Promotes Melanoma Progression Research Articles

LncRNA NEAT1 facilitates melanoma cell proliferation, migration, and invasion via regulating miR-495-3p and E2F3.

Melanoma contributes a lot to skin cancer-related deaths. lncRNAs are implicated in various diseases, including melanoma. lncRNA NEAT1 is frequently dysregulated and can play important roles in multiple cancers. Nevertheless, little has been studied about the function of NEAT1 in melanoma progression. In our present research, we displayed NEAT1 was overexpressed in melanoma cells. A series of functional assays showed that overexpression of NEAT1 promoted the proliferation, migration, and invasion of melanoma cells. By contrast, NEAT1 knockdown obviously restrained melanoma cell progression. Mechanistically, it was revealed that NEAT1 could directly bind with miR-495-3p, which led to a negative effect on miR-495-3p levels. In addition, miR-495-3p was significantly decreased in melanoma cells. Furthermore, E2F3 was postulated as the target of miR-495-3p and overexpression of this miR could suppress the levels of E2F3. Meanwhile, it was exhibited that melanoma cell proliferation, migration, and invasion induced by E2F3 silence was abrogated by miR-495-3p. Moreover, an in vivo xenograft nude mice model was established using A375 cells and it was indicated that NEAT1 promoted melanoma progression in vivo via regulating the miR-495-3p/E2F3 axis. In conclusion, we suggest that NEAT1 exerts an oncogenic effect on melanoma development via inhibition of miR-495-3p and induction of E2F3. NEAT1 might serve as a crucial prognostic biomarker of melanoma.

MiR-10b promoted melanoma progression through Wnt/β-catenin pathway by repressing ITCH expression

Dysregulation of microRNAs (miRNAs) have been reported to contribute to malignant progression in melanoma. However, the roles and mechanisms of several miRNAs in melanoma remain poorly understood. In our study, we showed that miR-10b was significantly up-regulated in melanoma tissues and cell lines, and was associated with overall survival of melanoma patients. Inhibition of miR-10b dramatically suppressed melanoma cell proliferation, migration and invasion in vitro and inhibited tumor growth in vivo. Moreover, we defined ITCH as a direct and functional downstream target of miR-10b, and showed that there was an inverse correlation between the expression of ITCH and miR-10b on melanoma tissues. Down-regulation of ITCH partially attenuated the inhibitory effects of miR-10b inhibition on melanoma cell proliferation, migration and invasion. Furthermore，we found that miR-10b exerted its effects on melanoma by regulating the Wnt/β-catenin pathway. Taken together, our results demonstrated that miR-10b was an important epigenetic modifier, promoting melanoma progression through regulating ITCH/Wnt/β-catenin pathway. These results offer a new strategy for epigenetic cancer therapy.

TRIM44 activates the AKT/mTOR signal pathway to induce melanoma progression by stabilizing TLR4

BackgroundThere is growing evidence that tripartite motif-containing protein 44 (TRIM44) plays crucial role in tumor development. However, the underlying mechanism of this deubiquitinating enzyme remains unclear.MethodsLarge clinical samples were used to detect TRIM44 expression and its associations with clinicopathological features and prognosis. Gain- and loss-of-function experiments in cell lines and mouse xenograft models were performed to elucidate the function and underlying mechanisms of TRIM44 induced tumor progression. Co-immunoprecipitation (Co-IP) assays and mass spectrometric analyses were applied to verify the interacting proteins of TRIM44.ResultsWe found that TRIM44 was commonly amplified in melanoma tissues compared with paratumoral tissues. TRIM44 expression also positively correlated with more aggressive clinicopathological features, such as Breslow depth (p = 0.025), distant metastasis (p = 0.012), and TNM stage (p = 0.002). Importantly, we found that TRIM44 was an independent indicator of prognosis for melanoma patients. Functionally, overexpression of TRIM44 facilitated cell invasion, migration, apoptosis resistance and proliferation in vitro, and promoted lung metastasis and tumorigenic ability in vivo. Importantly, high level of TRIM44 induced melanoma cell epithelial-mesenchymal transition (EMT), which is one of the most important mechanisms for the promotion of tumor metastasis. Mechanistically, high levels of TRIM44 increased the levels of p-AKT (T308) and p-mTOR (S2448), and a specific AKT inhibitor inhibited TRIM44-induced tumor progression. Co-IP assays and mass spectrometric analyses indicated that TRIM44 overexpression induces cell EMT through activating AKT/mTOR pathway via directly binding and stabilizing TOLL-like receptor 4 (TLR4), and TLR4 interference impeded TRIM44 induced tumor progression. Moreover, we demonstrated that TRIM44 is the target of miR-26b-5p, which is significantly downregulated in melanoma tissues and may be responsible for the overexpression of TRIM44.ConclusionsTRIM44, regulated by miR-26b-5p, promotes melanoma progression by stabilizing TLR4, which then activates the AKT/mTOR pathway. TRIM44 shows promise as a prognostic predictor and a therapeutic target for melanoma patients.

