FOXD2-AS1 correlates with the malignant status and regulates cell proliferation,migration, and invasion in cutaneous melanoma

Long noncoding RNA (lncRNA) FOXD2 adjacent opposite strand RNA 1 (FOXD2-AS1) has been shown to be dysregulated in several types of human cancer. However, the role of FOXD2-AS1 in cutaneous melanoma was still unclear. In our study, FOXD2-AS1 expression has been found to be upregulated in cutaneous melanoma tissue specimens and cell lines compared with that in normal tissue specimens and normal human epidermal melanocyte, respectively. Furthermore, high expression of FOXD2-AS1 was obviously correlated with deep Breslow thickness, present ulceration, high Clark level and distant metastasis in cutaneous melanoma patients. However, there were no statistical associations between FOXD2-AS1 expression and cutaneous melanoma patients’ disease-free survival and overall survival. The results of loss-of-function study showed that inhibition of FOXD2-AS1 suppresses cutaneous melanoma cell proliferation, migration and invasion through regulating phospho-Akt expression. In conclusion, FOXD2-AS1 is associated with clinical progression in cutaneous melanoma patients, and functions as oncogenic lncRNA in cutaneous melanoma cells.     

PSF3 marks malignant colon cancer and has a role in cancer cell proliferation

PSF3 (partner of Sld five 3) is a member of the tetrameric complex termed GINS, composed of SLD5, PSF1, PSF2, and PSF3, and well-conserved evolutionarily. Previous studies suggested that some GINS complex members are upregulated in cancer, but PSF3 expression in colon carcinoma has not been investigated. Here, we established a mouse anti-PSF3 antibody, and examined PSF3 expression in human colon carcinoma cell lines and colon carcinoma specimens. We found that PSF3 is expressed in the crypt region in normal colonic mucosa and that many PSF3-positive cells co-expressed Ki-67. This suggests that PSF3-positivity of normal mucosa is associated with cell proliferation. Expression of the PSF3 protein was greater in carcinoma compared with the adjacent normal mucosa, and even stronger in high-grade malignancies, suggesting that it may be associated with colon cancer progression. PSF3 gene knock-down in human colon carcinoma cell lines resulted in growth inhibition characterized by delayed S-phase progression. These results suggest that PSF3 is a potential biomarker for diagnosis of progression in colon cancer and could be a new target for cancer therapy.     

Involvement of p54(nrb), a PSF partner protein,in DNA double-strand break repair and radioresistance

Mammalian cells repair DNA double-strand breaks (DSBs) via efficient pathways of direct, nonhomologous DNA end joining (NHEJ) and homologous recombination (HR). Prior work has identified a complex of two polypeptides, PSF and p54(nrb), as a stimulatory factor in a reconstituted in vitro NHEJ system. PSF also stimulates early steps of HR in vitro. PSF and p54(nrb) are RNA recognition motif-containing proteins with well-established functions in RNA processing and transport, and their apparent involvement in DSB repair was unexpected. Here we investigate the requirement for p54(nrb) in DSB repair in vivo. Cells treated with siRNA to attenuate p54(nrb) expression exhibited a delay in DSB repair in a γ-H2AX focus assay. Stable knockdown cell lines derived by p54(nrb) miRNA transfection showed a significant increase in ionizing radiation-induced chromosomal aberrations. They also showed increased radiosensitivity in a clonogenic survival assay. Together, results indicate that p54(nrb) contributes to rapid and accurate repair of DSBs in vivo in human cells and that the PSF·p54(nrb) complex may thus be a potential target for radiosensitizer development.     

