Gene amplification resulting from double strand breaks (DSBs) is a typical genetic alteration in tumorigenesis and drug-resistant progression. Amplified oncogenes and drug-resistant genes are present on extrachromosomal DNAs (ecDNAs), or chromosomal homogeneously staining regions (HSRs). Considering the role of mismatch repair (MMR) as a sensor of DSBs, we hypothesized that MMR may be involved in gene amplification. We used two MTX-resistant HT-29 colorectal cancer cell lines, which served as models with amplified genes mainly in HSRs or ecDNAs. Expression of MSH2, a key protein in MMR, was increased following the acquisition of MTX-resistant. MMR inhibition was achieved by depleting MSH2. Suppression of MMR led to decreased copy numbers of amplified genes as well as the quantity of ecDNAs and HSR. This was caused by the decreased efficiency of DSBs repair, which resulted from the reduced ability of MMR to recruit DSBs repair proteins. Additionally, it accelerated the formation of micronuclei (MN)/nuclear buds (NBUDs), which functioned to eliminate the amplified genes. Furthermore, the suppression of MMR was capable of inhibiting cell proliferation and enhancing MTX-sensitivity in ecDNA-containing cells. Conversely, suppression of MMR had no effect on gene amplification in HSR-containing cells. Our findings demonstrate that MMR plays a pivotal role in gene amplification through mediating DSBs repair pathways and facilitating the formation of MN/NBUDs in ecDNA-containing cells. MMR is likely to emerge as a prime therapeutic target worthy of in-depth exploration in future clinical investigations.

During tumorigenesis, fibroblasts transition from homeostatic cells to cancer-promoting entities, a process potentially mediated by long non-coding RNAs (lncRNAs). While some oncogenic lncRNAs facilitate carcinogenesis, the role of downregulated homeostatic lncRNAs remains unclear. This study reveals that LINC01133, primarily expressed in fibroblasts, is significantly reduced in cancer-associated fibroblasts. The decreased expression of LINC01133 in gastric adenocarcinoma-associated fibroblasts diminishes its extracellular-vesicle-mediated uptake by tumor cells, leading to the acceleration of gastric cancer progression. KLF4 regulates the transcription of LINC01133 in fibroblasts. Mechanistically, binding of LINC01133 to the 173-191 residue region of CDC42 suppresses the activity of its downstream kinases PAK3 and MLK3, and LINC01133 was also the sponge of miR-199a-5p, thereby competing with PHLPP1. In vivo studies confirmed that either overexpression of LINC01133 or inhibition of miR-199a-5p suppresses gastric adenocarcinoma cell growth. In summary, LINC01133 re-activation may serve as a potential therapeutic strategy for inhibiting metastasis in gastric adenocarcinoma.

Gastric cancer is one of the most malignant digestive tract tumors worldwide and its progression is associated with gene expression and metabolic alteration. We revealed that the gastric cancer patients with lower expression level of TOB1 exhibited poorer overall survivals according to the data in Kaplan-Meier Plotter. The unphosphorylated TOB1 protein which is effective expressed lower in gastric cancer cells. The gastric cancer cells with TOB1 gene depletion performed higher abilities of proliferation, migration and invasion and lower ability of apoptosis in vitro. The TOB1 gene depletion also promoted the tumorigenesis of gastric cancer cells in vivo. The gastric cancer cells with TOB1 gene overexpression had the converse behaviors. The transcriptional and metabolic sequencing was performed. The analyzation results showed that genes correlate-expressed with TOB1 gene were enriched in the pathways related to ERK pathway, including focal adhesion pathway, which was verified using real-time quantitative PCR. After inhibiting ERK pathway, the proliferation, colony formation and migration abilities were reduced in gastric cancer cells with low phosphorylated TOB1 protein expression level. Moreover, Pearson correlation analysis was adopted to further analyze the correlation of enriched metabolic products and differentially expressed genes. The expression of Choline, UDP-N-acetylglucosamine, Adenosine and GMP were related to the function of TOB1. This study demonstrates the genes and metabolites related to focal adhesion pathway and ERK pathway are the potential diagnosis and therapeutic targets to gastric cancer with TOB1 depletion.

