AKT1 E17K ‐mutated meningioma cell lines respond to treatment with the AKT inhibitor AZD5363

2021 ◽  
Author(s):  
Peter John ◽  
Natalie Waldt ◽  
Josephine Liebich ◽  
Christoph Kesseler ◽  
Stefan Schnabel ◽  
...  
Keyword(s):  
Cell Lines ◽  
Akt Inhibitor ◽  
Meningioma Cell

scholarly journals Combined Application of Pan-AKT Inhibitor MK-2206 and BCL-2 Antagonist Venetoclax in B-Cell Precursor Acute Lymphoblastic Leukemia

2021 ◽  
Vol 22 (5) ◽  
pp. 2771
Author(s):  
Anna Richter ◽  
Elisabeth Fischer ◽  
Clemens Holz ◽  
Julia Schulze ◽  
Sandra Lange ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Tumor Cell ◽  
Cell Lines ◽  
Morphological Changes ◽  
Synergistic Effects ◽  
Lymphoblastic Leukemia ◽  
Tumor Cell Proliferation ◽  
Akt Inhibitor ◽  
Akt Phosphorylation ◽  
Combined Application

Aberrant PI3K/AKT signaling is a hallmark of acute B-lymphoblastic leukemia (B-ALL) resulting in increased tumor cell proliferation and apoptosis deficiency. While previous AKT inhibitors struggled with selectivity, MK-2206 promises meticulous pan-AKT targeting with proven anti-tumor activity. We herein, characterize the effect of MK-2206 on B-ALL cell lines and primary samples and investigate potential synergistic effects with BCL-2 inhibitor venetoclax to overcome limitations in apoptosis induction. MK-2206 incubation reduced AKT phosphorylation and influenced downstream signaling activity. Interestingly, after MK-2206 mono application tumor cell proliferation and metabolic activity were diminished significantly independently of basal AKT phosphorylation. Morphological changes but no induction of apoptosis was detected in the observed cell lines. In contrast, primary samples cultivated in a protective microenvironment showed a decrease in vital cells. Combined MK-2206 and venetoclax incubation resulted in partially synergistic anti-proliferative effects independently of application sequence in SEM and RS4;11 cell lines. Venetoclax-mediated apoptosis was not intensified by addition of MK-2206. Functional assessment of BCL-2 inhibition via Bax translocation assay revealed slightly increased pro-apoptotic signaling after combined MK-2206 and venetoclax incubation. In summary, we demonstrate that the pan-AKT inhibitor MK-2206 potently blocks B-ALL cell proliferation and for the first time characterize the synergistic effect of combined MK-2206 and venetoclax treatment in B-ALL.

scholarly journals Assessment of PI3K/mTOR/AKT Pathway Elements to Serve as Biomarkers and Therapeutic Targets in Penile Cancer

Cancers ◽  
2021 ◽  
Vol 13 (10) ◽  
pp. 2323
Author(s):  
Anita Thomas ◽  
Sascha Reetz ◽  
Philipp Stenzel ◽  
Katrin Tagscherer ◽  
Wilfried Roth ◽  
...  
Keyword(s):  
Cell Viability ◽  
Cell Lines ◽  
Penile Cancer ◽  
Multivariate Regression Analysis ◽  
Clinicopathological Characteristics ◽  
Akt Pathway ◽  
Akt Inhibitor ◽  
Free Survival ◽  
Kaplan Meier ◽  
Treatment Naïve

The PI3K/mTOR/AKT pathway might represent an intriguing option for treatment of penile cancer (PeCa). We aimed to assess whether members of this pathway might serve as biomarkers and targets for systemic therapy. Tissue of primary cancer from treatment-naïve PeCa patients was used for tissue microarray analysis. Immunohistochemical staining was performed with antibodies against AKT, pAKT, mTOR, pmTOR, pS6, pPRAS, p4EBP1, S6K1 and pp70S6K. Protein expression was correlated with clinicopathological characteristics as well as overall survival (OS), disease-specific survival (DSS), recurrence-free survival (RFS) and metastasis-free survival (MFS). AKT inhibition was tested in two primarily established, treatment-naïve PeCa cell lines by treatment with capivasertib and analysis of cell viability and chemotaxis. A total of 76 patients surgically treated for invasive PeCa were included. Higher expression of AKT was significantly more prevalent in high-grade tumors and predictive of DSS and OS in the Kaplan–Meier analysis, and an independent predictor of worse OS and DSS in the multivariate regression analysis. Treatment with pan-AKT inhibitor capivasertib in PeCa cell lines induced a significant downregulation of both total AKT and pAKT as well as decreased cell viability and chemotaxis. Selected protein candidates of the mTOR/AKT signaling pathway demonstrate association with histological and survival parameters of PeCa patients, whereas AKT appears to be the most promising one.

