Retraction Notice to: m6A demethylase FTO suppresses pancreatic cancer tumorigenesis by demethylating PJA2 and inhibiting Wnt signaling.

[This retracts the article DOI: 10.1016/j.omtn.2021.06.005.].

m6A demethylase FTO suppresses pancreatic cancer tumorigenesis by demethylating PJA2 and inhibiting Wnt signaling.

Pancreatic cancer is the deadliest malignancy of the digestive system and is the seventh most common cause of cancer-related deaths worldwide. The incidence and mortality of pancreatic cancer continue to increase, and its 5-year survival rate remains the lowest among all cancers. N6-methyladenine (m6A) is the most abundant reversible RNA modification in various eukaryotic messenger and long noncoding RNAs and plays crucial roles in the occurrence and development of cancers. However, the role of m6A in pancreatic cancer remains unclear. The present study aimed to explore the role of m6A and its regulators in pancreatic cancer and assess its underlying molecular mechanism associated with pancreatic cancer cell proliferation, invasion, and metastasis. Reduced expression of the m6A demethylase, fat mass and obesity-associated protein (FTO), was responsible for the high levels of m6A RNA modification in pancreatic cancer. Moreover, FTO demethylated the m6A modification of praja ring finger ubiquitin ligase 2 (PJA2), thereby reducing its mRNA decay, suppressing Wnt signaling, and ultimately restraining the proliferation, invasion, and metastasis of pancreatic cancer cells. Altogether, this study describes new, potential molecular therapeutic targets for pancreatic cancer that could pave the way to improve patient outcome.

Discovery of novel nitrogenous heterocyclic-containing quinoxaline-1,4-di-N-oxides as potent activator of autophagy in M.tb-infected macrophages.

As a continuation of our research on antimycobacterial agents, a series of novel quinoxaline-1,4-di-N-oxides (QdNOs) containing various nitrogenous heterocyclic moieties at the R6 position were designed and synthesized. Antimycobacterial activities, as well as the cytotoxic effects, of the compounds were assayed. Four compounds (6b, 6f, 6n, and 6o), characterized by 2-carboxylate ethyl or benzyl ester, 6-imidazolyl or 1,2,4-triazolyl, and a 7-fluorine group, exhibited the most potent antimycobacterial activity against M.tb strain H37Rv (MIC ≤ 0.25 µg/mL) with low toxicity in VERO cells (SI = 169.3-412.1). Compound 6o also exhibited excellent antimycobacterial activity in an M.tb-infected macrophage model and was selected for further exploration of the mode of antimycobacterial action of QdNOs. The results showed that compound 6o was capable of disrupting membrane integrity and disturbing energy homeostasis in M.tb. Furthermore, compound 6o noticeably increased cellular ROS levels and, subsequently, induced autophagy in M.tb-infected macrophages, possibly indicating the pathways of QdNOs-mediated inhibition of intracellular M.tb replication. The in vivo pharmacokinetic (PK) profiles indicated that compounds 6o was acceptably safe and possesses favorable PK properties. Altogether, these findings suggest that compound 6o is a promising antimycobacterial candidate for further research.

Genetic alterations and functional networks of m6A RNA methylation regulators in pancreatic cancer based on data mining.