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.

Background: Diesel exhaust particles (DEPs), a predominant component of ambient particulate matter (PM), are classified as ultrafine particles with the capacity to penetrate the cerebral blood-brain barrier (BBB). This penetration is implicated in the pathogenesis of central nervous system (CNS) disorders. The integrity of the BBB is inextricably linked to cerebrovascular homeostasis and the development of neurodegenerative disease, highlighting the importance of studying the effects and mechanisms of DEPs on BBB function damage. Methods and results: Utilizing mouse cerebral microvascular endothelial cells (bEnd.3 cells) as an in vitro model of the BBB, we explored the detrimental effects of DEPs exposure on BBB permeability and integrity, with particular focus on inflammation, cell apoptosis, and miRNA expression profiles. Our findings revealed that exposure to DEPs at varying concentrations for 48 h resulted in the inhibition of bEND.3 cell proliferation, induction of cell apoptosis, and an upregulation in the secretion of inflammatory cytokines/chemokines and adhesion molecules. The BBB integrity was further compromised, as evidenced by a decrease in trans-epithelial electrical resistance (TEER), a reduction in cytoskeletal F-actin, , and diminished tight junction (TJ) protein expression. Microarray analysis revealed that 23 miRNAs were upregulated and 11 were downregulated in response to a 50 mu g/mL DEPs treatment, with miR-466d-3p being notably differentially expressed. Wnt3 was identified as a target of miR466d-3p, with the Wnt signaling pathway being significantly enriched. We validated that miR-466d-3p expression was downregulated, and the protein expression levels of Wnt/beta-catenin and Wnt/PCP signaling components were elevated. The modulation of the Wnt signaling pathway by miR-466d-3p was demonstrated by the transfection of miR-466d-3p mimic, which resulted in a downregulation of Wnt3 and beta-catenin protein expression, and the mRNA level of Daam1, as well as an enhancement of TJ proteins ZO-1 and Claudin-5 expression. Conclusions: Our study further confirmed that DEPs can induce the disruption of BBB integrity through inflammatory processes. We identified alterations in the expression profile of microRNAs (miRNAs) in endothelial cells, with miR-466d-3p emerging as a key regulator of tight junction (TJ) proteins, essential for maintaining BBB integrity. Additionally, our findings primarily demonstrated that the Wnt/ beta-catenin and Wnt/PCP signaling pathway can be activated by DEPs and are regulated by miR-466d-3p. Under the combined effects of Wnt/PCP and inflammation, there is an ultimate increase in BBB hyperpermeability. Methods and results: Employing mouse cerebral microvascular endothelial cells (bEnd.3 cells) as an in vitro model of the BBB, we investigated the adverse effects of DEPs exposure on BBB permeability and integrity, with particular focus on inflammation, cell apoptosis, and miRNA expression profiles. Our findings revealed that exposure to DEPs at varying concentrations for 48 h resulted in the inhibition of bEND.3 cell proliferation, in- duction of cell apoptosis, and an increase in the release of inflammatory cytokines/chemokines and adhesion molecules. The BBB integrity was further compromised, as evidenced by a decrease in trans-epithelial electrical resistance(TEER), a reduction in cytoskeletal F-actin, loss of intercellular junctional organization, and diminished tight junction (TJ) protein expression. Microarray analysis disclosed that 23 miRNAs were upregulated and 11 were downregulated in bEND.3 cells treated with 50 mu g/mL DEPs compared to the controls. In particular, miR- 466d-3p was identified as a significantly differentially expressed miRNA. Wnt3 was predicted to be a target of miR-466d-3p, and the Wnt signaling pathway was identified as one of the most significantly enriched pathways. We validated that miR-466d-3p expression was downregulated, and the protein expression levels of Wnt/ (3- catenin and Wnt/PCP signaling components were elevated. The modulation of the Wnt signaling pathway by miR-466d-3p was demonstrated by the transfection of miR-466d-3p mimic, which resulted in a downregulation of Wnt3 and (3- catenin protein expression, and the mRNA level of Daam1, as well as an enhancement of TJ proteins ZO-1 and Claudin-5 expression. Conclusions: Our study further confirmed that DEPs can induce the disruption of BBB integrity by inflammation. We identified changes in the expression profile of microRNAs (miRNAs) in endothelial cells, with miR-466d-3p emerging as a regulator of tight junction (TJ) proteins, which are critical for maintaining BBB integrity. Addi- tionally, our findings primarily demonstrated that the Wnt/ (3- catenin and Wnt/PCP signaling pathway can be activated by DEPs and is regulated by miR-466d-3p, and under the combined effects of Wnt/PCP and inflam- mation ultimately led to hyperpermeability BBB.

