Role of Exosomes in Cancer and Aptamer-Modified Exosomes as a Promising Platform for Cancer Targeted Therapy.

Exosomes are increasingly recognized as important mediators of intercellular communication in cancer biology. Exosomes can be derived from cancer cells as well as cellular components in tumor microenvironment. After secretion, the exosomes carrying a wide range of bioactive cargos can be ingested by local or distant recipient cells. The released cargos act through a variety of mechanisms to elicit multiple biological effects and impact most if not all hallmarks of cancer. Moreover, owing to their excellent biocompatibility and capability of being easily engineered or modified, exosomes are currently exploited as a promising platform for cancer targeted therapy. In this review, we first summarize the current knowledge of roles of exosomes in risk and etiology, initiation and progression of cancer, as well as their underlying molecular mechanisms. The aptamer-modified exosome as a promising platform for cancer targeted therapy is then briefly introduced. We also discuss the future directions for emerging roles of exosome in tumor biology and perspective of aptamer-modified exosomes in cancer therapy.

EXT1 methylation promotes proliferation and migration and predicts the clinical outcome of non-small cell lung carcinoma via WNT signalling pathway.

DNA methylation is important for lung cancer prognosis. In this work, it is aimed to seek novel biomarkers with DNA methylation-expression-pathway pattern and explore its underlying mechanism. Prognostic DNA methylation sites and mRNAs were screened in NSCLC data set from TCGA, and further validated using the samples retrospectively collected, and EXT1 was identified as a potential target. Gene body methylation of three CpG sites (cg03276982, cg11592677, cg16286281) on EXT1 was significantly associated with clinical outcome, and the EXT1 gene expression also predicted prognosis. The expression level of EXT1 was also correlated with its DNA methylation level. This observation was further validated in a new data set consist of 170 samples. Knocking down of EXT1 resulted in decreased proliferation and migration. EXT1 targets were analysed using GSEA. It is found that the WNT signalling is the potential downstream target of EXT1. Further analyses revealed that the EXT1 targets the beta-catenin and effect migration rate of NSCLC cell lines. The WNT signalling inhibitor, XAV-939, effectively disrupted the migration promotion effect induced by EXT1. In summary, EXT1 methylation regulates the gene expression, effects the proliferation and migration via WNT pathway and predicted a poor prognosis for NSCLC.

Neuregulin Signaling in the Tumor Microenvironment.

Neuregulins, members of the largest subclass of growth factors of the epidermal growth factor family, mediate a myriad of cellular functions including survival, proliferation, and differentiation in normal tissues through binding to receptor tyrosine kinases of the ErbB family. However, aberrant neuregulin signaling in the tumor microenvironment is increasingly recognized as a key player in initiation and malignant progression of human cancers. In this chapter, we focus on the role of neuregulin signaling in the hallmarks of cancer, including cancer initiation and development, metastasis, as well as therapeutic resistance. Moreover, role of neuregulin signaling in the regulation of tumor microenvironment and targeting of neuregulin signaling in cancer from the therapeutic perspective are also briefly discussed.

Knockdown of CYP24A1 Aggravates 1α,25(OH)2D3-Inhibited Migration and Invasion of Mouse Ovarian Epithelial Cells by Suppressing EMT.

Epithelial-mesenchymal transition (EMT) bestows cancer cells with motile and invasive properties. But for ovarian tissues, EMT plays a physiological role in the postovulatory repair of ovary surface epithelial (OSE) cells. Accumulating data indicated that 1α,25(OH)2D3 decreased both the migration and invasion of various cancer cells by suppressing EMT. However, it remains unclear whether 1α,25(OH)2D3 inhibits the process of EMT during different stages of oncogenic transformation in mouse OSE (MOSE) cells. In present study, a spontaneous malignant transformation model of MOSE cells at three sequential stages (early, intermediate and late) was established in vitro first and then subjected to 1α,25(OH)2D3 treatment to investigate the effect of 1α,25(OH)2D3 on the oncogenic transformation of MOSE cells. We found that 1α,25(OH)2D3 significantly reduced the proliferation and invasion of late malignant transformed MOSE (M-L cells) cells by inhibiting EMT both in vitro and in vivo, but not in intermediate transformed (M-I) cells. Importantly, we found that the levels of CYP24A1 in M-I cells were dramatically higher than that in M-L cells following treatment with 1α,25(OH)2D3. Furthermore, we demonstrated that, in both M-I and M-L cells with CYP24A1 knockdown, 1α,25(OH)2D3 suppressed the proliferation and invasion, and reduced the expression of N-cadherin, Vimentin, ß-catenin and Snail. In addition, knockdown of CYP24A1 suppressed EMT by increasing E-cadherin while decreasing N-cadherin, Vimentin, ß-catenin and Snail. These findings provide support for inhibiting CYP24A1 as a potential approach to activate the vitamin D pathway in the prevention and therapy of ovarian cancer.