scholarly journals EXTH-64. SMALL GTPASES IN MENINGIOMAS: PROLIFERATION, MIGRATION, SURVIVAL, POTENTIAL TREATMENT AND INTERACTIONS

2020 ◽  
Vol 22 (Supplement_2) ◽  
pp. ii101-ii101
Author(s):  
Christoph Kesseler ◽  
Julian Kahr ◽  
Natalie Waldt ◽  
Nele Stroscher ◽  
Josephine Liebig ◽  
...  
Keyword(s):  
Overall Survival ◽  
Cell Lines ◽  
Small Gtpases ◽  
Potential Treatment ◽  
Rock Inhibitor ◽  
Meningioma Cell ◽  
And Migration ◽  
Proliferation And Migration

Abstract PURPOSE To evaluate the role of the small GTPases RhoA, Rac1 and Cdc42 in meningiomas as therapeutic targets and their interactions in meningiomas. EXPERIMENTAL DESIGN We analyzed expression of GTPases in human meningioma samples and meningioma cell lines of various WHO grades. Malignant IOMM-Lee meningioma cells were used to generate shRNA mediated knockdowns of GTPases RhoA, Rac1 or Cdc42 and to study knockdown effects on proliferation and migration, as well as analysis of cell morphology by confocal microscopy. The same tests were used to investigate effects of the two inhibitors Fasudil and EHT-1864 of malignant IOMM-Lee, KT21 and benign Ben-Men cells and the effects of these drugs on IOMM-Lee knockdown cells. The effects of GTPase knockdowns and Fasudil treatment were studied in terms of overall survival by intracranial xenografts of mice. Potential interactions of GTPases regarding NF2, mTOR and FAK-Paxillin were examined. RESULTS Small GTPases were upregulated in meningiomas of higher tumor grades. Reduced proliferation and migration could be achieved by GTPase knockdown in IOMM-Lee cells. Additionally, the ROCK-inhibitor Fasudil and Rac1-inhibitor EHT-1864 reduced proliferation in different meningioma cell lines and reduced proliferation and migration independent of GTPase knockdowns/status. Moreover, overall survival in vivo could also be increased by knockdowns of RhoA and Rac1 as well as Fasudil treatment. GTPase expression was affected dependent on the NF2 status but effects were not very distinct, indicating that NF2 is not strongly involved in GTPase regulation in meningiomas. In terms of mTOR and FAK-Paxillin signaling, each GTPase changes those pathways in a different manner. CONCLUSION Small GTPases are important effectors in meningioma proliferation and migration in vitro as well as survival in vivo and their inhibition should be considered as potential treatment option.

scholarly journals Identification of Ion Channels in Meningioma Cell Lines as Possible Targets for Combination Therapy

2021 ◽  
Vol 35 (S1) ◽  
Author(s):  
Erik White ◽  
Jesus Romero ◽  
Michael Prabhu ◽  
Samantha Beck ◽  
Vikram Prabhu ◽  
...  
Keyword(s):  
Ion Channels ◽  
Combination Therapy ◽  
Cell Lines ◽  
Meningioma Cell

Extracellular matrix-modulated differential invasion of human meningioma cell lines in vitro

1999 ◽  
Vol 263 (2-3) ◽  
pp. 214-216 ◽  
Author(s):  
Harcharan K Rooprai ◽  
Krishanthi Liyanage ◽  
Susan F.D Robinson ◽  
Apsara Kandanearatchi ◽  
Andrew F Dean ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Cell Lines ◽  
Human Meningioma ◽  
Meningioma Cell

Rituximab-Mediated Inhibition of the PI3K-Akt Survival Signaling Pathway in B-NHL Cell Lines Leading to Downregulation of Bclxl Expression and Chemosensitization.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 2830-2830
Author(s):  
Eriko Suzuki ◽  
Ali R. Jazirehi ◽  
Benjamin Bonavida
Keyword(s):  
Cell Survival ◽  
Signaling Pathway ◽  
Cell Lines ◽  
Kinase Inhibitor ◽  
Akt Pathway ◽  
Akt Inhibitor ◽  
Drug Induced ◽  
Similar Time ◽  
Cell Lysates ◽  
Induced Apoptosis

