LncRNA FAM83A-AS1 facilitates tumor proliferation and the migration via the HIF-1α/ glycolysis axis in lung adenocarcinoma

Background: Lung adenocarcinoma (LUAD), the major subtype of lung cancer, is among the leading cause of cancer-related death worldwide. Energy-related metabolic reprogramming metabolism is a hallmark of cancer shared by numerous cancer types, including LUAD. Nevertheless, the functional pathways and molecular mechanism by which FAM83A-AS1 acts in metabolic reprogramming in lung adenocarcinoma have not been fully elucidated.Methods: We used transwell, wound-healing scratch assay, and metabolic assays to explore the effect of FAM83A-AS1 in LUAD cell lines. Western blotting, Co-IP assays, and ubiquitination assays were used to detect the effects of FAM83A-AS1 on HIF-1α expression, degradation, and its binding to VHL. Moreover, an in vivo subcutaneous tumor formation assay was used to detect the effect of FAM83A-AS1 on LUAD.Results: Herein, we identified FAM83A-AS1 as a metabolism-related lncRNA, which was highly correlated with glycolysis, hypoxia, and OXPHOS pathways in LUAD patients using bioinformatics analysis. In addition, we uncovered that FAM83A-AS1 could promote the migration and invasion of LUAD cells, as well as influence the stemness of LUAD cells in vivo and vitro. Moreover, FAM83A-AS1 was shown to promote glycolysis in LUAD cell lines in vitro and in vivo, and was found to influence the expression of genes related to glucose metabolism. Besides, we revealed that FAM83A-AS1 could affect glycolysis by regulating HIF-1α degradation. Finally, we found that FAM83A-AS1 knockdown could inhibit tumor growth and suppress the expression of HIF-1α and glycolysis-related genes in vivo.Conclusion: Our study demonstrates that FAM83A-AS1 contributes to LUAD proliferation and stemness via the HIF-1α/glycolysis axis, making it a potential biomarker and therapeutic target in LUAD patients.     

Impact of DNA methyltransferases on the epigenetic regulation of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptor expression in malignant melanoma

Aberrant promoter methylation and resultant silencing of TRAIL decoy receptors were reported in a variety of cancers, but to date little is known about the relevance of this epigenetic modification in melanoma. In this study, we examined the methylation and the expression status of TRAIL receptor genes in cutaneous and uveal melanoma cell lines and specimens and their interaction with DNA methyltransferases (DNMTs) DNMT1, DNMT3a, and DNMT3b. DR4 and DR5 methylation was not frequent in cutaneous melanoma but on the contrary it was very frequent in uveal melanoma. No correlation between methylation status of DR4 and DR5 and gene expression was found. DcR1 and DcR2 were hypermethylated with very high frequency in both cutaneous and uveal melanoma. The concordance between methylation and loss of gene expression ranged from 91% to 97%. Here we showed that DNMT1 was crucial for DcR2 hypermethylation and that DNMT1 and DNMT3a coregulate the methylation status of DcR1. Our work also revealed the critical relevance of DcR1 and DcR2 expression in cell growth and apoptosis either in cutaneous or uveal melanoma. In conclusion, the results presented here claim for a relevant impact of aberrant methylation of decoy receptors in melanoma and allow to understand how the silencing of DcR1 and DcR2 is related to melanomagenesis.     

Downregulated long noncoding RNA LINC00313 inhibits the epithelial–mesenchymal transition,invasion, and migration of thyroid cancer cells through inhibiting the methylation of ALX4