BACKGROUND: Pancreatic cancer is a fatal malignancy of the digestive system and the 5-year survival rate remains low. Therefore, new molecular therapeutic targets are required to improve treatments, prognosis, and the survival of patients. N6-methyladenosine (m6A) is the most prevalent reversible methylation in mammalian messenger RNA (mRNA) and has critical roles in the tumorigenesis and metastasis of various malignancies. However, the role of m6A in pancreatic cancer is still unclear. Exploring genetic alterations and functional networks of m6A regulators in pancreatic cancer may provide new strategies for its treatment. METHODS: In this study, we used data from the Cancer Genome Atlas (TCGA) database and other public databases through cBioPortal, LinkedOmics, UALCAN, GEPIA, STRING, and the database for annotation, visualization, and integrated discovery (DAVID) to systematically analyze the molecular alterations and functions of 20 main m6A regulators in pancreatic cancer. RESULTS: We found that m6A regulators had widespread genetic alterations, and that their expression levels were significantly correlated with pancreatic cancer malignancy. Moreover, m6A regulators were associated with the prognosis of pancreatic cancer patients. CONCLUSIONS: m6A regulators play a crucial part in the occurrence and development of pancreatic cancer. Our study will guide further studies of m6A RNA modification in pancreatic cancer and could potentially provide new strategies for pancreatic cancer treatment.

ITGB1 enhances the Radioresistance of human Non-small Cell Lung Cancer Cells by modulating the DNA damage response and YAP1-induced Epithelial-mesenchymal Transition.

Objectives: Radiotherapy has played a limited role in the treatment of non-small cell lung cancer (NSCLC) due to the risk of tumour radioresistance. We previously established the radioresistant non-small cell lung cancer (NSCLC) cell line H460R. In this study, we identified differentially expressed genes between these radioresistant H460R cells and their radiosensitive parent line. We further evaluated the role of a differentially expressed gene, ITGB1, in NSCLC cell radioresistance and as a potential target for improving radiosensitivity. Materials and Methods: The radiosensitivity of NSCLC cells was evaluated by flow cytometry, colony formation assays, immunofluorescence, and Western blotting. Bioinformatics assay was used to identify the effect of ITGB1 and YAP1 expression in NSCLC tissues. Results: ITGB1 mRNA and protein expression levels were higher in H460R than in the parental H460 cells. We observed lower clonogenic survival and cell viability and a higher rate of apoptosis of ITGB1-knockdown A549 and H460R cells than of wild type cells post-irradiation. Transfection with an ITGB1 short hairpin (sh) RNA enhanced radiation-induced DNA damage and G2/M phase arrest. Moreover, ITGB1 induced epithelial-mesenchymal transition (EMT) of NSCLC cells. Silencing ITGB1 suppressed the expression and intracellular translocation of Yes-associated protein 1 (YAP1), a downstream effector of ITGB1. Conclusions: ITGB1 may induce radioresistance via affecting DNA repair and YAP1-induced EMT. Taken together, our data suggest that ITGB1 is an attractive therapeutic target to overcome NSCLC cell radioresistance.

A disposable acoustofluidic chip for nano/microparticle separation using unidirectional acoustic transducers.

Separation of nano/microparticles based on surface acoustic waves (SAWs) has shown great promise for biological, chemical, and medical applications ranging from sample purification to cancer diagnosis. However, the permanent bonding of a microchannel onto relatively expensive piezoelectric substrates and excitation transducers renders the SAW separation devices non-disposable. This limitation not only requires cumbersome cleaning and increased labor and material costs, but also leads to cross-contamination, preventing their implementation in many biological, chemical, and medical applications. Here, we demonstrate a high-performance, disposable acoustofluidic platform for nano/microparticle separation. Leveraging unidirectional interdigital transducers (IDTs), a hybrid channel design with hard/soft materials, and tilted-angle standing SAWs (taSSAWs), our disposable acoustofluidic devices achieve acoustic radiation forces comparable to those generated by existing permanently bonded, non-disposable devices. Our disposable devices can separate not only microparticles but also nanoparticles. Moreover, they can differentiate bacteria from human red blood cells (RBCs) with a purity of up to 96%. Altogether, we developed a unidirectional IDT-based, disposable acoustofluidic platform for micro/nanoparticle separation that can achieve high separation efficiency, versatility, and biocompatibility.

Transcription levels and prognostic significance of the NFI family members in human cancers.

