Effect of P‐Glycoprotein Expression Levels on the Concentration‐Dependent Permeability of Drugs to the Cell Membrane

2008 ◽  
Vol 97 (1) ◽  
pp. 553-565 ◽  
Author(s):  
Yoshiyuki Shirasaka ◽  
Toshiyasu Sakane ◽  
Shinji Yamashita
Keyword(s):  
Cell Membrane ◽  
Expression Levels ◽  
P Glycoprotein

P-Glycoprotein Contributes to Cell Membrane Depolarization of Hippocampus and Neocortex in a Model of Repetitive Seizures Induced by Pentylenetetrazole in Rats

2013 ◽  
Vol 19 (38) ◽  
pp. 6732-6738 ◽  
Author(s):  
Jerónimo A. Auzmendi ◽  
Sandra Orozco-Suárez ◽  
Ivette Bañuelos-Cabrera ◽  
María Eva González-Trujano ◽  
Eduardo Calixto González ◽  
...  
Keyword(s):  
Cell Membrane ◽  
Membrane Depolarization ◽  
P Glycoprotein

Correlation of expression levels of P-glycoprotein with resistance to adriamycin in a renal adenocarcinoma cell line

1997 ◽  
Vol 25 (6) ◽  
pp. 407-412
Author(s):  
S. Kawamoto ◽  
T. Deguchi ◽  
S. Nezasa ◽  
S. Yamada ◽  
M. Okano ◽  
...  
Keyword(s):  
Cell Line ◽  
Adenocarcinoma Cell Line ◽  
Adenocarcinoma Cell ◽  
Expression Levels ◽  
Renal Adenocarcinoma ◽  
P Glycoprotein

scholarly journals Altered Intestinal P-glycoprotein Expression Levels Affect Pharmacodynamics under Diabetic Condition

2012 ◽  
Vol 132 (2) ◽  
pp. 161-166
Author(s):  
Ayaka Nawa ◽  
Wakako Fujita-Hamabe ◽  
Shiroh Kisioka ◽  
Shogo Tokuyama
Keyword(s):  
Expression Levels ◽  
P Glycoprotein

scholarly journals The Establishment of Quantitatively Regulating Expression Cassette with sgRNA Targeting BIRC5 to Elucidate the Synergistic Pathway of Survivin with P-Glycoprotein in Cancer Multi-Drug Resistance

2022 ◽  
Vol 9 ◽  
Author(s):  
Changping Deng ◽  
Fabiao Hu ◽  
Zhangting Zhao ◽  
Yiwen Zhou ◽  
Yuping Liu ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Survivin Expression ◽  
Regulatory System ◽  
Multi Drug Resistance ◽  
Specific Gene ◽  
Expression Levels ◽  
P Glycoprotein ◽  
Biological Phenomena ◽  
Reversal Index ◽  
Mcf 7

Quantitative analysis and regulating gene expression in cancer cells is an innovative method to study key genes in tumors, which conduces to analyze the biological function of the specific gene. In this study, we found the expression levels of Survivin protein (BIRC5) and P-glycoprotein (MDR1) in MCF-7/doxorubicin (DOX) cells (drug-resistant cells) were significantly higher than MCF-7 cells (wild-type cells). In order to explore the specific functions of BIRC5 gene in multi-drug resistance (MDR), a CRISPR/Cas9-mediated knocking-in tetracycline (Tet)-off regulatory system cell line was established, which enabled us to regulate the expression levels of Survivin quantitatively (clone 8 named MCF-7/Survivin was selected for further studies). Subsequently, the determination results of doxycycline-induced DOX efflux in MCF-7/Survivin cells implied that Survivin expression level was opposite to DOX accumulation in the cells. For example, when Survivin expression was down-regulated, DOX accumulation inside the MCF-7/Survivin cells was up-regulated, inducing strong apoptosis of cells (reversal index 118.07) by weakening the release of intracellular drug from MCF-7/Survivin cells. Also, down-regulation of Survivin resulted in reduced phosphorylation of PI3K, Akt, and mTOR in MCF-7/Survivin cells and significantly decreased P-gp expression. Previous studies had shown that PI3K/Akt/mTOR could regulate P-gp expression. Therefore, we speculated that Survivin might affect the expression of P-gp through PI3K/Akt/mTOR pathway. In summary, this quantitative method is not only valuable for studying the gene itself, but also can better analyze the biological phenomena related to it.

scholarly journals R-2HG/KDM2B/RIPK1 Signaling Mediates Necroptosis in Myelodysplastic Syndromes

