Objective: : Radiation is known to induce autophagy in malignant glioma cells whether it is cytocidal or cytoprotective.Dexamethasone is frequently used to reduce tumor-associated brain edema, especially during radiation therapy. The purpose of the study was to determine whether and how dexamethasone affects autophagy in irradiated malignant glioma cells and to identify possible intervening molecular pathways. Methods: : We prepared p53 mutant U373 and LN229 glioma cell lines, which varied by phosphatase and tensin homolog (PTEN) mutational status and were used to make U373 stable transfected cells expressing GFP-LC3 protein. After performing cell survival assay after irradiation, the IC50 radiation dose was determined. Dexamethasone dose (10 µM) was determined from the literature and added to the glioma cells 24 hours before the irradiation. The effect of adding dexamethasone was evaluated by cell survival assay or clonogenic assay and cell cycle analysis. Measurement of autophagy was visualized by western blot of LC3-I/LC3-II and quantified by the GFP-LC3 punctuated pattern under fluorescence microscopy and acridine orange staining for acidic vesicle organelles by flow cytometry. Results: : Dexamethasone increased cell survival in both U373 and LN229 cells after irradiation. It interfered with autophagy after irradiation differently depending on the PTEN mutational status : the autophagy decreased in U373 (PTEN-mutated) cells but increased in LN229 (PTEN wild-type) cells. Inhibition of protein kinase B (AKT) phosphorylation after irradiation by LY294002 reversed the dexamethasone-induced decrease of autophagy and cell death in U373 cells but provoked no effect on both autophagy and cell survival in LN229 cells. After ATG5 knockdown, radiation-induced autophagy decreased and the effect of dexamethasone also diminished in both cell lines. The diminished autophagy resulted in a partial reversal of dexamethasone protection from cell death after irradiation in U373 cells; however, no significant change was observed in surviving fraction LN229 cells. Conclusion : Dexamethasone increased cell survival in p53 mutated malignant glioma cells and increased autophagy in PTENmutant malignant glioma cell but not in PTEN-wildtype cell. The difference of autophagy response could be mediated though the phosphatidylinositol 3-kinase/AKT/mammalian target of rapamycin signaling pathway.

Cancer is a complex, heterogeneic, and dynamic disease involving multiple gene-environment interactions, and affecting numerous biological pathways. As such, the development of reliable and robust non-invasive platforms constitutes a vital step toward realizing the potential of precision medicine. Distant metastases harbor unique genomic characteristics that are not detectable in the corresponding primary tumor of the same patient, and metastases located at different sites show considerable intra-patient heterogeneity. Thus, the analysis of the resected primary tumor alone or, if possible, re-evaluation of tumor characteristics based on the biopsy of the most accessible metastasis, may not reveal sufficient information for treatment decisions. Here, we propose that this dilemma can be solved by a new diagnostic concept: liquid biopsy, that is, the analysis of therapeutic targets and drug resistance-conferring gene mutations in or on circulating tumor cells (CTCs). Finally, the analysis of the resected primary tumor alone may provide misleading information with regard to the characteristics of metastases, the key target for systemic anticancer therapy. Liquid biopsies are noninvasive tests using blood or fluids that detect CTCs or the products of tumors, such as fragments of nucleotides or proteins that are shed into biological fluids from the primary or metastatic tumors. Such biopsies are expected to be informative or easily accessible tools to provide comprehensive information regarding cancers beyond conventional biopsies. Thus, this review addresses the use of CTCs in cancer detection, diagnosis and monitoring and discusses the direction of its clinical application in cancer patient care.
Biopsy ; Diagnosis ; Early Detection of Cancer ; Gene-Environment Interaction ; Humans ; Neoplasm Metastasis ; Neoplastic Cells, Circulating ; Nucleotides ; Patient Care ; Population Characteristics ; Precision Medicine