Prognostic significance of lncRNA DANCR expression in human cancers: a systematic review and meta-analysis

Several studies demonstrated that lncRNA differentiation antagonizing non-protein coding RNA (lncRNA DANCR) expression might have the potential capacity to predict the cancer prognosis; however, definite conclusion has not been obtained. The aim of this meta-analysis was to evaluate the prognostic value of lncRNA DANCR expression in cancers. PubMed, Web of Science, Scopus, and Embase were comprehensively searched for relevant studies. Studies meeting all inclusion standards were included into this meta-analysis. The analysis of overall survival (OS), disease-free survival (DFS), or clinicopathological features was conducted. Total 11 studies containing 1154 cancer patients were analyzed in this meta-analysis. The results showed, compared with low lncRNA DANCR expression, high lncRNA DANCR expression was significantly associated with shorter OS (hazard ratio [HR] = 1.85; 95% CI = 1.52–2.26; P<0.01) and DFS (HR = 1.82; 95% CI = 1.43–2.32; P<0.01) in cancers. Besides, high lncRNA DANCR expression predicted deeper tumor invasion (P<0.01), earlier lymph node metastasis (P<0.01), earlier distant metastasis (P<0.01), and more advanced clinical stage (P<0.01) compared with low lncRNA DANCR expression in cancer populations. High lncRNA DANCR expression was associated with worse prognosis compared with low lncRNA DANCR expression in cancers. LncRNA DANCR expression could serve as a prognostic factor of human cancers.     

LncRNA SRA mediates cell migration,invasion, and progression of ovarian cancer via NOTCH signaling and epithelial–mesenchymal transition

Long non-coding RNA (lncRNA) is a newly identified regulator of tumor formation and tumor progression. The function and expression of lncRNAs remain to be fully elucidated, but recent studies have begun to address their importance in human health and disease. The lncRNA, SRA, known as steroid receptor activator, acts as an important modulator of gynecological cancer, and its expression may affect biological functions including proliferation, apoptosis, steroid formation, and muscle development. However, it is still not well known whether SRA is involved in the regulation of ovarian cancer. The present study investigated the molecular function and association between SRA expression and clinicopathological factors. In ovarian cancer cell lines, SRA knockdown and overexpression regulated cell migration, proliferation, and invasion. Both in vivo and in vitro experiments using knockdown and overexpression showed that SRA potently regulated epithelial–mesenchymal transition (EMT) and NOTCH pathway components.Further, clinical data confirmed that SRA was a significant predictor of overall survival (OS) and progression-free survival and patients with ovarian cancer exhibiting high expression of SRA exhibited higher recurrence rates than patients with low SRA expression. In conclusion, the present study indicates that SRA has clinical significance as its expression can predict the prognosis of ovarian cancer patients. High expression of the lncRNA SRA is strongly correlated with recurrence-free survival of ovarian cancer patients.     

Human metastatic melanoma cell lines express high levels of growth hormone receptor and respond to GH treatment

Accumulating evidence implicates the growth hormone receptor (GHR) in carcinogenesis. While multiple studies show evidence for expression of growth hormone (GH) and GHR mRNA in human cancer tissue, there is a lack of quantification and only a few cancer types have been investigated. The National Cancer Institute’s NCI60 panel includes 60 cancer cell lines from nine types of human cancer: breast, CNS, colon, leukemia, melanoma, non-small cell lung, ovarian, prostate and renal. We utilized this panel to quantify expression of GHR, GH, prolactin receptor (PRLR) and prolactin (PRL) mRNA with real-time RT qPCR. Both GHR and PRLR show a broad range of expression within and among most cancer types. Strikingly, GHR expression is nearly 50-fold higher in melanoma than in the panel as a whole. Analysis of human metastatic melanoma biopsies confirmed GHR gene expression in melanoma tissue. In these human biopsies, the level of GHR mRNA is elevated in advanced stage IV tumor samples compared to stage III. Due to the novel finding of high GHR in melanoma, we examined the effect of GH treatment on three NCI60 melanoma lines (MDA-MB-435, UACC-62 and SK-MEL-5). GH increased proliferation in two out of three cell lines tested. Further analysis revealed GH-induced activation of STAT5 and mTOR in a cell line dependent manner. In conclusion, we have identified cell lines and cancer types that are ideal to study the role of GH and PRL in cancer, yet have been largely overlooked. Furthermore, we found that human metastatic melanoma tumors express GHR and cell lines possess active GHRs that can modulate multiple signaling pathways and alter cell proliferation. Based on this data, GH could be a new therapeutic target in melanoma.     

