Circular RNA circFOXO3 regulates KDM2A by targeting miR‐214 to promote tumor growth and metastasis in oral squamous cell carcinoma

Oral squamous cell carcinoma (OSCC) is a pathological type of oral cancer, which accounts for over 90% of oral cancers. It has been widely shown that circRNA is involved in the regulation of multiple malignant oral diseases including OSCC. However, the mechanism underlying how circRNA regulates OSCC is still not clearly elucidated. In this article, we report circFOXO3 promotes tumor growth and invasion of OSCC by targeting miR‐214 which specifically degrades the lysine demethylase 2A (KDM2A). CircRNA sequencing was conducted in OSCC tumor and tumor‐side tissues, and the expression of circFOXO3 is found to be markedly increased in tumor tissues. CircFOXO3 is also highly expressed in several OSCC cell lines compared with human oral keratinocytes. Transwell assay and colony formation showed that knockdown of circFOXO3 prevents the invasion and proliferation of oral cancer cells. Via bioinformatic research, miR‐214 was found to be the target of circFOXO3 and correlate well with circFOXO3 both in vitro and in vivo. KDM2A was then validated by database analysis and luciferase assay to be the direct target of miR‐214. KDM2A helps to promote tumor invasiveness and proliferation of OSCC. Collectively, our results proved that circFOXO3 sponges miR‐214 to up‐regulate the expression of KDM2A, thus promotes tumor progression in OSCC.     

Aberrant promoter hypermethylation of PBRM1, BAP1, SETD2, KDM6A and other chromatin-modifying genes is absent or rare in clear cell RCC

Recent sequencing studies of clear cell (conventional) renal cell carcinoma (ccRCC) have identified inactivating point mutations in the chromatin-modifying genes PBRM1, KDM6A/UTX, KDM5C/JARID1C, SETD2, MLL2 and BAP1. To investigate whether aberrant hypermethylation is a mechanism of inactivation of these tumor suppressor genes in ccRCC, we sequenced the promoter region within a bona fide CpG island of PBRM1, KDM6A, SETD2 and BAP1 in bisulfite-modified DNA of a representative series of 50 primary ccRCC, 4 normal renal parenchyma specimens and 5 RCC cell lines. We also interrogated the promoter methylation status of KDM5C and ARID1A in the Cancer Genome Atlas (TCGA) ccRCC Infinium data set. PBRM1, KDM6A, SETD2 and BAP1 were unmethylated in all tumor and normal specimens. KDM5C and ARID1A were unmethylated in the TCGA 219 ccRCC and 119 adjacent normal specimens. Aberrant promoter hypermethylation of PBRM1, BAP1 and the other chromatin-modifying genes examined here is therefore absent or rare in ccRCC.     

Adipocytes promote pancreatic cancer cell proliferation via glutamine transfer

Adipocytes promote progression of multiple cancers, but their role in pancreatic intraepithelial neoplasia (PanIN) and ductal adenocarcinoma (PDAC) is poorly defined. Nutrient transfer is a mechanism underlying stromal cell-cancer crosstalk. We studied the role of adipocytes in regulating in vitro PanIN and PDAC cell proliferation with a focus on glutamine metabolism. Murine 3T3L1 adipocytes were used to model adipocytes. Cell lines derived from PKCY mice were used to model PanIN and PDAC. Co-culture was used to study the effect of adipocytes on PanIN and PDAC cell proliferation in response to manipulation of glutamine metabolism. Glutamine secretion was measured with a bioanalyzer. Western blotting was used to study the effect of PanIN and PDAC cells on expression of glutamine-related enzymes in adipocytes. Adipocytes promote proliferation of PanIN and PDAC cells, an effect that was amplified in nutrient-poor conditions. Adipocytes secrete glutamine and rescue PanIN and PDAC cell proliferation in the absence of glutamine, an effect that was glutamine synthetase-dependent and involved PDAC cell-induced down-regulation of glutaminase expression in adipocytes. These findings suggest glutamine transfer as a potential mechanism underlying adipocyte-induced PanIN and PDAC cell proliferation.     

Histone demethylase KDM7A is required for stem cell maintenance and apoptosis inhibition in breast cancer

Histone demethylase KDM7A regulates neuronal differentiation and development in mammals. In this study, we found that KDM7A was also required for breast cancer stem cells (BCSCs) maintenance. Silencing KDM7A significantly reduced the BCSCs population and mamosphere formation in vitro, and inhibited breast tumor growth in vivo. Restoring KDM7A expression rescued the defect in stem cell maintenance. Our mechanism analysis suggested that KDM7A upregulated the stemness-associated factors KLF4 and c-MYC for BCSCs maintenance. In addition, KDM7A knockdown promoted apoptosis through decreasing BCL2 expression and BAD phosphorylation in breast cancer (BrCa). Furthermore, restoring KDM7A and BCL2 expression rescued apoptosis inhibition in breast cancer, suggesting that KDM7A inhibited apoptosis by upregulating the BCL2 level in breast cancer. In conclusion, KDM7A promotes cancer stem cell maintenance and apoptosis inhibition in breast cancer.     