PURPOSE: We sought to develop a matrix assisted laser desorption ionization-time of flight (MALDI-TOF)-based, ovarian cancer (OVC), low-mass-ion discriminant equation (LOME) and to evaluate a possible supportive role for triple-TOF mass analysis in identifying metabolic biomarkers. MATERIALS AND METHODS: A total of 114 serum samples from patients with OVC and benign ovarian tumors were subjected to MALDI-TOF analysis and a total of 137 serum samples from healthy female individuals and patients with OVC, colorectal cancer, hepatobiliary cancer, and pancreatic cancer were subjected to triple-TOF analysis. An OVC LOME was constructed by reference to the peak intensity ratios of discriminatory low-mass ion (LMI) pairs. Triple-TOF analysiswas used to select and identify metabolic biomarkers for OVC screening. RESULTS: Three OVC LOMEs were finally constructed using discriminatory LMI pairs (137.1690 and 84.4119 m/z; 496.5022 and 709.7642 m/z; and 524.5614 and 709.7642 m/z); all afforded accuracies of > 90%. The LMIs at 496.5022 m/z and 524.5614 m/z were those of lysophosphatidylcholine (LPC) 16:0 and LPC 18:0. Triple-TOF analysis selected seven discriminative LMIs; each LMI had a specificity > 90%. Of the seven LMIs, fourwith a 137.0455 m/z ion atretention times of 2.04-3.14 minuteswere upregulated in sera from OVC patients; the ion was identified as that derived from hypoxanthine. CONCLUSION: MALDI-TOF–based OVC LOMEs combined with triple-TOF–based OVC metabolic biomarkers allow reliable OVC screening; the techniques are mutually complementary both quantitatively and qualitatively.
Biomarkers ; Colorectal Neoplasms ; Female ; Humans ; Hypoxanthine ; Lysophosphatidylcholines ; Mass Screening ; Mass Spectrometry* ; Ovarian Neoplasms* ; Pancreatic Neoplasms ; Sensitivity and Specificity

PURPOSE: The serum carcinoembryonic antigen (CEA) level has been recognized as a prognostic factor in colorectal cancer, and associated with response of rectal cancer to radiotherapy. This study aimed to identify CEA-interacting proteins in colon cancer cells and observe post-irradiation changes in their expression. MATERIALS AND METHODS: CEA expression in colon cancer cells was examined by Western blot analysis. Using an anti-CEA antibody or IgG as a negative control, immunoprecipitation was performed in colon cancer cell lysates. CEA and IgG immunoprecipitates were used for liquid chromatography–tandem mass spectrometry (LC-MS/MS) analysis. Proteins identified in the CEA immunoprecipitates but not in the IgG immunoprecipitates were selected as CEA-interacting proteins. After radiation treatment, changes in expression of CEA-interacting proteins were monitored by Western blot analysis. RESULTS: CEA expression was higher in SNU-81 cells compared with LoVo cells. The membrane localization of CEA limited the immunoprecipitation results and thus the number of CEA-interacting proteins identified. Only the Ras-related protein Rab-6B and lysozyme C were identified as CEA-interacting proteins in LoVo and SNU-81 cells, respectively. Lysozyme C was detected only in SNU-81, and CEA expression was differently regulated in two cell lines; it was down-regulated in LoVo but up-regulated in SNU-81 in radiation dosage-dependent manner. CONCLUSION: CEA-mediated radiation response appears to vary, depending on the characteristics of individual cancer cells. The lysozyme C and Rab subfamily proteins may play a role in the link between CEA and tumor response to radiation, although further studies are needed to clarify functional roles of the identified proteins.
Blotting, Western ; Carcinoembryonic Antigen ; Cell Line ; Colon* ; Colonic Neoplasms* ; Colorectal Neoplasms ; Immunoglobulin G ; Immunoprecipitation ; Mass Spectrometry ; Membranes ; Muramidase ; Radiotherapy ; Rectal Neoplasms