Thyroid cancer represents one of the prevalent endocrine cancer with relatively high incidence rate around the world, accompanied by unchanged fatality rate. We probe into the specific role of LINC00313 in mediation of cellular processes of thyroid cancer including proliferation, migration, and invasion through the methylation of aristaless-like homeobox 4 (ALX4). Thyroid cancer-related long noncoding RNAs (lncRNAs) and genes were analyzed by microarray-based analysis. The antitumor effect of LINC00313 was examined with the gain- and loss-of-function experiments. In addition, the binding of LINC00313 and the promoter region of ALX4, and the interaction of LINC00313 with methylation-related proteins were detected. Later, xenograft tumors in nude mice were induced expecting to dig out the modulatory function of LINC00313 in tumor growth of thyroid carcinoma. The microarray-based analysis manifested that LINC00313 was overexpressed, whereas ALX4 was downregulated in thyroid cancer, the results of which were also verified in thyroid cancer tissues. Besides, our results demonstrated that LINC00313 bound to the ALX4 promoter region, and LINC00313 recruited DNMT1 and DNMT3B proteins to promote the methylation of ALX4 promoter region, thus suppressing the ALX4 expression. Finally, the downregulation of LINC00313 and upregulation of ALX4 repressed the AKT/mTOR signaling axis, thus inhibiting proliferative, migratory, invasive abilities as well as epithelial-to-mesenchymal transition (EMT) of thyroid cancer cells. Collectively, downregulated LINC00313 suppresses cell proliferation, migration, as well as invasion of thyroid cancer by inhibiting the methylation of ALX4 and increasing its expression by inactivation of the AKT/mTOR signaling pathway.     

DNMT3B7 Expression Promotes Tumor Progression to a More Aggressive Phenotype in Breast Cancer Cells

Epigenetic changes, such as DNA methylation, have been shown to promote breast cancer progression. However, the mechanism by which cancer cells acquire and maintain abnormal DNA methylation is not well understood. We have previously identified an aberrant splice form of a DNA methyltransferase, DNMT3B7, expressed in virtually all cancer cell lines but at very low levels in normal cells. Furthermore, aggressive MDA-MB-231 breast cancer cells have been shown to express increased levels of DNMT3B7 compared to poorly invasive MCF-7 cells, indicating that DNMT3B7 may have a role in promoting a more invasive phenotype. Using data gathered from The Cancer Genome Atlas, we show that DNMT3B7 expression is increased in breast cancer patient tissues compared to normal tissue. To determine the mechanism by which DNMT3B7 was functioning in breast cancer cells, two poorly invasive breast cancer cell lines, MCF-7 and T-47D, were stably transfected with a DNMT3B7 expression construct. Expression of DNMT3B7 led to hypermethylation and down-regulation of E-cadherin, altered localization of β-catenin, as well as increased adhesion turnover, cell proliferation, and anchorage-independent growth. The novel results presented in this study suggest a role for DNMT3B7 in the progression of breast cancer to a more aggressive state and the potential for future development of novel therapeutics.     

High FAM111B expression predicts aggressive clinicopathologic features and poor prognosis in ovarian cancer

BackgroundsOvarian cancer (OC) is the second most common gynecological tumor with the highest mortality rate worldwide. High FAM111B expression has been reported as a predictor of poor prognosis in other cancers, but its correlation with OC has not been reported.MethodsImmunohistochemistry of tissue microarrays was performed to detect FAM111B expression levels in 141 OC patient tissues. The prognostic value of FAM111B was determined by Kaplan–Meier survival analysis, and correlations between FAM111B expression and clinicopathologic features were investigated by the Clu-square test. The significance of FAM111B expression was verified bioinformatically using the Gene Expression Omnibus database. Protein-protein interaction were performed to explore downstream mechanisms of FAM111B in OC.ResultsAmong 141 OC patients, FAM111B was positively expressed in 87.23%, 58.16%, and 87.94%; and highly expressed in 8.51%, 17.02%, and 19.86%, as evaluated by cytoplasmic, nuclear, and combined cytoplasmic/nuclear staining. FAM111B expression was positively correlated with the expression of tumor protein markers KI67, EGFR, and PDL-1. Patients with high FAM111B expression had aggressive clinicopathologic features and shorter overall survival (P value 0.0428, 0.0050, 0.0029) and progression-free survival (P value 0.0251, 0.012, 0.0596) compared to the low FAM111B expression group for cytoplasmic, nuclear, and combined cytoplasmic/nuclear groups, respectively. These results were verified using patient data from the Gene Expression Omnibus. Seventeen genes co-expressed with FAM111B were primarily involved in “negative regulation of histone modification”, “hippo signaling” and “inner ear receptor cell differentiation”.ConclusionsHigh FAM111B expression may serve as a novel prognostic predictor and molecular therapeutic target for OC.     