Knockdown of KDM1A suppresses tumour migration and invasion by epigenetically regulating the TIMP1/MMP9 pathway in papillary thyroid cancer

Epigenetic dysregulation plays an important role in cancer. Histone demethylation is a well‐known mechanism of epigenetic regulation that promotes or inhibits tumourigenesis in various malignant tumours. However, the pathogenic role of histone demethylation modifiers in papillary thyroid cancer (PTC), which has a high incidence of early lymphatic metastasis, is largely unknown. Here, we detected the expression of common histone demethylation modifiers and found that the histone H3 lysine 4 (H3K4) and H3 lysine 9 (H3K9) demethylase KDM1A (or lysine demethylase 1A) is frequently overexpressed in PTC tissues and cell lines. High KDM1A expression correlated positively with age <55 years and lymph node metastasis in patients with PTC. Moreover, KDM1A was required for PTC cell migration and invasion. KDM1A knockdown inhibited the migration and invasive abilities of PTC cells both in vitro and in vivo. We also identified tissue inhibitor of metalloproteinase 1 (TIMP1) as a key KDM1A target gene. KDM1A activated matrix metalloproteinase 9 (MMP9) through epigenetic repression of TIMP1 expression by demethylating H3K4me2 at the TIMP1 promoter region. Rescue experiments clarified these findings. Altogether, we have uncovered a new mechanism of KDM1A repression of TIMP1 in PTC and suggest that KDM1A may be a promising therapeutic target in PTC.     

miR-329 suppresses the growth and motility of neuroblastoma by targeting KDM1A

MicroRNAs (miRNAs) act as key regulators of multiple cancers. miR-329 functions as a tumor suppressor in some malignancies. However, its role in neuroblastoma remains poorly understood. We found that miR-329 was decreased in metastatic tumor tissues compared with matched primary tumor tissues. Forced overexpression of miR-329 substantially suppressed cell proliferation, colony formation, migration, and invasion of neuroblastoma cells. Lysine-specific demethylase 1 (KDM1A) was found to be a target of miR-329. Furthermore, down-regulation of KDM1A by shRNA performed similar effects with overexpression of miR-329. Overexpression of KDM1A partially reversed the tumor suppressive effects of miR-329 in neuroblastoma cells. Collectively, miR-329 may suppress neuroblastoma cell growth and motility partially by targeting KDM1A.     

Circ_0005615 promotes cervical cancer cell growth and metastasis by modulating the miR-138-5p/KDM2A axis

Cervical cancer (CC) is a highly fatal gynecological malignancy due to its high metastasis and recurrence rate. Circular RNA (circRNA) has been regarded as a regulator of CC. However, the underlying molecular mechanism of circ_0005615 in CC remains unclear. The levels of circ_0005615, miR-138-5p, and lysine demethylase 2A (KDM2A) were measured using qRT-PCR or western blot. Cell proliferation was assessed by Cell Counting Kit-8, 5-ethynyl-2′-deoxyuridine, and colony formation experiments. Cell invasion and migration were tested by transwell assay and wound healing assay. Flow cytometry and Caspase-Glo 3/7 Assay kit were used to analyze cell apoptosis. The expression of proliferation-related and apoptosis-related markers was detected by western blot. The binding relationships among circ_0005615, miR-138-5p, and KDM2A were verified by dual-luciferase reporter assay or RNA immunoprecipitation assay. Xenograft assay was applied to detect the effect of circ_0005615 in vivo. Circ_0005615 and KDM2A were upregulated, while miR-138-5p was downregulated in CC tissues and cells. Circ_0005615 knockdown retarded cell proliferation, migration, and invasion, while promoting apoptosis. Besides, circ_0005615 sponged miR-138-5p, and miR-138-5p could target KDM2A. miR-138-5p inhibitor reversed the regulation of circ_0005615 knockdown on CC cell growth and metastasis, and KDM2A overexpression also abolished the inhibitory effect of miR-138-5p on CC cell growth and metastasis. In addition, we also discovered that circ_0005615 silencing inhibited CC tumor growth in vivo. Circ_0005615 acted as a tumor promoter in CC by regulating the miR-138-5p/KDM2A pathway.     

