resectable NSCLC and collected resected tumor (TUM), tumor-adjacent (ADJ) and tumor-distant normal (DIS) tissue samples, among whom 36 qualified for subsequent analyses. Methylation levels were profiled by bisulfite sequencing using a custom lung-cancer methylation panel. Results ADJ and DIS samples demonstrated similar methylation profiles, which were distinct from distinct from that of TUM. Comparison of TUM and DIS profiles led to identification of 1740 tumor-specific differential methylated regions (DMRs), including 1675 hypermethylated and 65 hypomethylated (adjusted P < 0.05). Six of the top 10 tumor-specific hypermethylated regions were associated with cancer development. We then compared the TUM, ADJ, and DIS to further identify the progressively aggravating aberrant methylations during cancer initiation and early development. A total of 332 DMRs were identified, including a predominant proportion of 312 regions showing stepwise increase in methylation levels as the sample drew nearer to the tumor (i.e. DIS < ADJ < TUM) and 20 regions showing a stepwise decrease pattern. Gene set enrichment analysis (GSEA) for KEGG and GO terms consistently suggested enrichment of DMRs located in transcription factor genes, suggesting a central role of epigenetic regulation of transcription factors in FC and tumorigenesis. Conclusion We revealed distinct methylation patterns between pre-malignant lesions and malignant tumors, suggesting the essential role of DNA methylation as an early step in pre-malignant field defects. Moreover, our study also identified differentially methylated genes, especially transcription factors, that could potentially be used as markers for lung cancer screening and for mechanistic studies of FC and early cancer development.

Since DNA methylation (DNAm) is associated with the carcinogenesis of various cancers, this study aimed to explore potential DNAm prognostic signatures of lung squamous cell carcinoma (LUSC). First, transcriptomic and methylation profiles of LUSC were obtained from The Cancer Genome Atlas database (TCGA). DNAm-related genes were screened by integrating DNAm and transcriptome profiles via MethylMix package. Subsequently, a prognostic signature was conducted with the least absolute shrinkage and selector operation (LASSO) Cox analysis. This signature combined with the clinicopathological parameters was then utilized to construct a prognostic nomogram via the rms package. A signature based on three DNAm-related genes claudin 1 (CLDN1), ATP-binding cassette subfamily C member 5 (ABCC5), and cystatin A (CSTA) that were hypomethylated and upregulated in LUSC was constructed. Univariate and multivariate Cox regression analysis suggested that this signature, combined with age and TNM.N stage, was significantly correlated with survival rate. Time-dependent receiver operating characteristics and calibration curves suggested the nomogram constructed with age and TNM.N stage variables could accurately evaluate the 3- and 5-year outcome of LUSC. Finally, the average mRNA and protein expression levels of CLDN1, ABCC5, and CSTA in LUSC were verified to be significantly higher than those in paracancerous tissues. Moreover, silencing CLDN1, ABCC5, and CSTA expressions could significantly reduce the carcinogenesis of the A549 cell line. The DNAm-driven prognostic signature consists of CLDN1, ABCC5, and CSTA incorporated with age and TNM. N stage could facilitate the prediction outcome of LUSC.

目的 基于DNA甲基化驱动基因开发预测肺鳞癌患者预后的列线图.方法 通过整合癌症基因组图谱(TCGA)肺鳞癌DNA甲基化和基因表达来鉴定甲基化驱动基因,采用Kaplan-Meier生存分析、LASSO回归和多变量Cox回归分析,建立风险评分模型,再对临床参数进行分析,构建列线图,Hosmer-Lemeshow检验绘制校准曲线对肺鳞癌患者生存率进行校准,受试者工作特征曲线(ROC曲线)评估列线图的预测效能.结果 通过MethylMix分析共筛选了45个DNA甲基化驱动基因,基因功能在丝氨酸家族氨基酸代谢过程、细胞和细胞间黏附连接、细胞和细胞间连接方面显著丰富(P<0.01).经Kaplan-Meier生存分析和LASSO回归分析显示,ATP结合家族亚家族C成员5(ABCC5)、封闭蛋白1重组蛋白(CLDN1)、半胱氨酸蛋白酶蛋白A(CSTA)基因与肺鳞癌患者预后相关(P<0.05).多因素Cox回归分析显示,年龄、TNM分期中N分期和风险评分与肺鳞癌患者的生存时间显著相关(HR=1.73,95％CI 1.09～2.75;HR=2.34,95％CI 1.21～4.52;HR=4.44,95％CI 1.61～12.26)(均P<0.05).构建包含这些变量的列线图,ROC曲线评估显示,该列线图预测肺鳞癌患者3年和5年生存率的准确性较高.结论 DNA甲基化驱动基因(ABCC5、CLDN1和CSTA)的改变与肺鳞癌患者的预后显著相关,构建的列线图模型可用于肺鳞癌患者预后的个体化预测.

