Erratum: MicroRNA-449a Inhibits Tumor Metastasis through AKT/ERK1/2 Inactivation by Targeting Steroid Receptor Coactivator (SRC) in Endometrial Cancer: Erratum.

[This corrects the article DOI: 10.7150/jca.27748.].

A stable atmospheric-pressure plasma for extreme-temperature synthesis.

Plasmas can generate ultra-high-temperature reactive environments that can be used for the synthesis and processing of a wide range of materials1,2. However, the limited volume, instability and non-uniformity of plasmas have made it challenging to scalably manufacture bulk, high-temperature materials3-8. Here we present a plasma set-up consisting of a pair of carbon-fibre-tip-enhanced electrodes that enable the generation of a uniform, ultra-high temperature and stable plasma (up to 8,000 K) at atmospheric pressure using a combination of vertically oriented long and short carbon fibres. The long carbon fibres initiate the plasma by micro-spark discharge at a low breakdown voltage, whereas the short carbon fibres coalesce the discharge into a volumetric and stable ultra-high-temperature plasma. As a proof of concept, we used this process to synthesize various extreme materials in seconds, including ultra-high-temperature ceramics (for example, hafnium carbonitride) and refractory metal alloys. Moreover, the carbon-fibre electrodes are highly flexible and can be shaped for various syntheses. This simple and practical plasma technology may help overcome the challenges in high-temperature synthesis and enable large-scale electrified plasma manufacturing powered by renewable electricity.

Sphingosine kinase 1 contributes to the metastatic potential of epithelial ovarian cancer to the adipocyte-rich niche.

Unlike many solid tumors, epithelial ovarian cancer (EOC) has a clear metastatic predilection to the adipocyte-rich niche, especially the omentum. However, the underlying mechanism driving this process remains incomplete. Here we show that SphK1 is over-expressed in omental metastases compared with ovarian primary tumors in EOC patients. In vitro, inhibition of SphK1 suppressed the metastatic ability of EOC induced by adipocytes. In vivo, blockage of SphK1 could attenuate the omental metastasis of EOC. Importantly, SphK1 modulates adipocyte-induced E/N-cadherin switch through Twist1, a key process in EOC metastasis. Our study reveals a previously unrecognized role of SphK1 in modulating the metastatic tropism of EOC to the adipocyte-rich niche, suggesting a new target for EOC therapy.

Activation of SphK2 contributes to adipocyte-induced EOC cell proliferation.

Epithelial ovarian cancer (EOC) is the leading cause of deaths due to cancer in women. Adipocytes have been suggested to play a key role in the stimulation of EOC growth. However, the mechanisms underlying the adipocyte-induced EOC proliferation remain undefined. Here, we provide the first evidence that adipocytes induce the activation of sphingosine kinase (SphK) 2 in EOC, which represents a novel pathway that mediates the adipocyte-induced EOC growth. SphK2 inhibition in EOC cells led to a remarkable inhibition of the adipocyte-induced cell proliferation. Moreover, the adipocyte-induced SphK2 activation in EOC cells was extracellular signal-regulated protein kinases (ERK) dependent. Furthermore, silencing SphK2 in EOC significantly inhibited the adipocyte-induced expression of phospho-ERK and c-Myc, two crucial players in EOC growth. Collectively, the current study unraveled a previously unrecognized role of SphK2 in the adipocyte-induced growth-promoting action in EOC, suggesting a novel target for EOC treatment.

Liposomal Delivery of MicroRNA-7 Targeting EGFR to Inhibit the Growth, Invasion, and Migration of Ovarian Cancer.