Upregulated KDM4B promotes prostate cancer cell proliferation by activating autophagy

Castration-resistant prostate cancer (CRPC) causes most of the deaths in patients with prostate cancer (PCa). The androgen receptor (AR) axis plays an important role in castration resistance. Emerging studies showed that the lysine demethylase KDM4B is a key molecule in AR signaling and turnover, and autophagy plays an important role in CRPC. However, little is known about whether KDM4B promotes CRPC progression by regulating autophagy. Here we used an androgen-independent LNCaP (LNCaP-AI) cell line to assay aberrant KDM4B expression using qPCR and western blot analysis and investigated the function of KDM4B in regulating cell proliferation. We found that KDM4B was markedly increased in LNCaP-AI cells compared with LNCaP cells. KDM4B level was significantly correlated with the Gleason score in PCa tissues. In vitro, KDM4B overexpression in CRPC cells promoted cell proliferation, whereas knockdown of KDM4B significantly inhibited cell proliferation. Upregulated KDM4B contributed to activate Wnt/β-catenin signaling and autophagy. Moreover, KDM4B activated autophagy by regulating the Wnt/β-catenin signaling. Finally, we demonstrated that autophagy inhibition attenuated KDM4B-induced CRPC cell proliferation. Our results provided novel insights into the function of KDM4B-driven CRPC development and indicated that KDM4B may be served as a potential target for CRPC therapy.     

Overexpression of let-7d explains down-regulated KDM3A and ENO2 in the pathogenesis of preeclampsia

Pre-eclampsia (PE) is the leading cause of maternal death; however, the causative molecular basis remains largely unknown. Recent studies have revealed the important role microRNAs (miRNAs) play in PE. We aimed to explore the effects of let-7d on trophoblast proliferation, migration, invasion and apoptosis in PE and its underlying mechanism. Placental tissues were collected from PE patients and healthy pregnant women, and it was found that let-7d expression was increased, while KDM3A and ENO2 expression was decreased in PE tissues and cells. Bioinformatics analysis indicated the interaction among let-7d, KDM3A and ENO2, confirmed by dual luciferase reporter gene assay; ChIP experiment identified methylated modification to ENO2 by KDM3A. With gain- and loss-function method, silencing of let-7d increased KDM3A expression and enhanced the binding between KDM3A and ENO2. Furthermore, overexpression of let-7d suppressed cell proliferation, migration and invasion of trophoblasts, and induced apoptosis of trophoblasts, while these capacities were restored upon additional treatment of overexpressed ENO2. PE rat models were established to explore the effects of let-7d and ENO2 on PE in vivo. The results established that the silencing of let-7d alleviated the tissue injury and PE-related symptoms when reducing urine protein, TUNEL-positive cells and increasing ENO2, and KDM3A expression in rats. Cumulatively, let-7d suppressed cell progression of trophoblasts, and induced apoptosis through the down-regulation of KDM3A to promote ENO2 methylation, thereby promoting progression of PE. Such an epigenetic network of let-7d, KDM3A and ENO2 in the pathogenesis of PE might provide novel insight into targeted therapy against this disorder.     

Long noncoding RNA POU3F3 promotes cancer cell proliferation in prostate carcinoma by upregulating rho-associated protein kinase 1

Long noncoding RNAs (lncRNAs) POU3F3 is overexpressed in esophageal squamous-cell carcinomas, while its role in other human cancers is unclear. In this study we found that POU3F3 and rho-associated protein kinase 1 (ROCK1) were both increased in tumor tissues than in adjacent healthy tissues of patients with prostate carcinoma. Expression levels of POU3F3 increased with increase in the diameter of tumor but were not significantly affected by lymph node metastasis or distant metastasis. Expression levels of POU3F3 and ROCK1 were positive correlated in tumor tissues but not in adjacent healthy tissues. POU3F3 and ROCK1 overexpression promoted, while ROCK1 knockdown inhibited the proliferation of prostate carcinoma cells. ROCK1 knockdown reduced the enhancing effect of POU3F3 overexpression on cancer cell proliferation. POU3F3 overexpression led to ROCK1 overexpression in prostate carcinoma cells, while ROCK1 overexpression did not significantly affect POU3F3 expression. Therefore, lncRNA POU3F3 may promote cancer cell proliferation in prostate carcinoma by upregulating ROCK1.     

Sumoylation of the histone demethylase KDM4A is required for binding to tumor suppressor p53 in HCT116 colon cancer cell lines

The histone demethylase lysine-specific demethylase 4A (KDM4A/Jmjd2A) has diverse functions, including involvement in gene regulation and cell cycle, and plays an oncogenic role in cancer cells. The modulation of KDM4A through post-translational modifications remains unclear. Here, we show that small ubiquitin-like modifier (SUMO) 1-mediated modification of KDM4A was required for interaction with tumor suppressor p53. Our data revealed that KDM4A is mainly sumoylated at lysine residue 471. However, the SUMO modification resulted in little change in subcellular localization, demethylase activity, or protein stability of KMD4A. Intriguingly, co-immunoprecipitation data revealed that sumoylation-defective mutants of KDM4A had a lower binding ability with p53 compared to that of wild-type KDM4A, suggesting a positive role for sumoylation in the interaction between KDM4A and p53. Together, these data suggest that KDM4A is post-translationally modified by SUMO, and this sumoylation may be a novel regulatory switch for controlling the interplay between KDM4A and p53.     