Mesenchymal stem cell-derived exosomes protect beta cells against hypoxia-induced apoptosis via miR-21 by alleviating ER stress and inhibiting p38 MAPK phosphorylation.

BACKGROUND: Hypoxia is a major cause of beta cell death and dysfunction after transplantation. The aim of this study was to investigate the effect of exosomes derived from mesenchymal stem cells (MSCs) on beta cells under hypoxic conditions and the potential underlying mechanisms. METHODS: Exosomes were isolated from the conditioned medium of human umbilical cord MSCs and identified by WB, NTA, and transmission electron microscopy. Beta cells (ßTC-6) were cultured in serum-free medium in the presence or absence of exosomes under 2% oxygen conditions. Cell viability and apoptosis were analysed with a CCK-8 assay and a flow cytometry-based annexin V-FITC/PI apoptosis detection kit, respectively. Endoplasmic reticulum stress (ER stress) proteins and apoptosis-related proteins were detected by the WB method. MiRNAs contained in MSC exosomes were determined by Illumina HiSeq, and treatment with specific miRNA mimics or inhibitors of the most abundant miRNAs was used to reveal the underlying mechanism of exosomes. RESULTS: Exosomes derived from MSC-conditioned culture medium were 40-100 nm in diameter and expressed the exosome markers CD9, CD63, CD81, HSP70, and Flotillin 1, as well as the MSC markers CD73, CD90, and CD105. Hypoxia significantly induced beta cell apoptosis, while MSC exosomes remarkably improved beta cell survival. The WB results showed that ER stress-related proteins, including GRP78, GRP94, p-eIF2α and CHOP, and the apoptosis-related proteins cleaved caspase 3 and PARP, were upregulated under hypoxic conditions but were inhibited by MSC exosomes. Moreover, the p38 MAPK signalling pathway was activated by hypoxia and was inhibited by MSC exosomes. The Illumina HiSeq results show that MSC exosomes were rich in miR-21, let-7 g, miR-1246, miR-381, and miR-100. After transfection with miRNA mimics, the viability of beta cells under hypoxia was increased significantly by miR-21 mimic, and the p38 MAPK and ER stress-related proteins in beta cells were downregulated. These changes were reversed after exosomes were pretreated with miR-21 inhibitor. CONCLUSIONS: Exosomes derived from MSCs could protect beta cells against apoptosis induced by hypoxia, largely by carrying miR-21, alleviating ER stress and inhibiting p38 MAPK signalling. This result indicated that MSC exosomes might improve encapsulated islet survival and benefit diabetes patients.

Hesperetin reverses P­glycoprotein­mediated cisplatin resistance in DDP­resistant human lung cancer cells via modulation of the nuclear factor­κB signaling pathway.

