The Correlation of Fluorescence of Protoporphyrinogen IX and Status of Isocitrate Dehydrogenase in Gliomas

Neurosurgery ◽  
2019 ◽  
Vol 87 (2) ◽  
pp. 408-417 ◽  
Author(s):  
Shigeo Ohba ◽  
Kazuhiro Murayama ◽  
Kiyonori Kuwahara ◽  
Eriel Sandika Pareira ◽  
Shunsuke Nakae ◽  
...  
Keyword(s):  
Isocitrate Dehydrogenase ◽  
Aminolevulinic Acid ◽  
Tumor Resection ◽  
Glioma Cells ◽  
World Health ◽  
Heme Oxygenase 1 ◽  
Vitro Assay ◽  
Mutant Idh1 ◽  
Intraoperative Fluorescence

Abstract BACKGROUND The extent of resection has been reported to be associated with overall survival in gliomas. The use of 5-aminolevulinic acid (5-ALA) has been recognized to increase the extent of tumor resection. OBJECTIVE To evaluate what factors affect the intraoperative fluorescence after administration of 5-ALA in gliomas. METHODS Correlation of intraoperative fluorescence and several clinical, radiographic, molecular biologic, and histopathologic characters was retrospectively evaluated in 104 patients (53 males and 51 females; mean age 54.2 yr) with gliomas at our institution. To clarify the mechanisms that mutant isocitrate dehydrogenase (IDH) affect the intraoperative fluorescence, in Vitro experiments using genetically engineered glioma cells harboring mutant IDH1 were performed. RESULTS Intraoperative fluorescence was observed in 82 patients (78.8%). In addition to age, magnetic resonance imaging enhancement, World Health Organization grades, and MIB-1 index, the status of IDH was revealed to be correlated with intraoperative fluorescence. In Vitro assay revealed that mutant IDH indirectly reduced the amount of exogenous 5-ALA-derived protoporphyrinogen IX in glioma cells by increasing activity of ferrochelatase and heme oxygenase 1. CONCLUSION Mutant IDH1/2-induced metabolite changes of exogenous 5-ALA were suggested to contribute to the lesser intraoperative fluorescence in gliomas with mutant IDH1/2 than in those without.

Identification of a Potential Inhibitor Targeting MurC Ligase of the Drug Resistant Pseudomonas aeruginosa Strain through Structure-Based Virtual Screening Approach and In Vitro Assay

2019 ◽  
Vol 20 (14) ◽  
pp. 1203-1212
Author(s):  
Abdelmonaem Messaoudi ◽  
Manel Zoghlami ◽  
Zarrin Basharat ◽  
Najla Sadfi-Zouaoui
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Virtual Screening ◽  
Homology Model ◽  
Generation Cephalosporin ◽  
World Health ◽  
Resistant Bacteria ◽  
Antimicrobial Drugs ◽  
Vitro Assay ◽  
Priority List

Background & Objective: Pseudomonas aeruginosa shows resistance to a large number of antibiotics, including carbapenems and third generation cephalosporin. According to the World Health Organization global report published in February 2017, Pseudomonas aeruginosa is on the priority list among resistant bacteria, for which new antibiotics are urgently needed. Peptidoglycan serves as a good target for the discovery of novel antimicrobial drugs. Methods: Biosynthesis of peptidoglycan is a multi-step process involving four mur enzymes. Among these enzymes, UDP-N-acetylmuramate-L-alanine ligase (MurC) is considered to be an excellent target for the design of new classes of antimicrobial inhibitors in gram-negative bacteria. Results: In this study, a homology model of Pseudomonas aeruginosa MurC ligase was generated and used for virtual screening of chemical compounds from the ZINC Database. The best screened inhibitor i.e. N, N-dimethyl-2-oxo-2,3-dihydro-1H-1,3-benzodiazole-5-sulfonamide was then validated experimentally through inhibition assay. Conclusion: The presented results based on combined computational and in vitro analysis open up new horizons for the development of novel antimicrobials against this pathogen.

Cadherin 13 overexpression as an important factor related to the absence of tumor fluorescence in 5-aminolevulinic acid–guided resection of glioma

2013 ◽  
Vol 119 (5) ◽  
pp. 1331-1339 ◽  
Author(s):  
Tomonari Suzuki ◽  
Satoru Wada ◽  
Hidetaka Eguchi ◽  
Jun-ichi Adachi ◽  
Kazuhiko Mishima ◽  
...  
Keyword(s):  
Gene Expression ◽  
Gene Expression Analysis ◽  
Aminolevulinic Acid ◽  
Glioma Cells ◽  
Mechanisms Of Action ◽  
Negative Regulators ◽  
Resection Rate ◽  
Fluorescent Signal ◽  
U251 Cells

