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Sample records for everolimus prevents mtor

  1. mTOR treatment in lymphangioleiomyomatosis: the role of everolimus.

    PubMed

    Yates, Deborah H

    2016-01-01

    The orphan lung disease lymphangioleiomyomatosis (LAM) has until recently been untreatable other than by lung transplantation. However, improved understanding of underlying disease mechanisms has revealed the central role of constitutive up-regulation of the mammalian target of rapamycin (mTOR) pathway in this disease. Although other pathways exist and are under investigation for treatment, several mTOR inhibitors are currently available and emerging information suggests that these may have some efficacy in preventing loss of lung function in LAM. This paper summarizes current understanding of treatment with mTOR inhibitors in LAM, and everolimus in particular. It outlines pharmacokinetics and pharmacodynamics relevant to the clinician, recent clinical studies, and issues with potential side effects. mTOR treatment is not yet available in most countries for LAM, but current data for treatment efficacy are impressive, and it is hoped that mTOR inhibition will soon be recognised as an important treatment of this disease.

  2. mTOR Inhibition by Everolimus Does Not Impair Closure of Punch Biopsy Wounds in Renal Transplant Patients

    PubMed Central

    Dutt, Shelley B.; Gonzales, Josephine; Boyett, Megan; Costanzo, Anne; Han, Peggy P.; Steinberg, Steven; McKay, Dianne B.; Jameson, Julie M.

    2017-01-01

    Background Mammalian target of rapamycin (mTOR) inhibitors are approved to prevent allograft rejection and control malignancy. Unfortunately, they are associated with adverse effects, such as wound healing complications that detract from more extensive use. There is a lack of prospective wound healing studies to monitor patients treated with mTOR inhibitors, such as everolimus or sirolimus, especially in nondiabetics. Methods Patients receiving everolimus with standard immunosuppressant therapy or standard immunosuppressant therapy without everolimus were administered 3-mm skin biopsy punch wounds in the left scapular region. Homeostatic gene expression was examined in the skin obtained from the biopsy and wound surface area was examined on day 7. Peripheral blood mononuclear cells were examined for cytokine production. Results There are no significant changes in autophagy related 13, epidermal growth factor, insulin-like growth factor binding protein 3, IL-2, kruppel-like factor 4, and TGFB1 gene expression in the skin suggesting that there is little impact of everolimus on these genes within nonwounded skin. Peripheral blood T cells are more sensitive to cell death in everolimus-treated patients, but they retain the ability to produce proinflammatory cytokines required for efficient wound repair. Importantly, there is no delay in the closure of biopsy wounds in patients receiving everolimus as compared to those not receiving mTOR inhibition. Conclusions Everolimus treatment is not associated with impaired closure of skin biopsy wounds in kidney transplant recipients. These data highlight the importance of exploring whether larger surgical wounds would show a similar result and how other factors, such as diabetes, impact wound healing complications associated with mTOR suppression.

  3. Delayed mTOR Inhibition with Low Dose of Everolimus Reduces TGFβ Expression, Attenuates Proteinuria and Renal Damage in the Renal Mass Reduction Model

    PubMed Central

    Kurdián, Melania; Herrero-Fresneda, Inmaculada; Lloberas, Nuria; Gimenez-Bonafe, Pepita; Coria, Virginia; Grande, María T.; Boggia, José; Malacrida, Leonel; Torras, Joan; Arévalo, Miguel A.; González-Martínez, Francisco; López-Novoa, José M.; Grinyó, Josep; Noboa, Oscar

    2012-01-01

    Background The immunosuppressive mammalian target of rapamycin (mTOR) inhibitors are widely used in solid organ transplantation, but their effect on kidney disease progression is controversial. mTOR has emerged as one of the main pathways regulating cell growth, proliferation, differentiation, migration, and survival. The aim of this study was to analyze the effects of delayed inhibition of mTOR pathway with low dose of everolimus on progression of renal disease and TGFβ expression in the 5/6 nephrectomy model in Wistar rats. Methods This study evaluated the effects of everolimus (0.3 mg/k/day) introduced 15 days after surgical procedure on renal function, proteinuria, renal histology and mechanisms of fibrosis and proliferation. Results Everolimus treated group (EveG) showed significantly less proteinuria and albuminuria, less glomerular and tubulointerstitial damage and fibrosis, fibroblast activation cell proliferation, when compared with control group (CG), even though the EveG remained with high blood pressure. Treatment with everolimus also diminished glomerular hypertrophy. Everolimus effectively inhibited the increase of mTOR developed in 5/6 nephrectomy animals, without changes in AKT mRNA or protein abundance, but with an increase in the pAKT/AKT ratio. Associated with this inhibition, everolimus blunted the increased expression of TGFβ observed in the remnant kidney model. Conclusion Delayed mTOR inhibition with low dose of everolimus significantly prevented progressive renal damage and protected the remnant kidney. mTOR and TGFβ mRNA reduction can partially explain this anti fibrotic effect. mTOR can be a new target to attenuate the progression of chronic kidney disease even in those nephropathies of non-immunologic origin. PMID:22427849

  4. Everolimus

    MedlinePlus

    ... Zortress) is used with other medications to prevent transplant rejection (attack of the transplanted organ by the ... organ) in certain adults who have received kidney transplants. Everolimus is in a class of medications called ...

  5. Differentiating the mTOR inhibitors everolimus and sirolimus in the treatment of tuberous sclerosis complex.

    PubMed

    MacKeigan, Jeffrey P; Krueger, Darcy A

    2015-12-01

    Tuberous sclerosis complex (TSC) is a genetic autosomal dominant disorder characterized by benign tumor-like lesions, called hamartomas, in multiple organ systems, including the brain, skin, heart, kidneys, and lung. These hamartomas cause a diverse set of clinical problems based on their location and often result in epilepsy, learning difficulties, and behavioral problems. TSC is caused by mutations within the TSC1 or TSC2 genes that inactivate the genes' tumor-suppressive function and drive hamartomatous cell growth. In normal cells, TSC1 and TSC2 integrate growth signals and nutrient inputs to downregulate signaling to mammalian target of rapamycin (mTOR), an evolutionarily conserved serine-threonine kinase that controls cell growth and cell survival. The molecular connection between TSC and mTOR led to the clinical use of allosteric mTOR inhibitors (sirolimus and everolimus) for the treatment of TSC. Everolimus is approved for subependymal giant cell astrocytomas and renal angiomyolipomas in patients with TSC. Sirolimus, though not approved for TSC, has undergone considerable investigation to treat various aspects of the disease. Everolimus and sirolimus selectively inhibit mTOR signaling with similar molecular mechanisms, but with distinct clinical profiles. This review differentiates mTOR inhibitors in TSC while describing the molecular mechanisms, pathogenic mutations, and clinical trial outcomes for managing TSC.

  6. Targeting the mTOR Complex by Everolimus in NRAS Mutant Neuroblastoma.

    PubMed

    Kiessling, Michael K; Curioni-Fontecedro, Alessandra; Samaras, Panagiotis; Lang, Silvia; Scharl, Michael; Aguzzi, Adriano; Oldrige, Derek A; Maris, John M; Rogler, Gerhard

    2016-01-01

    High-risk neuroblastoma remains lethal in about 50% of patients despite multimodal treatment. Recent attempts to identify molecular targets for specific therapies have shown that Neuroblastoma RAS (NRAS) is significantly mutated in a small number of patients. However, few inhibitors for the potential treatment for NRAS mutant neuroblastoma have been investigated so far. In this in-vitro study, we show that MEK inhibitors AZD6244, MEK162 and PD0325901 block cell growth in NRAS mutant neuroblastoma cell lines but not in NRAS wild-type cell lines. Several studies show that mutant NRAS leads to PI3K pathway activation and combined inhibitors of PI3K/mTOR effectively block cell growth. However, we observed the combination of MEK inhibitors with PI3K or AKT inhibitors did not show synergestic effects on cell growth. Thus, we tested single mTOR inhibitors Everolimus and AZD8055. Interestingly, Everolimus and AZD8055 alone were sufficient to block cell growth in NRAS mutant cell lines but not in wild-type cell lines. We found that Everolimus alone induced apoptosis in NRAS mutant neuroblastoma. Furthermore, the combination of mTOR and MEK inhibitors resulted in synergistic growth inhibition. Taken together, our results show that NRAS mutant neuroblastoma can be targeted by clinically available Everolimus alone or in combination with MEK inhibitors which could impact future clinical studies.

  7. Effects of the mTOR inhibitor everolimus and the PI3K/mTOR inhibitor NVP-BEZ235 in murine acute lung injury models.

    PubMed

    Üstün, Sevdican; Lassnig, Caroline; Preitschopf, Andrea; Mikula, Mario; Müller, Mathias; Hengstschläger, Markus; Weichhart, Thomas

    2015-09-01

    The mammalian target of rapamycin (mTOR) is a key signaling kinase associated with a variety of cellular functions including the regulation of immunological and inflammatory responses. Classic mTOR inhibitors such as rapamycin or everolimus are commonly used in transplant as well as cancer patients to prevent transplant rejection or cancer progression, respectively. Noninfectious drug-induced pneumonitis is a frequent side effect in mTOR-inhibitor-treated patients. Therefore, we tested the effects of the mTOR inhibitor everolimus and the novel dual PI3K/mTOR inhibitor NVP-BEZ235 in a murine lipopolysaccharide (LPS)-induced acute lung injury model. C57BL/6 mice were treated with either everolimus or NVP-BEZ235 on two consecutive days prior to intratracheal administration of LPS. LPS administration induced a significant increase in total cell, neutrophil and erythrocyte numbers in the bronchoalveolar lavage fluid. Histological examination revealed a serious lung injury as shown by interstitial edema, vascular congestion and mononuclear cell infiltration in these mice after 24h. Everolimus as well as NVP-BEZ235 did not noticeably affect overall histopathology of the lungs in the lung injury model. However, NVP-BEZ235 enhanced IL-6 and TNF-α expression after 24h. In contrast, everolimus did not affect IL-6 and TNF-α levels. Interestingly, both inhibitors reduced inflammatory cytokines in an LPS/oleic acid-induced lung injury model. In conclusion, the mTOR inhibitors did not worsen the overall histopathological severity, but they exerted distinct effects on proinflammatory cytokine expression in the lung depending on the lung injury model applied.

  8. Nanosecond Pulsed Electric Fields Enhance the Anti-tumour Effects of the mTOR Inhibitor Everolimus against Melanoma

    NASA Astrophysics Data System (ADS)

    Dai, Jie; Wu, Shan; Kong, Yan; Chi, Zhihong; Si, Lu; Sheng, Xinan; Cui, Chuanliang; Fang, Jing; Zhang, Jue; Guo, Jun

    2017-01-01

    The PI3K/mTOR/AKT pathway is activated in most melanomas, but mTOR inhibitors used singly have limited activity against advanced melanomas. The application of nanosecond pulsed electric fields (nsPEFs) is a promising cancer therapy approach. In this study, we evaluated the synergistic anti-tumour efficacy of the mTOR inhibitor everolimus in conjunction with nsPEFs against melanoma. The combined treatment of nsPEFs and everolimus gradually decreased cell growth concurrent with nsPEF intensity. nsPEFs alone or combined with everolimus could promote melanoma cell apoptosis, accompanied with a loss in cellular mitochondrial membrane potential and an increase in Ca2+ levels. In vivo experiments showed that a combination of the mTOR inhibitor everolimus and nsPEFs improved the inhibitory effect, and all skin lesions caused by nsPEFs healed in 1 week without any observed adverse effect. Combination treatment induced caspase-dependent apoptosis through the upregulation of the pro-apoptotic factor Bax and downregulation of the anti-apoptotic factor Bcl-2. Everolimus and nsPEFs synergistically inhibited angiogenesis by decreasing the expression of vascular endothelial growth factor (VEGF), VEGF receptor (VEGFR), and CD34. Our findings indicate that nsPEFs in combination with an mTOR inhibitor can be used as a potential treatment approach for advanced melanoma.

  9. Nanosecond Pulsed Electric Fields Enhance the Anti-tumour Effects of the mTOR Inhibitor Everolimus against Melanoma

    PubMed Central

    Dai, Jie; Wu, Shan; Kong, Yan; Chi, Zhihong; Si, Lu; Sheng, Xinan; Cui, Chuanliang; Fang, Jing; Zhang, Jue; Guo, Jun

    2017-01-01

    The PI3K/mTOR/AKT pathway is activated in most melanomas, but mTOR inhibitors used singly have limited activity against advanced melanomas. The application of nanosecond pulsed electric fields (nsPEFs) is a promising cancer therapy approach. In this study, we evaluated the synergistic anti-tumour efficacy of the mTOR inhibitor everolimus in conjunction with nsPEFs against melanoma. The combined treatment of nsPEFs and everolimus gradually decreased cell growth concurrent with nsPEF intensity. nsPEFs alone or combined with everolimus could promote melanoma cell apoptosis, accompanied with a loss in cellular mitochondrial membrane potential and an increase in Ca2+ levels. In vivo experiments showed that a combination of the mTOR inhibitor everolimus and nsPEFs improved the inhibitory effect, and all skin lesions caused by nsPEFs healed in 1 week without any observed adverse effect. Combination treatment induced caspase-dependent apoptosis through the upregulation of the pro-apoptotic factor Bax and downregulation of the anti-apoptotic factor Bcl-2. Everolimus and nsPEFs synergistically inhibited angiogenesis by decreasing the expression of vascular endothelial growth factor (VEGF), VEGF receptor (VEGFR), and CD34. Our findings indicate that nsPEFs in combination with an mTOR inhibitor can be used as a potential treatment approach for advanced melanoma. PMID:28054548

  10. Attributable Risk of Infection to mTOR Inhibitors Everolimus and Temsirolimus in the Treatment of Cancer.

    PubMed

    Garcia, Christine A; Wu, Shenhong

    2016-11-25

    The risk of infection attributable to mTOR inhibitors has not been determined. Databases from PubMed and abstracts presented at the American Society of Clinical Oncology meetings were searched. Eligible studies included randomized controlled trials, in which everolimus or temsirolimus was compared with placebo. A total of 12 trials were included. The attributable incidences of all-grade and high-grade infections to mTOR inhibitors were 9.3% (95% confidence interval (CI): 5.8-14.6%) and 2.3% (95% CI: 1.2-4.4%) respectively. The risk varied widely with tumor types (p <.001). There was substantial risk of infection attributable to mTOR inhibitors everolimus and temsirolimus.

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

    PubMed Central

    Xu, Song; Li, Li; Li, Min; Zhang, Mengli

    2017-01-01

    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.

  12. mTOR inhibitor-associated stomatitis (mIAS) in three patients with cancer treated with everolimus.

    PubMed

    Kalogirou, Eleni-Marina; Tosios, Konstantinos I; Piperi, Evangelia P; Sklavounou, Alexandra

    2015-01-01

    Mammalian targets of rapamycin inhibitors (mTOR inhibitors, mTORI) are indicated for the management of several cancer types, including hormone receptor--positive or HER2-negative breast cancer, advanced renal cell carcinoma, advanced neuroendocrine tumors of pancreatic origin, and tuberous sclerosis complex-related tumors. Among the most common adverse events of mTORI medication are discrete, large, solitary or multiple, superficial ulcers, almost exclusively situated on nonkeratinized oral mucosa, described as mTORI-associated stomatitis (mIAS). We describe the clinical presentation, course, and management of mIAS in three patients receiving the mTORI everolimus (Afinitor, Novartis, East Hanover, NJ). In two patients, mIAS manifested 9 and 30 days after first using everolimus, respectively, whereas in the third patient, it recurred 3 months after re-introduction of everolimus. Oral rinses with a "magic mouthwash" solution (dexamethasone oral drops solution 2 mg/mL × 10 mL, lidocaine gel 2% × 30 g, doxycycline suspension 50 mg/5 mL × 60 mL, and sucralfate oral suspension 1000 mg/5 mL × 150 mL, dissolved in sodium chloride 0.9% × 2000 mL) four times daily proved helpful in alleviating the symptoms, and the ulcers healed in 4 to 15 days. No side effects were recorded, and dose reduction or discontinuation of everolimus was not necessitated in two cases.

  13. Everolimus as an mTOR Inhibitor Suppresses Endometriotic Implants: an Experimental Rat Study

    PubMed Central

    Kacan, T.; Yildiz, C.; Baloglu Kacan, S.; Seker, M.; Ozer, H.; Cetin, A.

    2017-01-01

    Introduction Mammalian target of rapamycin is a pathway to block apoptosis. Recent studies showed that the activity of mammalian target of rapamycin pathway increases in endometriotic lesions. Aim of the present study was to study the effect of everolimus agent, a rapamycin analog, in an experimental endometriosis model. Materials and Methods Endometriosis established by the autotransplantation of uterine tissue in the peritoneal cavity was confirmed in 24 rats. The animals were then randomly divided into three groups to receive either everolimus (1.5 mg/kg/day, p. o.), anastrozole (0.004 mg/day, p. o.), or normal saline (0.1 mL, i. p.) for 14 days. Endometriotic foci were excised, stained with hematoxylin and eosin, and endometriosis was scored semiquantitatively. In addition, immunohistochemical examination were performed using primary antibodies of vascular endothelial growth factor, CD117, and Bax. Results Both anastrozole and everolimus lowered endometriosis scores. Significant decreases in ovarian follicles were observed following anastrozole treatment but not everolimus treatment. Conclusion Through its apoptosis-promoting effect, everolimus suppressed endometriotic foci without negatively affecting ovarian reserve. These findings support the hypothesis that everolimus merits further study on the way to developing a new endometriosis drug. PMID:28190891

  14. Phosphorylation of mTOR and S6RP predicts the efficacy of everolimus in patients with metastatic renal cell carcinoma

    PubMed Central

    2014-01-01

    Background The incidence of renal cell cancer (RCC) has been increasing for the past decade, and the 5-year survival for patients with metastatic RCC (mRCC) is rather low. Everolimus (RAD001), a new inhibitor for mammalian target of rapamycin (mTOR), is generally well tolerated, and demonstrates clinical benefit to patients with anti-VEGF-refractory mRCC. However, factors for selection of patients who may benefit from everolimus remain largely unknown. Here we aimed to explore potential molecular indicators for mRCC patients who may benefit from everolimus treatment. Methods Paraffin-embedded tumor tissue specimens derived from 18 mRCC patients before everolimus treatment, who participated the phase 1b trial of everolimus in VEGF receptor (VEGFR)-tyrosine kinase inhibitor (TKI)-refractory Chinese patients with mRCC (clinicaltrials.gov, NCT01152801), were examined for the expression levels of phosphorylated AKT, mTOR, eukaryotic initiation factor 4E (eIF4E) binding protein-1 (4EBP1) and 40S ribosomal protein S6 (S6RP) by immunohistochemistry. Clinical benefit rate (complete response [CR], partial response [PR], plus stable disease [SD] ≥ 6 months) and progression-free survival time (PFS) were correlated with expression levels of these mTOR-associated molecules. Results In these 18 patients, there were 1 PR, 15 SDs (including 9 SDs ≥ 6 months), and 2 progressive diseases (PD). The clinical benefit rate (CBR) was 55.6% (10/18), and the median PFS time was 8.4 months. Patients with positive expression of phospho-mTOR showed a better CBR (71.4% versus 0%, P = 0.023) and PFS time (11.3 versus 3.7 months, P = 0.001) than those patients with negative expression. The median PFS of patients with positive phospho-S6RP expression was longer (11.3 versus 3.7 months, P = 0.002) than that of patients negative for phospho-S6RP expression. However, expression levels of phospho-4EBP1 and phospho-AKT were unassociated to efficacy of everolimus treatment

  15. Profile of everolimus in the treatment of tuberous sclerosis complex: an evidence-based review of its place in therapy

    PubMed Central

    Capal, Jamie K; Franz, David Neal

    2016-01-01

    Tuberous sclerosis complex (TSC) is a relatively rare genetic disorder, affecting one in 6,000 births. Mammalian target of rapamycin (mTOR) inhibitors, such as everolimus, which have been previously used to prevent solid organ transplant rejection, augment anticancer treatment regimens, and prevent neovascularization of artificial cardiac stents, are now approved for treating TSC-related manifestations, such as subependymal giant cell astrocytomas and renal angiomyolipomas. The use of everolimus in treating subependymal giant cell astrocytomas is supported by long-term Phase II and III clinical trials. Seizures are a common feature in TSC, occurring in up to 96% of patients. While mTOR inhibitors currently do not have regulatory approval in treating this manifestation, small clinical studies have demonstrated beneficial outcomes with everolimus. Further evidence from a forthcoming Phase III clinical study may provide additional support for the use of everolimus for this indication. Also, there are no approved treatments for TSC-associated neuropsychiatric disorders, which include intellectual disability, behavioral difficulties, and autism spectrum disorder, but preclinical data and small studies have suggested that some neuropsychiatric symptoms may be improved through mTOR inhibition therapy. More evidence is needed, particularly regarding safety in young infants. This review focuses on the current evidence supporting the use of everolimus in neurologic and neuropsychiatric manifestations of TSC, and the place of everolimus in therapy. PMID:27601910

  16. Profile of everolimus in the treatment of tuberous sclerosis complex: an evidence-based review of its place in therapy.

    PubMed

    Capal, Jamie K; Franz, David Neal

    2016-01-01

    Tuberous sclerosis complex (TSC) is a relatively rare genetic disorder, affecting one in 6,000 births. Mammalian target of rapamycin (mTOR) inhibitors, such as everolimus, which have been previously used to prevent solid organ transplant rejection, augment anticancer treatment regimens, and prevent neovascularization of artificial cardiac stents, are now approved for treating TSC-related manifestations, such as subependymal giant cell astrocytomas and renal angiomyolipomas. The use of everolimus in treating subependymal giant cell astrocytomas is supported by long-term Phase II and III clinical trials. Seizures are a common feature in TSC, occurring in up to 96% of patients. While mTOR inhibitors currently do not have regulatory approval in treating this manifestation, small clinical studies have demonstrated beneficial outcomes with everolimus. Further evidence from a forthcoming Phase III clinical study may provide additional support for the use of everolimus for this indication. Also, there are no approved treatments for TSC-associated neuropsychiatric disorders, which include intellectual disability, behavioral difficulties, and autism spectrum disorder, but preclinical data and small studies have suggested that some neuropsychiatric symptoms may be improved through mTOR inhibition therapy. More evidence is needed, particularly regarding safety in young infants. This review focuses on the current evidence supporting the use of everolimus in neurologic and neuropsychiatric manifestations of TSC, and the place of everolimus in therapy.

  17. Prevention of Hepatitis B Virus Reactivation With Lamivudine in a Patient With Advanced Renal Cell Carcinoma Treated With Everolimus.

    PubMed

    D'Aniello, Carmine; Maruzzo, Marco; Basso, Umberto

    2016-01-01

    Anticancer agents may trigger reactivation of hepatitis B virus infection ensuing in asymptomatic to severe liver damage. Preemptive administration of antiviral agents such as lamivudine to patients receiving cytotoxic chemotherapy has been shown to inhibit viral replication and prevent such events. No data are available so far concerning the coadministration of antiviral agents and everolimus, an oral mammalian target of rapamycin inhibitor recently approved for the treatment of advanced renal cell carcinoma. We present in this study the first case to our knowledge of a hepatitis B surface antigen-positive patient with metastatic renal cell carcinoma who has been successfully treated with prophylactic lamivudine and everolimus. Long-term depletion of viral replication was obtained along with stabilization of lung and bone metastases. Hepatitis B surface antigen positivity may be found in up to 10% of cancer patients but should not be considered a contraindication to treatment with everolimus.

  18. Dosing and Safety Implications for Oncologists When Administering Everolimus to Patients With Hormone Receptor-Positive Breast Cancer.

    PubMed

    Rugo, Hope S

    2016-02-01

    Aberrations in the phosphoinositide 3-kinase/protein kinase B/mammalian target of rapamycin pathway are common abnormalities in breast cancer and are associated with the development of resistance to endocrine- and human epidermal growth factor receptor (HER)2-targeted therapies. Because of the significant improvement in progression-free survival for everolimus plus exemestane compared with exemestane plus placebo, everolimus, an mTOR inhibitor, was approved in the United States for the treatment of patients with hormone receptor-positive (HR+), HER-negative, advanced breast cancer whose disease had progressed while receiving letrozole or anastrozole. To provide optimal prevention and management strategies, it is crucial that clinicians are aware of the adverse events (AEs) associated with mTOR inhibition. Understanding the appropriate dose modifications will help reduce toxicity and improve drug tolerance, thus achieving the optimal benefit from everolimus. Analyses of data from the Breast Cancer Trials of Oral Everolimus 2 trial have shown that, despite a greater frequency of AEs in the everolimus plus exemestane treatment arm, the AEs were effectively managed with temporary dose reductions or interruptions. In some cases, the full dose of everolimus could be resumed. Despite a lower mean dose and duration of exposure in patients aged ≥ 70 versus < 70 years, everolimus plus exemestane was similarly efficacious, suggesting that appropriate dose reductions for toxicity will not adversely impact efficacy. Appropriate modification of the everolimus dose and dose delay according to the severity of AEs, with resumption of the optimal dose of everolimus when toxicity has improved, will positively affect patient outcomes in HR+ advanced breast cancer.

  19. Systemic and CNS activity of the RET inhibitor vandetanib combined with the mTOR inhibitor everolimus in KIF5B-RET re-arranged non-small cell lung cancer with brain metastases.

    PubMed

    Subbiah, Vivek; Berry, Jenny; Roxas, Michael; Guha-Thakurta, Nandita; Subbiah, Ishwaria Mohan; Ali, Siraj M; McMahon, Caitlin; Miller, Vincent; Cascone, Tina; Pai, Shobha; Tang, Zhenya; Heymach, John V

    2015-07-01

    In-frame fusion KIF5B (the-kinesin-family-5B-gene)-RET transcripts have been characterized in 1-2% of non-small cell lung cancers and are known oncogenic drivers. The RET tyrosine kinase inhibitor, vandetanib, suppresses fusion-induced, anchorage-independent growth activity. In vitro studies have shown that vandetanib is a high-affinity substrate of breast cancer resistance protein (Bcrp1/Abcg2) but is not transported by P-glycoprotein (P-gp), limiting its blood-brain barrier penetration. A co-administration strategy to enhance the brain accumulation of vandetanib by modulating P-gp/Abcb1- and Bcrp1/Abcg2-mediated efflux with mTOR inhibitors, specifically everolimus, was shown to increase the blood-brain barrier penetration. We report the first bench-to-bedside evidence that RET inhibitor combined with an mTOR inhibitor is active against brain-metastatic RET-rearranged lung cancer and the first evidence of blood-brain barrier penetration. A 74-year-old female with progressive adenocarcinoma of the lung (wild-type EGFR and no ALK rearrangement) presented for therapy options. A deletion of 5'RET was revealed by FISH assay, indicating RET-gene rearrangement. Because of progressive disease in the brain, she was enrolled in a clinical trial with vandetanib and everolimus (NCT01582191). Comprehensive genomic profiling revealed fusion of KIF5B (the-kinesin-family-5B-gene) and RET, in addition to AKT2 gene amplification. After two cycles of therapy a repeat MRI brain showed a decrease in the intracranial disease burden and PET/CT showed systemic response as well. Interestingly, AKT2 amplification seen is a critical component of the PI3K/mTOR pathway, alterations of which has been associated with both de novo and acquired resistance to targeted therapy. The addition of everolimus may have both overcome the AKT2 amplification to produce a response in addition to its direct effects on the RET gene. Our case report forms the first evidence of blood-brain barrier penetration by

  20. SAR405, a PIK3C3/Vps34 inhibitor that prevents autophagy and synergizes with MTOR inhibition in tumor cells.

    PubMed

    Pasquier, Benoit

    2015-04-03

    Autophagy plays an important role in cancer and it has been suggested that it functions not only as a tumor suppressor pathway to prevent tumor initiation, but also as a prosurvival pathway that helps tumor cells endure metabolic stress and resist death triggered by chemotherapeutic agents. We recently described the discovery of inhibitors of PIK3C3/Vps34 (phosphatidylinositol 3-kinase, catalytic subunit type 3), the lipid kinase component of the class III phosphatidylinositol 3-kinase (PtdIns3K). This PtdIns3K isoform has attracted significant attention in recent years because of its role in autophagy. Following chemical optimization we identified SAR405, a low molecular mass kinase inhibitor of PIK3C3, highly potent and selective with regard to other lipid and protein kinases. We demonstrated that inhibiting the catalytic activity of PIK3C3 disrupts vesicle trafficking from late endosomes to lysosomes. SAR405 treatment also inhibits autophagy induced either by starvation or by MTOR (mechanistic target of rapamycin) inhibition. Finally our results show that combining SAR405 with everolimus, the FDA-approved MTOR inhibitor, results in a significant synergy on the reduction of cell proliferation using renal tumor cells. This result indicates a potential therapeutic application for PIK3C3 inhibitors in cancer.

  1. Everolimus induces Met inactivation by disrupting the FKBP12/Met complex

    PubMed Central

    Raimondo, Lucia; D'Amato, Valentina; Servetto, Alberto; Rosa, Roberta; Marciano, Roberta; Formisano, Luigi; Mauro, Concetta Di; Orsini, Roberta Clara; Cascetta, Priscilla; Ciciola, Paola; De Maio, Ana Paula; Di Renzo, Maria Flavia; Cosconati, Sandro; Bruno, Agostino; Randazzo, Antonio; Napolitano, Filomena; Montuori, Nunzia; Veneziani, Bianca Maria; Placido, Sabino De; Bianco, Roberto

    2016-01-01

    Inhibition of the mechanistic target of rapamycin (mTOR) is a promising treatment strategy for several cancer types. Rapamycin derivatives such as everolimus are allosteric mTOR inhibitors acting through interaction with the intracellular immunophilin FKBP12, a prolyl isomerase with different cellular functions. Although mTOR inhibitors have significantly improved survival of different cancer patients, resistance and lack of predictive factors of response remain unsolved issues. To elucidate the mechanisms of resistance to everolimus, we evaluated Met activation in everolimus-sensitive/resistant human cancer cells, in vitro and in vivo. Biochemical and computational analyses were performed. Everolimus-resistant cells were xenografted into mice (10/group) and studied for their response to everolimus and Met inhibitors. The statistical significance of the in vitro results was evaluated by Student's t test. Everolimus reduced Met phosphorylation in everolimus-sensitive cells. This event was mediated by the formation of a Met-FKBP12 complex, which in turn is disrupted by everolimus. Aberrant Met activation in everolimus-resistant cells and overexpression of wild-type/mutant Met caused everolimus resistance. Pharmacological inhibition and RNA silencing of Met are effective in condition of everolimus resistance (P<0.01). In mice xenografted with everolimus-resistant cells, the combination of everolimus with the Met inhibitor PHA665752 reduced tumor growth and induced a statistically significant survival advantage (combination vs control P=0.0005). FKBP12 binding is required for full Met activation and everolimus can inhibit Met. Persistent Met activation might sustain everolimus resistance. These results identify a novel everolimus mechanism of action and suggest the development of clinical strategies based on Met inhibitors in everolimus-resistant cancers. PMID:27223077

  2. Targeting the mammalian target of rapamycin pathway with everolimus: implications for the management of metastatic breast cancer.

    PubMed

    Ng, Vin Cci; Johnson, Jeremy J; Cuellar, Sandra

    2015-12-01

    The inhibitors of mammalian target of rapamycin (mTOR) have documented antitumor activity via disruption of various signaling pathways leading to impaired cellular growth, proliferation, and survival. In preclinical studies, mTOR inhibitors use in combination with hormonal therapy has shown promising results in overcoming endocrine resistance in breast cancer cells. The role of everolimus in breast cancer was established in the Breast Cancer Trial of Oral Everolimus-2 (BOLERO-2) trial in combination with exemestane for patients with advanced metastatic hormone receptor-positive (HR+) breast cancer, who relapsed after initial hormonal manipulation. The study met its primary endpoint of significant improvement in progression free survival (PFS) with a median time to progression of 6.9 months in the combination group versus 2.8 months in exemestane group. Favorable improvements in PFS were reported across all patient subgroups regardless of age, Eastern Cooperative Oncology Group performance status, number of prior therapies, and presence of visceral metastases. Adverse events were mostly mild to moderate in severity and consistent with the known safety profile of everolimus. Major toxicities reported include stomatitis, non-infectious pneumonitis, and hyperglycemia. The purpose of this review is to discuss the role of everolimus as a valuable component in advanced metastatic breast cancer and delineate current strategies to prevent and manage the most common toxicities associated with this combination regimen.

  3. Everolimus and Malignancy after Solid Organ Transplantation: A Clinical Update

    PubMed Central

    De Simone, Paolo

    2016-01-01

    Malignancy after solid organ transplantation remains a major cause of posttransplant mortality. The mammalian target of rapamycin (mTOR) inhibitor class of immunosuppressants exerts various antioncogenic effects, and the mTOR inhibitor everolimus is licensed for the treatment of several solid cancers. In kidney transplantation, evidence from registry studies indicates a lower rate of de novo malignancy under mTOR inhibition, with some potentially supportive data from randomized trials of everolimus. Case reports and small single-center series have suggested that switch to everolimus may be beneficial following diagnosis of posttransplant malignancy, particularly for Kaposi's sarcoma and nonmelanoma skin cancer, but prospective studies are lacking. A systematic review has shown mTOR inhibition to be associated with a significantly lower rate of hepatocellular carcinoma (HCC) recurrence versus standard calcineurin inhibitor therapy. One meta-analysis has concluded that patients with nontransplant HCC experience a low but significant survival benefit under everolimus monotherapy, so far unconfirmed in a transplant population. Data are limited in heart transplantation, although observational data and case reports have indicated that introduction of everolimus is helpful in reducing the recurrence of skin cancers. Overall, it can be concluded that, in certain settings, everolimus appears a promising option to lessen the toll of posttransplant malignancy. PMID:27807479

  4. Everolimus in the management of metastatic neuroendocrine tumours

    PubMed Central

    Chan, David L.; Segelov, Eva; Singh, Simron

    2016-01-01

    Neuroendocrine tumours are increasing in incidence and cause a variety of symptoms. The mammalian target of rapamycin (mTOR) pathway plays a key role in neuroendocrine tumour (NET) pathogenesis, leading to increased lipid synthesis, protein synthesis and cellular growth. Upregulation of this pathway is noted in both hereditary and sporadic NETs. This understanding has led to investigations of mTOR inhibitors as therapy for metastatic NETs. After promising preclinical findings, everolimus, an mTOR inhibitor, was trialled in the RADIANT-1−4 studies on patients with advanced, well differentiated NETs. RADIANT-3 and RADIANT-4 established the efficacy of everolimus in improving progression-free survival (PFS) for metastatic NET of pancreatic, lung and gastrointestinal origin, leading to the US Food and Drug Administration (FDA) approval for its use in tumour control in those settings. Everolimus treatment is generally well tolerated; common adverse events include stomatitis, diarrhoea, rash and hyperglycaemia. Although discontinuation rates are low, many patients may require dose modification to successfully continue therapy. The combination of everolimus with somatostatin analogues (SSAs) (such as octreotide or pasireotide) or other targeted agents such as bevacizumab has not produced additional incremental benefit, and dual biologic therapy is not used widely. Ongoing trials are investigating everolimus compared with chemotherapy, optimal sequencing of therapy and combination of everolimus with radiotherapy. Future research should concentrate on identification of predictive biomarkers for benefit from mTOR therapy and include quality of life as a measure. PMID:28286565

  5. BRAFV600E-dependent Mcl-1 stabilization leads to everolimus resistance in colon cancer cells

    PubMed Central

    He, Kan; Chen, Dongshi; Ruan, Hang; Li, Xiangyun; Tong, Jingshan; Xu, Xiang; Zhang, Lin; Yu, Jian

    2016-01-01

    mTOR activation is commonly caused by oncogenic mutations in RAS/RAF/MAPK and PI3K/AKT pathways, and promotes cancer progression and therapeutic resistance. However, mTOR inhibitors show limited single agent efficacy in patients. mTOR inhibitors suppress tumor cell growth and angiogenesis, and have recently been shown to induce death receptor/FADD-dependent apoptosis in colon cancers. Using a panel of BRAF V600E and WT colorectal cancer cell lines and in vitro selected resistant culture, and xenograft models, we demonstrate here that BRAFV600E confers resistance to mTOR inhibitors. Everolimus treatment disrupts the S6K1-IRS-2/PI3K negative feedback loop, leading to BRAF V600E-dependent activation of ERK and Mcl-1 stabilization in colon cancer cells, which in turn blocks the crosstalk from the death receptor to mitochondria. Co-treatment with inhibitors to Mcl-1, PI3K, RAF or MEK restores mTOR inhibitor-induced apoptosis by antagonizing Mcl-1 or abrogating ERK activation in BRAFV600E cells. Our findings provide a rationale for genotype-guided patient stratification and potential drug combinations to prevent or mitigate undesired activation of survival pathways induced by mTOR inhibitors. PMID:27351224

  6. [Management of Stomatitis Associated with Treatment with Everolimus].

    PubMed

    Ota, Yoshihide; Kurita, Hiroshi; Umeda, Masahiro

    2016-02-01

    Stomatitis is a characteristic adverse event of everolimus and other mTOR inhibitors, and occurs at a high incidence and impairs QOL owing to pain. Most cases of stomatitis are mild to moderate. However, when stomatitis becomes serious, it can interfere with the continuation of medication. Therefore, it is important to place more emphasis on the prevention as well as early detection and treatment. In addition, patient education is also important. The possible occurrence of stomatitis, its signs and symptoms, as well as the importance of oral care need to be thoroughly explained prior to starting treatment. In order to smoothly carry out these measures, it will also be essential that cancer-treating physicians coordinate and collaborate with dentists, nurses, and pharmacists. It is desirable to establish appropriate prevention and management methods on the basis of the results of the Phase III prospective study, Oral Care-BC, currently ongoing in Japan.

  7. Interstitial Lung Disease Associated with mTOR Inhibitors in Solid Organ Transplant Recipients: Results from a Large Phase III Clinical Trial Program of Everolimus and Review of the Literature

    PubMed Central

    Lopez, Patricia

    2014-01-01

    Interstitial lung disease (ILD) has been reported with the use of mammalian target of rapamycin inhibitors (mTORi). The clinical and safety databases of three Phase III trials of everolimus in de novo kidney (A2309), heart (A2310), and liver (H2304) transplant recipients (TxR) were searched using a standardized MedDRA query (SMQ) search for ILD followed by a case-by-case medical evaluation. A literature search was conducted in MEDLINE and EMBASE. Out of the 1,473 de novo TxR receiving everolimus in Phase III trials, everolimus-related ILD was confirmed in six cases (one kidney, four heart, and one liver TxR) representing an incidence of 0.4%. Everolimus was discontinued in three of the four heart TxR, resulting in ILD improvement or resolution. Outcome was fatal in the kidney TxR (in whom everolimus therapy was continued) and in the liver TxR despite everolimus discontinuation. The literature review identified 57 publications on ILD in solid organ TxR receiving everolimus or sirolimus. ILD presented months or years after mTORi initiation and symptoms were nonspecific and insidious. The event was more frequent in patients with a late switch to mTORi. In most cases, ILD was reversed after prompt mTORi discontinuation. ILD induced by mTORi is an uncommon and potentially fatal event warranting early recognition and drug discontinuation. PMID:25580277

  8. Age-Related Neurodegeneration Prevention Through mTOR Inhibition: Potential Mechanisms and Remaining Questions

    PubMed Central

    Jahrling, Jordan B.; Laberge, Remi-Martin

    2016-01-01

    With the global aging population, Alzheimer's disease, Parkinson's disease and mild cognition impairment are increasing in prevalence. The success of rapamycin as an agent to extend lifespan in various organisms, including mice, brings hope that chronic mTOR inhibition could also refrain age-related neurodegeneration. Here we review the evidence suggesting that mTOR inhibition - mainly with rapamycin - is a valid intervention to delay age-related neurodegeneration. We discuss the potential mechanisms by which rapamycin may facilitate neurodegeneration prevention or restoration of cognitive function. We also discuss the known side effects of rapamycin and provide evidence to alleviate exaggerated concerns regarding its wider clinical use. We explore the small molecule alternatives to rapamycin and propose future directions for their development, mainly by exploring the possibility of targeting the downstream effectors of mTOR: S6K1 and especially S6K2. Finally, we discuss the strengths and weaknesses of the models used to determine intervention efficacy for neurodegeneration. We address the difficulties of interpreting data using the common way of investigating the efficacy of interventions to delay/prevent neurodegeneration by observing animal behavior while these animals are under treatment. We propose an experimental design that should isolate the variable of aging in the experimental design and resolve the ambiguity present in recent literature. PMID:26059360

  9. Pelvic lymphangioleiomyomatosis treated successfully with everolimus

    PubMed Central

    Wahid, Sharjil; Chiang, Ping Chia; Luo, Hao Lun; Huang, Shun-Chen; Tsai, Eing-Mei; Chiang, Po Hui

    2017-01-01

    Abstract Background: Lymphangioleiomyomatosis (LAM) is a rare disease affecting young women caused by abnormal proliferation of smooth muscle-like cells (LAM cells) in the lungs and extrapulmonary sites (extrapulmonary LAM). The objective of this case series is to demonstrate marked regression in 2 cases of retroperitoneal LAM after treatment with everolimus, an mTOR inhibitor. Methods: We enrolled 2 cases with large volume, extrapulmonary pelvic LAM, and evaluated them with contrast-enhanced abdominal computed tomographic (CT) scans at presentation and serially during treatment with everolimus. Results were objectively quantified using the Response Evaluation Criteria in Solid Tumors, RECIST, Version 1.1. Results: After 12 to 18 months of treatment with everolimus, both patients showed substantial reduction in the volume of their tumors. The first had about 50% regression of the pelvic LAM and renal angiomyolipoma (AML). The second patient had extensive abdomino-pelvic LAM which after treatment showed complete remission. Both patients have not demonstrated disease progression after nearly 4 and 2 years of follow-up, respectively. Conclusions: This case series demonstrates the enormous value of mTOR inhibitors (specifically everolimus) in the management of extrapulmonary pelvic LAM, of which there is no effective treatment currently available. PMID:28272193

  10. Inhibition of mTOR by apigenin in UVB-irradiated keratinocytes: A new implication of skin cancer prevention.

    PubMed

    Bridgeman, Bryan B; Wang, Pu; Ye, Boping; Pelling, Jill C; Volpert, Olga V; Tong, Xin

    2016-05-01

    Ultraviolet B (UVB) radiation is the major environmental risk factor for developing skin cancer, the most common cancer worldwide, which is characterized by aberrant activation of Akt/mTOR (mammalian target of rapamycin). Importantly, the link between UV irradiation and mTOR signaling has not been fully established. Apigenin is a naturally occurring flavonoid that has been shown to inhibit UV-induced skin cancer. Previously, we have demonstrated that apigenin activates AMP-activated protein kinase (AMPK), which leads to suppression of basal mTOR activity in cultured keratinocytes. Here, we demonstrated that apigenin inhibited UVB-induced mTOR activation, cell proliferation and cell cycle progression in mouse skin and in mouse epidermal keratinocytes. Interestingly, UVB induced mTOR signaling via PI3K/Akt pathway, however, the inhibition of UVB-induced mTOR signaling by apigenin was not Akt-dependent. Instead, it was driven by AMPK activation. In addition, mTOR inhibition by apigenin in keratinocytes enhanced autophagy, which was responsible, at least in part, for the decreased proliferation in keratinocytes. In contrast, apigenin did not alter UVB-induced apoptosis. Taken together, our results indicate the important role of mTOR inhibition in UVB protection by apigenin, and provide a new target and strategy for better prevention of UV-induced skin cancer.

  11. Symptomatic Control of Neuroendocrine Tumours with Everolimus.

    PubMed

    Bainbridge, Hannah E; Larbi, Emmanuel; Middleton, Gary

    2015-12-01

    Everolimus, a mammalian target of rapamycin (mTOR) inhibitor, increases progression-free survival in patients with advanced neuroendocrine tumours. Patients with neuroendocrine tumours and symptomatic carcinoid have inferior health-related quality of life than those without symptoms. We aimed to evaluate the effect of everolimus on symptomatic control of neuroendocrine tumours. Fifteen patients with metastatic neuroendocrine disease pre-treated with depot octreotide received combination everolimus and octreotide (midgut = 8, pancreatic = 3, other = 4). Reasons for initiation of everolimus were progressive disease (PD) by response evaluation criteria in solid tumours (n = 5), worsening syndromic symptomology (n = 5), or both (n = 5). Symptomatic and objective response and toxicity were evaluated using standard criteria. 7/10 patients who were syndromic had improvements in symptomology, with a mean duration of symptom control 13.9 months (range 1-39). All 10 symptomatic patients had non pancreatic neuroendocrine (pNET) primaries, and with everolimus, 6/10 had reduced stool frequency, 3/7 had a reduction of asthenia, and 5/7 had reduced frequency and severity of flushing. Sixty percent of patients experienced any grade toxicities, including the following: 40% grade 1/2 stomatitis, 7% grade 3/4 stomatitis, 20% grade 1/2 rash, 13% diarrhoea, and one case of pneumonitis. In this cohort of 15 patients, we demonstrated that 70% of non pNET individuals with common carcinoid syndrome symptoms resistant to depot octreotide had improvement in these symptoms on institution of everolimus, with meaningful durations of symptom control. Although this data is observational, to our knowledge, this represents the largest analysis of carcinoid syndrome control with combined everolimus and octreotide.

  12. Combined treatment by octreotide and everolimus: Octreotide enhances inhibitory effect of everolimus in aggressive meningiomas.