BACKGROUND: The nuclear factor I (NFI) is a family of transcription factors consisting of four distinct but closely related genes, NFIA, NFIB, NFIC and NFIX, which are important in the development of various tissues and organs in mammals. Recent study results have shown that NFI family may play a critical role in the progression of various human tumors and have been identified as key tumor suppressors and oncogenes for many cancers. However, the expression levels and distinctive prognostic values of the NFI family remain poorly explored in most cancers. MATERIALS AND METHODS: In the present study, the differences in mRNA expression of the NFI family in various cancers were investigated using the Oncomine and TCGA databases, and the mRNA expression, genetic alteration and DNA methylation of the NFI family members in various cancers were examined using cBioPortal for Cancer Genomics. In addition, the prognostic significance of the NFI family was assessed in multiple cancers using the Kaplan-Meier plotter (KM plotter) and SurvExpress databases. RESULTS: The mRNA expression levels in the NFI family were significantly downregulated in most cancers compared with normal tissues and DNA hypermethylation might downregulate the NFI family expression. Although NFIX expression was not downregulated in kidney, colorectal and prostate cancers. Furthermore, NFIB expression was upregulated in gastric cancer. Further survival analyses based on the KM plotter and SurvExpress databases showed dysregulations of the NFI genes were significantly correlated with survival outcomes in breast, lung, and head and neck cancers. Decreased expression levels of NFIA, NFIB and NFIC were associated with poor overall survival (OS) in head and neck cancer. Low mRNA expression of NFIA and NFIB was significantly associated with OS and first progression in lung adenocarcinoma, but not in lung squamous cell carcinoma. In addition, potential correlations between NFI family members and survival outcomes were also observed in liver, esophageal, kidney and cervical cancer. CONCLUSION: The results from the present study indicated certain members of the NFI family could be promising therapeutic targets and novel prognostic biomarkers for human cancers.

Multiple metachronous rare primary malignant tumors: A case report.

Multiple primary malignant tumors (MPMTs) are rarely seen among the patients with malignant neoplasms. Moreover, the existence of five MPMTs in the same patient is an extremely rare phenomenon. In this case, a 42-year-old male patient developed five metachronous MPMTs within 16 years and the duration between each malignant tumor shortened with the progression of the disease. Multidisciplinary treatments were used on this patient and he fought against the cancers until the end of his life. Our report provides us with a new awareness of MPMTs, which should be considered when we come across with cancer patients who develop various unexplainable symptoms after the diagnosis of the first neoplasm.

A novel role for NFIA in restoring radiosensitivity in radioresistant NSCLC cells by downregulating the AKT and ERK pathways.

Radioresistance remains the most challenging issue leading to radiotherapy failure in the treatment of non-small cell lung cancer (NSCLC). The nuclear factor IA (NFIA) is associated with tumor response to treatments in many cancers, but its role in NSCLC radioresistance remains unclear. Here, we established two radioresistant NSCLC cell lines, H226R and H460R, by dose-gradient irradiation to investigate the function of NFIA in NSCLC radioresistance. The results showed a dramatically reduced expression of NFIA in radioresistant cells accompanied with elevated phosphorylation of AKT and ERK, when compared with their parental cells. Overexpression of NFIA restored the sensitivity of radioresistant cells to radiation through increased ionizing radiation (IR)-induced apoptosis and DNA damage by downregulating p-AKT and p-ERK, whereas knockdown of NFIA promoted radioresistance of the parental cells. Our findings suggested that NFIA enhanced cell radiosensitivity by downregulating p-AKT and p-ERK in NSCLC. Our study fills a gap in the field of NFIA and radioresistance, and establishes a mechanistic foundation to improve radiotherapy efficiency in NSCLC patients.

ceRNA network analysis reveals prognostic markers for glioblastoma.