Lung cancer is the leading cause of cancer­associated mortality worldwide. Cisplatin (DDP) is a first­line chemotherapeutic drug for the treatment of lung cancer; however, the majority of patients develop resistance to DDP. P­glycoprotein (P­gp), also referred to as multidrug resistance (MDR) protein 1, is associated with an MDR phenotype, which results in failure of cancer chemotherapy; thus, identifying effective MDR pump inhibitors may improve the outcomes of patients who develop resistance to treatment. Hesperetin is a derivative of hesperidin, which is extracted from tangerine peel and exhibits multiple antitumor properties. In the present study, human lung adenocarcinoma A549 and A549/DDP cells were treated with different concentrations of hesperetin and DDP, respectively. Furthermore, rhodamine 123 efflux assays, Cell Counting Kit­8 assays, immunofluorescence, reverse transcription­quantitative PCR and western blot analysis were used to elucidate the mechanisms underlying the effects of hesperetin On A549/DDP cells. Additionally, a xenograft model of lung cancer in nude mice was established to explore the effects of hesperetin on A549/DDP cell growth in vivo. The results demonstrated that hesperetin sensitized A549/DDP cells to DDP. In vivo, hesperetin pretreatment significantly inhibited tumor growth. Mechanistically, hesperetin markedly decreased the expression of P­gp and increased the intracellular accumulation of the P­gp substrate, rhodamine 123, in A549/DDP cells. In addition, pretreatment of A549/DDP cells with hesperetin significantly inhibited nuclear factor (NF)­κB (p65) activity and its nuclear translocation. Taken together, the results of the present study suggest that hesperetin reversed P­gp­mediated MDR by decreasing P­gp expression in A549/DDP cells, which was associated with inhibition of the NF­κB signaling pathway. These findings may provide the basis for the use of hesperetin clinically to reverse MDR.

ABAT and ALDH6A1, regulated by transcription factor HNF4A, suppress tumorigenic capability in clear cell renal cell carcinoma.

BACKGROUND: Clear cell renal cell carcinoma (ccRCC) is a malignancy characterized by metabolic reprogramming. ABAT and ALDH6A1 are metabolic enzymes. In this study, we aim to investigate the associations of ABAT and ALDH6A1 with the malignancy of ccRCC cells. METHODS: The gene expression levels of ABAT and ALDH6A1 in ccRCC were analyzed from gene expression microarray datasets and RNA sequencing data. Clinical information was analyzed from The Cancer Genome Atlas (TCGA) data. The distributions of ABAT and ALDH6A1 in ccRCC clinical tissues were screened by reverse transcription-quantitative polymerase chain reaction (RT-QPCR) and immunohistochemical assays. The effect of overexpression of ABAT or ALDH6A1 was measured by detecting the cell viability, migration ability, and the ratio of lactate and nicotinamide adenine dinucleotide phosphate (NADPH). Chromatin immunoprecipitation (ChIP) and luciferase reporter assays were carried out to investigate the transcript regulation of HNF4A in ABAT and ALDH6A1. RESULTS: Remarkable downregulated ABAT and ALDH6A1 expression levels were observed in ccRCC patients and low expression of ABAT and ALDH6A1 was correlated with poor survival. Overexpression of ABAT or ALDH6A1 significantly attenuated cell proliferation and migration, and impaired lactate production. In ABAT increased ccRCC cells, the ratio of NADPH/NADP+ was reduced. Finally, we demonstrated that ABAT and ALDH6A1 were directly regulated by a tumor suppressor, HNF4A. CONCLUSIONS: These observations identified HNF4A-regulated low-expressed ABAT and ALDH6A1 as promising diagnostic and prognostic biomarkers for ccRCC.

Pazopanib as salvage therapy in metastatic renal cell carcinoma with hypercalcemic crisis and renal insufficiency: a case report and literature review.

A hypercalcemic crisis in renal cell carcinoma (RCC) is an extremely rare and life-threatening condition for advanced RCC patients. It is considered nearly intractable for treatment and a poor-risk category by Memorial Sloan Kettering Cancer Center (MSKCC) criteria. In our case, best supportive care was regularly administered according to the related guidelines and consensuses but with little high-quality, prospective clinical trial data to support the therapeutic strategy. Indeed, determining the individual etiological treatment for a given patient can be challenging. Here, we present a typical case with hypercalcemic crisis, reduced renal function (chronic kidney disease, CKD4), and poor performance status. The patient, who was treated with pazopanib of an individual lower dose of 200 mg daily as salvage therapy, had significantly improved quality of life (QOL) and prolonged progression-free survival (PFS) and overall survival (OS). These are the first results of their kind to be reported of a clinical benefit being generally observed with single doses of 800 mg. How to individually control the primary disease and concurrently relieve the symptoms in clinic to improve QOL and prolong the patient's PFS and OS is worthy of exploration.

