mpm cell line nci h2452  (ATCC)

Journal: Translational Oncology

Article Title: The highly selective and oral phosphoinositide 3-kinase delta (PI3K-δ) inhibitor roginolisib induces apoptosis in mesothelioma cells and increases immune effector cell composition

doi: 10.1016/j.tranon.2023.101857

Figure Lengend Snippet: Association between PI3K-δ expression and clinicopathological characteristics in 89 MPM patients.

Article Snippet: MPM patient-derived xenograft (PDX) cell lines PXF698 and PXF1118 (both from epithelioid tumors) and PXF1752 (from sarcomatoid MPM) were from Charles River Germany GmbH (Freiburg, Germany) and cultured in RPMI medium supplemented with 10% Fetal Bovine Serum (FBS) (Biochrom AG, Berlin, Germany).

Techniques: Expressing

Journal: Translational Oncology

Article Title: The highly selective and oral phosphoinositide 3-kinase delta (PI3K-δ) inhibitor roginolisib induces apoptosis in mesothelioma cells and increases immune effector cell composition

doi: 10.1016/j.tranon.2023.101857

Figure Lengend Snippet: Roginolisib impairs mesothelioma cell viability and induces apoptotic cell death. (A) MPM cell lines PXF698, PXF1118 and PXF1752 were exposed to the indicated concentrations of roginolisib. The viability of the cells was measured after 72 h of treatment by a MTT cytotoxicity assay. Data points, mean ± SD of independent triplicate experiments. Immunohistochemistry analysis of PI3K-δ protein expression in PXF698, PXF1118 and PXF1752 cells. Scale, 50 µm. (B) PXF698 cells were exposed to roginolisib (100 µM), paclitaxel (400 nM) or DMSO in the presence of the RealTime-Glo Annexin V Apoptosis and Necrosis Assay. Luminescence (phosphatidylsprrerine:annexin V binding – apoptosis; solid line) and fluorescence (loss of membrane integrity – necrosis; dashed line) were recorded for 40 h at the indicated time points. A clear temporal lag between phosphatidylserine exposure and lack of membrane integrity is indicative of apoptotic cell death leading to secondary necrosis. Data points, mean of duplicate samples, representative of independent duplicate experiments. (C) Caspase-3/7 activity after treatment of cells with 100 µM roginolisib or 0.1% DMSO for 24 h. Data points, mean ± SD of independent triplicate experiments. 2way ANOVA test (Dunnett‘s multiple comparison); ****, p<0.0001; ***, p=0.0002. (D) Apoptotic membrane blebbing after treatment of PXF698 cells with 100 µM roginolisib for 24 h, detected by phase-contrast microscopy. (E) PARP cleavage and levels of Mcl-1 and BIM in PXF698 and PXF1752 cells after 100 µM roginolisib treatment at 0, 4, 24 h. (F) Immunoblotting shows the levels of p-AKT (S473) and p-ERK1/2 (T202/204) in PXF1118 and PXF1752 cells after 100 µM roginolisib treatment at 0, 4, 24 h. Bar graphs represent semi-quantitative analysis of phospho-protein expression relative to total protein, normalized to protein levels at 0 h. (G) Schematic illustration of PI3K/AKT/mTOR and ERK signaling.

Article Snippet: MPM patient-derived xenograft (PDX) cell lines PXF698 and PXF1118 (both from epithelioid tumors) and PXF1752 (from sarcomatoid MPM) were from Charles River Germany GmbH (Freiburg, Germany) and cultured in RPMI medium supplemented with 10% Fetal Bovine Serum (FBS) (Biochrom AG, Berlin, Germany).

Techniques: Cytotoxicity Assay, Immunohistochemistry, Expressing, Binding Assay, Fluorescence, Membrane, Activity Assay, Comparison, Microscopy, Western Blot

Journal: Translational Oncology

Article Title: The highly selective and oral phosphoinositide 3-kinase delta (PI3K-δ) inhibitor roginolisib induces apoptosis in mesothelioma cells and increases immune effector cell composition

doi: 10.1016/j.tranon.2023.101857

Figure Lengend Snippet: Synergistic cytotoxicity of PI3K-δ inhibitors and sapanisertib (mTORC1/2 inhibitor) or ipatasertib (AKT inhibitor). (A) MPM cells were treated with PI3K-δ inhibitors roginolisib (Rog) or idelalisib (Idel) as single agents and in combination with sapanisertib (Sapa) (PXF698) or ipatasertib (Ipa) (PXF1118) for 72 h, after which cell viability was assessed using an MTT cytotoxicity assay. Data points, mean of quintuplets of two independent experiments. (B) Schematic illustration of PI3K/AKT/mTOR and ERK signaling. Growth factors and hormones activate PI3 kinase catalyzing the production of PIP3, which in turn coordinates cell proliferation, metabolism and survival via activating AKT and mTOR signaling pathways. mTOR kinase serves as core component of two distinct protein complexes, mTORC1 and mTORC2. Activated Ras triggers both PI3K/AKT and ERK signaling, regulating cell proliferation and survival. (C) Molecular effects of simultaneous PI3K-δ/mTOR and PI3K-δ/AKT inhibition. Immunoblotting shows the levels of p-AKT (S473), p-PRAS40 p-PRAS40 (T246), p-RPS6 (S235/236) and p-ERK1/2 (T202/204) in PXF1118 cells treated with 100 µM roginolisib (Rog), 100 µM idelalisib (Idel), 30 nM sapanisertib (Sapa), 10 µM ipatasertib (Ipa), or their combinations as indicated for 4 h.