PurposeThis study aimed to explore the circular RNA (circRNA/circ) profile engaged in non-small cell lung cancer (NSCLC) development and metastasis and to investigate potentially key carcinogenic circRNAs related to NSCLC. MethodsCircRNA profiles between 10 NSCLC tissues and 10 adjacent tissues and between five NSCLC tissues with lymph node metastasis (LNM) and five NSCLC tissues without LNM were detected by Arraystar Human circRNA Array followed by bioinformatics. Circ_0008594 knockdown, circ_0004293 overexpression, and circ_0003832 overexpression plasmids were transfected into H23 and H460 cells to sort potential oncogenic circRNA. Then circ_0008594 overexpression and knockdown plasmids were transfected, followed by that circ_0008594 knockdown plus miR-760 knockdown plasmids were transfected into these cells. Cell proliferation, apoptosis, invasion, stemness, and pathways were detected. In addition, xenograft mice models were constructed via injecting H23 cells with circ_0008594 overexpression or knockdown to validate the findings. ResultsA total of 455 dysregulated circRNAs in NSCLC tissues versus adjacent tissues and 353 dysregulated circRNAs in NSCLC tissues with LNM versus those without LNM were discovered. Via cross-analysis, 19 accordant circRNAs were uncovered, among which three candidate circRNAs (circ_0008594, circ_0004293, circ_0003832) were chosen for functional experiments, during which it was observed that circ_0008549 affected H23 and H460 cell proliferation and apoptosis more obviously than circ_0004293 and circ_0003832. Subsequent experiments showed that circ_0008594 promoted H23 and H460 cell proliferation and invasion but affected stemness less and negatively regulated miR-760 via direct binding. Furthermore, miR-760 attenuated the effect of circ_0008549 on regulating H23 and H460 cell functions and the PI3K/AKT and MEK/ERK pathways. In vivo experiments further confirmed that circ_0008549 increased tumor volume, epithelial-mesenchymal transition, and the PI3K/AKT and MEK/ERK pathways while reducing tumor apoptosis and miR-760 NSCLC xenograft models. ConclusionOur study identifies several valuable circRNAs related to NSCLC development and LNM. Furthermore, as a key functional circRNA, circ_0008594 was observed to promote NSCLC progression by regulating the miR-760-mediated PI3K/AKT and MEK/ERK pathways.

Background: Thymic epithelial tumors (TETs), originating from the thymic epithelial cells, are the most common primary neoplasms of the anterior mediastinum. Emerging evidence demonstrated that the competing endogenous RNAs (ceRNAs) exerted a crucial effect on tumor development. Hence, it is urgent to understand the regulatory mechanism of ceRNAs in TETs and its impact on tumor prognosis. Methods: TETs datasets were harvested from the UCSC Xena as the training cohort, followed by differentially expressed mRNAs (DEmRNAs), lncRNAs (DElncRNAs), and miRNAs (DEmiRNAs) at different pathologic type (A, AB, B, and TC) identified via DESeq2 package. clusterProfiler package was utilized to carry out gene ontology and Kyoto encyclopedia of genes and genomes functional analysis on the DEmRNAs. Subsequently, the lncRNA-miRNA-mRNA regulatory network was constructed to screen the key DEmRNAs. After the key DEmRNAs were verified in the external cohort from Gene Expression Omnibus database, their associated-ceRNAs modules were used to perform the K-M and Cox regression analysis to build a prognostic significance for TETs. Lastly, the feasibility of the prognostic significance was validated by receiver operating characteristic (ROC) curves and the area under the curve. Results: Finally, a total of 463 DEmRNAs, 87 DElncRNAs, and 20 DEmiRNAs were obtained from the intersection of differentially expressed genes in different pathological types of TETs. Functional enrichment analysis showed that the DEmRNAs were closely related to cell proliferation and tumor development. After lncRNA-miRNA-mRNA network construction and external cohort validation, a total of 4 DEmRNAs DOCK11, MCAM, MYO10, and WASF3 were identified and their associated-ceRNA modules were significantly associated with prognosis, which contained 3 lncRNAs (lncRNA LINC00665, lncRNA NR2F1-AS1, and lncRNA RP11-285A1.1), 4 mRNAs (DOCK11, MCAM, MYO10, and WASF3), and 4 miRNAs (hsa-mir-143, hsa-mir-141, hsa-mir-140, and hsa-mir-3199). Meanwhile, ROC curves verified the accuracy of prediction ability of the screened ceRNA modules for prognosis of TETs. Conclusion: Our study revealed that ceRNAs modules might exert a crucial role in the progression of TETs. The mRNA associated-ceRNA modules could effectively predict the prognosis of TETs, which might be the potential prognostic and therapeutic markers for TETs patients.

According to the new optimal game analysis on the innovation ability of Chinese enterprises, the best strategy to compete and cooperate in the game is cooperative innovation, which can attribute the advantage resources among to different parts in the cooperative innovation system, and increase the innovation ability of the enterprises and further improve the whole industry. In the research, a pattern of cooperative innovation system is built for any enterprise as an innovation subject, and the structure and function of the system is revealed from three levels as micro-scope, mid-scope and macro-scope, in doing which, the basic conditions for a healthy development of a cooperative innovation system is discussed. Then basic standards to measuring the state of a cooperative innovation system are established by quantization of the knowledge management entropy and dissipative structure negative entropy. Finally, a possible road is pointed out for the orderly development of the cooperative innovation system in the relation to the environment.