Role of autophagy in temozolomide-induced cytotoxicity for malignant glioma cells

Kanzawa, Takao; Germano, Isabelle M.; Komata, Tadashi; Ito, Hiroki; Kondo, Yasuko; Kondo, Shinji

doi:10.1038/sj.cdd.4401359

Cited by 912 publications

(877 citation statements)

References 35 publications

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“…To begin to examine the antitumor effect of TMZ on malignant glioma cell lines, unsychronized U251 GBM cells were first treated with TMZ (100 mM for 3 h) and further incubated for 3-14 days in the absence of TMZ, after which cells were analyzed by FACS for DNA content. Consistent with previous reports, [11][12][13] cells accumulated with 4N DNA content at the G2-M boundary, with ratio of 4N/2N DNA content peaking 3 days after TMZ treatment and decreasing gradually to base levels by 10 days post-TMZ exposure (Figures 1a and b).…”

Section: Resultssupporting

confidence: 92%

“…Consistent with previous reports, no apoptotic cells were detected microscopically or by cell cycle analysis (lack of cells with sub-G1 DNA content). [11][12][13] Finally, because recent investigations also demonstrated that radiation or chemotherapeutic agents including TMZ induce autophagy in malignant glioma cells, 13,14 we measured conversion of microtubule-associated protein 1 lightchain 3 (MAP1-LC3) (LC3-I, 18 kDa), to a lipidized form (LC3-II, 16 kDa) in TMZ-treated cells as a measure of autophagy. As shown in Figure 1d, TMZ induced an accumulation of the LC3-II form of MAP1-LC3 as early as 3 days after TMZ exposure, similar to what was noted following a 1-day exposure of the cells to the known inducer of autophagy, rapamycin.…”

Section: Resultsmentioning

confidence: 99%

“…11,12 In a previous study, it was shown that TMZ induces autophagy, and that pharmacological inhibition of autophagy could influence cellular oucome. 13 The exact means by which this occurred, however, and the potential role of autophagy -induced ATP production in the process, were not determined. In the present study, we investigated the relationship between autophagy, autophagy-associated ATP production, and cell death in response to TMZ and a second DNA damaging agent Etoposide.…”

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DNA damaging agent-induced autophagy produces a cytoprotective adenosine triphosphate surge in malignant glioma cells

et al. 2006

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Although autophagy enhances cell survival in nutrient-deprived cells by increasing adenosine triphosphate (ATP) production, it remains unclear if autophagy functions similarly in cells treated with cytotoxic chemotherapy agents. To address this issue, we measured both the ability of DNA damaging agents (Temozolomide, and Etoposide) to induce an autophagy-dependent production of ATP, and the effects of modulation of autophagy on drug-induced cell death. Both drugs induced an autophagyassociated increase in ATP production in multiple glioma cell lines. The drug-induced ATP surge could not be blocked by glucose starvation, but could be blocked by preincubation with the autophagy inhibitor 3-methyladenine (3-MA), an siRNA targeting beclin 1, or the mitochondrial inhibitor oligomycin. Inhibition of autophagy-induced ATP production increased nonapoptotic cell death associated with micronucleation, while restoration of the 3-MA-inhibited ATP surge by addition of pyruvate suppressed cell death. These results show that DNA damaging agents induce an autophagy-associated ATP surge that protects cells and may contribute to drug resistance. Autophagy is a process by which subcellular constituents are degraded in autophagosomes/autolysosomes in response to stress. 1 Whereas growth factors enhance the ability of metazoan cells to take up extracelluar nutrients and suppress autophagy, deprivation of either growth factors or extracellular nutrients leads to decreased nutrient uptake and derepression of autophagy. The catabolic autophagy process is believed to allow starving cells to support adenosine triphosphate (ATP) production and avoid cell death. 2,3 The autophagic process is regulated by both class I and class III phosphatidylinositol 3-kinase (PI3K) pathways. 2,4,5 Mammalian target of Rapamycin (mTOR) functions downstream of the class-I PI3K/Akt, and is a key negative regulator of autophagy. In this role, mTOR serves as a metabolic sensor that coordinates crosstalk between nutrient availability and autophagy. 6,7 In contrast, class III PI3K positively regulates autophagy. The formation of a preautophagosomal isolation membrane that encloses cytoplasmic cargo is positively regulated by phosphatidylinositol 3-phosphate, the products of class III PI3K phosphorylation. 4 It has also been suggested that the trafficking of autophagosomes is regulated by class III PI3Ks, 4,5 an idea supported by the observation that 3-methyladenine (3-MA), which inhibits class III PI3K activity, also inhibits autophagosome formation and is widely used as an autophagy inhibitor. 5 It is also known that autophagy is induced in cancer cells derived from tissues such as breast, colon, prostate and brain in response to a variety of anticancer therapies. 3,8 However, autophagy in response to these drugs can produce different outcomes, in some cases promoting cell adaptation and survival similar to what is seen in nutrient deprived cells, while in other cases, inducing programmed cell death type. The role of drug-induced autophagy in cell death or surv...