Glioblastoma (GBM) is a common aggressive cancer that originates in the brain, which has a poor prognosis. It is therefore crucial to understand its underlying genetic mechanisms in order to develop novel therapies. The present study aimed to identify some prognostic markers and candidate therapeutic targets for GBM. To do so, RNA expression levels in tumor and normal tissues were compared by microarray analysis. The differential expression of RNAs in normal and cancer tissues was analyzed, and a competing endogenous RNA (ceRNA) network was constructed for pathway analysis. The results revealed that RNA expression patterns were considerably different between normal and tumor samples. A ceRNA network was therefore constructed with the differentially expressed RNAs. ETS variant 5 (ETV5), myocyte enhancer factor 2C and ETS transcription factor (ELK4) were considerably enriched in the significant pathway of 'transcriptional misregulation in cancer'. In addition, prognostic analysis demonstrated that ETV5 and ELK4 expression levels were associated with the survival time of patients with GBM. These results suggested that ELK4 and ETV5 may be prognostic markers for GBM, and that their microRNAs may be candidate therapeutic targets.

Subgroup analysis reveals molecular heterogeneity and provides potential precise treatment for pancreatic cancers.

BACKGROUND: The relationship between molecular heterogeneity and clinical features of pancreatic cancer remains unclear. In this study, pancreatic cancer was divided into different subgroups to explore its specific molecular characteristics and potential therapeutic targets. PATIENTS AND METHODS: Expression profiling data were downloaded from The Cancer Genome Atlas database and standardized. Bioinformatics techniques such as unsupervised hierarchical clustering was used to explore the optimal molecular subgroups in pancreatic cancer. Clinical pathological features and pathways in each subgroup were also analyzed to find out the potential clinical applications and initial promotive mechanisms of pancreatic cancer. RESULTS: Pancreatic cancer was divided into three subgroups based on different gene expression features. Patients included in each subgroup had specific biological features and responded significantly different to chemotherapy. CONCLUSION: Three distinct subgroups of pancreatic cancer were identified, which means that patients in each subgroup might benefit from targeted individual management.

A Convenient and Sensitive LC-MS/MS Method for Simultaneous Determination of Carbadox- and Olaquindox-Related Residues in Swine Muscle and Liver Tissues.

This paper presents a convenient and sensitive LC-MS/MS method for the simultaneous determination of carbadox and olaquindox residues, including desoxyolaquindox (DOLQ), desoxycarbadox (DCBX), quinoxaline-2-carboxylic acid (QCA), 3-methyl-quinoxaline-2-carboxylic acid (MQCA), and the glycine conjugates of QCA and MQCA (namely, QCA-glycine and MQCA-glycine, resp.) in swine muscle and liver tissues. Tissue samples were extracted with 2% metaphosphoric acid in 20% methanol and cleaned up by solid-phase extraction (SPE) on a mixed-mode anion-exchange column (Oasis MAX). Analysis was performed on a C18 column by detection with mass spectrometry in the multiple reaction monitoring (MRM) mode. The limits of detection (LODs) of the six analytes were determined to be 0.01 µg·kg-1 to 0.25 µg·kg-1, and the limits of quantification (LOQs) were 0.02 µg·kg-1 to 0.5 µg·kg-1. The total recoveries of the six analytes in all tissues were higher than 79.1% with the RSD% less than 9.2%. The developed method can determine the real residue level of QCA and MQCA, whether they are present in free form or as glycine conjugates in tissues, together with the carcinogenic desoxy metabolites DCBX and DOLQ with high recovery. Therefore, this method was suitable for routine analysis of residue control programmes and the residue depletion study of CBX and OLQ on swine.

Aggregation of lipid rafts activates c-met and c-Src in non-small cell lung cancer cells.