DNA甲基化与癌症

DNA甲基化是重要的表观遗传修饰,主要发生在DNA的CpG岛. DNA的甲基化通过DNA甲基转移酶（DNA methyltransferases, DNMTs）完成. DNA甲基化参与了细胞分化、基因组稳定性、X染色体失活、基因印记等多种细胞生物学过程.单基因水平及基因组范围内的DNA甲基化改变在肿瘤发生发展中亦发挥重要作用. 抑癌基因的异常甲基化引起的表达抑制,可导致肿瘤细胞的增殖失控和侵袭转移,并参与肿瘤组织的血管生成过程.在许多肿瘤的研究中都发现了基因组整体DNA低甲基化所导致的染色体不稳定性. 本文从DNA的异常高甲基化和低甲基化两方面论述了DNA甲基化在细胞恶变发生发展过程中的改变及其影响,并阐述了DNA甲基化改变在肿瘤诊断和治疗中的作用.     

PFKFB4 negatively regulated the expression of histone acetyltransferase GCN5 to mediate the tumorigenesis of thyroid cancer

Thyroid cancer (TC) is the most common malignant endocrine tumor, and its incidence has progressively increased over several decades. Accumulating evidence has suggested that PFKFB4, a critical regulatory enzyme of glycolysis, has been implicated in various solid cancers. However, the exact effect of PFKFB4 on TC remains unclear. Hence, the objective of this work was to investigate the role of PFKFB4 in TC and explore the underlying regulatory mechanisms. Here, we provide evidence that mRNA levels of PFKFB4 were upregulated in TC patients’ thyroids and cell lines. Downregulation of PFKFB4 reduced TC cell viability and inhibited colony formation. In addition, the migration and invasion of TC cells were suppressed by PFKFB4 knockdown, suggesting that PFKFB4 is positively correlated with tumorigenesis of TC. Molecularly, knockdown of PFKFB4 significantly inhibited expression of GCN5 and phosphorylation of PI3K/AKT. Moreover, the suppressive role of shPFKFB4 in TC cell growth was reversed by upregulation of GCN5. Finally, the in vivo experiment indicated that downregulation of PFKF4B suppressed tumor growth in xenografts TC model mice. In total, our results suggested that PFKFB4-mediated TC tumorigenesis by positively regulating GCN5 and PI3K/AKT signaling. These findings provide new research directions and therapeutic options considering PFKF4B as a novel diagnosis marker and therapeutic target.     

Role of DNMT3B in the regulation of early neural and neural crest specifiers

The de novo DNA methyltransferase DNMT3B functions in establishing DNA methylation patterns during development. DNMT3B missense mutations cause immunodeficiency, centromere instability and facial anomalies (ICF) syndrome. The restriction of Dnmt3b expression to neural progenitor cells, as well as the mild cognitive defects observed in ICF patients, suggests that DNMT3B may play an important role in early neurogenesis. We performed RNAi knockdown of DNMT3B in human embryonic stem cells (hESCs) in order to investigate the mechanistic contribution of DNMT3B to DNA methylation and early neuronal differentiation. While DNMT3B was not required for early neuroepithelium specification, DNMT3B deficient neuroepithelium exhibited accelerated maturation with earlier expression, relative to normal hESCs, of mature neuronal markers (such as NEUROD1) and of early neuronal regional specifiers (such as those for the neural crest). Genome-wide analyses of DNA methylation by MethylC-seq identified novel regions of hypomethylation in the DNMT3B knockdowns along the X chromosome as well as pericentromeric regions, rather than changes to promoters of specific dysregulated genes. We observed a loss of H3K27me3 and the polycomb complex protein EZH2 at the promoters of early neural and neural crest specifier genes during differentiation of DNMT3B knockdown but not normal hESCs. Our results indicate that DNMT3B mediates large-scale methylation patterns in hESCs and that DNMT3B deficiency in the cells alters the timing of their neuronal differentiation and maturation.     