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Section: Resultssupporting

confidence: 92%

Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

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DNA damaging agent-induced autophagy produces a cytoprotective adenosine triphosphate surge in malignant glioma cells

et al. 2006

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“…For example, to block autophagy at early stages and prevent autophagosome formation, one could inhibit the Type III PI3 kinase complex with 3-methyl adenine or knockdown Beclin 1 or Atg5 with siRNAs, while to inhibit autophagy at later stages one could prevent fusion between the autophagosome and lysosome by knocking down or inhibiting Rab7 or use pharmacological agents such as the lysomotropic agent chloroquine or bafilomycin A1, an inhibitor of the vacuolar type H + ATPase. It may be important to discriminate between blocking autophagy at early or later steps because differences in outcome for the cell have been noted when this has been done (42). When the later stages of autophagy are blocked, the cell may accumulate large numbers of autophagosomes.…”

Section: Regulation Of Autophagymentioning

confidence: 99%

Apoptosis and autophagy: regulatory connections between two supposedly different processes

2007

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Apoptosis and autophagy are genetically-regulated, evolutionarily-conserved processes that regulate cell fate. Both apoptosis and autophagy are important in development and normal physiology and in a wide range of diseases. Recent studies show that despite the marked differences between these two processes, their regulation is intimately connected and the same regulators can sometimes control both apoptosis and autophagy. In this review, I discuss some of these findings, which provide possible molecular mechanisms for crosstalk between apoptosis and autophagy and suggest that it may be useful to think of these processes as different facets of the same cell death continuum rather than completely separate processes. KeywordsAutophagy; Apoptosis; Bcl-2, FADD; Atg 5In the early to mid 1990s there was a rapid increase in our understating about the regulation of apoptosis. Fifteen or so years later we are in the midst of a similarly exciting period for understanding autophagy with very rapid growth of publications on the regulation and function of this process (1). Autophagy (the form of autophagy discussed here is macroautophagy, other forms-microautophagy and chaperone-mediated autophagy share the same lysosomal degradation mechanism but differ in the way that material is delivered to the lysosome) is a degradative process involving sequestration of parts of the cytoplasm in double membrane vesicles (autophagic vesicles or autophagosomes) that fuse with lysosomes forming the autophagolysosome. In the autophagolysosome, the cytoplasmic material that was engulfed is hydrolyzed and the resulting amino acids and other macromolecular precursors can be recycled (2-4), see Fig 1 for a schematic of the process. Autophagy is a mechanism by which organelles are removed and is the primary degradation mechanism for long-lived proteins and thus maintains quality control for proteins and organelles. Autophagy is ubiquitous in eukaryotic cells and important in development and in diverse pathophysiological conditions (5)-for example providing protection against neurodegeneration (6,7), infections (8,9) and tumor development (10-13). Cells that are deficient in autophagy also demonstrate enhanced chromosomal instability (14,15), which may be related to the tumorigenesis associated with autophagy deficiency.Autophagy is important in cell death decisions and can protect cells by preventing them from undergoing apoptosis. For example, increased autophagy in nutrient deprived or growth factorwithdrawn cells allows cell survival (16,17) by inhibiting apoptosis. Autophagy also protects cells from various other apoptotic stimuli (18). It is not clear how autophagy stops cells from undergoing apoptosis; one suggested mechanism is the sequestration of damaged mitochondria (18) thus preventing released cytochrome c from being able to form a functional apoptosome NIH Public Access Autophagic cell death (also known as Type II programmed cell death to distinguish it from apoptosis or Type I programmed cell death) (20-22) has been describ...

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“…The cytotoxic effect of TMZ can be affected by complex molecular pathways 20. Herein, we observed that the expression of FBW7 was markedly inhibited at both protein and mRNA levels in U251/TMZ, a cell line which was isolated in our previous experiment with 4‐fold 50% inhibition rate (IC50; data not shown), compared to the parental U251 cell line ( P ‐value <.01 for quantitative RT‐PCR, Figure 3A,B), suggesting the potential role of FBW7 in mediating TMZ response.…”

Section: Resultsmentioning

confidence: 99%

FBW7 is associated with prognosis, inhibits malignancies and enhances temozolomide sensitivity in glioblastoma cells

Lin¹,

Qiu

et al. 2018

Cancer Science

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F‐box and WD repeat domain‐containing 7 (FBW7) is a SCF‐type E3 ubiquitin ligase targeting a multitude of oncoproteins for degradation. Acting as one of the most important tumor suppressors, it is frequently inactivated in various tumors. In this study we aimed to evaluate the relationship of FBW7 with glioma pathology and prognosis, and examine its effect in glioma malignancies and temozolomide (TMZ)‐based therapy. Clinical tissues and TCGA database analysis revealed that FBW7 expression was correlated inversely with glioma histology and positively with patient survival time. Lentivirus transfection‐induced FBW7 overexpression significantly suppressed proliferation, invasion and migration of U251 and U373 cells, whereas knockdown of FBW7 by targeted shRNA promoted proliferation, invasion and migration of glioma cells. Most importantly, the expression level of FBW7 was found to affect the 50% inhibitory concentration (IC50) of U251 and the TMZ‐resistant variant. Combining TMZ with FBW7 overexpression notably increased the cytotoxicity compared to TMZ treatment alone, which was conversely attenuated by FBW7 knockdown. Moreover, flow cytometry (FC) analysis showed overexpression of FBW7, TMZ or the combination‐increased proportion of G2/M arrest and the apoptotic rate, whereas FBW7 inhibition reduced G2/M arrest and apoptosis in U251 cells. Finally, mechanistic study found that FBW7 overexpression downregulated Aurora B, Mcl1 and Notch1 levels in a time‐dependent pattern and this expressional suppression was independent of TMZ. These findings collectively demonstrate the critical role of FBW7 as a prognostic factor and a potential target to overcome chemoresistance of glioblastoma.

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Role of autophagy in temozolomide-induced cytotoxicity for malignant glioma cells

Cited by 912 publications

References 35 publications

DNA damaging agent-induced autophagy produces a cytoprotective adenosine triphosphate surge in malignant glioma cells

DNA damaging agent-induced autophagy produces a cytoprotective adenosine triphosphate surge in malignant glioma cells

Apoptosis and autophagy: regulatory connections between two supposedly different processes

FBW7 is associated with prognosis, inhibits malignancies and enhances temozolomide sensitivity in glioblastoma cells

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