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 5049-5049
Author(s):  
Shuanghong Zhu ◽  
Chen Mei ◽  
Hongyan Tong ◽  
Jie Jin
Keyword(s):  
Bone Marrow ◽  
Cell Death ◽  
Cell Membrane ◽  
Membrane Permeability ◽  
Cell Lines ◽  
Myelodysplastic Syndromes ◽  
Poor Prognosis ◽  
Cell Membrane Permeability ◽  
Expression Levels ◽  
Low Expression

Introduction: Myelodysplastic syndromes (MDS) are a group of heterogeneous hematopoietic stem cell disorders and manifested as ineffective hematopoiesis, refractory cytopenia and a propensity to evolve into acute myeloid leukemia (AML). Isocitrate dehydrogenase 1/2 (IDH1/2) mutations are common in both MDS and AML. Mutated IDH produces R-2-hydroxyglutarate (R-2HG) which inhibits multiple α-ketoglutarate/α-KG-dependent dioxygenases by competing against α-KG binding. Recent studies have demonstrated that R-2-HG can abrogates leukemic growth and induce leukemia cell death. Several nonapoptotic cell death have been identified, including phosphoribosyl pyrophosphate (PPRP)-1 mediated necrosis, pyroptosis, ferroptosis and necroptosis. By now, the specific type of cell death by which R-2-HG exerts anti-tumor effects is still unknown. Results: (1) (2R)-Octyl-2-HG exhibited anti-tumor effect on SKM-1, THP-1, Molm-13, HL-60 cell lines and bone marrow mononuclear cells from MDS and AML patients in a dose- and time-dependent manner. The overexpression of IDH2 mutation induced by doxycycline significantly decreased the viability of AML cell lines, and the inhibition rate was related to the dose of doxycycline. (2R)-Octyl-2-HG promotes apoptosis and causes G0/G1 phase arrest. (2) R-2-HG leads to increased expression of RIPK1 in high-risk MDS cells. The results of gene enrichment analysis indicated that the apoptotic pathway was enriched in (2R)-Octyl-2-HG groups, and the expression of RIPK1 gene was increased in all three (SKM-1, NOMO-1 and MA9.3ITD) (2R)-Octyl-2-HG groups. After treatment with (2R)-Octyl-2-HG, the mRNA and protein expression levels of RIPK1 gene were increased. (3) R-2-HG triggers RIPK1-dependent necroptosis and occurs earlier than apoptosis. Within 10 hours after treatment with (2R)-Octyl-2-HG, cell membrane permeability was disrupted, interactions between RIPK1 and caspase 8 increased, as well as phosphorylated MLKL level. However, caspase activity did not increase significantly, suggesting that necroptosis occurs earlier than apoptosis. With RIPK1 inhibitor Necrostatin-1 and (2R)-Octyl-2-HG co-treating cells, proliferation inhibition was reduced, cell membrane permeability was more stable and RIPK1-caspase8 complex was difficult to form. The same phenomenon occurs in SKM-1 cells stably transfected with RIPK1 shRNA virus, suggesting that RIPK1-dependent necroptosis is involved in cell death caused by (2R)-Octyl-2-HG. (4) In vivo experiments demonstrated that necroptosis by R-2-HG is dependent on the expression of RIPK. In RIPK1 shRNA MDS mice, the tumor burden showed a decreasing trend, but did not show a significant change in the R-2-HG treatment group. Treatment of scramble shRNA MDS mice with R-2-HG resulted in significantly smaller tumors in the spleen and less engrafment of CD45+ cells in bone marrow. (5) Low RIPK1 expression predicts poor prognosis in MDS patients. Data from the TCGA and GEO public databases indicate that RIPK1 expression is reduced in MDS and AML patients compared to healthy controls. Survival analysis showed that patients with lower RIPK1 expression levels had significantly shorter overall survival (OS) than patients with higher RIPK1 expression levels. MDS patients with lower RIPK1 expression levels progress to leukemia more frequently. (6) Inhibition of KDM2B induces necroptosis independently. Western blot assay shows the knockdown of KDM2B, upregulation of RIPK1 and increased levels of p-MLKL. Analysis of cell numbers showed that proliferation ceased from 4 days after doxycycline treatment onwards in shKDM2B cells. Co-IP assay shows the formation of RIPK1-caspase8 complex. Conclusion: This study confirmed that R-2-HG inhibited the viability of MDS and AML cells. R-2-HG increased the expression of RIPK1 in MDS cells, inducing necroptosis. Necroptosis occurred earlier than apoptosis. Inhibition of RIPK1 can alleviate the inhibitory effect of R-2-HG on MDS and AML cells. Clinical studies have shown that low expression of the RIPK1 gene is associated with poor prognosis in patients with MDS and AML. MDS patients with low expression of RIPK1 was more likely to progress to leukemia. Inhibition of KDM2B can induce necroptosis independently. Disclosures No relevant conflicts of interest to declare.