PURPOSE: The serum carcinoembryonic antigen (CEA) level has been recognized as a prognostic factor in colorectal cancer, and associated with response of rectal cancer to radiotherapy. This study aimed to identify CEA-interacting proteins in colon cancer cells and observe post-irradiation changes in their expression. MATERIALS AND METHODS: CEA expression in colon cancer cells was examined by Western blot analysis. Using an anti-CEA antibody or IgG as a negative control, immunoprecipitation was performed in colon cancer cell lysates. CEA and IgG immunoprecipitates were used for liquid chromatography–tandem mass spectrometry (LC-MS/MS) analysis. Proteins identified in the CEA immunoprecipitates but not in the IgG immunoprecipitates were selected as CEA-interacting proteins. After radiation treatment, changes in expression of CEA-interacting proteins were monitored by Western blot analysis. RESULTS: CEA expression was higher in SNU-81 cells compared with LoVo cells. The membrane localization of CEA limited the immunoprecipitation results and thus the number of CEA-interacting proteins identified. Only the Ras-related protein Rab-6B and lysozyme C were identified as CEA-interacting proteins in LoVo and SNU-81 cells, respectively. Lysozyme C was detected only in SNU-81, and CEA expression was differently regulated in two cell lines; it was down-regulated in LoVo but up-regulated in SNU-81 in radiation dosage-dependent manner. CONCLUSION: CEA-mediated radiation response appears to vary, depending on the characteristics of individual cancer cells. The lysozyme C and Rab subfamily proteins may play a role in the link between CEA and tumor response to radiation, although further studies are needed to clarify functional roles of the identified proteins.
Blotting, Western ; Carcinoembryonic Antigen ; Cell Line ; Colon* ; Colonic Neoplasms* ; Colorectal Neoplasms ; Immunoglobulin G ; Immunoprecipitation ; Mass Spectrometry ; Membranes ; Muramidase ; Radiotherapy ; Rectal Neoplasms

(E)-3-Phenyl-1-(2-pyrrolyl)-2-propenone (PPP) is a pyrrole derivative of chalcone, in which the B-ring of chalcone linked to β-carbon is replaced by pyrrole group. While pyrrole has been studied for possible Src inhibition activity, chalcone, especially the substituents on the B-ring, has shown pharmaceutical, anti-inflammatory, and anti-oxidant properties via inhibition of NF-κB activity. Our study is aimed to investigate whether this novel synthetic compound retains or enhances the pharmaceutically beneficial activities from the both structures. For this purpose, inflammatory responses of lipopolysaccharide (LPS)-treated RAW264.7 cells were analyzed. Nitric oxide (NO) production, inducible NO synthase (iNOS) and tumor necrosis factor-α (TNF-α) mRNA expression, and the intracellular inflammatory signaling cascade were measured. Interestingly, PPP strongly inhibited NO release in a dose-dependent manner. To further investigate this anti-inflammatory activity, we identified molecular pathways by immunoblot analyses of nuclear fractions and whole cell lysates prepared from LPS-stimulated RAW264.7 cells with or without PPP pretreatment. The nuclear levels of p50, c-Jun, and c-Fos were significantly inhibited when cells were exposed to PPP. Moreover, according to the luciferase reporter gene assay after cotransfection with either TRIF or MyD88 in HEK293 cells, NF-κB-mediated luciferase activity dose-dependently diminished. Additionally, it was confirmed that PPP dampens the upstream signaling cascade of NF-κB and AP-1 activation. Thus, PPP inhibited Syk, Src, and TAK1 activities induced by LPS or induced by overexpression of these genes. Therefore, our results suggest that PPP displays anti-inflammatory activity via inhibition of Syk, Src, and TAK1 activity, which may be developed as a novel anti-inflammatory drug.
Chalcone* ; Genes, Reporter ; HEK293 Cells ; Luciferases ; Macrophages ; Necrosis ; Nitric Oxide ; Nitric Oxide Synthase ; Phosphotransferases* ; RNA, Messenger ; Transcription Factor AP-1

