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Rationale: In breast cancer, high intratumor DNA methylation heterogeneity can lead to drug-resistant, metastasis and poor prognosis of tumors, which increases the complexity of cancer diagnosis and treatment. However, most studies are limited to average DNA methylation level of individual CpGs and ignore heterogeneous DNA methylation patterns of cell subpopulations within the tumor. Thus, quantifying the variability in DNA methylation pattern in sequencing reads is valuable for understanding intratumor heterogeneity. Methods: We performed Reduced Representation Bisulfite Sequencing and RNA sequencing for tumor core and tumor periphery regions within one breast tumor. By developing a method named “epialleJS” based on Jensen-Shannon divergence, we detected the differential epialleles between tumor core and tumor periphery (CPDEs). We then explored the correlation between intratumor methylation heterogeneity and hypoxic microenvironment in TCGA breast cancer cohort. Results: More than 70% of CPDEs had higher epipolymorphism in tumor core than tumor periphery, and these CPDEs had lower methylation in tumor core. The CPDEs with lower methylation in tumor core may associate with hypoxic tumor microenvironment. Moreover, we identified a signature of five hypoxia-related DNA methylation markers which can predict the prognosis of breast cancer patients, including a CpG site cg15190451 in gene SLC16A5. Furthermore, immunohistochemical analysis confirmed that the expression of SLC16A5 was associated with clinicopathological characteristics and survival of breast cancer patients. Conclusions: The analysis of intratumor DNA methylation heterogeneity based on epialleles reveals that disordered methylation patterns in tumor core are associated with hypoxic microenvironment, which provides a framework for understanding biological heterogeneous behavior and guidance for developing effective treatment schemes for breast cancer patients.

Aim: We aim to predict transcription factor (TF) binding events from knowledge of gene expression and epigenetic modifications. Materials & methods: TF-binding events based on the Encode project and The Cancer Genome Atlas data were analyzed by the random forest method. Results: We showed the high performance of TF-binding predictive models in GM12878, HeLa, HepG2 and K562 cell lines and applied them to other cell lines and tissues. The genes bound by the top TFs ( MAX and MAZ) were significantly associated with cancer-related processes such as cell proliferation and DNA repair. Conclusion: We successfully constructed TF-binding predictive models in cell lines and applied them in tissues.

Background An increasing number of studies have shown that long noncoding RNAs (lncRNAs) play essential roles in tumor initiation and progression. LncRNAs act as tumor promoters or suppressors by targeting specific genes via epigenetic modifications and competing endogenous RNA (ceRNA) mechanisms. In this study, we explored the function and detailed mechanisms of long intergenic nonprotein coding RNA 673 (LINC00673) in breast cancer progression. Methods Quantitative real-time PCR (qRT-PCR) was used to examine the expression of LINC00673 in breast cancer tissues and in adjacent normal tissues. Gain-of-function and loss-of function experiments were conducted to investigate the biological functions of LINC00673 in vitro and in vivo. We also explored the potential role of LINC00673 as a therapeutic target using antisense oligonucleotide (ASO) in vivo. RNA sequencing (RNA-seq), dual-luciferase reporter assays, chromatin immunoprecipitation (ChIP) assay, and rescue experiments were performed to uncover the detailed mechanism of LINC00673 in promoting breast cancer progression. Results In the present study, LINC00673 displayed a trend of remarkably increased expression in breast cancer tissues and was associated with poor prognosis in breast cancer patients. Importantly, LINC00673 depletion inhibited breast cancer cell proliferation by inhibiting the cell cycle and increasing apoptosis. Furthermore, ASO therapy targeting LINC00673 substantially suppressed breast cancer cell proliferation in vivo. Mechanistically, LINC00673 was found to act as a ceRNA by sponging miR-515-5p to regulate MARK4 expression, thus inhibiting the Hippo signaling pathway. Finally, ChIP assay showed that the transcription factor Yin Yang 1 (YY1) could bind to the LINC00673 promoter and increase its transcription in cis. Conclusions YY1-activated LINC00673 may exert an oncogenic function by acting as a sponge for miR-515-5p to upregulate the MARK4 and then inhibit Hippo signaling pathway, and may serve as a potential therapeutic target.

Advances in the molecular characteristics of cancers have facilitated the classification system from morphology to molecular characteristic-based subtypes. Cancer profiling has expanded in its focus from protein-coding genes to noncoding RNAs, with advances in the depth and quality of transcriptome sequencing. Here, we examined the profiles of long noncoding RNAs (lncRNAs) according to breast cancer subtype categories in The Cancer Genome Atlas (TCGA) database to identify a cohort of breast cancer- and oestrogen receptor (ER)-negative-associated lncRNAs. According to the prioritization of variation in ER-negative-associated lncRNAs, we identified and investigated the role of LINC00511 in breast cancer. We determined that high LINC00511 expression was an unfavourable prognostic factor for patients with breast cancer. Furthermore, LINC00511 promoted tumour growth by accelerating the G1/S transition and inhibiting apoptosis. At the transcriptional level, ER deficiency directly affected the expression of LINC00511 activated by transcription factor AP-2 (TFAP-2) in breast cancer cells. Moreover, mechanistic investigations demonstrated that ER-negative-associated LINC00511 interacted with enhancer of zeste homologue 2 (EZH2, the catalytic subunit of polycomb repressive complex 2, PRC2) and recruited PRC2 to mediate histone methylation, contributing to the repression of CDKN1B in the nucleus. This process resulted in altered ER-negative breast cancer cell biology. By highlighting the oncogenic function of LINC00511, we revealed the role of lncRNAs in regulating the network of cell cycle control in ER-negative breast cancer and suggested the exploitation of LINC00511 as an anticancer therapy in the future.