LINC00261 elevation inhibits angiogenesis and cell cycle progression of pancreatic cancer cells by upregulating SCP2 via targeting FOXP3

Long non-coding RNAs (lncRNAs) biological functions and molecular mechanisms associated with pancreatic cancer (PC) remain to be poorly elucidated. We aimed to clarify the role of lncRNA LINC00261 (LINC00261) in PC and confirm its regulatory mechanisms. Bioinformatics analysis, RNA pull-down and RIP assays were performed to investigate relationship between LINC00261 and forkhead box P3 (FOXP3). Further, dual-luciferase reporter gene and ChIP assays were employed to confirm the relationship among LINC00261, FOXP3 and sterol carrier protein-2 (SCP2). PC cells were introduced with a series of vectors to verify the effects of LINC00261 and SCP2 on the viability, cell cycle progression, migration and angiogenesis of PC cells. Nude mice with the xenograft tumour were used to evaluate the effects LINC00261 on the tumourigenicity. LINC00261 was lowly expressed in PC tissues and cells. SCP2 was inhibited by LINC00261 through FOXP3. Functionally, upregulated LINC00261 or downregulated SCP2 led to reduced cell viability, migration, angiogenesis and tumourigenicity potentials. This study demonstrated the inhibitory role of LINC00261 in the angiogenesis and cell cycle progression of PC cells. It acts through the negative regulation of SCP2 via targeting FOXP3. Findings in this study highlight a potentially biomarker for PC treatment.     

PELP1 is a reader of histone H3 methylation that facilitates oestrogen receptor‐α target gene activation by regulating lysine demethylase 1 specificity

Histone methylation has a key role in oestrogen receptor (ERα)‐mediated transactivation of genes. Proline glutamic acid and leucine‐rich protein 1 (PELP1) is a new proto‐oncogene that functions as an ERα co‐regulator. In this study, we identified histone lysine demethylase, KDM1, as a new PELP1‐interacting protein. These proteins, PELP1 and KDM1, were both recruited to ERα target genes, and PELP1 depletion affected the dimethyl histone modifications at ERα target genes. Dimethyl‐modified histones H3K4 and H3K9 are recognized by PELP1, and PELP1 alters the substrate specificity of KDM1 from H3K4 to H3K9. Effective demethylation of dimethyl H3K9 by KDM1 requires a KDM1–ERα–PELP1 functional complex. These results suggest that PELP1 is a reader of H3 methylation marks and has a crucial role in modulating the histone code at the ERα target genes.     

Inhibition of the histone demethylase KDM4B leads to activation of KDM1A,attenuates bacterial-induced pro-inflammatory cytokine release,and reduces osteoclastogenesis

Periodontal disease (PD) afflicts 46% of Americans with no effective adjunctive therapies available. While most pharmacotherapy for PD targets bacteria, the host immune response is responsible for driving tissue damage and bone loss in severe disease. Herein, we establish that the histone demethylase KDM4B is a potential drug target for the treatment of PD. Immunohistochemical staining of diseased periodontal epithelium revealed an increased abundance of KDM4B that correlates with inflammation. In murine calvarial sections exposed to Aggregatibacter actinomycetemcomitans lipopolysaccharide (Aa-LPS), immunohistochemical staining revealed a significant increase in KDM4B protein expression. The 8-hydroxyquinoline ML324 is known to inhibit the related demethylase KDM4E in vitro, but has not been evaluated against any other targets. Our studies indicate that ML324 also inhibits KDM4B (IC50: 4.9 μM), and decreases the pro-inflammatory cytokine response to an Aa-LPS challenge in vitro. Our results suggest that KDM4B inhibition-induced immunosuppression works indirectly, requiring new protein synthesis. In addition, fluorescence-stained macrophages exhibited a significant decrease in global monomethyl histone 3 lysine 4 (H3K4me) levels following an Aa-LPS challenge that was prevented by KDM4B inhibition, suggesting this effect is produced through KDM1A-mediated demethylation of H3K4. Finally, ML324 inhibition of KDM4B in osteoclast progenitors produced a significant reduction in Aa-LPS-induced osteoclastogenesis. These data link histone methylation with host immune response to bacterial pathogens in PD, and suggest a previously unreported, alternative mechanism for epigenetic control of the host inflammatory environment. As such, KDM4B represents a new therapeutic target for treating hyper-inflammatory diseases that result in bone destruction.