Development of a versatile DNMT and HDAC inhibitor C02S modulating multiple cancer hallmarks for breast cancer therapy

DNMT and HDAC are closely related to each other and involved in various human diseases especially cancer. These two enzymes have been widely recognized as antitumor targets for drug discovery. Besides, research has indicated that combination therapy consisting of DNMT and HDAC inhibitors exhibited therapeutic advantages. We have reported a DNMT and HDAC dual inhibitor 15a of which the DNMT enzymatic inhibitory potency needs to be improved. Herein we reported the development of a novel dual DNMT and HDAC inhibitor C02S which showed potent enzymatic inhibitory activities against DNMT1, DNMT3A, DNMT3B and HDAC1 with IC₅₀ values of 2.05, 0.93, 1.32, and 4.16 µM, respectively. Further evaluations indicated that C02S could inhibit DNMT and HDAC at cellular levels, thereby inversing mutated methylation and acetylation and increasing expression of tumor suppressor proteins. Moreover, C02S regulated multiple biological processes including inducing apoptosis and G0/G1 cell cycle arrest, inhibiting angiogenesis, blocking migration and invasion, and finally suppressing tumor cells proliferation in vitro and tumor growth in vivo.     

UPF1甲基化和miR-744-5p/CCND1在甲状腺乳头状癌中的作用机制研究

摘要 目的：探讨UPF1甲基化和miR-744-5p/CCND1在甲状腺乳头状癌中的作用机制研究。方法：将人甲状腺乳头状癌细胞株TCP-1和正常甲状腺上皮细胞Nthy-ori-3分别用去甲基化试剂5-Aza-CdR进行干预,分别在干预前后采用甲基化特异性PCR技术检测UPF1基因甲基化变化,采用Western-Blotting 检测干预UPF1、DNMT1、miR-744-5p、CCND1蛋白相对表达,采用transwell细胞侵袭实验检测细胞侵袭情况。结果：PCR扩增显示,UPF1基因在Nthy-ori-3组仅出现非甲基化引物扩增条带（U条带）,在TCP-1组仅出现甲基化引物扩增条带（M条带）。经5-Aza-Cdr作用后,UPF1基因甲基化扩增条带减少,甲基化表达降低。各组UPF1、DNMT1、miR-744-5p、CCND1蛋白相对表达差异具有统计学意义（P＜0.05）。与Nthy-ori-3组比较,TCP-1组DNMT1、UPF1蛋白相对表达明显提高,miR-744-5p、CCND1蛋白相对表达明显降低（P＜0.05）;与TCP-1组比较,TCP-1干预组DNMT1、UPF1蛋白相对表达明显降低,miR-744-5p、CCND1蛋白相对表达明显提高（P＜0.05）。与TCP-1组比较,TCP-1干预组细胞侵袭、迁移数量明显减少（P＜0.05）。结论：UPF1甲基化存在于甲状腺乳头状癌中,UPF1基因甲基化的表达缺失可能抑制miR-744-5p/CCND1轴,在甲状腺乳头状癌发生发展中发挥关键作用。     

Silencing of lncRNA LINC00514 inhibits the malignant behaviors of papillary thyroid cancer through miR-204–3p/CDC23 axis

Numerous studies have provided that long noncoding RNAs (lncRNAs) possess important roles in regulating tumorigenesis. However, up to data, the role of LINC00514 in cancer, including thyroid cancer, remains unknown. In the present study, we found that LINC00514 expression was significantly upregulated in papillary thyroid cancer (PTC) tissues by bioinformatics analysis. Loss-of-function studies revealed that LINC00514 silencing inhibited the proliferation, migration and invasion of PTC cells while promoting apoptosis in vitro. Moreover, LINC00514 knockdown suppressed PTC growth in vivo. RNA-FISH showed that LINC00514 mainly locates in the nucleus of PTC cells. Through bioinformatics prediction, we identified that LINC00514 served as the sponge for miR-204–3p, and miR-204–3p directly targeted CDC23. Thus, LINC00514 promoted CDC23 expression via restraining miR-204–3p activity, leading to PTC progression. In sum, our findings demonstrated that LINC00514 contributes to PTC progression and might be a potential target for PTC therapy.