Background The landscape and biological functions of tumor suppressor long noncoding RNAs in breast cancer are still unknown. Methods Data from whole transcriptome sequencing of 33 breast specimens in the Harbin Medical University Cancer Center cohort and The Cancer Genome Atlas was applied to identify and validate the landscape of tumor suppressor long noncoding RNAs, which was further validated by The Cancer Genome Atlas pancancer data including 33 cancer types and 12,839 patients. Next, the expression model, prognostic roles, potential biological functions and epigenetic regulation of tumor suppressor long noncoding RNAs were investigated and validated in the breast cancer and pancancer cohorts. Finally, EPB41L4A-AS2 was selected to validate our novel finding, and the tumor suppressive roles of EPB41L4A-AS2 in breast cancer were examined. Results We identified and validated the landscape of tumor suppressor long noncoding RNAs in breast cancer. The expression of the identified long noncoding RNAs was downregulated in cancer tissue samples compared with normal tissue samples, and these long noncoding RNAs correlated with a favorable prognosis in breast cancer patients and the patients in the pancancer cohort. Multiple carcinogenesis-associated biological functions were predicted to be regulated negatively by these long noncoding RNAs. Moreover, these long noncoding RNAs were transcriptionally regulated by epigenetic modification, including DNA methylation and histone methylation modification. Finally, EPB41L4A-AS2 inhibited breast cancer cell proliferation, migration and invasion and induced cell apoptosis in vitro. Mechanistically, EPB41L4A-AS2, acting at least in part as a tumor suppressor, upregulated tumor suppressor gene expression. Moreover, ZNF217 recruited EZH2 to the EPB41L4A-AS2 locus and suppressed the expression of EPB41L4A-AS2 by epigenetically increasing H3K27me3 enrichment. Conclusions This work enlarges the functional landscape of known long noncoding RNAs in human cancer and provides novel insights into the suppressive roles of these long noncoding RNAs. Electronic supplementary material The online version of this article (10.1186/s13046-019-1096-0) contains supplementary material, which is available to authorized users.

Background: Several studies have found that DNA methylation is associated with transcriptional regulation and affect sponge regulation of non-coding RNAs in cancer. The integration of circRNA, miRNA, DNA methylation and gene expression data to identify sponge circRNAs is important for revealing the role of DNA methylation-mediated regulation of sponge circRNAs in cancer progression. Method: We established a DNA methylation-mediated circRNA crosstalk network by integrating gene expression, DNA methylation and non-coding RNA data of breast cancer in TCGA. Four modules (26 candidate circRNAs) were mined. Next, 10 DNA methylation-mediated sponge circRNAs (sp_circRNAs) and 5 sponge driver genes (sp_driver genes) in breast cancer were identified in the CMD network using a computational process. Findings: Among the identified genes, ERBB2 was associated with six sponge circRNAs, which illustrates its better sponge regulatory function. Survival analysis showed that DNA methylations of 10 sponge circRNA host genes are potential prognostic biomarkers in the TCGA dataset (p=0.0239) and GSE78754 dataset (p=0.0377). In addition, the DNA methylation of two sponge circRNA host genes showed a significant negative correlation with their driver gene expressions. Interpretation: We developed a strategy to predict sponge circRNAs by DNA methylation mediated with playing the role of regulating breast cancer sponge driver genes. Funding Statement: This work was supported by National Natural Science Foundation of China [grant number 81573021], the Natural Science Foundation of Heilongjiang Province [grant number ZD2015003], and the Heilongjiang Science and Technology Plan Project [YS17C22]. Declaration of Interests: We declare that we have no financial and personal relationships with other people or organizations that can inappropriately influence our work, there is no professional or other personal interest of any nature or kind in any product, service and company that could be construed as influencing the position presented in, or the review of, this manuscript. Ethics Approval Statement: The authors declare: "Not applicable."

Intervertebral disc (IVD) degeneration is known to aggravate with age and oxidative stress is implicated in the pathogenesis of many age-related diseases. Nuclear factor (erythroid-derived-2)-like 2 (Nrf2) can confer adaptive protection against oxidative and proteotoxic stress in cells. In this study, we assessed whether Nrf2 can protect against oxidative stress in nucleus pulposus (NP) cells. In addition, we investigated Nrf2 expression in NP tissue samples from patients with different degrees of IVD degeneration and a mouse model of aging and IVD degeneration and the influence of H2O2-induced oxidative stress on autophagic pathways in NP cells. Autophagy was assessed by measuring levels of autophagy-related protein (ATG) family members and the autophagic markers, p62 and LC3. We found that expression of Nrf2 progressively decreased in human NP tissue samples of patients with increasing degrees of IVD degeneration. Nrf2 deficiency leads to the degeneration of IVDs during aging. Nrf2 knockout also aggravates IVD degeneration and reduces autophagic gene expression in an induced mouse model of IVD degeneration. The detrimental effects of H2O2-induced oxidative stress were increased in autophagy-deficient cells via reduced expression of Atg7 and the Keap1–Nrf2–p62 autophagy pathway. Taken together, these results suggest that excessive oxidative stress causes the upregulation of autophagy, and autophagy acts as an antioxidant feedback response activated by a Keap1-Nrf2-p62 feedback loop in IVD degeneration.