染色体外环状DNA(eccDNA)是一类独立于染色体的环状闭合DNA分子,呈现单链或双链,长度主要在1 kb以下,广泛存在于自然界中.目前认为,eccDNA的起源并非随机而是具有明显的序列倾向性,并提示其具有潜在的调控基因表达和影响基因组稳定性和可塑性等作用.eccDNA的生成是一个多机制参与的复杂过程,主要包括染色体微缺失、细胞凋亡和凋亡介导的DNA片段化以及DNA损伤修复等过程.eccDNA染色体起源和亚细胞定位的不同,决定了eccDNA具有调控相关基因表达、产生特异转录本、激活免疫炎症反应等不同的功能.eccDNA在孕妇外周血、成神经管细胞瘤、晚期慢性肾病等不同生理或病理状态下,表现出长度、丰度和表观遗传学等方面的特异性分子特征,同时它的环状结构赋予其较高的稳定性,提示其可能具有成为新型生物标志物的潜力,在液体活检等领域可能具有应用价值.本综述旨在阐明eccDNA的生物学特性、生物学功能以及潜在应用等方面的最新研究进展,为了解eccDNA的发生机制、特征和作用以及为后续研究提供思路.

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.

Extrachromosomal DNAs (ecDNAs), also known as double minutes (DMs), can induce a fast increase in gene copy numbers and promote the development of cancer, including drug resistance. MutS homolog 3 (MSH3), a key protein in mismatch repair, has been indicated to participate in the regulation of DNA double-strand break (DSB) repair, which has been reported to be associated with the formation of ecDNAs. However, it remains unclear whether MSH3 can influence drug resistance via ecDNAs in cancer. In the present study, high MSH3 expression was observed in methotrexate (MTX)-resistant HT29 cells [DM- and homogeneously staining region (HSR)-containing cells] compared with parental HT29 cells. Additionally, decreased amounts of ecDNAs, HSRs and amplified genes locating on ecDNAs and HSRs were detected following depletion of MSH3 and this could be reversed by overexpressing MSH3 in DM-containing cells. No corresponding changes were found in HSR-containing cells. The present study further verified the involvement of MSH3-regulated DNA DSB repair pathways in the formation of ecDNAs by detecting the expression of core proteins and pathway activity. Furthermore, expulsion of ecDNAs/HSRs was detected and increased frequencies of micronuclei/nuclear buds with dihydrofolate reductase (DHFR) signals were observed in MSH3-depleted DM-containing cells. Finally, changes in MSH3 expression could affect DHFR amplification-derived DHFR expression and cell sensitivity to MTX, suggesting that MSH3 may influence cancer drug resistance by altering the amount of ecDNAs. In conclusion, the present study revealed a novel mechanism involving MSH3 in the regulation of ecDNAs by DSB repair, which will have clinical value in the treatment of ecDNA-based drug resistance in cancer.

