scholarly journals Integrated Proteomic and Phosphoproteomics Analysis of DKK3 Signaling Reveals Activated Kinase in the Most Aggressive Gallbladder Cancer

Cells ◽  
2021 ◽  
Vol 10 (3) ◽  
pp. 511
Author(s):  
Kirti Gondkar ◽  
Gajanan Sathe ◽  
Neha Joshi ◽  
Bipin Nair ◽  
Akhilesh Pandey ◽  
...  
Keyword(s):  
Tumor Suppressor ◽  
Gallbladder Cancer ◽  
Cell Lines ◽  
High Resolution Mass Spectrometry ◽  
Cell Cycle Control ◽  
Complex Nature ◽  
Chromosomal Replication ◽  
Metal Affinity ◽  
Tandem Mass Tag ◽  
First Time

DKK3 is a secreted protein, which belongs to a family of Wnt antagonists and acts as a potential tumor suppressor in gallbladder cancer. To further understand its tumor suppressor functions, we overexpressed DKK3 in 3 GBC cell lines. We have employed high-resolution mass spectrometry and tandem mass tag (TMT) multiplexing technology along with immobilized metal affinity chromatography to enrich phosphopeptides to check the downstream regulators. In this study, we reported for the first time the alteration in the phosphorylation of 14 kinases upon DKK3 overexpression. In addition, we observed DKK3 induced hyper phosphorylation of 2 phosphatases: PPP1R12A and PTPRA, which have not been reported previously. Canonical pathway analysis of altered molecules indicated differential enrichment of signaling cascades upon DKK3 overexpression in all the 3 cell lines. Protein kinase A signaling, Sirtuin signaling pathway, and Cell Cycle Control of Chromosomal Replication were observed to be differentially activated in the GBC cell lines. Our study revealed, DKK3 overexpression has differential effect based on the aggressive behavior of the cell lines. This study expands the understanding of DKK3-mediated signaling events and can be used as a primary factor for understanding the complex nature of this molecule.

Polyethylene Glycol - Polyethylenimine Nanoparticle-Mediated Transfection of Mir-150 Into the Leukemia Cells and Determination of the Expression Pattern Changes in Apoptosis and Cell Cycle Genes

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 4679-4679
Author(s):  
Tugce Balci ◽  
Cigir Biray Avci ◽  
Ipek Ozcan ◽  
Sunde Yilmaz Susluer ◽  
Cagla Kayabasi ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Polyethylene Glycol ◽  
Tumor Suppressor ◽  
Expression Pattern ◽  
Cell Lines ◽  
Cell Model ◽  
Cell Cycle Control ◽  
Leukemia Cell ◽  
Gene Expression Pattern ◽  
Leukemia Cells

Abstract Abstract 4679 The aims of this study are transfection of tumor suppressor miR-150 that is down-regulated in leukemogenesis into leukemia cells mediated by Polyethylene Glycol - Polyethylenimine (PEG-PEI) nanoparticle and determine the changes in gene expression pattern in chronic myeloid leukemia model cell lines; K-562 and KU812 and non-leukemia cell lines NCI-BL2347 and NCI-BL2171. Characterization studies and stability studies of PEG-PEI copolymers were performed and by using these copolymers 9 nanoparticle formulations were prepared. Three copolymers (named T1, T3 and T9) were eligible for further analysis and nanoparticle complex (named F1, F2 and F3) formulations, particles that their size less than 300 nM have been evaluated. Nanoparticle-mediated substitution of miR-150 reduced the expressions of cell cycle control genes CDK2 and CCNG2, CDK6 7-fold and 2-fold, respectively in K562 leukemia cell model. Nanoparticle-mediated substitution of miR-150 has been evaluated KU812 leukemia cell model and reduced the expression of cell cycle control genes CHEK2, CDK5RAP1 and CCNG2 and CDKN1A 6- fold 5 –fold and 2-fold, respectively. Substitution of deregulated tumor suppressor miR-150 to leukemia cells with non-viral transfection yielded promising results for the treatment of leukemia. Taking into account these results, it should be supported by preclinical studies in animal models, which would add benefit to current treatment protocols in clinical application. Disclosures: No relevant conflicts of interest to declare.