    PubMed

    Graillon, Thomas; Defilles, Céline; Mohamed, Amira; Lisbonis, Christophe; Germanetti, Anne-Laure; Chinot, Olivier; Figarella-Branger, Dominique; Roche, Pierre-Hugues; Adetchessi, Tarek; Fuentes, Stéphane; Metellus, Philippe; Dufour, Henry; Enjalbert, Alain; Barlier, Anne

    2015-08-01

    Treatment for recurrent and aggressive meningiomas remains an unmet medical need in neuro-oncology, and chemotherapy exhibits limited clinical activity, if any. Merlin expression, encoded by the NF2 gene, is lost in a majority of meningiomas, and merlin is a negative regulator of mTORC1. The sst2 somatostatin receptor, targeted by octreotide, is highly expressed in meningiomas. To investigate new therapeutic strategies, we evaluated the activity of everolimus (mTOR inhibitor), BKM-120 and BEZ-235 (new Pi3K/Akt/mTOR inhibitors), octreotide and a combined treatment (octreotide plus everolimus), on cell proliferation, signaling pathways, and cell cycle proteins, respectively. The in vitro study was conducted on human meningioma primary cells extracted from fresh tumors, allowing the assessment of somatostatin analogs at the concentration levels used in patients. The results were correlated to WHO grades. Further, everolimus decreased cell viability of human meningiomas, but concomitantly, induced Akt activation, reducing the antiproliferative effect of the drug. The new Pi3K inhibitors were not more active than everolimus alone, limiting their clinical relevance. In contrast, a clear cooperative inhibitory effect of octreotide and everolimus was observed on cell proliferation in all tested meningiomas, including WHO grades II-III. Octreotide not only reversed everolimus-induced Akt phosphorylation but also displayed additive and complementary effects with everolimus on downstream proteins involved in translation (4EB-P1), and controlling cell cycle (p27Kip1 and cyclin D1). We have demonstrated a co-operative action between everolimus and octreotide on cell proliferation in human meningiomas, including aggressive ones, establishing the basis for a clinical trial.

  13. Everolimus exhibits anti-tumorigenic activity in obesity-induced ovarian cancer

    PubMed Central

    Guo, Hui; Zhong, Yan; Jackson, Amanda L.; Clark, Leslie H.; Kilgore, Josh; Zhang, Lu; Han, Jianjun; Sheng, Xiugui; Gilliam, Timothy P.; Gehrig, Paola A.; Zhou, Chunxiao; Bae, Victoria L.

    2016-01-01

    Everolimus inhibits mTOR kinase activity and its downstream targets by acting on mTORC1 and has anti-tumorigenic activity in ovarian cancer. Clinical and epidemiologic data find that obesity is associated with worse outcomes in ovarian cancer. In addition, obesity leads to hyperactivation of the mTOR pathway in epithelial tissues, suggesting that mTOR inhibitors may be a logical choice for treatment in obesity-driven cancers. However, it remains unclear if obesity impacts the effect of everolimus on tumor growth in ovarian cancer. The present study was aimed at evaluating the effects of everolimus on cytotoxicity, cell metabolism, apoptosis, cell cycle, cell stress and invasion in human ovarian cancer cells. A genetically engineered mouse model of serous ovarian cancer fed a high fat diet or low fat diet allowed further investigation into the inter-relationship between everolimus and obesity in vivo. Everolimus significantly inhibited cellular proliferation, induced cell cycle G1 arrest and apoptosis, reduced invasion and caused cellular stress via inhibition of mTOR pathways in vitro. Hypoglycemic conditions enhanced the sensitivity of cells to everolimus through the disruption of glycolysis. Moreover, everolimus was found to inhibit ovarian tumor growth in both obese and lean mice. This reduction coincided with a decrease in expression of Ki-67 and phosphorylated-S6, as well as an increase in cleaved caspase 3 and phosphorylated-AKT. Metabolite profiling revealed that everolimus was able to alter tumor metabolism through different metabolic pathways in the obese and lean mice. Our findings support that everolimus may be a promising therapeutic agent for obesity-driven ovarian cancers. PMID:26959121

  14. Potential therapeutic effects of the MTOR inhibitors for preventing ageing and progeria‐related disorders

    PubMed Central

    Evangelisti, Camilla; Cenni, Vittoria

    2016-01-01

    The mammalian target of rapamycin (mTOR) pathway is an highly conserved signal transduction axis involved in many cellular processes, such as cell growth, survival, transcription, translation, apoptosis, metabolism, motility and autophagy. Recently, this signalling pathway has come to the attention of the scientific community owing to the unexpected finding that inhibition of mTOR by rapamycin, an antibiotic with immunosuppressant and chemotherapeutic properties, extends lifespan in diverse animal models. Moreover, rapamycin has been reported to rescue the cellular phenotype in a progeroid syndrome [Hutchinson–Gilford Progeria syndrome (HGPS)] that recapitulates most of the traits of physiological ageing. The promising perspectives raised by these results warrant a better understanding of mTOR signalling and the potential applications of mTOR inhibitors to counteract ageing‐associated diseases and increase longevity. This review is focused on these issues. PMID:26952863

  15. SiRNA Targeting mTOR Effectively Prevents the Proliferation and Migration of Human Lens Epithelial Cells

    PubMed Central

    Zhang, Chunmei; Liu, Jingjing; Jin, Na; Zhang, Guiming; Xi, Yahui; Liu, Hongling

    2016-01-01

    Posterior capsule opacification (PCO) is the most common complication that causes visual decrease after extracapsular cataract surgery. The primary cause of PCO formation is the proliferation of the residual lens epithelial cells (LECs). The mammalian target of rapamycin (mTOR) plays an important role in the growth and migration of LECs. In the current study, we used small interfering RNA (siRNA) to specifically attenuate mTOR in human lens epithelial B3 cells (HLE B3). We aimed to examine the effect of mTOR-siRNA on the proliferation, migration and epithelial-to-mesenchymal transition (EMT) of HLE B3 cells and explore the underlying mechanisms. The mTOR-siRNA was transfected into HLE B3 cells using lipofectamine 2000. The mRNA and protein levels of mTOR were examined to confirm the efficiency of mTOR-siRNA. The levels of mRNA and protein as well as the activity of mTOR down-stream effectors p70 ribosomal protein S6 kinase (p70S6K) and protein kinase B (PKB, AKT) were examined using real-time PCR or Western blot, respectively. The cell proliferation was determined using cell counting kit (CCK) 8 and cell growth curve assay. The cell migration was examined using Transwell system and Scratch assay. MTOR-siRNA effectively eliminated mTOR mRNA and protein. The proliferation and migration were significantly suppressed by mTOR-siRNA transfection. mTOR-siRNA reduced the mRNA of p70S6K and AKT in a time-dependent manner. Furthermore, the phosphorylation of p70S6K and AKT was decreased by mTOR-siRNA. MTOR-siRNA also eliminated the formation of mTORC1 and mTORC2 protein complex and blocked the transforming growth factor (TGF)-β-induced EMT. Our results suggested that mTOR-siRNA could effectively inhibit the proliferation, migration and EMT of HLE B3 cells through the inhibition of p70S6K and AKT. These results indicated that mTOR-siRNA might be an effective agent inhibiting HLE cells growth and EMT following cataract surgery and provide an alternative therapy for preventing

  16. Targeting Focal Adhesion Kinase and Resistance to mTOR Inhibition in Pancreatic Neuroendocrine Tumors

    PubMed Central

    François, Rony A.; Maeng, Kyungah; Nawab, Akbar; Kaye, Frederic J.; Hochwald, Steven N.; Zajac-Kaye, Maria

    2015-01-01

    Background: Focal adhesion kinase (FAK) mediates survival of normal pancreatic islets through activation of AKT. Upon malignant transformation of islet cells into pancreatic neuroendocrine tumors (PanNETs), AKT is frequently overexpressed and mutations in the AKT/mTOR pathway are detected. Because mTOR inhibitors rarely induce PanNET tumor regression, partly because of feedback activation of AKT, novel combination strategies are needed to target FAK/AKT/mTOR signaling. Methods: We characterized the activation of FAK in PanNETs using immunohistochemistry and Western blot analysis and tested the FAK inhibitor PF-04554878 in human PanNET cells in vitro and in vivo (at least three mice per group). In addition, we evaluated the effect of combined FAK and mTOR inhibition on PanNET viability and apoptosis. All statistical tests were two-sided. Results: We found that FAK is overexpressed and hyperphosphorylated in human PanNETs and that PF-04554878 strongly inhibited FAK (Tyr397) autophosphorylation in a dose-dependent manner. We found that PF-04554878 inhibited cell proliferation and clonogenicity and induced apoptosis in PanNET cells. Moreover, oral administration of PF-04554878 statistically significantly reduced tumor growth in a patient-derived xenograft model of PanNET (P = .02) and in a human PanNET xenograft model of peritoneal carcinomatosis (P = .03). Importantly, PF-04554878 synergized with the mTOR inhibitor everolimus by preventing feedback AKT activation. Conclusions: We demonstrate for the first time that FAK is overexpressed in PanNETs and that inhibition of FAK activity induces apoptosis and inhibits PanNET proliferation. We found that the novel FAK inhibitor PF-04554878 synergizes with everolimus, a US Food and Drug Administration–approved agent for PanNETs. Our findings warrant the clinical investigation of combined FAK and mTOR inhibition in PanNETs. PMID:25971297

  17. Use of Everolimus After Multivisceral Transplantation: A Report of Two Cases.

    PubMed

    Rao, B; Segovia, M C; Kazimi, M; Parekh, R; Raoufi, M; Jafri, S-M

    2016-03-01

    Inhibitors of mechanistic target of rapamycin are used in solid organ transplant procedures to avoid calcineurin inhibitor complications, including nephrotoxicity and malignancy. We present 2 cases of multivisceral transplantation for neuroendocrine tumor (NET) for which everolimus was implemented for its potential to prevent NET recurrence as well as preserve renal function. The first case was complicated by NET recurrence in the liver before initiation of everolimus. After initiation of everolimus, the patient developed a ventral hernia and elevated aminotransferase levels with nonspecific biopsy findings. The second case was complicated by cytomegalovirus infection with elevated everolimus trough levels as well as acute cellular rejection. Everolimus was reinitiated in both cases in addition to decreasing the dosage of tacrolimus, and there were no further complications. Everolimus was beneficial in stabilizing renal function in both patients and has the theoretical potential to prevent recurrence of NET.

  18. Long-term acquired everolimus resistance in pancreatic neuroendocrine tumours can be overcome with novel PI3K-AKT-mTOR inhibitors

    PubMed Central

    Vandamme, Timon; Beyens, Matthias; de Beeck, Ken Op; Dogan, Fadime; van Koetsveld, Peter M; Pauwels, Patrick; Mortier, Geert; Vangestel, Christel; de Herder, Wouter; Van Camp, Guy; Peeters, Marc; Hofland, Leo J

    2016-01-01

    Background: The mTOR-inhibitor everolimus improves progression-free survival in advanced pancreatic neuroendocrine tumours (PNETs). However, adaptive resistance to mTOR inhibition is described. Methods: QGP-1 and BON-1, two human PNET cell lines, were cultured with increasing concentrations of everolimus up to 22 weeks to reach a dose of 1 μM everolimus, respectively, 1000-fold and 250-fold initial IC50. Using total DNA content as a measure of cell number, growth inhibitory dose–response curves of everolimus were determined at the end of resistance induction and over time after everolimus withdrawal. Response to ATP-competitive mTOR inhibitors OSI-027 and AZD2014, and PI3K-mTOR inhibitor NVP-BEZ235 was studied. Gene expression of 10 PI3K-Akt-mTOR pathway-related genes was evaluated using quantitative real-time PCR (RT–qPCR). Results: Long-term everolimus-treated BON-1/R and QGP-1/R showed a significant reduction in everolimus sensitivity. During a drug holiday, gradual return of everolimus sensitivity in BON-1/R and QGP-1/R led to complete reversal of resistance after 10–12 weeks. Treatment with AZD2014, OSI-027 and NVP-BEZ235 had an inhibitory effect on cell proliferation in both sensitive and resistant cell lines. Gene expression in BON-1/R revealed downregulation of MTOR, RICTOR, RAPTOR, AKT and HIF1A, whereas 4EBP1 was upregulated. In QGP-1/R, a downregulation of HIF1A and an upregulation of ERK2 were observed. Conclusions: Long-term everolimus resistance was induced in two human PNET cell lines. Novel PI3K-AKT-mTOR pathway-targeting drugs can overcome everolimus resistance. Differential gene expression profiles suggest different mechanisms of everolimus resistance in BON-1 and QGP-1. PMID:26978006

  19. Evaluation of the impact of the cancer therapy everolimus on the central nervous system in mice.

    PubMed

    Dubois, Martine; Le Joncour, Vadim; Tonon, Marie-Christine; Anouar, Youssef; Proust, François; Morin, Fabrice; Gandolfo, Pierrick; Joly, Florence; Hilber, Pascal; Castel, Hélène

    2014-01-01

    Cancer and treatments may induce cognitive impairments in cancer patients, and the causal link between chemotherapy and cognitive dysfunctions was recently validated in animal models. New cancer targeted therapies have become widely used, and their impact on brain functions and quality of life needs to be explored. We evaluated the impact of everolimus, an anticancer agent targeting the mTOR pathway, on cognitive functions, cerebral metabolism, and hippocampal cell proliferation/vascular density in mice. Adult mice received everolimus daily for 2 weeks, and behavioral tests were performed from 1 week after the last treatment. Everolimus-treated mice displayed a marked reduction in weight gain from the last day of the treatment period. Ex vivo analysis showed altered cytochrome oxidase activity in selective cerebral regions involved in energy balance, food intake, reward, learning and memory modulation, sleep/wake cycle regulation, and arousal. Like chemotherapy, everolimus did not alter emotional reactivity, learning and memory performances, but in contrast to chemotherapy, did not affect behavioral flexibility or reactivity to novelty. In vivo hippocampal neural cell proliferation and vascular density were also unchanged after everolimus treatments. In conclusion, two weeks daily everolimus treatment at the clinical dose did not evoke alteration of cognitive performances evaluated in hippocampal- and prefrontal cortex-dependent tasks that would persist at one to four weeks after the end of the treatment completion. However, acute everolimus treatment caused selective CO modifications without altering the mTOR effector P70S6 kinase in cerebral regions involved in feeding behavior and/or the sleep/wake cycle, at least in part under control of the solitary nucleus and the parasubthalamic region of the hypothalamus. Thus, this area may represent a key target for everolimus-mediating peripheral modifications, which has been previously associated with symptoms such as

  20. [Level of evidence for therapeutic drug monitoring of everolimus].

    PubMed

    Goirand, Françoise; Royer, Bernard; Hulin, Anne; Saint-Marcoux, Franck

    2011-01-01

    Everolimus has proven efficacy for prevention of rejection in adult de novo renal and cardiac transplant recipient in combination with ciclosporine and corticosteroids. Therapeutic drug monitoring (TDM) with target trough concentration (C0) value from 3 to 8 µg/L has been proposed. Through a systematic review of the literature, this work explored a level of recommendation for this TDM. Everolimus exhibits both wide interindividual pharmacokinetic variability and poor relationship between dose and exposure. A good relationship has been reported between C0 values and global exposure to the drug (i.e. AUC). Although C0 > 3 µg/L has been associated with a decreased incidence of rejection, the upper limit of 8 µg/L has never been formally validated. No clinical trial testing other exposure indices or comparing efficacy and/or toxicity of everolimus therapy with and without TDM has been published so far. Consequently the level of recommendation for everolimus monitoring is "recommended".

  1. Pivotal Role of mTOR Signaling in Hepatocellular Carcinoma

    PubMed Central

    Villanueva, Augusto; Chiang, Derek Y.; Newell, Pippa; Peix, Judit; Thung, Swan; Alsinet, Clara; Tovar, Victoria; Roayaie, Sasan; Minguez, Beatriz; Sole, Manel; Battiston, Carlo; van Laarhoven, Stijn; Fiel, Maria I; Di Feo, Analisa; Hoshida, Yujin; Yea, Steven; Toffanin, Sara; Ramos, Alex; Martignetti, John A.; Mazzaferro, Vincenzo; Bruix, Jordi; Waxman, Samuel; Schwartz, Myron; Meyerson, Matthew; Friedman, Scott L.; Llovet, Josep M.

    2008-01-01

    BACKGROUND The advent of targeted therapies in hepatocellular carcinoma (HCC) has underscored the importance of pathway characterization to identify novel molecular targets for treatment. Based on its role in cell growth and differentiation, we evaluated mTOR signaling activation in human HCC, as well as the anti-tumoral effect of a dual-level blockade of the mTOR pathway. METHODS The mTOR pathway was assessed using integrated data from mutation analysis (direct sequencing), DNA copy number changes (SNP-array), mRNA levels (qRT-PCR and gene expression microarray), and protein activation (immunostaining) in 351 human samples, including HCC (n=314), and non-tumoral tissue (n=37). Effects of dual blockade of mTOR signaling using a rapamycin analog (everolimus) and an EGFR/VEGFR inhibitor (AEE788) were evaluated in liver cancer cell lines, and in a tumor xenograft model. RESULTS Aberrant mTOR signaling (phosphorylated-RPS6) was present in half of the cases, associated with IGF pathway activation, EGF upregulation, and PTEN dysregulation. PTEN and PI3KCA-B mutations were rare events. Chromosomal gains in RICTOR (25% of patients) and positive pRPS6 staining correlated with recurrence. RICTOR-specific siRNA downregulation reduced tumor cell viability in vitro. Blockage of mTOR signaling with everolimus in vitro and in a xenograft model decelerated tumor growth and increased survival. This effect was enhanced in vivo after EGFR blockade. CONCLUSIONS MTOR signaling has a critical role in the pathogenesis of HCC, with evidence for the role of RICTOR in tumor oncogenesis. MTOR blockade with everolimus is effective in vivo. These findings establish a rationale for targeting mTOR pathway in clinical trials in HCC. PMID:18929564

  2. Combination of everolimus and tacrolimus: a potentially effective regimen for recalcitrant psoriasis.

    PubMed

    Wei, Kai-Che; Lai, Ping-Chin

    2015-01-01

    Severe forms of psoriasis that are refractory to conventional therapies are often difficult to manage. The mammalian target of rapamycin (mTOR) inhibitors potentially have versatile effects toward putative psoriatic pathologic pathways. Therefore, mTOR inhibitors may offer a range of new therapeutic options for patients with psoriasis. We describe a 55-year-old male renal transplant patient with refractory psoriasis. We adjusted his antirejection regimen and put him on everolimus (Certican(®); Novartis, Basel, Switzerland; a semisynthetic macrolide, belonging to the mTOR inhibitors family) with low-dose tacrolimus. This combination regimen maintained his graft function and successfully controlled his psoriasis. His skin lesions never recurred in the next 18 months. To our knowledge, this is the first report showing that the combination of everolimus and tacrolimus could be used to treat recalcitrant psoriasis. The relative benefit-risk profiles of such therapies worth further investigation.

  3. Everolimus: in patients with subependymal giant cell astrocytoma associated with tuberous sclerosis complex.

    PubMed

    Curran, Monique P

    2012-02-01

    Everolimus is an orally administered inhibitor of the mammalian target of rapamycin (mTOR). Everolimus (starting dosage 3.0 mg/m(2)) was associated with a significant reduction in the volume of the largest subependymal giant cell astrocytoma (SEGA) in 28 patients aged ≥3 years with tuberous sclerosis complex (TSC) in a phase II trial (C2485). At 6 months, 32% of patients treated with everolimus had a ≥50% reduction in the volume of their largest SEGA lesion (assessed via an independent central radiology review); 75% had a ≥30% reduction. No patients developed new lesions. During the extension phase of this trial (median duration 34 months), the reduction in SEGA volume was maintained, with no everolimus recipient requiring surgery or other therapy for SEGA or hydrocephalus. In a phase III trial (EXIST-1) in 117 patients with SEGA associated with TSC, 35% of everolimus recipients (starting dosage 4.5 mg/m(2)) versus none of the placebo recipients (p < 0.0001) had an overall response (a reduction in the sum of all target SEGA volumes of ≥50% relative to baseline, nonworsening of non-target SEGA lesions, no new SEGA lesions, and no new/worsening hydrocephalus). Everolimus was generally well tolerated in patients with SEGA associated with TSC; most drug-related adverse reactions were mild to moderate in severity.

  4. Everolimus in the management of metastatic renal cell carcinoma: an evidence-based review of its place in therapy

    PubMed Central

    Buti, Sebastiano; Leonetti, Alessandro; Dallatomasina, Alice; Bersanelli, Melissa

    2016-01-01

    Introduction Renal cell carcinoma (RCC) is the most common type of kidney cancer in adults, and its pathogenesis is strictly related to altered cellular response to hypoxia, in which mTOR signaling pathway is implicated. Everolimus, an mTOR serine/threonine kinase inhibitor, represents a therapeutic option for the treatment of advanced RCC. Aim The objective of this article is to review the evidence for the treatment of metastatic RCC with everolimus. Evidence review Everolimus was approved for second- and third-line therapy in patients with advanced RCC according to the results of a Phase III pivotal trial that demonstrated a benefit in median progression-free survival of ~2 months compared to placebo after failure of previous lines of therapy, of which at least one was an anti-VEGFR tyrosine kinase inhibitor (TKI). The role of this drug in first-line setting has been investigated in Phase II trials, with no significant clinical benefit, even in combination with bevacizumab. Everolimus activity in non-clear cell RCC is supported by two randomized Phase II trials that confirmed the benefit in second-line setting but not in first line. Recently, two randomized Phase III trials (METEOR and CheckMate 025) demonstrated the inferiority of everolimus in second-line setting compared to the TKI cabozantinib and to the immune checkpoint inhibitor nivolumab, respectively. Moreover, a recent Phase II study demonstrated a significant benefit for the second-line combination treatment with everolimus plus lenvatinib (a novel TKI) in terms of progression-free survival and overall survival compared to the single-agent everolimus. Basing on preclinical data, the main downstream effectors of mTOR cascade, S6RP and its phosphorylated form, could be good predictive biomarkers of response to everolimus. The safety profile of the drug is favorable, with a good cost-effectiveness compared to second-line sorafenib or axitinib, and no significant impact on the quality of life of treated

  5. mTOR inhibitors in urinary bladder cancer.

    PubMed

    Pinto-Leite, R; Arantes-Rodrigues, R; Sousa, Nuno; Oliveira, P A; Santos, L

    2016-09-01

    Despite the great scientific advances that have been made in cancer treatment, there is still much to do, particularly with regard to urinary bladder cancer. Some of the drugs used in urinary bladder cancer treatment have been in use for more than 30 years and show reduced effectiveness and high recurrence rates. There have been several attempts to find new and more effective drugs, to be used alone or in combination with the drugs already in use, in order to overcome this situation.The biologically important mammalian target of rapamycin (mTOR) pathway is altered in cancer and mTOR inhibitors have raised many expectations as potentially important anticancer drugs. In this article, the authors will review the mTOR pathway and present their experiences of the use of some mTOR inhibitors, sirolimus, everolimus and temsirolimus, in isolation and in conjunction with non-mTOR inhibitors cisplatin and gemcitabine, on urinary bladder tumour cell lines. The non-muscle-invasive cell line, 5637, is the only one that exhibits a small alteration in the mTOR and AKT phosphorylation after rapalogs exposure. Also, there was a small inhibition of cell proliferation. With gemcitabine plus everolimus or temsirolimus, the results were encouraging as a more effective response was noticed with both combinations, especially in the 5637 and T24 cell lines. Cisplatin associated with everolimus or temsirolimus also gave promising results, as an antiproliferative effect was observed when the drugs were associated, in particular on the 5637 and HT1376 cell lines. Everolimus or temsirolimus in conjunction with gemcitabine or cisplatin could have an important role to play in urinary bladder cancer treatment, depending on the tumour grading.

  6. Ablation of PGC1 beta prevents mTOR dependent endoplasmic reticulum stress response

    PubMed Central

    Camacho, Alberto; Rodriguez-Cuenca, Sergio; Blount, Margaret; Prieur, Xavier; Barbarroja, Nuria; Fuller, Maria; Hardingham, Giles E.; Vidal-Puig, Antonio

    2012-01-01

    Mitochondria dysfunction contributes to the pathophysiology of obesity, diabetes, neurodegeneration and ageing. The peroxisome proliferator-activated receptor-gamma coactivator-1β (PGC-1β) coordinates mitochondrial biogenesis and function as well as fatty acid metabolism. It has been suggested that endoplasmic reticulum (ER) stress may be one of the mechanisms linking mitochondrial dysfunction and these pathologies. Here we investigate whether PGC-1β ablation affects the ER stress response induced by specific nutritional and pharmacological challenges in the CNS. By using flow cytometry, western blot, real time PCR and several pharmacological and nutritional interventions in PGC-1β knock out and WT mice, we confirmed that PGC-1β coordinates mitochondria function in brain and reported for the first time that a) ablation of PGC-1β is associated with constitutive activation of mTORC1 pathway associated with increased basal GRP78 protein levels in hypothalamus and cortex of animals fed chow diet; and b) in animals fed chronically with high fat diet (HFD) or high protein diet (HPD), we observed a failure to appropriately induce ER stress response in the absence of PGC-1β, associated with an increase in mTOR pathway phosphorylation. This contrasted with the appropriate upregulation of ER stress response observed in wild type littermates. Additionally, inefficient in vitro induction of ER stress by thapsigargin seems result in apoptotic neuronal cell death in PGC-1β KO. Our data indicate that PGC-1β is required for a neuronal ER response to nutritional stress imposed by HFD and HPD diets and that genetic ablation of PGC-1β might increase the susceptibility to neuronal damage and cell death. PMID:22771762

  7. Effects of preset sequential administrations of sunitinib and everolimus on tumour differentiation in Caki-1 renal cell carcinoma

    PubMed Central

    Santos, C D; Tijeras-Raballand, A; Serova, M; Sebbagh, S; Slimane, K; Faivre, S; de Gramont, A; Raymond, E

    2015-01-01

    Background: Sunitinib (VEGFR/PDGFR inhibitor) and everolimus (mTOR inhibitor) are both approved for advanced renal cell carcinoma (RCC) as first-line and second-line therapy, respectively. In the clinics, sunitinib treatment is limited by the emergence of acquired resistance, leading to a switch to second-line treatment at progression, often based on everolimus. No data have been yet generated on programmed alternating sequential strategies combining alternative use of sunitinib and everolimus before progression. Such strategy is expected to delay the emergence of acquired resistance and improve tumour control. The aim of our study was to assess the changes in tumours induced by three different sequences administration of sunitinib and everolimus. Methods: In human Caki-1 RCC xenograft model, sunitinib was alternated with everolimus every week, every 2 weeks, or every 3 weeks. Effects on necrosis, hypoxia, angiogenesis, and EMT status were assessed by immunohisochemistry and immunofluorescence. Results: Sunitinib and everolimus programmed sequential regimens before progression yielded longer median time to tumour progression than sunitinib and everolimus monotherapies. In each group of treatment, tumour growth control was associated with inhibition of mTOR pathway and changes from a mesenchymal towards an epithelial phenotype, with a decrease in vimentin and an increase in E-cadherin expression. The sequential combinations of these two agents in a RCC mouse clinical trial induced antiangiogenic effects, leading to tumour necrosis. Conclusions: In summary, our study showed that alternate sequence of sunitinib and everolimus mitigated the development of mesenchymal phenotype compared with sunitinib as single agent. PMID:25422908

  8. Everolimus and pancreatic neuroendocrine tumors (PNETs): Activity, resistance and how to overcome it.

    PubMed

    Capozzi, Monica; Caterina, Ieranò; De Divitiis, Chiara; von Arx, Claudia; Maiolino, Piera; Tatangelo, Fabiana; Cavalcanti, Ernesta; Di Girolamo, Elena; Iaffaioli, Rosario Vincenzo; Scala, Stefania; Tafuto, Salvatore

    2015-09-01

    Neuroendocrine tumors (NET) are rare malignancies, with the most common site of origin being from the gastrointestinal tract, particularly the pancreas, small bowel and appendix. Pancreatic neuroendocrine tumors (PNETs) can be functional, hormone secreting tumors, and can have distinctive symptoms leading to the diagnosis. In contrast nonfunctional tumors, the majority of PNETs, usually present later either incidentally or due to tumor bulk symptoms. Currently Everolimus, an inhibitor of mammalian target of rapamycin (mTOR), is the most promising drug for patients with unresectable, metastatic disease, in progressive well-differentiated PNETs and many studies are ongoing to demonstrate its effects on the other neuroendocrine histotipes. Food and Drug Administration (FDA) and European Medicines Agency (EMA) registered Everolimus in advanced/metastatic breast cancer, in advanced/metastatic renal cell carcinoma and in well/moderately differentiated pancreatic neuroendocrine tumors. Nevertheless only a subset of patients respond to the therapy due to the development of drug resistance. Thus the powerful Everolimus antitumor activity have prompted extensive efforts to overcome drug resistance and to maximize clinical benefit. In this review we aim to summarize current knowledge on mechanisms of Everolimus and other mTOR inhibitors molecules resistance with the intent to overcome it.

  9. [The drug of the month: everolimus (Afinitor) for the treatment of metastatic breast cancer].

    PubMed

    Jerusalem, G; Rorive, A; Collignon, J

    2014-09-01

    Sequential endocrine treatments are recommended for estrogen receptor (ER) positive human epidermal growth factor receptor 2 (HER 2) negative metastatic breast cancers except in the case of symptomatic visceral disease. However, patients who suffer from disease progression while receiving a non-steroidal aromatase inhibitor (NSAI) have a very poor prognosis with standard endocrine therapy alone. Recently, based onthe results of the BOLERO 2 trial, the mammalian target of rapamycin (mTOR) inhibitor everolimus, combined with exemestane, a steroidal aromatase inhibitor, has been approved in Europe and the US for patients suffering from ER positive HER2 negative advanced breast cancer previously treated by a NSAI. The median progression-free survival (PFS) increased from 3.2 to 7.8 months in patients receiving everolimus and exemestane compared to placebo and exemestane. The magnitude of benefit was consistent in all pre-specified subgroups. Side effects were manageable and the quality of life was at least maintained. Everolimus has also beenrecently studied in HER2 positive locally advanced or metastatic disease in heavily pretreated patients (BOLERO 3 trial). This trial met its primary endpoint. The median PFS was increased in patients receiving trastuzumab, vinorelbine and everolimus compared to patients receiving trastuzumab, vinorelbine and placebo. We review pharmacological data and side effects of the drug. We also review the most important clinical trials leading to reimbursement of everolimus in metastatic breast cancer.

  10. Antagonistic effects of chloroquine on autophagy occurrence potentiate the anticancer effects of everolimus on renal cancer cells.

    PubMed

    Grimaldi, A; Santini, D; Zappavigna, S; Lombardi, A; Misso, G; Boccellino, M; Desiderio, V; Vitiello, P P; Di Lorenzo, G; Zoccoli, A; Pantano, F; Caraglia, M

    2015-01-01

    Renal cell carcinoma is an aggressive disease often asymptomatic and weakly chemo-radiosensitive. Currently, new biologic drugs are used among which everolimus, an mTOR inhibitor, that has been approved for second-line therapy. Since mTOR is involved in the control of autophagy, its antitumor capacity is often limited. In this view, chloroquine, a 4-alkylamino substituted quinoline family member, is an autophagy inhibitor that blocks the fusion of autophagosomes and lysosomes. In the present study, we evaluated the effects of everolimus alone or in combination with chloroquine on renal cancer cell viability and verified possible synergism. Our results demonstrate that renal cancer cells are differently sensitive to everolimus and chloroquine and the pharmacological combination everolimus/chloroquine was strongly synergistic inducing cell viability inhibition. In details, the pharmacological synergism occurs when chloroquine is administered before everolimus. In addition, we found a flow autophagic block and shift of death mechanisms to apoptosis. This event was associated with decrease of Beclin-1/Bcl(-)2 complex and parallel reduction of anti-apoptotic protein Bcl(-)2 in combined treatment. At last, we found that the enhancement of apoptosis induced by drug combination occurs through the intrinsic mitochondrial apoptotic pathway activation, while the extrinsic pathway is involved only partly following its activation by chloroquine. These results provide the basis for new therapeutic strategies for the treatment of renal cell carcinoma after appropriate clinical trial.

  11. Antagonistic effects of chloroquine on autophagy occurrence potentiate the anticancer effects of everolimus on renal cancer cells

    PubMed Central

    Grimaldi, A; Santini, D; Zappavigna, S; Lombardi, A; Misso, G; Boccellino, M; Desiderio, V; Vitiello, P P; Di Lorenzo, G; Zoccoli, A; Pantano, F; Caraglia, M

    2015-01-01

    Renal cell carcinoma is an aggressive disease often asymptomatic and weakly chemo-radiosensitive. Currently, new biologic drugs are used among which everolimus, an mTOR inhibitor, that has been approved for second-line therapy. Since mTOR is involved in the control of autophagy, its antitumor capacity is often limited. In this view, chloroquine, a 4-alkylamino substituted quinoline family member, is an autophagy inhibitor that blocks the fusion of autophagosomes and lysosomes. In the present study, we evaluated the effects of everolimus alone or in combination with chloroquine on renal cancer cell viability and verified possible synergism. Our results demonstrate that renal cancer cells are differently sensitive to everolimus and chloroquine and the pharmacological combination everolimus/chloroquine was strongly synergistic inducing cell viability inhibition. In details, the pharmacological synergism occurs when chloroquine is administered before everolimus. In addition, we found a flow autophagic block and shift of death mechanisms to apoptosis. This event was associated with decrease of Beclin-1/Bcl-2 complex and parallel reduction of anti-apoptotic protein Bcl-2 in combined treatment. At last, we found that the enhancement of apoptosis induced by drug combination occurs through the intrinsic mitochondrial apoptotic pathway activation, while the extrinsic pathway is involved only partly following its activation by chloroquine. These results provide the basis for new therapeutic strategies for the treatment of renal cell carcinoma after appropriate clinical trial. PMID:25866016

  12. mTOR function and therapeutic targeting in breast cancer

    PubMed Central

    Hare, Stephen H; Harvey, Amanda J

    2017-01-01

    The mTOR pathway was discovered in the late 1970s after the compound and natural inhibitor of mTOR, rapamycin was isolated from the bacterium Streptomyces hygroscopicus. mTOR is serine/threonine kinase belonging to the phosphoinositide 3-kinase related kinase (PIKK) family. It forms two distinct complexes; mTORC1 and mTORC2. mTORC1 has a key role in regulating protein synthesis and autophagy whilst mTORC2 is involved in regulating kinases of the AGC family. mTOR signaling is often over active in multiple cancer types including breast cancer. This can involve mutations in mTOR itself but more commonly, in breast cancer, this is related to an increase in activity of ErbB family receptors or alterations and mutations of PI3K signaling. Rapamycin and its analogues (rapalogues) bind to the intercellular receptor FKBP12, and then predominantly inhibit mTORC1 signaling via an allosteric mechanism. Research has shown that inhibition of mTOR is a useful strategy in tackling cancers, with it acting to slow tumor growth and limit the spread of a cancer. Rapalogues have now made their way into the clinic with the rapalogue everolimus (RAD-001/Afinitor) approved for use in conjunction with exemestane, in post-menopausal breast cancer patients with advanced disease who are HER-2 negative (normal expression), hormone receptor positive and whose prior treatment with non-steroidal aromatase inhibitors has failed. Testing across multiple trials has proven that everolimus and other rapalogues are a viable way of treating certain types of cancer. However, rapalogues have shown some drawbacks both in research and clinically, with their use often activating feedback pathways that counter their usefulness. As such, new types of inhibitors are being explored that work via different mechanisms, including inhibitors that are ATP competitive with mTOR and which act to perturb signaling from both mTOR complexes.

  13. MTOR downregulates iodide uptake in thyrocytes.

    PubMed

    de Souza, Elaine Cristina Lima; Padrón, Alvaro Souto; Braga, William Miranda Oliveira; de Andrade, Bruno Moulin; Vaisman, Mário; Nasciutti, Luiz Eurico; Ferreira, Andrea Claudia Freitas; de Carvalho, Denise Pires

    2010-07-01

    Phosphoinositide-3-kinase (PI3K) inhibition increases functional sodium iodide symporter (NIS) expression in both FRTL-5 rat thyroid cell line and papillary thyroid cancer lineages. In several cell types, the stimulation of PI3K results in downstream activation of the mechanistic target of rapamycin (MTOR), a serine-threonine protein kinase that is a critical regulator of cellular metabolism, growth, and proliferation. MTOR activation is involved in the regulation of thyrocyte proliferation by TSH. Here, we show that MTOR inhibition by rapamycin increases iodide uptake in TSH-stimulated PCCL3 thyroid cell line, although the effect of rapamycin was less pronounced than PI3K inhibition. Thus, NIS inhibitory pathways stimulated by PI3K might also involve the activation of proteins other than MTOR. Insulin downregulates iodide uptake and NIS protein expression even in the presence of TSH, and both effects are counterbalanced by MTOR inhibition. NIS protein expression levels were correlated with iodide uptake ability, except in cells treated with TSH in the absence of insulin, in which rapamycin significantly increased iodide uptake, while NIS protein levels remained unchanged. Rapamycin avoids the activation of both p70 S6 and AKT kinases by TSH, suggesting the involvement of MTORC1 and MTORC2 in TSH effect. A synthetic analog of rapamycin (everolimus), which is clinically used as an anticancer agent, was able to increase rat thyroid iodide uptake in vivo. In conclusion, we show that MTOR kinase participates in the control of thyroid iodide uptake, demonstrating that MTOR not only regulates cell survival, but also normal thyroid cell function both in vitro and in vivo.

  14. Clinical presentation and management of mTOR inhibitor-associated stomatitis.

    PubMed

    de Oliveira, Marcio Augusto; Martins E Martins, Fabiana; Wang, Qian; Sonis, Stephen; Demetri, George; George, Suzanne; Butrynski, James; Treister, Nathaniel S

    2011-10-01

    Anti-cancer agents that inhibit the mTOR pathway are associated with a number of unique toxicities, with one of the most significant and potentially dose-limiting being stomatitis. The objective of this study was to report the clinical features and management outcomes of a series of cancer patients who developed painful mTOR inhibitor-associated stomatitis (mIAS). Seventeen cancer patients developed mIAS while being treated with everolimus- or ridaforolimus-containing protocols at the Dana-Farber Cancer Institute and were referred to the oral medicine clinic for evaluation and management. Clinical characteristics, toxicity management, and outcomes were summarized. In addition, the frequency and rationale for dose reductions and therapy discontinuation were assessed. The median duration of mTOR inhibitor therapy was 80 days (range 9-187 days). The median time to development of mouth ulcers was 10 days (range 4-25 days). Five patients required protocol-directed dose reductions due to grades 2 and 3 stomatitis and one patient discontinued cancer treatment due to mouth ulcers. Clinical improvement and pain relief was reported in 86.6% of patients following topical, intralesional, or systemic corticosteroid therapy, with side effects limited to secondary candidiasis (n=2). Mouth ulcers are a common and potentially dose limiting toxicity associated with the use of mTOR inhibitors in cancer treatment. This case series demonstrates that local and systemic corticosteroid therapy is an effective approach to managing patients with symptomatic mIAS. Prospective studies are necessary to evaluate the effectiveness of treatment and prevention strategies with the ultimate goal of improving overall cancer treatment outcomes.

  15. Lipopolysaccharide Exposure Induces Maternal Hypozincemia, and Prenatal Zinc Treatment Prevents Autistic-Like Behaviors and Disturbances in the Striatal Dopaminergic and mTOR Systems of Offspring.

    PubMed

    Kirsten, Thiago Berti; Chaves-Kirsten, Gabriela P; Bernardes, Suene; Scavone, Cristoforo; Sarkis, Jorge E; Bernardi, Maria Martha; Felicio, Luciano F

    2015-01-01

    Autism is characterized by social deficits, repetitive behaviors, and cognitive inflexibility. The risk factors appear to include genetic and environmental conditions, such as prenatal infections and maternal dietary factors. Previous investigations by our group have demonstrated that prenatal exposure to lipopolysaccharide (LPS), which mimics infection by gram-negative bacteria, induces autistic-like behaviors. To understand the causes of autistic-like behaviors, we evaluated maternal serum metal concentrations, which are involved in intrauterine development and infection/inflammation. We identified reduced maternal levels of zinc, magnesium, selenium and manganese after LPS exposure. Because LPS induced maternal hypozincemia, we treated dams with zinc in an attempt to prevent or ease the impairments in the offspring. We evaluated the social and cognitive autistic-like behaviors and brain tissues of the offspring to identify the central mechanism that triggers the development of autism. Prenatal LPS exposure impaired play behaviors and T-maze spontaneous alternations, i.e., it induced autistic-like behaviors. Prenatal LPS also decreased tyrosine hydroxylase levels and increased the levels of mammalian target of rapamycin (mTOR) in the striatum. Thus, striatal dopaminergic impairments may be related to autism. Moreover, excessive signaling through the mTOR pathway has been considered a biomarker of autism, corroborating our rat model of autism. Prenatal zinc treatment prevented these autistic-like behaviors and striatal dopaminergic and mTOR disturbances in the offspring induced by LPS exposure. The present findings revealed a possible relation between maternal hypozincemia during gestation and the onset of autism. Furthermore, prenatal zinc administration appears to have a beneficial effect on the prevention of autism.

  16. Lipopolysaccharide Exposure Induces Maternal Hypozincemia, and Prenatal Zinc Treatment Prevents Autistic-Like Behaviors and Disturbances in the Striatal Dopaminergic and mTOR Systems of Offspring

    PubMed Central

    Kirsten, Thiago Berti; Chaves-Kirsten, Gabriela P.; Bernardes, Suene; Scavone, Cristoforo; Sarkis, Jorge E.; Bernardi, Maria Martha; Felicio, Luciano F.

    2015-01-01

    Autism is characterized by social deficits, repetitive behaviors, and cognitive inflexibility. The risk factors appear to include genetic and environmental conditions, such as prenatal infections and maternal dietary factors. Previous investigations by our group have demonstrated that prenatal exposure to lipopolysaccharide (LPS), which mimics infection by gram-negative bacteria, induces autistic-like behaviors. To understand the causes of autistic-like behaviors, we evaluated maternal serum metal concentrations, which are involved in intrauterine development and infection/inflammation. We identified reduced maternal levels of zinc, magnesium, selenium and manganese after LPS exposure. Because LPS induced maternal hypozincemia, we treated dams with zinc in an attempt to prevent or ease the impairments in the offspring. We evaluated the social and cognitive autistic-like behaviors and brain tissues of the offspring to identify the central mechanism that triggers the development of autism. Prenatal LPS exposure impaired play behaviors and T-maze spontaneous alternations, i.e., it induced autistic-like behaviors. Prenatal LPS also decreased tyrosine hydroxylase levels and increased the levels of mammalian target of rapamycin (mTOR) in the striatum. Thus, striatal dopaminergic impairments may be related to autism. Moreover, excessive signaling through the mTOR pathway has been considered a biomarker of autism, corroborating our rat model of autism. Prenatal zinc treatment prevented these autistic-like behaviors and striatal dopaminergic and mTOR disturbances in the offspring induced by LPS exposure. The present findings revealed a possible relation between maternal hypozincemia during gestation and the onset of autism. Furthermore, prenatal zinc administration appears to have a beneficial effect on the prevention of autism. PMID:26218250

  17. Circadian regulation of mTOR by the ubiquitin pathway in renal cell carcinoma.

    PubMed

    Okazaki, Hiroyuki; Matsunaga, Naoya; Fujioka, Takashi; Okazaki, Fumiyasu; Akagawa, Yui; Tsurudome, Yuuya; Ono, Mayumi; Kuwano, Michihiko; Koyanagi, Satoru; Ohdo, Shigehiro

    2014-01-15

    Circadian clock systems regulate many biologic functions, including cell division and hormone secretion in mammals. In this study, we explored the effects of circadian control on the pivot cell growth regulatory mTOR, the activity of which is deregulated in tumor cells compared with normal cells. Specifically, we investigated whether the antitumor effect of an mTOR inhibitor could be improved by changing its dosing schedule in RenCa tumor-bearing mice. Active, phosphorylated mTOR displayed a 24-hour rhythm, and levels of total mTOR protein (but not mRNA) also showed a circadian rhythm in RenCa tumor masses. Through investigations of the oscillation mechanism for mTOR expression, we identified the ubiquitination factor Fbxw7 as an mTOR regulator that oscillated in its expression in a manner opposite from mTOR. Fbxw7 transcription was regulated by the circadian regulator D-site-binding protein. Notably, administration of the mTOR inhibitor everolimus during periods of elevated mTOR improved survival in tumor-bearing mice. Our findings demonstrate that the circadian oscillation of mTOR activity is regulated by circadian clock systems, which influence the antitumor effect of mTOR inhibitors.

  18. Effects of mTOR and calcineurin inhibitors combined therapy in Epstein-Barr virus positive and negative Burkitt lymphoma cells.

    PubMed

    Wowro, Sylvia J; Schmitt, Katharina R L; Tong, Giang; Berger, Felix; Schubert, Stephan

    2016-01-01

    Post-transplant lymphoproliferative disorder is a severe complication in solid organ transplant recipients, which is highly associated with Epstein-Barr virus infection in pediatric patients and occasionally presents as Burkitt- or Burkitt-like lymphoma. The mammalian target of rapamycin (mTOR) pathway has been described as a possible antitumor target whose inhibition may influence lymphoma development and proliferation after pediatric transplantation. We treated Epstein-Barr virus positive (Raji and Daudi) and negative (Ramos) human Burkitt lymphoma derived cells with mTOR inhibitor everolimus alone and in combination with clinically relevant immunosuppressive calcineurin inhibitors (tacrolimus or cyclosporin A). Cell proliferation, toxicity, and mitochondrial metabolic activity were analyzed. The effect on mTOR Complex 1 downstream targets p70 S6 kinase, eukaryotic initiation factor 4G, and S6 ribosomal protein activation was also investigated. We observed that treatment with everolimus alone significantly decreased Burkitt lymphoma cell proliferation and mitochondrial metabolic activity. Everolimus in combination with cyclosporin A had a stronger suppressive effect in Epstein-Barr virus negative but not in Epstein-Barr virus positive cells. In contrast, tacrolimus completely abolished the everolimus-mediated suppressive effects. Moreover, we showed a significant decrease in activation of mTOR Complex 1 downstream targets after treatment with everolimus that was attenuated when combined with tacrolimus, but not with cyclosporin A. For the first time we showed the competitive effect between everolimus and tacrolimus when used as combination therapy on Burkitt lymphoma derived cells. Thus, according to our in vitro data, the combination of calcineurin inhibitor cyclosporin A with everolimus is preferred to the combination of tacrolimus and everolimus.