Icariin induces apoptosis of human lung adenocarcinoma cells by activating the mitochondrial apoptotic pathway.

Lung cancer is the largest cause of morbidity and mortality among tumor diseases. Traditional first-line chemotherapeutic drugs are frequently accompanied by serious side effects when used to treat tumors, thus, novel drugs with reduced toxic effects may improve a patients' quality of life. Icariin, an extract of herba epimedii, has been demonstrated to exhibit multiple antitumor effects with low toxicity. In the present study, cell cycle analysis, apoptosis assays, DAPI staining, CCK8 assays, xenograft tumor models, mitochondrial membrane potential analysis, western blotting and reverse transcription-quantitative PCR were performed to determine the molecular mechanism underlying icariin activity in the human lung adenocarcinoma cell lines, A549 and H1975. The results showed that icariin reduced proliferation of A549 and H1975 cells in a time- and dose-dependent manner in vitro to a greater degree than the control BEAS-2B cells, and this was associated with increased apoptosis, but not with cell cycle progression. In vivo experiments showed that icariin treatment significantly decreased proliferation of H1975 cells in a xenograft mouse model. Mechanistically, icariin activated the mitochondrial pathway by inhibiting the activation of the PI3K-Akt pathway-associated kinase, Akt, resulting in the activation of members of the caspase family of proteins, and thus inducing apoptosis of A549 cells. Taken together, the results revealed that icariin has anti-cancer properties in lung cancer in vitro and in vivo without any noticeable toxic effects on normal lung epithelial cells. Icariin in combination with conventional anti-cancer agents may be an effective therapeutic strategy for treatment of lung carcinoma.

LAT, HOXD3 and NFE2L3 identified as novel DNA methylation-driven genes and prognostic markers in human clear cell renal cell carcinoma by integrative bioinformatics approaches.

Background: Abnormal DNA methylation of is one of the important mechanisms leading to tumor pathogenesis. The purpose of this study was to explore differentially methylated genes that may drive the development of renal clear cell carcinoma through a comprehensive analysis of the TCGA database. Materials and methods: Methylation data and RNA-seq data for clear cell renal cell carcinoma were downloaded from The Cancer Genome Atlas (TCGA). Differentially methylated genes and the differential genes associated with survival were then screened by MethylMix R package and univariate Cox proportional-hazards model, respectively. Their common genes were then intersected and obtained for further analysis. Correlation of gene expression and methylation levels, gene set enrichment analysis (GSEA) enrichments, survival curve, and ROC curve plotting for DNA methylation-driven genes were finally performed. The methylation alterations of the three genes were validated via two GEO datasets (GSE70303 and GSE113501), and the genes expression level was verified through two GEO datasets (GSE6344 and GSE53757). Results: Three novel DNA methylation-driven genes LAT, HOXD3 and NFE2L3 were identified in clear cell renal cell carcinoma. Expression analysis further revealed that hypomethylation levels of LAT and NFE2L3 showed higher gene expression levels, while HOXD3 exhibited opposite methylation-expression pattern. The CpG sites of LAT (cg16462073), HOXD3 (cg24000528) and NFE2L3 (cg16882373) that may affect respective gene expressions were also identified. For the survival analysis, we found that hypomethylation and over-expression of LAT and NFE2L3 were correlated with poor survival, while hypermethylation and low-expression HOXD3 was correlated with poor survival of clear cell renal cell carcinoma patients. In addition, GSEA KEGG analysis and biological processes of these genes were also enriched for functional analysis. Kaplan-Meier survival and ROC analyses of these genes showed an average risk score of 0.9140593, AUC = 0.692, which suggested a good clinical application value. Finally, the opposite methylation-expression pattern of these three genes were verified in GEO datasets. Conclusions: In this study, we successfully exhibited the potential DNA methylation-driven genes LAT, HOXD3, and NFE2L3 involved in clear cell renal cell carcinoma. Moreover, gene functions and prognostic risk models were also elucidated, which facilitated the expansion of the current study on the role of methylation in the pathology process of clear cell renal cell carcinoma.