Phytochemical Constituents of Annona reticulata and their Cytotoxic Activity

2020 ◽  
Vol 17 (3) ◽  
pp. 206-210
Author(s):  
Ty Viet Pham ◽  
Thang Quoc Le ◽  
Anh Tuan Le ◽  
Hung Quoc Vo ◽  
Duc Viet Ho
Keyword(s):  
Cytotoxic Activity ◽  
Cell Lines ◽  
Structural Determination ◽  
Phytochemical Investigation ◽  
Ic50 Values ◽  
Phytochemical Constituents ◽  
First Time ◽  
Annona Reticulata

A phytochemical investigation of the leaves of Annona reticulata led to the isolation and structural determination of β-sitosterol (1), ent-pimara-8(14),15-dien-19-oic acid (2), ent-pimara- 8(14),15-dien-19-ol (3), quercetin (4), quercetin 3-O-α-L-arabinopyranoside (5), and a mixture of quercetin 3-O-β-D-galactopyranoside (6a) and quercetin 3-O-β-D-glucopyranoside (6b). Of these, compounds 2 and 3 were isolated from the genus Annona for the first time. Compound 3 showed strong cytotoxicity against SK-LU-1 and SW626 cell lines with IC50 values of 17.64 ± 1.07 and 19.79 ± 1.41 μg mL-1, respectively.

Apoptosis Caused by Triterpenes and Phytosterols and Antioxidant Activity of an Enriched Flavonoid Extract from Passiflora mucronata

2019 ◽  
Vol 18 (10) ◽  
pp. 1405-1416 ◽  
Author(s):  
Isabel C.V. da Silva ◽  
Goran N. Kaluđerović ◽  
Pollyana F. de Oliveira ◽  
Denise O. Guimarães ◽  
Carla H. Quaresma ◽  
...  
Keyword(s):  
Antioxidant Activity ◽  
Cell Lines ◽  
Oleanolic Acid ◽  
Folk Medicine ◽  
Hexane Fraction ◽  
Cytotoxicity Test ◽  
B16f10 Cell ◽  
Dpph Method ◽  
Melanoma B16f10 ◽  
First Time

Background: P. mucronata (Pm) comes from South America, Brazil and is characterized as “Maracujá de Restinga”. It is used in folk medicine for its soothing properties and in treating insomnia. Objective: The present study for the first time analyzed the antioxidant and cytotoxicity of the hydroalcoholic leaves extract and fractions from Pm. Method: The cytotoxicity test will be evaluated by different assays (MTT and CV) against human prostate cancer (PC3) and mouse malignant melanoma (B16F10) cell lines, and the antioxidant test by DPPH method. Results: β-Amyrin, oleanolic acid, β-sitosterol and stigmasterol were isolated of the most active, hexane fraction. These substances were tested against the tumor cell lines: β-sitosterol and stigmasterol showed the most relevant activity to PC3 in CV assay and, oleanolic acid to B16F10 by the MTT assay. In addition, it was possible to indicate that the mode of cell death for stigmasterol, presumably is apoptosis. In terms of antioxidant activity, the hydroalcoholic leaves extract presented higher activity (EC50 133.3 µg/mL) compared to the flower (EC50 152.3 µg/mL) and fruit (EC50 207.9 µg/mL) extracts. By the HPLC-MS, it was possible to identify the presence of flavones in the leaf extract (isoschaftoside, schaftoside, isovitexin, vitexin, isoorientin, orientin). Conclusions: P. mucronata hexane fraction showed promising cytotoxic effect against cancer cell lines, and stigmasterol contributes to this activity, inducing apoptosis of these cells. Furthermore, as other Passiflora species, Pm extract showed antioxidant activity and flavones are its major phenolic compounds.

scholarly journals Analysis of Pembrolizumab in Human Plasma by LC-MS/HRMS. Method Validation and Comparison with Elisa

Biomedicines ◽  
2021 ◽  
Vol 9 (6) ◽  
pp. 621
Author(s):  
Aurélien Millet ◽  
Nihel Khoudour ◽  
Jérôme Guitton ◽  
Dorothée Lebert ◽  
François Goldwasser ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Human Plasma ◽  
High Resolution Mass Spectrometry ◽  
Internal Standard ◽  
Therapeutic Drug ◽  
Limit Of Quantification ◽  
Immunoglobulin G4 ◽  
Positive Mode ◽  
Surrogate Peptide ◽  
First Time