Abstract Rituximab (chimeric anti-CD20 monoclonal antibody) has been used in the treatment of B-NHL. We have reported in vitro that rituximab treatment signals B-NHL cell lines Ramos and Daudi and inhibits both the ERK 1/2 MAPK and NF-κB signaling pathways leading to selective inhibition of Bclxl expression and sensitization to drug-induced apoptosis. The inhibition of the NF-κB signaling pathway by rituximab was shown to be due, in part, to the induction of the Raf Kinase Inhibitor Protein (RKIP) (Jazirehi, et al., 2005 Cancer Research 65:264–276). The PI3K-Akt signaling pathway is a key regulator of cell survival and aberrant activation of the PI3K-Akt pathway has been implicated in both drug resistance and resistance to apoptosis-inducing stimuli. Akt can promote cell survival by indirectly activating the proximal transcription factor NF-κB through the phosphorylation of I-kappa B kinase (I-κB) (Ozes et. al. Nature401:82–85, 1999). This study investigated whether NF-κB inhibition by rituximab and downregulation of Bclxl expression was also the result of rituximab-mediated inhibition of the PI3K-Akt pathway. Ramos and Daudi B-NHL cell lines were treated with rituximab (20 ug/ml) and cell lysates were prepared and both Akt and phospho-Akt (p-Akt) expression were examined by western blot. The findings demonstrate that both cell lines show constitutively activated p-Akt and treatment with rituximab significantly inhibited p-Akt but not Akt. Time kinetics analysis demonstrated that inhibition of p-Akt was first detected at 3–6 hours following rituximab treatment and inhibition was maintained up to 24 hours. Concomitantlly, a similar time kinetics revealed inhibition of NF-κB activity as assessed by EMSA. Since the inhibition of NF-kB activity resulted in significant downregulation of Bclxl expression, we also examined the role of the Akt pathway in the regulation of Bclxl expression. Tumor cells were treated with the Akt inhibitor LY294002 and analysis of cell lysates showed significant downregulation of Bclxl expression. Rituximab was previously shown to sensitize B-NHL cells to drug-induced apoptosis via inhibition of NF-κB activity and Bclxl expression. We examined if inhibition of the Akt pathway also chemosensitized the cells. Treatment of Ramos cells with the Akt inhibitor LY294002 significantly sensitized the cells to CDDP-induced apoptosis and synergy was achieved. Altogether, these findings demonstrate, for the first time, that rituximab inhibits the Akt pathway and that this pathway is involved in the regulation of tumor- cell resistance to chemotherapeutic drugs. This study also proposes that the Akt pathway is a potential targeting pathway for therapeutic intervention in the treatment of rituximab and drug-resistant B-NHL.

Proteomic Analysis Identifies Differences in Multiple Myeloma (MM) Cells Sensitive and Resistant to AKT Inhibition.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3402-3402
Author(s):  
Alissa Huston ◽  
Alexey Leontovich ◽  
Michael Timm ◽  
Yazan Alsayed ◽  
Ujjal Singha ◽  
...  
Keyword(s):  
Heat Shock ◽  
Heat Shock Protein ◽  
Cell Lines ◽  
Proteomic Analysis ◽  
Molecular Mechanisms ◽  
Treatment Time ◽  
Control Sample ◽  
Differentially Expressed ◽  
Akt Inhibitor ◽  
The Mean

Abstract Prior studies have demonstrated that MM cells with PTEN mutation and high AKT activity are more sensitive to inhibitors of the PI3K/Akt/mTOR pathway. However, the molecular mechanisms that regulate the differential response to these agents are not well characterized. The objective of this study was to determine proteins that are different between MM cell lines with higher AKT activity (OPM2) and lower AKT activity (MM.1S) in response to the AKT inhibitor, perifosine. Methods: MM cell lines (MM.1S and OPM2) were treated with serial concentration of perifosine (KRX-0401, Keryx, NY, NY, provided by the NCI). Proteomic analysis using the nanoscale BD Clontech antibody-based protein microarray technique was performed using cells treated with perifosine (10uM for 16 hrs) or vehicle (sterile water) as control. Apoptosis was determined using Annexin V/PI FACS analysis at 24 and 48 hrs. The treatment time and concentration were chosen so that it did not induce more than 25% apoptosis to ensure adequate analysis of changes in signaling pathways. The antibody microarray is a technique that detects differences in protein abundance between the treated and control sample with each experiment by hybridizing fluorescently labeled (Cy3 and Cy5) protein mixtures onto slides spotted with 512 human monoclonal antibodies. Two microarray slides were used for each experiment. The slides were scanned using the Axon GenePix 4000B scanner. Two ratios were generated from the spot images for each protein target. The mean of the ratios of Cy5/Cy3 of both slides were analyzed using Clontech software and used to calculate an Internally Normalized Ratio (INR = (Ratio1/Ratio2, ratios 1 and 2 correspond to slides 1 and 2) for each spot on the array. The INR values were input into GeneSpring 6.0 software (Silicon Genetics, CA). The data was normalized to the mean INR of the two cell lines. Proteins whose expression level changed relative to control greater than 1.3 fold were determined. Unsupervised clustering demonstrated a different protein signature between MM.1S and OPM2 in response to perifosine. There were 144 proteins differentially expressed by 1.3 fold between MM.1S and OPM2. Proteins that were downregulated in OPM2 as compared to MM.1S included those in the PI3K pathway and cell cycle regulation such as PTEN, p70S6Kinase, the AKT substrate GSK-3, eEF-2 kinase, eIF-4g, Ku-80, cyclin A, E2F-2, CDK2, CDK7, and c-myc; proteins involved in apoptosis such as p21WAF, caspase 4 and 8, FADD, and PARP; kinases such as PKAc, PKA RI, ERK2, and JNK1; and other proteins regulating apoptosis and proliferation including p53, the NF-kB inhibitor IkB, and the heat shock protein HSP70. The nanoscale protein array is a useful and rapid technique that may be used to identify differences between resistant and sensitive cells to novel therapeutic agents. We identified proteins that are differentially expressed between MM cells sensitive and relatively resistant to the AKT inhibitor perifosine. Further analysis of the role of these proteins in the mechanism of resistance/sensitivity to perifosine is being performed. Future use of inhibitors of NF-kB (bortezomib) or heat shock protein inhibitors in conjunction with perifosine may overcome resistance induced by these proteins in MM cells with low AKT activity. Supported in part by an ASH scholar award and an MMRF grant.

