scholarly journals A SYSTEM TO IDENTIFY INHIBITORS OF MTOR SIGNALING USING HIGH-RESOLUTION GROWTH ANALYSIS IN S. CEREVISIAE

2019 ◽  
Vol 3 (Supplement_1) ◽  
pp. S101-S102
Author(s):  
Michael Kiflezghi ◽  
Matt Kaeberlein
Keyword(s):  
Risk Factor ◽  
Specific Inhibitor ◽  
Mtor Inhibitors ◽  
Mtor Signaling ◽  
Differential Sensitivity ◽  
Model Organisms ◽  
Mtor Inhibition ◽  
Transplant Patients ◽  
Pain And Suffering ◽  
Mtor Complex 1

Abstract Age is the main risk factor for cancer, cardiovascular disease, neurodegeneration and other diseases prevalent in the world’s aging population. These diseases increase the pain and suffering and result in billions of dollars in healthcare costs. Addressing the common risk factor may allow for simultaneous amelioration of these diseases providing personal and economic relief to people and societies around the globe. The mTOR signaling pathway has been shown to be a robust target in the aging process with its inhibition resulting in increased lifespan in model organisms from yeast to mice. Rapamycin is an FDA approved drug for use in transplant patients and is a potent and specific inhibitor of mTOR complex 1. Rapamycin’s use as an anti-aging therapeutic in otherwise healthy individuals is complicated by the occurrence of side effects. As such, groups studying inhibition of the mTOR pathway are searching for alternative inhibitors that may be able decouple the deleterious effects of rapamycin administration from the lifespan extending effects. To address this issue, we have developed a high-throughput yeast-based assay for the identification of novel mTOR inhibitors. By utilizing mutant strains with differential sensitivity to mTOR inhibition, comparative growth kinetics of microcultures exposed to an inhibitor can be used to discern the mTOR inhibitory status of a compound. Furthermore, the assay can provide mechanistic insight on a compound’s mode of inhibition providing a rich, fast readout of a compound’s potential for inhibiting mTOR. This approach allows for the screening of large libraries of compounds speeding the discovery process.

scholarly journals Impact on Autophagy and Ultraviolet B Induced Responses of Treatment with the MTOR Inhibitors Rapamycin, Everolimus, Torin 1, and pp242 in Human Keratinocytes

2017 ◽  
Vol 2017 ◽  
pp. 1-21 ◽  
Author(s):  
Song Xu ◽  
Li Li ◽  
Min Li ◽  
Mengli Zhang ◽  
Mei Ju ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Cellular Response ◽  
Mtor Inhibitors ◽  
Mtor Signaling ◽  
Ultraviolet B ◽  
Epidermal Keratinocytes ◽  
Human Epidermal Keratinocytes ◽  
Mtor Inhibition ◽  
The Impact ◽  
Uva Radiation

The mechanistic target of Rapamycin (MTOR) protein is a crucial signaling regulator in mammalian cells that is extensively involved in cellular biology. The function of MTOR signaling in keratinocytes remains unclear. In this study, we detected the MTOR signaling and autophagy response in the human keratinocyte cell line HaCaT and human epidermal keratinocytes treated with MTOR inhibitors. Moreover, we detected the impact of MTOR inhibitors on keratinocytes exposed to the common carcinogenic stressors ultraviolet B (UVB) and UVA radiation. As a result, keratinocytes were sensitive to the MTOR inhibitors Rapamycin, everolimus, Torin 1, and pp242, but the regulation of MTOR downstream signaling was distinct. Next, autophagy induction only was observed in HaCaT cells treated with Rapamycin. Furthermore, we found that MTOR signaling was insensitive to UVB but sensitive to UVA radiation. UVB treatment also had no impact on the inhibition of MTOR signaling by MTOR inhibitors. Finally, MTOR inhibition by Rapamycin, everolimus, or pp242 did not affect the series of biological events in keratinocytes exposed to UVB, including the downregulation of BiP and PERK, activation of Histone H2A and JNK, and cleavage of caspase-3 and PARP. Our study demonstrated that MTOR inhibition in keratinocytes cannot always induce autophagy, and the MTOR pathway does not play a central role in the UVB triggered cellular response.