Ovarian cancer is highly aggressive and has high rates of recurrence and metastasis. Due to the limited effects of current treatments, it is necessary to conduct research and develop new treatment options. The application of gene therapy in tumor therapy is gradually increasing and has exciting prospects. MicroRNA-7 (miR-7) has been reported to inhibit the growth, invasion, and metastasis of a variety of solid tumors. Cationic liposomes are safe and effective gene delivery systems for transfection in vivo and in vitro. To realize the application of miR-7 in the treatment of ovarian cancer, cationic liposomes were prepared with 1,2-dioleoyl-3-trimethylammonium-propane, 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine, and cholesterol. The miR-7 liposomes had a suitable particle size, potential, and a high cellular uptake rate. MiR-7 encapsulated by liposomes could be effectively delivered to ovarian cancer cells and successfully targeted to the tumor site in a mouse xenograft model of ovarian cancer. In vitro and in vivo experiments revealed that the miR-7 liposomes had a significant ability to inhibit the growth, invasion, and migration of ovarian cancer, probably by inhibiting the expression of the epidermal growth factor receptor. Our studies of miR-7 liposomes demonstrated a safe and efficient microRNA delivery system for the gene therapy of ovarian cancer.

Activation of SphK1 by adipocytes mediates epithelial ovarian cancer cell proliferation.

BACKGROUND: Adipocytes, active facilitators of epithelial ovarian cancer (EOC) growth, have been implicated in the link between obesity and EOC. However, the current understanding of the mechanisms underlying adipocyte-induced EOC cell proliferation remains incomplete. RESULTS: We provide the first evidence showing that sphingosine kinase (SphK) 1 is critical for adipocyte-induced EOC cell proliferation. Adipocytes are capable of activating SphK1, which then leads to extracellular signal-regulated kinase (ERK) phosphorylation. Moreover, adipocyte-induced SphK1 activation is ERK dependent. Furthermore, sphingosine 1-phosphate receptor (S1PR) 1 and S1PR3, key components of the SphK1 signalling pathway, participate in adipocyte-mediated growth-promoting action in EOC cells. CONCLUSIONS: Our study reveals a previously unrecognized role of SphK1 in adipocyte-induced growth-promoting action in EOC, suggesting a new target for EOC therapy.

Inhibition of sphingosine kinase 2 down-regulates ERK/c-Myc pathway and reduces cell proliferation in human epithelial ovarian cancer.

BACKGROUND: Epithelial ovarian cancer (EOC) is the leading cause of death from female cancers. In our previous study, sphingosine kinase 2 (SphK2) inhibitor was shown to display anti-EOC activities. The purpose of this study was to evaluate further the expression characteristics and clinical significance of SphK2 in EOC and to explore the roles and underlying mechanisms of SphK2 in EOC cell survival. METHODS: The expression of SphK2 was examined by immunohistochemistry (IHC) and Western blot, and its clinical implications and prognostic significance were analyzed. We performed a cellular proliferation assay, and a mouse xenograft model was established to confirm the roles of SphK2 in vitro and in vivo. Cell cycle analysis, apoptosis assay, and Western blot were performed to examine cell cycle progression and apoptosis rate. Gene set enrichment analysis (GSEA), and Western blot were used to investigate the downstream signaling pathways related to SphK2 function. RESULTS: The expression level of SphK2 was shown to be associated with stage, histological grade, lymph node metastasis, and ascites status. More importantly, a high SphK2 expression level was a prognostic indicator of overall survival (OS) and relapse-free survival (RFS). Moreover, knockdown of SphK2 arrested cell cycle progression and inhibited EOC cell proliferation both in vitro and in vivo. Furthermore, ERK/c-Myc, the key pathway in EOC progression, was important for SphK2-mediated mitogenic action in EOC cells. CONCLUSIONS: Our findings provided the first evidence that SphK2 played a crucial role in EOC proliferation by regulating the ERK/c-Myc pathway. This indicated that SphK2 might serve as a prognostic marker and potential therapeutic target in EOC.

Adipocytes: active facilitators in epithelial ovarian cancer progression?

There is growing evidence that adipocytes play important roles in the progression of multiple cancers. Moreover, in obesity, adipocytes alter their original functions and contribute to the metabolic and inflammatory changes of adipose tissue microenvironment, which can further enhance tumor development. At present, the roles of adipocytes in the pathogenesis of epithelial ovarian cancer (EOC) are far from being fully elucidated. Herein, we summarized the recent advances in understanding the roles of adipocytes in EOC progression. Adipocytes, close neighbors of EOC tissue, promote EOC growth, invasion, metastasis and angiogenesis through adipokine secretion, metabolic remodeling and immune microenvironment modulation. Moreover, adipocytes are important therapeutic targets and may work as useful anticancer drug delivery depot for EOC treatment. Furthermore, adipocytes also act as a therapeutic obstacle for their involvement in EOC treatment resistance. Hence, better characterization of the adipocytes in EOC microenvironment and the crosstalk between adipocytes and EOC cells may provide insights into EOC progression and suggest novel therapeutic opportunities.