Object Gliomas contain aggressive malignant cancer, and resection rate remains an important factor in treatment. Currently, fluorescence-guided resection using orally administered 5-aminolevulinic acid (5-ALA) has proved to be beneficial in improving the prognosis of patients with gliomas. 5-ALA is metabolized to protoporphyrin IX (PpIX) that accumulates selectively in the tumor and exhibits strong fluorescence upon excitation, but glioma cells do not always respond to 5-ALA, which can result in incomplete or excessive resection. Several possible mechanisms for this phenomenon have been suggested, but they remain poorly understood. To clarify the probable mechanisms underlying the variable induction of fluorescence and to improve fluorescence-guided surgery, the authors searched for key negative regulators of fluorescent signal induced by 5-ALA. Methods A comprehensive gene expression analysis was performed using microarrays in 11 pairs of tumor specimens, fluorescence-positive and fluorescence-negative tumors, and screened genes overexpressed specifically in fluorescence-negative tumors as the possible candidates for key negative regulators of 5-ALA–induced fluorescence. The most possible candidate was selected through annotation analysis in combination with a comparison of expression levels, and the relevance of expression of the selected gene to 5-ALA–induced fluorescence in tumor tissues was confirmed in the quantified expression levels. The biological significance of an identified gene in PpIX accumulation and 5-ALA–induced fluorescence was evaluated by in vitro PpIX fluorescence intensity analysis and in vitro PpIX fluorescence molecular imaging in 4 human glioblastoma cell lines (A1207, NMCG1, U251, and U373). Knockdown analyses using a specific small interfering RNA in U251 cells was also performed to determine the mechanisms of action and genes working as partners in the 5-ALA metabolic pathway. Results The authors chose 251 probes that showed remarkably high expression only in fluorescent-negative tumors (median intensity of expression signal > 1.0), and eventually the cadherin 13 gene (CDH13) was selected as the most possible determinant of 5-ALA–induced fluorescent signal in gliomas. The mean expression level of CDH13 in the fluorescence-negative gliomas was statistically higher than that in positive ones (p = 0.027), and knockdown of CDH13 expression enhanced the fluorescence image and increased the amount of PpIX 13-fold over controls (p < 0.001) in U251 glioma cells treated with 5-ALA. Comprehensive gene expression analysis of the CDH13-knockdown U251 cells demonstrated another two genes possibly involved in the PpIX biosynthesis: ATP-binding cassette transporter (ABCG2) significantly decreased in the CDH13 knockdown, while oligopeptide transporter 1 (PEPT1) increased. Conclusions The cadherin 13 gene might play a role in the PpIX accumulation pathway and act as a negative regulator of 5-ALA–induced fluorescence in glioma cells. Although further studies to clarify the mechanisms of action in the 5-ALA metabolic pathway would be indispensable, the results of this study might lead to a novel fluorescent marker able to overcome the obstacles of existing fluorescence-guided resection and improve the limited resection rate.

5-Aminolevulinic Acid Fluorescence-Guided Resection of Intramedullary Ependymoma: Report of 9 Cases

2012 ◽  
Vol 72 (2) ◽  
pp. ons159-ons168 ◽  
Author(s):  
Tomoo Inoue ◽  
Toshiki Endo ◽  
Kenichi Nagamatsu ◽  
Mika Watanabe ◽  
Teiji Tominaga
Keyword(s):  
Spinal Cord ◽  
Aminolevulinic Acid ◽  
Tumor Resection ◽  
Extent Of Resection ◽  
Labeling Index ◽  
World Health ◽  
Histopathological Diagnosis ◽  
Glioma Surgery ◽  
Ki 67 ◽  
Grade Ii

Abstract BACKGROUND: Resection guided by 5-aminolevulinic acid (5-ALA) fluorescence has proved to be useful in intracranial glioma surgery. However, the effects of 5-ALA on spinal cord tumors remain unknown. OBJECTIVE: To evaluate the usefulness of 5-ALA fluorescence-guided resection of intramedullary ependymoma for achieving maximum tumor resection. METHODS: This study included 10 patients who underwent surgical resection of an intramedullary ependymoma. Nine patients were orally administered 5-ALA (20 mg/kg) 2 hours before the induction of anesthesia. 5-ALA fluorescence was visualized with an operating microscope. Tumors were removed in a standardized manner with electro-physiological monitoring. The extent of resection was evaluated on the basis of intra-operative findings and postoperative magnetic resonance imaging. Histopathological diagnosis was established according to World Health Organization 2007 criteria. Cell proliferation was assessed by Ki-67 labeling index. RESULTS: 5-ALA fluorescence was positive in 7 patients (6 grade II and 1 grade III) and negative in 2 patients (grade II). Intraoperative findings were dichotomized: Tumors covered by the cyst were easily separated from the normal parenchyma, whereas tumors without the cyst appeared to be continuous to the spinal cord. In these cases, 5-ALA fluorescence was especially valuable in delineating the ventral and cranial and caudal margins. Ki-67 labeling index was significantly higher in 5-ALA-positive cases compared with 5-ALA-negative cases. All patients improved neurologically or stabilized after surgery. CONCLUSION: 5-ALA fluorescence was useful for detecting tumor margins during surgery for intramedullary ependymoma. When combined with electrophysiological monitoring, fluorescence-guided resection could help to achieve maximum tumor resection safely.