Pembrolizumab is a humanized immunoglobulin G4-kappa anti-PD1 antibody used in the treatment of different solid tumors or haematological malignancies. A liquid chromatography coupled with a high resolution mass spectrometry (orbitrap technology) method was fully developed, optimized, and validated for quantitative analysis of pembrolizumab in human plasma. A mass spectrometry assay was used for the first time a full-length stable isotope-labelled pembrolizumab-like (Arginine 13C6-15N4 and Lysine 13C6-15N2) as an internal standard; the sample preparation was based on albumin depletion and trypsin digestion and, finally, one surrogate peptide was quantified in positive mode. The assay showed good linearity over the range of 1–100 μg/mL, a limit of quantification at 1 μg/mL, excellent accuracy from 4.4% to 5.1%, and also a between-day precision below 20% at the limit of quantification. In parallel, an in-house ELISA was developed with a linearity range from 2.5 to 50 µg/mL. Then, results were obtained from 70 plasma samples of cancer patients that were treated with pembrolizumab and quantified with both methods were compared using the Passing-Bablok regression analysis and Bland-Altman plotting. The LC-MS/HRMS method is easy to implement in the laboratory for use in the context of PK/PD studies, clinical trials, or therapeutic drug monitoring.

scholarly journals An E3 Ubiquitin Ligase RNF139 Serves as a Tumor-Suppressor in Glioma

2021 ◽  
Author(s):  
Xiaofeng Chen ◽  
Weiping Kuang ◽  
Yong Zhu ◽  
Bin Zhou ◽  
Xiaosong Li ◽  
...  
Keyword(s):  
Tumor Suppressor ◽  
Cell Lines ◽  
Ubiquitin Ligase ◽  
Glioma Cell ◽  
Protein Modification ◽  
E3 Ubiquitin Ligase ◽  
Ring Finger ◽  
Glioma Cells ◽  
Migration And Invasion ◽  
Tissue Samples

AbstractGlioma is highly lethal because of its high malignancy. Ubiquitination, a type of ubiquitin-dependent protein modification, has been reported to play an oncogenic or tumor-suppressive role in glioma development, depending on the targets. Ring finger protein 139 (RNF139) is a membrane-bound E3 ubiquitin ligase serving as a tumor suppressor by ubiquitylation-dependently suppressing cell growth. Herein, we firstly confirmed the abnormal downregulation of RNF139 in glioma tissues and cell lines. In glioma cells, ectopic RNF139 overexpression could inhibit, whereas RNF139 knockdown could aggravate the aggressive behaviors of glioma cells, including hyperproliferation, migration, and invasion. Moreover, in two glioma cell lines, RNF139 overexpression inhibited, whereas RNF139 knockdown enhanced the phosphorylation of phosphatidylinositol 3-kinase (PI3K) and AKT serine/threonine kinase 1 (AKT). In a word, we demonstrate the aberration in RNF139 expression in glioma tissue samples and cell lines. RNF139 serves as a tumor-suppressor in glioma by inhibiting glioma cell proliferation, migration, and invasion and promoting glioma cell apoptosis through regulating PI3K/AKT signaling.

scholarly journals Expression of BARD1 β Isoform in Selected Pediatric Tumors

Genes ◽  
2021 ◽  
Vol 12 (2) ◽  
pp. 168
Author(s):  
Anna Jasiak ◽  
Natalia Krawczyńska ◽  
Mariola Iliszko ◽  
Katarzyna Czarnota ◽  
Kamil Buczkowski ◽  
...  
Keyword(s):  
Pediatric Cancer ◽  
Genome Stability ◽  
Cell Cycle Control ◽  
Reverse Transcriptase Gene ◽  
Neoplastic Tissue ◽  
Adult Type ◽  
Qpcr Method ◽  
New Treatments ◽  
First Time ◽  
Work Supports

Currently, many new possible biomarkers and mechanisms are being searched and tested to analyse pathobiology of pediatric tumours for the development of new treatments. One such candidate molecular factor is BARD1 (BRCA1 Associated RING Domain 1)—a tumour-suppressing gene involved in cell cycle control and genome stability, engaged in several types of adult-type tumours. The data on BARD1 significance in childhood cancer is limited. This study determines the expression level of BARD1 and its isoform beta (β) in three different histogenetic groups of pediatric cancer—neuroblastic tumours, and for the first time in chosen germ cell tumours (GCT), and rhabdomyosarcoma (RMS), using the qPCR method. We found higher expression of beta isoform in tumour compared to healthy tissue with no such changes concerning BARD1 full-length. Additionally, differences in expression of BARD1 β between histological types of neuroblastic tumours were observed, with higher levels in ganglioneuroblastoma and ganglioneuroma. Furthermore, a higher expression of BARD1 β characterized yolk sac tumours (GCT type) and RMS when comparing with non-neoplastic tissue. These tumours also showed a high expression of the TERT (Telomerase Reverse Transcriptase) gene. In two RMS cases we found deep decrease of BARD1 β in post-chemotherapy samples. This work supports the oncogenicity of the beta isoform in pediatric tumours, as well as demonstrates the differences in its expression depending on the histological type of neoplasm, and the level of maturation in neuroblastic tumours.