BACKGROUND: Activation of c-Met, a receptor tyrosine kinase, induces radiation therapy resistance in non-small cell lung cancer (NSCLC). The activated residual of c-Met is located in lipid rafts (Duhon et al. Mol Carcinog 49:739-49, 2010). Therefore, we hypothesized that disturbing the integrity of lipid rafts would restrain the activation of the c-Met protein and reverse radiation resistance in NSCLC. In this study, a series of experiments was performed to test this hypothesis. METHODS: NSCLC A549 and H1993 cells were incubated with methyl-ß-cyclodextrin (MßCD), a lipid raft inhibitor, at different concentrations for 1 h before the cells were X-ray irradiated. The following methods were used: clonogenic (colony-forming) survival assays, flow cytometry (for cell cycle and apoptosis analyses), immunofluorescence microscopy (to show the distribution of proteins in lipid rafts), Western blotting, and biochemical lipid raft isolation (purifying lipid rafts to show the distribution of proteins in lipid rafts). RESULTS: Our results showed that X-ray irradiation induced the aggregation of lipid rafts in A549 cells, activated c-Met and c-Src, and induced c-Met and c-Src clustering to lipid rafts. More importantly, MßCD suppressed the proliferation of A549 and H1993 cells, and the combination of MßCD and radiation resulted in additive increases in A549 and H1993 cell apoptosis. Destroying the integrity of lipid rafts inhibited the aggregation of c-Met and c-Src to lipid rafts and reduced the expression of phosphorylated c-Met and phosphorylated c-Src in lipid rafts. CONCLUSIONS: X-ray irradiation induced the aggregation of lipid rafts and the clustering of c-Met and c-Src to lipid rafts through both lipid raft-dependent and lipid raft-independent mechanisms. The lipid raft-dependent activation of c-Met and its downstream pathways played an important role in the development of radiation resistance in NSCLC cells mediated by c-Met. Further studies are still required to explore the molecular mechanisms of the activation of c-Met and c-Src in lipid rafts induced by radiation.

Sp1-driven up-regulation of miR-19a decreases RHOB and promotes pancreatic cancer.

Cancer treatment alters microRNA (miRNA) expression, revealing potential therapeutic targets (oncotarget). Here we treated pancreatic cancer (ASPC-1) cells with either recombinant human endostatin (rh-endostatin) or gemcitabine. Then high-throughput sequencing assay was performed to screen for altered miRNAs. Both treatments decreased levels of MiR-19a. We found that miR-19a stimulated cell proliferation, migration, invasion in vitro and tumor growth in vivo. High levels of miR-19a correlated with poor prognosis in patients. Ras homolog family member B (RHOB) was identified as a direct target of miR-19a. Furthermore, RHOB was down-regulated in human pancreatic cancer samples. Restoration of RHOB induced apoptosis, inhibited proliferation and migration of ASPC-1 cells. SP-1 was identified as an upstream transcription factor of miR-19a gene, promoting miR-19a transcription. Rh-endostatin decreased miR-19a expression by down-regulating SP-1. These findings suggest that miR-19a is a potential therapeutic target in pancreatic cancer.

Cancer stem cells from a rare form of glioblastoma multiforme involving the neurogenic ventricular wall.