Article Snippet: MPM patient-derived xenograft (PDX) cell lines PXF698 and PXF1118 (both from epithelioid tumors) and PXF1752 (from sarcomatoid MPM) were from Charles River Germany GmbH (Freiburg, Germany) and cultured in RPMI medium supplemented with 10% Fetal Bovine Serum (FBS) (Biochrom AG, Berlin, Germany).

Techniques: Cytotoxicity Assay, Protein-Protein interactions, Inhibition, Western Blot

Journal: Translational Oncology

Article Title: The highly selective and oral phosphoinositide 3-kinase delta (PI3K-δ) inhibitor roginolisib induces apoptosis in mesothelioma cells and increases immune effector cell composition

doi: 10.1016/j.tranon.2023.101857

Figure Lengend Snippet: Roginolisib enhances the cytotoxicity of cisplatin and nivolumab in patient-derived tumor cells in the presence of stromal cells. (A) BAP1-wild-type epithelioid (BAP + epi) and BAP1-null sarcomatoid (BAP − sar) MPM cells were co-cultured with peripheral mononuclear cells (PBMCs) and exposed to the indicated concentrations of roginolisib in the absence (-MRC5) or presence of fibroblasts (+MRC5). The viability of the cells was measured after 72 hours by WST-1 staining. Data points, mean of independent quadruplicate experiments. (B) MPM cells were treated with 5 µM cisplatin (Pt) and 1 µg/ml nivolumab (Nivo) for 72 h in the absence and presence of MRC5 cells. (C, D) The combination of roginolisib with 5 µM cisplatin plus 1 µg/ml nivolumab, either as combined/concomitant treatment (C) or in sequential treatment (D), was investigated in the absence (-MRC5) and presence of fibroblasts (+MRC5). 0: untreated cells. Nonparametric Kruskal–Wallis test followed by Dunn's multiple comparison test; ***p<0.001: vs. untreated cells; °p<0.05, °°°p<0.001: MRC5+ vs MRC5-.

Article Snippet: MPM patient-derived xenograft (PDX) cell lines PXF698 and PXF1118 (both from epithelioid tumors) and PXF1752 (from sarcomatoid MPM) were from Charles River Germany GmbH (Freiburg, Germany) and cultured in RPMI medium supplemented with 10% Fetal Bovine Serum (FBS) (Biochrom AG, Berlin, Germany).

Techniques: Derivative Assay, Cell Culture, Staining, Comparison

Journal: Translational Oncology

Article Title: The highly selective and oral phosphoinositide 3-kinase delta (PI3K-δ) inhibitor roginolisib induces apoptosis in mesothelioma cells and increases immune effector cell composition

doi: 10.1016/j.tranon.2023.101857

Figure Lengend Snippet: Roginolisib exerts immunomodulatory activity. BAP1-wild-type epithelioid (A) and BAP1-null sarcomatoid (B) MPM cells were co-cultured with peripheral mononuclear cells (PBMCs) in the absence or presence of MRC5 fibroblasts and treated with roginolisib alone, co-incubated with cisplatin/nivolumab or in sequential treatment (cisplatin/nivolumab followed by roginolisib). The immunophenotype of immune cells was analyzed by flow cytometry (n = 2, two independent experiments). *p<0.05, **p<0.01, ***p<0.001.

Article Snippet: MPM patient-derived xenograft (PDX) cell lines PXF698 and PXF1118 (both from epithelioid tumors) and PXF1752 (from sarcomatoid MPM) were from Charles River Germany GmbH (Freiburg, Germany) and cultured in RPMI medium supplemented with 10% Fetal Bovine Serum (FBS) (Biochrom AG, Berlin, Germany).

Techniques: Activity Assay, Cell Culture, Incubation, Flow Cytometry

Journal: Translational Cancer Research

Article Title: Induction of metallothionein expression by supplementation of zinc induces resistance against platinum-based treatment in malignant pleural mesothelioma

doi: 10.21037/tcr-22-2651

Figure Lengend Snippet: Gene expression of MT2A after supplementation of different zinc concentrations. The data were measured 77 h after supplementation of zinc. The difference in MT2A expression of the sample between zinc-treatment and control is shown on the y-axis (2 −ΔCp ) (Cp means the crossing point describing the cycle at which the fluorescence first rises significantly above the background fluorescence). All MPM cell lines showed elevated expression of MT2A by adding higher concentrations of zinc [(A) MSTO211H (P=0.0833), (B) NCI-H2452 (P=1), and (C) NCI-H2052 (P=0.0833)]. P values were calculated by using the Spearman’s rank correlation test. MT2A , metallothionein IIA; MPM, malignant pleural mesothelioma.

Article Snippet: All MPM cell lines [NCI-H2052: CRL-5915; NCI-H2452: CRL-5946; MSTO-211H: CRL-2081; American Type Culture Collection (ATCC), Virginia, USA] were cultured in Roswell Park Memorial Institute 1640 medium (RPMI) (Thermo Fisher Scientific, Massachusetts, USA), supplemented with 10% fetal calf serum (FCS) and 1% penicillin and streptomycin (P/S, Thermo Fisher Scientific, Massachusetts, USA).

Techniques: Gene Expression, Expressing, Control, Fluorescence