  19. Prevention of tumor growth driven by PIK3CA and HPV oncogenes by targeting mTOR signaling with metformin in oral squamous carcinomas expressing OCT3

    PubMed Central

    Madera, Dmitri; Vitale-Cross, Lynn; Martin, Daniel; Schneider, Abraham; Molinolo, Alfredo A.; Gangane, Nitin; Carey, Thomas E.; McHugh, Jonathan B.; Komarck, Christine M.; Walline, Heather M.; William, William N.; Seethala, Raja R.; Ferris, Robert; Gutkind, J. Silvio

    2015-01-01

    Most head and neck squamous cell carcinomas (HNSCC) exhibit a persistent activation of the PI3K-mTOR signaling pathway. We have recently shown that metformin, an oral antidiabetic drug that is also used to treat lipodystrophy in HIV-infected (HIV+) individuals, diminishes mTOR activity and prevents the progression of chemically-induced experimental HNSCC premalignant lesions. Here, we explored the preclinical activity of metformin in HNSCCs harboring PIK3CA mutations and HPV oncogenes, both representing frequent HNSCC alterations, aimed at developing effective targeted preventive strategies. The biochemical and biological effects of metformin were evaluated in representative HNSCC cells expressing mutated PIK3CA or HPV oncogenes (HPV+). The oral delivery of metformin was optimized to achieve clinical relevant blood levels. Molecular determinants of metformin sensitivity were also investigated, and their expression levels examined in a large collection of HNSCC cases. We found that metformin inhibits mTOR signaling and tumor growth in HNSCC cells expressing mutated PIK3CA and HPV oncogenes, and that these activities require the expression of organic cation transporter 3 (OCT3/SLC22A3), a metformin uptake transporter. Co-expression of OCT3 and the mTOR pathway activation marker pS6 were observed in most HNSCC cases, including those arising in HIV+ patients. Activation of the PI3K-mTOR pathway is a widespread event in HNSCC, including HPV− and HPV+ lesions arising in HIV+ patients, all of which co-express OCT3. These observations may provide a rationale for the clinical evaluation of metformin to halt HNSCC development from precancerous lesions, including in HIV+ individuals at risk of developing HPV-associated cancers. PMID:25681087

  20. Common occurrence of everolimus-associated aphthous stomatitis in Japanese heart transplant recipients.

    PubMed

    Sasaoka, T; Kato, T S; Oda, N; Wada, K; Komamura, K; Asakura, M; Hashimura, K; Ishibashi-Ueda, H; Nakatani, T; Isobe, M; Kitakaze, M

    2010-11-01

    Mammalian target of rapamycin (mTOR) inhibitors display antiproliferative effects with less nephrotoxicity than calcineurin inhibitors. However, clinical use of mTOR inhibitors can be associated with a series of adverse events. We experienced cases of aphthous stomatitis associated with everolimus (EVL) in four Japanese heart transplant recipients treated at the target trough EVL blood level after a switch from mycophenolate mofetil between April and December 2007. All four patients developed aphthous stomatitis; three required reduction of the exposure and one, EVL discontinuation due to stomatitis as well as other side effects. All patients recovered from stomatitis after reduction or withdrawal of EVL. Thus, we considered that EVL-related stomatitis might occur commonly among the Japanese population. The proper dosage, effects, and frequency of the side effects of mTOR inhibitors may vary by ethnic population.

  1. Targeting of Tumor Growth and Angiogenesis Underlies the Enhanced Antitumor Activity of Lenvatinib in Combination with Everolimus.

    PubMed

    Matsuki, Masahiro; Adachi, Yusuke; Ozawa, Yoichi; Kimura, Takayuki; Hoshi, Taisuke; Okamoto, Kiyoshi; Tohyama, Osamu; Mitsuhashi, Kaoru; Yamaguchi, Atsumi; Matsui, Junji; Funahashi, Yasuhiro

    2017-01-20

    The combination of lenvatinib-a multiple receptor tyrosine kinase (RTK) inhibitor-plus everolimus-a mammalian target of rapamycin (mTOR) inhibitor-significantly improved clinical outcomes versus everolimus monotherapy in a phase 2 clinical study of metastatic renal cell carcinoma (RCC). Here, we investigated potential mechanisms underlying the antitumor activity of the combination treatment in preclinical RCC models. Lenvatinib plus everolimus showed greater antitumor activity than either monotherapy in 3 human RCC xenograft mouse models (A-498, Caki-1, and Caki-2). In particular, the combination led to tumor regression in the A-498 and Caki-1 models. In the A-498 model, everolimus demonstrated antiproliferative activity, whereas lenvatinib showed antiangiogenic effects. The antiangiogenic activity was potentiated by the lenvatinib plus everolimus combination in Caki-1 xenografts, where FGF-driven angiogenesis may contribute to tumor growth. The combination showed mostly additive activity in VEGF-activated, and synergistic activity against FGF-activated endothelial cells in cell proliferation and tube formation assays, as well as strongly suppressed mTOR-S6K-S6 signaling. Enhanced antitumor activities of the combination versus each monotherapy were also observed in mice bearing human pancreatic KP-1 xenografts overexpressing VEGF or FGF. Our results indicated that simultaneous targeting tumor cell growth and angiogenesis by lenvatinib plus everolimus resulted in enhanced antitumor activity. The enhanced inhibition of both VEGF- and FGF-signaling pathways by the combination underlies its superior antiangiogenic activity in human RCC xenograft models. This article is protected by copyright. All rights reserved.

  2. The dual targeting of insulin and insulin-like growth factor 1 receptor enhances the mTOR inhibitor-mediated antitumor efficacy in hepatocellular carcinoma

    PubMed Central

    Pivonello, Claudia; Negri, Mariarosaria; De Martino, Maria Cristina; Napolitano, Maria; de Angelis, Cristina; Provvisiero, Donatella Paola; Cuomo, Gaia; Auriemma, Renata Simona; Simeoli, Chiara; Izzo, Francesco; Colao, Annamaria; Hofland, Leo J.; Pivonello, Rosario

    2016-01-01

    Deregulation of mTOR and IGF pathways is frequent in hepatocellular carcinoma (HCC), thus mTOR and IGF1R represent suitable therapeutic targets in HCC. The aim of this study was to evaluate the effects of mTOR inhibitors (mTORi) and OSI-906, blocker of IGF1R/IR, on HCC cell proliferation, viability, migration and invasion, and alpha-fetoprotein (α-FP) secretion. In HepG2 and HuH-7 we evaluated, the expression of mTOR and IGF pathway components; the effects of Sirolimus, Everolimus, Temsirolimus and OSI-906 on cell proliferation; the effects of Sirolimus, OSI-906, and their combination, on cell secretion, proliferation, viability, cell cycle, apoptosis, invasion and migration. Moreover, intracellular mechanisms underlying these cell functions were evaluated in both cell lines. Our results show that HepG2 and HuH-7 present with the same mRNA expression profile with high levels of IGF2. OSI-906 inhibited cell proliferation at high concentration, while mTORi suppressed cell proliferation in a dose-time dependent manner in both cell lines. The co-treatment showed an additive inhibitory effect on cell proliferation and viability. This effect was not related to induction of apoptosis, but to G0/G1 phase block. Moreover, the co-treatment prevented the Sirolimus-induced AKT activation as escape mechanism. Both agents demonstrated to be differently effective in inhibiting α-FP secretion. Sirolimus, OSI-906, and their combination, blocked cell migration and invasion in HuH-7. These findings indicate that, co-targeting of IGF1R/IR and mTOR pathways could be a novel therapeutic approach in the management of HCC, in order to maximize antitumoral effect and to prevent the early development of resistance mechanisms. PMID:26756219

  3. Role of mTOR Inhibitors in Kidney Disease

    PubMed Central

    Kajiwara, Moto; Masuda, Satohiro

    2016-01-01

    The first compound that inhibited the mammalian target of rapamycin (mTOR), sirolimus (rapamycin) was discovered in the 1970s as a soil bacterium metabolite collected on Easter Island (Rapa Nui). Because sirolimus showed antiproliferative activity, researchers investigated its molecular target and identified the TOR1 and TOR2. The mTOR consists of mTOR complex 1 (mTORC1) and mTORC2. Rapalogues including sirolimus, everolimus, and temsirolimus exert their effect mainly on mTORC1, whereas their inhibitory effect on mTORC2 is mild. To obtain compounds with more potent antiproliferative effects, ATP-competitive inhibitors of mTOR targeting both mTORC1 and mTORC2 have been developed and tested in clinical trials as anticancer drugs. Currently, mTOR inhibitors are used as anticancer drugs against several solid tumors, and immunosuppressive agents for transplantation of various organs. This review discusses the role of mTOR inhibitors in renal disease with a particular focus on renal cancer, diabetic nephropathy, and kidney transplantation. PMID:27338360

  4. A randomized, controlled trial of everolimus-based dual immunosuppression versus standard of care in de novo kidney transplant recipients.

    PubMed

    Chadban, Steven J; Eris, Josette Marie; Kanellis, John; Pilmore, Helen; Lee, Po Chang; Lim, Soo Kun; Woodcock, Chad; Kurstjens, Nicol; Russ, Graeme

    2014-03-01

    Kidney transplant recipients receiving calcineurin inhibitor-based immunosuppression incur increased long-term risks of cancer and kidney fibrosis. Switch to mammalian target of rapamycin (mTOR) inhibitors may reduce these risks. Steroid or Cyclosporin Removal After Transplant using Everolimus (SOCRATES), a 36-month, prospective, multinational, open-label, randomized controlled trial for de novo kidney transplant recipients, assessed whether everolimus switch could enable elimination of mycophenolate plus either steroids or CNI without compromising efficacy. Patients received cyclosporin, mycophenolate and steroids for the first 14 days then everolimus with mycophenolate and CNIwithdrawal (CNI-WD); everolimus with mycophenolate and steroid withdrawal (steroid-WD); or cyclosporin, mycophenolate and steroids (control). 126 patients were randomized. The steroid WD arm was terminated prematurely because of excess discontinuations. Mean eGFR at month 12 for CNI-WD versus control was 65.1 ml/min/1.73 m2 vs. 67.1 ml/min/1.73 m2 by ITT, which met predefined noninferiority criteria (P=0.026). The CNI-WD group experienced a higher rate of BPAR(31% vs. control 13%, P=0.048) and showed a trend towards higher composite treatment failure (BPAR, graft loss, death, loss to follow-up). The 12 month results from SOCRATES show noninferiority in eGFR, but a significant excess of acute rejection when everolimus was commenced at week 2 to enable a progressive withdrawal of mycophenolate and cyclosporin in kidney transplant recipients.

  5. Conversion from cyclosporine to everolimus at 4.5 months posttransplant: 3-year results from the randomized ZEUS study.

    PubMed

    Budde, K; Lehner, F; Sommerer, C; Arns, W; Reinke, P; Eisenberger, U; Wüthrich, R P; Scheidl, S; May, C; Paulus, E-M; Mühlfeld, A; Wolters, H H; Pressmar, K; Stahl, R; Witzke, O

    2012-06-01

    The long-term effect of conversion from calcineurin inhibitor (CNI) therapy to an mTOR inhibitor requires clarification. Following completion of the 12-month, open-label, multicenter ZEUS study, in which 300 kidney transplant recipients were randomized to continue cyclosporine (CsA) or convert to everolimus at 4.5 months posttransplant, outcomes were assessed at month 36 (n = 284; 94.7%). CNI therapy was reintroduced in 28.4% of everolimus patients by month 36. The primary efficacy endpoint, estimated glomerular filtration rate (Nankivell, ANCOVA) was significantly higher with everolimus versus the CsA group at month 24 (7.6 mL/min/1.73 m(2) , 95%CI 4.3, 11.0 mL/min/1.73 m(2) ; p < 0.001) and month 36 (7.5 mL/min/1.73 m(2) , 95%CI 3.6, 11.4 mL/min/1.73 m(2) ; p < 0.001). The incidence of biopsy-proven acute rejection from randomization to month 36 was 13.0% in the everolimus arm and 4.8% in the CsA arm (p = 0.015). Patient and graft survival, as well as incidences of malignancy, severe infections and hospitalization, were similar between groups. Kidney transplant patients who are converted from CsA to everolimus at month 4.5 and who remain on everolimus thereafter may achieve a significant improvement in renal function that is maintained to 3 years. There was a significantly higher rate of rejection in the everolimus arm but this did not exert a deleterious effect by 3 years posttransplant.

  6. Everolimus in the treatment of neuroendocrine tumors of the respiratory and gastroenteropancreatic systems.

    PubMed

    Flaum, Nicola; Valle, Juan W; Mansoor, Wasat; McNamara, Mairéad G

    2016-11-01

    Neuroendocrine tumors (NETs) are a rare diverse group of malignancies occurring most commonly in the gastroenteropancreatic system and the lungs. The incidence of NETs is increasing worldwide; median survival for patients with metastatic NETs is 5-65 months. A growing body of evidence shows survival benefit in patients with advanced NETs (gastroenteropancreatic and lung) treated with mTOR inhibitor everolimus, with improvement in survival being demonstrated in the clinical trial and real-world setting. Everolimus has been shown to have a manageable safety profile, with the most common adverse events being stomatitis, rash, diarrhea, fatigue and infections. Due to the rarity of the condition, there are challenges in conducting clinical trials in these patients. Further research is required to clarify the role of adjuvant therapy, treatment sequencing and the use of multimodality treatments.

  7. Significant cytostatic effect of everolimus on a gefitinib-resistant anaplastic thyroid cancer cell line harboring PI3KCA gene mutation

    PubMed Central

    ONODA, NAOYOSHI; NAKAMURA, MASANORI; AOMATSU, NAOKI; NODA, SATORU; KASHIWAGI, SHINICHIRO; KURATA, KENTO; UCHINO, SHINYA; HIRAKAWA, KOSEI

    2015-01-01

    We previously demonstrated the efficacy of gefitinib, a tyrosine kinase inhibitor of the epidermal growth factor receptor (EGFR), on an anaplastic thyroid cancer (ATC) cell line. We also observed that gefitinib was not effective in regulating cell growth in a different ATC cell line that exhibited an altered EGFR-initiated signal transduction pathway. In the present study, we attempted to regulate the downstream effector of EGFR-Akt-mammalian target of rapamycin (mTOR) pathway by an mTOR inhibitor, everolimus. A total of 8 ATC cell lines were employed, 7 of which were established in our institute. OCUT-2 was known to carry a mutation in the phosphoinositide-3-kinase, catalytic, α polypeptide gene (PI3KCA) and to be gefitinib-resistant, whereas ACT-1 exhibited a remarkable growth arrest by gefitinib. All the cell lines were tested for the cytotoxic effect of everolimus. The mechanisms of cellular toxicity were investigated by EGFR stimulation, cell cycle and concurrent exposure to paclitaxel. In OCUT-2, but not in any of the other cell lines, everolimus achieved a significant growth inhibition (inhibition of 30 and 50% was achieved by concentrations of 0.8 and 5 nM, respectively). The growth in OCUT-2 was inhibited by everolimus, even with concordant EGFR stimulation. This effect was demonstrated by a G2M cell cycle arrest. An additive effect of everolimus onto the cytotoxic effect of paclitaxel was demonstrated at a dose of 1–2 nM. A significant growth inhibitory effect of everolimus on the gefitinib-resistant ATC cell line was demonstrated, suggesting a possible correlation between the efficacy of everolimus and PI3KCA gene mutation and the significance of molecular-targeted therapy in the management of ATC. PMID:26137260

  8. Long-Term Use of Everolimus in Patients with Tuberous Sclerosis Complex: Final Results from the EXIST-1 Study

    PubMed Central

    Franz, David N.; Belousova, Elena; Sparagana, Steven; Bebin, E. Martina; Frost, Michael D.; Kuperman, Rachel; Witt, Olaf; Kohrman, Michael H.; Flamini, J. Robert; Wu, Joyce Y.; Curatolo, Paolo; de Vries, Petrus J.; Berkowitz, Noah; Niolat, Julie; Jóźwiak, Sergiusz

    2016-01-01

    Background Everolimus, a mammalian target of rapamycin (mTOR) inhibitor, has demonstrated efficacy in treating subependymal giant cell astrocytomas (SEGAs) and other manifestations of tuberous sclerosis complex (TSC). However, long-term use of mTOR inhibitors might be necessary. This analysis explored long-term efficacy and safety of everolimus from the conclusion of the EXIST-1 study (NCT00789828). Methods and Findings EXIST-1 was an international, prospective, double-blind, placebo-controlled phase 3 trial examining everolimus in patients with new or growing TSC-related SEGA. After a double-blind core phase, all remaining patients could receive everolimus in a long-term, open-label extension. Everolimus was initiated at a dose (4.5 mg/m2/day) titrated to a target blood trough of 5–15 ng/mL. SEGA response rate (primary end point) was defined as the proportion of patients achieving confirmed ≥50% reduction in the sum volume of target SEGA lesions from baseline in the absence of worsening nontarget SEGA lesions, new target SEGA lesions, and new or worsening hydrocephalus. Of 111 patients (median age, 9.5 years) who received ≥1 dose of everolimus (median duration, 47.1 months), 57.7% (95% confidence interval [CI], 47.9–67.0) achieved SEGA response. Of 41 patients with target renal angiomyolipomas at baseline, 30 (73.2%) achieved renal angiomyolipoma response. In 105 patients with ≥1 skin lesion at baseline, skin lesion response rate was 58.1%. Incidence of adverse events (AEs) was comparable with that of previous reports, and occurrence of emergent AEs generally decreased over time. The most common AEs (≥30% incidence) suspected to be treatment-related were stomatitis (43.2%) and mouth ulceration (32.4%). Conclusions Everolimus use led to sustained reduction in tumor volume, and new responses were observed for SEGA and renal angiomyolipoma from the blinded core phase of the study. These findings support the hypothesis that everolimus can safely reverse

  9. Long-term mTOR inhibitors administration evokes altered calcium homeostasis and platelet dysfunction in kidney transplant patients

    PubMed Central

    López, Esther; Berna-Erro, Alejandro; Bermejo, Nuria; Brull, José María; Martinez, Rocío; Garcia Pino, Guadalupe; Alvarado, Raul; Salido, Ginés María; Rosado, Juan Antonio; Cubero, Juan José; Redondo, Pedro Cosme

    2013-01-01

    The use of the mammal target of rapamycin (mTOR) inhibitors has been consolidated as the therapy of election for preventing graft rejection in kidney transplant patients, despite their immunosuppressive activity is less strong than anti-calcineurin agents like tacrolimus and cyclosporine A. Furthermore, as mTOR is widely expressed, rapamycin (a macrolide antibiotic produced by Streptomyces hygroscopicus) is recommended in patients presenting neoplasia due to its antiproliferative actions. Hence, we have investigated whether rapamycin presents side effects in the physiology of other cell types different from leucocytes, such as platelets. Blood samples were drawn from healthy volunteers and kidney transplant patients long-term medicated with rapamycin: sirolimus and everolimus. Platelets were either loaded with fura-2 or directly stimulated, and immunoassayed or fixed with Laemmli's buffer to perform the subsequent analysis of platelet physiology. Our results indicate that rapamycin evokes a biphasic time-dependent alteration in calcium homeostasis and function in platelets from kidney transplant patients under rapamycin regime, as demonstrated by the reduction in granule secretion observed and subsequent impairment of platelet aggregation in these patients compared with healthy volunteers. Platelet count was also reduced in these patients, thus 41% of patients presented thrombocytopenia. All together our results show that long-term administration of rapamycin to kidney transplant patients evokes alteration in platelet function. PMID:23577651

  10. Targeting mTOR pathway inhibits tumor growth in different molecular subtypes of triple-negative breast cancers

    PubMed Central

    Hatem, Rana; Botty, Rania El; Chateau-Joubert, Sophie; Servely, Jean-Luc; Labiod, Dalila; de Plater, Ludmilla; Assayag, Franck; Coussy, Florence; Callens, Céline; Vacher, Sophie; Reyal, Fabien; Cosulich, Sabina; Diéras, Véronique

    2016-01-01

    Triple-negative breast cancers (TNBC) are characterized by frequent alterations in the PI3K/AKT/mTOR signaling pathway. In this study, we analyzed PI3K pathway activation in 67 patient-derived xenografts (PDX) of breast cancer and investigated the anti-tumor activity of the mTOR inhibitor everolimus in 15 TNBC PDX with different expression and mutational status of PI3K pathway markers. Expression of the tumor suppressors PTEN and INPP4B was lost in 55% and 76% of TNBC PDX, respectively, while mutations in PIK3CA and AKT1 genes were rare. In 7 PDX treatment with everolimus resulted in a tumor growth inhibition higher than 50%, while 8 models were classified as low responder or resistant. Basal-like, LAR (Luminal AR), mesenchymal and HER2-enriched tumors were present in both responder and resistant groups, suggesting that tumor response to everolimus is not restricted to a specific TNBC subtype. Analysis of treated tumors showed a correlation between tumor response and post-treatment phosphorylation of AKT, increased in responder PDX, while PI3K pathway markers at baseline were not sufficient to predict everolimus response. In conclusion, targeting mTOR decreased tumor growth in 7 out of 15 TNBC PDX tested. Response to everolimus occurred in different TNBC subtypes and was associated with post-treatment increase of P-AKT. PMID:27374081

  11. Adjuvant Everolimus for Resected Kidney Cancer

    Cancer.gov

    In this clinical trial, patients with renal cell cancer who have undergone partial or complete nephrectomy will be randomly assigned to take everolimus tablets or matching placebo tablets daily for 54 weeks.

  12. L744,832 and Everolimus Induce Cytotoxic and Cytostatic Effects in Non-Hodgkin Lymphoma Cells.

    PubMed

    Mendes, José; Gonçalves, Ana Cristina; Alves, Raquel; Jorge, Joana; Pires, Ana; Ribeiro, Ana; Sarmento-Ribeiro, Ana Bela

    2016-04-01

    Non-Hodgkin Lymphoma (NHL) constitutes a very heterogeneous group of diseases with different aggressiveness. Diffuse large B-cell lymphoma (DLBCL) and Burkitt's lymphoma (BL) are two clinically aggressive lymphomas from the germinal center, very heterogeneous and with different genetic signatures. Several intracellular pathways are involved in lymphomagenesis, being BCR/PI3K/AKT/mTOR and RAS/RAF pathways the most frequently ones. In this context the therapeutic potential of a mTOR inhibitor--everolimus--and a RAS/RAF pathway inhibitor--L744,832--was evaluated in two NHL cell lines. Farage and Raji cells were cultured in the absence and presence of several concentrations of everolimus and L744,832 in monotherapy and in combination with each other, as well as in association with the conventional chemotherapy drug vincristine. Our results show that everolimus and L744,832 induce antiproliferative and cytotoxic effect in a time-, dose-, and cell line-dependent manner, inducing cell death mainly by apoptosis. A potentiation effect was observed when the drugs were used in combination. In conclusion, the results suggest that everolimus and L744,832, alone or in combination, could provide therapeutic benefits in these subtypes of NHL.

  13. Everolimus Combined With Gefitinib in Patients with Metastatic Castration-Resistant Prostate Cancer: Phase I/II Results and Signaling Pathway Implications

    PubMed Central

    Rathkopf, Dana E.; Larson, Steven M.; Anand, Aseem; Morris, Michael J.; Slovin, Susan F.; Shaffer, David R.; Heller, Glenn; Carver, Brett; Rosen, Neal; Scher, Howard I.

    2015-01-01

    Background The effects of mammalian target of rapamycin (mTOR) inhibition are limited by feedback reactivation of receptor tyrosine kinase signaling in PTEN-null tumors, thus we tested the combination of mTOR inhibition (everolimus) and EGFR inhibition (gefitinib) in castration-resistant prostate cancer (CRPC). Methods In phase I, 12 patients (10 CRPC, 2 glioblastoma) received daily gefitinib (250 mg) with weekly everolimus (30, 50, or 70 mg). In phase II, 27 CRPC patients received gefitinib with everolimus 70 mg. Results Phase I revealed no pharmacokinetic interactions and no dose-limiting toxicities. In phase II, 18 of 27 (67%) patients discontinued treatment before the 12-week evaluation due to progression as evidenced by prostate-specific antigen (PSA) levels (n=6) or imaging (n=5), or grade ≥2 toxicity (n=7). Thirteen of the total 37 (35%) CRPC patients exhibited a rapidly rising PSA after starting treatment which declined upon discontinuation. Fluorodeoxyglucose positron emission tomography at 24 to 72 hours after starting treatment showed a decrease in standardized uptake value consistent with mTOR inhibition in 27 of 33 (82%) evaluable patients; there was a corresponding rise in PSA in 20 of these 27 patients (74%). Conclusions The combination of gefitinib and everolimus did not result in significant antitumor activity. The induction of PSA in tumors treated with mTOR inhibitors was consistent with preclinical data that PI3K pathway signaling feedback inhibits the androgen receptor (AR). This clinical evidence of relief of feedback inhibition promoting enhanced AR activity supports future studies combining PI3K pathway inhibitors and second-generation AR inhibitors in CRPC. PMID:26178426

  14. mTOR inhibitors in the treatment of breast cancer.

    PubMed

    Vinayak, Shaveta; Carlson, Robert W

    2013-01-01

    The phosphatidylinositol 3-kinase/mammalian target of rapamycin (PI3K/mTOR) pathway is commonly dysregulated in breast cancer. In preclinical studies, hyperactivation of the PI3K pathway has been linked to resistance to both endocrine therapy and trastuzumab (Herceptin). Rapalogs, agents that primarily inhibit mTOR-raptor complex 1, have been studied in combination with endocrine therapy to overcome endocrine resistance.Trials of combination endocrine therapy and rapalogs in metastatic hormone receptor-positive breast cancer have demonstrated variable results. However, two independent trials have recently shown that combination everolimus (Afinitor) and tamoxifen or combination everolimus and exemestane (Aromasin) is more effective than either endocrine agent alone. These trials selected patients with cancer refractory to endocrine therapy, which may be important in sensitizing tumors to inhibition of this pathway. In human epidermal growth factor receptor 2 (HER2)-positive breast cancer, the early clinical data with combinations of PI3K/mTOR inhibitors and anti-HER2 therapies are encouraging. Efforts to identify clinical biomarkers of response or resistance to mTOR inhibitors are ongoing. This review will summarize results of preclinical and clinical studies aswell as ongoing clinical trials with mTOR or dual PI3K/mTOR inhibitors.

  15. Phase II Study of Everolimus and Letrozole in Patients With Recurrent Endometrial Carcinoma

    PubMed Central

    Slomovitz, Brian M.; Jiang, Yunyun; Yates, Melinda S.; Soliman, Pamela T.; Johnston, Taren; Nowakowski, Maureen; Levenback, Charles; Zhang, Qian; Ring, Kari; Munsell, Mark F.; Gershenson, David M.; Lu, Karen H.; Coleman, Robert L.

    2015-01-01

    Purpose The phosphoinositol-3 kinase (PI3K) pathway is frequently dysregulated in endometrial cancer (EC). Hormonal manipulation leads to response in some patients with EC, but resistance derived from PI3K pathway activation has been documented. Targeting mammalian target of rapamycin (mTOR) may overcome endocrine resistance. We conducted a two-institution phase II trial of everolimus and letrozole in women with recurrent EC. Patients and Methods Patients were considered incurable, had measurable disease, and were treated with up to two prior cytotoxic regimens. Everolimus was administered orally at 10 mg daily and letrozole was administered orally at 2.5 mg daily. Each cycle consisted of 4 weeks of therapy. Patients were treated until progression, toxicity, or complete response (CR). The primary end point was the clinical benefit rate (CBR), which was defined as CR, partial response, or stable disease (≥ 16 weeks) by RECIST 1.0 criteria. Translational studies were performed to correlate biomarkers with response. Results Thirty-eight patients were enrolled (median age, 62 years; range, 24 to 82 years). Thirty-five patients were evaluable for response. The CBR was 40% (14 of 35 patients); the median number of cycles among responders was 15 (range, seven to 29 cycles). The confirmed objective response rate (RR) was 32% (11 of 35 patients; nine CRs and two partial responses; median, 15 cycles; range, eight to 29 cycles). Twenty percent of patients (seven of 35 patients) were taken off treatment after a prolonged CR and at the discretion of the treating clinician. None of the patients discontinued treatment as a result of toxicity. Serous histology was the best predictor of lack of response. Patients with endometrioid histology and CTNNB1 mutations responded well to everolimus and letrozole. Conclusion Everolimus plus letrozole results in a high CBR and RR in patients with recurrent EC. Further development of this combination in recurrent endometrioid EC is under way

  16. Efficacy and safety of conversion from cyclosporine to everolimus in living-donor kidney transplant recipients: an analysis from the ZEUS study.

    PubMed

    Lehner, Frank; Budde, Klemens; Zeier, Martin; Wüthrich, Rudolf P; Reinke, Petra; Eisenberger, Ute; Mühlfeld, Anja; Arns, Wolfgang; Stahl, Rolf; Heller, Katharina; Witzke, Oliver; Wolters, Heiner H; Suwelack, Barbara; Klehr, Hans Ulrich; Stangl, Manfred; Hauser, Ingeborg A; Nadalin, Silvio; Porstner, Martina; May, Christoph; Paulus, Eva-Maria; Sommerer, Claudia

    2014-11-01

    Conversion of living-donor kidney transplant patients from calcineurin inhibitor therapy to an mTOR inhibitor is poorly documented. In the prospective, multicentre ZEUS study, 300 kidney transplant recipients without prior rejection (Banff grade >1) and serum creatinine ≤265 μmol/l were randomized to continue cyclosporine or convert to everolimus at 4.5 months post-transplant. In a post hoc analysis of 80 living-donor recipients, adjusted estimated GFR (Nankivell) at month 12 (the primary endpoint) was 74.3 (95% CI [70.7, 77.9]) ml/min/1.73 m(2) with everolimus versus 63.8 (95% CI [60.0, 67.7]) ml/min/1.73 m(2) ) with cyclosporine, a difference of 10.5 ml/min/1.73 m(2) in favour of everolimus (P < 0.001). From randomization to month 12, adjusted estimated GFR increased by a mean of 9.8 (95% CI [6.2, 13.4]) ml/min/1.73 m(2) with everolimus versus -0.7 (95% CI [-4.6, 3.1]) ml/min/1.73 m(2) ) (P < 0.001) with cyclosporine. There were six biopsy-proven acute rejection episodes in everolimus-treated patients (five Banff grade I) and one episode in cyclosporine-treated patients (Banff grade 1). Overall safety profile was similar between groups. Discontinuation due to adverse events occurred in three everolimus patients (7.1%) and five cyclosporine patients (13.2%) between randomization and month 12. Initiation of everolimus with early elimination of calcineurin therapy is associated with a significant renal benefit at 12 months post-transplant that is observed in both living and deceased-donor recipients. (clinicaltrials.gov NCT00154310).

  17. Molecular analysis of a male breast cancer patient with prolonged stable disease under mTOR/PI3K inhibitors BEZ235/everolimus

    PubMed Central

    Brannon, A. Rose; Frizziero, Melissa; Chen, David; Hummel, Jennifer; Gallo, Jorge; Riester, Markus; Patel, Parul; Cheung, Wing; Morrissey, Michael; Carbone, Carmine; Cottini, Silvia; Tortora, Giampaolo; Melisi, Davide

    2016-01-01

    The mTORC1 inhibitor everolimus (Afinitor/RAD001) has been approved for multiple cancer indications, including ER+/HER2− metastatic breast cancer. However, the combination of everolimus with the dual PI3K/mTOR inhibitor BEZ235 was shown to be more efficacious than either everolimus or BEZ235 alone in preclinical models. Herein, we describe a male breast cancer (MBC) patient who was diagnosed with hormone receptor-positive (HR+)/HER2− stage IIIA invasive ductal carcinoma and sequentially treated with chemoradiotherapy and hormonal therapy. Upon the development of metastases, the patient began a 200 mg twice-daily BEZ235 and 2.5 mg weekly everolimus combination regimen. The patient sustained a prolonged stable disease of 18 mo while undergoing the therapy, before his tumor progressed again. Therefore, we sought to both better understand MBC and investigate the underlying molecular mechanisms of the patient's sensitivity and subsequent resistance to the BEZ235/everolimus combination therapy. Genomic and immunohistochemical analyses were performed on samples collected from the initial invasive ductal carcinoma pretreatment and a metastasis postprogression on the BEZ235/everolimus combination treatment. Both tumors were relatively quiet genomically with no overlap to recurrent MBC alterations in the literature. Markers of PI3K/mTOR pathway hyperactivation were not identified in the pretreatment sample, which complements previous reports of HR+ female breast cancers being responsive to mTOR inhibition without this activation. The postprogression sample, however, demonstrated greater than fivefold increased estrogen receptor and pathogenesis-related protein expression, which could have constrained the PI3K/mTOR pathway inhibition by BEZ235/everolimus. Overall, these analyses have augmented the limited episteme on MBC genetics and treatment. PMID:27148582

  18. [A Case of Renal Cell Carcinoma with High Everolimus Blood Concentrations and Hyperglycemia Due to Everolimus-Induced Hepatic Dysfunction].

    PubMed

    Takasaki, Shinya; Kikuchi, Masafumi; Kawasaki, Yoshihide; Ito, Akihiro; Arai, Yoichi; Yamaguchi, Hiroaki; Mano, Nariyasu

    2017-01-01

    We report the case of a patient who had renal cell carcinoma with high everolimus blood concentrations and hyperglycemia due to everolimus-induced hepatic dysfunction. A 74-year-old man who underwent right nephrectomy for renal cell carcinoma was administered everolimus for multiple lung metastases. Everolimus caused grade 3 hepatic dysfunction and hyperglycemia; hence, high blood levels of everolimus were observed. Although the patient was re-administrated everolimus after recovering from hepatic dysfunction, hepatic function test values worsened again. Everolimus was discontinued before its blood concentration increased, and the patient was switched to axitinib treatment. Therefore, the measurement of everolimus blood level is considered useful for the management of adverse events in renal cell carcinoma.

  19. The Role of mTOR Inhibitors in the Treatment of Patients with Tuberous Sclerosis Complex: Evidence-based and Expert Opinions.

    PubMed

    Curatolo, Paolo; Bjørnvold, Marit; Dill, Patricia E; Ferreira, José Carlos; Feucht, Martha; Hertzberg, Christoph; Jansen, Anna; Jóźwiak, Sergiusz; Kingswood, J Christopher; Kotulska, Katarzyna; Macaya, Alfons; Moavero, Romina; Nabbout, Rima; Zonnenberg, Bernard A

    2016-04-01

    Tuberous sclerosis complex (TSC) is a genetic disorder arising from mutations in the TSC1 or TSC2 genes. The resulting over-activation of the mammalian target of rapamycin (mTOR) signalling pathway leaves patients with TSC susceptible to the growth of non-malignant tumours in multiple organs. Previously, surgery was the main therapeutic option for TSC. However, pharmacological therapy with mTOR inhibitors such as everolimus and sirolimus is now emerging as an alternate approach. Everolimus and sirolimus have already been shown to be effective in treating subependymal giant cell astrocytoma (SEGA) and renal angiomyolipoma (AML), and everolimus is currently being evaluated in treating TSC-related epilepsy. In November 2013 a group of European experts convened to discuss the current options and practical considerations for treating various manifestations of TSC. This article provides evidence-based recommendations for the treatment of SEGA, TSC-related epilepsy and renal AML, with a focus on where mTOR inhibitor therapy may be considered alongside other treatment options. Safety considerations regarding mTOR inhibitor therapy are also reviewed. With evidence of beneficial effects in neurological and non-neurological TSC manifestations, mTOR inhibitors may represent a systemic treatment for TSC.

  20. Preclinical evaluation of the combination of mTOR and proteasome inhibitors with radiotherapy in malignant peripheral nerve sheath tumors.

    PubMed

    Yamashita, A S; Baia, G S; Ho, J S Y; Velarde, E; Wong, J; Gallia, G L; Belzberg, A J; Kimura, E T; Riggins, G J

    2014-05-01

    About one half of malignant peripheral nerve sheath tumors (MPNST) have Neurofibromin 1 (NF1) mutations. NF1 is a tumor suppressor gene essential for negative regulation of RAS signaling. Survival for MPNST patients is poor and we sought to identify an effective combination therapy. Starting with the mTOR inhibitors rapamycin and everolimus, we screened for synergy in 542 FDA approved compounds using MPNST cells with a native NF1 loss in both alleles. We further analyzed the cell cycle and signal transduction. In vivo growth effects of the drug combination with local radiation therapy (RT) were assessed in MPNST xenografts. The synergistic combination of mTOR inhibitors with bortezomib yielded a reduction in MPNST cell proliferation. The combination of mTOR inhibitors and bortezomib also enhanced the anti-proliferative effect of radiation in vitro. In vivo, the combination of mTOR inhibitor (everolimus) and bortezomib with RT decreased tumor growth and proliferation, and augmented apoptosis. The combination of approved mTOR and proteasome inhibitors with radiation showed a significant reduction of tumor growth in an animal model and should be investigated and optimized further for MPNST therapy.

  1. Novel agents and associated toxicities of inhibitors of the pi3k/Akt/mtor pathway for the treatment of breast cancer

    PubMed Central

    Chia, S.; Gandhi, S.; Joy, A.A.; Edwards, S.; Gorr, M.; Hopkins, S.; Kondejewski, J.; Ayoub, J.P.; Califaretti, N.; Rayson, D.; Dent, S.F.

    2015-01-01

    The pi3k/Akt/mtor (phosphatidylinositol 3 kinase/ Akt/mammalian target of rapamycin) signalling pathway is an established driver of oncogenic activity in human malignancies. Therapeutic targeting of this pathway holds significant promise as a treatment strategy. Everolimus, an mtor inhibitor, is the first of this class of agents approved for the treatment of hormone receptor–positive, human epidermal growth factor receptor 2–negative advanced breast cancer. Everolimus has been associated with significant improvements in progression-free survival; however, it is also associated with increased toxicity related to its specific mechanism of action. Methods A comprehensive review of the literature conducted using a focused medline search was combined with a search of current trials at http://ClinicalTrials.gov/. Summary tables of the toxicities of the various classes of pi3k/Akt/mtor inhibitors were created. A broad group of Canadian health care professionals was assembled to review the data and to produce expert opinion and summary recommendations for possible best practices in managing the adverse events associated with these pathway inhibitors. Results Differing toxicities are associated with the various classes of pi3k/Akt/mtor pathway inhibitors. The most common unique adverse events observed in everolimus clinical trials in breast cancer include stomatitis (all grades: approximately 60%), noninfectious pneumonitis (15%), rash (40%), hyperglycemia (15%), and immunosuppression (40%). To minimize grades 3 and 4 toxicities and to attempt to attain optimal outcomes, effective management of those adverse events is critical. Management should be interdisciplinary and should use approaches that include education, early recognition, active intervention, and potentially prophylactic strategies. Discussion Everolimus likely represents the first of many complex oral targeted therapies for the treatment of breast cancer. Using this agent as a template, it is essential to

  2. [Response to everolimus in patients with giant cell astrocytoma associated to tuberous sclerosis complex].

    PubMed

    Mateos-González, M Elena; López-Laso, Eduardo; Vicente-Rueda, Josefina; Camino-León, Rafael; Fernández-Ramos, Joaquín A; Baena-Gómez, M Auxiliadora; Peña-Rosa, M José

    2014-12-01

    Introduccion. Los astrocitomas subependimarios de celulas gigantes (SEGA) se presentan en el 5-20% de los pacientes con complejo esclerosis tuberosa (CET) y son los tumores cerebrales mas comunes en el CET. Son tumores benignos, de estirpe glioneural, que se desarrollan fundamentalmente en las primeras dos decadas de la vida, en general cercanos al foramen de Monro, y pueden ocasionar hidrocefalia e hipertension intracraneal. Constituyen la principal causa de muerte en el CET. Recientemente, los inhibidores mTOR han demostrado ser una alternativa terapeutica a la reseccion quirurgica. Objetivo. Describir nuestra experiencia con everolimus para el tratamiento de pacientes con SEGA y CET. Pacientes y metodos. Estudio prospectivo de la respuesta de los pacientes con CET y al menos un SEGA en crecimiento. Resultados. Recibieron tratamiento tres mujeres y tres varones con una edad media de 12,3 años. Un paciente habia sido previamente intervenido quirurgicamente por SEGA con hidrocefalia. El diametro maximo medio del SEGA al inicio del tratamiento era de 15,3 mm (rango: 11,3-24,8 mm). Se inicio tratamiento con everolimus, 2,5 mg/dia por via oral en pacientes con superficie corporal < 1,2 m2 y 5 mg/dia en pacientes con superficie corporal > 1,2 m2. Dos pacientes presentaron hipertrigliceridemia; uno, anorexia; otro, un afta; y una paciente, amenorrea. La reduccion media del volumen del SEGA a los tres meses de tratamiento fue del 46%, y la reduccion se mantuvo estable en controles posteriores (6-25 meses). Conclusiones. El tratamiento con everolimus disminuye el tamaño de los SEGA asociados a CET con un perfil de seguridad adecuado, y constituye una alternativa a la cirugia en casos seleccionados.

  3. Phase I Study of Everolimus in Combination with Gemcitabine and Split-Dose Cisplatin in Advanced Urothelial Carcinoma

    PubMed Central

    Abida, Wassim; Milowsky, Matthew I.; Ostrovnaya, Irina; Gerst, Scott R.; Rosenberg, Jonathan E.; Voss, Martin H.; Apolo, Andrea B.; Regazzi, Ashley M.; McCoy, Asia S.; Boyd, Mariel E.; Bajorin, Dean F.

    2016-01-01

    Background: Cisplatin-based combination chemotherapy is standard first-line treatment for patients with advanced urothelial carcinoma (UC). Molecular profiling studies reveal that the PI3K/AKT/mTOR pathway is altered in a significant percentage of UCs. Objective: We conducted a phase I trial to evaluate the feasibility of combining the mTOR inhibitor everolimus with gemcitabine and split-dose cisplatin (GC) in advanced UC in the first-line setting. Methods: Patients received gemcitabine 800 mg/m2 and cisplatin 35 mg/m2 on days 1 and 8 of 21-day cycles for a total of 6 cycles in combination with everolimus at increasing dose levels (DL1:5 mg QOD, DL2:5 mg daily, DL3:10 mg daily) following a standard 3+3 design. Responses were assessed every 2 cycles. Patients with at least stable disease (SD) continued everolimus until progression. Goals were to establish dose-limiting toxicities (DLTs) and maximum tolerated dose (MTD) for the combination. Results: 12 patients were enrolled, 3 at DL1, 3 at DL2, and an additional 6 at DL1 *(DL1 following de-escalation). 3/3 patients at DL2 had DLTs during cycle 1. 2/8 evaluable patients at DL1/DL1 * had DLTs during cycle 1. DLTs were primarily hematologic. Further toxicities, also primarily hematologic, were observed during later treatment cycles, leading to 8 chemotherapy dose reductions overall. Partial responses were observed in 4/10 evaluable patients, and SD in 5/10. Median overall survival was 10.8 months (95% CI 6.9, not reached). Conclusions: The maximum tolerated dose was reached at the lowest dose level, 5 mg QOD, for everolimus in combination with gemcitabine and split-dose cisplatin in advanced UC. The regimen was limited by hematologic toxicity. PMID:27376132

  4. Rationale and protocol of the MetNET-1 trial, a prospective, single center, phase II study to evaluate the activity and safety of everolimus in combination with octreotide LAR and metformin in patients with advanced pancreatic neuroendocrine tumors.

    PubMed

    Pusceddu, Sara; de Braud, Filippo; Concas, Laura; Bregant, Cristina; Leuzzi, Livia; Formisano, Barbara; Buzzoni, Roberto

    2014-01-01

    Abnormal PI3K-AKT-mTOR pathway signalling and autocrine activation of the mTOR pathway, mediated through insulin-like growth factor-1, have been implicated in the proliferation of pancreatic neuroendocrine tumor (pNET) cells. Everolimus, an mTOR inhibitor, has shown antitumor benefit in pNETs alone and in combination with octreotide LAR in RADIANT-1 and RADIANT-3 studies. Although everolimus-based phase II/III trials have improved progression-free survival for pNET, its use has not impacted on prolonging overall survival. Metformin has recently shown some anti-cancer activity in both in vitro and in vivo studies by its indirect properties to decrease insulin and insulin-like growth factor-1 (IGF-1) levels and by its antitumour effect to promote AMPK activation and consequently inhibition to TSC1-2/mTOR complex. In light of even more retrospective evidence of metformin's anticancer activity, a prospective evaluation is required to either confirm or discard these preliminary findings. With the aim to evaluate the antiproliferative effect of metformin in combination with everolimus and octreotide LAR in pancreatic well-differentiated neuroendocrine tumor patients, a single arm, prospective, single center phase II study was designed (MetNET-1 trial, NCT 02294006). Forty-three patients are expected to be evaluated. The study is ongoing, and recruitment is estimated to be completed in August 2016. The results will be anticipated in 2017.

  5. Everolimus in patients with metastatic renal cell carcinoma previously treated with bevacizumab: a prospective multicenter study CRAD001LRU02T.