Follicle-stimulating hormone promotes the proliferation of epithelial ovarian cancer cells by activating sphingosine kinase.

Follicle-stimulating hormone (FSH) is closely related to the pathogenesis and progression of epithelial ovarian cancer (EOC). However, until now, knowledge relating to FSH-driven signalling pathways that lead to the growth of EOC remained incomplete. We sought to explore whether sphingosine kinase (SphK) could mediate FSH-induced ovarian cancer cell proliferation and which pathway might be involved in this process. The expression of phospho-SphK1 and phospho-SphK2 was detected in sections of EOC tissues by Immunohistochemical staining, and clinical significances were analyzed by statistical analysis. EOC cells were treated with FSH or/and SKI-II. CCK8 assays and colony formation assays were used to investigate cell proliferation. Western blot was carried out to detect protein expression in EOC cell line after treated with FSH. Here, for the first time, we provide evidence that high expression levels of phospho-SphK1 and phospho-SphK2 were both prognostic indicators of overall survival (OS) in EOC. Additionally, the expression levels of both phospho-SphK1 and phospho-SphK2 were closely correlated with the expression level of follicle-stimulating hormone receptor (FSHR) in ovarian cancer tissues. FSH stimulated the phosphorylation of both SphK1 and SphK2 and was able to regulate the survival and growth of ovarian cancer cells by activating SphK1 and SphK2 through ERK1/2. Both isoenzymes of SphK were equally responsible for FSH-induced cell proliferation of EOC. Both Erk1/2 and Akt activation play important roles in mediating FSH-induced cell proliferation after phosphorylation of SphK. Moreover, our data demonstrated that S1P receptor 1 (S1PR1) and S1PR3, key components of the SphK signalling system, were involved in FSH-mediated proliferation of EOC. Taken together, the results of the current study revealed that SphK is an essential mediator in FSH-induced proliferation of ovarian cancer cells in EOC, which indicates a new signalling pathway that controls FSH-mediated growth in EOC and suggests a new strategy that pharmaceutically targets both isoenzymes of SphK for the management of ovarian cancer.

Long non-coding RNA GAS5 inhibits DDP-resistance and tumor progression of epithelial ovarian cancer via GAS5-E2F4-PARP1-MAPK axis.

BACKGROUND: Epithelial ovarian cancer (EOC) is the malignant tumor of the female reproductive system with the highest fatality rate. Tolerance of chemotherapeutic drugs like cisplatin (DDP) occurring in very early stage is one of the important factors of the poor prognosis of epithelial ovarian cancer. Here we aim to study the dysregulation of a particular long noncoding RNA, lncRNA GAS5, and its role in EOC progression. METHODS: The low expression of lncRNA GAS5 in EOC tissues and OC cell lines was determined by microarray analyses and Real-Time qPCR. Flow cytometer assays were used to detect cell cycle and apoptosis of OC cells. CCK8 assay were performed to investigate the DDP sensitivity of OC cells. Western blot was carried out to detect cell growth markers, apoptotic markers, PARP1, E2F4, MAPK pathway protein expression and other protein expression in OC cell lines. The binding of GAS5 and E2F4 were proved by RNA pull-down and RIP assay. The effect of E2F4 on PARP1 were determined by CHIP-qPCR assay and luciferase reporter assay. The effect of lncRNA GAS5 on OC cells was assessed in vitro and in vivo. RESULTS: By microarray (3 EOC tissues νs. 3 normal ovary tissues) and RT- qPCR (53 EOC tissues νs. 10 normal ovary tissues) we identified lncRNA GAS5 to be dramatically low expressed in EOC samples and correlated with prognosis. Compared with sensitive cell lines, GAS5 was also low expressed in DDP resistant OC cell lines, and over-expression of GAS5 significantly enhanced the sensitivity of OC cells to DDP in vivo and in vitro. Meanwhile the over-expression of GAS5 also caused OC cells G0/G1 arrest and apoptosis increase. Mechanistically, GAS5 might regulate PARP1 expression by recruiting the transcription factor E2F4 to its promoter, and then affect the MAPK pathway activity. Due to the 5'TOP structure, GAS5 could be regulated by transcription inhibitor rapamycin in OC cells. CONCLUSION: Here we explored the specific mechanisms of EOC cisplatin resistance and tumor progress due to lncRNA-GAS5, presented the GAS5-E2F4-PARP1-MAPK axis and its role in OC drug-sensitivity and progression for the first time, and the results may provide experimental basis for clinical application.