SAT-212-Treatment with mTOR inhibitors is an independent risk factor for chronic hepatitis E in transplant patients with increased transaminases levels

2019 ◽  
Vol 70 (1) ◽  
pp. e723
Author(s):  
Mar Riveiro-Barciela ◽  
Judit Vidal-González ◽  
Luisa Roade ◽  
Joan Martínez Camprecios ◽  
Basilio Rodríguez ◽  
...  
Keyword(s):  
Chronic Hepatitis ◽  
Risk Factor ◽  
Independent Risk Factor ◽  
Mtor Inhibitors ◽  
Hepatitis E ◽  
Transplant Patients

mTOR Signaling Is Required for the Survival and Proliferation of Hematopoietic Stem/Progenitor Cells

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 4787-4787
Author(s):  
Xia Xiao ◽  
Hongmei Luo ◽  
Amanda C. LaRue ◽  
Bradley A. Schulte ◽  
Yong Wang
Keyword(s):  
Progenitor Cells ◽  
Cell Transformation ◽  
Conflicts Of Interest ◽  
Mtor Inhibitors ◽  
Mtor Signaling ◽  
Immunofluorescent Staining ◽  
Mouse Bone Marrow ◽  
Mtor Inhibition ◽  
Hematopoietic Stem

Abstract Abstract 4787 The mammalian target of rapamycin (mTOR) is a serine/threonine kinase that regulates protein synthesis, gene transcription, cell growth and cell proliferation. Previous studies have demonstrated that abnormalities in the Pten-mTOR pathway may contribute to the development of leukemia and lead to premature exhaustion of hematopoietic stem cells (HSCs). These findings suggest a role for mTOR in the regulation of HSC self-renewal and cell transformation. The aim of this study was to investigate the involvement of mTOR signaling in the survival and proliferation of HSCs and hematopoietic progenitor cells (HPCs). Immunofluorescent staining with antibodies against phosphorylated mTOR and S6 kinase revealed that Thrombopoietin (Tpo) activates mTOR in cultured mouse bone marrow (BM) lineage negative cells, suggesting that Tpo may promote the survival and proliferation of HSCs/HPCs via activation of the mTOR signaling pathway. Further studies revealed that treatment with an mTOR specific small molecule inhibitor (Ku-69734) significantly suppressed the colony-forming ability of HPCs as evidenced by a dose-dependent decrease in the production of CFU-GM, BFU-E and CFU-GEMM. We also examined the clonogenic function of HSCs using cobblestone-area forming cell (CAFC) assays and found that Ku-69734 treatment markedly reduced the number of CAFCs in long-term BM culture. Moreover, immunophenotyping and flow cytometric analyses showed that inhibition of mTOR induced apoptosis primarily in HSCs and to a lesser degree in HPCs, indicating that mTOR inhibition may suppress the clonogenic function of HSCs and HPCs via the induction of apoptosis. Together, these data demonstrate that mTOR signaling is required for the survival and proliferation of HSCs and HPCs. Given that many mTOR inhibitors are currently in clinical trials for the treatment of cancers, our findings provide the rationale to further evaluate the potential toxicity of mTOR inhibitors in HSCs/HPCs in vivo. Disclosures: No relevant conflicts of interest to declare.

scholarly journals mTOR Inhibitors for the Treatment of Severe Congenital Hyperinsulinism: Perspectives on Limited Therapeutic Success

2016 ◽  
Vol 101 (12) ◽  
pp. 4719-4729 ◽  
Author(s):  
Marie Szymanowski ◽  
Maria Salomon Estebanez ◽  
Raja Padidela ◽  
Bing Han ◽  
Karolina Mosinska ◽  
...  
Keyword(s):  
Glycemic Control ◽  
Medical Therapy ◽  
Mtor Inhibitor ◽  
Pancreatic Insufficiency ◽  
Safety Information ◽  
Mtor Inhibitors ◽  
Mtor Signaling ◽  
Congenital Hyperinsulinism ◽  
Mtor Inhibition ◽  
Fasting Tolerance