    PubMed

    Tsimafeyeu, Ilya; Snegovoy, Anton; Varlamov, Sergei; Safina, Sufia; Varlamov, Ilya; Gurina, Ludmila; Manzuk, Ludmila

    2015-09-01

    Everolimus is an orally administered inhibitor of the mammalian target of rapamycin (mTOR) recommended for patients with metastatic renal cell carcinoma (mRCC) who progressed on previous vascular endothelial growth factor (VEGF) receptor-tyrosine kinase inhibitor therapy. Efficacy of everolimus in patients who progressed on anti-VEGF monoclonal antibody bevacizumab is unknown. We did a multicenter prospective trial of everolimus in patients with mRCC whose disease had progressed on bevacizumab ± interferon alpha (IFN). Patients with clear-cell mRCC which had progressed on bevacizumab ± IFN received everolimus 10 mg once daily. The primary end point was the proportion of patients remaining progression-free for 56 days, and a two-stage Simon design was used, with 80% power and an alpha risk of 5%. This study is registered with ClinicalTrials.gov, number NCT02056587. From December 2011 to October 2013, a total of 37 patients (28 M, 9 F) were enrolled. Median age was 60.5 years (range 41-66), 1% had Eastern Cooperative Oncology Group Performance Status (ECOG PS) >2, and Memorial Sloan-Kettering Cancer Center (MSKCC) favorable/intermediate risk was 38/62%. Five (14%) patients had a confirmed partial response and 26 (70%) patients had a stable disease. Median progression-free survival was 11.5 months (95% CI, 8.8-14.2). Median overall survival was not reached. No grade 3 or 4 treatment-related toxicities were observed. The most common grade 2 adverse events were fatigue (19%) and pneumonitis (8%). Everolimus demonstrated a favorable toxicity profile and promising anti-tumor activity as a second-line therapy in metastatic renal cell carcinoma (RCC) patients previously treated with bevacizumab ± IFN.

  6. The Role of Everolimus in Renal Cell Carcinoma

    PubMed Central

    Valdivieso, Roger; Dell’Oglio, Paolo; Trudeau, Vincent; Larcher, Alessandro; Karakiewicz, Pierre I.

    2015-01-01

    Everolimus (RAD001) is an orally administered agent that inhibits the mammalian target of rapamycin serine-threonine kinase. A phase III pivotal trial on everolimus, published in 2008, provided the first evidence for the efficacy of sequential therapy for patients with metastatic clear cell renal cell carcinoma (RCC). In this study, everolimus was used after failure of one or several previous lines of therapy, and it demonstrated a 3-month survival benefit relative to placebo. Currently, based on the level 1 evidence, everolimus represents the molecule of choice for third-line therapy after failure of previous two tyrosine kinase inhibitors (TKIs). However, second-line use after failure of one TKI is challenged by two new molecules (nivolumab and cabozantinib), which proved to have better efficacy with similar toxicity profile. In non-clear cell metastatic RCC, the current evidence recommends everolimus as a second-line therapy after failure of previous first-line sunitinib.

  7. Addition of Everolimus Post VEGFR Inhibition Treatment Failure in Advanced Sarcoma Patients Who Previously Benefited from VEGFR Inhibition: A Case Series

    PubMed Central

    Hays, John L.; Chen, James L.

    2016-01-01

    Background Patients with metastatic sarcoma who progress on vascular endothelial growth factor receptor inhibitors (VEGFRi) have limited treatment options. Upregulation of the mTOR pathway has been demonstrated to be a means of resistance to targeted VEGFRi in metastatic sarcoma. Patients and methods Retrospective cohort study to evaluate the clinical benefit at four months of combining mTOR inhibition (mTORi) via everolimus with VEGFRi in patients who have derived benefit from single-agent VEGFRi but have progressed. Patients with recurrent, metastatic soft tissue or bone sarcomas who progressed after deriving clinical benefit to VEGFRi beyond 12 weeks were continued on VEGFRi with the addition of everolimus (5 mg daily). Progression free survival was measured from start of VEGFRi to disease progression on single agent VEGFRi as well as from the addition of everolimus therapy to disease progression or drug discontinuation due to toxicity. Clinical benefit was defined as stable disease or partial response at 4 months. Results Nine patients were evaluated. Two patients did not tolerate therapy due to GI toxicity and one elected to discontinue therapy. Of the remaining six patients, the clinical benefit rate at four months was 50%. Progression free survival (PFS) for these patients was 3.1 months ranging from 0.5 to 7.2 months with one patient remaining on combination therapy. Conclusion In this heavily pre-treated, advanced sarcoma population, the addition of mTOR inhibition to VEGFRi based therapy resulted in a clinical benefit for a subset of patients. Prospective studies will be needed to verify these results. PMID:27295141

  8. The Effect of Everolimus in an In Vitro Model of Triple Negative Breast Cancer and Osteoclasts

    PubMed Central

    Mercatali, Laura; Spadazzi, Chiara; Miserocchi, Giacomo; Liverani, Chiara; De Vita, Alessandro; Bongiovanni, Alberto; Recine, Federica; Amadori, Dino; Ibrahim, Toni

    2016-01-01

    Metastatic bone disease has a major impact on morbidity of breast cancer (BC) patients. Alterations in mTOR signaling are involved both in cancer progression and in osteoclast differentiation. The purpose of this study was to assess the role of mTOR inhibitor Everolimus (Eve) on osteoclastogenesis induced by triple negative BC cells. To this aim, we developed an in vitro human model of osteoclastogenesis from peripheral blood monocytes co-cultured with the triple negative SCP2 and the hormonal receptor positive MCF7 cell lines. Osteoclastogenesis was evaluated by TRAP staining, evaluation of F actin rings and Calcitonin Receptor expression. Eve significantly reduced differentiation induced by cancer cells and resulted more effective when evaluated in combination with Denosumab and Zoledronic Acid (Zol). Combination with Zol showed a total abrogation of osteoclast differentiation induced by the triple negative cell line, not by MCF7. Finally, we observed that Eve was active in the inhibition of the crosstalk between cancer cells and osteoclasts reproduced by our model, highlighting a new therapeutic choice for the subsetting of triple negative BC patients. We observed a difference in the response to bone-targeted therapy with respect to BC subtypes. Our model may represent a valid platform for preclinical trials on bone-targeted drugs and for the study of the interplay of BC with bone stromal cells. PMID:27809291

  9. Efficacy and safety of concentration-controlled everolimus with reduced-dose cyclosporine in Japanese de novo renal transplant patients: 12-month results

    PubMed Central

    2013-01-01

    Background No study to date has evaluated the efficacy and safety of everolimus with reduced-exposure cyclosporine in Japanese de-novo renal transplant (RTx) patients. Methods This 12-month, multicenter, open-label study randomized (1:1) 122 Japanese de-novo RTx patients to either an everolimus regimen (1.5 mg/day starting dose (target trough: 3 to 8 ng/ml) + reduced-dose cyclosporine) or a mycophenolate mofetil (MMF) regimen (2 g/day + standard dose cyclosporine). All patients received basiliximab and corticosteroids. Key endpoints at month 12 were composite efficacy failure (treated biopsy-proven acute rejection, graft loss, death, or loss to follow-up) and renal function (estimated glomerular filtration rate; Modification of Diet in Renal Disease-4). Results Clear cyclosporine exposure reduction was achieved in the everolimus group throughout the study (52% reduction at month 12). Month 12 efficacy failure rates showed everolimus 1.5 mg to be non-inferior to MMF (11.5% vs. 11.5%). The median estimated glomerular filtration rate at month 12 was 58.00 ml/minute/1.73 m2 in the everolimus group versus 55.25 ml/minute/1.73 m2 in the MMF group (P = 0.063). Overall, the incidence of adverse events was comparable between the groups with some differences in line with the known safety profile of the treatments. The everolimus group had a higher incidence of wound healing events and edema, whereas a higher rate of cytomegalovirus infections was reported in the MMF group. Conclusions This study confirmed the efficacy of everolimus 1.5 mg/day (target trough: 3 to 8 ng/ml) in Japanese RTx patients for preventing acute rejection, while allowing for substantial cyclosporine sparing. Renal function and safety findings were comparable with previous reports from other RTx populations. Trial registration ClinicalTrials.gov number: NCT00658320 PMID:23866828

  10. Everolimus immunosuppression in kidney transplantation: What is the optimal strategy?

    PubMed

    Witzke, Oliver; Sommerer, Claudia; Arns, Wolfgang

    2016-01-01

    Two main everolimus-based strategies have been pursued to facilitate calcineurin inhibitor (CNI) reduction after kidney transplantation: (i) everolimus with reduced CNI exposure from time of transplant and (ii) pre-emptive introduction of everolimus with CNI reduction or withdrawal at some point post-transplant. Randomized trials have shown no loss of immunosuppressive efficacy for everolimus (targeting 3-8 ng/mL) with reduced-exposure CNI versus standard-exposure CNI and mycophenolic acid (MPA) in low-to-moderate risk patients. Renal function has tended to be numerically, but not significantly, higher with everolimus and reduced-CNI versus MPA and standard-CNI. One study which used very low CsA exposure in everolimus-treated patients reported a substantial improvement in estimated GFR compared to controls, but this requires confirmation. Pre-emptive conversion to everolimus at three to six months after kidney transplantation significantly improves long-term renal function, but with an increased rate of mild acute rejection. Earlier conversion (up to two months post-transplant) can lead to an increase in rejection risk, while later conversion (more than six months post-transplant) is unproductive unless baseline renal function is good. This article considers the risks and benefits associated with either strategy, and reviews specific clinical situations that influence the optimal approach in individual patients. The balance of evidence suggests two options. De novo everolimus with reduced CNI, steroids and induction therapy ensures immunosuppressive efficacy in low- or standard-risk populations, and investigations into this strategy are ongoing. Conversion to everolimus with CNI withdrawal between three and six months post-transplant offers a long-term renoprotective effect if baseline graft function is good.

  11. Paclitaxel, bevacizumab, and everolimus/placebo as first-line treatment for patients with metastatic HER2-negative breast cancer: a randomized placebo-controlled phase II trial of the Sarah Cannon Research Institute.

    PubMed

    Yardley, Denise A; Bosserman, Linda D; O'Shaughnessy, Joyce A; Harwin, William N; Morgan, Susan K; Priego, Victor M; Peacock, Nancy W; Bass, J David; Burris, Howard A; Hainsworth, John D

    2015-11-01

    Amplified PI3K/Akt/mTOR signaling is common in metastatic breast cancer (MBC). The mTOR inhibitor everolimus improves progression-free survival (PFS) when added to steroidal aromatase inhibitor therapy. This randomized phase II trial compares the efficacy of paclitaxel/bevacizumab/everolimus and paclitaxel/bevacizumab/placebo as first-line treatment for MBC. Patients with untreated HER2-negative MBC were randomized (1:1) to receive 28-day cycles of paclitaxel 90 mg/m(2) IV (days 1, 8, and 15) and bevacizumab 10 mg/kg IV (days 1, 15) with either everolimus 10 mg (Arm 1) or placebo (Arm 2) daily. Treatment continued (evaluation every 8 weeks) until progression or unacceptable toxicity. Treatment of 110 patients allowed detection of an improvement in median PFS from 11 to 16 months (70 % power, α = 0.10). Between August 2009 and June 2011, 113 patients (median age 58 years; 88 % ER or PR positive) were randomized (Arm 1, 56; Arm 2, 57). Patients in both arms received a median of six treatment cycles. Median PFS (95 % CI) was 9.1 months (6.8-18.8) for Arm 1, and 7.1 months (5.6-10.8) for Arm 2 (p = 0.89). Comparisons of other efficacy endpoints were also similar in the two treatment arms. Patients receiving everolimus had more anemia, stomatitis, diarrhea, rash, and arthralgia/myalgia, although the overall incidence of severe (grade 3/4) toxicity was similar. The addition of everolimus did not improve the efficacy of weekly paclitaxel/bevacizumab as first-line treatment for patients with HER2-negative MBC. These results contrast with the demonstrated efficacy of adding everolimus to either hormonal or HER2-targeted therapy in previously treated patients.

  12. Phosphorylation of mTOR Ser2481 is a key target limiting the efficacy of rapalogs for treating hepatocellular carcinoma

    PubMed Central

    Shibata, Tomohiro; Noda, Masaki; Kawahara, Akihiko; Akiba, Jun; Murakami, Yuichi; Yano, Hirohisa; Kuwano, Michihiko; Ono, Mayumi

    2016-01-01

    Hepatocellular carcinoma (HCC) is one of the most common cancers worldwide. Although recent studies facilitate the identification of crucial genes and relevant regulatory pathways, therapeutic approaches against advanced HCC are insufficiently effective. Therefore, we aimed here to develop potent therapeutics to provide a reliable biomarker for the therapeutic efficacy in patients with HCC. To this end, we first compared the cytotoxic effects of various anti-cancer drugs between well differentiated (HAK-1A) and poorly differentiated (HAK-1B) cell lines established from a single HCC tumor. Of various drug screened, HAK-1B cells were more sensitive by a factor of 2,000 to the mTORC1 inhibitors (rapalogs), rapamycin and everolimus, than HAK-1A cells. Although rapalogs inhibited phosphorylation of mTOR Ser2448 in HAK-1A and HAK-1B cells, phosphorylation of mTOR Ser2481 was specifically inhibited only in HAK-1B cells. Silencing of Raptor induced apoptosis and inhibited the growth of only HAK-1B cells. Further, three other cell lines established independently from the tumors of three patients with HCC were also approximately 2,000-fold times more sensitive to rapamycin, which correlated closely with the inhibition of mTOR Ser2481 phosphorylation by rapamycin. Treatment with everolimus markedly inhibited the growth of tumors induced by poorly differentiated HAK-1B and KYN-2 cells and phosphorylation of mTOR Ser2481 in vivo. To our knowledge, this is the first study showing that the phosphorylation of mTOR Ser2481 is selectively inhibited by rapalogs in mTORC1-addicted HCC cells and may be a potential reliable biomarker for the therapeutic efficacy of rapalogs for treating HCC patients. PMID:27329724

  13. Granuloma-forming interstitial pneumonia occurring one year after the start of everolimus therapy.

    PubMed

    Saito, Yoshinobu; Kunugi, Shinobu; Suzuki, Yasutomo; Narita, Kousuke; Miura, Yukiko; Minegishi, Yuji; Kimura, Go; Kondo, Yukihiro; Azuma, Arata; Fukuda, Yuh; Gemma, Akihiko

    2013-01-01

    We experienced a case of interstitial lung disease (ILD) that occurred one year after the start of everolimus therapy for renal cell carcinoma. The pathological features included interstitial pneumonia with granuloma formation. Everolimus is known to cause ILD; however, its pathology is unclear. Granuloma-forming interstitial pneumonia associated with everolimus is uncommon, although it may be one of the pathological patterns associated with everolimus-induced ILD. This is a slow-onset case of everolimus-induced ILD in a patient with renal cell carcinoma. Physicians should thus be aware of the potential for the development of ILD at any time during the administration of everolimus therapy.

  14. Analytical performance of a new liquid chromatography/tandem mass spectrometric method for determination of everolimus concentrations in whole blood.

    PubMed

    McMillin, Gwendolyn A; Johnson-Davis, Kamisha; Dasgupta, Amitava

    2012-04-01

    The immunosuppressant everolimus was recently approved for prophylactic use in the United States, to prevent organ rejection in adult kidney transplant recipients. The currently accepted therapeutic range for everolimus is 3-8 ng/mL. Therapeutic drug monitoring (TDM) using predose EDTA whole blood samples is required to optimize dose. We describe a simple extraction method and analysis by liquid chromatography tandem mass spectrometry (LC-MS/MS) to support routine TDM of everolimus. Samples were prepared by protein precipitation and filtration. The first quadrupole was set to select the ammonium adducts (Equation is included in full-text article.)of everolimus (m/z 975.62) and rapamycin-d3 (m/z 934.70), the internal standard. The second quadrupole was used as a collision chamber, and the third quadrupole was then used to select characteristic product ions of everolimus (m/z 908.50 and 890.50) and rapamycin-d3 (m/z 864.60 and 846.50). The method had an analytical measurement range of 2.0-150 ng/mL. Total imprecision, expressed as percent coefficient of variation (mean concentration), was 19.1% (3.3 ng/mL), 10.6% (5.9 ng/mL), 8.1% (19.2 ng/mL), 5.7% (25.8 ng/mL), and 9.1% (34.2 ng/mL). The new method was compared with 2 other everolimus methods also based on LC-MS/MS, with 64 residual patient specimens. Agreement, based on simple linear regression, was excellent. Method A comparison: y = 0.96x - 1.12 (r = 0.99), n = 20, 2.5-44.7 ng/mL. Method B comparison: y = 0.96x + 0.49 (r = 0.99), n = 44, 2.1-85.6 ng/mL. We conclude that this method could support TDM of everolimus for a wide range of clinical indications.

  15. The Changes of Lipid Metabolism in Advanced Renal Cell Carcinoma Patients Treated with Everolimus: A New Pharmacodynamic Marker?

    PubMed Central

    Pantano, Francesco; Santoni, Matteo; Procopio, Giuseppe; Rizzo, Mimma; Iacovelli, Roberto; Porta, Camillo; Conti, Alessandro; Lugini, Antonio; Milella, Michele; Galli, Luca; Ortega, Cinzia; Guida, Francesco Maria; Silletta, Marianna; Schinzari, Giovanni; Verzoni, Elena; Modica, Daniela; Crucitti, Pierfilippo; Rauco, Annamaria; Felici, Alessandra; Ballatore, Valentina; Cascinu, Stefano; Tonini, Giuseppe; Carteni, Giacomo; Russo, Antonio; Santini, Daniele

    2015-01-01

    Background Everolimus is a mammalian target of rapamycin (mTOR) inhibitor approved for the treatment of metastatic renal cell carcinoma (mRCC). We aimed to assess the association between the baseline values and treatmentrelated modifications of total serum cholesterol (C), triglycerides (T), body mass index (BMI), fasting blood glucose level (FBG) and blood pressure (BP) levels and the outcome of patients treated with everolimus for mRCC. Methods 177 patients were included in this retrospective analysis. Time to progression (TTP), clinical benefit (CB) and overall survival (OS) were evaluated. Results Basal BMI was significantly higher in patients who experienced a CB (p=0,0145). C,T and C+T raises were significantly associated with baseline BMI (p=0.0412, 0.0283 and 0.0001). Median TTP was significantly longer in patients with T raise compared to patients without T (10 vs 6, p=0.030), C (8 vs 5, p=0.042) and C+T raise (10.9 vs 5.0, p=0.003). At the multivariate analysis, only C+T increase was associated with improved TTP (p=0.005). T raise (21.0 vs 14.0, p=0.002) and C+T increase (21.0 vs 14.0, p=0.006) were correlated with improved OS but were not significant at multivariate analysis. Conclusion C+T raise is an early predictor for everolimus efficacy for patients with mRCC. PMID:25885920

  16. Osteonecrosis of the jaw associated with everolimus: A case report

    PubMed Central

    Yamamoto, Daigo; Tsubota, Yu; Utsunomiya, Toshiki; Sueoka, Noriko; Ueda, Aiko; Endo, Kayoko; Yoshikawa, Katsuhiro; Kon, Masanori

    2017-01-01

    Everolimus, a mammalian target of rapamycin inhibitor, has recently been approved for the treatment of metastatic estrogen receptor-positive breast cancer, at a daily dose of 10 mg in combination with exemestane. Osteonecrosis of the jaw (ONJ) is a rare but severe condition, characterized by exposed necrotic bone, and is associated with various drugs that are often used to treat advanced malignancies. We herein report the case of a patient with breast cancer who developed ONJ during treatment with everolimus, which improved after discontinuation of the drug. To the best of our knowledge, this is the first reported case of everolimus-associated ONJ in a patient receiving everolimus for metastatic breast cancer. In 2014, an 80-year-old woman was started on treatment with everolimus and exemestane for stage IIB estrogen receptor-positive breast cancer. Within 2 months, the left side of her face became edematous, with localized heat and tenderness of the left mandibular region and a 3-mm round area of exposed bone. There was purulent discharge and the surrounding gingiva was edematous and erythematous. The left mandible exhibited a low signal intensity area on T1-weighted magnetic resonance imaging. Treatment was discontinued and ONJ showed improvement after 2 months. Therefore, when prescribing everolimus for metastatic breast cancer, oncologists should be aware of the possibility of ONJ as a complication. PMID:28357105

  17. Activating the translational repressor 4E-BP or reducing S6K-GSK3β activity prevents accelerated axon growth induced by hyperactive mTOR in vivo

    PubMed Central

    Gong, Xuan; Zhang, Longbo; Huang, Tianxiang; Lin, Tiffany V.; Miyares, Laura; Wen, John; Hsieh, Lawrence; Bordey, Angélique

    2015-01-01

    Abnormal axonal connectivity and hyperactive mTOR complex 1 (mTORC1) are shared features of several neurological disorders. Hyperactive mTORC1 alters axon length and polarity of hippocampal neurons in vitro, but the impact of hyperactive mTORC1 on axon growth in vivo and the mechanisms underlying those effects remain unclear. Using in utero electroporation during corticogenesis, we show that increasing mTORC1 activity accelerates axon growth without multiple axon formation. This was prevented by counteracting mTORC1 signaling through p70S6Ks (S6K1/2) or eukaryotic initiation factor 4E-binding protein (4E-BP1/2), which both regulate translation. In addition to regulating translational targets, S6K1 indirectly signals through GSK3β, a regulator of axogenesis. Although blocking GSK3β activity did not alter axon growth under physiological conditions in vivo, blocking it using a dominant-negative mutant or lithium chloride prevented mTORC1-induced accelerated axon growth. These data reveal the contribution of translational and non-translational downstream effectors such as GSK3β to abnormal axon growth in neurodevelopmental mTORopathies and open new therapeutic options for restoring long-range connectivity. PMID:26220974

  18. The Effect of Everolimus Initiation and Calcineurin Inhibitor Elimination on Cardiac Allograft Vasculopathy in De Novo Recipients: One-Year Results of a Scandinavian Randomized Trial.

    PubMed

    Arora, S; Andreassen, A K; Andersson, B; Gustafsson, F; Eiskjaer, H; Bøtker, H E; Rådegran, G; Gude, E; Ioanes, D; Solbu, D; Sigurdardottir, V; Dellgren, G; Erikstad, I; Solberg, O G; Ueland, T; Aukrust, P; Gullestad, L

    2015-07-01

    Early initiation of everolimus with calcineurin inhibitor therapy has been shown to reduce the progression of cardiac allograft vasculopathy (CAV) in de novo heart transplant recipients. The effect of de novo everolimus therapy and early total elimination of calcineurin inhibitor therapy has, however, not been investigated and is relevant given the morbidity and lack of efficacy of current protocols in preventing CAV. This 12-month multicenter Scandinavian trial randomized 115 de novo heart transplant recipients to everolimus with complete calcineurin inhibitor elimination 7-11 weeks after HTx or standard cyclosporine immunosuppression. Ninety-five (83%) patients had matched intravascular ultrasound examinations at baseline and 12 months. Mean (± SD) recipient age was 49.9 ± 13.1 years. The everolimus group (n = 47) demonstrated significantly reduced CAV progression as compared to the calcineurin inhibitor group (n = 48) (ΔMaximal Intimal Thickness 0.03 ± 0.06 and 0.08 ± 0.12 mm, ΔPercent Atheroma Volume 1.3 ± 2.3 and 4.2 ± 5.0%, ΔTotal Atheroma Volume 1.1 ± 19.2 mm(3) and 13.8 ± 28.0 mm(3) [all p-values ≤ 0.01]). Everolimus patients also had a significantly greater decline in levels of soluble tumor necrosis factor receptor-1 as compared to the calcineurin inhibitor group (p = 0.02). These preliminary results suggest that an everolimus-based CNI-free can potentially be considered in suitable de novo HTx recipients.

  19. mTOR inhibitors counteract tamoxifen-induced activation of breast cancer stem cells.

    PubMed

    Karthik, Govindasamy-Muralidharan; Ma, Ran; Lövrot, John; Kis, Lorand Levente; Lindh, Claes; Blomquist, Lennart; Fredriksson, Irma; Bergh, Jonas; Hartman, Johan

    2015-10-10

    Breast cancer cells with stem cell characteristics (CSC) are a distinct cell population with phenotypic similarities to mammary stem cells. CSCs are important drivers of tumorigenesis and the metastatic process. Tamoxifen is the most widely used hormonal therapy for estrogen receptor (ER) positive cancers. In our study, tamoxifen was effective in reducing proliferation of ER + adherent cancer cells, but not their CSC population. We isolated, expanded and incubated CSC from seven breast cancers with or without tamoxifen. By genome-wide transcriptional analysis we identified tamoxifen-induced transcriptional pathways associated with ribosomal biogenesis and mRNA translation, both regulated by the mTOR-pathway. We observed induction of the key mTOR downstream targets S6K1, S6RP and 4E-BP1 in-patient derived CSCs by tamoxifen on protein level. Using the mTOR inhibitors rapamycin, everolimus and PF-04691502 (a dual PI3K/mTOR inhibitor) and in combination with tamoxifen, significant reduction in mammosphere formation was observed. Hence, we suggest that the CSC population play a significant role during endocrine resistance through activity of the mTOR pathway. In addition, tamoxifen further stimulates the mTOR-pathway but can be antagonized using mTOR-inhibitors.

  20. Mutant p53 proteins counteract autophagic mechanism sensitizing cancer cells to mTOR inhibition.

    PubMed

    Cordani, Marco; Oppici, Elisa; Dando, Ilaria; Butturini, Elena; Dalla Pozza, Elisa; Nadal-Serrano, Mercedes; Oliver, Jordi; Roca, Pilar; Mariotto, Sofia; Cellini, Barbara; Blandino, Giovanni; Palmieri, Marta; Di Agostino, Silvia; Donadelli, Massimo

    2016-08-01

    Mutations in TP53 gene play a pivotal role in tumorigenesis and cancer development. Here, we report that gain-of-function mutant p53 proteins inhibit the autophagic pathway favoring antiapoptotic effects as well as proliferation of pancreas and breast cancer cells. We found that mutant p53 significantly counteracts the formation of autophagic vesicles and their fusion with lysosomes throughout the repression of some key autophagy-related proteins and enzymes as BECN1 (and P-BECN1), DRAM1, ATG12, SESN1/2 and P-AMPK with the concomitant stimulation of mTOR signaling. As a paradigm of this mechanism, we show that atg12 gene repression was mediated by the recruitment of the p50 NF-κB/mutant p53 protein complex onto the atg12 promoter. Either mutant p53 or p50 NF-κB depletion downregulates atg12 gene expression. We further correlated the low expression levels of autophagic genes (atg12, becn1, sesn1, and dram1) with a reduced relapse free survival (RFS) and distant metastasis free survival (DMFS) of breast cancer patients carrying TP53 gene mutations conferring a prognostic value to this mutant p53-and autophagy-related signature. Interestingly, the mutant p53-driven mTOR stimulation sensitized cancer cells to the treatment with the mTOR inhibitor everolimus. All these results reveal a novel mechanism through which mutant p53 proteins promote cancer cell proliferation with the concomitant inhibition of autophagy.

  1. Metabolic response to everolimus in patient-derived triple negative breast cancer xenografts.

    PubMed

    Euceda, Leslie R; Hill, Deborah K; Stokke, Endre; Hatem, Rana; Botty, Rania El; Bièche, Ivan; Marangoni, Elisabetta; Bathen, Tone F; Moestue, Siver A

    2017-03-14

    Patients with triple negative breast cancer (TNBC) are unresponsive to endocrine and anti-HER2 pharmacotherapy, limiting their therapeutic options to chemotherapy. TNBC is frequently associated with abnormalities in the PI3K/AKT/mTOR signaling pathway; drugs targeting this pathway are currently being evaluated in these patients. However, response is variable, partly due to heterogeneity within TNBC, conferring a need to identify biomarkers predicting response and resistance to targeted therapy. In this study, we used a metabolomics approach to assess response to the mTOR inhibitor everolimus in a panel of TNBC patient-derived xenografts (PDX) (n=103 animals). Tumor metabolic profiles were acquired using high-resolution magic angle spinning magnetic resonance spectroscopy. Partial least squares-discriminant analysis on relative metabolite concentrations discriminated treated xenografts from untreated controls with an accuracy of 67% (p=0.003). Multilevel linear mixed-effects models (LMM) indicated reduced glycolytic lactate production and glutaminolysis after treatment, consistent with PI3K/AKT/mTOR pathway inhibition. Although inherent metabolic heterogeneity between different PDX models seemed to hinder prediction of treatment response, the metabolic effects following treatment were more pronounced in responding xenografts compared to non-responders. Additionally, the metabolic information predicted p53 mutation status, which may provide complimentary insight into the interplay between PI3K signaling and other drivers of disease progression.

  2. Sirolimus and Everolimus Pathway: Reviewing Candidate Genes Influencing Their Intracellular Effects

    PubMed Central

    Granata, Simona; Dalla Gassa, Alessandra; Carraro, Amedeo; Brunelli, Matteo; Stallone, Giovanni; Lupo, Antonio; Zaza, Gianluigi

    2016-01-01

    Sirolimus (SRL) and everolimus (EVR) are mammalian targets of rapamycin inhibitors (mTOR-I) largely employed in renal transplantation and oncology as immunosuppressive/antiproliferative agents. SRL was the first mTOR-I produced by the bacterium Streptomyces hygroscopicus and approved for several medical purposes. EVR, derived from SRL, contains a 2-hydroxy-ethyl chain in the 40th position that makes the drug more hydrophilic than SRL and increases oral bioavailability. Their main mechanism of action is the inhibition of the mTOR complex 1 and the regulation of factors involved in a several crucial cellular functions including: protein synthesis, regulation of angiogenesis, lipid biosynthesis, mitochondrial biogenesis and function, cell cycle, and autophagy. Most of the proteins/enzymes belonging to the aforementioned biological processes are encoded by numerous and tightly regulated genes. However, at the moment, the polygenic influence on SRL/EVR cellular effects is still not completely defined, and its comprehension represents a key challenge for researchers. Therefore, to obtain a complete picture of the cellular network connected to SRL/EVR, we decided to review major evidences available in the literature regarding the genetic influence on mTOR-I biology/pharmacology and to build, for the first time, a useful and specific “SRL/EVR genes-focused pathway”, possibly employable as a starting point for future in-depth research projects. PMID:27187382

  3. Recent Advances and Challenges of mTOR Inhibitors Use in the Treatment of Patients with Tuberous Sclerosis Complex

    PubMed Central

    Robalo, Conceição

    2017-01-01

    Tuberous sclerosis complex (TSC) is a genetic condition characterized by the presence of benign, noninvasive, and tumor-like lesions called hamartomas that can affect multiple organ systems and are responsible for the clinical features of the disease. In the majority of cases, TSC results from mutations in the TSC1 and TSC2 genes, leading to the overactivation of the mammalian target of rapamycin (mTOR) signalling pathway, which controls several cell functions, including cell growth, proliferation, and survival. The establishment of a connection between TSC and mTOR led to the clinical use of drugs known as mTOR inhibitors (like rapamycin, also known as sirolimus and everolimus), which are becoming an increasingly interesting tool in the management of TSC-associated features, such as subependymal giant cell astrocytomas, renal angiomyolipomas, and also epilepsy. However, the intrinsic characteristics of these drugs and their systemic effects in such a heterogeneous condition pose many challenges in clinical practice, so that some questions remain unanswered. This article provides an overview of the pharmacological aspects of mTOR inhibitors about the clinical trials leading to their approval in TSC-related conditions and exposes current challenges and future directions associated with this promising therapeutic line. PMID:28386314

  4. PTEN Loss Does Not Predict for Response to RAD001 (Everolimus) in a Glioblastoma Orthotopic Xenograft Test Panel

    PubMed Central

    Yang, Lin; Clarke, Michelle J.; Carlson, Brett L.; Mladek, Ann C.; Schroeder, Mark A.; Decker, Paul; Wu, Wenting; Kitange, Gaspar J.; Grogan, Patrick T.; Goble, Jennie M.; Uhm, Joon; Galanis, Evanthia; Giannini, Caterina; Lane, Heidi A.; James, C. David; Sarkaria, Jann N.

    2014-01-01

    Purpose Hyperactivation of the phosphatidylinositol 3-kinase/Akt signaling through disruption of PTEN function is common in glioblastoma multiforme, and these genetic changes are predicted to enhance sensitivity to mammalian target of rapamycin (mTOR) inhibitors such as RAD001 (everolimus). Experimental Design To test whether PTEN loss could be used as a predictive marker for mTOR inhibitor sensitivity, the response of 17 serially transplantable glioblastoma multiforme xenografts was evaluated in an orthotopic therapy evaluation model. Of these 17 xenograft lines, 7 have either genomic deletion or mutation of PTEN. Results Consistent with activation of Akt signaling, there was a good correlation between loss of PTEN function and elevated levels of Akt phosphorylation. However, of the 7 lines with disrupted PTEN function, only 1 tumor line (GBM10) was significantly sensitive to RAD001 therapy (25% prolongation in median survival), whereas1 of 10 xenograft lines with wild-type PTEN was significantly sensitive to RAD001 (GS22; 34% prolongation in survival). Relative to placebo, 5 days of RAD001 treatment was associated with a marked 66% reduction in the MIB1 proliferation index in the sensitive GBM10 line (deleted PTEN) compared with a 25% and 7% reduction in MIB1 labeling index in the insensitive GBM14 (mutant PTEN) and GBM15 (wild-type PTEN) lines, respectively. Consistent with a cytostatic antitumor effect, bioluminescent imaging of luciferase-transduced intracranial GBM10 xenografts showed slowed tumor growth without significant tumor regression during RAD001 therapy. Conclusion These data suggest that loss of PTEN function is insufficient to adequately predict responsiveness to mTOR inhibitors in glioblastoma multiforme. PMID:18559622

  5. Conversion from a calcineurin inhibitor-based immunosuppressive regimen to everolimus in renal transplant recipients: effect on renal function and proteinuria.

    PubMed

    Morales, J; Fierro, A; Benavente, D; Zehnder, C; Ferrario, M; Contreras, L; Herzog, C; Buckel, E

    2007-04-01

    New immunosuppressive agents are being actively researched to avoid complications of chronic allograft nephropathy (CAN), calcineurin inhibitor (CNI) nephrotoxicity, and posttransplantation cancer. The family of mTOR inhibitors offers a unique immunosuppressive opportunity to avoid CNI toxicity and reduce the incidence of malignancy. Nevertheless, increasing data have demonstrated that sirolimus (SRL), the first mTOR introduced in the treatment of solid organ transplant recipients, induces proteinuria, an adverse event that could produce deterioration of long-term renal function. In this short-term study of patients followed for 1 to 16 months, we examined changes in renal function and proteinuria among renal transplant recipients converted from a CNI-based regimen to an everolimus (EVL)-based one, a recently introduced mTOR inhibitor. Our data showed that renal function can be optimized after conversion to EVL by up to 42% in recipients showing CAN grade 1 or 2, or CNI nephrotoxicity. Importantly, patients who improved their creatinine clearance did not show increased proteinuria measured in a voided specimen as the ratio of urinary protein and creatinine concentration (P/C). These results, if confirmed with long-term follow-up and a larger number of patients, would allow us to consider EVL as a promising agent for maintenance immunosuppressive regimens in kidney transplantation.

  6. Everolimus-associated stomatitis in a patient who had renal transplant.

    PubMed

    Ji, Yisi D; Aboalela, Ali; Villa, Alessandro

    2016-10-19

    Everolimus is used as an immunosuppressant in renal allograft transplant rejection and in metastatic breast cancer treatment. One side effect of everolimus is stomatitis, referred to as mammalian target of rapamycin inhibitor-associated stomatitis. This side effect can affect treatment course and contribute to discontinuation of therapy or dose reduction, previously reported in the treatment of metastatic breast cancer. Here, we present a case of everolimus-associated stomatitis with a novel management method with intralesional triamcinolone that allows for continuous course of everolimus.

  7. Adenosine triphosphate-competitive mTOR inhibitors: a new class of immunosuppressive agents that inhibit allograft rejection.

    PubMed

    Rosborough, B R; Raïch-Regué, D; Liu, Q; Venkataramanan, R; Turnquist, H R; Thomson, A W

    2014-09-01

    The mechanistic/mammalian target of rapamycin (mTOR) is inhibited clinically to suppress T cell function and prevent allograft rejection. mTOR is the kinase subunit of two mTOR-containing complexes, mTOR complex (mTORC) 1 and 2. Although mTORC1 is inhibited by the macrolide immunosuppressant rapamycin (RAPA), its efficacy may be limited by its inability to block mTORC1 completely and its limited effect on mTORC2. Adenosine triphosphate (ATP)-competitive mTOR inhibitors are an emerging class of mTOR inhibitors that compete with ATP at the mTOR active site and inhibit any mTOR-containing complex. Since this class of compounds has not been investigated for their immunosuppressive potential, our goal was to determine the influence of a prototypic ATP-competitive mTOR inhibitor on allograft survival. AZD8055 proved to be a potent suppressor of T cell proliferation. Moreover, a short, 10-day course of the agent successfully prolonged murine MHC-mismatched, vascularized heart transplant survival. This therapeutic effect was associated with increased graft-infiltrating regulatory T cells and reduced CD4(+) and CD8(+) T cell interferon-γ production. These studies establish for the first time, that ATP-competitive mTOR inhibition can prolong organ allograft survival and warrant further investigation of this next generation mTOR inhibitors.

  8. Angiotensin II type I receptor (AT1R) is an independent prognosticator of esophageal squamous cell carcinoma and promotes cells proliferation via mTOR activation

    PubMed Central

    Li, Shau-Hsuan; Lu, Hung-I; Chang, Alice Y.W.; Huang, Wan-Ting; Lin, Wei-Che; Lee, Ching-Chang; Tien, Wan-Yu; Lan, Ya-Chun; Tsai, Hsin-Ting; Chen, Chang-Han

    2016-01-01

    Background The aim of this study was to investigate the effects of the angiotensin II/ angiotensin II type I receptor (AT1R) and angiotensin II type II receptor (AT2R) signaling pathway in esophageal squamous cell carcinoma (ESCC). Methods Immunohistochemistry was performed to evaluate the expression levels of AT1R and AT2R in tissues from 152 surgically resected ESCC patients, and those expression levels were then correlated with treatment outcomes. The angiotensin II/AT1R/AT2R signaling pathway and its biological effects in the context of ESCC were investigated in vitro and in vivo. Results In human samples, AT1R overexpression was univariately associated with inferior overall survival and remained multivariately independent (hazard ratio=1.812). In vitro, angiotensin II stimulated the growth of ESCC cells in a dose-dependent manner. Treatment with irbesartan or AT1R-RNAi knockdown but not treatment with PD123319 significantly decreased the level of angiotensin II-induced ESCC cell proliferation. Angiotensin II also caused mTOR activation in a dose-dependent manner, and everolimus or mTOR-RNAi knockdown significantly suppressed the level of angiotensin II-induced ESCC cell proliferation. Furthermore, AT1R-RNAi knockdown suppressed the activation of mTOR. Clinically, AT1R expression was also correlated with phosphorylated mTOR expression. In a xenograft model, local angiotensin II injection enhanced tumor growth, and this effect could be decreased by treatment with irbesartan or everolimus. In a 4-NQO-induced-ESCC murine model, irbesartan significantly decreased the incidence of esophageal tumor. Conclusions These findings suggest that AT1R overexpression is an independent adverse prognosticator for patients with ESCC and that angiotensin II/AT1R signaling stimulates ESCC growth, in part through mTOR activation. PMID:27564102

  9. Novel nervous and multi-system regenerative therapeutic strategies for diabetes mellitus with mTOR

    PubMed Central

    Maiese, Kenneth

    2016-01-01

    Throughout the globe, diabetes mellitus (DM) is increasing in incidence with limited therapies presently available to prevent or resolve the significant complications of this disorder. DM impacts multiple organs and affects all components of the central and peripheral nervous systems that can range from dementia to diabetic neuropathy. The mechanistic target of rapamycin (mTOR) is a promising agent for the development of novel regenerative strategies for the treatment of DM. mTOR and its related signaling pathways impact multiple metabolic parameters that include cellular metabolic homeostasis, insulin resistance, insulin secretion, stem cell proliferation and differentiation, pancreatic β-cell function, and programmed cell death with apoptosis and autophagy. mTOR is central element for the protein complexes mTOR Complex 1 (mTORC1) and mTOR Complex 2 (mTORC2) and is a critical component for a number of signaling pathways that involve phosphoinositide 3-kinase (PI 3-K), protein kinase B (Akt), AMP activated protein kinase (AMPK), silent mating type information regulation 2 homolog 1 (Saccharomyces cerevisiae) (SIRT1), Wnt1 inducible signaling pathway protein 1 (WISP1), and growth factors. As a result, mTOR represents an exciting target to offer new clinical avenues for the treatment of DM and the complications of this disease. Future studies directed to elucidate the delicate balance mTOR holds over cellular metabolism and the impact of its broad signaling pathways should foster the translation of these targets into effective clinical regimens for DM. PMID:27127460

  10. mTOR inhibitors in cancer therapy

    PubMed Central

    Xie, Jianling; Wang, Xuemin; Proud, Christopher G.

    2016-01-01

    The mammalian target of rapamycin, mTOR, plays key roles in cell growth and proliferation, acting at the catalytic subunit of two protein kinase complexes: mTOR complexes 1 and 2 (mTORC1/2). mTORC1 signaling is switched on by several oncogenic signaling pathways and is accordingly hyperactive in the majority of cancers. Inhibiting mTORC1 signaling has therefore attracted great attention as an anti-cancer therapy. However, progress in using inhibitors of mTOR signaling as therapeutic agents in oncology has been limited by a number of factors, including the fact that the classic mTOR inhibitor, rapamycin, inhibits only some of the effects of mTOR; the existence of several feedback loops; and the crucial importance of mTOR in normal physiology. PMID:27635236

  11. Phase II Study of Temozolomide (TMZ) and Everolimus (RAD001) Therapy for Metastatic Melanoma

    PubMed Central

    Dronca, Roxana S.; Allred, Jacob B.; Perez, Domingo G.; Nevala, Wendy K.; Lieser, Elizabeth A.T.; Thompson, Michael; Maples, William J.; Creagan, Edward T.; Pockaj, Barbara A.; Kaur, Judith S.; Moore, Timothy D.; Marchello, Benjamin T.; Markovic, Svetomir N.

    2014-01-01

    Objective Mammalian target of rapamycin (mTOR) pathway is activated in malignant melanoma and in situ lesions as opposed to benign nevi. Inhibition of PI3K-Akt-mTOR signaling is implicated in sensitization of melanoma cells to alkylating agents [temozolomide (TMZ)] and inhibition of tumor angiogenesis. Methods We conducted a single-arm phase II multi-institution cooperative group study to assess the antitumor activity and safety profile of the combination of TMZ and the rapamycin derivative everolimus in patients with metastatic unresectable malignant melanoma. Patients received 10 mg/d of RAD001 for 5 of 7 days (ie, 50 mg/ wk) and 200 mg/m2/d of TMZ for 5 days each cycle. Results Of the first 39 eligible patients, 17 were PFS-9 successes, for a predetermined threshold of 18/39 patients for a positive trial. Overall, 21 of 48 patients were progression free at 9 weeks, for an event-free survival rate of 44% (95% confidence interval, 29%–59%). The median progression-free survival was 2.4 months and the median overall survival was 8.6 months. Four patients achieved a partial response; the median duration of response was 15.1 months. No complete remissions were observed. Treatment was in general well tolerated with only 1 patient discontinuing therapy due to toxicity (hyperlipidemia). Conclusions The combination of TMZ and RAD001 was well tolerated but failed to meet/exceed our study threshold for promising clinical activity in patients with metastatic melanoma. PMID:23357973

  12. DDX5 promotes gastric cancer cell proliferation in vitro and in vivo through mTOR signaling pathway

    PubMed Central

    Du, Cheng; Li, Dan-qi; Li, Na; Chen, Li; Li, Shi-sen; Yang, Yang; Hou, Ming-xiao; Xie, Man-jiang; Zheng, Zhen-dong

    2017-01-01

    DEAD (Asp-Glu-Ala-Asp) box helicase 5 (DDX5) is an ATP-dependent RNA helicase that is overexpressed in various malignancies. Increasing evidence suggests that DDX5 participates in carcinogenesis and cancer progression via promoting cell proliferation and metastasis. However, the functional role of DDX5 in gastric cancer is largely unknown. In this study, we observed that DDX5 was significantly up-regulated in gastric cancer tissues compared with the paired adjacent normal tissues. The expression of DDX5 correlated strongly with Ki67 index and pathological stage of gastric cancer. In vitro and in vivo studies suggested that knockdown of DDX5 inhibited gastric cancer cell proliferation, colony formation and xenografts growth, whereas ectopic expression of DDX5 promoted these cellular functions. Mechanically, DDX5 induced gastric cancer cell growth by activating mTOR/S6K1. Treatment of everolimus, the specific mTOR inhibitor, significantly attenuated DDX5-mediated cell proliferation. Interestingly, the expression of DDX5 and p-mTOR in gastric cancer tissues demonstrated a positive correlation. Taken together, these results revealed a novel role of DDX5 in gastric cancer cell proliferation via the mTOR pathway. Therefore, DDX5 may serve as a therapeutic target in gastric cancer. PMID:28216662

  13. Everolimus-eluting stents: update on current clinical studies.

    PubMed

    Allocco, Dominic J; Joshi, Anita A; Dawkins, Keith D

    2011-01-01

    Everolimus-eluting stents (EES) have become the most commonly implanted coronary stents worldwide. This review describes and analyzes the clinical data supporting the use of EES, focusing primarily on published, randomized, controlled trials. Everolimus-eluting stents have been shown to have less restenosis, stent thrombosis, and periprocedural myocardial infarction compared with earlier generation paclitaxel-eluting stents (PES). Lower rates of adverse events for EES compared with PES were generally seen in all subgroups, with the notable exception of patients with diabetes mellitus. There have been fewer, randomized, clinical trials comparing EES with either sirolimus-eluting stents or zotarolimus-eluting stents, although very good results with EES have been observed in the trials that have been performed. Recent clinical trial data suggest that this excellent safety and efficacy profile is maintained in a next-generation EES designed to have improved mechanical properties and radiopacity.