Sphingosine kinase 2 inhibitor ABC294640 displays anti-epithelial ovarian cancer activities in vitro and in vivo.

Background: ABC294640 is a non-lipid competitive inhibitor of SphK2. It exhibited anti-proliferative activities in many human malignancies, including ovarian cancer. However, its potential mechanism of action remains poorly understood. Methods: In this paper, epithelial ovarian cancer (EOC) cell lines SKOV3 and HO8910 were treated with ABC294640. In order to explore the effect of ABC294640 on the behavior of ovarian cancer cells in vitro, we used cell counting kit-8 (CCK-8) assays, colony formation assays, flow cytometry, quantitative real-time PCR (qRT-PCR), Western blot analysis and immunohistochemistry to detect the effect of ABC294640 on cell proliferation, cell cycle distribution, cell apoptosis, the expression of related factors at mRNA levels, and the expression of related factors at protein level. An intra-abdominal xenograft tumor model of EOC was set up to assess the tumor growth in nude mice. Results: The results obtained indicate that EOC cell proliferation was noticeably inhibited in a concentration-dependent manner by ABC294640. ABC294640 caused cell cycle arrest in S phase and increased cell apoptosis rate in EOC cells. Also, the proteins, including phosphorylated retinoblastoma protein (P -Rb), cyclin D1, cyclin B1, and Bcl-2 were significantly inhibited, while cleaved-caspase 3 was activated. ABC294640 inhibited the expression of c-Myc in EOC. The in vivo assay showed an inhibitory effect of ABC294640 on tumor growth. Conclusions: ABC294640 could downregulate the expression of c-Myc in EOC both in vitro and in vivo. ABC294640 inhibited tumor growth in EOC via cell cycle arrest and inducing cell apoptosis both in vitro and in vivo, partially by decreasing the expression of cell cycle-associated proteins (such as P-Rb, cyclin B1, and cyclin D1) and promoting caspase 3 activation via downregulation expression of c-Myc. It suggested that ABC294640 had the potential to serve as an agent in EOC treatment.

MicroRNA-449a Inhibits Tumor Metastasis through AKT/ERK1/2 Inactivation by Targeting Steroid Receptor Coactivator (SRC) in Endometrial Cancer.

Endometrial cancer represents the leading frequency in gynecological malignancy in developed countries. Even with early detection, metastasis and recurrence remain the main reasons for a high death rate. MicroRNA-449a (miR-449a) has been reported to function as a tumor suppressor, yet the role of miR-449a in endometrial cancer metastasis has not been investigated. The present study identified that miR-449a was downregulated in advanced endometrial cancer. Overexpression of miR-449a decreased the migration and invasion of KLE and AN3CA endometrial cancer cells. Using luciferase reporter assays, we identified that miR-449a directly targeted the steroid receptor coactivator (SRC) by binding to sites in the 3' untranslated regions. Elevated expressions of SRC have been witnessed in advanced endometrial cancer tissues and have promoted tumor metastasis. We also identified that the suppressive effect of miR-449a on metastasis could be mediated by downregulating SRC and that miR-449a could suppress AKT and ERK1/2 pathway activation in endometrial cancer cells. These findings contribute to the current understanding of the function of miR-449a in endometrial cancer.