Context: Congenital hyperinsulinism (CHI) is the most common cause of persistent hypoglycemia in neonates and infants. In medically unresponsive CHI, subtotal pancreatectomy is performed to achieve euglycemia with consequent diabetes in later life. Sirolimus, a mammalian target of rapamycin (mTOR) inhibitor, has been reported to obviate the need for pancreatectomy, but experience is limited. Objective: We have investigated the efficacy and adverse effect profile of mTOR inhibitors in the treatment of severe CHI. Design, Setting, and Patients: This was an observational review of 10 severe CHI patients treated with mTOR inhibitors, in France and the United Kingdom, with the intention of achieving glycemic control without pancreatectomy. Safety information was recorded. Main Outcome Measure(s): We examined whether mTOR inhibitors achieved glycemic control, fasting tolerance, and weaning of supportive medical therapy. Results: mTOR inhibition achieved euglycemia, fasting tolerance, and reduced medical therapy in only three patients (30%). Triglyceride levels were elevated in five patients (50%). One child required a blood transfusion for anemia, four had stomatitis, two had sepsis, one developed varicella zoster, and two patients developed gut dysmotility in association with exocrine pancreatic insufficiency. In silico analysis of transcriptome arrays from CHI patients revealed no significant association between mTOR signaling and disease. Pancreatic tissue from two patients who did not respond to sirolimus showed no reduction in cell proliferation, further suggesting that mTOR signaling did not down-regulate proliferation in the CHI pancreas. Conclusion: mTOR inhibitor treatment is associated with very limited success and must be used with caution in children with severe CHI.

scholarly journals Inhibition of Mammalian Target of Rapamycin Induces Phosphatidylinositol 3-Kinase-Dependent and Mnk-Mediated Eukaryotic Translation Initiation Factor 4E Phosphorylation

2007 ◽  
Vol 27 (21) ◽  
pp. 7405-7413 ◽  
Author(s):  
Xuerong Wang ◽  
Ping Yue ◽  
Chi-Bun Chan ◽  
Keqiang Ye ◽  
Takeshi Ueda ◽  
...  
Keyword(s):  
Translation Initiation ◽  
Mammalian Target Of Rapamycin ◽  
Mtor Inhibitors ◽  
Mtor Signaling ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Phosphatidylinositol 3 Kinase ◽  
Mtor Inhibition ◽  
Eukaryotic Translation ◽  
Eukaryotic Translation Initiation

ABSTRACT The initiation factor eukaryotic translation initiation factor 4E (eIF4E) plays a critical role in initiating translation of mRNAs, including those encoding oncogenic proteins. Therefore, eIF4E is considered a survival protein involved in cell cycle progression, cell transformation, and apoptotic resistance. Phosphorylation of eIF4E (usually at Ser209) increases its binding affinity for the cap of mRNA and may also favor its entry into initiation complexes. Mammalian target of rapamycin (mTOR) inhibitors suppress cap-dependent translation through inhibition of the phosphorylation of eIF4E-binding protein 1. Paradoxically, we have shown that inhibition of mTOR signaling increases eIF4E phosphorylation in human cancer cells. In this study, we focused on revealing the mechanism by which mTOR inhibition increases eIF4E phosphorylation. Silencing of either mTOR or raptor could mimic mTOR inhibitors’ effects to increase eIF4E phosphorylation. Moreover, knockdown of mTOR, but not rictor or p70S6K, abrogated rapamycin's ability to increase eIF4E phosphorylation. These results indicate that mTOR inhibitor-induced eIF4E phosphorylation is secondary to mTOR/raptor inhibition and independent of p70S6K. Importantly, mTOR inhibitors lost their ability to increase eIF4E phosphorylation only in cells where both Mnk1 and Mnk2 were knocked out, indicating that mTOR inhibitors increase eIF4E phosphorylation through a Mnk-dependent mechanism. Given that mTOR inhibitors failed to increase Mnk and eIF4E phosphorylation in phosphatidylinositol 3-kinase (PI3K)-deficient cells, we conclude that mTOR inhibition increases eIF4E phosphorylation through a PI3K-dependent and Mnk-mediated mechanism. In addition, we also suggest an effective therapeutic strategy for enhancing mTOR-targeted cancer therapy by cotargeting mTOR signaling and Mnk/eIF4E phosphorylation.

Cellular and molecular effects of the mTOR inhibitor everolimus

2015 ◽  
Vol 129 (10) ◽  
pp. 895-914 ◽  
Author(s):  
Uttara Saran ◽  
Michelangelo Foti ◽  
Jean-François Dufour
Keyword(s):  
Mtor Inhibitor ◽  
Mtor Inhibitors ◽  
Cancer Therapies ◽  
Breast Cancers ◽  
Mtor Inhibition ◽  
Growth And Survival ◽  
Molecular Effects ◽  
Mtor Complex 1