  14. Therapeutic Drug Monitoring of Everolimus: A Consensus Report.

    PubMed

    Shipkova, Maria; Hesselink, Dennis A; Holt, David W; Billaud, Eliane M; van Gelder, Teun; Kunicki, Paweł K; Brunet, Mercè; Budde, Klemens; Barten, Markus J; De Simone, Paolo; Wieland, Eberhard; López, Olga Millán; Masuda, Satohiro; Seger, Christoph; Picard, Nicolas; Oellerich, Michael; Langman, Loralie J; Wallemacq, Pierre; Morris, Raymond G; Thompson, Carol; Marquet, Pierre

    2016-04-01

    In 2014, the Immunosuppressive Drugs Scientific Committee of the International Association of Therapeutic Drug Monitoring and Clinical Toxicology called a meeting of international experts to provide recommendations to guide therapeutic drug monitoring (TDM) of everolimus (EVR) and its optimal use in clinical practice. EVR is a potent inhibitor of the mammalian target of rapamycin, approved for the prevention of organ transplant rejection and for the treatment of various types of cancer and tuberous sclerosis complex. EVR fulfills the prerequisites for TDM, having a narrow therapeutic range, high interindividual pharmacokinetic variability, and established drug exposure-response relationships. EVR trough concentrations (C0) demonstrate a good relationship with overall exposure, providing a simple and reliable index for TDM. Whole-blood samples should be used for measurement of EVR C0, and sampling times should be standardized to occur within 1 hour before the next dose, which should be taken at the same time everyday and preferably without food. In transplantation settings, EVR should be generally targeted to a C0 of 3-8 ng/mL when used in combination with other immunosuppressive drugs (calcineurin inhibitors and glucocorticoids); in calcineurin inhibitor-free regimens, the EVR target C0 range should be 6-10 ng/mL. Further studies are required to determine the clinical utility of TDM in nontransplantation settings. The choice of analytical method and differences between methods should be carefully considered when determining EVR concentrations, and when comparing and interpreting clinical trial outcomes. At present, a fully validated liquid chromatography tandem mass spectrometry assay is the preferred method for determination of EVR C0, with a lower limit of quantification close to 1 ng/mL. Use of certified commercially available whole-blood calibrators to avoid calibration bias and participation in external proficiency-testing programs to allow continuous cross

  15. Activation of MTOR in pulmonary epithelium promotes LPS-induced acute lung injury.

    PubMed

    Hu, Yue; Lou, Jian; Mao, Yuan-Yuan; Lai, Tian-Wen; Liu, Li-Yao; Zhu, Chen; Zhang, Chao; Liu, Juan; Li, Yu-Yan; Zhang, Fan; Li, Wen; Ying, Song-Min; Chen, Zhi-Hua; Shen, Hua-Hao

    2016-12-01

    MTOR (mechanistic target of rapamycin [serine/threonine kinase]) plays a crucial role in many major cellular processes including metabolism, proliferation and macroautophagy/autophagy induction, and is also implicated in a growing number of proliferative and metabolic diseases. Both MTOR and autophagy have been suggested to be involved in lung disorders, however, little is known about the role of MTOR and autophagy in pulmonary epithelium in the context of acute lung injury (ALI). In the present study, we observed that lipopolysaccharide (LPS) stimulation induced MTOR phosphorylation and decreased the expression of MAP1LC3B/LC3B (microtubule-associated protein 1 light chain 3 β)-II, a hallmark of autophagy, in mouse lung epithelium and in human bronchial epithelial (HBE) cells. The activation of MTOR in HBE cells was mediated by TLR4 (toll-like receptor 4) signaling. Genetic knockdown of MTOR or overexpression of autophagy-related proteins significantly attenuated, whereas inhibition of autophagy further augmented, LPS-induced expression of IL6 (interleukin 6) and IL8, through NFKB signaling in HBE cells. Mice with specific knockdown of Mtor in bronchial or alveolar epithelial cells exhibited significantly attenuated airway inflammation, barrier disruption, and lung edema, and displayed prolonged survival in response to LPS exposure. Taken together, our results demonstrate that activation of MTOR in the epithelium promotes LPS-induced ALI, likely through downregulation of autophagy and the subsequent activation of NFKB. Thus, inhibition of MTOR in pulmonary epithelial cells may represent a novel therapeutic strategy for preventing ALI induced by certain bacteria.

  16. The FGFR4-G388R single-nucleotide polymorphism alters pancreatic neuroendocrine tumor progression and response to mTOR inhibition therapy.

    PubMed

    Serra, Stefano; Zheng, Lei; Hassan, Manal; Phan, Alexandria T; Woodhouse, Linda J; Yao, James C; Ezzat, Shereen; Asa, Sylvia L

    2012-11-15

    Pancreatic neuroendocrine tumors (pNET), also known as islet cell tumors, exhibit a wide range of biologic behaviors ranging from long dormancy to rapid progression. Currently, there are few molecular biomarkers that can be used to predict recurrence/metastasis or response to therapy. This study examined the predictive and prognostic value of a single nucleotide polymorphism substituting an arginine (R) for glycine (G) in codon 388 of the FGFR4 transmembrane domain. We established the FGFR4 genotype of 71 patients with pNETs and correlated genotype with biologic behavior. We created an in vivo model of pNET with BON1 cells and transfected them with either FGFR4-G388 or FGFR4-R388 to determine the mechanism of action and to examine response to the mTOR inhibitor everolimus. We then validated the predictive results of experimental studies in a group of patients treated with everolimus. FGFR4-R388 is associated with more aggressive clinical behavior in patients with pNETs with a statistically significant higher risk of advanced tumor stage and liver metastasis. Using an orthotopic mouse xenograft model, we show that FGFR4-R388 promotes tumor progression by increasing intraperitoneal spread and metastatic growth within the liver. Unlike FGFR4-G388, FGFR4-R388 BON1 tumors exhibited diminished responsiveness to everolimus. Concordantly, there was a statistically significant reduction in response to everolimus in patients with FGFR4-R388. Our findings highlight the importance of the FGFR4 allele in pNET progression and identify a predictive marker of potential therapeutic importance in this disease.

  17. Amenorrhea as a rare drug-related adverse event associated with everolimus for pancreatic neuroendocrine tumors.

    PubMed

    Kawaguchi, Yoshiaki; Maruno, Atsuko; Kawashima, Yohei; Ito, Hiroyuki; Ogawa, Masami; Mine, Tetsuya

    2014-11-14

    The patient was an asymptomatic 43-year-old woman. Abdominal ultrasonography and enhanced computed tomography showed a tumor lesion accompanied by multiple cystic changes in the liver and the pancreatic tail. Endoscopic ultrasound-fine needle aspiration was performed on the pancreatic tumor lesion and revealed pancreatic neuroendocrine tumor (PNET). As it was unresectable due to multiple liver metastases, the decision was made to initiate treatment with everolimus and transcatheter arterial chemoembolization. The patient ceased menstruating after the start of everolimus administration. When the administration was discontinued due to interstitial lung disease, menstruation resumed, but then again stopped with everolimus resumption. An association between everolimus and amenorrhea was highly suspected. Amenorrhea occurred as a rare adverse event of everolimus. As the younger women might be included in PNETs patients, we should put this adverse event into consideration.

  18. Co-administration strategy to enhance brain accumulation of vandetanib by modulating P-glycoprotein (P-gp/Abcb1) and breast cancer resistance protein (Bcrp1/Abcg2) mediated efflux with m-TOR inhibitors

    PubMed Central

    Minocha, Mukul; Khurana, Varun; Qin, Bin; Pal, Dhananjay; Mitra, Ashim K

    2012-01-01

    The objectives of this study were (i) to characterize the interaction of vandetanib with P-glycoprotein (P-gp) and breast cancer resistance protein (Bcrp1) in vitro and in vivo (ii) to study the modulation of P-gp and BCRP mediated efflux of vandetanib with specific transport inhibitors and m-TOR inhibitors, everolimus and temsirolimus. Cellular accumulation and bi-directional transport studies in MDCKII cell monolayers were conducted to delineate the role of efflux transporters on disposition of vandetanib. Brain distribution studies were conducted in male FVB wild-type mice with vandetanib administered intravenously either alone or in the presence of specific inhibitors and m-TOR inhibitors. In vitro studies suggested that vandetanib is a high affinity substrate of Bcrp1 but is not transported by P-gp. Interestingly, in vivo brain distribution studies in FVB wild type mice indicated that vandetanib penetration into the brain is restricted by both Bcrp1 and P-gp mediated active efflux at the blood brain barrier (BBB). Co-administration of elacridar, a dual P-gp/BCRP inhibitor increased the brain to plasma concentration ratio of vandetanib upto 5 fold. Of the two m-TOR pathway inhibitors examined; everolimus showed potent effect on modulating vandetanib brain penetration whereas no significant affect on vandetanib brain uptake was observed following temsirolimus co-administration. This finding could be clinically relevant as everolimus can provide synergistic pharmacological effect in addition to primary role of vandetanib efflux modulation at BBB for the treatment of brain tumors. PMID:22633931

  19. A role for Raptor phosphorylation in the mechanical activation of mTOR signaling.

    PubMed

    Frey, John W; Jacobs, Brittany L; Goodman, Craig A; Hornberger, Troy A

    2014-02-01

    The activation of mTOR signaling is necessary for mechanically-induced changes in skeletal muscle mass, but the mechanisms that regulate the mechanical activation of mTOR signaling remain poorly defined. In this study, we set out to determine if changes in the phosphorylation of Raptor contribute to the mechanical activation of mTOR. To accomplish this goal, mouse skeletal muscles were subjected to mechanical stimulation via a bout of eccentric contractions (EC). Using mass spectrometry and Western blot analysis, we found that ECs induced an increase in Raptor S696, T706, and S863 phosphorylation, and this effect was not inhibited by rapamycin. This observation suggested that changes in Raptor phosphorylation might be an upstream event in the pathway through which mechanical stimuli activate mTOR. To test this, we employed a phospho-defective mutant of Raptor (S696A/T706A/S863A) and found that the EC-induced activation of mTOR signaling was significantly blunted in muscles expressing this mutant. Furthermore, mutation of the three phosphorylation sites altered the interactions of Raptor with PRAS40 and p70(S6k), and it also prevented the EC-induced dissociation of Raptor from p70(S6k). Combined, these results suggest that changes in the phosphorylation of Raptor play an important role in the pathway through which mechanical stimuli activate mTOR signaling.

  20. Multiple roles for mTOR signaling in both glutamatergic and GABAergic synaptic transmission

    PubMed Central

    Weston, Matthew C.; Chen, Hongmei; Swann, John W.

    2012-01-01

    Summary The mammalian target of rapamycin (mTOR) signaling pathway in neurons integrates a variety of extracellular signals to produce appropriate translational responses. mTOR signaling is hyperactive in neurological syndromes in both humans and mouse models that are characterized by epilepsy, autism and cognitive disturbances. In addition, rapamycin, a clinically important immunosuppressant, is a specific and potent inhibitor of mTOR signaling. While mTOR is known to regulate growth and synaptic plasticity of glutamatergic neurons, its effects on basic parameters of synaptic transmission are less well studied, and its role in regulating GABAergic transmission is unexplored. We therefore performed an electrophysiological and morphological comparison of glutamatergic and GABAergic neurons in which mTOR signaling was either increased by loss of the repressor Pten or decreased by treatment with rapamycin. We found that hyperactive mTOR signaling increased evoked synaptic responses in both glutamatergic and GABAergic neurons by approximately 50%, due to an increase in the number of synaptic vesicles available for release, the number of synapses formed and the miniature event size. Prolonged (72 hours) rapamycin treatment prevented these abnormalities and also decreased synaptic transmission in wild-type glutamatergic, but not GABAergic, neurons. Further analyses suggested that hyperactivation of the mTOR pathway also impairs presynaptic function, possibly by interfering with vesicle fusion. Despite this presynaptic impairment, the net effect of Pten loss is enhanced synaptic transmission in both GABAergic and glutamatergic neurons, which has numerous implications – depending on where in the brain mutations of an mTOR suppressor gene takes place during development. PMID:22895726

  1. FTY720 and everolimus in de novo renal transplant patients at risk for delayed graft function: results of an exploratory one-yr multicenter study.

    PubMed

    Tedesco-Silva, H; Lorber, M I; Foster, C E; Sollinger, H W; Mendez, R; Carvalho, D B; Shapiro, R; Rajagopalan, P R; Mayer, H; Slade, J; Kahan, B D

    2009-01-01

    This exploratory, multicenter, open-label study evaluated the efficacy and safety of FTY720, as a part of an immunosuppressive regimen, in combination with everolimus and steroids in de novo renal transplant recipients at increased risk of delayed graft function (DGF). Patients received FTY720 (5 mg) and everolimus (4 mg) 2-12 h pre-transplantation, followed by 2.5 mg/d FTY720 and concentration-controlled everolimus (4-8 ng/mL) post-transplant for 12 months. Induction therapy was prohibited. After enrollment of 56 of the planned 200 patients between 2000 and 2002, the recruitment was terminated. The primary endpoint, rate of graft loss, or death at three months was 15.4% and the biopsy-confirmed acute rejection was 42.3%. Death or graft loss at 12 months in the DGF and non-DGF arms was 36.0% and 25.9%, respectively. The mean estimated creatinine clearance at three months was 63 and 55 mL/min in the non-DGF and DGF groups, respectively, while at 12 months it was 56 mL/min in both the groups. Although there was no comparator arm, the results from this exploratory study (compared with data from other phases II and III trials) indicated no apparent benefits of FTY720-based regimens for prevention of acute rejection and preservation of renal function in renal transplant recipients at high risk of DGF.

  2. mTOR and the health benefits of exercise.

    PubMed

    Watson, Kurt; Baar, Keith

    2014-12-01

    Exercise is the greatest physiological stress that our bodies experience. For example, during maximal endurance exercise in elite athlete's cardiac output can increase up to 8-fold and the working muscles receive 21-times more blood each minute than at rest. Given the physiological stress associated with exercise and the adaptations that occur to handle this stress, it is not surprising that exercise training is known to prevent or effectively treat a multitude of degenerative conditions including cardiovascular disease, cancer, diabetes, depression, Alzheimer's disease, Parkinson's disease, and many others. Many of the health benefits of exercise are mediated by the mammalian/mechanistic target of rapamycin (mTOR), either in complex 1 or 2, not only within the working muscle, but also in distant tissues such as fat, liver, and brain. This review will discuss how exercise activates mTOR in diverse tissues and the ways that mTOR is important in the adaptive response that makes us bigger, stronger, and healthier as a result of exercise.

  3. Everolimus Treatment for an Early Infantile Subependymal Giant Cell Astrocytoma With Tuberous Sclerosis Complex.

    PubMed

    Fukumura, Shinobu; Watanabe, Toshihide; Takayama, Rumiko; Minagawa, Kimio; Tsutsumi, Hiroyuki

    2015-08-01

    Subependymal giant cell astrocytomas are benign tumors often observed with tuberous sclerosis complex. These tumors are rarely diagnosed during fetal life or early infancy. Until recently, the only available treatment has been surgical resection. Current clinical research has demonstrated that everolimus can induce these tumors' regression. We report a 19-month-old boy with tuberous sclerosis complex. At 2 months of age, he presented with congenital subependymal giant cell astrocytoma that was complicated by refractory epilepsy and severe mental retardation. Treatment with everolimus was started when he was 10 months old. Three months after initiating everolimus, the tumor was significantly reduced in size, and the reduction was subsequently maintained. His seizures decreased and he showed cognitive and developmental improvement. No severe adverse events have been observed to date. Everolimus has promise as an effective alternative to surgery for subependymal giant cell astrocytomas during early infancy.

  4. Everolimus-induced Pneumonitis after Drug-eluting Stent Implantation: A Case Report

    SciTech Connect

    Sakamoto, Susumu Kikuchi, Naoshi; Ichikawa, Atsuo; Sano, Go; Satoh, Keita; Sugino, Keishi; Isobe, Kazutoshi; Takai, Yujiro; Shibuya, Kazutoshi; Homma, Sakae

    2013-08-01

    Despite the wide use of everolimus as an antineoplastic coating agent for coronary stents to reduce the rate of restenosis, little is known about the health hazards of everolimus-eluting stents (EES). We describe a case of pneumonitis that developed 2 months after EES implantation for angina. Lung pathology demonstrated an organizing pneumonia pattern that responded to corticosteroid therapy. Although the efficacy of EES for ischemic heart disease is well established, EES carries a risk of pneumonitis.

  5. mTOR inhibitors induce apoptosis in colon cancer cells via CHOP-dependent DR5 induction on 4E-BP1 dephosphorylation.

    PubMed

    He, K; Zheng, X; Li, M; Zhang, L; Yu, J

    2016-01-14

    The mammalian target of rapamycin (mTOR) is commonly activated in colon cancer. mTOR complex 1 (mTORC1) is a major downstream target of the PI3K/ATK pathway and activates protein synthesis by phosphorylating key regulators of messenger RNA translation and ribosome synthesis. Rapamycin analogs Everolimus and Temsirolimus are non-ATP-competitive mTORC1 inhibitors, and suppress proliferation and tumor angiogenesis and invasion. We now show that apoptosis plays a key role in their anti-tumor activities in colon cancer cells and xenografts through the DR5, FADD and caspase-8 axis, and is strongly enhanced by tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and 5-fluorouracil. The induction of DR5 by rapalogs is mediated by the ER stress regulator and transcription factor CHOP, but not the tumor suppressor p53, on rapid and sustained inhibition of 4E-BP1 phosphorylation, and attenuated by eIF4E expression. ATP-competitive mTOR/PI3K inhibitors also promote DR5 induction and FADD-dependent apoptosis in colon cancer cells. These results establish activation of ER stress and the death receptor pathway as a novel anticancer mechanism of mTOR inhibitors.

  6. mTOR inhibitors induce apoptosis in colon cancer cells via CHOP-dependent DR5 induction upon 4E-BP1 dephosphorylation

    PubMed Central

    He, Kan; Zheng, Xingnan; Li, Mei; Zhang, Lin; Yu, Jian

    2015-01-01

    The mammalian target of rapamycin (mTOR) is commonly activated in colon cancer. mTOR complex 1 (mTORC1) is a major downstream target of the PI3K/ATK pathway and activates protein synthesis by phosphorylating key regulators of mRNA translation and ribosome synthesis. Rapamycin analogs Everolimus and Temsirolimus are non-ATP-competitive mTORC1 inhibitors, and suppress proliferation and tumor angiogenesis and invasion. We now show that apoptosis plays a key role in their anti-tumor activities in colon cancer cells and xenografts through the DR5, FADD and caspase-8 axis, and is strongly enhanced by TRAIL and 5-fluorouracil. The induction of DR5 by rapalogs is mediated by the ER stress regulator and transcription factor CHOP, but not the tumor suppressor p53, upon rapid and sustained inhibition of 4E-BP1 phosphorylation, and attenuated by eIF4E expression. ATP-competitive mTOR/PI3K inhibitors also promote DR5 induction and FADD-dependent apoptosis in colon cancer cells. These results establish activation of ER stress and the death receptor pathway as a novel anticancer mechanism of mTOR inhibitors. PMID:25867072

  7. Everolimus affects vasculogenic mimicry in renal carcinoma resistant to sunitinib

    PubMed Central

    Serova, Maria; Tijeras-Raballand, Annemilaï; Santos, Celia Dos; Martinet, Matthieu; Neuzillet, Cindy; Lopez, Alfred; Mitchell, Dianne C.; Bryan, Brad A.; Gapihan, Guillaume; Janin, Anne; Bousquet, Guilhem; Riveiro, Maria Eugenia; Bieche, Ivan; Faivre, Sandrine

    2016-01-01

    Angiogenesis is hallmark of clear cell renal cell carcinogenesis. Anti-angiogenic therapies have been successful in improving disease outcome; however, most patients treated with anti-angiogenic agents will eventually progress. In this study we report that clear cell renal cell carcinoma was associated with vasculogenic mimicry in both mice and human with tumor cells expressing endothelial markers in the vicinity of tumor vessels. We show that vasculogenic mimicry was efficiently targeted by sunitinib but eventually associated with tumor resistance and a more aggressive phenotype both in vitro and in vivo. Re-challenging these resistant tumors in mice, we showed that second-line treatment with everolimus particularly affected vasculogenic mimicry and tumor cell differentiation compared to sorafenib and axitinib. Finally, our results highlighted the phenotypic and genotypic changes at the tumor cell and microenvironment levels during sunitinib response and progression and the subsequent improvement second-line therapies bring to the current renal cell carcinoma treatment paradigm. PMID:27509260

  8. Dissociation of the pharmacological effects of THC by mTOR blockade.

    PubMed

    Puighermanal, Emma; Busquets-Garcia, Arnau; Gomis-González, Maria; Marsicano, Giovanni; Maldonado, Rafael; Ozaita, Andrés

    2013-06-01

    The potential therapeutic benefits of cannabinoid compounds have raised interest in understanding the molecular mechanisms that underlie cannabinoid-mediated effects. We previously showed that the acute amnesic-like effects of delta9-tetrahydrocannabinol (THC) were prevented by the subchronic inhibition of the mammalian target of rapamycin (mTOR) pathway. In the present study, we assess the relevance of the mTOR pathway in other acute and chronic pharmacological effects of THC. The rapamycin derivative temsirolimus, an inhibitor of the mTOR pathway approved by the Food and Drug Administration, prevents both the anxiogenic- and the amnesic-like effects produced by acute THC. In contrast, THC-induced anxiolysis, hypothermia, hypolocomotion, and antinociception are not sensitive to the mTOR inhibition. In addition, a clear tolerance to THC-induced anxiolysis, hypothermia, hypolocomotion, and antinociception was observed after chronic treatment, but not to its anxiogenic- and amnesic-like effects. Temsirolimus pre-treatment prevented the amnesic-like effects of chronic THC without affecting the downregulation of CB1 receptors (CB1R) induced by this chronic treatment. Instead, temsirolimus blockade after chronic THC cessation did not prevent the residual cognitive deficit produced by chronic THC. Using conditional knockout mice lacking CB1R in GABAergic or glutamatergic neurons, we found that GABAergic CB1Rs are mainly downregulated under chronic THC treatment conditions, and CB1-GABA-KO mice did not develop cognitive deficits after chronic THC exposure. Therefore, mTOR inhibition by temsirolimus allows the segregation of the potentially beneficial effects of cannabinoid agonists, such as the anxiolytic and antinociceptive effects, from the negative effects, such as anxiogenic- and amnesic-like responses. Altogether, these results provide new insights for targeting the endocannabinoid system in order to prevent possible side effects.

  9. Dissociation of the Pharmacological Effects of THC by mTOR Blockade

    PubMed Central

    Puighermanal, Emma; Busquets-Garcia, Arnau; Gomis-González, Maria; Marsicano, Giovanni; Maldonado, Rafael; Ozaita, Andrés

    2013-01-01

    The potential therapeutic benefits of cannabinoid compounds have raised interest in understanding the molecular mechanisms that underlie cannabinoid-mediated effects. We previously showed that the acute amnesic-like effects of delta9-tetrahydrocannabinol (THC) were prevented by the subchronic inhibition of the mammalian target of rapamycin (mTOR) pathway. In the present study, we assess the relevance of the mTOR pathway in other acute and chronic pharmacological effects of THC. The rapamycin derivative temsirolimus, an inhibitor of the mTOR pathway approved by the Food and Drug Administration, prevents both the anxiogenic- and the amnesic-like effects produced by acute THC. In contrast, THC-induced anxiolysis, hypothermia, hypolocomotion, and antinociception are not sensitive to the mTOR inhibition. In addition, a clear tolerance to THC-induced anxiolysis, hypothermia, hypolocomotion, and antinociception was observed after chronic treatment, but not to its anxiogenic- and amnesic-like effects. Temsirolimus pre-treatment prevented the amnesic-like effects of chronic THC without affecting the downregulation of CB1 receptors (CB1R) induced by this chronic treatment. Instead, temsirolimus blockade after chronic THC cessation did not prevent the residual cognitive deficit produced by chronic THC. Using conditional knockout mice lacking CB1R in GABAergic or glutamatergic neurons, we found that GABAergic CB1Rs are mainly downregulated under chronic THC treatment conditions, and CB1–GABA–KO mice did not develop cognitive deficits after chronic THC exposure. Therefore, mTOR inhibition by temsirolimus allows the segregation of the potentially beneficial effects of cannabinoid agonists, such as the anxiolytic and antinociceptive effects, from the negative effects, such as anxiogenic- and amnesic-like responses. Altogether, these results provide new insights for targeting the endocannabinoid system in order to prevent possible side effects. PMID:23358238

  10. RAD001 (everolimus) induces dose-dependent changes to cell cycle regulation and modifies the cell cycle response to vincristine.

    PubMed

    Saunders, P O; Weiss, J; Welschinger, R; Baraz, R; Bradstock, K F; Bendall, L J

    2013-10-01

    More than 50% of adults and ~20% of children with pre-B acute lymphoblastic leukemia (ALL) relapse following treatment. Dismal outcomes for patients with relapsed or refractory disease mandate novel approaches to therapy. We have previously shown that the combination of the mTOR inhibitor RAD001 (everolimus) and the chemotherapeutic agent vincristine increases the survival of non-obese diabetic/severe combined immuno-deficient (NOD/SCID) mice bearing human ALL xenografts. We have also shown that 16 μM RAD001 synergized with agents that cause DNA damage or microtubule disruption in pre-B ALL cells in vitro. Here, we demonstrate that RAD001 has dose-dependent effects on the cell cycle in ALL cells, with 1.5 μM RAD001 inhibiting pRb, Ki67 and PCNA expression and increasing G0/1 cell cycle arrest, whereas 16 μM RAD001 increases pRb, cyclin D1, Ki67 and PCNA, with no evidence of an accumulation of cells in G0/1. Transition from G2 into mitosis was promoted by 16 μM RAD001 with reduced phosphorylation of cdc2 in cells with 4 N DNA content. However, 16 μM RAD001 preferentially induced cell death in cells undergoing mitosis. When combined with vincristine, 16 μM RAD001 reduced the vincristine-induced accumulation of cells in mitosis, probably as a result of increased death in this population. Although 16 μM RAD001 weakly activated Chk1 and Chk2, it suppressed strong vincristine-induced activation of these cell cycle checkpoint regulators. We conclude that RAD001 enhances chemosensitivity at least in part through suppression of cell cycle checkpoint regulation in response to vincristine and increased progression from G2 into mitosis.

  11. Anti-PD-L1 treatment enhances antitumor effect of everolimus in a mouse model of renal cell carcinoma.

    PubMed

    Hirayama, Yukiyoshi; Gi, Min; Yamano, Shotaro; Tachibana, Hirokazu; Okuno, Takahiro; Tamada, Satoshi; Nakatani, Tatsuya; Wanibuchi, Hideki

    2016-12-01

    Immunotherapy based on blockade of the programmed death-1 (PD-1)/programmed death-ligand 1 (PD-L1) axis has shown promising clinical activity for renal cell carcinoma (RCC) patients; however, the most effective use of these agents in combination with conventional targeted therapy remains to be resolved. Here we evaluated the therapeutic efficacy of the combination of the mTOR inhibitor everolimus (EVE) and anti-PD-L1 using an immunocompetent mouse model of RCC. We first assessed the in vitro effect of EVE on PD-L1 expression in the human 786-O and mouse RENCA RCC cell lines and found that EVE upregulated PD-L1 expression in these RCC cell lines. We then treated RENCA tumor-bearing mice with EVE and found that PD-L1 expression was also increased in tumor cells after EVE treatment. To determine the antitumor effects of EVE alone, anti-PD-L1 alone, and EVE in combination with anti-PD-L1, we evaluated their antitumor effects on RENCA tumor-bearing mice. A significant decrease in the tumor burden was observed in the EVE alone but not in the anti-PD-L1 alone treatment group compared with the control group. Importantly, the combination of EVE with anti-PD-L1 significantly reduced tumor burden compared with the EVE alone treatment, increasing tumor infiltrating lymphocytes (TILs) and the ratio of cytotoxic CD8(+) T cells to TILs. The results of the present study demonstrated that anti-PD-L1 treatment enhanced the antitumor effect of EVE in a mouse model, supporting a direct translation of this combination strategy to the clinic for the treatment of RCC.

  12. Multiple indications for everolimus after liver transplantation in current clinical practice

    PubMed Central

    Bilbao, Itxarone; Dopazo, Cristina; Lazaro, Jose; Castells, Lluis; Caralt, Mireia; Sapisochin, Gonzalo; Charco, Ramon

    2014-01-01

    AIM: To assess our experience with the use and management of everolimus-based regimens post-liver transplantation and to redefine the potential role of this drug in current clinical practice. METHODS: From October 1988 to December 2012, 1023 liver transplantations were performed in 955 patients in our Unit. Seventy-four patients (7.74%) received immunosuppression with everolimus at some time post-transplantation. Demographic characteristics, everolimus indication, time elapsed from transplantation to the introduction of everolimus, doses and levels administered, efficacy, side effects, discontinuation and post-conversion survival were analyzed. RESULTS: Mean age at the time of conversion to everolimus was 57.7 ± 10 years. Indications for conversion were: refractory rejection 31.1%, extended hepatocellular carcinoma in explanted liver 19%, post-transplant hepatocellular carcinoma recurrence 8.1%, de novo tumour 17.6%, renal insufficiency 8.1%, severe neurotoxicity 10.8%, and others 5.4%. Median time from transplantation to introduction of everolimus was 6 mo (range: 0.10-192). Mean follow-up post-conversion was 22 ± 19 mo (range: 0.50-74). The event for which the drug was indicated was resolved in 60.8% of patients, with the best results in cases of refractory rejection, renal insufficiency and neurotoxicity. Results in patients with cancer were similar to those of a historical cohort treated with other immunosuppressants. The main side effects were dyslipidemia and infections. Post-conversion acute rejection occurred in 14.9% of cases. The drug was discontinued in 28.4% of patients. CONCLUSION: Everolimus at low doses in combination with tacrolimus is a safe immunosuppressant with multiple early and late indications post-liver transplantation. PMID:25032101

  13. Finding a better drug for epilepsy: The mTOR pathway as an antiepileptogenic target

    PubMed Central

    Galanopoulou, Aristea S.; Gorter, Jan A.; Cepeda, Carlos

    2012-01-01

    Summary The mTOR signaling pathway regulates cell growth, differentiation, proliferation and metabolism. Loss of function mutations in upstream regulators of mTOR have been highly associated with dysplasias, epilepsy and neurodevelopmental disorders. These include tuberous sclerosis, which is due to mutations in TSC1 or TSC2 genes, mutations in phosphatase and tensin homolog (PTEN) as in Cowden syndrome, polyhydramnios, megalencephaly, symptomatic epilepsy syndrome (PMSE) due to mutations in the STE20-related kinase adaptor alpha (STRADalpha), and neurofibromatosis type 1 attributed to neurofibromin 1 mutations. Inhibition of the mTOR pathway with rapamycin may prevent epilepsy and improve the underlying pathology in mouse models with disrupted mTOR signaling, due to PTEN or TSC mutations. However the timing and duration of its administration appear critical in defining the seizure and pathology-related outcomes. Rapamycin application in human cortical slices from patients with cortical dysplasias reduces the 4-aminopyridine induced oscillations. In the multiple-hit model of infantile spasms, pulse high dose rapamycin administration can reduce the cortical overactivation of the mTOR pathway, suppresses spasms and has disease-modifying effects by partially improving cognitive deficits. In post-status epilepticus models of temporal lobe epilepsy, rapamycin may ameliorate the development of epilepsy-related pathology and reduce the expression of spontaneous seizures, but its effects depend on the timing and duration of administration, and possibly the model used. The observed recurrence of seizures and epilepsy-related pathology after rapamycin discontinuation suggests the need for continuous administration to maintain the benefit. However, the use of pulse administration protocols may be useful in certain age-specific epilepsy syndromes, like infantile spasms, whereas repetitive pulse rapamycin protocols may suffice to sustain a long-term benefit in genetic disorders

  14. The role of mTOR signaling in the regulation of protein synthesis and muscle mass during immobilization in mice

    PubMed Central

    You, Jae-Sung; Anderson, Garrett B.; Dooley, Matthew S.; Hornberger, Troy A.

    2015-01-01

    ABSTRACT The maintenance of skeletal muscle mass contributes substantially to health and to issues associated with the quality of life. It has been well recognized that skeletal muscle mass is regulated by mechanically induced changes in protein synthesis, and that signaling by mTOR is necessary for an increase in protein synthesis and the hypertrophy that occurs in response to increased mechanical loading. However, the role of mTOR signaling in the regulation of protein synthesis and muscle mass during decreased mechanical loading remains largely undefined. In order to define the role of mTOR signaling, we employed a mouse model of hindlimb immobilization along with pharmacological, mechanical and genetic means to modulate mTOR signaling. The results first showed that immobilization induced a decrease in the global rates of protein synthesis and muscle mass. Interestingly, immobilization also induced an increase in mTOR signaling, eIF4F complex formation and cap-dependent translation. Blocking mTOR signaling during immobilization with rapamycin not only impaired the increase in eIF4F complex formation, but also augmented the decreases in global protein synthesis and muscle mass. On the other hand, stimulating immobilized muscles with isometric contractions enhanced mTOR signaling and rescued the immobilization-induced decrease in global protein synthesis through a rapamycin-sensitive mechanism that was independent of ribosome biogenesis. Unexpectedly, the effects of isometric contractions were also independent of eIF4F complex formation. Similar to isometric contractions, overexpression of Rheb in immobilized muscles enhanced mTOR signaling, cap-dependent translation and global protein synthesis, and prevented the reduction in fiber size. Therefore, we conclude that the activation of mTOR signaling is both necessary and sufficient to alleviate the decreases in protein synthesis and muscle mass that occur during immobilization. Furthermore, these results indicate

  15. Potential biomarkers for the therapeutic efficacy of sorafenib, sunitinib and everolimus.

    PubMed

    Nakai, Yasushi; Miyake, Makito; Morizawa, Yosuke; Hori, Shunta; Tatsumi, Yoshihiro; Anai, Satoshi; Onishi, Sayuri; Tanaka, Nobumichi; Fujimoto, Kiyohide

    2017-01-01

    We examined extracellular signal‑regulated kinase (ERK), 4E‑binding protein 1 (4EBP1) and p70 ribosomal S6 kinase (p70) as potential biomarkers for pretreatment prediction of the prognosis of patients with metastatic renal cell carcinoma (RCC) treated with sorafenib, sunitinib or everolimus. 786‑O and 769‑P cells were treated with sorafenib, sunitinib and everolimus. The expression of phosphorylated/total ERK, phosphorylated/total 4EBP1 and phosphorylated/total p70 was evaluated using western blotting. ERK, 4EBP1 and p70 were knocked down by siRNA in 786‑O and 769‑P cells. Then, the viability after treatment with each drug was assessed. Expression of phosphorylated (phospho)‑ERK, -4EBP1 and -p70 was immunohistochemically evaluated in radical nephrectomy specimens and correlated with progression‑free survival during treatment with each molecular targeting agent. Sorafenib inhibited the expression of phospho-ERK and -4EBP1 in 769‑P cells; sunitinib, phospho-ERK and -4EBP1 in 786‑O and 769‑P cells; and everolimus, phospho-p70 in 786‑O and 769‑P cells. Knockdown of ERK reduced sensitivity to sorafenib in both cell lines, knockdown of ERK and 4EBP1 reduced sensitivity to sunitinib in 769‑P cells, and knockdown of 4EBP1 and p70 reduced sensitivity to everolimus in 786‑O cells. High expression of phospho-ERK, -4EBP1 and -p70 correlated with better progression‑free survival in patients treated with sorafenib, sunitinib and everolimus, respectively. Our results indicate that phospho-ERK, -4EBP1 and/or -ERK, and phospho-p70 can be used as biomarkers for the therapeutic efficacy of sorafenib, sunitinib and everolimus, respectively.

  16. A case of pneumocystis pneumonia associated with everolimus therapy for renal cell carcinoma.

    PubMed

    Saito, Yoshinobu; Nagayama, Mikie; Miura, Yukiko; Ogushi, Satoko; Suzuki, Yasutomo; Noro, Rintaro; Minegishi, Yuji; Kimura, Go; Kondo, Yukihiro; Gemma, Akihiko

    2013-05-01

    A 76-year-old female with advanced renal cell carcinoma had been treated with everolimus for 3 months. She visited our hospital because of a cough and fever lasting a few days. Chest X-rays showed bilateral infiltrative shadows, and a chest computed tomography scan showed homogeneous ground-glass opacities with mosaic patterns, especially in the apical region. The laboratory results revealed a decreased white blood cell count with lymphocytopenia and high levels of lactate dehydrogenase, C-reactive protein and KL-6. Pneumonitis was suspected and, therefore, everolimus therapy was interrupted. At that time, the pneumonitis was thought to be drug-induced interstitial lung disease. However, it was not possible to rule out pneumocystis pneumonia, because the patient was immunocompromised and the computed tomography findings suggested the possibility of pneumocystis pneumonia. The pneumonitis progressed rapidly and the patient developed respiratory failure, so we performed bronchoalveolar lavage to make a definitive diagnosis, and simultaneously started treatment with prednisolone and trimethoprim-sulfamethoxazole to cover both interstitial lung disease and pneumocystis pneumonia. A polymerase chain reaction assay of the bronchoalveolar lavage fluid was positive for Pneumocystis carinii DNA, and the serum level of β-d-glucan was significantly elevated. Thus, the patient was diagnosed with pneumocystis pneumonia, which was cured by the treatment. Interstitial lung disease is a major adverse drug reaction associated with everolimus, and interstitial lung disease is the first condition suspected when a patient presents with pneumonitis during everolimus therapy. Pneumocystis pneumonia associated with everolimus therapy is rare, but our experience suggests that pneumocystis pneumonia should be considered as a differential diagnosis when pneumonitis is encountered in patients receiving everolimus therapy.

  17. Combining cytochrome P-450 3A4 modulators and cyclosporine or everolimus in transplantation is successful

    PubMed Central

    González, Fernando; Valjalo, Ricardo

    2015-01-01

    AIM: To describe the long term follow-up of kidney allograft recipients receiving ketoconazole with calcineurin inhibitors (CNI) alone or combined with everolimus. METHODS: This is an open-label, prospective observational clinical trial in low immunologic risk patients who, after signing an Institutional Review Board approved consent form, were included in one of two groups. The first one (n = 59) received everolimus (target blood level, 3-8 ng/mL) and the other (n = 114) azathioprine 2 mg/kg per day or mycophenolate mofetyl (MMF) 2 g/d. Both groups also received tapering steroids, the cytochrome P-450 3A4 (CYP3A4) modulator, ketoconazole 50-100 mg/d, and cyclosporine with C0 targets in the everolimus group of 200-250 ng/mL in 1 mo, 100-125 ng/mL in 2 mo, and 50-65 ng/mL thereafter, and in the azathioprine or MMF group of 250-300 ng/mL in 1 mo, 200-250 ng/mL in 2 mo, 180-200 ng/mL until 3-6 mo, and 100-125 ng/mL thereafter. Clinical visits were performed monthly the first year and quarterly thereafter by treating physicians and all data was extracted by the investigators. RESULTS: The clinical characteristics of these two cohorts were similar. During the follow up (66 + 31 mo), both groups showed comparable clinical courses, but the biopsy proven acute rejection rate during the full follow-up period seemed to be lower in the everolimus group (20% vs 36%; P = 0.04). The everolimus group did not show a higher surgical complication rate than the other group. By the end of the follow-up period, the everolimus group tended to show a higher glomerular filtration rate. Nevertheless, we found no evidence of a consistent negative slope of the temporal allograft function estimated by the modification of the diet in renal disease formula in any of both groups. At 6 years of follow-up, the uncensored and death-censored graft survivals were 91% and 93%, and 91% and 83% in the everolimus plus cyclosporine, and cyclosporine alone groups, respectively. The addition of ketoconazole

  18. New drugs, new challenges for dermatologists: mucocutaneous ulcers secondary to everolimus.

    PubMed

    Pasin, Victor Pavan; Pereira, Amanda Regio; Carvalho, Kalline Andrade de; Paiva, João Marcos Góes de; Enokihara, Milvia Maria Simões e Silva; Porro, Adriana Maria

    2015-01-01

    Everolimus, a mammalian target of rapamycin inhibitor, is an emerging drug, which is being increasingly applied in oncology and solid organ transplantation. Oral ulcers are a frequent side effect associated with this immunosupressor. We report the case of a renal transplant recipient who developed disfiguring oral and perianal ulcers secondary to everolimus's toxicity. This is probably the first report of perianal involvement. Dermatologists need to be aware of the potential mucocutaneous adverse effects related to these new drugs that are becoming evermore common in our clinical practice.

  19. Inhibition of mTOR Pathway by Rapamycin Reduces Brain Damage in Rats Subjected to Transient Forebrain Ischemia

    PubMed Central

    Yang, Xiao; Hei, Changhun; Liu, Ping; Song, Yaozu; Thomas, Taylor; Tshimanga, Sylvie; Wang, Feng; Niu, Jianguo; Sun, Tao; Li, P. Andy

    2015-01-01

    The aims of this study are to clarify the role of mTOR in mediating cerebral ischemic brain damage and the effects of rapamycin on ischemic outcomes. Ten minutes of forebrain ischemia was induced in rats, and their brains were sampled after 3 h, 16 h, and 7 days reperfusion for histology, immunohistochemistry and biochemical analysis. Our data demonstrated that cerebral ischemia resulted in both apoptotic and necrotic neuronal death; cerebral ischemia and reperfusion led to significant increases of mRNA and protein levels of p-mTOR and its downstream p-P70S6K and p-S6; elevation of LC3-II, and release of cytochrome c into the cytoplasm in both the cortex and hippocampus. Inhibition of mTOR by rapamycin markedly reduced ischemia-induced damage; suppressed p-Akt, p-mTOR, p-P70S6K and p-S6 protein levels; decreased LC3-II and Beclin-1; and prevented cytochrome c release in the two structures. All together, these data provide evidence that cerebral ischemia activates mTOR and autophagy pathways. Inhibition of mTOR deactivates the mTOR pathway, suppresses autophagy, prevents cytochrome c release and reduces ischemic brain damage. PMID:26681922

  20. Inhibition of Histone Deacetylases (HDACs) and mTOR Signaling: Novel Strategies Towards the Treatment of Prostate Cancer

    DTIC Science & Technology

    2012-04-01

    of the combination of the HDAC inhibitor panobinostat with the mTORC1 inhibitor everolimus . Panobinostat/ everolimus combination treatment resulted in...to single treatments. We identified that panobinostat/ everolimus combination resulted in enhanced anti-tumor activity mediated by decreased tumor...that low dose concurrent panobinostat/ everolimus combination therapy is well tolerated and results in greater anti-tumor activity and therapeutic

  1. The mTOR inhibitor sirolimus suppresses renal, hepatic, and cardiac tissue cellular respiration.

    PubMed

    Albawardi, Alia; Almarzooqi, Saeeda; Saraswathiamma, Dhanya; Abdul-Kader, Hidaya Mohammed; Souid, Abdul-Kader; Alfazari, Ali S

    2015-01-01

    The purpose of this in vitro study was to develop a useful biomarker (e.g., cellular respiration, or mitochondrial O2 consumption) for measuring activities of mTOR inhibitors. It measured the effects of commonly used immunosuppressants (sirolimus-rapamycin, tacrolimus, and cyclosporine) on cellular respiration in target tissues (kidney, liver, and heart) from C57BL/6 mice. The mammalian target of rapamycin (mTOR), a serine/ threonine kinase that supports nutrient-dependent cell growth and survival, is known to control energy conversion processes within the mitochondria. Consistently, inhibitors of mTOR (e.g., rapamycin, also known as sirolimus or Rapamune®) have been shown to impair mitochondrial function. Inhibitors of the calcium-dependent serine/threonine phosphatase calcineurin (e.g., tacrolimus and cyclosporine), on the other hand, strictly prevent lymphokine production leading to a reduced T-cell function. Sirolimus (10 μM) inhibited renal (22%, P=0.002), hepatic (39%, P<0.001), and cardiac (42%, P=0.005) cellular respiration. Tacrolimus and cyclosporine had no or minimum effects on cellular respiration in these tissues. Thus, these results clearly demonstrate that impaired cellular respiration (bioenergetics) is a sensitive biomarker of the immunosuppressants that target mTOR.

  2. The role of mTOR signalling in the regulation of skeletal muscle mass in a rodent model of resistance exercise

    PubMed Central

    Ogasawara, Riki; Fujita, Satoshi; Hornberger, Troy A.; Kitaoka, Yu; Makanae, Yuhei; Nakazato, Koichi; Naokata, Ishii

    2016-01-01

    Resistance exercise (RE) activates signalling by the mammalian target of rapamycin (mTOR), and it has been suggested that rapamycin-sensitive mTOR signalling controls RE-induced changes in protein synthesis, ribosome biogenesis, autophagy, and the expression of peroxisome proliferator gamma coactivator 1 alpha (PGC-1α). However, direct evidence to support the aforementioned relationships is lacking. Therefore, in this study, we investigated the role of rapamycin-sensitive mTOR in the RE-induced activation of muscle protein synthesis, ribosome biogenesis, PGC-1α expression and hypertrophy. The results indicated that the inhibition of rapamycin-sensitive mTOR could prevent the induction of ribosome biogenesis by RE, but it only partially inhibited the activation of muscle protein synthesis. Likewise, the inhibition of rapamycin-sensitive mTOR only partially blocked the hypertrophic effects of chronic RE. Furthermore, both acute and chronic RE promoted an increase in PGC-1α expression and these alterations were not affected by the inhibition of rapamycin-sensitive mTOR. Combined, the results from this study not only establish that rapamycin-sensitive mTOR plays an important role in the RE-induced activation of protein synthesis and the induction of hypertrophy, but they also demonstrate that additional (rapamycin-sensitive mTOR-independent) mechanisms contribute to these fundamentally important events. PMID:27502839

  3. Assessing Metabolic Changes in Response to mTOR Inhibition in a Mantle Cell Lymphoma Xenograft Model Using AcidoCEST MRI

    PubMed Central

    Akhenblit, Paul J.; Hanke, Neale T.; Gill, Alexander; Persky, Daniel O.; Howison, Christine M.; Pagel, Mark D.; Baker, Amanda F.