Postprandial increase in serum CA125 as a surrogate biomarker for early diagnosis of ovarian cancer.

BACKGROUND: CA125 is a prevalently used serum biomarker for detecting ovarian cancer over the last three decades. However, it has a significant deficiency in screening for early-stage cancer. With the purpose of exploring an effective approach to improve its performance in early diagnosis, we investigated the postprandial fluctuation pattern of cancer-derived CA125 and the underlying mechanism. METHODS: In two medical centers, 551 patients sonographically diagnosed with ovarian (adnexal) cysts (< 5 cm in diameter) were enrolled and divided into five disease groups (pelvic inflammatory cysts, retention cysts, endometrioma, benign/borderline cystadenoma and malignant cysts). The subtle differences in 1-h postprandial serum CA125 increases were compared between disease groups. A support vector machine (SVM)-based algorithm was used for refining the performance of CA125 postprandial increment. Ovarian cancer xenograft animal and cancer cell models were used to recapitulate the clinical findings and reveal the molecular basis of postprandial blood glucose and insulin in invoking the synthesis/secretion/re-absorption of CA125. RESULTS: Patients with ovarian cancer presented the highest postprandial increment 13.3 ± 0.7% (mean ± standard deviation) among the five disease groups. Using a CA125 increment ≥ 10% criterion, the sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) reached 83.3, 96.3, 61.1 and 98.8%, respectively, for early-stage ovarian cancer. This performance was further improved by the SVM-based CA125-increment algorithm, which exhibited 91.7% sensitivity, 99.2% specificity, 89.2% PPV and 99.4% NPV. Both modalities manifested diagnostic advantages over the traditional CA125 test (75.0% sensitivity, 25.4% specificity, 6.6% PPV and 93.6% NPV at the cut-off of 35 U/mL). Regarding the molecular basis, the postprandial blood glucose and insulin-invoked overexpression of Mucin 16 (encoding CA125) were demonstrated in animal and cancer cell models, which were mediated by the PI3K-Akt pathway. Nevertheless, a Mesothelin-based CA125 re-absorption behavior was noted in the treated cancer cells, which contributed to the over-drop following the postprandial peak of serum CA125. CONCLUSIONS: Cancer-derived serum CA125 possesses a unique and distinctive postprandial pattern, that distinguishes it from the common CA125 elevation in a benign disease condition. The dynamic measurement/assessment strategy can achieve a discriminatory power superior to that of a static test.

Enhanced Chemotherapeutic Efficacy of Paclitaxel Nanoparticles Co-delivered with MicroRNA-7 by Inhibiting Paclitaxel-Induced EGFR/ERK pathway Activation for Ovarian Cancer Therapy.

Chemotherapy-induced activation of cell survival pathways leads to drug resistance. MicroRNAs (miRNAs) post-transcriptionally regulate gene expression in many biological pathways. Paclitaxel (PTX) is one of the first-line chemotherapy drugs for ovarian cancer, and it induces the activation of the epidermal growth factor receptor (EGFR)/extracellular signal-regulated kinase (ERK) pathway that leads to tumor cell proliferation, survival, invasion, and drug resistance. MicroRNA-7 (miR-7) has the ability to suppress the EGFR/ERK pathway. To sensitize chemotherapy, we developed monomethoxy(poly(ethylene glycol))-poly(d,l-lactide- co-glycolide)-poly(l-lysine) nanoparticles for the simultaneous co-delivery of PTX and miR-7. The resulting PTX/miR-7 nanoparticles (P/MNPs) protect miRNA from degradation, possess a sequential and controlled release of drugs, improve the transfection efficiency of miRNA, decrease the half-maximal inhibitory concentration of PTX, and increase the apoptosis of ovarian cancer cells. The chemotherapeutic efficacy of PTX is prominently enhanced in vitro and in vivo via the inhibition of PTX-induced EGFR/ERK pathway activation by miR-7. Our studies in P/MNPs reveal a novel paradigm for a dual-drug-delivery system of chemotherapeutics and gene therapy in treating cancers.