Downregulation of RNF128 activates Wnt/\u03b2-catenin signaling to induce cellular EMT and stemness via CD44 and CTTN ubiquitination in melanoma

BackgroundRing finger proteins (RNFs) were involved in carcinogenesis. Here, we aimed to explore the detailed mechanism of RNF128 in the progression of melanoma.MethodsWe reanalyzed several gene expression profiles from the Gene Expression Omnibus (GEO) database and obtained the overlapped differential expressed RNF genes. Among them, RNF128 was selected to further explore its expression, the biological significance, and the underlying molecular mechanism, as well as the clinical relevance in melanoma patients.ResultsRNF128 was found to be significantly downregulated in the selected datasets, which was further verified in our melanoma tissues. Moreover, RNF128 downregulation was shown to correlate with the malignant phenotype of melanoma, and further functional assays demonstrated that low levels of RNF128 promoted melanoma progression via inducing cell epithelial-mesenchymal transition (EMT) and the acquisition of stemness. Mechanistically, RNF128 interference activated the Wnt pathway via simultaneously ubiquitinating CD44/cortactin (CTTN), resulting in CD44 and c-Myc transcription, thus revealed that RNF128 participated in a positive feedback of the Wnt pathway-CD44 loop. Clinically, we found that patients expressing low RNF128 and high CD44/CTTN levels had a poor prognosis.ConclusionDownregulated RNF128 activates Wnt signaling to induce cellular EMT and stemness by ubiquitinating and degrading CD44/CTTN, and RNF128 is a reliable diagnostic and prognostic biomarker, and a deeper understanding of RNF128 may contribute to the treatment of melanoma.

Elevated nuclear auto-antigenic sperm protein promotes melanoma progression by inducing cell proliferation.

BackgroundNuclear auto-antigenic sperm protein (NASP) has been implicated in tumorigenesis. However, its role in melanoma is still unclear.Materials and methodsIn the present study, we detected the mRNA and protein level of NASP in melanoma cell lines and tissues. Then the role of NASP was investigated by transfecting with NASP siRNAs. Finally, the prognosis of NASP was analyzed in 100 melanoma patients through Cox regression and Kaplan–Meier analyses.ResultsWe showed that NASP was significantly overexpressed in melanoma tissues, and unregulated NASP promoted melanoma cell proliferation via promoting cell cycle G1/S phase transition. Additionally, the expression of NASP was closely related to proliferating cell nuclear antigen, a widely accepted biomarker for cell proliferation. Clinically, we found that a high level of NASP predicated poor overall survival and high cumulative recurrence rates. Multivariate analysis revealed that NASP was a risk biomarker for predicting the prognosis of melanoma patients.ConclusionElevated NASP plays an important role in melanoma cell proliferation and tumor progression, and it can be used as an independent prognostic biomarker for melanoma patients.

Autophagy-independent p62 in metastatic melanoma

By pairing up with RNA-binding proteins, p62 promotes melanoma progression.

Negative Regulatory Loop between Microphthalmia-Associated Transcription Factor (MITF) and Notch Signaling.

