mk-2206 and Glioma

Research over the past decade has suggested important roles for pseudogenes in glioma. This study aimed to show that pseudogene PRELI domain-containing 1 pseudogene 6 (PRELID1P6) promotes glioma progression. Aberrant expression of genes was screened using The Cancer Genome Atlas database. We found that mRNA level of PRELID1P6 was highly upregulated in glioma and was associated with a shorter survival time. Functional studies showed that the knockdown of PRELID1P6 decreased cell proliferation, sphere formation, and clone formation ability and blocked the cell cycle transition at G0/G1, while overexpression of PRELID1P6 had the opposite effects. Mechanistically, knockdown of PRELID1P6 changed the cellular localization of heterogeneous nuclear ribonucleoprotein H1 (hnRNPH1) from nucleus to cytoplasm, which promoted ubiquitin-mediated degradation of hnRNPH1. RNA-sequence and gene set enrichment analysis suggested that knockdown of PRELID1P6 regulates the apoptosis signaling pathway. Western blotting showed that PRELID1P6 increased TRF2 expression by hnRNPH1-mediated alternative splicing effect and activated the Akt/mTOR pathway. Furthermore, Akt inhibitor MK2206 treatment reversed the oncogenic function of PRELID1P6. PRELID1P6 was also found to be negatively regulated by miR-1825. Our result showed that PRELID1P6 promotes glioma progression through the hnHNPH1-Akt/mTOR pathway. These findings shed new light on the important role of PRELID1P6 as a novel oncogene for glioma.

Topics: Animals; Brain Neoplasms; Cell Line, Tumor; Female; Glioma; Heterocyclic Compounds, 3-Ring; Heterogeneous-Nuclear Ribonucleoproteins; Humans; Male; Mice; Mice, Inbred BALB C; Mice, Nude; Mitochondrial Proteins; Proto-Oncogene Proteins c-akt; Pseudogenes; Signal Transduction; TOR Serine-Threonine Kinases

Vasculogenic mimicry (VM), the formation of vessel-like structures by highly invasive tumor cells, has been considered one of several mechanisms responsible for the failure of anti-angiogenesis therapy in glioma patients. Therefore, inhibiting VM formation might be an effective therapeutic method to antagonize the angiogenesis resistance. This study aimed to show that an extracellular protein called Tenascin-c (TNC) is involved in VM formation and that TNC knockdown inhibits VM in glioma. TNC was upregulated with an increase in glioma grade. TNC and VM formation are potential independent predictors of survival of glioma patients. TNC upregulation was correlated with VM formation, and exogenous TNC stimulated VM formation. Furthermore, TNC knockdown significantly suppressed VM formation and proliferation in glioma cells in vitro and in vivo, with a reduction in cellular invasiveness and migration. Mechanistically, TNC knockdown decreased Akt phosphorylation at Ser

Topics: Animals; Apoptosis; Brain Neoplasms; Cell Line, Tumor; Cell Movement; Cell Proliferation; Female; Glioma; Heterocyclic Compounds, 3-Ring; Heterografts; Humans; Male; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Mice; Mice, Inbred BALB C; Mice, Nude; Middle Aged; Neoplasm Grading; Neovascularization, Pathologic; Phosphorylation; Proto-Oncogene Proteins c-akt; Tenascin; Up-Regulation

microRNA-489 (miR-489), a newly identified tumor-related miRNA, functions as an oncogene or tumor suppressor via regulating growth and metastasis of human cancers. But, the clinical significance, biological function and underlying mechanisms of miR-489 in glioma remain rarely known. Here, we showed that the levels of miR-489 in glioma tissues were notably underexpressed compared to corresponding non-tumor tissues. In accordance, the relative levels of miR-489 were decreased in glioma cell lines compared with NHA cells. Kaplan-Meier plots indicated that miR-489 low expressing glioma patients showed a prominent shorter overall survival. In addition, miR-489 overexpression prohibited proliferation and cell cycle progression, and promoted apoptosis in U251 cells. While, miR-489 knockdown showed opposite effects on these cellular processes of U87 cells. In vivo experiments demonstrated that miR-489 restoration reduced the tumor volume and weight of subcutaneous glioma xenografts in nude mice. Notably, Spindlin 1 (SPIN1) was inversely and directly regulated by miR-489 in glioma cells. A negative correlation between the expression of miR-489 and SPIN1 mRNA was confirmed in glioma tissues. Interestingly, miR-489 inversely modulated activation of PI3K/AKT pathway and expression of downstream targets including p-mTOR, Cyclin D1 and BCL-XL. SPIN1 re-expression abolished the effects of miR-489 on U251 cells with enhanced activation of PI3K/AKT pathway and malignant phenotype. Meanwhile, AKT inhibitor MK-2206 blocked activation of PI3K/AKT pathway and resulted in reduced proliferation, cell cycle arrest and increased apoptosis in miR-489 down-regulating U87 cells. Altogether, our data support that miR-489 loss facilitates malignant phenotype of glioma cells probably via SPIN1-mediated PI3K/AKT pathway.