PURPOSE: Patients show variable responses to chemoradiotherapy (CRT), which is generally administered before surgery for locally advanced rectal cancer (LARC). The aim of this study was to identify molecular markers predictive of CRT responses by analysis of low-mass ions (LMIs) in serum of LARC patients. MATERIALS AND METHODS: LMIs (< 1,000 m/z) in serum obtained before CRT from 73 LARC (cT3-4) patients were profiled using matrix-assisted laser desorption/ionization mass spectrometry. LMIs with higher weighting factors in discriminating CRT responses were selected using principal components analysis and discriminant analysis. Selected LMIs were identified using the Human Metabolome Database. The concentrations of identified LMIs were determined by colorimetric enzyme assay, and compared according to post-CRT pathological stage (ypStage) or Dworak's tumor regression grade (TRG). RESULTS: The nine highest-ranking LMIs were selected. Among them, two LMIs with 137.08 and 169.04 m/z were identified as hypoxanthine (HX) and phosphoenolpyruvic acid (PEP), respectively. Higher HX concentration was observed in patients with ypStage 0-1 compared to ypStage 2-4 (p=0.034) or ypStage 3-4 (p=0.030); a similar difference was observed between TRG 4-3 and TRG 1 (p=0.035). HX > 16.0 muM showed significant association with ypStage 0-1 or TRG 4-3 than ypStage 3-4 (p=0.009) or TRG 1 (p=0.024), respectively. In contrast, a significantly lower concentration of PEP was observed in TRG 4-3 compared with TRG 2-1 (p=0.012). CONCLUSION: Findings of this study demonstrated that serum concentrations of HX and PEP, identified using LMI profiling, may be useful for predicting the CRT response of LARC patients before treatment.
Biological Markers* ; Chemoradiotherapy* ; Enzyme Assays ; Humans ; Hypoxanthine* ; Ions ; Mass Spectrometry ; Metabolome ; Phosphoenolpyruvate ; Rectal Neoplasms*

Overexpression of amyloid precursor protein with the Swedish mutation causes abnormal hyperphosphorylation of the microtubule-associated protein tau. Hyperphosphorylated isoforms of tau are major components of neurofibrillary tangles, which are histopathological hallmarks of Alzheimer's disease. Protein phosphatase 2A (PP2A), a major tau protein phosphatase, consists of a structural A subunit, catalytic C subunit, and a variety of regulatory B subunits. The B subunits have been reported to modulate function of the PP2A holoenzyme by regulating substrate binding, enzyme activity, and subcellular localization. In the current study, we characterized regulatory B subunit-specific regulation of tau protein phosphorylation. We showed that the PP2A B subunit PPP2R2A mediated dephosphorylation of tau protein at Ser-199, Ser-202/Thr-205, Thr-231, Ser-262, and Ser-422. Down-regulation of PPP2R5D expression decreased tau phosphorylation at Ser-202/Thr-205, Thr-231, and Ser-422, which indicates activation of the tau kinase glycogen synthase kinase 3 beta (GSK3beta) by PP2A with PPP2R5D subunit. The level of activating phosphorylation of the GSK3beta kinase Akt at Thr-308 and Ser-473 were both increased by PPP2R5D knockdown. We also characterized B subunit-specific phosphorylation sites in tau using mass spectrometric analysis. Liquid chromatography-mass spectrometry revealed that the phosphorylation status of the tau protein may be affected by PP2A, depending on the specific B subunits. These studies further our understanding of the function of various B subunits in mediating site-specific regulation of tau protein phosphorylation.
Alzheimer Disease ; Amyloid ; Catalytic Domain ; Down-Regulation ; Glycogen Synthase Kinase 3 ; Negotiating ; Neurofibrillary Tangles ; Phosphorylation* ; Phosphotransferases ; Protein Isoforms ; Protein Phosphatase 2* ; Spectrum Analysis ; tau Proteins*