5-fluorouracil(5FU)is a pyrimidine antimetabolite, which binds to thymidine synthase(TS)after transformation in vivo and interferes with DNA synthesis, thereby leading to cell damage and death.5FU is widely used in the chemotherapy for colorectal cancer, breast cancer, gastric cancer and other cancers.However, there is still an important limitation of the clinical application of 5FU that patients are prone to develop drug resistance, which leads to low efficacy or failure of chemotherapy.Non-coding RNA(ncRNA)is a class of RNAs that does not encode proteins.Recent studies have found that ncRNA plays important roles in the 5FU resistance mechanism by regulating the expression of downstream target genes.In this review, the mechanisms underlying 5FU resistance, such as nucleotide metabolism, gene repair and tumor microenvironment, are summarized, and how ncRNAs play a role in these mechanisms are illustrated.It provides supports for further research on the mechanisms of 5FU drug resistance and exploring the treatment plans for reversing drug resistance.

N6-methyladenosine(m 6A)modification is one of the most common and abundant modification form in eukaryotes, and it can regulate the post-transcriptional expression levels of genes without changing the base sequences.The modification process is dynamic and reversible and is coordinately regulated by methyltransferases, demethylases and corresponding reader proteins.Abnormality of enzymes for m 6A modificationcan cause a series of diseases, such as tumors.This article reviews the composition and functions of m 6A modification-related proteins, and the functions and regulatory mechanisms of m 6A modification in the processes of tumor proliferation, invasion and metastasis, angiogenesis, inflammatory response, immune response, genomic instability, and cell metabolism.

DNMT1 is not only an important DNA methyltransferase, which maintains the unity of methylation modification between offspring and parents, but also contains specific sequences at the N-terminus, which make it play an important role in DNA damage response(DDR). DNA damage response is the general term for the response of cells to DNA damage, involving the recognition and amplification of DNA damage signals, the recruitment of DNA repair factors to initiate repair pathways, coordination of cycle regulation, and induction of cell apoptosis.Here, we review the functions of DNMT1, the process of DDR and DNMT1’s involvement in the DDR process by participating in the perception and transmission of damage signals, regulating DNA damage repair, arresting cycle checkpoints and inducing cell apoptosis.

微小RNA (microRNA,miRNA)是一类由19～24个核苷酸构成的内源性的非编码单链RNA,通过与靶基因完全或不完全结合降解mRNA或抑制mRNA的翻译,在体内起到转录后调控作用,参与多种生理和病理过程.新近报道发现,miR-338-5P在黑色素瘤、消化系统肿瘤、肺癌等各种类型的癌症中异常表达,其通过结合一系列的靶基因来调控细胞增殖、转移、侵袭和凋亡,在不同肿瘤中发挥抑癌或促癌的不同作用,并可能成为癌症诊断的生物标志物和治疗靶点.在其他炎性相关疾病中,比如病理性心肌肥厚、类风湿性关节炎等,也发现miR-338-5P可以通过靶向不同基因,参与疾病的发生发展.现综述miR-338-5P在肿瘤、肿瘤放化疗以及其他炎性相关疾病中的作用和其在疾病诊断、预后判断等方面的应用,为肿瘤等疾病相关标志物以及更有效的治疗靶点的开发提供新思路.

Epigenetic modifications which have significant changes in the early stage of tumorigenesis play a critical role in the process of normal cell growth and development as well as in tumorigenesis. Genomic instability is one of the important features of tumors. Studying the relationship between epigenetic modifications and genomic instability will help to further understand the mechanisms of tumorigenesis, which will provide reasonable evidence for the early clinical diagnoses and therapies of cancers. This article summarizes the functions and mechanisms of epigenetic modifications, and the relationship between the alterations of these modifications and genomic instability of tumor cells.

Hypoxia as a common feature in the microenvironment of solid tumors, plays a key role in the development, progression, metastasis and prognosis of tumors. The DNA damage response system is an efficient repair mechanism to maintain genomic stability. Hypoxia affects genomic stability by regulating the process of DNA damage responses. This review summarizes the multifacet regulation of hypoxia on DNA damage responses, including changes in cell cycle checkpoint activation and triggering of downstream cascade reactions, down-regulation of transcription and translation of DNA repair pathways, and epigenetic regulation of DNA damage responses. These are of great significance for understanding the response to hypoxia and DNA damage in tumors, discovering potential therapeutic targets of tumors, and preventing the malignant progression of tumors.