Integrative gene expression profiling reveals that dysregulated triple microRNAs confer paclitaxel resistance in non-small cell lung cancer via co-targeting MAPT.

BACKGROUND: Paclitaxel has shown significant anti-tumor activity against non-small cell lung cancer (NSCLC); however, resistance to paclitaxel frequently occurs and represents a significant clinical problem and its underlying molecular mechanism remains elusive. METHODS: Long-term treatment of culture cell with paclitaxel was carried out to mimic the development of acquired drug resistance in NSCLC. Cell proliferation and clonogenic assay and apoptosis evaluation were carried out to determine the efficacy of paclitaxel on NSCLC cells. Western blot analyses were performed to determine the expression and activation of proteins. Apoptosis enzyme-linked immunosorbent assay was used to quantify cytoplasmic histone-associated DNA fragments. Microarray analyses were applied to explore both mRNA and miRNA expression profiles in NSCLC cells followed by integrative analysis. qRT-PCR was carried out to verify the differentially expressed mRNAs and miRNAs. RESULTS: The expression of 652 genes was shown to be changed at least 2-fold in paclitaxel-resistant NSCLC (H460_TaxR) cells with 511 upregulated and 141 downregulated as compared with that in parental H460 cells. The differentially expressed genes were functionally enriched in regulating the cell proliferation, cell death, and response to endogenous stimulus, and clustered in pathways such as cancer and signaling by the G protein-coupled receptor (GPCR). Moreover, 43 miRNAs were shown to be differentially expressed in H460_TaxR cells with 15 upregulated and 28 downregulated as compared with parental H460 cells. A total of 289 pairs of miRNA-potential target gene were revealed in H460_TaxR cells by bioinformatics analysis. Furthermore, integrative analysis of miRNAs and gene expression profiles revealed that dysregulated miR-362-3p, miR-766-3p, and miR-6507-3p might confer paclitaxel resistance in NSCLC via targeting MAPT simultaneously. CONCLUSION: Our findings suggested that specific manipulation of MAPT-targeting miRNAs may be a novel strategy to overcome paclitaxel resistance in patients with NSCLC especially large-cell lung carcinoma.

miR-192/215-5p act as tumor suppressors and link Crohn's disease and colorectal cancer by targeting common metabolic pathways: An integrated informatics analysis and experimental study.

MicroRNAs have emerged as key regulators involved in a variety of biological processes. Previous studies have demonstrated that miR-192/215 participated in progression of Crohn's disease and colorectal cancer. However, their concrete relationships and regulation networks in diseases remain unclear. Here, we used bioinformatics methods to expound miR-192/215-5p macrocontrol regulatory networks shared by two diseases. For data mining and figure generation, several miRNA prediction tools, Human miRNA tissue atlas, FunRich, miRcancer, MalaCards, STRING, GEPIA, cBioPortal, GEO databases, Pathvisio, Graphpad Prism 6 software, etc . are extensively applied. miR-192/215-5p were specially distributed in colon tissues and enriched biological pathways were closely associated with human cancers. Emerging role of miR-192/215-5p and their common pathways in Crohn's disease and colorectal cancer was also analyzed. Based on results derived from multiple approaches, we identified the biological functions of miR-192/215-5p as a tumor suppressor and link Crohn's disease and colorectal cancer by targeting triglyceride synthesis and extracellular matrix remodeling pathways.

Valproic acid exhibits anti-tumor activity selectively against EGFR/ErbB2/ErbB3-coexpressing pancreatic cancer via induction of ErbB family members-targeting microRNAs.