    2016-01-01

    AcidoCEST magnetic resonance imaging (MRI) has previously been shown to measure tumor extracellular pH (pHe) with excellent accuracy and precision. This study investigated the ability of acidoCEST MRI to monitor changes in tumor pHe in response to therapy. To perform this study, we used the Granta 519 human mantle cell lymphoma cell line, which is an aggressive B-cell malignancy that demonstrates activation of the phosphatidylinositol-3-kinase/Akt/mammalian target of rapamycin (mTOR) pathway. We performed in vitro and in vivo studies using the Granta 519 cell line to investigate the efficacy and associated changes induced by the mTOR inhibitor, everolimus (RAD001). AcidoCEST MRI studies showed a statistically significant increase in tumor pHe of 0.10 pH unit within 1 day of initiating treatment, which foreshadowed a decrease in tumor growth of the Granta 519 xenograft model. AcidoCEST MRI then measured a decrease in tumor pHe 7 days after initiating treatment, which foreshadowed a return to normal tumor growth rate. Therefore, this study is a strong example that acidoCEST MRI can be used to measure tumor pHe that may serve as a marker for therapeutic efficacy of anticancer therapies. PMID:27140422

  4. Modulating mTOR in aging and health.

    PubMed

    Johnson, Simon C; Sangesland, Maya; Kaeberlein, Matt; Rabinovitch, Peter S

    2015-01-01

    The physiological responses to nutrient availability play a central role in aging and disease. Genetic and pharmacological studies have identified highly conserved cellular signaling pathways that influence aging by regulating the interface between nutrient and hormone cues and cellular growth and maintenance. Among these pathways, the mechanistic target of rapamycin (mTOR) has been most reproducibly shown to modulate aging in evolutionarily diverse organisms as reduction in mTOR activity extends life span from yeast to rodents. mTOR has been shown to play a role in a broad range of diseases, and is of particular interest to human health and aging due to the availability of clinically approved pharmacological agents targeting the mTOR complexes and other components of the mTOR signaling network. Characterizing the role of mTOR in aging and health promises to provide new avenues for intervention in human aging and disease through modulation of this signaling pathway.

  5. Combination of clopidogrel and everolimus dramatically reduced the development of transplant arteriosclerosis in murine aortic allografts.

    PubMed

    Eckl, Sebastian; Heim, Christian; Abele-Ohl, Silke; Hoffmann, Julia; Ramsperger-Gleixner, Martina; Weyand, Michael; Ensminger, Stephan M

    2010-09-01

    Our group has shown that platelet inhibition with clopidogrel, an antagonist of the P2Y12 adenosine diphosphate receptor on platelets, reduced the formation of transplant arteriosclerosis. The aim of this study was to investigate whether a combination of cyclosporin or everolimus with clopidogrel has a beneficial effect on the development of transplant arteriosclerosis. Fully MHC mismatched C57Bl/6 (H2(b)) donor aortas were transplanted into CBA.J (H2(k)) recipients and mice received either clopidogrel alone (1 mg/kg/day) or in combination with cyclosporin (2 mg/kg/day) or everolimus (0.05 mg/kg/day). Grafts were analysed by histology and morphometry on day 30 after transplantation. In mice treated with clopidogrel alone, transplant arteriosclerosis was significantly reduced [intima proliferation 56 +/- 11% vs. 81 +/- 7% (control)/n = 7]. Daily application of everolimus reduced the development of transplant arteriosclerosis compared with untreated controls [intima proliferation of 29 +/- 9% vs. 81 +/- 7% (control)/n = 7]. Strikingly, combination of clopidogrel and everolimus almost abolished the formation of transplant arteriosclerosis [intima proliferation: 11 +/- 8% vs. 81 +/- 7% (control)/n = 7]. By contrast, combination of cyclosporin and clopidogrel compared with clopidogrel alone showed no additive effect. These results demonstrate that combination of platelet- and mammalian target of Rapamycin-inhibition can dramatically reduce the development of transplant arteriosclerosis.

  6. Phase I Study of Capecitabine, Oxaliplatin, Bevacizumab, and Everolimus in Advanced Solid Tumors

    PubMed Central

    Rangwala, F.; Bendell, J.; Kozloff, M.; Arrowood, C.; Dellinger, A.; Meadows, J.; Tourt-Uhlig, S.; Murphy, J.; Meadows, K.L.; Starr, A.; Broderick, S.; Brady, J.C.; Cushman, S. M.; Morse, M.; Uronis, H.; Hsu, S.D.; Zafar, S.Y.; Wallace, J.; Starodub, A.; Strickler, J.; Pang, H.; Nixon, A.B.; Hurwitz, H.

    2014-01-01

    Purpose To define maximum tolerated dose (MTD), toxicities, and pharmacodynamics of capecitabine, oxaliplatin, bevacizumab, and everolimus in advanced solid tumor patients. Design This was a standard “3+3” dose-escalation trial. All subjects received bevacizumab 7.5mg/kg on day one of each cycle. Doses for capecitabine, oxaliplatin and everolimus were modified per dose limiting toxicity (DLT). Baseline and on-treatment plasma biomarkers were analyzed. Archived tumor mRNA levels were evaluated for NRP1, NRP2 and VEGF-A isoforms. Results Twenty-nine patients were evaluable for toxicity and 30 for efficacy. Two DLTs were observed in cohort 1 and one DLT each was observed in cohort -1 and -1b. Grade ≥3 toxicities included neutropenia, hypertension, perforation/fistula/hemorrhage, hypertriglyceridemia, diarrhea, and thromboembolism. Twelve subjects experienced partial response (PR); 12 had stable disease as best response. Three of seven chemorefractory metastatic colorectal cancer (mCRC) subjects experienced PR; eight of 15 chemonaive mCRC subjects experienced PR. Plasma TβRIII and IL-6 increased on treatment but without correlation to outcome. Increased VEGF165 levels significantly correlated with longer progression free survival. Conclusions Everolimus with full dose capecitabine, oxaliplatin, and bevacizumab had unacceptable toxicity. MTD was: everolimus 5mg daily; capecitabine 680mg/m2 BID days 1-14; oxaliplatin 100mg/m2 and bevacizumab 7.5mg/kg, day one. Activity was noted in mCRC. PMID:24711126

  7. Emerging evidence of a link between the polycystins and the mTOR pathways.

    PubMed

    Boletta, Alessandra

    2009-10-28

    Autosomal dominant polycystic kidney disease (ADPKD) is a genetic disease characterized by the formation of renal cysts. This disease can be caused by mutations in two genes, PKD1 and PKD2, which encode polycystin-1 (PC-1) and -2 (PC-2), respectively.PC-1 is a large plasma membrane receptor involved in the regulation of several biological functions and signaling pathways, and PC-2 is a calcium channel of the TRP family. The two proteins associate in a complex to prevent cyst formation, but the precise mechanism(s) involved remain largely unknown.This review will focus on recent advances in our understanding of the functions of polycystins and their role in signal transduction.Increased activity of the mammalian target of rapamycin (mTOR) kinase has been observed in cysts found in ADPKD tissues. Rapamycin has been shown to have beneficial effects in rodent models of polycystic kidney disease, prompting the initiation of pilot clinical trials with human patients. Furthermore, a direct role for PC-1 in the regulation of cell growth (size) via mTOR has recently been demonstrated.Major advancements in the study of mTOR biology have highlighted that this kinase exists in association with two different complexes, mTOR complex 1 (mTORC1) and mTOR complex 2 (mTORC2). The mTORC1 complex regulates cell growth (size), proliferation, translation and autophagy, and mTORC2 regulates the actin cytoskeleton and apoptosis. Interestingly, mTORC2 has been shown to contain the kinase responsible for the phosphorylation of Akt at Serine 473. Previous studies have shown that PC-1 controls the PI 3-kinase/Akt cascade to regulate apoptosis and the actin cytoskeleton, suggesting that this receptor might regulate mTOR at several levels.This review aims to discuss three different, inter-related themes emerging from the literature: (i) studies performed in our and other laboratories collectively suggest that PC-1 might be able to differentially regulate the two mTOR complexes; (ii) several

  8. The mechanical activation of mTOR signaling: an emerging role for late endosome/lysosomal targeting.

    PubMed

    Jacobs, Brittany L; Goodman, Craig A; Hornberger, Troy A

    2014-02-01

    It is well recognized that mechanical signals play a critical role in the regulation of skeletal muscle mass, and the maintenance of muscle mass is essential for mobility, disease prevention and quality of life. Furthermore, over the last 15 years it has become established that signaling through a protein kinase called the mammalian (or mechanistic) target of rapamycin (mTOR) is essential for mechanically-induced changes in protein synthesis and muscle mass, however, the mechanism(s) via which mechanical stimuli regulate mTOR signaling have not been defined. Nonetheless, advancements are being made, and an emerging body of evidence suggests that the late endosome/lysosomal (LEL) system might play a key role in this process. Therefore, the purpose of this review is to summarize this body of evidence. Specifically, we will first explain why the Ras homologue enriched in brain (Rheb) and phosphatidic acid (PA) are considered to be direct activators of mTOR signaling. We will then describe the process of endocytosis and its involvement in the formation of LEL structures, as well as the evidence which indicates that mTOR and its direct activators (Rheb and PA) are all enriched at the LEL. Finally, we will summarize the evidence that has implicated the LEL in the regulation of mTOR by various growth regulatory inputs such as amino acids, growth factors and mechanical stimuli.

  9. Chronic Repression of mTOR Complex 2 Induces Changes in the Gut Microbiota of Diet-induced Obese Mice

    PubMed Central

    Jung, Mi-Ja; Lee, Jina; Shin, Na-Ri; Kim, Min-Soo; Hyun, Dong-Wook; Yun, Ji-Hyun; Kim, Pil Soo; Whon, Tae Woong; Bae, Jin-Woo

    2016-01-01

    Alterations in the gut microbiota play a crucial role in host physiology and metabolism; however, the molecular pathways underlying these changes in diet-induced obesity are unclear. Mechanistic target of rapamycin (mTOR) signaling pathway is associated with metabolic disorders such as obesity and type 2 diabetes (T2D). Therefore, we examined whether changes in the regulation of mTOR signaling induced by diet (a high-fat diet [HFD] or normal-chow diet) and/or therapeutics (resveratrol [a specific inhibitor of mTOR complex 1] or rapamycin [an inhibitor of both mTOR complex 1 and 2]) altered the composition of the gut microbiota in mice. Oral administration of resveratrol prevented glucose intolerance and fat accumulation in HFD-fed mice, whereas rapamycin significantly impaired glucose tolerance and exacerbated intestinal inflammation. The abundance of Lactococcus, Clostridium XI, Oscillibacter, and Hydrogenoanaerobacterium increased under the HFD condition; however, the abundance of these species declined after resveratrol treatment. Conversely, the abundance of unclassified Marinilabiliaceae and Turicibacter decreased in response to a HFD or rapamycin. Taken together, these results demonstrated that changes in the composition of intestinal microbiota induced by changes in mTOR activity correlate with obese and diabetic phenotypes. PMID:27471110

  10. Everolimus in combination with cyclosporin a as pre- and posttransplantation immunosuppressive therapy in nonmyeloablative allogeneic hematopoietic stem cell transplantation.

    PubMed

    Junghanss, Christian; Rathsack, Susanne; Wacke, Rainer; Weirich, Volker; Vogel, Heike; Drewelow, Bernd; Mueller, Sabrina; Altmann, Simone; Freund, Mathias; Lange, Sandra

    2012-07-01

    Everolimus (RAD001) is an mTOR inhibitor that has been successfully used as an immunosuppressant in solid-organ transplantation. Data in allogeneic hematopoietic stem cell transplantation (HSCT) is limited. This study aimed to investigate pharmacokinetics, safety, and efficacy of RAD001 in a canine allogeneic HSCT model. First, pharmacokinetics of RAD001 were performed in healthy dogs in order to determine the appropriate dosing. Doses of 0.25 mg RAD001 twice daily in combination with 15 mg/kg cyclosporin A (CsA) twice daily were identified as appropriate starting doses to achieve the targeted range of RAD001 (3-8 μg/L) when orally administered. Subsequently, 10 dogs were transplanted using 2 Gy total body irradiation (TBI) for conditioning and 0.25 mg RAD001 twice daily plus 15 mg/kg CsA twice daily for pre- and posttransplantation immunosuppression. Seven of the 10 transplanted dogs were maintained at the starting RAD001 dose throughout the study. For the remaining 3 dogs, dose adjustments were necessary. RAD001 accumulation over time did not occur. All dogs initially engrafted. Five dogs eventually rejected the graft (weeks 10, 10, 13, 27, and 56). Two dogs died of pneumonia (weeks 8 and 72) but were chimeric until then. Total cholesterol rose from median 4.1 mmol/L (3.5-5.7 mmol/L) before HSCT to 6.0 mmol/l (5.0-8.5 mmol/l) at day 21 after HSCT, but remained always within normal range. Changes in creatinine and triglyceride values were not observed. Long-term engraftment rates were inferior to sirolimus/CsA and mycophenolate mofetil (MMF)/CsA regimen, respectively. RAD001/CsA caused a more pronounced reduction of platelet counts to median 2 × 10(9)/L (range: 0-21 × 10(9)/L) and longer time to platelet recovery of 21 days (range: 14-24 days) compared with MMF/CsA. CsA c(2h) levels were significantly enhanced in the RAD001/CsA regimen, but c(0h) and area under the curve from 0 to 12 hours (AUC(0-12h)) values did not differ compared with an MMF

  11. Renal function improvement in liver transplant recipients after early everolimus conversion: A clinical practice cohort study in Spain.

    PubMed

    Bilbao, Itxarone; Salcedo, Magdalena; Gómez, Miguel Angel; Jimenez, Carlos; Castroagudín, Javier; Fabregat, Joan; Almohalla, Carolina; Herrero, Ignacio; Cuervas-Mons, Valentín; Otero, Alejandra; Rubín, Angel; Miras, Manuel; Rodrigo, Juan; Serrano, Trinidad; Crespo, Gonzalo; De la Mata, Manuel; Bustamante, Javier; Gonzalez-Dieguez, M Luisa; Moreno, Antonia; Narvaez, Isidoro; Guilera, Magda

    2015-08-01

    A national, multicenter, retrospective study was conducted to assess the results obtained for liver transplant recipients with conversion to everolimus in daily practice. The study included 477 recipients (481 transplantations). Indications for conversion to everolimus were renal dysfunction (32.6% of cases), hepatocellular carcinoma (HCC; 30.2%; prophylactic treatment for 68.9%), and de novo malignancy (29.7%). The median time from transplantation to conversion to everolimus was 68.7 months for de novo malignancy, 23.8 months for renal dysfunction, and 7.1 months for HCC and other indications. During the first year of treatment, mean everolimus trough levels were 5.4 (standard deviation [SD], 2.7) ng/mL and doses remained stable (1.5 mg/day) from the first month after conversion. An everolimus monotherapy regimen was followed by 28.5% of patients at 12 months. Patients with renal dysfunction showed a glomerular filtration rate (4-variable Modification of Diet in Renal Disease) increase of 10.9 mL (baseline mean, 45.8 [SD, 25.3] versus 57.6 [SD, 27.6] mL/minute/1.73 m(2) ) at 3 months after everolimus initiation (P < 0.001), and 6.8 mL at 12 months. Improvement in renal function was higher in patients with early conversion (<1 year). Adverse events were the primary reason for discontinuation in 11.2% of cases. The probability of survival at 3 years after conversion to everolimus was 83.0%, 71.1%, and 59.5% for the renal dysfunction, de novo malignancy, and HCC groups, respectively. Everolimus is a viable option for the treatment of renal dysfunction, and earlier conversion is associated with better recovery of renal function. Prospective studies are needed to confirm advantages in patients with malignancy.

  12. Five-year outcomes in kidney transplant patients converted from cyclosporine to everolimus: the randomized ZEUS study.

    PubMed

    Budde, K; Lehner, F; Sommerer, C; Reinke, P; Arns, W; Eisenberger, U; Wüthrich, R P; Mühlfeld, A; Heller, K; Porstner, M; Veit, J; Paulus, E-M; Witzke, O

    2015-01-01

    ZEUS study was an open-label, 12-month, multicenter study in which 300 de novo kidney transplant recipients were randomized to continue receiving cyclosporine (CsA) or convert to everolimus at 4.5 months posttransplant. Five-year follow-up data were available for 245/269 patients (91.1%) who completed the core 12-month study (123 everolimus, 109 CsA). At 5 years, adjusted estimated GFR was 66.2 mL/min/1.73 m(2) with everolimus versus 60.9 mL/min/1.73 m(2) with CsA; the mean difference was 5.3 mL/min/1.73 m(2) in favor of everolimus (95% CI 2.4, 8.3; p < 0.001 [intent-to-treat population]). In a post hoc analysis of patients remaining on study drug at 5 years (everolimus 77, CsA 86), mean difference was 8.2 mL/min/1.73 m(2) (95% CI 4.3, 12.1; p < 0.001) in favor of everolimus. The cumulative incidence of biopsy-proven acute rejection postrandomization was 13.6% with everolimus versus 7.5% with CsA (p = 0.095), largely accounted for by grade I rejection (16/21 patients and 7/11 patients, respectively). Postrandomization, graft loss, mortality, serious adverse events and neoplasms were similar in both arms. In conclusion, conversion of kidney transplant patients to everolimus at 4.5 months posttransplant is associated with a significant improvement in renal function that is maintained to at least 5 years. The increase in early mild acute rejection did not affect long-term graft function.

  13. mTOR Inhibitors Alone and in Combination with JAK2 Inhibitors Effectively Inhibit Cells of Myeloproliferative Neoplasms

    PubMed Central

    Martinelli, Serena; Tozzi, Lorenzo; Guglielmelli, Paola; Bosi, Alberto; Vannucchi, Alessandro M.

    2013-01-01

    Background Dysregulated signaling of the JAK/STAT pathway is a common feature of chronic myeloproliferative neoplasms (MPN), usually associated with JAK2V617F mutation. Recent clinical trials with JAK2 inhibitors showed significant improvements in splenomegaly and constitutional symptoms in patients with myelofibrosis but meaningful molecular responses were not documented. Accordingly, there remains a need for exploring new treatment strategies of MPN. A potential additional target for treatment is represented by the PI3K/AKT/mammalian target of rapamycin (mTOR) pathway that has been found constitutively activated in MPN cells; proof-of-evidence of efficacy of the mTOR inhibitor RAD001 has been obtained recently in a Phase I/II trial in patients with myelofibrosis. The aim of the study was to characterize the effects in vitro of mTOR inhibitors, used alone and in combination with JAK2 inhibitors, against MPN cells. Findings Mouse and human JAK2V617F mutated cell lines and primary hematopoietic progenitors from MPN patients were challenged with an allosteric (RAD001) and an ATP-competitive (PP242) mTOR inhibitor and two JAK2 inhibitors (AZD1480 and ruxolitinib). mTOR inhibitors effectively reduced proliferation and colony formation of cell lines through a slowed cell division mediated by changes in cell cycle transition to the S-phase. mTOR inhibitors also impaired the proliferation and prevented colony formation from MPN hematopoietic progenitors at doses significantly lower than healthy controls. JAK2 inhibitors produced similar antiproliferative effects in MPN cell lines and primary cells but were more potent inducers of apoptosis, as also supported by differential effects on cyclinD1, PIM1 and BcLxL expression levels. Co-treatment of mTOR inhibitor with JAK2 inhibitor resulted in synergistic activity against the proliferation of JAK2V617F mutated cell lines and significantly reduced erythropoietin-independent colony growth in patients with polycythemia vera

  14. Discovery – Targeted Treatments and mTOR Inhibitors

    Cancer.gov

    Thanks to discovering the anticancer effects of mTOR inhibitors, cancer treatment for pNet, a rare type of pancreatic cancer, were revolutionized. Through clinical trials, NCI continues to investigate the life-saving potential of mTOR inhibitors.

  15. Transcriptional regulation of the stress response by mTOR.

    PubMed

    Aramburu, Jose; Ortells, M Carmen; Tejedor, Sonia; Buxadé, Maria; López-Rodríguez, Cristina

    2014-07-01

    The kinase mammalian target of rapamycin (mTOR) is a central regulator of cell growth and proliferation that integrates inputs from growth factor receptors, nutrient availability, intracellular ATP (adenosine 5'-triphosphate), and a variety of stressors. Since early works in the mid-1990s uncovered the role of mTOR in stimulating protein translation, this kinase has emerged as a rather multifaceted regulator of numerous processes. Whereas mTOR is generally activated by growth- and proliferation-stimulating signals, its activity can be reduced and even suppressed when cells are exposed to a variety of stress conditions. However, cells can also adapt to stress while maintaining their growth capacity and mTOR function. Despite knowledge accumulated on how stress represses mTOR, less is known about mTOR influencing stress responses. In this review, we discuss the capability of mTOR, in particular mTOR complex 1 (mTORC1), to activate stress-responsive transcription factors, and we outline open questions for future investigation.

  16. TGF-β inhibits the activation and functions of NK cells by repressing the mTOR pathway.

    PubMed

    Viel, Sébastien; Marçais, Antoine; Guimaraes, Fernando Souza-Fonseca; Loftus, Roisin; Rabilloud, Jessica; Grau, Morgan; Degouve, Sophie; Djebali, Sophia; Sanlaville, Amélien; Charrier, Emily; Bienvenu, Jacques; Marie, Julien C; Caux, Christophe; Marvel, Jacqueline; Town, Liam; Huntington, Nicholas D; Bartholin, Laurent; Finlay, David; Smyth, Mark J; Walzer, Thierry

    2016-02-16

    Transforming growth factor-β (TGF-β) is a major immunosuppressive cytokine that maintains immune homeostasis and prevents autoimmunity through its antiproliferative and anti-inflammatory properties in various immune cell types. We provide genetic, pharmacologic, and biochemical evidence that a critical target of TGF-β signaling in mouse and human natural killer (NK) cells is the serine and threonine kinase mTOR (mammalian target of rapamycin). Treatment of mouse or human NK cells with TGF-β in vitro blocked interleukin-15 (IL-15)-induced activation of mTOR. TGF-β and the mTOR inhibitor rapamycin both reduced the metabolic activity and proliferation of NK cells and reduced the abundances of various NK cell receptors and the cytotoxic activity of NK cells. In vivo, constitutive TGF-β signaling or depletion of mTOR arrested NK cell development, whereas deletion of the TGF-β receptor subunit TGF-βRII enhanced mTOR activity and the cytotoxic activity of the NK cells in response to IL-15. Suppression of TGF-β signaling in NK cells did not affect either NK cell development or homeostasis; however, it enhanced the ability of NK cells to limit metastases in two different tumor models in mice. Together, these results suggest that the kinase mTOR is a crucial signaling integrator of pro- and anti-inflammatory cytokines in NK cells. Moreover, we propose that boosting the metabolic activity of antitumor lymphocytes could be an effective strategy to promote immune-mediated tumor suppression.

  17. A role for everolimus in post-transplant encapsulating peritoneal sclerosis: first case report.

    PubMed

    Sud, Rahul; Garry, Lorraine; Spicer, Stephen Timothy; Allen, Richard D M; Eris, Josette M; Wyburn, Kate; Verran, Deborah; Cooper, Caroline Louise; Chadban, Steve

    2014-04-01

    Encapsulating peritoneal sclerosis (EPS) is a rare complication of peritoneal dialysis (PD) that carries a high morbidity and mortality. The 'two hit theory' suggests that long term deterioration of the peritoneum combined with intraperitoneal inflammation is needed in the pathogenesis of EPS. For unclear reasons, post transplantation EPS is being increasingly reported in patients previously on PD. To date, there is no proven effective therapy with an absence of randomised controlled trials. Individual case reports and small case series have reported on the use of tamoxifen and corticosteroids for medical management of EPS. The use of everolimus has been reported in a single case, and never in the setting of renal transplantation. Here, we present the first case of post-transplant encapsulating peritoneal sclerosis treated successfully with a combination of everolimus, tamoxifen, low dose corticosteroid and surgery.

  18. Metformin with everolimus and octreotide in pancreatic neuroendocrine tumor patients with diabetes.

    PubMed

    Pusceddu, Sara; Buzzoni, Roberto; Vernieri, Claudio; Concas, Laura; Marceglia, Sara; Giacomelli, Luca; Milione, Massimo; Leuzzi, Livia; Femia, Daniela; Formisano, Barbara; Mazzaferro, Vincenzo; de Braud, Filippo

    2016-05-01

    A bidirectional relationship seems to exist between diabetes mellitus and development of pancreatic tumors. Metformin, the most widely used drug in the treatment of Type 2 diabetes mellitus, has recently emerged as a potentially active agent in cancer chemoprevention and treatment. In this article, we discuss the potential correlation between glycemic status, administration of antiglycemic treatments, such as metformin or insulin, and prognosis of pancreatic neuroendocrine tumors patients treated with everolimus and octreotide, on the basis of existing evidence and our experience.

  19. Safety, Efficacy, and Patient Acceptability of Everolimus in the Treatment of Breast Cancer.

    PubMed

    Lousberg, Laurence; Jerusalem, Guy

    2016-01-01

    Everolimus combined with exemestane is an important treatment option for patients suffering from estrogen receptor-positive, human epidermal growth factor receptor 2-negative, advanced breast cancer (ABC) who have been previously treated with a nonsteroidal aromatase inhibitor (NSAI). After presentation of phase III registration trial BOLERO-2, several phase IIIb trials have been started to evaluate this regimen in a more real-world setting. Here, we review the efficacy and safety data published or presented at selected international meetings. These studies confirmed the outcome observed in the BOLERO-2 trial. Patient acceptance rate is also discussed by focusing on the permanent everolimus discontinuation rate in these trials. Factors influencing the safety profile are also reported, including the impact of age. The optimal sequence of combined therapy approaches associating targeted and endocrine therapy (ET) has yet to be determined as new treatment options such as cyclin-dependent kinase inhibitors become available. However, everolimus-exemestane remains an important treatment option with a major impact on progression-free survival (PFS) and an acceptable safety profile.

  20. Combination of Leflunomide and Everolimus for treatment of BK virus nephropathy.

    PubMed

    Jaw, Juli; Hill, Prue; Goodman, David

    2017-04-01

    BK nephropathy (BKN) is a common cause of graft dysfunction following kidney transplantation. Minimization of immunosuppressive therapy remains the first line of therapy, but this may lead to rejection and graft loss. In some cases, despite lowering immunosuppression, BK infection can persist, leading to chronic damage and kidney failure. Currently, there is no specific anti-BK viral therapy. Recent in vitro experiments have demonstrated a reduction in BK viral replication when infected cells are treated with the combination of Leflunomide and Everolimus. This study aims to explore the effect of this drugs combination on viral clearance and graft function in patients with persistent disease despite reduction in immunosuppression. We treated three patients with combination Leflunomide and Everolimus. Data on medical history, biochemical parameters and viral loads were collected. Significant improvement in viral loads was observed in two cases with resolution of viremia in another (Table 1). Two recipients had preserved allograft function. The remaining graft was lost because of combination of obstruction and BKN. No adverse reactions such as bone marrow toxicity were observed. Combination of Leflunomide and Everolimus is safe and should be considered as a rescue therapy in treatment of BKN, especially in those who fail to clear this infection despite reduction of immunosuppressive therapy.

  1. Deregulation of mTOR signaling is involved in thymic lymphoma development in Atm-/- mice

    SciTech Connect

    Kuang, Xianghong; Shen, Jianjun; Wong, Paul K.Y.; Yan, Mingshan

    2009-06-05

    Abnormal thymocyte development with thymic lymphomagenesis inevitably occurs in Atm-/- mice, indicating that ATM plays a pivotal role in regulating postnatal thymocyte development and preventing thymic lymphomagenesis. The mechanism for ATM controls these processes is unclear. We have shown previously that c-Myc, an oncoprotein regulated by the mammalian target of rapamycin (mTOR), is overexpressed in Atm-/- thymocytes. Here, we show that inhibition of mTOR signaling with its specific inhibitor, rapamycin, suppresses normal thymocyte DNA synthesis by downregulating 4EBP1, but not S6K, and that 4EBP1 phosphorylation and cyclin D1 expression are coordinately increased in Atm-/- thymocytes. Administration of rapamycin to Atm-/- mice attenuates elevated phospho-4EBP1, c-Myc and cyclin D1 in their thymocytes, and delays thymic lymphoma development. These results indicate that mTOR downstream effector 4EBP1 is essential for normal thymocyte proliferation, but deregulation of 4EBP1 in Atm deficiency is a major factor driving thymic lymphomagenesis in the animals.

  2. Regulation of mTOR by mechanically induced signaling events in skeletal muscle.

    PubMed

    Hornberger, Troy Alan; Sukhija, Kunal Balu; Chien, Shu

    2006-07-01

    Mechanical stimuli play a major role in the regulation of skeletal muscle mass, and the maintenance of muscle mass contributes significantly to disease prevention and the quality of life. Although a link between mechanical stimuli and the regulation of muscle mass has been recognized for decades, the mechanisms involved in converting mechanical information into the molecular events that control this process have not been defined. Nevertheless, significant advancements are being made in this field, and it has recently been established that signaling through a rapamycin-sensitive pathway is necessary for mechanically induced growth of skeletal muscle. Since rapamycin is a highly specific inhibitor of a protein kinase called the mammalian target of rapamycin (mTOR), many investigators have concluded that mTOR signaling is necessary for the mechanically induced growth of skeletal muscle. In this review, we have summarized the current knowledge regarding how mechanical stimuli activate mTOR signaling, discussed the newly discovered role of phospholipase D (PLD) and phosphatidic acid (PA) in this pathway, and considered the potential roles of PLD and PA in the mechanical regulation of skeletal muscle mass.

  3. Activation of mTOR (mechanistic target of rapamycin) in rheumatic diseases

    PubMed Central

    Perl, Andras

    2017-01-01

    Mechanistic target of rapamycin (mTOR, also known as mammalian target of rapamycin) is a ubiquitous serine/threonine kinase that regulates cell growth, proliferation and survival. These effects are cell-type-specific, and are elicited in response to stimulation by growth factors, hormones and cytokines, as well as to internal and external metabolic cues. Rapamycin was initially developed as an inhibitor of T-cell proliferation and allograft rejection in the organ transplant setting. Subsequently, its molecular target (mTOR) was identified as a component of two interacting complexes, mTORC1 and mTORC2, that regulate T-cell lineage specification and macrophage differentiation. mTORC1 drives the proinflammatory expansion of T helper (TH) type 1, TH17, and CD4−CD8− (double-negative, DN) T cells. Both mTORC1 and mTORC2 inhibit the development of CD4+CD25+FoxP3+ T regulatory (TREG) cells and, indirectly, mTORC2 favours the expansion of Tfollicular helper (TFH) cells which, similarly to DN T cells, promote B-cell activation and autoantibody production. In contrast to this proinflammatory effect of mTORC2, mTORC1 favours, to some extent, an anti-inflammatory macrophage polarization that is protective against infections and tissue inflammation. Outside the immune system, mTORC1 controls fibroblast proliferation and chondrocyte survival, with implications for tissue fibrosis and osteoarthritis, respectively. Rapamycin (which primarily inhibits mTORC1), ATP-competitive, dual mTORC1/mTORC2 inhibitors and upstream regulators of the mTOR pathway are being developed to treat autoimmune, hyperproliferative and degenerative diseases. In this regard, mTOR blockade promises to increase life expectancy through treatment and prevention of rheumatic diseases. PMID:26698023

  4. mTOR signaling in growth control and disease

    PubMed Central

    Laplante, Mathieu; Sabatini, David M.

    2012-01-01

    The mammalian target of rapamycin (mTOR) signaling pathway senses and integrates a variety of environmental cues to regulate organismal growth and homeostasis. The pathway regulates many major cellular processes and is implicated in an increasing number of pathological conditions, including cancer, obesity, type 2 diabetes, and neurodegeneration. Here, we review recent advances in our understanding of the mTOR pathway and its role in health and disease as well as aging. We further discuss pharmacological approaches to treat human pathologies linked to mTOR deregulation. PMID:22500797

  5. The MANDELA study: A multicenter, randomized, open-label, parallel group trial to refine the use of everolimus after heart transplantation.

    PubMed

    Deuse, Tobias; Bara, Christoph; Barten, Markus J; Hirt, Stephan W; Doesch, Andreas O; Knosalla, Christoph; Grinninger, Carola; Stypmann, Jörg; Garbade, Jens; Wimmer, Peter; May, Christoph; Porstner, Martina; Schulz, Uwe

    2015-11-01

    In recent years a series of trials has sought to define the optimal protocol for everolimus-based immunosuppression in heart transplantation, with the goal of minimizing exposure to calcineurin inhibitors (CNIs) and harnessing the non-immunosuppressive benefits of everolimus. Randomized studies have demonstrated that immunosuppressive potency can be maintained in heart transplant patients receiving everolimus despite marked CNI reduction, although very early CNI withdrawal may be inadvisable. A potential renal advantage has been shown for everolimus, but the optimal time for conversion and the adequate reduction in CNI exposure remain to be defined. Other reasons for use of everolimus include a substantial reduction in the risk of cytomegalovirus infection, and evidence for inhibition of cardiac allograft vasculopathy, a major cause of graft loss. The ongoing MANDELA study is a 12-month multicenter, randomized, open-label, parallel-group study in which efficacy, renal function and safety are compared in approximately 200 heart transplant patients. Patients receive CNI therapy, steroids and everolimus or mycophenolic acid during months 3 to 6 post-transplant, and are then randomized at month 6 post-transplant (i) to convert to CNI-free immunosuppression with everolimus and mycophenolic acid or (ii) to continue reduced-exposure CNI, with concomitant everolimus. Patients are then followed to month 18 post-transplant The rationale and expectations for the trial and its methodology are described herein.

  6. [Experience with everolimus therapy for patients with metastatic renal cancer in Hungary].

    PubMed

    Maráz, Anikó; Bodoky, György; Dank, Magdolna; Géczi, Lajos; Kahán, Zsuzsanna; Mangel, László; Révész, János; Szűcs, Miklós

    2014-03-01

    Everolimus is indicated for the therapy of adults with advanced renal cell carcinoma after failure of treatment with vascular endothelial growth factor receptor (VEGFR) tyrosine kinase inhibitor (TKI). The aim of the study was a multicenter evaluation of efficiency and toxicity of everolimus in patients with metastatic renal carcinoma who received one line of VEGFR-TKI therapy. Data of one hundred and one patients were analyzed retrospectively. Patients received everolimus therapy between January 2010 and July 2013. Data were collected in 7 different oncology institutes in Hungary. Starting daily dose of everolimus was 10 mg in 28-day cycles. Physical and laboratory examinations were done monthly. Imaging tests were performed every 3 months. Tumor response and toxicity were evaluated according to RECIST 1.0 and NCI CTCAE 3.0, respectively. Statistical analysis was performed with SPPS version 20.0 for Windows. Currently 26 (27%) patients are being treated, 52 (54.1%) patients are alive. Median progression-free survival (PFS) was 5.7 months (95% CI 4.07-7.33). Partial remission, stable disease and progression occurred in 6 (6%), 71 (74%) and 19 (20%) patients, respectively. Median overall survival (OS) was 14.3 months (95% CI 6.99-19.81). PFS and OS results were more favorable in patients with ECOG 0-1. Survival was poorer in case of anemia, while better if PFS was longer than 12 months. In anemic patients with ECOG 0-1 and ECOG 2-3 OS was 30.9 and 7.7 months, respectively (p=0.031). Dose reduction and treatment delay happened in 8 (7.9%) and 12 (11.9%) cases, respectively. The most common side effects were the following: exanthema, edema, stomatitis, pneumonitis, anemia and abnormal kidney-, liver functions, blood sugar and cholesterol levels. According to the Hungarian experience, everolimus can safely be administered. PFS and OS results representing the centers' everyday practice, are similar to the results of the respective subgroups in the registration study.

  7. mTOR is a Promising Therapeutic Target Both in Cisplatin-Sensitive and Cisplatin-Resistant Clear Cell Carcinoma of the Ovary

    PubMed Central

    Mabuchi, Seiji; Kawase, Chiaki; Altomare, Deborah A.; Morishige, Kenichirou; Sawada, Kenjiro; Hayashi, Masami; Tsujimoto, Masahiko; Yamoto, Mareo; Klein-Szanto, Andres J.; Schilder, Russell J.; Ohmichi, Masahide; Testa, Joseph R.; Kimura, Tadashi

    2009-01-01

    Translational Relevance Clear cell carcinoma (CCC) of the ovary is a distinctive subtype of epithelial ovarian cancer associated with a poorer sensitivity to platinum-based chemotherapy and a worse prognosis than the more common serous adenocarcinoma (SAC). To improve survival, the development of new treatment strategies that target CCC more effectively is necessary. Our results show that mTOR is more frequently activated in CCCs than in SACs. Our data have relevance for the design of future clinical studies of first-line treatment for patients with CCC of the ovary. Moreover, the finding of increased expression of phospho-mTOR and greater sensitivity to RAD001 in cisplatin-resistant CCC cells than in cisplatin-sensitive cells suggests a novel treatment option for patients with recurrent disease after cisplatin-based first-line chemotherapy. Purpose mTOR (mammalian target of rapamycin) plays a central role in cell proliferation and is regarded as a promising target in cancer therapy including for ovarian cancer. This study aims to examine the role of mTOR as a therapeutic target in clear cell carcinoma (CCC) of the ovary which is regarded as aggressive, chemo-resistant histological subtype. Experimental Design Using tissue microarrays of 98 primary ovarian cancers (52 clear cell carcinomas and 46 serous adenocarcinomas), the expression of phospho-mTOR was assessed by immunohistochemistry. Then, the growth-inhibitory effect of mTOR inhibition by RAD001 (everolimus) was examined using 2 pairs of cisplatin-sensitive parental (RMG1 and KOC7C) and cisplatin-resistant human CCC cell lines (RMG1-CR and KOC7C-CR) both in vitro and in vivo. Results Immunohistochemical analysis demonstrated mTOR was more frequently activated in CCCs than in serous adenocarcinomas (86.6% vs 50%). Treatment with RAD001 markedly inhibited the growth of both RMG1 and KOC7C cells both in vitro and in vivo. Increased expression of phospho-mTOR was observed in cisplatin-resistant RMG1-CR and KOC7C

  8. Tuberin, p27 and mTOR in different cells.

    PubMed

    Burgstaller, S; Rosner, M; Lindengrün, C; Hanneder, M; Siegel, N; Valli, A; Fuchs, C; Hengstschläger, M

    2009-02-01

    Mutations in the genes TSC1 or TSC2 cause the autosomal dominantly inherited tumor suppressor syndrome tuberous sclerosis, which is characterized by the development of tumors, named hamartomas, in different organs. The TSC gene products, hamartin and tuberin, form a complex, of which tuberin is assumed to be the functional component. Both, hamartin and tuberin have been implicated in the control of the cell cycle by activating the cyclin-dependent kinase inhibitor p27 and in cell size regulation by inhibiting the mammalian target of rapamycin (mTOR) a regulator of the p70 ribosomal protein S6 kinase (p70S6K) and its target the ribosomal protein S6. The tuberin/hamartin complex was shown to protect p27 from protein degradation. Within the mTOR signaling pathway tuberin harbors GTPase activating (GAP) potential toward Rheb, which is a potent regulator of mTOR. In this study, we have analyzed the protein levels of tuberin, p27, cyclin D1, mTOR and phospho mTOR Ser2448 (activated mTOR), S6 and phospho S6 Ser240/244 (activated S6) and as controls alpha-tubulin and topoisomerase IIbeta, in ten different cells, including primary normal cells, immortalized and transformed cell lines.

  9. mTOR complex 2 signaling and functions.

    PubMed

    Oh, Won Jun; Jacinto, Estela

    2011-07-15

    The mechanistic target of rapamycin (mTOR) plays a central role in cellular growth and metabolism. mTOR forms two distinct protein complexes, mTORC1 and mTORC2. Much is known about the regulation and functions of mTORC1 due to availability of a natural compound, rapamycin, that inhibits this complex. Studies that define mTORC2 cellular functions and signaling have lagged behind. The development of pharmacological inhibitors that block mTOR kinase activity, and thereby inhibit both mTOR complexes, along with availability of mice with genetic knockouts in mTOR complex components have now provided new insights on mTORC2 function and regulation. Since prolonged effects of rapamycin can also disrupt mTORC2, it is worth re-evaluating the contribution of this less-studied mTOR complex in cancer, metabolic disorders and aging. In this review, we focus on recent developments on mammalian mTORC2 signaling mechanisms and its cellular and tissue-specific functions.

  10. Prevention

    MedlinePlus

    ... our e-newsletter! Aging & Health A to Z Prevention Basic Facts & Information Some factors that affect your ... control of the things that you can change. Preventive Recommendations for Adults Aged 65 and Older The ...

  11. Prevention

    MedlinePlus

    ... Is Strong Error processing SSI file About Heart Disease & Stroke Prevention Heart disease and stroke are an epidemic in ... to avoid secondhand smoke. Barriers to Effective Heart Disease & Stroke Prevention Many people with key risk factors for heart ...

  12. Efficacy and Safety of Everolimus for Maintenance Immunosuppression of Kidney Transplantation: A Meta-Analysis of Randomized Controlled Trials

    PubMed Central

    Liu, Jinyu; Liu, Dong; Li, Juan; Zhu, Lan; Zhang, Chengliang; Lei, Kai; Xu, Qiling; You, Ruxu

    2017-01-01

    Background Conversion to everolimus is often used in kidney transplantation to overcome calcineurin inhibitor (CNI) nephrotoxicity but there is conflicting evidence for this approach. Objectives To investigate the benefits and harm from randomized clinical trials (RCTs) involving the conversion from CNI to everolimus after kidney transplantation. Methods Databases were searched up to March 2016. Two reviewers independently assessed trials for eligibility and quality, and extracted data. Results are expressed as risk ratio (RR) or mean difference (MD) with 95% confidence intervals (CI). Results Eleven RCTs, with a total of 1,633 patients, met the final inclusion criteria. Patients converted to everolimus had improved renal function at 1 year posttransplant with an estimated glomerular filtration rate (eGFR) of 5.36 mL/min per 1.73 m2 greater than patients remaining on CNI (p = 0.0005) and the longer-term results (> 1 year) of renal function was identical to that of 1 year. There was not a substantial difference in graft loss, mortality, and the occurrence of adverse events (AEs) or serious AEs. However, the risks of acute rejection and trial termination due to AEs with everolimus are respectively 1.82 and 2.63 times greater than patients staying on CNI at 1 year posttransplant (p = 0.02, p = 0.03, respectively). Further, those patients who converted to everolimus had a substantially greater risk of anemia, hyperlipidemia, hypercholesterolemia, hypokalemia, proteinuria, stomatitis, mouth ulceration, and acne. Conclusions Conversion from CNI to everolimus after kidney transplantation is associated with improved renal function in the first 5 years posttransplant but increases the risk of acute rejection at 1 year posttransplant and may not be well endured. PMID:28107397

  13. Platelet reactivity over time in coronary artery disease patients treated with a bioabsorbable everolimus-eluting scaffold.

    PubMed

    Tello-Montoliu, Antonio; Rivera, José; Hernández-Romero, Diana; Silvente, Ana; Jover, Eva; Quintana, Miriam; Orenes-Piñero, Esteban; Hurtado, José; Ferreiro, José Luis; Marín, Francisco; Valdés, Mariano

    2016-12-01

    Everolimus-eluting bioabsorbable scaffolds (BVSs) have exhibited similar long-term clinical outcomes compared to its everolimus-eluting metallic counterparts. However, reports from earlier studies have shown a signal for an increased rate of stent thrombosis. The aim of the current investigation is to describe the platelet reactivity profiles over time in patients treated with everolimus-eluting BVS in comparison to everolimus-eluting metallic stents. This is a pilot study in which patients on aspirin and clopidogrel with at least 1 everolimus-eluting BVS were included (n = 24). Patients with at least 1 everolimus-eluting metallic stent implanted were included as control group (n = 25). Blood samples were taken at time of discharge and at 3- and 6-month follow-up. Platelet function tests included VerifyNow (VN-P2Y12), multiplate aggregometry (MEA), and light transmission aggregometry (LTA). There was no difference in platelet reactivity at discharge, 3- and 6-month visits (unadjusted p = 0.733 and p = 0.582; p = 0.432 and p = 0.899 after adjusting for discharge value platelet reactivity0, respectively) using VN-P2Y12. Similar findings were observed with LTA. However, patients with BVS showed significantly higher platelet reactivity than patients with metallic stents at 3 and 6 months in the crude analysis (p = 0.003) and after adjusting for discharge value (p = 0.013) measured with ADP-MEA. There were no differences in platelet reactivity mediated by the T × A2 pathway between both groups. Finally, there is no statistical difference in high on-clopidogrel platelet reactivity (HPR) rate between both groups. The results of this pilot study suggest that BVS might have different platelet reactivity profiles, and warrants further investigation in dedicated clinical studies.