Melanoma, a melanocyte-origin neoplasm, is a highly metastatic and treatment-resistance cancer. While it is well established that notch signaling activation promotes melanoma progression, little is known about the reciprocal interactions between Notch signaling and melanoma-specific pathways. Here we reveal a negative regulatory loop between Notch signaling and microphthalmia-associated transcription factor (MITF), the central regulator of melanoma progression and the driver of melanoma plasticity. We further demonstrate that Notch signaling activation, in addition to the known competition-based repression mechanism of MITF transcriptional activity, inhibits the transcription of MITF, leading to a decrease in MITF expression. We also found that MITF binds to the promoter of the gene encoding the master regulator of Notch signaling, recombination signal binding protein J kappa (RBPJK), leading to its upregulation. Our findings suggest that, once activated, Notch signaling represses MITF signaling to maintain the melanoma invasiveness and metastatic phenotype.

The FOXC2 Transcription Factor Promotes Melanoma Outgrowth and Regulates Expression of Genes Associated With Drug Resistance and Interferon Responsiveness.

The FOXC2 transcription factor promotes the progression of several cancer types, but has not been investigated in the context of melanoma cells. To study FOXC2's influence on melanoma progression, we generated a FOXC2-deficient murine melanoma cell line and evaluated The Cancer Genome Atlas (TCGA) patient datasets. We compared tumor growth kinetics and RNA-seq/qRT-PCR gene expression profiles from wild-type versus FOXC2-deficient murine melanomas. We also performed Kaplan-Meier survival analysis of TCGA data to assess the influence of FOXC2 gene expression on melanoma patients' response to chemotherapy and immunotherapy. FOXC2 promotes melanoma progression and regulates the expression of genes associated with multiple oncogenic pathways, including the oxidative stress response, xenobiotic metabolism, and interferon responsiveness. FOXC2 expression in melanoma correlates negatively with patient response to chemotherapy and immunotherapy. FOXC2 drives a tumor-promoting gene expression program in melanoma and is a prognostic indicator of patient response to multiple cancer therapies.

Long non-coding RNA ZEB1-AS1 indicates poor prognosis and promotes melanoma progression through targeting miR-1224-5p.

Long non-coding RNAs (lncRNAs) have critical roles in various types of cancer, but their roles in the development of melanoma and the underlying molecular mechanisms remain to be fully elucidated. In the present study, the role of zinc finger E-box binding homeobox 1 antisense RNA 1 (ZEB1-AS1) in melanoma was assessed. The expression levels of ZEB1-AS1 were increased in melanoma cell lines and tumor tissues as indicated by reverse transcription-quantitative polymerase chain reaction analysis. Kaplan-Meier analysis suggested that higher expression of ZEB1-AS1 predicts poor prognosis of melanoma patients. Furthermore, knockdown of ZEB1-AS1 inhibited the proliferation, migration and invasion of melanoma, suggesting a role of ZEB1-AS1 in the development and progression of melanoma. In addition, a luciferase reporter assay confirmed that the expression of miR-1224-5p was directly regulated by ZEB1-AS1. Transfection with miR-1224-5p mimics reduced the levels of ZEB1-AS1 and the proliferation, migration and invasion of melanoma. In conclusion, ZEB1-AS1 enhances the proliferation, migration and invasion of melanoma, at least in part by inhibiting the expression of miR-1224-5p, and its overexpression is associated with poor survival of melanoma patients. In addition, the ZEB1-AS1/miR-1224-5p interaction may be a promising therapeutic target for melanoma treatment.

MiR-25 Promotes Melanoma Progression by regulating RNA binding motif protein 47.

Melanoma is the most aggressive skin cancer, and accounts for the major part of skin cancer-related deaths in the world. In addition, the underlying mechanism of tumor progression in melanoma remains far from being elucidated. In this study, we have evaluated the function of miR-25 in melanoma. First, we examined the expression of miR-25 in four melanoma cell lines (A875, MV3, M14 and uacc-257) and in a normal melanocyte cell line (HEM-a). Then, we overexpressed miR-25 in M14 cells. Our results show that miR-25 promotes M14 cell proliferation and migration. We found that miR-25 up-regulates the PI3K/Akt/mTOR signaling pathway in these tumor cells. Furthermore, a luciferase-based reporter gene assay showed that miR-25 could directly target the RNA-binding motif protein 47 (RBM47). Taken together, our findings suggest that RBM47 is a promising target for the treatment of melanoma.