scholarly journals Autophagy augments the self-renewal of lung cancer stem cells by the degradation of ubiquitinated p53

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Jianyu Wang ◽  
Doudou Liu ◽  
Zhiwei Sun ◽  
Ting Ye ◽  
Jingyuan Li ◽  
...  
Keyword(s):  
Lung Cancer ◽  
Stem Cells ◽  
Cancer Stem Cells ◽  
Cell Lines ◽  
Human Cancer ◽  
Suppressor Activity ◽  
Self Renewal ◽  
Tumor Suppressor Activity ◽  
Lung Cancer Stem Cells ◽  
First Time

AbstractIt has been postulated that cancer stem cells (CSCs) are involved in all aspects of human cancer, although the mechanisms governing the regulation of CSC self-renewal in the cancer state remain poorly defined. In the literature, both the pro- and anti-oncogenic activities of autophagy have been demonstrated and are context-dependent. Mounting evidence has shown augmentation of CSC stemness by autophagy, yet mechanistic characterization and understanding are lacking. In the present study, by generating stable human lung CSC cell lines with the wild-type TP53 (A549), as well as cell lines in which TP53 was deleted (H1229), we show, for the first time, that autophagy augments the stemness of lung CSCs by degrading ubiquitinated p53. Furthermore, Zeb1 is required for TP53 regulation of CSC self-renewal. Moreover, TCGA data mining and analysis show that Atg5 and Zeb1 are poor prognostic markers of lung cancer. In summary, this study has elucidated a new CSC-based mechanism underlying the oncogenic activity of autophagy and the tumor suppressor activity of p53 in cancer, i.e., CSCs can exploit the autophagy-p53-Zeb1 axis for self-renewal, oncogenesis, and progression.

scholarly journals Oxonium Ion Guided Analysis of Quantitative Proteomics Data Reveals Site-Specific O-Glycosylation of Anterior Gradient Protein 2 (AGR2)

2021 ◽  
Vol 22 (10) ◽  
pp. 5369
Author(s):  
Martina Pirro ◽  
Yassene Mohammed ◽  
Arnoud H. de Ru ◽  
George M. C. Janssen ◽  
Rayman T. N. Tjokrodirijo ◽  
...  
Keyword(s):  
Cell Lines ◽  
Quantitative Proteomics ◽  
Glycosylation Site ◽  
Total Cell ◽  
Tandem Mass ◽  
Proteomics Data ◽  
Site Specific ◽  
Oxonium Ions ◽  
Tandem Mass Tag ◽  
Colorectal Cancer Cell Lines

Developments in mass spectrometry (MS)-based analyses of glycoproteins have been important to study changes in glycosylation related to disease. Recently, the characteristic pattern of oxonium ions in glycopeptide fragmentation spectra had been used to assign different sets of glycopeptides. In particular, this was helpful to discriminate between O-GalNAc and O-GlcNAc. Here, we thought to investigate how such information can be used to examine quantitative proteomics data. For this purpose, we used tandem mass tag (TMT)-labeled samples from total cell lysates and secreted proteins from three different colorectal cancer cell lines. Following automated glycopeptide assignment (Byonic) and evaluation of the presence and relative intensity of oxonium ions, we observed that, in particular, the ratio of the ions at m/z 144.066 and 138.055, respectively, could be used to discriminate between O-GlcNAcylated and O-GalNAcylated peptides, with concomitant relative quantification between the different cell lines. Among the O-GalNAcylated proteins, we also observed anterior gradient protein 2 (AGR2), a protein which glycosylation site and status was hitherto not well documented. Using a combination of multiple fragmentation methods, we then not only assigned the site of modification, but also showed different glycosylation between intracellular (ER-resident) and secreted AGR2. Overall, our study shows the potential of broad application of the use of the relative intensities of oxonium ions for the confident assignment of glycopeptides, even in complex proteomics datasets.

scholarly journals CSIG-25. MiR-338-3p AS A TUMOR SUPPRESSOR THROUGH Pttg MODULATION IN GH3 CELL LINES

2016 ◽  
Vol 18 (suppl_6) ◽  
pp. vi46-vi46
Author(s):  
Jin Mo Cho ◽  
Yangjong Lee ◽  
Cheol Ryong Ku ◽  
Wonjin Kim ◽  
Sun Ho Kim ◽  
...  
Keyword(s):  
Tumor Suppressor ◽  
Cell Lines ◽  
Gh3 Cell
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