mTOR (mechanistic target of rapamycin) functions as the central regulator for cell proliferation, growth and survival. Up-regulation of proteins regulating mTOR, as well as its downstream targets, has been reported in various cancers. This has promoted the development of anti-cancer therapies targeting mTOR, namely fungal macrolide rapamycin, a naturally occurring mTOR inhibitor, and its analogues (rapalogues). One such rapalogue, everolimus, has been approved in the clinical treatment of renal and breast cancers. Although results have demonstrated that these mTOR inhibitors are effective in attenuating cell growth of cancer cells under in vitro and in vivo conditions, subsequent sporadic response to rapalogues therapy in clinical trials has promoted researchers to look further into the complex understanding of the dynamics of mTOR regulation in the tumour environment. Limitations of these rapalogues include the sensitivity of tumour subsets to mTOR inhibition. Additionally, it is well known that rapamycin and its rapalogues mediate their effects by inhibiting mTORC (mTOR complex) 1, with limited or no effect on mTORC2 activity. The present review summarizes the pre-clinical, clinical and recent discoveries, with emphasis on the cellular and molecular effects of everolimus in cancer therapy.

Adaptation to chronic mTOR inhibition in cancer and in aging

2013 ◽  
Vol 41 (4) ◽  
pp. 956-961 ◽  
Author(s):  
Rebecca Gilley ◽  
Kathryn Balmanno ◽  
Claire L. Cope ◽  
Simon J. Cook
Keyword(s):  
Drug Target ◽  
Kinase Inhibitors ◽  
Mtor Inhibitors ◽  
Allosteric Inhibitors ◽  
Mtor Inhibition ◽  
Cell Responses ◽  
Downstream Effector ◽  
Signalling Network ◽  
Effector Pathways ◽  
Mtor Complex 1

The mTOR [mammalian (or mechanistic) target of rapamycin] protein kinase co-ordinates catabolic and anabolic processes in response to growth factors and nutrients and is a validated anticancer drug target. Rapamycin and related allosteric inhibitors of mTORC1 (mTOR complex 1) have had some success in specific tumour types, but have not exhibited broad anticancer activity, prompting the development of new ATP-competitive mTOR kinase inhibitors that inhibit both mTORC1 and mTORC2. In common with other targeted kinase inhibitors, tumours are likely to adapt and acquire resistance to mTOR inhibitors. In the present article, we review studies that describe how tumour cells adapt to become resistant to mTOR inhibitors. mTOR is a central signalling hub which responds to an array of signalling inputs and activates a range of downstream effector pathways. Understanding how this signalling network is remodelled and which pathways are invoked to sustain survival and proliferation in the presence of mTOR inhibitors can provide new insights into the importance of the various mTOR effector pathways and may suggest targets for intervention to combine with mTOR inhibitors. Finally, since chronic mTOR inhibition by rapamycin can increase lifespan and healthspan in nematodes, fruitflies and mice, we contrast these studies with tumour cell responses to mTOR inhibition.

scholarly journals Functional Impacts of the BRCA1-mTORC2 Interaction in Breast Cancer

2019 ◽  
Vol 20 (23) ◽  
pp. 5876 ◽  
Author(s):  
Kimiko L. Krieger ◽  
Wen-Feng Hu ◽  
Tyler Ripperger ◽  
Nicholas T. Woods
Keyword(s):  
Breast Cancer ◽  
Dna Damage ◽  
Cancer Cells ◽  
Breast Cancer Cells ◽  
Mtor Inhibitors ◽  
Mtor Signaling ◽  
Protein Domain ◽  
Mtor Inhibition ◽  
Increased Risk ◽  
The Impact

Deleterious mutations in Breast Cancer 1 (BRCA1) are associated with an increased risk of breast and ovarian cancer. Mutations in the tandem BRCA1 C-terminal (tBRCT) protein domain disrupt critical protein interactions required for the faithful repair of DNA through homologous recombination, which contributes to oncogenesis. Our studies have identified RICTOR, PRR5, and SIN1 subunits of the mammalian target of rapamycin complex 2 (mTORC2) as interacting partners with the tBRCT domain of BRCA1 leading to the disruption of the mTORC2 complex. However, the interplay between mTORC2 signaling and BRCA1 function in the DNA damage response (DDR) remains to be determined. In this study, we used protein interaction assays to determine the binary interactions between the tBRCT domain and mTORC2 subunits, evaluated the impact of mTOR inhibition on the transcriptional function of the tBRCT, evaluated the impact of mTOR signaling on BRCA1 recruitment to DNA damage-induced foci and determined the breast cancer cell line response to mTOR inhibition dependent upon BRCA1 expression and mutation. This study determined that PRR5, RICTOR, and SIN1 could each independently interact with the BRCA1 tBRCT. Inhibition of mTORC1, but not mTORC1/2, increases BRCA1 transcriptional activation activity. Treatment with pan-mTOR inhibitor PP242 diminishes DNA damage-induced γH2AX and BRCA1 foci formation. Breast cancer cells lacking expression of functional BRCA1 are more sensitive to mTOR inhibitors. These data suggest that mTOR signaling is required for BRCA1 response to DNA damage and breast cancer cells lacking BRCA1 are more sensitive to pan-mTOR inhibition. This work suggests chemotherapeutic strategies using mTOR inhibitors could be tailored for patients that lack functional BRCA1.