Bosutinib is a Substrate of the ABC Efflux Transporter P-Glycoprotein.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3763-3763
Author(s):  
Pietro Perini ◽  
Sara Redaelli ◽  
Rocco Piazza ◽  
Michela Viltadi ◽  
Frank Boschelli ◽  
...  
Keyword(s):  
Western Blot ◽  
Cell Lines ◽  
Kinase Inhibitor ◽  
Biological Effects ◽  
Intracellular Concentration ◽  
Apoptosis Induction ◽  
Altered Expression ◽  
Expression Levels ◽  
Abl Tyrosine Kinase ◽  
P Glycoprotein

Abstract Abstract 3763 Poster Board III-699 Bosutinib (Bos) (Wyeth Pharmaceuticals) is a dual Src-Abl tyrosine kinase inhibitor developed to inhibit the activity of BCR-ABL in chronic myeloid leukemia. P-glycoprotein (P-gp) is an ATP-binding cassette (ABC) transporter responsible of Imatinib (Im) efflux, and its increased expression can be related to Im resistance. The aim of the study was to define if P-gp is also responsible for the cellular efflux of Bos, and if P-gp altered expression can be related to Bos resistance. Four different K562 Ph+ cell lines were used. 1) K562S, Im sensitive; 2) K562DOX, cells resistant to Doxorubicin that overexpress P-gp (kind gift of JP Marie, Université Pierre et Marie Curie, Paris); 3) K562DOX siP-gp, K562DOX cells carrying a stable silencing of P-gp (kind gift of E Gunsilius, Innsbruck Medical University, Austria); 4) K562DOX H1, K562DOX cells carrying a control vector (kind gift of E Gunsilius). Real Time PCR confirmed that K562DOX and K562DOX H1 express higher levels of P-gp than K562DOX siP-gp (approximately 10 fold) and K562S (approximately 10000 fold). Western blot α-P-gp showed similar results. We then assessed Bos IC50 on the cell lines with a proliferation assay. K562DOX (IC50=175.3nM) and K562DOX H1 (IC50=102 nM) are resistant to Bos if compared to K562DOX siP-gp (IC50=9.1nM) and K562S (IC50=8.7nM). This data confirm that the P-gp overexpression is related to Bos resistance. Using C-14 radiolabeled Bosutinib (C-14 Bos) (Wyeth Pharmaceuticals) we set up an Intracellular Uptake and Retention (IUR) assay on K562S, K562DOX and K562DOX siP-gp cells. Cells were pre-treated for an hour with 0-60μM of Verapamil (Ver), a P-gp inhibitor, and then treated for two hours with 1μM C-14 Bos. In presence of Ver, K562DOX showed a significant increased amount of intracellular C-14 Bos ([] C-14 Bos Ver60μM/ [] C-14 Bos Ver0μM=5.8) that was not observed in K562S or K562DOX siP-gp. These data confirm that the intracellular concentration of Bos is strictly related to the expression levels and to the activity of P-gp. To test the biological effect of P-gp inhibition, we evaluated Bos IC50 by proliferation and apoptosis induction in K562DOX and K562S cells pre-treated with increasing concentrations of Ver. K562S did not show a significant decrease of Bos IC50 even at the highest concentration of Ver (22.5μM). In K562DOX the same concentration of Ver led to a 20 fold decrease of Bos IC50 (9nM), down to a level comparable to Bos IC50 in K562S cells (8.7nM). Thus, the inhibition of P-gp seems to restore the efficacy of Bos on proliferation inhibition and apoptosis induction in BCR-ABL+ cells. Finally, we evaluated the effect of P-gp inhibition on the phosphorylation levels of BCR-ABL with an α-phosphoTyrosine western blot. K562DOX and K562S cells were treated with Bos concentrations ranging from 0 to 80nM, and in absence or presence of Ver (5μM). Phosphorylation levels of BCR-ABL in K562S treated with Ver didn't show any difference when compared to untreated samples. In K562DOX, in absence of Ver, BCR-ABL was phosphorylated even in presence of high Bos concentration; the treatment with Ver restored the sensitivity to Bos, thus leading to a phosphorylation pattern similar to the one of K562S cells. This is compatible with increasing intracellular concentration of Bos after P-gp inhibition which results in increased molecular effect of Bos on BCR-ABL tyrosine phosphorylation. All together, these data confirm the relevance of expression levels and activity of P-gp for the intracellular concentration of Bos. The intracellular concentration of Bos is strictly related to its molecular activity on BCR-ABL and to its biological effects on Ph+ cells. Disclosures: Boschelli: Wyeth Pharmaceuticals: Employment.

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