BACKGROUND: Deregulated ErbB signaling plays an important role in tumorigenesis of pancreatic cancer. However, patients with pancreatic cancer benefit little from current existed therapies targeting the ErbB signaling. Here, we explore the potential anti-tumor activity of Valproic acid against pancreatic cancer via targeting ErbB family members. METHODS: Cell viability assay and apoptosis evaluation were carried out to determine the efficacy of VPA on pancreatic cancer cells. Western blot analyses were performed to determine the expression and activation of proteins. Apoptosis enzyme-linked immunosorbent assay was used to quantify cytoplasmic histone associated DNA fragments. Lentiviral expression system was used to introduce overexpression of exogeneous genes or gene-targeting short hairpin RNAs (shRNAs). qRT-PCR was carried out to analyze the mRNAs and miRNAs expression levels. Tumor xenograft model was established to evaluate the in vivo anti-pancreatic cancer activity of VPA. RESULTS: VPA preferentially inhibited cell proliferation/survival of, and induced apoptosis in EGFR/ErbB2/ErbB3-coexpressing pancreatic cancer cells within its clinically achievable range [40~100 mg/L (0.24~0.6 mmol/L)]. Mechanistic investigations revealed that VPA treatment resulted in simultaneous significant down-regulation of EGFR, ErbB2, and ErbB3 in pancreatic cancer cells likely via induction of ErbB family members-targeting microRNAs. Moreover, the anti-pancreatic cancer activity of VPA was further validated in tumor xenograft model. CONCLUSIONS: Our data strongly suggest that VPA may be added to the treatment regimens for pancreatic cancer patients with co-overexpression of the ErbB family members.

Prognostic values of GMPS, PR, CD40, and p21 in ovarian cancer.

Early detection and prediction of prognosis and treatment responses are all the keys in improving survival of ovarian cancer patients. This study profiled an ovarian cancer progression model to identify prognostic biomarkers for ovarian cancer patients. Mouse ovarian surface epithelial cells (MOSECs) can undergo spontaneous malignant transformation in vitro cell culture. These were used as a model of ovarian cancer progression for alterations in gene expression and signaling detected using the Illumina HiSeq2000 Next-Generation Sequencing platform and bioinformatical analyses. The differential expression of four selected genes was identified using the gene expression profiling interaction analysis (http://gepia.cancer-pku.cn/) and then associated with survival in ovarian cancer patients using the Cancer Genome Atlas dataset and the online Kaplan-Meier Plotter (http://www.kmplot.com) data. The data showed 263 aberrantly expressed genes, including 182 up-regulated and 81 down-regulated genes between the early and late stages of tumor progression in MOSECs. The bioinformatic data revealed four genes (i.e., guanosine 5'-monophosphate synthase (GMPS), progesterone receptor (PR), CD40, and p21 (cyclin-dependent kinase inhibitor 1A)) to play an important role in ovarian cancer progression. Furthermore, the Cancer Genome Atlas dataset validated the differential expression of these four genes, which were associated with prognosis in ovarian cancer patients. In conclusion, this study profiled differentially expressed genes using the ovarian cancer progression model and identified four (i.e., GMPS, PR, CD40, and p21) as prognostic markers for ovarian cancer patients. Future studies of prospective patients could further verify the clinical usefulness of this four-gene signature.

MiR-30a-5p frequently downregulated in prostate cancer inhibits cell proliferation via targeting PCLAF.

MicroRNAs (miRNAs) have vastly expanded our view of RNA world in intracellular signal regulating networks. Here, we functionally characterized a normally highly expressed miRNA, miR-30a-5p (MIMAT0000087), which exhibits downregulated expression profiles in prostate cancer samples. MiR-30a-5p knockdown and overexpression in PC-3 cell line alters cell proliferation supporting a tumour suppressor role. We also discovered that PCLAF is the direct target of miR-30a-5p. Notably, PC-3 cell proliferation is inhibited by miR-30a-5p/PCLAF axis. miR-30a-5p represents a novel molecule of functionally important miRNAs which may shed light on the novel therapeutic targets for prostate cancer.

Functional Long Noncoding RNAs (lncRNAs) in Clear Cell Kidney Carcinoma Revealed by Reconstruction and Comprehensive Analysis of the lncRNA-miRNA-mRNA Regulatory Network.