scholarly journals mTOR Inhibition: From Aging to Autism and Beyond

Scientifica ◽  
2013 ◽  
Vol 2013 ◽  
pp. 1-17 ◽  
Author(s):  
Matt Kaeberlein
Keyword(s):  
Mtor Inhibitors ◽  
Cellular Level ◽  
Mtor Signaling ◽  
Target Of Rapamycin ◽  
Mtor Inhibition ◽  
Pharmacological Modulation ◽  
Related Disease ◽  
Mechanistic Target Of Rapamycin ◽  
Age Related ◽  
Growing Body

The mechanistic target of rapamycin (mTOR) is a highly conserved protein that regulates growth and proliferation in response to environmental and hormonal cues. Broadly speaking, organisms are constantly faced with the challenge of interpreting their environment and making a decision between “grow or do not grow.” mTOR is a major component of the network that makes this decision at the cellular level and, to some extent, the tissue and organismal level as well. Although overly simplistic, this framework can be useful when considering the myriad functions ascribed to mTOR and the pleiotropic phenotypes associated with genetic or pharmacological modulation of mTOR signaling. In this review, I will consider mTOR function in this context and attempt to summarize and interpret the growing body of literature demonstrating interesting and varied effects of mTOR inhibitors. These include robust effects on a multitude of age-related parameters and pathologies, as well as several other processes not obviously linked to aging or age-related disease.

scholarly journals Sirolimus and Other Mechanistic Target of Rapamycin Inhibitors Directly Activate Latent Pathogenic Human Polyomavirus Replication

2020 ◽  
Author(s):  
Jennifer Alvarez Orellana ◽  
Hyun Jin Kwun ◽  
Sara Artusi ◽  
Yuan Chang ◽  
Patrick S Moore
Keyword(s):  
Dna Replication ◽  
Jc Virus ◽  
Mtor Inhibitors ◽  
Bk Virus ◽  
Target Of Rapamycin ◽  
Human Polyomavirus ◽  
Merkel Cell ◽  
Mtor Inhibition ◽  
Mechanistic Target Of Rapamycin ◽  
Transplant Patients

Abstract Background Human polyomaviruses can reactivate in transplant patients, causing nephropathy, progressive multifocal leukoencephalopathy, Merkel cell carcinoma, pruritic, rash or trichodysplasia spinulosa. Sirolimus and related mechanistic target of rapamycin (mTOR) inhibitors are transplant immunosuppressants. It is unknown if they directly reactivate polyomavirus replication from latency beyond their general effects on immunosuppression. Methods In vitro expression and turnover of large T (LT) proteins from BK virus, JC virus (JCV), Merkel cell polyomavirus (MCV), human polyomavirus 7 (HPyV7), and trichodysplasia spinulosa polyomavirus (TSV) after drug treatment were determined by immunoblotting, proximity ligation, replicon DNA replication, and whole virus immunofluorescence assays. Results mTOR inhibition increased LT protein expression for all 5 pathogenic polyomaviruses tested. This correlated with LT stabilization, decrease in the S-phase kinase-associated protein 2 (Skp2) E3 ligase targeting these LT proteins for degradation, and increase in virus replication for JCV, MCV, TSV, and HPyV7. Treatment with sirolimus, but not the calcineurin inhibitor tacrolimus, at levels routinely achieved in patients, resulted in a dose-dependent increase in viral DNA replication for BKV, MCV, and HPyV7. Conclusions mTOR inhibitors, at therapeutic levels, directly activate polyomavirus replication through a Skp2-dependent mechanism, revealing a proteostatic latency mechanism common to polyomaviruses. Modifying existing drug regimens for transplant patients with polyomavirus-associated diseases may reduce symptomatic polyomavirus replication while maintaining allograft-sparing immunosuppression.

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