BACKGROUND/AIMS: We performed a prospective study to compare the feasibility, safety and tolerance among ultrathin transnasal (UT-N), thin transnasal (T-N) and ultrathin oral (UT-O) esophagogastroduodenoscopy. METHODS: Two narrow diameter endoscopes (phi=5.2 mm for UT-N and UT-O, phi=6.5 mm for T-N) were used. The operator factors and patient factors were quantified by a visual analogue scale. RESULTS: The procedure was successfully completed in 100 of 100 patients in the UT-O group. The T-N group, when compared with the UT-N group, accounted for a significantly higher portion of failure (14% vs 3%, respectively, p=0.000), more cases of epistaxis (11% vs 3%, respecttively, p=0.013) and more complaints of nasal pain (17% vs 6%, p=0.016). The overall quality of the exam was significantly higher in the UT-N group (UT-N, 8.7; T-N, 8.1; UT-O, 8.2, p=0.04). The frequency of a incurring a gag reflex was significantly lower in the UT-N group (UT-N, 1.26; T-N, 1.48; UT-O, 2.94, p= 0.000). The patients' score for overall general satisfaction was higher in the UT-N group (UT-N, 8.5; T-N, 7.8; UT-O, 7.7, p=0.006). Nausea was significantly reduced the in UT-N group (UT-N, 8.2; T-N, 7.8; UT-O, 7.3, p= 0.003). Patients in the UT-N group were more willing to repeat the same procedure (UT-N, 82%; T-N, 65%; UT-O, 71%, p=0.046). CONCLUSIONS: Ultrathin transnasal endoscopy is more feasible, safe and comfortable compared with the thin transnasal endoscopy or when compared with either instrument that was passed orally.
Endoscopes ; Endoscopy ; Epistaxis ; Humans ; Nausea ; Prospective Studies ; Reflex

BACKGROUND/AIMS: Despite of increasing numbers of reports on intraductal papillary mucinous tumor (IPMT), there is still difficulty in its' diagnosis, treatment and prediction of prognosis. The purpose of this multicenter study was to evaluate the clinico-pathological features of IPMT in Korea and suggest the prediction criteria of malignancy in IPMT. METHODS: We retrospectively reviewed the clinico-pathological data of 208 patients who underwent operations with IPMT between 1993 and 2002 at 28 institutes in Korea. RESULTS: Of the 208 patients with a mean age of 60.5+/-9.7 years, 147 were men and 61 were women. 124 patients underwent pancreatoduodenectomy, 42 distal pancreatectomy, 17 total pancreatectomy, 25 limited pancreas resection. Benign cases were 128 (adenoma (n=62), borderline (n=66)) and malignant cases were 80 (non-invasive (n=29), invasive (n=51)). A significant difference in 5-year survival was observed between benign and malignant group (92.6% vs. 65.3%; p=0.006). Of the 6 factors (age, location, duct dilatation, tumor appearance, main duct type, and tumor size) that showed the statistical difference in univariate analysis between benign and malignant group, we found three significant factors (tumor appearance (p=0.009), tumor size (p=0.023), and dilated duct size (p=0.010)) by multivariate analysis. CONCLUSION: Although overall prognosis of IPMT is superior to ordinary pancreatic cancer, more curative surgery is recommended in malignant IPMT. Tumor appearance (papillary), tumor size (> or =30 mm) and dilated duct size (> or = 12 mm) can be used as preoperative indicators of malig-nancy in IPMT.
Academies and Institutes ; Diagnosis ; Dilatation ; Female ; Humans ; Korea* ; Male ; Mucins* ; Multivariate Analysis ; Pancreas* ; Pancreatectomy ; Pancreatic Neoplasms ; Pancreaticoduodenectomy ; Prognosis ; Retrospective Studies