BACKGROUND: The cancer stem cell (CSC) hypothesis posits that deregulated neural stem cells (NSCs) form the basis of brain tumors such as glioblastoma multiforme (GBM). GBM, however, usually forms in the cerebral white matter while normal NSCs reside in subventricular and hippocampal regions. We attempted to characterize CSCs from a rare form of glioblastoma multiforme involving the neurogenic ventricular wall. METHODS: We described isolating CSCs from a GBM involving the lateral ventricles and characterized these cells with in vitro molecular biomarker profiling, cellular behavior, ex vivo and in vivo techniques. RESULTS: The patient's MRI revealed a heterogeneous mass with associated edema, involving the left subventricular zone. Histological examination of the tumor established it as being a high-grade glial neoplasm, characterized by polygonal and fusiform cells with marked nuclear atypia, amphophilic cytoplasm, prominent nucleoli, frequent mitotic figures, irregular zones of necrosis and vascular hyperplasia. Recurrence of the tumor occurred shortly after the surgical resection. CD133-positive cells, isolated from the tumor, expressed stem cell markers including nestin, CD133, Ki67, Sox2, EFNB1, EFNB2, EFNB3, Cav-1, Musashi, Nucleostemin, Notch 2, Notch 4, and Pax6. Biomarkers expressed in differentiated cells included Cathepsin L, Cathepsin B, Mucin18, Mucin24, c-Myc, NSE, and TIMP1. Expression of unique cancer-related transcripts in these CD133-positive cells, such as caveolin-1 and -2, do not appear to have been previously reported in the literature. Ex vivo organotypic brain slice co-culture showed that the CD133+ cells behaved like tumor cells. The CD133-positive cells also induced tumor formation when they were stereotactically transplanted into the brains of the immune-deficient NOD/SCID mice. CONCLUSIONS: This brain tumor involving the neurogenic lateral ventricular wall was comprised of tumor-forming, CD133-positive cancer stem cells, which are likely the driving force for the rapid recurrence of the tumor in the patient.

Transcriptome analysis of neural progenitor cells by a genetic dual reporter strategy.

Global analysis of stem/progenitor cells promises new insight into mechanisms that govern self-renewal and cellular potential, an unresolved question of stem/progenitor cell biology. Despite rapid advance of genome-wide profiling methods, the difficulty in cell purification remains a major challenge for global analysis of somatic stem/progenitor cells. Genetic tagging with a reporter provides a powerful tool for identification and isolation of a specific mature cell type; however, for stem/progenitor cells, reporter retention by progeny may be a concern for impurity. Here, we describe a genetic system combining a progenitor cell specific label with a second tag for marking differentiation. We present evidence that differential labeling of neural progenitor cells and their progeny enables prospective purification of these two cell types, whereas isolation based on a single marker compromises the purity of the intended progenitor population. Comparative expression profiling between the purified progenitors and progeny documents a neural progenitor cell transcriptome and uncovers an important role of Tyro3/Axl/Mer receptor tyrosine kinases in the maintenance of neural progenitor cells. This study establishes a general strategy for isolation of somatic stem/progenitor cells and provides a transcriptome database of neural progenitor cells useful for identification of causal factors of neural progenitor cell state, global dissection of epigenetic control of cellular potential, as well as for developing biomarkers or targets of brain cancer stem/initiating cells for therapeutic interventions.

An alternative processing of integrin alpha(v) subunit in tumor cells by membrane type-1 matrix metalloproteinase.

Membrane type-1 matrix metalloproteinase (MT1-MMP) and alpha(v)beta(3) integrin are both essential to cell invasion. Maturation of integrin pro-alpha(v)chain (pro-alpha(v)) involves its cleavage by proprotein convertases (PC) to form the disulfide-bonded 125-kDa heavy and 25-kDa light alpha chains. Our report presents evidence of an alternative pathway of pro-alpha(v) processing involving MT1-MMP. In breast carcinoma MCF7 cells deficient in MT1-MMP, pro-alpha(v) is processed by a conventional furin-like PC, and the mature alpha(v) integrin subunit is represented by the 125-kDa heavy chain and the 25-kDa light chain commencing from the N-terminal Asp(891). In contrast, in cells co-expressing alpha(v)beta(3) and MT1-MMP, MT1-MMP functions as an integrin convertase. MT1-MMP specifically cleaves pro-alpha(v), generating a 115-kDa heavy chain with the truncated C terminus and a 25-kDa light chain commencing from the N-terminal Leu(892). PC-cleavable alpha(3) and alpha(5) but not the PC-resistant alpha(2) integrin subunit are also susceptible to MT1-MMP cleavage. These novel mechanisms involved in the processing of integrin alpha subunits underscore the significance and complexity of interactions between MT1-MMP and adhesion receptors and suggest that regulation of integrin functionality may be an important role of MT1-MMP in migrating tumor cells.