scholarly journals BET degrader inhibits tumor progression and stem-like cell growth via Wnt/β-catenin signaling repression in glioma cells

2020 ◽  
Vol 11 (10) ◽  
Author(s):  
Tao Tian ◽  
Tongqi Guo ◽  
Wei Zhen ◽  
Jianjun Zou ◽  
Fuyong Li
Keyword(s):  
Stem Cell ◽  
Glioma Cells ◽  
World Health ◽  
Tumor Type ◽  
Cycle Arrest ◽  
Bet Inhibitors ◽  
Health Organization ◽  
The Brain

Abstract Based on their histological appearance, gliomas are a very common primary tumor type of the brain and are classified into grades, Grade I to Grade IV, of the World Health Organization. Treatment failure is due to the cancer stem cells (CSC) phenotype maintenance and self-renewal. BET degraders such as ZBC260 represents a novel class of BET inhibitors that act by inducing BET proteins degradation. This study explores the mode of action and effects of ZBC260 in vivo and in vitro against glioma. By inhibiting cell proliferation and inducting cell cycle arrest, the fact that glioma cell lines show sensitivity to ZBC260. Notably, ZBC260 targeted glioma without side effects in vivo. In addition, the stem cell-like properties of glioma cells were inhibited upon ZBC260 treatment. When the mechanism was examined, our findings indicated that Wnt/β-catenin pathway repression is required for ZBC260-induced stem cell-like properties and tumor growth suppression. In conclusion, the growth of tumors and stem cell-like properties were inhibited by ZBC260 via Wnt/β-catenin repression, which suggests ZBC260 as a potential therapeutic agent for glioma.

scholarly journals Metabolism-based isolation of invasive glioblastoma cells with specific gene signatures and tumorigenic potential

2020 ◽  
Vol 2 (1) ◽  
Author(s):  
Stuart James Smith ◽  
Jonathan Rowlinson ◽  
Maria Estevez-Cebrero ◽  
David Onion ◽  
Alison Ritchie ◽  
...  
Keyword(s):  
Cell Population ◽  
Aminolevulinic Acid ◽  
Specific Gene ◽  
Normal Brain ◽  
Vitro Assay ◽  
Molecular Abnormalities ◽  
Molecular Features ◽  
Background Population ◽  
Sirna Knockdown

Abstract Background Glioblastoma (GBM) is a highly aggressive brain tumor with rapid subclonal diversification, harboring molecular abnormalities that vary temporospatially, a contributor to therapy resistance. Fluorescence-guided neurosurgical resection utilizes the administration of 5-aminolevulinic acid (5-ALA) generating individually fluorescent tumor cells within a background population of non-neoplastic cells in the invasive tumor region. The aim of the study was to specifically isolate and interrogate the invasive GBM cell population using a novel 5-ALA-based method. Methods We have isolated the critical invasive GBM cell population by developing 5-ALA-based metabolic fluorescence-activated cell sorting. This allows purification and study of invasive cells from GBM without an overwhelming background “normal brain” signal to confound data. The population was studied using RNAseq, real-time PCR, and immunohistochemistry, with gene targets functionally interrogated on proliferation and migration assays using siRNA knockdown and known drug inhibitors. Results RNAseq analysis identifies specific genes such as SERPINE1 which is highly expressed in invasive GBM cells but at low levels in the surrounding normal brain parenchyma. siRNA knockdown and pharmacological inhibition with specific inhibitors of SERPINE1 reduced the capacity of GBM cells to invade in an in vitro assay. Rodent xenografts of 5-ALA-positive cells were established and serially transplanted, confirming tumorigenicity of the fluorescent patient-derived cells but not the 5-ALA-negative cells. Conclusions Identification of unique molecular features in the invasive GBM population offers hope for developing more efficacious targeted therapies compared to targeting the tumor core and for isolating tumor subpopulations based upon intrinsic metabolic properties.

scholarly journals Simple In Vitro Assay for Determining the Sensitivity of Plasmodium vivax Isolates from Fresh Human Blood to Antimalarials in Areas where P. vivax Is Endemic