Akt Inhibitor Perifosine-Induced Cytotoxicity Is Associated with Significant Downregulation of Survivin in Human Multiple Myeloma (MM) Cells.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 3410-3410
Author(s):  
Teru Hideshima ◽  
Hiroshi Yasui ◽  
Laurence Catley ◽  
Noopur Raje ◽  
Dharminder Chauhan ◽  
...  
Keyword(s):  
Cell Lines ◽  
Molecular Mechanisms ◽  
Caspase 3 ◽  
Single Agent ◽  
Survivin Expression ◽  
Mek Inhibitor ◽  
Parp Cleavage ◽  
Caspase 8 ◽  
Akt Inhibitor ◽  
Apoptotic Proteins

Abstract Perifosine (NSC 639966; Keryx Biopharmaceuticals, New York, NY) is a synthetic novel alkylphospholipid, a new class of anti-tumor agents which potently inhibits Akt (PKB) activity. Our previous studies have shown that Perifosine induces significant cytotoxicity in MM cells triggered by c-Jun NH2-terminal kinase (JNK) activation followed by caspase-8, caspase-9, and PARP cleavage even in the presence of cytokines (ie, IL-6 and IGF-1) or bone marrow stromal cell (BMSCs). Importantly, MEK inhibitor and bortezomib enhance Perifosine-induced cytotoxicity. It has also shown significant anti-tumor activity in a human MM cell xenograft mouse model (Hideshima et al. Blood2006, 107:4053–4062). In this study, we further delineated molecular mechanisms whereby Perifosine triggers cytotoxicity as a single agent and in combination with bortezomib in MM cells. In most MM cell lines, the IC50 for Perifosine-induced cytotoxicity is 5–10 μM range assessed by MTT assay at 24h; however, apoptosis assessed by APO2.7 staining, varied in each cell line. Moreover, neither the degree of JNK phosphorylation nor caspase-8/9/PARP cleavage correlated with Perifosine-induced cytotoxicity. Therefore we further examined expression level of anti-apoptotic proteins in MM cell lines and found that survivin, which has a crucial role in regulation of caspase-3 activity, was markedly downregulated by Perifosine treatment in a time- and dose-dependent fashion, without affecting expression of other anti-apoptotic proteins (ie, cIAP, XIAP, Bcl-2, Bcl-xL). Since survivin is a known downstream protein of β-catenin/TCF-4 cascade, we next hypothesized that Perifosine may inhibit β-catenin activity. As expected, Perifosine significantly downregulated both phosphorylation and protein expression of β-catenin, associated with downregulation of survivin and enhanced caspase-3 cleavage. Real-time PCR confirmed that gene expression of survivin was suppressed 35% and 55% after 3h and 6h Perifosine treatment, respectively. Since β-catenin is a substrate of proteasomes, we further examined whether bortezomib could augment survivin expression by blocking its degradation. Importantly, bortezomib significantly upregulated β-catenin and survivin, which was blocked in the presence of Perifosine. These results suggest that inhibition of bortezomib-induced survivin expression, at least in part, accounts for enhanced bortezomib-induced cytotoxicity by Perifosine. Based upon these preclinical studies, a rational combination trial of bortezomib with Perifosine to treat relapsed refractory MM is currently ongoing.

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