BACKGROUND A variety of treatment strategies have been developed for clear cell kidney carcinoma (KIRC); however, there is still a need for effective therapeutic targets and prognostic molecular biomarkers. Given that long noncoding RNAs (lncRNAs) has been emerging as an important regulator in tumorigenesis, we explored potential functional lncRNAs in KIRC by comprehensively analyzing the lncRNA-miRNA-mRNA regulatory network with bioinformatics processing tools. MATERIAL AND METHODS RNA-seq/miRNA-seq data of KIRC in The Cancer Genome Atlas (TCGA) were obtained and analyzed. The "edgeR" package in R software was used to identify differentially expressed lncRNAs (DElncRNAs, differentially expressed long noncoding RNAs), miRNAs (DEmiRNAs, differentially expressed micro RNAs), and mRNAs (DEmRNAs, differentially expressed messenger RNAs) in KIRC and normal samples. A global triple network was conducted based on the competing endogenous RNA (ceRNA) theory, and survival analysis was conducted by "survival" package in R software. RESULTS A total of 4246 DElncRNAs, 179 DEmiRNAs, and 5758 DEmRNAs were identified, among which a subset of them (321 lncRNAs, 26 miRNAs, and 1068 mRNAs) were found to constitute a global ceRNA network in KIRC. Four lncRNAs (ENTPD3-AS1, FGD5-AS1, LIFR-AS1, and UBAC2-AS1) were revealed to be potential therapeutic targets as well as prognostic biomarkers of KIRC by our extensive functional analysis. CONCLUSIONS We reported here the identification of functional lncRNAs in KIRC via a TCGA data-based bioinformatics analysis. We believe that this study might contribute to improving the comprehension of the lncRNA-mediated ceRNA regulatory mechanisms in the tumorigenesis of KIRC. Meanwhile, our results suggested that 4 lncRNAs might act as potential therapeutic targets or candidate prognostic biomarkers in KIRC.

Mesenchymal stem cells drive paclitaxel resistance in ErbB2/ErbB3-coexpressing breast cancer cells via paracrine of neuregulin 1.

We had previously demonstrated that increased expression of ErbB3 is required for ErbB2-mediated paclitaxel resistance in breast cancer cells. In the present study, we have explored the possible role of mesenchymal stem cells (MSCs) in regulating the paclitaxel-sensitivity of ErbB2/ErbB3-coexpressing breast cancer cells. We show that human umbilical cord-derived MSCs express significantly higher level of neuregulin-1 as compared with ErbB2/ErbB3-coexpressing breast cancer cells themselves. Coculture or treatment with conditioned medium of MSCs not only decreases the anti-proliferation effect of paclitaxel on ErbB2/ErbB3-coexpressing breast cancer cells, but also significantly inhibits paclitaxel-induced apoptosis. We further demonstrate that this MSCs-drived paclitaxel resistance in ErbB2/ErbB3-coexpressing breast cancer cells could be attributed to upregulation of Survivin via paracrine effect of NRG-1/ErbB3/PI-3K/Akt signaling, as either specific knockdown expression of ErbB3, or blocking of downstream PI-3K/Akt signaling, or specific inhibition of Survivin can completely reverse this effect. Moreover, targeted knockdown of NRG-1 expression in MSCs abrogates theirs effect on paclitaxel sensitivity of ErbB2/ErbB3-coexpressing breast cancer cells. Taken together, our study indicate that paracrine of NRG-1 by MSCs induces paclitaxel resistance in ErbB2/ErbB3-coexpressing breast cancer cells through PI-3K/Akt signaling-dependent upregulation of Survivin. Our findings suggest that simultaneously targeting mesenchymal stem cells in tumor microenvironment may be a novel strategy to overcome paclitaxel resistance in patients with ErbB2/ErbB3-coexpressing breast cancer.

Survivin-targeting miR-542-3p overcomes HER3 signaling-induced chemoresistance and enhances the antitumor activity of paclitaxel against HER2-overexpressing breast cancer.