MicroRNA-373 promotes cell migration via targeting salt-inducible kinase 1 expression in melanoma.

It is well established that altered expression of microRNAs (miRs) is critical in numerous human cancer types. Nevertheless, the molecular mechanisms of many miRs are yet to be elucidated. In the present study, reverse transcription-quantitative polymerase chain reaction and western blot analyses, and cell migration assays were performed to verify dysregulation of miR-373 in melanoma and its biological function. The transcriptional level of miR-373 was identified to be upregulated in melanoma tissues and cell lines compared with nevus and normal melanocytes. miR-373 was identified to function as an oncomiR, promoting melanoma cell migration. Notably, miR-373 was observed to suppress its downstream gene salt-inducible kinase 1 (SIK1) through directly binding the 3'-untranslated region of SIK1 expression. Furthermore, reduced SIK1 expression was identified to be responsible for the oncogenic effect of miR-373. In conclusion, the present study indicates that miR-373 functions as an oncomiR to promote melanoma progression through targeting SIK1 expression. This may provide a new therapeutic approach for melanoma.

Long non-coding RNA CASC15 promotes melanoma progression by epigenetically regulating PDCD4

BackgroundLong non-coding RNAs (LncRNAs) have been identified as critical regulators in a variety of cancer types. Cancer susceptibility candidate 15 (CASC15), a lncRNA located at chromosome 6p22.3, has been discovered to participate in melanoma progression and phenotype switching. Nevertheless, the roles and molecular mechanisms of CASC15 in melanoma are far from being understood.ResultsWe found that CASC15 expression was up-regulated in melanoma tissues and associated with advanced pathological stages. Function experiments displayed that CASC15 knockdown hindered proliferation, facilitated apoptosis and suppressed invasion, while CASC15 overexpression facilitated proliferation and invasion in melanoma cells. Further mechanistic analysis showed that CASC15 epigenetically silenced the expression of programmed cell death 4 (PDCD4) by recruiting EZH2 and increasing H3K27me3 level at the promoter region of PDCD4. Additionally, PDCD4 overexpression inhibited proliferation, enhanced apoptosis and decreased invasion of melanoma cells. Moreover, CASC15-knockdown-induced anti-cancer effects were abated by PDCD4 down-regulation. Furthermore, depletion of CASC15 blocked tumor growth of melanoma by up-regulating PDCD4 in vivo.ConclusionsCASC15 acts as an oncogene by negatively regulating PDCD4 expression via recruiting EZH2 and subsequently increasing H3K27me3 level. Together, our study indicates that CASC15/EZH2/PDCD4 may serve as a promising therapeutic target for melanoma intervention.

The Ectodysplasin receptor EDAR acts as a tumor suppressor in melanoma by conditionally inducing cell death.

Ectodysplasin receptor EDAR is seen as a typical Tumor Necrosis Factor receptor (TNFR) family member known to interact with its ligand Eda-A1, and signaling mainly through the nuclear factor-kappaB (NF-κB) and c-jun N-terminal kinases pathways. Mutations in genes that encode proteins involved in EDAR transduction cascade cause anhidrotic ectodermal dysplasia. Here, we report an unexpected pro-apoptotic activity of EDAR when unbound to its ligand Eda-A1, which is independent of NF-κB pathway. Contrarily to other death receptors, EDAR does recruit caspase-8 to trigger apoptosis but solely upon ligand withdrawal, thereby behaving as the so-called dependence receptors. We propose that pro-apoptotic activity of unbound EDAR confers it a tumor suppressive activity. Along this line, we identified loss-of-pro-apoptotic function mutations in EDAR gene in human melanoma. Moreover, we show that the invalidation of EDAR in mice promotes melanoma progression in a B-Raf mutant background. Together, these data support the view that EDAR constrains melanoma progression by acting as a dependence receptor.

Critical role for non‑GAP function of Gαs in RGS1‑mediated promotion of melanoma progression through AKT and ERK phosphorylation.