LncRNA TDRG1 enhances tumorigenicity in endometrial carcinoma by binding and targeting VEGF-A protein

Endometrial carcinoma is one of the most frequently diagnosed cancers in females. Long non-coding RNAs (lncRNAs) have been associated with cancer; its role in endometrial carcinoma is an emerging area of research. In this article, lncRNA TDRG1 expression in human endometrial carcinoma tissues and normal endometrial tissues was quantified by qRT-PCR. LncRNA TDRG1 was overexpressed or knocked-down in neither HEC-1B nor Ishikawa endometrial carcinoma cells, respectively, to assess cellular phenotype and expression of related molecules. Our results showed that lncRNA TDRG1 was significantly overexpressed in endometrial carcinoma tissues. Overexpression of lncRNA TDRG1 promoted endometrial carcinoma cell viability, invasion and migratory ability, inhibited apoptosis, and upregulated VEGF-A, PI3K, Bcl-2, MMP2 and survivin; knockdown of lncRNA TDRG1 had the opposite effects. LncRNA TDRG1 overexpression increased tumorigenicity in vivo and was associated with the upregulation of VEGF-A. RNA binding protein immunoprecipitation (RIP) assays confirmed that lncRNA TDRG1 directly binds to VEGF-A protein. Furthermore, knockdown of VEGFA in lncRNA TDRG1-overexpressing endometrial carcinoma cells reversed the effects of lncRNA TDRG1 on cell proliferation, invasion, migration and apoptosis. In conclusion, lncRNA TDRG1 may promote endometrial carcinoma cell proliferation and invasion by positively targeting VEGF-A and modulating relative genes.     

Variability in Melanoma Metalloproteinase Expression Profiling

The proteolytic activities of a disintegrin and metalloproteinase (ADAM); a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS), and matrix metalloproteinase (MMP) families play important roles in normal and multiple pathological conditions. These metalloproteases have potential roles in the degradation of the ECM and in the processing of bioactive molecules. In the present study, RNA was isolated from multiple normal fibroblast and metastatic melanoma cell lines, as well as the isogenic normal tissue and tumor samples, and the gene expression levels of six ADAMs, eight MMPs, and four ADAMTSs were analyzed by real-time PCR. This approach allowed for detected changes in mRNA expression of the individual metalloproteinase genes to be compared between normal and metastatic states and also between tissue and cultured cells. Increased gene expression of several ADAM and MMP family members (MMP1, MMP8, MMP15, and ADAM15) occurred in melanoma tissue and was replicated in tissue cultures. In general, the level of ADAM and MMP mRNA expression was several-fold higher in cultured cells compared with the isogenic tissue from which they were derived. Passage-dependent expression patterns were observed for MMP8 and MMP9 in in-house-derived metastatic melanoma cell lines. This reiterates earlier suggestions that experiments using cells that have been maintained in culture should be interpreted with great care.     

MicroRNA signatures differentiate melanoma subtypes

Melanoma is an aggressive cancer that is highly resistance to therapies once metastasized. We studied microRNA (miRNA) expression in clinical melanoma subtypes and evaluated different miRNA signatures in the background of gain of function somatic and inherited mutations associated with melanoma. Total RNA from 42 patient derived primary melanoma cell lines and three independent normal primary melanocyte cell cultures was evaluated by miRNA array. MiRNA expression was then analyzed comparing subtypes and additional clinicopathologic criteria including somatic mutations. The prevalence and association of an inherited variant in a miRNA binding site in the 3′UTR of the KRAS oncogene, referred to as the KRAS-variant, was also evaluated. We show that seven miRNAs, miR-142-3p, miR-486, miR-214, miR-218, miR-362, miR-650 and miR-31, were significantly correlated with acral as compared to non-acral melanomas (p < 0.04). In addition, we discovered that the KRAS-variant was enriched in non-acral melanoma (25%), and that miR-137 under expression was significantly associated with melanomas with the KRAS-variant. Our findings indicate that miRNAs are differentially expressed in melanoma subtypes and that their misregulation can be impacted by inherited gene variants, supporting the hypothesis that miRNA misregulation reflects biological differences in melanoma.Key words: melanoma, microRNA profiling, biomarker, acral, KRAS-variant, SNP     