Topics: Animals; Apoptosis; Brain Neoplasms; Cell Cycle; Cell Cycle Proteins; Cell Line, Tumor; Cell Proliferation; Female; Gene Expression Regulation, Neoplastic; Gene Knockdown Techniques; Glioma; Heterocyclic Compounds, 3-Ring; Humans; Male; Mice, Nude; MicroRNAs; Microtubule-Associated Proteins; Middle Aged; Multivariate Analysis; Phenotype; Phosphatidylinositol 3-Kinases; Phosphoproteins; Phosphorylation; Prognosis; Proportional Hazards Models; Proto-Oncogene Proteins c-akt; Signal Transduction

Molecular targeted therapy can potentially provide more effective treatment for patients with high-grade gliomas. Notch and Akt are notable target molecules as they play important roles in a variety of cellular processes, such as regeneration, differentiation, proliferation, migration, and invasion. Here, we assessed the therapeutic possibility of inhibiting Notch and Akt in gliomas using the clinically available, selective small molecule inhibitors MRK003 and MK-2206. We evaluated their efficacy individually and as a combination therapy in U251 and U87 glioma cell lines. We confirmed that MK-2206 effectively inhibits Akt phosphorylation in a dose-dependent manner, whereas MRK003 inhibits Notch signaling and Akt phosphorylation. Both MRK003 and MK-2206 significantly inhibited cell growth, migration, and invasion in a dose-dependent manner. Akt dephosphorylation was enhanced by combination therapy with MRK003 and MK-2206. However, the effect of combination treatment did not exceed that of MK-2206 monotherapy in proliferation assay. Inhibition of invasion, further enhanced by combination therapy, correlated with increased Akt inactivation. In summary, combination therapy with MRK003 and MK-2206 may be effective for inhibiting invasion but not proliferation.

Topics: Antineoplastic Agents; Brain Neoplasms; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cyclic S-Oxides; Drug Interactions; Glioma; Heterocyclic Compounds, 3-Ring; Humans; Molecular Targeted Therapy; Neoplasm Invasiveness; Phosphorylation; Proto-Oncogene Proteins c-akt; Receptors, Notch; Signal Transduction; Thiadiazoles

Gefitinib, a small molecule inhibitor of the epidermal growth factor receptor tyrosine kinase, has been shown to induce autophagy as well as apoptosis in tumor cells. Yet, how to use autophagy and apoptosis to improve therapeutic efficacy of this drug against cancer remains to be explored. We reported here that MK-2206, a potent allosteric Akt inhibitor currently in phase I trials in patients with solid tumors, could reinforce the cytocidal effect of gefitinib against glioma. We found that cotreatment with gefitinib and MK-2206 increased the cytotoxicity of this growth factor receptor inhibitor in the glioma cells, and the CompuSyn synergism/antagonism analysis showed that MK-2206 acted synergistically with gefitinib. The benefit of the combinatorial treatment was also shown in an intracranial glioma mouse model. In the presence of MK-2206, there was a significant increase in apoptosis in glioma cells treated with gefitinib. MK-2206 also augmented the autophagy-inducing effect of gefitinib, as evidenced by increased levels of the autophagy marker, LC3-II. Inhibition of autophagy by silencing of the key autophagy gene, beclin 1 or 3-MA, further increased the cytotoxicity of this combinatorial treatment, suggesting that autophagy induced by these agents plays a cytoprotective role. Notably, at 48 hours following the combinatorial treatment, the level of LC3-II began to decrease but Bim was significantly elevated, suggesting a switch from autophagy to apoptosis. On the basis of the synergistic effect of MK-2206 on gefitinib observed in this study, the combination of these two drugs may be utilized as a new therapeutic regimen for malignant glioma.

Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Apoptosis Regulatory Proteins; Autophagy; Bcl-2-Like Protein 11; Beclin-1; Brain Neoplasms; Cell Line, Tumor; Drug Synergism; ErbB Receptors; Gefitinib; Glioma; Heterocyclic Compounds, 3-Ring; Humans; Male; Membrane Proteins; Mice; Mice, Inbred BALB C; Microtubule-Associated Proteins; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-akt; Quinazolines; RNA Interference; RNA, Small Interfering

Eukaryotic elongation factor-2 (eEF-2) kinase, also known as calmodulin-dependent protein kinase III, is a unique calcium/calmodulin-dependent enzyme. eEF-2 kinase can act as a negative regulator of protein synthesis and a positive regulator of autophagy under environmental or metabolic stresses. Akt, a key downstream effector of the PI3K signaling pathway that regulates cell survival and proliferation, is an attractive therapeutic target for anticancer treatment. Akt inhibition leads to activation of both apoptosis, type I programmed cell death and autophagy, a cellular degradation process via lysosomal machinery (also termed type II programmed cell death). However, the underlying mechanisms that dictate functional relationship between autophagy and apoptosis in response to Akt inhibition remain to be delineated. Our recent study demonstrated that inhibition of eEF-2 kinase can suppress autophagy but promote apoptosis in tumor cells subjected to Akt inhibition, indicating a role of eEF-2 kinase as a controller in the crosstalk between autophagy and apoptosis. Furthermore, inhibition of eEF-2 kinase can reinforce the efficacy of a novel Akt inhibitor, MK-2206, against human glioma. These findings may help optimize the use of Akt inhibitors in the treatment of cancer and other diseases.

Topics: Antineoplastic Agents; Apoptosis; Autophagy; Brain Neoplasms; Cell Line, Tumor; Cell Lineage; Cell Survival; eIF-2 Kinase; Enzyme Inhibitors; Gene Expression Regulation, Enzymologic; Glioma; Heterocyclic Compounds, 3-Ring; Humans; Models, Biological; Proto-Oncogene Proteins c-akt