2003 ◽  
Vol 47 (1) ◽  
pp. 170-173 ◽  
Author(s):  
Bruce M. Russell ◽  
Rachanee Udomsangpetch ◽  
Karl H. Rieckmann ◽  
Barbara M. Kotecka ◽  
Russell E. Coleman ◽  
...  
Keyword(s):  
Plasmodium Vivax ◽  
Poor Response ◽  
In Vitro Assay ◽  
World Health ◽  
Vitro Assay ◽  
Malaria Clinic ◽  
Drug Assays ◽  
Tak Province ◽  
Health Organization

ABSTRACT The aim of this study was to develop a simple, field-practical, and effective in vitro method for determining the sensitivity of fresh erythrocytic Plasmodium vivax isolates to a range of antimalarials. The method used is a modification of the standard World Health Organization (WHO) microtest for determination of P. falciparum drug sensitivity. The WHO method was modified by removing leukocytes and using a growth medium supplemented with AB+ serum. We successfully carried out 34 in vitro drug assays on 39 P. vivax isolates collected from the Mae Sod malaria clinic, Tak Province, Thailand. The mean percentage of parasites maturing to schizonts (six or more merozoites) in control wells was 66.5% ± 5.9% (standard deviation). This level of growth in the control wells enabled rapid microscopic determination (5 min per isolate per drug) of the MICs of chloroquine, dihydroartemisinin, WR238605 (tafenoquine), and sulfadoxine. P. vivax was relatively sensitive to chloroquine (MIC = 160 ng/ml, 50% inhibitory concentration [IC50] = 49.8 ng/ml) and dihydroartemisinin (MIC = 0.5 ng/ml, IC50 = 0.47 ng/ml). The poor response of P. vivax to both tafenoquine (MIC = 14,000 ng/ml, IC50 = 9,739 ng/ml) and sulfadoxine (MIC = 500,000 ng/ml, IC50 = 249,000 ng/ml) was due to the slow action of these drugs and the innate resistance of P. vivax to sulfadoxine. The in vitro assay developed in our study should be useful both for assessing the antimalarial sensitivity of P. vivax populations and for screening new antimalarials in the absence of long-term P. vivax cultures.

scholarly journals 5-Aminolevulinic acid-mediated sonosensitization of rat RG2 glioma cells in vitro

2016 ◽  
Vol 3 ◽  
pp. 234-240 ◽  
Author(s):  
Krzysztof Bilmin ◽  
Tamara Kujawska ◽  
Wojciech Secomski ◽  
Andrzej Nowicki ◽  
Paweł Grieb
Keyword(s):  
Aminolevulinic Acid ◽  
Glioma Cells

scholarly journals Radiosensitization and a Less Aggressive Phenotype of Human Malignant Glioma Cells Expressing Isocitrate Dehydrogenase 1 (IDH1) Mutant Protein: Dissecting the Mechanisms

Cancers ◽  
2019 ◽  
Vol 11 (6) ◽  
pp. 889 ◽  
Author(s):  
Jacqueline Kessler ◽  
Tim Hohmann ◽  
Antje Güttler ◽  
Marina Petrenko ◽  
Christian Ostheimer ◽  
...  
Keyword(s):  
Malignant Glioma ◽  
Isocitrate Dehydrogenase ◽  
Idh1 Mutation ◽  
Cell Stiffness ◽  
Diffuse Glioma ◽  
Isocitrate Dehydrogenase 1 ◽  
Aggressive Phenotype ◽  
Γh2ax Foci ◽  
Mutant Idh1

The presence of an isocitrate dehydrogenase 1 (IDH1) mutation is associated with a less aggressive phenotype, increased sensitivity to radiation, and increased overall survival in patients with diffuse glioma. Based on in vitro experimentations in malignant glioma cell lines, the consequences on cellular processes of IDH1R132H expression were analyzed. The results revealed that IDH1R132H expression enhanced the radiation induced accumulation of residual γH2AX foci and decreased the amount of glutathione (GSH) independent of the oxygen status. In addition, expression of the mutant IDH1 caused a significant increase of cell stiffness and induced an altered organization of the cytoskeleton, which has been shown to reinforce cell stiffness. Furthermore, IDH1R132H expression decreased the expression of vimentin, an important component of the cytoskeleton and regulator of the cell stiffness. The results emphasize the important role of mutant IDH1 in treatment of patients with diffuse gliomas especially in response to radiation. Hence, detection of the genetic status of IDH1 before therapy massively expands the utility of immunohistochemistry to accurately distinguish patients with a less aggressive and radiosensitive IDH1-mutant diffuse glioma suitable for radiotherapy from those with a more aggressive IDH1-wildtype diffuse glioma who might benefit from an individually intensified therapy comprising radiotherapy and alternative medical treatments.

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