Adiponectin AS lncRNA inhibits adipogenesis by transferring from nucleus to cytoplasm and attenuating Adiponectin mRNA translation

Adiponectin (AdipoQ) is an adipocyte-derived hormone with positive function on systemic glucose and lipid metabolism. Long noncoding RNA (lncRNA) is emerging as a vital regulator of adipogenesis. However, AdipoQ-related lncRNAs in lipid metabolism have not been explored. Here, AdipoQ antisense (AS) lncRNA was first identified, and we further found that it inhibited adipogenesis. The half-life of AdipoQ AS lncRNA was 10?h, whereas that of AdipoQ mRNA was 4?h. During adipogenic differentiation, AdipoQ AS lncRNA translocated from nucleus to cytoplasm. AdipoQ AS lncRNA and AdipoQ mRNA formed an RNA duplex. Moreover, AdipoQ AS lncRNA delivered via injection of adenovirus expressing AdipoQ AS lncRNA decreases white adipose tissue (WAT), brown adipose tissue (BAT) and liver triglycerides (TG) in mice consuming a high fat diet (HFD). Interestingly, the non-overlapping region of AdipoQ AS lncRNA improved serum glucose tolerance and insulin sensitivity in HFD mice, but not AdipoQ AS lncRNA. In conclusion, AdipoQ AS lncRNA transfer from nucleus to cytoplasm inhibits adipogenesis through formation of an AdipoQ AS lncRNA/AdipoQ mRNA duplex to suppress the translation of AdipoQ mRNA. Taken together, we suggest that AdipoQ AS lncRNA is a novel therapeutic target for obesity-related metabolic diseases.     

Long noncoding RNA DGCR5 suppresses gastric cancer progression by acting as a competing endogenous RNA of PTEN and BTG1

Long noncoding RNA (lncRNA) DiGeorge syndrome critical region gene 5 (DGCR5) has been reported to correlate with a variety of cancers, with its expression pattern and potential mechanism not clarified in gastric cancer (GC). In this study, we demonstrated that DGCR5 was downregulated in cancerous tissues and plasma samples from patients with GC, and its downregulation was associated with advanced TNM stage and positive lymphatic metastasis. Plasma DGCR5 had an area under the receiver operating characteristic curve (AUC) of 0.722 for diagnosis of GC. Gain- and loss-of-function of DGCR5 revealed that DGCR5 functioned as a competing endogenous RNA for miR-23b to suppress GC cell proliferation, invasion and migration, and facilitate apoptosis by regulating PTEN and BTG1 in vitro. Furthermore, the overexpression of DGCR5 suppressed tumor growth, and inhibited the expression of miR-23b and proliferation antigen Ki-67, but increased the expression of PTEN and BTG1 in vivo. In conclusion, our results show that DGCR5 is a tumor-suppressive lncRNA that regulates PTEN and BTG1 expression through directly binding to miR-23b. This mechanism may contribute to a better understanding of GC pathogenesis and provide a potential therapeutic strategy for GC.     