Regulator of G‑protein signaling1 (RGS1) has been found to be a critical factor in melanoma and other malignancies. However, the mechanism involved in the RGS1‑mediated promotion of melanoma progression is not clear. We based our study on samples collected from pathological specimens of melanoma patients. We found by immunohistochemistry that RGS1 expression was significantly higher in melanoma than that noted in nevus tissue (P<0.05). Kaplan‑Meier analysis demonstrated a significant correlation between increased RGS1 expression and reduced disease‑specific survival (P<0.05). RGS1 expression was also found to be related to the proliferation and migration of melanoma cells. RGS1 was able to bind to the Gαs in immunoprecipitation, but this interaction did not accelerate GTP hydrolysis in our experiment. Furthermore, we found that RGS1 may promote melanoma progression through the downstream effects of Gαs signaling, such as the increased phosphorylation of AKT and ERK by western blotting. Our results demonstrated that RGS1 promotes melanoma progression through regulation of Gαs‑mediated inactivation of AKT and ERK. Therefore, RGS1 is a novel therapeutic target for melanoma treatment.

Postdiagnosis aspirin use and overall survival in patients with melanoma

Mouse studies show that tumor-derived prostaglandins and platelets promote melanoma progression and immune evasion. Determine whether aspirin confers longer survival in patients with melanoma. A retrospective cohort study of 1522 patients at Indiana University Health who had melanoma diagnosed between 2000 and 2014 and were followed up through September 2016. Aspirin use was associated with longer overall survival in univariate analysis and after controlling for age, sex, stage, and treatment modalities (hazard ratio [HR], 0.58; 95% confidence interval [CI], 0.45-0.75). Aspirin use was not associated with survival in patients with in situ and stage I melanoma but was associated with better survival in stages II (HR, 0.45; 95% CI, 0.24-0.82) and III (HR, 0.57; 95% CI; 0.34-0.96). No statistical significance was observed in stage IV patients (HR, 0.55; 95% CI, 0.27-1.13). Inturn, melanoma in patients using aspirin before diagnosis was less likely to be diagnosed in stages III orIV. Observational study. Aspirin could provide a survival advantage in melanoma. Clinical trials investigating the therapeutic potential of aspirin are warranted.

Melanocyte-specific gene 1 promotes melanoma progression by enhancing the expression of Bcl-2.

Malignant melanoma is a class of highly malignant tumors derived from melanocytes. At present, the dysregulated gene expression involved in the progression of melanoma has attracted much attention. In the present study, the gene expression profile of human melanoma tissue was screened using a cDNA microarray, and it was identified that melanocyte-specific gene 1 (MSG1) was significantly overexpressed in melanoma tissue compared with paired nevus tissues. The overexpression of MSG1 in melanoma was subsequently confirmed using immunohistochemistry in a set of melanoma tissues. It was additionally identified that the overexpression of MSG1 may promote cell viability and inhibit cell apoptosis in human melanoma A375 cells, thus promoting melanoma progression. Mechanistically, following screening of the expression of apoptosis-associated proteins, MSG1 was demonstrated to enhance the expression of the apoptosis inhibitor B-cell lymphoma 2 (Bcl-2) to inhibit melanoma cell apoptosis. Therefore, it was concluded that the overexpression of MSG1 inhibits apoptosis by enhancing Bcl-2 expression in malignant melanoma, thus promoting melanoma progression.

Ligand-activated BMP signaling inhibits cell differentiation and death to promote melanoma.

Oncogenomic studies indicate that copy number variation (CNV) alters genes involved in tumor progression; however, identification of specific driver genes affected by CNV has been difficult, as these rearrangements are often contained in large chromosomal intervals among several bystander genes. Here, we addressed this problem and identified a CNV-targeted oncogene by performing comparative oncogenomics of human and zebrafish melanomas. We determined that the gene encoding growth differentiation factor 6 (GDF6), which is the ligand for the BMP family, is recurrently amplified and transcriptionally upregulated in melanoma. GDF6-induced BMP signaling maintained a trunk neural crest gene signature in melanomas. Additionally, GDF6 repressed the melanocyte differentiation gene MITF and the proapoptotic factor SOX9, thereby preventing differentiation, inhibiting cell death, and promoting tumor growth. GDF6 was specifically expressed in melanomas but not melanocytes. Moreover, GDF6 expression levels in melanomas were inversely correlated with patient survival. Our study has identified a fundamental role for GDF6 and BMP signaling in governing an embryonic cell gene signature to promote melanoma progression, thus providing potential opportunities for targeted therapy to treat GDF6-positive cancers.