  14. Evodiamine Inhibits Insulin-Stimulated mTOR-S6K Activation and IRS1 Serine Phosphorylation in Adipocytes and Improves Glucose Tolerance in Obese/Diabetic Mice

    PubMed Central

    Wang, Ting; Kusudo, Tatsuya; Takeuchi, Tamaki; Yamashita, Yukari; Kontani, Yasuhide; Okamatsu, Yuko; Saito, Masayuki; Mori, Nozomu; Yamashita, Hitoshi

    2013-01-01

    Evodiamine, an alkaloid extracted from the dried unripe fruit of the tree Evodia rutaecarpa Bentham (Rutaceae), reduces obesity and insulin resistance in obese/diabetic mice; however, the mechanism underlying the effect of evodiamine on insulin resistance is unknown. This study investigated the effect of evodiamine on signal transduction relating to insulin resistance using obese/diabetic KK-Ay mice and an in vitro adipocyte culture. There is a significant decrease in the mammalian target of rapamycin (mTOR) and ribosomal S6 protein kinase (S6K) signaling in white adipose tissue (WAT) in KK-Ay mice treated with evodiamine, in which glucose tolerance is improved. In addition, reduction of insulin receptor substrate 1 (IRS1) serine phosphorylation, an indicator of insulin resistance, was detected in their WAT, suggesting suppression of the negative feedback loop from S6K to IRS1. As well as the stimulation of IRS1 and Akt serine phosphorylation, insulin-stimulated phosphorylation of mTOR and S6K is time-dependent in 3T3-L1 adipocytes, whereas evodiamine does not affect their phosphorylation except for an inhibitory effect on mTOR phosphorylation. Moreover, evodiamine inhibits the insulin-stimulated phosphorylation of mTOR and S6K, leading to down-regulation of IRS1 serine phosphorylation in the adipocytes. Evodiamine also stimulates phosphorylation of AMP-activated protein kinase (AMPK), an important regulator of energy metabolism, which may cause down-regulation of mTOR signaling in adipocytes. A similar effect on AMPK, mTOR and IRS1 phosphorylation was found in adipocytes treated with rosiglitazone. These results suggest evodiamine improves glucose tolerance and prevents the progress of insulin resistance associated with obese/diabetic states, at least in part, through inhibition of mTOR-S6K signaling and IRS1 serine phosphorylation in adipocytes. PMID:24391749

  15. mTOR Overactivation in Mesenchymal cells Aggravates CCl4− Induced liver Fibrosis

    PubMed Central

    Shan, Lanlan; Ding, Yan; Fu, You; Zhou, Ling; Dong, Xiaoying; Chen, Shunzhi; Wu, Hongyuan; Nai, Wenqing; Zheng, Hang; Xu, Wanfu; Bai, Xiaochun; Jia, Chunhong; Dai, Meng

    2016-01-01

    Hepatic stellate cells are of mesenchymal cell type located in the space of Disse. Upon liver injury, HSCs transactivate into myofibroblasts with increase in expression of fibrillar collagen, especially collagen I and III, leading to liver fibrosis. Previous studies have shown mTOR signaling is activated during liver fibrosis. However, there is no direct evidence in vivo. The aim of this study is to examine the effects of conditional deletion of TSC1 in mesenchymal on pathogenesis of liver fibrosis. Crossing mice bearing the floxed TSC1 gene with mice harboring Col1α2-Cre-ER(T) successfully generated progeny with a conditional knockout of TSC1 (TSC1 CKO) in collagen I expressing mesenchymal cells. TSC1 CKO and WT mice were subjected to CCl4, oil or CCl4+ rapamycin treatment for 8 weeks. TSC1 CKO mice developed pronounced liver fibrosis relative to WT mice, as examined by ALT, hydroxyproline, histopathology, and profibrogenic gene. Absence of TSC1 in mesenchymal cells induced proliferation and prevented apoptosis in activated HSCs. However, there were no significant differences in oil-treated TSC1 CKO and WT mice. Rapamycin, restored these phenotypic changes by preventing myofibroblasts proliferation and enhancing their apoptosis. These findings revealed mTOR overactivation in mesenchymal cells aggravates CCl4− induced liver fibrosis and the rapamycin prevent its occurance. PMID:27819329

  16. Effects of Salidroside on cobalt chloride-induced hypoxia damage and mTOR signaling repression in PC12 cells.

    PubMed

    Zhong, Xiaoyong; Lin, Ruhui; Li, Zuanfang; Mao, Jingjie; Chen, Lidian

    2014-01-01

    Salidroside (SA), a phenylpropanoid glycoside isolated from Rhodiola rosea L., has been documented to exert a broad spectrum of pharmacological properties, including protective effects against neuronal death induced by various stresses. To provide further insights into the neuroprotective functions of SA, this study examined whether SA can attenuate cobalt chloride (CoCl2)-induced hypoxia damage and mammalian target of rapamycin (mTOR) signaling repression in PC12 differentiated cells. Differentiated PC12 cells were exposed to CoCl2 for 12 h to mimic hypoxic/ischemic conditions and treated with SA at the same time, followed by electron microscopy and analysis of cell viability, intracellular reactive oxygen species (ROS) level, hypoxia-inducible factor-1α (HIF-1α) level, and the regulated in development and DNA damage responses (REDD1)/mTOR/ p70 ribosomal S6 kinase (p70S6K) signaling pathway. Our data indicated that SA can dramatically attenuate the ultrastructural damage of mitochondria induced by CoCl2 and significantly decrease CoCl2-induced ROS production. Moreover, phosphorylated mammalian target of rapamycin (p-mTOR) was significantly reduced by CoCl2, and this inhibition was relieved by the treatment of SA in PC12 cells, as evidenced by immunoblot and quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analyses. The SA effects were blocked by pretreatment of RAD001. The results indicate that SA can rescue CoCl2-induced repression of REDD1/mTOR/ p70S6K signal transduction in PC12 cells. Our data demonstrate that SA is able to attenuate CoCl2-induced hypoxia damage and mTOR signaling repression, suggesting that SA may protect brain neurons from ischemic injury through mTOR signaling, and provide new insights into the prevention and treatment of cerebral ischemic.

  17. mTOR inhibitor for the treatment of hepatocellular carcinoma.

    PubMed

    Kudo, Masatoshi

    2011-01-01

    Mammalian target of rapamycin (mTOR) plays a central role in the regulation of cellular growth, proliferation, and survival via a cytoplasmic serine/threonine kinase. mTOR also works as a nutrition sensor to monitor cellular metabolism. mTOR is located downstream in the PI3K/Akt pathway, in which Akt and the tuberous sclerosis complex (TSC) 1/2 are involved, to form a signal transduction pathway. New anticancer agents that target mTOR in the PI3K/Akt pathway of the signal transduction pathways involved in cell proliferation control have recently been developed and are already commercially available. A phase III clinical trial of mTOR inhibitor for hepatocellular carcinoma (HCC) is now ongoing worldwide to expand indications. RAD001 is a signal-transduction inhibitor (STI) that targets mTOR (more specifically, mTORC1). mTORC1 signaling is intricately regulated by mitogens, growth factors, energy, and nutrients. mTORC1 is a regulator essential for general protein synthesis, located downstream of the PI3K/AKT/mTOR pathway, which is dysregulated in most human cancers. Inhibiting mTOR with molecules, such as RAD001, generates additive effects that accompany upstream and downstream target inhibition; alternatively, upstream receptor inhibition is compensated for by inhibiting the downstream pathway, even if some resistance develops against receptor inhibition regardless of initial or acquired resistance. In conclusion, RAD001 is a potential targeted agent for HCC and therefore final results of a phase III study are awaited.

  18. The mTOR Complex Controls HIV Latency.

    PubMed

    Besnard, Emilie; Hakre, Shweta; Kampmann, Martin; Lim, Hyung W; Hosmane, Nina N; Martin, Alyssa; Bassik, Michael C; Verschueren, Erik; Battivelli, Emilie; Chan, Jonathan; Svensson, J Peter; Gramatica, Andrea; Conrad, Ryan J; Ott, Melanie; Greene, Warner C; Krogan, Nevan J; Siliciano, Robert F; Weissman, Jonathan S; Verdin, Eric

    2016-12-14

    A population of CD4 T lymphocytes harboring latent HIV genomes can persist in patients on antiretroviral therapy, posing a barrier to HIV eradication. To examine cellular complexes controlling HIV latency, we conducted a genome-wide screen with a pooled ultracomplex shRNA library and in vitro system modeling HIV latency and identified the mTOR complex as a modulator of HIV latency. Knockdown of mTOR complex subunits or pharmacological inhibition of mTOR activity suppresses reversal of latency in various HIV-1 latency models and HIV-infected patient cells. mTOR inhibitors suppress HIV transcription both through the viral transactivator Tat and via Tat-independent mechanisms. This inhibition occurs at least in part via blocking the phosphorylation of CDK9, a p-TEFb complex member that serves as a cofactor for Tat-mediated transcription. The control of HIV latency by mTOR signaling identifies a pathway that may have significant therapeutic opportunities.

  19. Regulation on Beclin-1 expression by mTOR in CoCl2-induced HT22 cell ischemia-reperfusion injury.

    PubMed

    Yang, Tao; Li, Dongbo; Liu, Feng; Qi, Lei; Yan, Ge; Wang, Maode

    2015-07-21

    It has been reported that cerebral ischemia/reperfusion (I/R) injury can activate autophagy. However, the role of autophagy in cerebral I/R injury remains controversy. Two major proteins, mTOR and Beclin-1, govern the formation of autophagosomes to regulate autophagy activity. However, the cross-talking between Beclin-1 and mTOR in cerebral I/R injury remains elusive. In this study, global cerebral I/R injury animal model and focal cerebral I/R injury animal model were induced to test the variation of Beclin-1 level in vivo. To further confirm the variation of Beclin-1 level and investigate the cross-talking between Beclin-1 and mammalian target of rapamycin (mTOR) in I/R injury, we used cobalt chloride (CoCl2) to develop an I/R injury cell model in HT22 cell line. Our data showed that the levels of Beclin-1 and phosphorylated mammalian target of rapamycin (p-mTOR) were clearly induced by I/R injury in vitro. And the time course studies suggested that the Beclin-1 and mTOR may have coordinated regulation in ischemia stages but not in reperfusion stages. Moreover, inhibitor of mTOR could prevent Beclin-1 decreasing, but this prevention may play opposite roles in different stages of I/R injury. We conclude that this study represents a major advance in our understanding of the cross-talking of two key proteins, Beclin-1 and mTOR, in autophagy and the role of autophagy in cerebral I/R injury.

  20. mTOR pathway activation is a favorable prognostic factor in human prostate adenocarcinoma

    PubMed Central

    Stelloo, Suzan; Sanders, Joyce; Nevedomskaya, Ekaterina; de Jong, Jeroen; Peters, Dennis; van Leenders, Geert J.L.H.; Jenster, Guido; Bergman, Andries M.; Zwart, Wilbert

    2016-01-01

    Prostate cancer patients with localized disease are treated with curative intent. However, the disease will recur in approximately 30% of patients with a high incidence of morbidity and mortality. Prognostic biomarkers are needed to identify patients with high risk of relapse. mTOR pathway activation is reported in prostate cancer, but clinical trials testing efficacy of mTOR inhibitors were unsuccessful. To explain this clinical observation, we studied the expression and prognostic impact of mTOR-S2448 phosphorylation in localized prostate carcinomas. mTOR-S2448 phosphorylation is indicative for an activated mTOR pathway in prostate cancer. Surprisingly, the mTOR signaling pathway is activated specifically in prostate cancer patients with a favorable outcome. Since tumors from poor-outcome patients have low levels of mTOR-S2448 phosphorylation, this may explain why mTOR inhibitors proved unsuccessful in prostate cancer trials. PMID:27096957

  1. Risk of fatigue in patients with solid tumors treated with everolimus, temsirolimus or ridaforolimus: a comparative meta-analysis.

    PubMed

    Abdel-Rahman, Omar; Fouad, Mona

    2015-04-01

    We performed a meta-analysis of fatigue associated with the use of everolimus, temsirolimus or ridaforolimus in patients with solid tumors. Eligible studies included randomized trials of patients with solid tumors on everolimus, temsirolimus or ridaforolimus describing events of fatigue. A total of 18 clinical trials including 8143 patients were considered eligible for the meta-analysis. On the basis of random-effects model, we found that the relative risk of all-grade and high-grade fatigue were 1.26 [95% CI: 1.09-1.46; p < 0.0001], 1.49 [95% CI: 0.99, 2.24; p = 0.05], respectively. On subgroup analysis, we cannot identify any difference between everolimus and temsirolimus in the risk of fatigue. Thus, our meta-analysis has demonstrated that regimens containing everolimus, temsirolimus or ridaforolimus for the treatment of solid tumors are associated with an increased risk of all-grade fatigue, whereas the risk of high-grade fatigue did not reach the threshold of statistical significance. Close clinical monitoring and pre-emptive treatment for fatigue are recommended.

  2. Development and Validation of Stability-indicating High Performance Liquid Chromatographic Method for the Estimation of Everolimus in Tablets

    PubMed Central

    Sharmila, D.; Rao, A. Lakshmana; Kalyani, L.

    2015-01-01

    The present study depicts the development of a validated reversed-phase high performance liquid chromatographic method for the determination of the everolimus in presence of degradation products or pharmaceutical excipients. Stress study was performed on everolimus and it was found that it degrade sufficiently in oxidizing and acidic conditions but less degradation was found in alkaline, neutral, thermal and photolytic conditions. The separation was carried out on Hypersil BDS C18 column (100×4.6 mm, 5 μ) column having particle size 5 μ using acetate buffer:acetonitrile (50:50 v/v) with pH 6.5 adjusted with orthophosphoric acid as mobile phase at flow rate of 1 ml/min. The wavelength of the detection was 280 nm. A retention time (Rt) nearly 3.110 min was observed. The calibration curve for everolimus was linear (r2=0.999) from range of 25-150 μg/ml with limit of detection and limit of quantification of 0.036 μg/ml and 0.109 μg/ml, respectively. Analytical validation parameters such as selectivity, specificity, linearity, accuracy and precision were evaluated and relative standard deviation value for all the key parameters were less than 2.0%. The recovery of the drug after standard addition was found to be 100.55%. Thus, the developed RP-HPLC method was found to be suitable for the determination of everolimus in tablets containing various excipients. PMID:26798176

  3. Ultra fast liquid chromatography-tandem mass spectrometry routine method for simultaneous determination of cyclosporin A, tacrolimus, sirolimus, and everolimus in whole blood using deuterated internal standards for cyclosporin A and everolimus.

    PubMed

    Meinitzer, Andreas; Gartner, Gabriele; Pilz, Stefan; Stettin, Mariana

    2010-02-01

    Specific chromatographic methods for the measurement of cyclosporin A, tacrolimus, sirolimus, and everolimus blood levels in patients with organ transplants are time consuming when large numbers of samples must be processed. The authors developed a robust and fast (1 minute) online solid-phase extraction liquid chromatography/tandem mass spectrometry method for the simultaneous quantification of cyclosporin A, tacrolimus, sirolimus, and everolimus. After protein precipitation of the whole blood with zinc sulphate and methanol, the supernatant was loaded on a wide pore reversed-phase column and cleansed of potential interferences with high flow for 20 seconds. After column switching, the analytes were transferred within 20 seconds in the back-flush mode to a short phenyl-hexyl column. The valve was then returned to its initial position and the chromatographic separation performed within 20 seconds. In the meantime, the loading column was prepared for the next injection. Ammoniated adducts of protonated molecules were used as precursor ions for all analytes. Multiple-reaction mode transitions for each immunosuppressant and the internal standards were used for quantification. The working range of the method was 10-1500 microg/L for cyclosporin A, 1.0-44 microg/L for tacrolimus, 1.0-48 microg/L for sirolimus, and 1.2-48 microg/L for everolimus. Within and between-run assay coefficients of variation ranged from 1.8% to 13.0%. The described liquid chromatography/tandem mass spectrometry method shows best performance using the internal standards cyclosporin A-d4 for cyclosporin A, everolimus-d4 for everolimus and ascomycin for tacrolimus and sirolimus. In conclusion, the authors present a very fast, robust, and economical analytical method for therapeutic monitoring of multiple immunosuppressants in daily clinical practice.

  4. Targeting mTOR in Pancreatic Ductal Adenocarcinoma

    PubMed Central

    Iriana, Sentia; Ahmed, Shahzad; Gong, Jun; Annamalai, Alagappan Anand; Tuli, Richard; Hendifar, Andrew Eugene

    2016-01-01

    Treatment options for advanced pancreatic ductal adenocarcinoma (PDAC) are limited; however, new therapies targeting specific tumor-related molecular characteristics may help certain patient cohorts. Emerging preclinical data have shown that inhibition of mammalian target of rapamycin (mTOR) in specific KRAS-dependent PDAC subtypes leads to inhibition of tumorigenesis in vitro and in vivo. Early phase II studies of mono-mTOR inhibition have not shown promise. However, studies have shown that combined inhibition of multiple steps along the mTOR signaling pathway may lead to sustained responses by targeting mechanisms of tumor resistance. Coordinated inhibition of mTOR along with specific KRAS-dependent mutations in molecularly defined PDAC subpopulations may offer a viable alternative for treatment in the future. PMID:27200288

  5. Safety, Efficacy, and Patient Acceptability of Everolimus in the Treatment of Breast Cancer

    PubMed Central

    Lousberg, Laurence; Jerusalem, Guy

    2016-01-01

    Everolimus combined with exemestane is an important treatment option for patients suffering from estrogen receptor-positive, human epidermal growth factor receptor 2-negative, advanced breast cancer (ABC) who have been previously treated with a nonsteroidal aromatase inhibitor (NSAI). After presentation of phase III registration trial BOLERO-2, several phase IIIb trials have been started to evaluate this regimen in a more real-world setting. Here, we review the efficacy and safety data published or presented at selected international meetings. These studies confirmed the outcome observed in the BOLERO-2 trial. Patient acceptance rate is also discussed by focusing on the permanent everolimus discontinuation rate in these trials. Factors influencing the safety profile are also reported, including the impact of age. The optimal sequence of combined therapy approaches associating targeted and endocrine therapy (ET) has yet to be determined as new treatment options such as cyclin-dependent kinase inhibitors become available. However, everolimus–exemestane remains an important treatment option with a major impact on progression-free survival (PFS) and an acceptable safety profile. PMID:28096680

  6. Use of Everolimus in Liver Transplantation: Recommendations From a Working Group

    PubMed Central

    De Simone, Paolo; Fagiuoli, Stefano; Cescon, Matteo; De Carlis, Luciano; Tisone, Giuseppe; Volpes, Riccardo; Cillo, Umberto

    2017-01-01

    Abstract Immunosuppression after liver transplantation (LT) is presently based on use of calcineurin inhibitors (CNI), although they are associated with an increased incidence of renal dysfunction, cardiovascular complications, and de novo and recurrent malignancies. Over the past decade, mammalian target of rapamycin inhibitors have received considerable attention as immunosuppressants because they are associated with a more favorable renal profile versus CNI, as well as antiproliferative activity in clinical studies. Comprehensive guidelines on use of everolimus (EVR) in LT are still lacking. In Italy, a project, named Everolimus: the road to long-term functioning, was initiated to collect the experience on EVR after LT with the aim of providing guidance for transplant clinicians. Herein, recommendations by this national consensus group, based on Delphi methodology, are presented. Consensus was reached on 20 of the 23 statements proposed, and their level of evidence, grade of recommendation, and percent of agreement are reported. Statements are grouped into 4 areas: (A) renal function; (B) time of EVR introduction, CNI reduction and elimination, and risk for graft rejection; (C) antiproliferative effects of EVR; and (D) management of EVR-related adverse events. The high level of consensus shows that there is good agreement on the routine use of EVR in predefined clinical scenarios, especially in light of posttransplant nephrotoxicity and other adverse events associated with long-term administration of CNIs. PMID:27495768

  7. Prevention

    MedlinePlus

    ... Ban For Clinicians Clinical Recognition Specimen Collection Treatment Smallpox Vaccine Guidance Infection Control: Hospital Infection Control: Home ... Mouth Infection) Poxvirus and Rabies Branch Travelers’ Health: Smallpox & Other Orthopoxvirus-Associated Infections Poxvirus Prevention Recommend on ...

  8. A phase I/II trial of BNC105P with everolimus in metastatic renal cell carcinoma (mRCC)

    PubMed Central

    Pal, Sumanta; Azad, Arun; Bhatia, Shailender; Drabkin, Harry; Costello, Brian; Sarantopoulos, John; Kanesvaran, Ravindran; Lauer, Richard; Starodub, Alexander; Hauke, Ralph; Sweeney, Christopher J.; Hahn, Noah M.; Sonpavde, Guru; Richey, Stephen; Breen, Timothy; Kremmidiotis, Gabriel; Leske, Annabell; Doolin, Elizabeth; Bibby, David C.; Simpson, Jeremy; Iglesias, Jose; Hutson, Thomas

    2015-01-01

    Purpose BNC105P inhibits tubulin polymerization, and preclinical studies suggest possible synergy with everolimus. In this phase I/II study, efficacy and safety of the combination were explored in patients with metastatic renal cell carcinoma (mRCC). Experimental Design A phase I study in patients with clear cell mRCC and any prior number of therapies was conducted using a classical 3+3 design to evaluate standard doses of everolimus with increasing doses of BNC105P. At the recommended phase II dose (RP2D), patients with clear cell mRCC and 1-2 prior therapies (including ≥1 VEGF-TKI) were randomized to BNC105P with everolimus (Arm A) or everolimus alone (Arm B). The primary endpoint of the study was 6-month progression-free survival (6MPFS). Secondary endpoints included response rate, PFS, overall survival (OS) and exploratory biomarker analyses. Results In the phase I study (n=15), a dose of BNC105P at 16 mg/m2 with everolimus at 10 mg daily was identified as the RP2D. In the phase II study, 139 patients were randomized, with 69 and 67 evaluable patients in Arms A and B, respectively. 6MPFS was similar in the treatment arms (Arm A: 33.82% v Arm B: 30.30%, P=0.66) and no difference in median PFS was observed (Arm A: 4.7 mos v Arm B: 4.1 mos; P=0.49). Changes in matrix metalloproteinase-9, stem cell factor, sex hormone binding globulin and serum amyloid A protein were associated with clinical outcome with BNC105P. Conclusions Although the primary endpoint was not met in an unselected population, correlative studies suggest several biomarkers that warrant further prospective evaluation. PMID:25788492

  9. A Recollection of mTOR Signaling in Learning and Memory

    ERIC Educational Resources Information Center

    Graber, Tyson E.; McCamphill, Patrick K.; Sossin, Wayne S.

    2013-01-01

    Mechanistic target of rapamcyin (mTOR) is a central player in cell growth throughout the organism. However, mTOR takes on an additional, more specialized role in the developed neuron, where it regulates the protein synthesis-dependent, plastic changes underlying learning and memory. mTOR is sequestered in two multiprotein complexes (mTORC1 and…

  10. Activation of mTOR signaling leads to orthopedic surgery-induced cognitive decline in mice through β-amyloid accumulation and tau phosphorylation.

    PubMed

    Shen, Wenzhen; Lu, Keliang; Wang, Jiawan; Wu, Anshi; Yue, Yun

    2016-10-01

    Postoperative cognitive dysfunction (POCD) is a serious complication following surgery, however, the mechanism of POCD remains to be elucidated. Previous evidence has revealed that POCD may be associated with the pathogenesis of neurodegenerative processes. The mammalian target of rapamycin (mTOR) signaling pathway has been reported to be crucial in the pathophysiology of neurodegenerative diseases. However, the implications of mTOR in POCD remains to be fully elucidated. In the present study, western blotting and enzyme‑linked immunosorbent assay were used to determine the expression of mTOR and any associated downstream targets; contextual fear conditioning was used to estimate the learning and memory ability of mice. Using an animal model of orthopedic surgery, it was found that surgical injury impaired hippocampal‑dependent memory and enhanced the levels of phosphorylated mTOR at Serine‑2448, phosphorylated 70‑kDa ribosomal protein S6 kinase (p70S6K) at Threonine‑389 with accumulation of β‑amyloid (Aβ) and hyperphosphorylated tau at Serine-396, compared with the control group. Pretreatment with rapamycin, an mTOR inhibitor, restored the abnormal mTOR/p70S6K signaling induced by surgery, attenuated the accumulation of Aβ and reduced the phosphorylation of tau protein. Rapamycin also reversed the surgery‑induced cognitive dysfunction. The results of the present study suggested that the surgical stimulus activated mTOR/p70S6K signaling excessively, and that the inhibition of mTOR signaling with rapamycin may prevent postoperative cognitive deficits, partly through attenuating the accumulation of Aβ and hyperphosphorylation of tau protein.

  11. Ubiquitin hydrolase UCH-L1 destabilizes mTOR complex 1 by antagonizing DDB1-CUL4-mediated ubiquitination of raptor.

    PubMed

    Hussain, Sajjad; Feldman, Andrew L; Das, Chittaranjan; Ziesmer, Steven C; Ansell, Stephen M; Galardy, Paul J

    2013-03-01

    Mammalian target of rapamycin (mTOR) is a serine/threonine kinase that regulates processes including mRNA translation, proliferation, and survival. By assembling with different cofactors, mTOR forms two complexes with distinct biological functions. Raptor-bound mTOR (mTORC1) governs cap-dependent mRNA translation, whereas mTOR, rictor, and mSin1 (mTORC2) activate the survival and proliferative kinase Akt. How the balance between the competing needs for mTORC1 and -2 is controlled in normal cells and deregulated in disease is poorly understood. Here, we show that the ubiquitin hydrolase UCH-L1 regulates the balance of mTOR signaling by disrupting mTORC1. We find that UCH-L1 impairs mTORC1 activity toward S6 kinase and 4EBP1 while increasing mTORC2 activity toward Akt. These effects are directly attributable to a dramatic rearrangement in mTOR complex assembly. UCH-L1 disrupts a complex between the DDB1-CUL4 ubiquitin ligase complex and raptor and counteracts DDB1-CUL4-mediated raptor ubiquitination. These events lead to mTORC1 dissolution and a secondary increase in mTORC2. Experiments in Uchl1-deficient and transgenic mice suggest that the balance between these pathways is important for preventing neurodegeneration and the development of malignancy. These data establish UCH-L1 as a key regulator of the dichotomy between mTORC1 and mTORC2 signaling.

  12. Comparative effectiveness of everolimus and axitinib as second targeted therapies for metastatic renal cell carcinoma in the US: a retrospective chart review.

    PubMed

    Vogelzang, Nicholas J; Pal, Sumanta K; Signorovitch, James E; Reichmann, William M; Li, Nanxin; Yang, Chelsey; Liu, Zhimei; Perez, Jose Ricardo; Jonasch, Eric

    2016-01-01

    Background Second targeted therapies for metastatic renal cell carcinoma (mRCC) include mammalian target of rapamycin inhibitors (mTORis) and tyrosine kinase inhibitors (TKIs). This observational study compares overall survival (OS) and progression-free survival (PFS) of patients treated with everolimus (an mTORi) and axitinib (a TKI) following first TKI, and assesses the impact of type and duration of first TKI on the relative effectiveness of these second targeted therapies. Methods Retrospective reviews of medical records were conducted by medical oncologists or hematologists/oncologists recruited from a nationwide panel. Included patients with mRCC were required to have discontinued a first TKI (sunitinib, sorafenib, or pazopanib) for medical reasons, and to have initiated everolimus or axitinib as second targeted therapy between February 2012 and January 2013. OS and PFS were compared between patients treated with everolimus vs. axitinib using multivariable Cox proportional hazards regression models. Comparative results were also stratified by type and duration of first TKI. Results Included patients (n = 325 for everolimus and n = 127 for axitinib) had a mean age of 61 years and 31% were female. Sunitinib was the most commonly used first TKI (73%). After adjusting for patient characteristics, no statistically significant differences were observed in OS or PFS between everolimus and axitinib. When stratifying by type and duration of first TKI, there was no statistically significant difference in OS between everolimus and axitinib in all subgroups except for patients with <6 months on sunitinib or sorafenib as first TKI. No significant difference in PFS was observed in any subgroup. Limitations Important limitations include potential missing or inaccurate data in medical charts, and confounding due to unobserved factors. Conclusions In this retrospective chart review, no significant differences were detected in OS or PFS between axitinib and

  13. The DPYSL2 gene connects mTOR and schizophrenia

    PubMed Central

    Pham, X; Song, G; Lao, S; Goff, L; Zhu, H; Valle, D; Avramopoulos, D

    2016-01-01

    We previously reported a schizophrenia-associated polymorphic CT di-nucleotide repeat (DNR) at the 5′-untranslated repeat (UTR) of DPYSL2, which responds to mammalian target of Rapamycin (mTOR) signaling with allelic differences in reporter assays. Now using microarray analysis, we show that the DNR alleles interact differentially with specific proteins, including the mTOR-related protein HuD/ELAVL4. We confirm the differential binding to HuD and other known mTOR effectors by electrophoretic mobility shift assays. We edit HEK293 cells by CRISPR/Cas9 to carry the schizophrenia risk variant (13DNR) and observe a significant reduction of the corresponding CRMP2 isoform. These edited cells confirm the response to mTOR inhibitors and show a twofold shortening of the cellular projections. Transcriptome analysis of these modified cells by RNA-seq shows changes in 12.7% of expressed transcripts at a false discovery rate of 0.05. These transcripts are enriched in immunity-related genes, overlap significantly with those modified by the schizophrenia-associated gene, ZNF804A, and have a reverse expression signature from that seen with antipsychotic drugs. Our results support the functional importance of the DPYSL2 DNR and a role for mTOR signaling in schizophrenia. PMID:27801893

  14. The CECARI Study: Everolimus (Certican®) Initiation and Calcineurin Inhibitor Withdrawal in Maintenance Heart Transplant Recipients with Renal Insufficiency: A Multicenter, Randomized Trial

    PubMed Central

    Derthoo, David; Van Caenegem, Olivier; De Pauw, Michel; Nellessen, Eric; Duerinckx, Nathalie; Droogne, Walter; Vörös, Gábor; Meyns, Bart; Belmans, Ann; Janssens, Stefan; Vanhaecke, Johan

    2017-01-01

    In this 3-year, open-label, multicenter study, 57 maintenance heart transplant recipients (>1 year after transplant) with renal insufficiency (eGFR 30–60 mL/min/1.73 m2) were randomized to start everolimus with CNI withdrawal (N = 29) or continue their current CNI-based immunosuppression (N = 28). The primary endpoint, change in measured glomerular filtration rate (mGFR) from baseline to year 3, did not differ significantly between both groups (+7.0 mL/min in the everolimus group versus +1.9 mL/min in the CNI group, p = 0.18). In the on-treatment analysis, the difference did reach statistical significance (+9.4 mL/min in the everolimus group versus +1.9 mL/min in the CNI group, p = 0.047). The composite safety endpoint of all-cause mortality, major adverse cardiovascular events, or treated acute rejection was not different between groups. Nonfatal adverse events occurred in 96.6% of patients in the everolimus group and 57.1% in the CNI group (p < 0.001). Ten patients (34.5%) in the everolimus group discontinued the study drug during follow-up due to adverse events. The poor adherence to the everolimus therapy might have masked a potential benefit of CNI withdrawal on renal function. PMID:28316834

  15. Determination and validation of mTOR kinase-domain 3D structure by homology modeling.

    PubMed

    Lakhlili, Wiame; Chevé, Gwénaël; Yasri, Abdelaziz; Ibrahimi, Azeddine

    2015-01-01

    The AKT/mammalian target of rapamycin (mTOR) pathway is considered as one of the commonly activated and deregulated signaling pathways in human cancer. mTOR is associated with other proteins in two molecular complexes: mTOR complex 1/Raptor and the mTOR complex 2/Rictor. Using the crystal structure of the related lipid kinase PI3Kγ, we built a model of the catalytic region of mTOR. The modeling of the three-dimensional (3D) structure of the mTOR was performed by homology modeling program SWISS-MODEL. The quality and validation of the obtained model were performed using PROCHECK and PROVE softwares. The overall stereochemical property of the protein was assessed by the Ramachandran plot. The model validation was also done by docking of known inhibitors. In this paper, we describe and validate a 3D model for the mTOR catalytic site.

  16. mTOR is necessary for proper satellite cell activity and skeletal muscle regeneration

    SciTech Connect

    Zhang, Pengpeng; Liang, Xinrong; Shan, Tizhong; Jiang, Qinyang; Deng, Changyan; Zheng, Rong; Kuang, Shihuan

    2015-07-17

    The serine/threonine kinase mammalian target of rapamycin (mTOR) is a key regulator of protein synthesis, cell proliferation and energy metabolism. As constitutive deletion of Mtor gene results in embryonic lethality, the function of mTOR in muscle stem cells (satellite cells) and skeletal muscle regeneration remains to be determined. In this study, we established a satellite cell specific Mtor conditional knockout (cKO) mouse model by crossing Pax7{sup CreER} and Mtor{sup flox/flox} mice. Skeletal muscle regeneration after injury was severely compromised in the absence of Mtor, indicated by increased number of necrotic myofibers infiltrated by Evans blue dye, and reduced number and size of regenerated myofibers in the Mtor cKO mice compared to wild type (WT) littermates. To dissect the cellular mechanism, we analyzed satellite cell-derived primary myoblasts grown on single myofibers or adhered to culture plates. The Mtor cKO myoblasts exhibited defective proliferation and differentiation kinetics when compared to myoblasts derived from WT littermates. At the mRNA and protein levels, the Mtor cKO myoblasts expressed lower levels of key myogenic determinant genes Pax7, Myf5, Myod, Myog than did the WT myoblasts. These results suggest that mTOR is essential for satellite cell function and skeletal muscle regeneration through controlling the expression of myogenic genes. - Highlights: • Pax7{sup CreER} was used to delete Mtor gene in satellite cells. • Satellite cell specific deletion of Mtor impairs muscle regeneration. • mTOR is necessary for satellite cell proliferation and differentiation. • Deletion of Mtor leads to reduced expression of key myogenic genes.

  17. Everolimus-eluting stent platforms in percutaneous coronary intervention: comparative effectiveness and outcomes

    PubMed Central

    Panoulas, Vasileios F; Mastoris, Ioannis; Konstantinou, Klio; Tespili, Maurizio; Ielasi, Alfonso

    2015-01-01

    Despite the remarkable benefits obtained following the introduction of the first-generation drug-eluting stent (DES), concerns were raised over its long-term safety, particularly with regard to very late (beyond 1 year) stent thrombosis. Newer-generation DESs have been developed to overcome this limitation using novel stent platforms, new drugs, more biocompatible durable polymers, and bioabsorbable polymers or backbones. To date, new-generation DESs have virtually replaced the use of first-generation DESs worldwide. In this review article, we discuss in detail the design, pharmacology, and mechanism of action of the newer-generation permanent and bioresorbable everolimus-eluting platforms. Furthermore, we present and evaluate the current evidence on the performance and safety of these devices compared to those of other available stent platforms. PMID:26244031

  18. Dexamethasone and BCAA Failed to Modulate Muscle Mass and mTOR Signaling in GH-Deficient Rats

    PubMed Central

    Nishida, Hikaru; Ikegami, Ayaka; Kaneko, Chiaki; Kakuma, Hitomi; Nishi, Hisano; Tanaka, Noriko; Aoyama, Michiko; Usami, Makoto; Okimura, Yasuhiko

    2015-01-01

    Branched-chain amino acids (BCAAs) and IGF-I, the secretion of which is stimulated by growth hormone (GH), prevent muscle atrophy. mTOR plays a pivotal role in the protective actions of BCAA and IGF-1. The pathway by which BCAA activates mTOR is different from that of IGF-1, which suggests that BCAA and GH work independently. We tried to examine whether BCAA exerts a protective effect against dexamethasone (Dex)-induced muscle atrophy independently of GH using GH-deficient spontaneous dwarf rats (SDRs). Unexpectedly, Dex did not induce muscle atrophy assessed by the measurement of cross-sectional area (CSA) of the muscle fibers and did not increase atrogin-1, MuRF1 and REDD1 expressions, which are activated during protein degradation. Glucocorticoid (GR) mRNA levels were higher in SDRs compared to GH-treated SDRs, indicating that the low expression of GR is not the reason of the defect of Dex’s action in SDRs. BCAA did not stimulate the phosphorylation of p70S6K or 4E-BP1, which stimulate protein synthesis. BCAA did not decrease the mRNA level of atrogin-1 or MuRF1. These findings suggested that Dex failed to modulate muscle mass and that BCAA was unable to activate mTOR in SDRs because these phosphorylations of p70S6K and 4E-BP1 and the reductions of these mRNAs are regulated by mTOR. In contrast, after GH supplementation, these responses to Dex were normalized and muscle fiber CSA was decreased by Dex. BCAA prevented the Dex-induced decrease in CSA. BCAA increased the phosphorylation of p70S6K and decreased the Dex-induced elevations of atrogin-1 and Bnip3 mRNAs. However, the amount of mTORC1 components including mTOR was not decreased in the SDRs compared to the normal rats. These findings suggest that GH increases mTORC1 activity but not its content to recover the action of BCAA in SDRs and that GH is required for actions of Dex and BCAA in muscles. PMID:26086773

  19. Dexamethasone and BCAA Failed to Modulate Muscle Mass and mTOR Signaling in GH-Deficient Rats.

    PubMed

    Nishida, Hikaru; Ikegami, Ayaka; Kaneko, Chiaki; Kakuma, Hitomi; Nishi, Hisano; Tanaka, Noriko; Aoyama, Michiko; Usami, Makoto; Okimura, Yasuhiko

    2015-01-01

    Branched-chain amino acids (BCAAs) and IGF-I, the secretion of which is stimulated by growth hormone (GH), prevent muscle atrophy. mTOR plays a pivotal role in the protective actions of BCAA and IGF-1. The pathway by which BCAA activates mTOR is different from that of IGF-1, which suggests that BCAA and GH work independently. We tried to examine whether BCAA exerts a protective effect against dexamethasone (Dex)-induced muscle atrophy independently of GH using GH-deficient spontaneous dwarf rats (SDRs). Unexpectedly, Dex did not induce muscle atrophy assessed by the measurement of cross-sectional area (CSA) of the muscle fibers and did not increase atrogin-1, MuRF1 and REDD1 expressions, which are activated during protein degradation. Glucocorticoid (GR) mRNA levels were higher in SDRs compared to GH-treated SDRs, indicating that the low expression of GR is not the reason of the defect of Dex's action in SDRs. BCAA did not stimulate the phosphorylation of p70S6K or 4E-BP1, which stimulate protein synthesis. BCAA did not decrease the mRNA level of atrogin-1 or MuRF1. These findings suggested that Dex failed to modulate muscle mass and that BCAA was unable to activate mTOR in SDRs because these phosphorylations of p70S6K and 4E-BP1 and the reductions of these mRNAs are regulated by mTOR. In contrast, after GH supplementation, these responses to Dex were normalized and muscle fiber CSA was decreased by Dex. BCAA prevented the Dex-induced decrease in CSA. BCAA increased the phosphorylation of p70S6K and decreased the Dex-induced elevations of atrogin-1 and Bnip3 mRNAs. However, the amount of mTORC1 components including mTOR was not decreased in the SDRs compared to the normal rats. These findings suggest that GH increases mTORC1 activity but not its content to recover the action of BCAA in SDRs and that GH is required for actions of Dex and BCAA in muscles.

  20. Dual-Blocking of PI3K and mTOR Improves Chemotherapeutic Effects on SW620 Human Colorectal Cancer Stem Cells by Inducing Differentiation

    PubMed Central

    Kim, Buyun

    2016-01-01

    Cancer stem cells (CSCs) have tumor initiation, self-renewal, metastasis and chemo-resistance properties in various tumors including colorectal cancer. Targeting of CSCs may be essential to prevent relapse of tumors after chemotherapy. Phosphatidylinositol-3-kinase (PI3K) and mammalian target of rapamycin (mTOR) signals are central regulators of cell growth, proliferation, differentiation, and apoptosis. These pathways are related to colorectal tumorigenesis. This study focused on PI3K and mTOR pathways by inhibition which initiate differentiation of SW620 derived CSCs and investigated its effect on tumor progression. By using rapamycin, LY294002, and NVP-BEZ235, respectively, PI3K and mTOR signals were blocked independently or dually in colorectal CSCs. Colorectal CSCs gained their differentiation property and lost their stemness properties most significantly in dual-blocked CSCs. After treated with anti-cancer drug (paclitaxel) on the differentiated CSCs cell viability, self-renewal ability and differentiation status were analyzed. As a result dual-blocking group has most enhanced sensitivity for anti-cancer drug. Xenograft tumorigenesis assay by using immunodeficiency mice also shows that dual-inhibited group more effectively increased drug sensitivity and suppressed tumor growth compared to single-inhibited groups. Therefore it could have potent anti-cancer effects that dual-blocking of PI3K and mTOR induces differentiation and improves chemotherapeutic effects on SW620 human colorectal CSCs. PMID:26955235

  1. mTOR is necessary for proper satellite cell activity and skeletal muscle regeneration.

    PubMed

    Zhang, Pengpeng; Liang, Xinrong; Shan, Tizhong; Jiang, Qinyang; Deng, Changyan; Zheng, Rong; Kuang, Shihuan

    The serine/threonine kinase mammalian target of rapamycin (mTOR) is a key regulator of protein synthesis, cell proliferation and energy metabolism. As constitutive deletion of Mtor gene results in embryonic lethality, the function of mTOR in muscle stem cells (satellite cells) and skeletal muscle regeneration remains to be determined. In this study, we established a satellite cell specific Mtor conditional knockout (cKO) mouse model by crossing Pax7(CreER) and Mtor(flox/flox) mice. Skeletal muscle regeneration after injury was severely compromised in the absence of Mtor, indicated by increased number of necrotic myofibers infiltrated by Evans blue dye, and reduced number and size of regenerated myofibers in the Mtor cKO mice compared to wild type (WT) littermates. To dissect the cellular mechanism, we analyzed satellite cell-derived primary myoblasts grown on single myofibers or adhered to culture plates. The Mtor cKO myoblasts exhibited defective proliferation and differentiation kinetics when compared to myoblasts derived from WT littermates. At the mRNA and protein levels, the Mtor cKO myoblasts expressed lower levels of key myogenic determinant genes Pax7, Myf5, Myod, Myog than did the WT myoblasts. These results suggest that mTOR is essential for satellite cell function and skeletal muscle regeneration through controlling the expression of myogenic genes.

  2. mTOR Enhances Foam Cell Formation by Suppressing the Autophagy Pathway

    PubMed Central

    Li, Lingxia; Niu, Xiaolin; Dang, Xiaoyan; Li, Ping; Qu, Li; Bi, Xiaoju; Gao, Yanxia; Hu, Yanfen; Li, Manxiang; Qiao, Wanhai; Peng, Zhuo; Pan, Longfei

    2014-01-01

    Recently, autophagy has drawn more attention in cardiovascular disease as it has important roles in lipid metabolism. Mammalian target of rapamycin (mTOR) is a key regulator of autophagy; however, its effect on atherosclerosis and the underlying mechanism remains undefined. In this study, an obvious upregulation of mTOR and p-mTOR protein was observed in macrophage-derived foam cells. Blocking mTOR expression with specific small interference RNA (siRNA) dramatically suppressed foam cell formation, accompanied by a decrease of lipid deposition. Further mechanistic analysis indicated that suppressing mTOR expression significantly upregulated autophagic marker LC3 expression and downregulated autophagy substrate p62 levels, indicating that mTOR silencing triggered autophagosome formation. Moreover, blocking mTOR expression obviously accelerated neutral lipid delivery to lysosome and cholesterol efflux from foam cells, implying that mTOR could induce macrophage foam cell formation by suppressing autophagic pathway. Further, mTOR silencing significantly upregulated ULK1 expression, which was accounted for mTOR-induced foam cell formation via autophagic pathway as treatment with ULK1 siRNA dampened LC3-II levels and increased p62 expression, concomitant with lipid accumulation and decreased cholesterol efflux from foam cells. Together, our data provide an insight into how mTOR accelerates the pathological process of atherosclerosis. Accordingly, blocking mTOR levels may be a promising therapeutic agent against atherosclerotic complications. PMID:24512183

  3. mTOR signaling and its roles in normal and abnormal brain development.

    PubMed

    Takei, Nobuyuki; Nawa, Hiroyuki

    2014-01-01

    Target of rapamycin (TOR) was first identified in yeast as a target molecule of rapamycin, an anti-fugal and immunosuppressant macrolide compound. In mammals, its orthologue is called mammalian TOR (mTOR). mTOR is a serine/threonine kinase that converges different extracellular stimuli, such as nutrients and growth factors, and diverges into several biochemical reactions, including translation, autophagy, transcription, and lipid synthesis among others. These biochemical reactions govern cell growth and cause cells to attain an anabolic state. Thus, the disruption of mTOR signaling is implicated in a wide array of diseases such as cancer, diabetes, and obesity. In the central nervous system, the mTOR signaling cascade is activated by nutrients, neurotrophic factors, and neurotransmitters that enhances protein (and possibly lipid) synthesis and suppresses autophagy. These processes contribute to normal neuronal growth by promoting their differentiation, neurite elongation and branching, and synaptic formation during development. Therefore, disruption of mTOR signaling may cause neuronal degeneration and abnormal neural development. While reduced mTOR signaling is associated with neurodegeneration, excess activation of mTOR signaling causes abnormal development of neurons and glia, leading to brain malformation. In this review, we first introduce the current state of molecular knowledge of mTOR complexes and signaling in general. We then describe mTOR activation in neurons, which leads to translational enhancement, and finally discuss the link between mTOR and normal/abnormal neuronal growth during development.

  4. A Phase 1 Study of Everolimus + Weekly Cisplatin + Intensity Modulated Radiation Therapy in Head-and-Neck Cancer

    SciTech Connect

    Fury, Matthew G.; Lee, Nancy Y.; Sherman, Eric; Ho, Alan L.; Rao, Shyam; Heguy, Adriana; Shen, Ronglai; Korte, Susan; Lisa, Donna; Ganly, Ian; Patel, Snehal; Wong, Richard J.; Shaha, Ashok; Shah, Jatin; Haque, Sofia; Katabi, Nora; Pfister, David G.

    2013-11-01

    Purpose: Elevated expression of eukaryotic protein synthesis initiation factor 4E (eIF4E) in histologically cancer-free margins of resected head and neck squamous cell carcinomas (HNSCCs) is mediated by mammalian target of rapamycin complex 1 (mTORC1) and has been associated with increased risk of disease recurrence. Preclinically, inhibition of mTORC1 with everolimus sensitizes cancer cells to cisplatin and radiation. Methods and Materials: This was single-institution phase 1 study to establish the maximum tolerated dose of daily everolimus given with fixed dose cisplatin (30 mg/m{sup 2} weekly × 6) and concurrent intensity modulated radiation therapy for patients with locally and/or regionally advanced head-and-neck cancer. The study had a standard 3 + 3 dose-escalation design. Results: Tumor primary sites were oral cavity (4), salivary gland (4), oropharynx (2), nasopharynx (1), scalp (1), and neck node with occult primary (1). In 4 of 4 cases in which resected HNSCC surgical pathology specimens were available for immunohistochemistry, elevated expression of eIF4E was observed in the cancer-free margins. The most common grade ≥3 treatment-related adverse event was lymphopenia (92%), and dose-limiting toxicities (DLTs) were mucositis (n=2) and failure to thrive (n=1). With a median follow up of 19.4 months, 2 patients have experienced recurrent disease. The maximum tolerated dose was everolimus 5 mg/day. Conclusions: Head-and-neck cancer patients tolerated everolimus at therapeutic doses (5 mg/day) given with weekly cisplatin and intensity modulated radiation therapy. The regimen merits further evaluation, especially among patients who are status post resection of HNSCCs that harbor mTORC1-mediated activation of eIF4E in histologically negative surgical margins.