NVP-LDE225, a Potent and Selective SMOOTHENED Antagonist Reduces Melanoma Growth In Vitro and In Vivo

Melanoma is one of the most aggressive cancers and its incidence is increasing worldwide. So far there are no curable therapies especially after metastasis. Due to frequent mutations in members of the mitogen-activated protein kinase (MAPK) signaling pathway, this pathway is constitutively active in melanoma. It has been shown that the SONIC HEDGEHOG (SHH)-GLI and MAPK signaling pathway regulate cell growth in many tumors including melanoma and interact with each other in the regulation of cell proliferation and survival.Here we show that the SHH-GLI pathway is active in human melanoma cell lines as they express downstream target of this pathway GLI1. Expression of GLI1 was significantly higher in human primary melanoma tissues harboring BRAF^V600E mutation than those with wild type BRAF. Pharmacologic inhibition of BRAF^V600E in human melanoma cell lines resulted in decreased expression of GLI1 thus demonstrating interaction of SHH-GLI and MAPK pathways. Inhibition of SHH-GLI pathway by the novel small molecule inhibitor of smoothened NVP-LDE225 was followed by inhibition of cell growth and induction of apoptosis in human melanoma cell lines, interestingly with both BRAF^V600E and BRAF^{Wild Type} status. NVP-LDE225 was potent in reducing cell proliferation and inducing tumor growth arrest in vitro and in vivo, respectively and these effects were superior to the natural compound cyclopamine.Finally, we conclude that inhibition of SHH-GLI signaling pathway in human melanoma by the specific smoothened inhibitor NVP-LDE225 could have potential therapeutic application in human melanoma even in the absence of BRAF^V600E mutation and warrants further investigations.     

长非编码RNA SNHG18对人胃癌细胞BGC823增殖和凋亡的影响

目的:探究长非编码RNA SNHG18对胃癌细胞增殖和凋亡的影响。方法:采用实时定量PCR(qRT-PCR)技术检测人胃癌组织及癌旁组织和胃癌细胞系中lncRNA SNHG18的表达;采用MTT和克隆形成试验观察转染SNHG18过表达质粒后胃癌细胞BGC823增殖活力的变化;通过流式细胞术检测lncRNA SNHG18对胃癌细胞BGC823凋亡的影响。结果:相较于癌旁组织和胃正常粘膜上皮细胞系GSE-1,胃癌组织及胃癌细胞系中SNHG18的表达水平显著降低(P0.05);胃癌细胞过表达SNHG18增殖活力以及克隆形成的能力均显著降低(P0.05),而细胞凋亡率明显升高(P0.05)。结论:胃癌组织中长非编码RNA SNHG18呈低表达,可促进胃癌细胞增殖并抑制其凋亡,可能在胃癌发生发展过程中发挥重要作用。     

LncRNA CRNDE promotes the progression and angiogenesis of pancreatic cancer via miR-451a/CDKN2D axis

BackgroundThe lncRNA colorectal neoplasia differentially expressed (lncRNA CRNDE) has been reported to play a pivotal role in various cancers. However, the expression and function of CRNDE in pancreatic cancer remain unclear. The objective of this study was to investigate the effects of CRNDE on pancreatic cancer and the underlying mechanisms.MethodsThe expression of CRNDE in pancreatic cancer tissues and cell lines was determined by RT-qPCR. Proliferation and angiogenesis were detected by MTT, colony formation, transwell and tube formation assays in vitro and in vivo. ELISA assay was used to detect the secretion of VEGFA. IHC was performed to test the expression levels of Ki67 and CD31. The binding sites between CRNDE, CDKN2D and miR-451a were predicted by bioinformatics analysis. Dual luciferase reporter and RNA immunoprecipitation assays were conducted to confirm the interaction with each other.ResultsThe results showed that CRNDE was significantly up-regulated in pancreatic cancer tissues as well as cell lines. CRNDE overexpression promoted the progression and angiogenesis of pancreatic cancer cells in vitro and in vivo. Moreover, we identified that CRNDE functioned as a sponge for miR-451a and CRNDE overexpression inhibited the expression of miR-451a. Furthermore, we confirmed that miR-451a directly interacted with CDKN2D and negatively regulated CDKN2D expression. In addition, CRNDE was found to positively regulate CDKN2D expression and mediate pancreatic cancer cell proliferation and angiogenesis through miR-451a/CDKN2D axis.ConclusionCRNDE modulates cell proliferation and angiogenesis via miR-451a/CDKN2D axis in pancreatic cancer, which provides a potential therapeutic target for pancreatic cancer treatment.