The unfolded protein response impacts melanoma progression by enhancing FGF expression and can be antagonized by a chemical chaperone

The mechanisms hallmarking melanoma progression are insufficiently understood. Here we studied the impact of the unfolded protein response (UPR) - a signalling cascade playing ambiguous roles in carcinogenesis - in melanoma malignancy. We identified isogenic patient-derived melanoma cell lines harboring BRAFV600E-mutations as a model system to study the role of intrinsic UPR in melanoma progression. We show that the activity of the three effector pathways of the UPR (ATF6, PERK and IRE1) was increased in metastatic compared to non-metastatic cells. Increased UPR-activity was associated with increased flexibility to cope with ER stress. The activity of the ATF6- and the PERK-, but not the IRE-pathway, correlated with poor survival in melanoma patients. Using whole-genome expression analysis, we show that the UPR is an inducer of FGF1 and FGF2 expression and cell migration. Antagonization of the UPR using the chemical chaperone 4-phenylbutyric acid (4-PBA) reduced FGF expression and inhibited cell migration and viability. Consistently, FGF expression positively correlated with the activity of ATF6 and PERK in human melanomas. We conclude that chronic UPR stimulates the FGF/FGF-receptor signalling axis and promotes melanoma progression. Hence, the development of potent chemical chaperones to antagonize the UPR might be a therapeutic approach to target melanoma.

Paracrine Activin-A Signaling Promotes Melanoma Growth and Metastasis through Immune Evasion

The secreted growth factor Activin-A of the transforming growth factor β family and its receptors can promote or inhibit several cancer hallmarks including tumor cell proliferation and differentiation, vascularization, lymphangiogenesis and inflammation. However, a role in immune evasion and its relationship with tumor-induced muscle wasting and tumor vascularization, and the relative contributions of autocrine versus paracrine Activin signaling remain to be evaluated. To address this, we compared the effects of truncated soluble Activin receptor IIB as a ligand trap, or constitutively active mutant type IB receptor versus secreted Activin-A or the related ligand Nodal in mouse and human melanoma cell lines and tumor grafts. We found that although cell-autonomous receptor activation arrested tumor cell proliferation, Activin-A secretion stimulated melanoma cell dedifferentiation and tumor vascularization by functional blood vessels, and it increased primary and metastatic tumor burden and muscle wasting. Importantly, in mice with impaired adaptive immunity, the tumor-promoting effect of Activin-A was lost despite sustained vascularization and cachexia, suggesting that Activin-A promotes melanoma progression by inhibiting antitumor immunity. Paracrine Activin-A signaling emerges as a potential target for personalized therapies, both to reduce cachexia and to enhance the efficacy of immunotherapies.

The DNA Methylcytosine Dioxygenase Tet2 Sustains Immunosuppressive Function of Tumor-Infiltrating Myeloid Cells to Promote Melanoma Progression

Ten-Eleven-Translocation-2 (Tet2) is a DNA methylcytosine dioxygenase that functions as a tumorsuppressor in hematopoietic malignancies. We examined the role of Tet2 in tumor-tissue myeloid cells and found that Tet2 sustains theimmunosuppressive function of these cells. We found that Tet2 expression is increased in intratumoral myeloid cells both in mouse models of melanoma and in melanoma patients and that this increased expression is dependent on an IL-1R-MyD88 pathway. Ablation of Tet2 in myeloid cells suppressed melanoma growth invivo and shifted the immunosuppressive gene expression program in tumor-associated macrophages to a proinflammatory one, with a concomitant reduction of the immunosuppressive function. This resulted in increased numbers of effector Tcells in the tumor, and Tcell depletion abolished the reduced tumor growth observed upon myeloid-specific deletion of Tet2. Our findings reveal a non-cell-intrinsic, tumor-promoting function for Tet2 and suggest that Tet2 may present a therapeutic targetfor the treatment of non-hematologic malignancies.