DNA double-strand breaks are severe DNA lesions and are tightly related to gene am-plification secondary to injury . Their repair pathways include homologous recombination ( HR ) and non-ho-mologous end joining ( NHEJ ) . NHEJ can be divided into classical non-homologous end joining ( C-NHEJ ) and alternative non-homologous end joining ( A-NHEJ ) . They are relevant , but more importantly they compete with each other in most of conditions . This review briefly introduces the differences and connection between the two kinds of NHEJ in DNA recognition , processing , and ligation . And the latest progression in clinic therapy of these two kinds of DSB repair pathways .

DNA double strand break ( DSB ) is one of the most serious DNA damages . Homolo-gous recombination and non-homologous end joining are the two pathways repairing the DSBs in cells . Mismatch repair ( MMR ) plays a key role in maintaining genomic stability through correcting base-base mismatches and insertion/deletion mispairs generated during DNA replication and recombination . Addition-ally , MMR can recruit key factors in HR pathway , refuse the recombination between heteroduplex and cor-rect the mismatches in non-homologous end joining . Therefore , MMR has an important role in DSBs re-pair , ensuring the accuracy of DSBs repair . This review summarizes the mechanism of MMR and DSBs re-pair , and discusses the role of MMR in DSBs repair .

Extrachromosomal circular DNAs ( eccDNAs ) can be originated from the sequences of chromosomes , and have high heterogeneity in origin . In different surroundings , the production of eccDNAs involves various generation model and repair mechanisms . Technologies as high throughput sequencing ( HTS ) greatly help to identify more eccDNAs . Recently , eccDNAs were found in human circulating sys-tem . EccDNAs can affect cell life activities , promote tumor cell adaptive evolution and increase the ge-nome plasticity and instability . EccDNAs show great potential in diagnosis , prognosis prediction and treat-ment of tumors and in liquid biopsy as well . The purpose of this review is to elucidate the research pro-gress on the origin , function , study strategy and potential utility of human eccDNAs .

Gene amplification, which involves the two major topographical structures double minutes (DMs) and homegeneously stained region (HSR), is a common mechanism of treatment resistance in cancer and is initiated by DNA double‐strand breaks. NHEJ, one of DSB repair pathways, is involved in gene amplification as we demonstrated previously. However, the involvement of homologous recombination, another DSB repair pathway, in gene amplification remains to be explored. To better understand the association between HR and gene amplification, we detected HR activity in DM‐ and HSR‐containing MTX‐resistant HT‐29 colon cancer cells. In DM‐containing MTX‐resistant cells, we found increased homologous recombination activity compared with that in MTX‐sensitive cells. Therefore, we suppressed HR activity by silencing BRCA1, the key player in the HR pathway. The attenuation of HR activity decreased the numbers of DMs and DM‐form amplified gene copies and increased the exclusion of micronuclei and nuclear buds that contained DM‐form amplification; these changes were accompanied by cell cycle acceleration and increased MTX sensitivity. In contrast, BRCA1 silencing did not influence the number of amplified genes and MTX sensitivity in HSR‐containing MTX‐resistant cells. In conclusion, our results suggest that the HR pathway plays different roles in extrachromosomal and intrachromosomal gene amplification and may be a new target to improve chemotherapeutic outcome by decreasing extrachromosomal amplification in cancer.

Circular RNA (circRNA) is a new type of non-coding RNA.It is mainly composed of exons,and prevalent and stable presence among living organisms.Research has revealed that circRNA has many functions such as microRNA (miRNA) sponges,and regulation of alternative splicing and gene expression.Recent evidence suggests that circRNA plays an important role in many diseases.This review summarizes our current understanding of theclassifications,characteristics,functions,and the relationships of circRNA with diseases,in order to aid diagnosis,treatment,and prevention of clinical related diseases.