Elevated expression of HER3, which interacts with HER2 in breast cancer cells, confers chemoresistance via phosphoinositide 3-kinase (PI-3K)/Akt-dependent upregulation of Survivin. However, the underlying mechanism is not clear. Ectopic expression or specific knockdown of HER3 in HER2-overexpressing breast cancer cells did not alter Survivin mRNA levels and Survivin protein stability, supporting the notion that HER3 signaling may regulate specific miRNAs that target Survivin to alter its protein translation. Here we showed that overexpression and specific knockdown of HER3 reduced and enhanced expression of two Survivin-targeting miRNAs, miR-203 and miR-542-3p, in breast cancer cells, respectively. While the specific inhibitor of either miR-203 or miR-542-3p attenuated an anti-HER3 antibody-induced downregulation of Survivin, inhibition of miR-542-3p exhibited a better efficacy than miR-203 inhibition did. Consistently, miR-542-3p mimic was much more effective than miR-203 mimic not only in inhibition of Survivin, but also in enhancement of paclitaxel-induced apoptosis in HER2-overexpressing breast cancer cells. Moreover, the combination of miR-542-3p mimic and paclitaxel, as compared with either agent alone, significantly inhibited in vivo tumor growth of HER2-overexpressing breast cancer cells. Collectively, our data indicated that the HER3/PI-3K/Akt signaling upregulates Survivin via suppression of miR-203 and miR-542-3p. Because miR-542-3p has three binding sites on the 3'-UTR of Survivin mRNA, its mimic was able to effectively downregulate Survivin in vitro and in vivo. Thus, miR-542-3p-replacement therapy is an excellent approach to overcome HER3-mediated paclitaxel resistance and significantly enhances the antitumor activity of paclitaxel against HER2-overexpressing breast cancer.

CHOP favors endoplasmic reticulum stress-induced apoptosis in hepatocellular carcinoma cells via inhibition of autophagy.

C/EBP-homologous protein (CHOP) is an important component of the endoplasmic reticulum (ER) stress response. We demonstrated the induction of ER stress in response to tunicamycin stimulation, as evidenced by increased expression of chaperone proteins Grp78, Grp94, and enhanced eukaryotic initiation factor 2 subunit 1 (eIF2α) phosphorylation in hepatocellular carcinoma cells. Tunicamycin-induced ER stress resulted in apoptosis and autophagy simultaneously. While inhibition of autophagy mediated by 3-methyladenine pretreatment or direct knockdown of LC3B promoted cell apoptosis, activation of autophagy with rapamycin decreased tunicamycin- induced apoptosis in HCC cells. Furthermore, CHOP was shown to be significantly upregulated upon treatment with tunicamycin in HCC cells. Specific knockdown of CHOP not only enhanced tunicamycin-induced autophagy, but also significantly attenuated ER stress-induced apoptosis in HCC cells. Accordingly, simultaneous inhibition of autophagy in HCC cells with CHOP-knockdown could partially resensitize ER stress-induced apoptosis. Taken together, our data indicate that CHOP may favor ER stress-induced apoptosis in HCC cells via inhibition of autophagy in vitro.

MicroRNA-mediated epigenetic targeting of Survivin significantly enhances the antitumor activity of paclitaxel against non-small cell lung cancer.

Elevated expression of Survivin correlates with poor prognosis, tumor recurrence, and drug resistance in various human cancers, including non-small cell lung cancer (NSCLC). The underlying mechanism of Survivin upregulation in cancer cells remains elusive. To date, no Survivin-targeted therapy has been approved for cancer treatment. Here, we explored the molecular basis resulting in Survivin overexpression in NSCLC and investigated the antitumor activity of the class I HDAC inhibitor entinostat in combination with paclitaxel. Our data showed that entinostat significantly enhanced paclitaxel-mediated anti-proliferative/anti-survival effects on NSCLC cells in vitro and in vivo. Mechanistically, entinostat selectively decreased expression of Survivin via induction of miR-203 (in vitro and in vivo) and miR-542-3p (in vitro). Moreover, analysis of NSCLC patient samples revealed that the expression levels of miR-203 were downregulated due to promoter hypermethylation in 45% of NSCLC tumors. In contrast, increased expression of both DNA methytransferase I (DNMT1) and Survivin was observed and significantly correlated with the reduced miR-203 in NSCLC. Collectively, these data shed new lights on the molecular mechanism of Survivin upregulation in NSCLC. Our findings also support that the combinatorial treatments of entinostat and paclitaxel will likely exhibit survival benefit in the NSCLC patients with overexpression of DNMT1 and/or Survivin. The DNMT1-miR-203-Survivin signaling axis may provide a new avenue for the development of novel epigenetic approaches to enhance the chemotherapeutic efficacy against NSCLC.