  5. Efficacy of everolimus with reduced-exposure cyclosporine in de novo kidney transplant patients at increased risk for efficacy events: analysis of a randomized trial.

    PubMed

    Carmellini, Mario; Garcia, Valter; Wang, Zailong; Vergara, Marcela; Russ, Graeme

    2015-10-01

    The efficacy of de novo everolimus with reduced-exposure calcineurin inhibitor (CNI) was examined in kidney transplant subpopulations from the A2309 study that were identified to be at increased risk for efficacy events. A2309 was a 24-month, multicenter, open-label trial in which 833 de novo kidney transplant recipients were randomized to everolimus targeting 3-8 or 6-12 ng/ml with reduced-exposure cyclosporine (CsA), or mycophenolic acid (MPA) with standard-exposure CsA, all with basiliximab induction. The composite efficacy endpoint was treated biopsy-proven acute rejection (BPAR), graft loss, death, or loss to follow-up. Cox proportional hazard modeling showed male gender, younger recipient age, black race, delayed graft function, human leukocyte antigen (HLA) mismatch ≥3 and increasing donor age to be significantly predictive for the composite efficacy endpoint at months 12 or 24 post-transplant. CsA exposure was 53-75 % lower, and 46-75 % lower, in patients receiving everolimus 3-8 ng/ml or receiving everolimus 6-12 ng/ml, respectively, versus MPA-treated patients. The incidence of the composite endpoint was similar in all three treatment groups within each subpopulation analyzed. The incidence of treated BPAR was similar with everolimus 3-8 ng/ml or MPA in all subpopulations, but less frequent with everolimus 6-12 ng/ml versus MPA in patients with HLA mismatch ≥3 (p = 0.049). This post hoc analysis of a large, randomized trial suggests that a de novo regimen of everolimus with reduced-exposure CsA maintains immunosuppressive efficacy even in kidney transplant patients at increased risk for efficacy events despite substantial reductions in CsA exposure.

  6. Everolimus Initiation With Early Calcineurin Inhibitor Withdrawal in De Novo Heart Transplant Recipients: Three-Year Results From the Randomized SCHEDULE Study.

    PubMed

    Andreassen, A K; Andersson, B; Gustafsson, F; Eiskjaer, H; Rådegran, G; Gude, E; Jansson, K; Solbu, D; Karason, K; Arora, S; Dellgren, G; Gullestad, L

    2016-04-01

    In a randomized, open-label trial, de novo heart transplant recipients were randomized to everolimus (3-6 ng/mL) with reduced-exposure calcineurin inhibitor (CNI; cyclosporine) to weeks 7-11 after transplant, followed by increased everolimus exposure (target 6-10 ng/mL) with cyclosporine withdrawal or standard-exposure cyclosporine. All patients received mycophenolate mofetil and corticosteroids. A total of 110 of 115 patients completed the 12-month study, and 102 attended a follow-up visit at month 36. Mean measured GFR (mGFR) at month 36 was 77.4 mL/min (standard deviation [SD] 20.2 mL/min) versus 59.2 mL/min (SD 17.4 mL/min) in the everolimus and CNI groups, respectively, a difference of 18.3 mL/min (95% CI 11.1-25.6 mL/min; p < 0.001) in the intention to treat population. Multivariate analysis showed treatment to be an independent determinant of mGFR at month 36. Coronary intravascular ultrasound at 36 months revealed significantly reduced progression of allograft vasculopathy in the everolimus group compared with the CNI group. Biopsy-proven acute rejection grade ≥2R occurred in 10.2% and 5.9% of everolimus- and CNI-treated patients, respectively, during months 12-36. Serious adverse events occurred in 37.3% and 19.6% of everolimus- and CNI-treated patients, respectively (p = 0.078). These results suggest that early CNI withdrawal after heart transplantation supported by everolimus, mycophenolic acid and steroids with lymphocyte-depleting induction is safe at intermediate follow-up. This regimen, used selectively, may offer adequate immunosuppressive potency with a sustained renal advantage.

  7. Therapeutic drug monitoring of everolimus using the dried blood spot method in combination with liquid chromatography-mass spectrometry.

    PubMed

    van der Heijden, J; de Beer, Y; Hoogtanders, K; Christiaans, M; de Jong, G J; Neef, C; Stolk, L

    2009-11-01

    An assay of everolimus based on finger prick sampling and consecutive application as a blood spot on sampling paper has been developed. We explored several methods [K. Hoogtanders, J. van der Heijden, M. Christiaans, P. Edelbroek, J. van Hooff, L. Stolk, J. Pharm. Biomed. Anal. 44 (2006) 658-664; A. Allanson, M. Cotton, J. Tettey, et al., J. Pharm. Biomed. Anal. 44 (2007) 963-969] and developed a new method, namely the impregnation of sampling paper with a solution of plasma-protein, formic acid and ammonium acetate, in combination with the extraction of the blood spot by filter filtration. This kind of sample preparation provides new possibilities for blood spot sampling especially if analytes are adsorbed to the paper. The dried blood spot was analysed using the HPLC-electrospray-tandem mass spectrometry method, with 32-desmethoxyrapamycin as the internal standard. The working range of our study was 2-30 microg/l. Within this range, intra-and inter-assay variability for precision and accuracy was <15%. Everolimus blood spot samples proved stable for 3 days at 60 degrees C and for 32 days at 4 degrees C. Everolimus concentrations of one stable out-patient were compared after both blood spot sampling and conventional venous sampling on various occasions. Results indicate that this new method is promising for therapeutic drug monitoring in stable renal transplant patients.

  8. Benzofuran derivatives as anticancer inhibitors of mTOR signaling.

    PubMed

    Salomé, Christophe; Ribeiro, Nigel; Chavagnan, Thierry; Thuaud, Frédéric; Serova, Maria; de Gramont, Armand; Faivre, Sandrine; Raymond, Eric; Désaubry, Laurent

    2014-06-23

    A series of 32 derivatives and isosteres of the mTOR inhibitor 2 were synthesized and compared for their cytotoxicity in radioresistant SQ20B cancer cell line. Several of these compounds, in particular 30b, were significantly more cytotoxic than 2. Importantly, 30b was shown to block both mTORC1 and Akt signaling, suggesting insensitivity to the resistance associated to Akt overactivation observed with rapamycin derivatives currently used in clinic.

  9. On the participation of mTOR in recognition memory.

    PubMed

    Myskiw, Jociane C; Rossato, Janine I; Bevilaqua, Lia R M; Medina, Jorge H; Izquierdo, Iván; Cammarota, Martín

    2008-03-01

    Evidence indicates that activation of the neuronal protein synthesis machinery is required in areas of the brain relevant to memory for consolidation and persistence of the mnemonic trace. Here, we report that inhibition of hippocampal mTOR, a protein kinase involved in the initiation of mRNA translation, immediately or 180min but not 540min after training impairs consolidation of long-term object recognition memory without affecting short-term memory retention or exploratory behavior. When infused into dorsal CA1 after long-term memory reactivation in the presence of familiar objects the mTOR inhibitor rapamycin (RAP) did not affect retention. However, when given immediately after exposing animals to a novel and a familiar object, RAP impaired memory for both of them. The amnesic effect of the post-retrieval administration of RAP was long-lasting, did not happen after exposure to two novel objects or following exploration of the training arena in the absence of other stimuli, suggesting that it was contingent with reactivation of the consolidated trace in the presence of a behaviorally relevant and novel cue. Our results indicate that mTOR activity is required in the dorsal hippocampus for consolidation of object recognition memory and suggest that inhibition of this kinase after memory retrieval in the presence of a particular set of cues hinders persistence of the original recognition memory trace.

  10. Targeting the mTOR pathway in hepatocellular carcinoma: Current state and future trends

    PubMed Central

    Matter, Matthias S.; Decaens, Thomas; Andersen, Jesper B.; Thorgeirsson, Snorri S.

    2014-01-01

    Summary Mechanistic target of rapamycin (mTOR) regulates cell growth, metabolism and aging in response to nutrients, cellular energy stage and growth factors. mTOR is frequently up-regulated in cancer including hepatocellular carcinoma (HCC) and is associated with bad prognosis, poorly differentiated tumors, and earlier recurrence. Blocking mTOR with rapamycin and first generation mTOR inhibitors, called rapalogs, has shown promising reduction of HCC tumors growth in preclinical models. Currently, rapamycin/rapalogs are used in several clinical trials for the treatment of advanced HCC, and as adjuvant therapy in HCC patients after liver transplantation and TACE. A second generation of mTOR pathway inhibitors has been developed recently, and is being tested in various clinical trials of solid cancers and has been used in preclinical HCC models. The results of series of clinical trials using mTOR inhibitors in HCC treatment will emerge in the near future. PMID:24308993

  11. Cost-effectiveness analysis of everolimus plus exemestane versus exemestane alone for treatment of hormone receptor positive metastatic breast cancer.

    PubMed

    Diaby, Vakaramoko; Adunlin, Georges; Zeichner, Simon B; Avancha, Kiran; Lopes, Gilberto; Gluck, Stefan; Montero, Alberto J

    2014-09-01

    Everolimus in combination with exemestane significantly improved progression-free survival compared to exemestane alone in patients previously treated with non-steroidal aromatase inhibitors in the BOLERO-2 trial. As a result, this combination has been approved by the food and drug administration to treat postmenopausal women with hormone receptor positive and HER2 negative metastatic breast cancer. A cost-effectiveness analysis was conducted to determine whether everolimus represents good value for money, utilizing data from BOLERO-2. A decision-analytic model was used to estimate the incremental cost-effectiveness ratio between treatment arms of the BOLERO-2 trial. Costs were obtained from the Center for Medicare Services drug payment table and physician fee schedule. Benefits were expressed as quality-adjusted progression-free survival weeks (QAPFW) and quality-adjusted progression-free years (QAPFY), with utilities/disutilities derived from the literature. Deterministic and probabilistic sensitivity analyses were performed. A willingness to pay threshold of 1-3 times the per capita gross domestic product was adopted, as per the definition of the World Health Organization. The U.S. per capita gross domestic product in 2013 was $49,965; thus, a threshold varying between $49,965 and $149,895 was considered. Everolimus/exemestane had an incremental benefit of 11.88 QAPFW (0.22 QAPFY) compared to exemestane and an incremental cost of $60,574. This translated into an ICER of $265,498.5/QAPFY. Univariate sensitivity analyses showed important variations of the ICER, ranging between $189,836.4 and $530,947/QAPFY. A tornado analysis suggested that the key drivers of our model, by order of importance, included health utility value for stable disease, everolimus acquisition costs, and transition probabilities from the stable to the progression states. The Monte-Carlo simulation showed results that were similar to the base-case analysis. This cost-effectiveness analysis

  12. The ketogenic diet inhibits the mammalian target of rapamycin (mTOR) pathway.

    PubMed

    McDaniel, Sharon S; Rensing, Nicholas R; Thio, Liu Lin; Yamada, Kelvin A; Wong, Michael

    2011-03-01

    The ketogenic diet (KD) is an effective treatment for epilepsy, but its mechanisms of action are poorly understood. We investigated the hypothesis that the KD inhibits mammalian target of rapamycin (mTOR) pathway signaling. The expression of pS6 and pAkt, markers of mTOR pathway activation, was reduced in hippocampus and liver of rats fed KD. In the kainate model of epilepsy, KD blocked the hippocampal pS6 elevation that occurs after status epilepticus. Because mTOR signaling has been implicated in epileptogenesis, these results suggest that the KD may have anticonvulsant or antiepileptogenic actions via mTOR pathway inhibition.

  13. Adapting the Stress Response: Viral Subversion of the mTOR Signaling Pathway.

    PubMed

    Le Sage, Valerie; Cinti, Alessandro; Amorim, Raquel; Mouland, Andrew J

    2016-05-24

    The mammalian target of rapamycin (mTOR) is a central regulator of gene expression, translation and various metabolic processes. Multiple extracellular (growth factors) and intracellular (energy status) molecular signals as well as a variety of stressors are integrated into the mTOR pathway. Viral infection is a significant stress that can activate, reduce or even suppress the mTOR signaling pathway. Consequently, viruses have evolved a plethora of different mechanisms to attack and co-opt the mTOR pathway in order to make the host cell a hospitable environment for replication. A more comprehensive knowledge of different viral interactions may provide fruitful targets for new antiviral drugs.

  14. Resveratrol induces autophagy by directly inhibiting mTOR through ATP competition

    PubMed Central

    Park, Dohyun; Jeong, Heeyoon; Lee, Mi Nam; Koh, Ara; Kwon, Ohman; Yang, Yong Ryoul; Noh, Jungeun; Suh, Pann-Ghill; Park, Hwangseo; Ryu, Sung Ho

    2016-01-01

    Resveratrol (RSV) is a natural polyphenol that has a beneficial effect on health, and resveratrol-induced autophagy has been suggested to be a key process in mediating many beneficial effects of resveratrol, such as reduction of inflammation and induction of cancer cell death. Although various resveratrol targets have been suggested, the molecule that mediates resveratrol-induced autophagy remains unknown. Here, we demonstrate that resveratrol induces autophagy by directly inhibiting the mTOR-ULK1 pathway. We found that inhibition of mTOR activity and presence of ULK1 are required for autophagy induction by resveratrol. In line with this mTOR dependency, we found that resveratrol suppresses the viability of MCF7 cells but not of SW620 cells, which are mTOR inhibitor sensitive and insensitive cancer cells, respectively. We also found that resveratrol-induced cancer cell suppression occurred ULK1 dependently. For the mechanism of action of resveratrol on mTOR inhibition, we demonstrate that resveratrol directly inhibits mTOR. We found that resveratrol inhibits mTOR by docking onto the ATP-binding pocket of mTOR (i.e., it competes with ATP). We propose mTOR as a novel direct target of resveratrol, and inhibition of mTOR is necessary for autophagy induction. PMID:26902888

  15. The mTOR signalling cascade: paving new roads to cure neurological disease.

    PubMed

    Crino, Peter B

    2016-07-01

    Defining the multiple roles of the mechanistic (formerly 'mammalian') target of rapamycin (mTOR) signalling pathway in neurological diseases has been an exciting and rapidly evolving story of bench-to-bedside translational research that has spanned gene mutation discovery, functional experimental validation of mutations, pharmacological pathway manipulation, and clinical trials. Alterations in the dual contributions of mTOR - regulation of cell growth and proliferation, as well as autophagy and cell death - have been found in developmental brain malformations, epilepsy, autism and intellectual disability, hypoxic-ischaemic and traumatic brain injuries, brain tumours, and neurodegenerative disorders. mTOR integrates a variety of cues, such as growth factor levels, oxygen levels, and nutrient and energy availability, to regulate protein synthesis and cell growth. In line with the positioning of mTOR as a pivotal cell signalling node, altered mTOR activation has been associated with a group of phenotypically diverse neurological disorders. To understand how altered mTOR signalling leads to such divergent phenotypes, we need insight into the differential effects of enhanced or diminished mTOR activation, the developmental context of these changes, and the cell type affected by altered signalling. A particularly exciting feature of the tale of mTOR discovery is that pharmacological mTOR inhibitors have shown clinical benefits in some neurological disorders, such as tuberous sclerosis complex, and are being considered for clinical trials in epilepsy, autism, dementia, traumatic brain injury, and stroke.

  16. Anti-tumor activity of selective inhibitor of nuclear export (SINE) compounds, is enhanced in non-Hodgkin lymphoma through combination with mTOR inhibitor and dexamethasone.

    PubMed

    Muqbil, Irfana; Aboukameel, Amro; Elloul, Sivan; Carlson, Robert; Senapedis, William; Baloglu, Erkan; Kauffman, Michael; Shacham, Sharon; Bhutani, Divaya; Zonder, Jeffrey; Azmi, Asfar S; Mohammad, Ramzi M

    2016-12-28

    In previous studies we demonstrated that targeting the nuclear exporter protein exportin-1 (CRM1/XPO1) by a selective inhibitor of nuclear export (SINE) compound is a viable therapeutic strategy against Non-Hodgkin Lymphoma (NHL). Our studies along with pre-clinical work from others led to the evaluation of the lead SINE compound, selinexor, in a phase 1 trial in patients with CLL or NHL (NCT02303392). Continuing our previous work, we studied combinations of selinexor-dexamethasone (DEX) and selinexor-everolimus (EVER) in NHL. Combination of selinexor with DEX or EVER resulted in enhanced cytotoxicity in WSU-DLCL2 and WSU-FSCCL cells which was consistent with enhanced apoptosis. Molecular analysis showed enhancement in the activation of apoptotic signaling and down-regulation of XPO1. This enhancement is consistent with the mechanism of action of these drugs in that both selinexor and DEX antagonize NF-κB (p65) and mTOR (EVER target) is an XPO1 cargo protein. SINE compounds, KPT-251 and KPT-276, showed activities similar to CHOP (cyclophosphamide-hydroxydaunorubicin-oncovin-prednisone) regimen in subcutaneous and disseminated NHL xenograft models in vivo. In both animal models the anti-lymphoma activity of selinexor is enhanced through combination with DEX or EVER. The in vivo activity of selinexor and related SINE compounds relative to 'standard of care' treatment is consistent with the objective responses observed in Phase I NHL patients treated with selinexor. Our pre-clinical data provide a rational basis for testing these combinations in Phase II NHL trials.

  17. mTOR pathway activation in multiple myeloma cell lines and primary tumour cells: pomalidomide enhances cytoplasmic-nuclear shuttling of mTOR protein

    PubMed Central

    Guglielmelli, Tommasina; Giugliano, Emilia; Brunetto, Vanessa; Rapa, Ida; Cappia, Susanna; Giorcelli, Jessica; Rrodhe, Sokol; Papotti, Mauro; Saglio, Giuseppe

    2015-01-01

    mTOR is a protein kinase that plays a central role in regulating critical cellular processes. We evaluated the activation and cellular localization of the mTOR pathway in multiple myeloma (MM) and analyzed the role of pomalidomide in regulating mTOR. By immunohistochemistry cytoplasmic p-mTOR stained positive in 57 out 101 (57.6%) cases with a nuclear p-mTOR localization in 14 out 101 samples (13.8%). In the 70 MM samples analyzed for the entire pathway, p-mTOR expression significantly correlated with p-AKT, p-P70S6K, and p-4E-BP1 suggesting that the AKT/mTOR pathway is activated in a subset of MM patients. Immunofluorescence assays demonstrated that mTOR protein is distributed throughout the cytoplasm and the nucleus at baseline in MM cell lines and in plasma cells of 13 MM patients and that pomalidomide facilitated the shift of the mTOR protein in the nucleus. By western blotting, treatment with pomalidomide increased nuclear mTOR and p-mTOR expression levels in the nucleus with a concomitant decrease of the cytoplasmic fractions while does not seem to affect significantly AKT phosphorylation status. In MM cells the anti-myeloma activity of pomalidomide may be mediated by the regulation of the mTOR pathway. PMID:26097872

  18. Mechanical stimulation induces mTOR signaling via an ERK-independent mechanism: implications for a direct activation of mTOR by phosphatidic acid.

    PubMed

    You, Jae Sung; Frey, John W; Hornberger, Troy A

    2012-01-01

    Signaling by mTOR is a well-recognized component of the pathway through which mechanical signals regulate protein synthesis and muscle mass. However, the mechanisms involved in the mechanical regulation of mTOR signaling have not been defined. Nevertheless, recent studies suggest that a mechanically-induced increase in phosphatidic acid (PA) may be involved. There is also evidence which suggests that mechanical stimuli, and PA, utilize ERK to induce mTOR signaling. Hence, we reasoned that a mechanically-induced increase in PA might promote mTOR signaling via an ERK-dependent mechanism. To test this, we subjected mouse skeletal muscles to mechanical stimulation in the presence or absence of a MEK/ERK inhibitor, and then measured several commonly used markers of mTOR signaling. Transgenic mice expressing a rapamycin-resistant mutant of mTOR were also used to confirm the validity of these markers. The results demonstrated that mechanically-induced increases in p70(s6k) T389 and 4E-BP1 S64 phosphorylation, and unexpectedly, a loss in total 4E-BP1, were fully mTOR-dependent signaling events. Furthermore, we determined that mechanical stimulation induced these mTOR-dependent events, and protein synthesis, through an ERK-independent mechanism. Similar to mechanical stimulation, exogenous PA also induced mTOR-dependent signaling via an ERK-independent mechanism. Moreover, PA was able to directly activate mTOR signaling in vitro. Combined, these results demonstrate that mechanical stimulation induces mTOR signaling, and protein synthesis, via an ERK-independent mechanism that potentially involves a direct interaction of PA with mTOR. Furthermore, it appears that a decrease in total 4E-BP1 may be part of the mTOR-dependent mechanism through which mechanical stimuli activate protein synthesis.

  19. Nanoscale mechanical measurement determination of the glass transition temperature of poly(lactic acid)/everolimus coated stents in air and dissolution media.

    PubMed

    Wu, Ming; Kleiner, Lothar; Tang, Fuh-Wei; Hossainy, Syed; Davies, Martyn C; Roberts, Clive J

    2009-03-02

    Localized atomic force microscopy (AFM) force analysis on poly(lactic acid) (PLA) and poly(lactic acid)/everolimus coated stents has been performed under ambient conditions. Similar Young's modulus were derived from both PLA and PLA/everolimus stent surface, namely 2.25+/-0.46 and 2.04+/-0.39GPa, respectively, indicating that the drug, everolimus does not significantly effect the mechanical properties of PLA up to a 1:1 (w/w) drug loading. Temperature controlled force measurements on PLA only coated stents in air and in a 1% Triton surfactant solution allowed the glass transition temperature (T(g)) of the polymer to be determined. A significant drop of the Young's modulus in solution was observed at 36 degrees C, suggests that in vivo the T(g) of the polymer is below body temperature. The possible consequences on drug release and the mechanisms by which this may occur are considered.

  20. Skeletal myocyte hypertrophy requires mTOR kinase activity and S6K1

    SciTech Connect

    Park, In-Hyun . E-mail: ihpark@uiuc.edu; Erbay, Ebru; Nuzzi, Paul; Chen Jie

    2005-09-10

    The protein kinase mammalian target of rapamycin (mTOR) is a central regulator of cell proliferation and growth, with the ribosomal subunit S6 kinase 1 (S6K1) as one of the key downstream signaling effectors. A critical role of mTOR signaling in skeletal muscle differentiation has been identified recently, and an unusual regulatory mechanism independent of mTOR kinase activity and S6K1 is revealed. An mTOR pathway has also been reported to regulate skeletal muscle hypertrophy, but the regulatory mechanism is not completely understood. Here, we report the investigation of mTOR's function in insulin growth factor I (IGF-I)-induced C2C12 myotube hypertrophy. Added at a later stage when rapamycin no longer had any effect on normal myocyte differentiation, rapamycin completely blocked myocyte hypertrophy as measured by myotube diameter. Importantly, a concerted increase of average myonuclei per myotube was observed in IGF-I-stimulated myotubes, which was also inhibited by rapamycin added at a time when it no longer affected normal differentiation. The mTOR protein level, its catalytic activity, its phosphorylation on Ser2448, and the activity of S6K1 were all found increased in IGF-I-stimulated myotubes compared to unstimulated myotubes. Using C2C12 cells stably expressing rapamycin-resistant forms of mTOR and S6K1, we provide genetic evidence for the requirement of mTOR and its downstream effector S6K1 in the regulation of myotube hypertrophy. Our results suggest distinct mTOR signaling mechanisms in different stages of skeletal muscle development: While mTOR regulates the initial myoblast differentiation in a kinase-independent and S6K1-independent manner, the hypertrophic function of mTOR requires its kinase activity and employs S6K1 as a downstream effector.

  1. Everolimus-Based Therapy versus Chemotherapy among Patients with HR+/HER2- Metastatic Breast Cancer: Comparative Effectiveness from a Chart Review Study.

    PubMed

    Li, Nanxin; Hao, Yanni; Xie, Jipan; Lin, Peggy L; Koo, Valerie; Ohashi, Erika; Wu, Eric Q

    2015-01-01

    Objective. To compare the real-world effectiveness of everolimus-based therapy and chemotherapy in postmenopausal women with hormone-receptor-positive/human-epidermal-growth-factor-receptor-2-negative (HR+/HER2-) metastatic breast cancer (mBC). Methods. This retrospective chart review examined a nationwide sample of postmenopausal HR+/HER2- mBC women in community-based oncology practices. Patients received everolimus-based therapy or chemotherapy for mBC between 07/01/2012 and 04/15/2013, after failure of a non-steroidal aromatase inhibitor. Overall survival (OS), progression-free survival (PFS), and time on treatment (TOT) were compared using Kaplan-Meier analysis and Cox proportional hazards models adjusting for line of therapy and baseline characteristics. Results. 234 and 137 patients received everolimus-based therapy and chemotherapy. Patients treated with everolimus-based therapy tended to have less aggressive mBC than patients treated with chemotherapy. Multivariate-adjusted Cox models showed that everolimus-based therapy was associated with significantly longer OS [hazard ratio (HR) = 0.37, 95% confidence interval (CI): 0.22-0.63], PFS (HR = 0.70, 95% CI = 0.50-0.97), and TOT (HR = 0.34, 95% CI: 0.25-0.45) than chemotherapy. Adjusted comparative effectiveness results were generally consistent across lines of therapy. Conclusion. In this retrospective chart review of postmenopausal HR+/HER2- mBC patients, treatment with everolimus-based therapy was associated with longer OS, PFS, and TOT than chemotherapy.

  2. RTOG 0913: A Phase 1 Study of Daily Everolimus (RAD001) in Combination With Radiation Therapy and Temozolomide in Patients With Newly Diagnosed Glioblastoma

    SciTech Connect

    Chinnaiyan, Prakash; Won, Minhee; Wen, Patrick Y.; Wendland, Merideth; Dipetrillo, Thomas A.; Corn, Benjamin W.; Mehta, Minesh P.

    2013-08-01

    Purpose: To determine the safety of the mammalian target of rapamycin inhibitor everolimus (RAD001) administered daily with concurrent radiation and temozolomide in newly diagnosed glioblastoma patients. Methods and Materials: Everolimus was administered daily with concurrent radiation (60 Gy in 30 fractions) and temozolomide (75 mg/m{sup 2} per day). Everolimus was escalated from 2.5 mg/d (dose level 1) to 5 mg/d (dose level 2) to 10 mg/d (dose level 3). Adjuvant temozolomide was delivered at 150 to 200 mg/m{sup 2} on days 1 to 5, every 28 days, for up to 12 cycles, with concurrent everolimus at the previously established daily dose of 10 mg/d. Dose escalation continued if a dose level produced dose-limiting toxicities (DLTs) in fewer than 3 of the first 6 evaluable patients. Results: Between October 28, 2010, and July 2, 2012, the Radiation Therapy Oncology Group 0913 protocol initially registered a total of 35 patients, with 25 patients successfully meeting enrollment criteria receiving the drug and evaluable for toxicity. Everolimus was successfully escalated to the predetermined maximum tolerated dose of 10 mg/d. Two of the first 6 eligible patients had a DLT at each dose level. DLTs included gait disturbance, febrile neutropenia, rash, fatigue, thrombocytopenia, hypoxia, ear pain, headache, and mucositis. Other common toxicities were grade 1 or 2 hypercholesterolemia and hypertriglyceridemia. At the time of analysis, there was 1 death reported, which was attributed to tumor progression. Conclusions: Daily oral everolimus (10 mg) combined with both concurrent radiation and temozolomide followed by adjuvant temozolomide is well tolerated, with an acceptable toxicity profile. A randomized phase 2 clinical trial with mandatory correlative biomarker analysis is currently under way, designed to both determine the efficacy of this regimen and identify molecular determinants of response.

  3. The tug-of-war over MTOR in Legionella infections

    PubMed Central

    Ivanov, Stanimir S.

    2017-01-01

    A ruptured bacteria-containing organelle within the cytosol of an infected eukaryotic cell frequently initiates host defense responses that restrict pathogen replication. Therefore, source for lipids must be found to accommodate the organelle membrane expansion required to support bacterial replication. How host cells are manipulated to provide lipids for the expansion of pathogen-occupied organelles is not well understood. By investigating the interaction between macrophages and the human pulmonary pathogen Legionella pneumophila we uncovered that the host metabolic checkpoint kinase Mechanistic target of rapamycin (MTOR) is a central regulator of the pathogen niche expansion program. PMID:28357391

  4. Activation of Both the Calpain and Ubiquitin-Proteasome Systems Contributes to Septic Cardiomyopathy through Dystrophin Loss/Disruption and mTOR Inhibition

    PubMed Central

    Freitas, Ana Caroline Silva; Figueiredo, Maria Jose; Campos, Erica Carolina; Soave, Danilo Figueiredo; Ramos, Simone Gusmao; Tanowitz, Herbert B.

    2016-01-01

    Cardiac dysfunction caused by the impairment of myocardial contractility has been recognized as an important factor contributing to the high mortality in sepsis. Calpain activation in the heart takes place in response to increased intracellular calcium influx resulting in proteolysis of structural and contractile proteins with subsequent myocardial dysfunction. The purpose of the present study was to test the hypothesis that increased levels of calpain in the septic heart leads to disruption of structural and contractile proteins and that administration of calpain inhibitor-1 (N-acetyl-leucinyl-leucinyl-norleucinal (ALLN)) after sepsis induced by cecal ligation and puncture prevents cardiac protein degradation. We also tested the hypothesis that calpain plays a role in the modulation of protein synthesis/degradation through the activation of proteasome-dependent proteolysis and inhibition of the mTOR pathway. Severe sepsis significantly increased heart calpain-1 levels and promoted ubiquitin and Pa28β over-expression with a reduction in the mTOR levels. In addition, sepsis reduced the expression of structural proteins dystrophin and β-dystroglycan as well as the contractile proteins actin and myosin. ALLN administration prevented sepsis-induced increases in calpain and ubiquitin levels in the heart, which resulted in decreased of structural and contractile proteins degradation and basal mTOR expression levels were re-established. Our results support the concept that increased calpain concentrations may be part of an important mechanism of sepsis-induced cardiac muscle proteolysis. PMID:27880847

  5. Overactive mTOR signaling leads to endometrial hyperplasia in aged women and mice.

    PubMed

    Bajwa, Preety; Nielsen, Sarah; Lombard, Janine M; Rassam, Loui; Nahar, Pravin; Rueda, Bo R; Wilkinson, J Erby; Miller, Richard A; Tanwar, Pradeep S

    2017-01-31

    During aging, uncontrolled epithelial cell proliferation in the uterus results in endometrial hyperplasia and/or cancer development. The mTOR signaling pathway is one of the major regulators of aging as suppression of this pathway prolongs lifespan in model organisms. Genetic alterations in this pathway via mutations and/or amplifications are often encountered in endometrial cancers. However, the exact contribution of mTOR signaling and uterine aging to endometrial pathologies is currently unclear. This study examined the role of mTOR signaling in uterine aging and its implications in the development of endometrial hyperplasia. The hyperplastic endometrium of both postmenopausal women and aged mice exhibited elevated mTOR activity as seen with increased expression of the pS6 protein. Analysis of uteri from Pten heterozygous and Pten overexpressing mice further confirmed that over-activation of mTOR signaling leads to endometrial hyperplasia. Pharmacological inhibition of mTOR signaling using rapamycin treatment suppressed endometrial hyperplasia in aged mice. Furthermore, treatment with mTOR inhibitors reduced colony size and proliferation of a PTEN negative endometrial cancer cell line in 3D culture. Collectively, this study suggests that hyperactivation of the mTOR pathway is involved in the development of endometrial hyperplasia in aged women and mice.

  6. Metformin inhibits thyroid cancer cell growth, migration, and EMT through the mTOR pathway.

    PubMed

    Han, Baiyu; Cui, Hanzhi; Kang, Lei; Zhang, Xuelin; Jin, Zhitao; Lu, Lanmin; Fan, Zhongyi

    2015-08-01

    Mammalian target of rapamycin (mTOR) signaling pathways have been shown to be activated in thyroid cancer. Recent evidences have demonstrated that the antidiabetic agent metformin, an activator of 5'-AMP-activated protein kinase, can impair the proliferation and migration of cancer cells via inhibition of mTOR. However, the underlying mechanisms remain unclear. In this study, we show that metformin can inhibit mTOR pathway to impair growth and migration of the thyroid cancer cell lines. Cyclin D1 and c-Myc are important regulators of cancer cell growth, and we observed that treatment of thyroid cancer cells with metformin reduced c-Myc and cyclin D1 expression through suppression of mTOR and subsequent inhibition of P70S6K1 and 4E-BP1 phosphorylation. Metformin reduced epithelial to mesenchymal transition (EMT) in thyroid carcinoma cells. Moreover, metformin regulated expression of the EMT-related markers E-cadherin, N-cadherin, and Snail. Additionally, knockdown of TSC2, the upstream regulatory molecule of mTOR pathway, or treatment of rapamycin, the mTOR inhibitor, could abolish the effects of metformin to regulate thyroid cancer cell proliferation, migration, EMT, and mTOR pathway molecules. These results indicate that metformin can suppress the proliferation, migration, and EMT of thyroid cancer cell lines by inhibiting mTOR signaling. These findings suggest that metformin and its molecular targets may be useful in thyroid carcinoma therapy.

  7. The ever-evolving role of mTOR in translation.

    PubMed

    Fonseca, Bruno D; Smith, Ewan M; Yelle, Nicolas; Alain, Tommy; Bushell, Martin; Pause, Arnim

    2014-12-01

    Control of translation allows for the production of stoichiometric levels of each protein in the cell. Attaining such a level of fine-tuned regulation of protein production requires the coordinated temporal and spatial control of numerous cellular signalling cascades impinging on the various components of the translational machinery. Foremost among these is the mTOR signalling pathway. The mTOR pathway regulates both the initiation and elongation steps of protein synthesis through the phosphorylation of numerous translation factors, while simultaneously ensuring adequate folding of nascent polypeptides through co-translational degradation of misfolded proteins. Perhaps most remarkably, mTOR is also a key regulator of the synthesis of ribosomal proteins and translation factors themselves. Two seminal studies have recently shown in translatome analysis that the mTOR pathway preferentially regulates the translation of mRNAs encoding ribosomal proteins and translation factors. Therefore, the role of the mTOR pathway in the control of protein synthesis extends far beyond immediate translational control. By controlling ribosome production (and ultimately ribosome availability), mTOR is a master long-term controller of protein synthesis. Herein, we review the literature spanning the early discoveries of mTOR on translation to the latest advances in our understanding of how the mTOR pathway controls the synthesis of ribosomal proteins.

  8. Promotion of ovarian follicle growth following mTOR activation: synergistic effects of AKT stimulators.

    PubMed

    Cheng, Yuan; Kim, Jaehong; Li, Xiao Xiao; Hsueh, Aaron J

    2015-01-01

    Mammalian target of rapamycin (mTOR) is a serine/threonine kinase and mTOR signaling is important in regulating cell growth and proliferation. Recent studies using oocyte- and granulosa cell-specific deletion of mTOR inhibitor genes TSC1 or TSC2 demonstrated the important role of mTOR signaling in the promotion of ovarian follicle development. We now report that treatment of ovaries from juvenile mice with an mTOR activator MHY1485 stimulated mTOR, S6K1 and rpS6 phosphorylation. Culturing ovaries for 4 days with MHY1485 increased ovarian explant weights and follicle development. In vivo studies further demonstrated that pre-incubation of these ovaries with MHY1485 for 2 days, followed by allo-grafting into kidney capsules of adult ovariectomized hosts for 5 days, led to marked increases in graft weights and promotion of follicle development. Mature oocytes derived from MHY1485-activated ovarian grafts could be successfully fertilized, leading the delivery of healthy pups. We further treated ovaries with the mTOR activator together with AKT activators (PTEN inhibitor and phosphoinositol-3-kinase stimulator) before grafting and found additive enhancement of follicle growth. Our studies demonstrate the ability of an mTOR activator in promoting follicle growth, leading to a potential strategy to stimulate preantral follicle growth in infertile patients.

  9. Promotion of Ovarian Follicle Growth following mTOR Activation: Synergistic Effects of AKT Stimulators

    PubMed Central

    Cheng, Yuan; Kim, Jaehong; Li, Xiao Xiao; Hsueh, Aaron J.

    2015-01-01

    Mammalian target of rapamycin (mTOR) is a serine/threonine kinase and mTOR signaling is important in regulating cell growth and proliferation. Recent studies using oocyte- and granulosa cell-specific deletion of mTOR inhibitor genes TSC1 or TSC2 demonstrated the important role of mTOR signaling in the promotion of ovarian follicle development. We now report that treatment of ovaries from juvenile mice with an mTOR activator MHY1485 stimulated mTOR, S6K1 and rpS6 phosphorylation. Culturing ovaries for 4 days with MHY1485 increased ovarian explant weights and follicle development. In vivo studies further demonstrated that pre-incubation of these ovaries with MHY1485 for 2 days, followed by allo-grafting into kidney capsules of adult ovariectomized hosts for 5 days, led to marked increases in graft weights and promotion of follicle development. Mature oocytes derived from MHY1485-activated ovarian grafts could be successfully fertilized, leading the delivery of healthy pups. We further treated ovaries with the mTOR activator together with AKT activators (PTEN inhibitor and phosphoinositol-3-kinase stimulator) before grafting and found additive enhancement of follicle growth. Our studies demonstrate the ability of an mTOR activator in promoting follicle growth, leading to a potential strategy to stimulate preantral follicle growth in infertile patients. PMID:25710488

  10. Everolimus With Reduced Tacrolimus Improves Renal Function in De Novo Liver Transplant Recipients: A Randomized Controlled Trial

    PubMed Central

    De Simone, P; Nevens, F; De Carlis, L; Metselaar, H J; Beckebaum, S; Saliba, F; Jonas, S; Sudan, D; Fung, J; Fischer, L; Duvoux, C; Chavin, K D; Koneru, B; Huang, M A; Chapman, W C; Foltys, D; Witte, S; Jiang, H; Hexham, J M; Junge, G

    2012-01-01

    In a prospective, multicenter, open-label study, de novo liver transplant patients were randomized at day 30±5 to (i) everolimus initiation with tacrolimus elimination (TAC Elimination) (ii) everolimus initiation with reduced-exposure tacrolimus (EVR+Reduced TAC) or (iii) standard-exposure tacrolimus (TAC Control). Randomization to TAC Elimination was terminated prematurely due to a higher rate of treated biopsy-proven acute rejection (tBPAR). EVR+Reduced TAC was noninferior to TAC Control for the primary efficacy endpoint (tBPAR, graft loss or death at 12 months posttransplantation): 6.7% versus 9.7% (−3.0%; 95% CI −8.7, 2.6%; p<0.001 for noninferiority [12% margin]). tBPAR occurred in 2.9% of EVR+Reduced TAC patients versus 7.0% of TAC Controls (p = 0.035). The change in adjusted estimated GFR from randomization to month 12 was superior with EVR+Reduced TAC versus TAC Control (difference 8.50 mL/min/1.73 m2, 97.5% CI 3.74, 13.27 mL/min/1.73 m2, p<0.001 for superiority). Drug discontinuation for adverse events occurred in 25.7% of EVR+Reduced TAC and 14.1% of TAC Controls (relative risk 1.82, 95% CI 1.25, 2.66). Relative risk of serious infections between the EVR+Reduced TAC group versus TAC Controls was 1.76 (95% CI 1.03, 3.00). Everolimus facilitates early tacrolimus minimization with comparable efficacy and superior renal function, compared to a standard tacrolimus exposure regimen 12 months after liver transplantation. PMID:22882750

  11. Synergistic activity of ALK and mTOR inhibitors for the treatment of NPM-ALK positive lymphoma

    PubMed Central

    Redaelli, Sara; Ceccon, Monica; Antolini, Laura; Rigolio, Roberta; Pirola, Alessandra; Peronaci, Marco; Gambacorti-Passerini, Carlo; Mologni, Luca

    2016-01-01

    ALK-positive Anaplastic Large Cell Lymphoma (ALCL) represents a subset of Non-Hodgkin Lymphoma whose treatment benefited from crizotinib development, a dual ALK/MET inhibitor. Crizotinib blocks ALK-triggered pathways such as PI3K/AKT/mTOR, indispensable for survival of ALK-driven tumors. Despite the positive impact of targeted treatment in ALCL, resistant clones are often selected during therapy. Strategies to overcome resistance include the design of second generation drugs and the use of combined therapies that simultaneously target multiple nodes essential for cells survival. We investigated the effects of combined ALK/mTOR inhibition. We observed a specific synergistic effect of combining ALK inhibitors with an mTOR inhibitor (temsirolimus), in ALK+ lymphoma cells. The positive cooperation resulted in an increased inhibition of mTOR effectors, compared to single treatments, a block in G0/G1 phase and induction of apoptosis. The combination was able to prevent the selection of resistant clones, while long-term exposure to single agents led to the establishment of resistant cell lines, with either ALK inhibitor or temsirolimus. In vivo, mice injected with Karpas 299 cells and treated with low dose combination showed complete regression of tumors, while only partial inhibition was obtained in single agents-treated mice. Upon treatment stop the combination was able to significantly delay tumor relapses. Re-challenge of relapsed tumors at a higher dose led to full regression of xenografts in the combination group, but not in mice treated with lorlatinib alone. In conclusion, our data suggest that the combination of ALK and mTOR inhibitors could be a valuable therapeutic option for ALK+ ALCL patients. PMID:27662658

  12. Placental mTOR links maternal nutrient availability to fetal growth.

    PubMed

    Roos, Sara; Powell, Theresa L; Jansson, Thomas

    2009-02-01

    The mTOR (mammalian target of rapamycin) signalling pathway functions as a nutrient sensor, both in individual cells and, more globally, in organs such as the fat body in Drosophila and the hypothalamus in the rat. The activity of placental amino acid transporters is decreased in IUGR (intrauterine growth restriction), and recent experimental evidence suggests that these changes contribute directly to the restricted fetal growth. We have shown that mTOR regulates the activity of the placental L-type amino acid transporter system and that placental mTOR activity is decreased in IUGR. The present review summarizes the emerging evidence implicating placental mTOR signalling as a key mechanism linking maternal nutrient and growth factor concentrations to amino acid transport in the human placenta. Since fetal growth is critically dependent on placental nutrient transport, placental mTOR signalling plays an important role in the regulation of fetal growth.

  13. Targeting bone metastatic cancer: Role of the mTOR pathway.

    PubMed

    Bertoldo, Francesco; Silvestris, Franco; Ibrahim, Toni; Cognetti, Francesco; Generali, Daniele; Ripamonti, Carla Ida; Amadori, Dino; Colleoni, Marco Angelo; Conte, Pierfranco; Del Mastro, Lucia; De Placido, Sabino; Ortega, Cinzia; Santini, Daniele

    2014-04-01

    One of the great challenges of cancer medicine is to develop effective treatments for bone metastatic cancer. Most patients with advanced solid tumors will develop bone metastasis and will suffer from skeletal related events associated with this disease. Although some therapies are available to manage symptoms derived from bone metastases, an effective treatment has not been developed yet. The mammalian target of rapamycin (mTOR) pathway regulates cell growth and survival. Alterations in mTOR signaling have been associated with pathological malignancies, including bone metastatic cancer. Inhibition of mTOR signaling might therefore be a promising alternative for bone metastatic cancer management. This review summarizes the current knowledge on mTOR pathway signaling in bone tissue and provides an overview on the known effects of mTOR inhibition in bone cancer, both in in vitro and in vivo models.

  14. Characterization of pomiferin triacetate as a novel mTOR and translation inhibitor

    PubMed Central

    Bajer, Magdalena M.; Kunze, Michael M.; Blees, Johanna S.; Bokesch, Heidi R.; Chen, Hanyong; Brauß, Thilo F.; Dong, Zigang; Gustafson, Kirk R.; Biondi, Ricardo M.; Henrich, Curtis J.; McMahon, James B.; Colburn, Nancy H.; Schmid, Tobias; Brüne, Bernhard

    2014-01-01

    Deregulation of the phosphatidylinositol 3-kinase (PI3K)-Akt-mammalian target of rapamycin (mTOR)-70 kDa ribosomal protein S6 kinase 1 (p70S6K) pathway is commonly observed in many tumors. This pathway controls proliferation, survival, and translation, and its overactivation is associated with poor prognosis for tumor-associated survival. Current efforts focus on the development of novel inhibitors of this pathway. In a cell-based high-throughput screening assay of 15 272 pure natural compounds, we identified pomiferin triacetate as a potent stabilizer of the tumor suppressor programmed cell death 4 (Pdcd4). Mechanistically, pomiferin triacetate appeared as a general inhibitor of the PI3K-Akt-mTOR-p70S6K cascade. Interference with this pathway occurred downstream of Akt but upstream of p70S6K. Specifically, mTOR kinase emerged as the molecular target of pomiferin triacetate, with similar activities against mTOR complexes 1 and 2. In an in vitro mTOR kinase assay pomiferin triacetate dose-dependently inhibited mTOR with an IC50 of 6.2 µM. Molecular docking studies supported the interaction of the inhibitor with the catalytic site of mTOR. Importantly, pomiferin triacetate appeared to be highly selective for mTOR compared to a panel of 17 lipid and 50 protein kinases tested. As a consequence of the mTOR inhibition, pomiferin triacetate efficiently attenuated translation. In summary, pomiferin triacetate emerged as a novel and highly specific mTOR inhibitor with strong translation inhibitory effects. Thus, it might be an interesting lead structure for the development of mTOR- and translation-targeted anti-tumor therapies. PMID:24513322

  15. The emerging role of m-TOR up-regulation in brain Astrocytoma.

    PubMed

    Ryskalin, Larisa; Limanaqi, Fiona; Biagioni, Francesca; Frati, Alessandro; Esposito, Vincenzo; Calierno, Maria Teresa; Lenzi, Paola; Fornai, Francesco

    2017-05-01

    The present manuscript is an overview of various effects of mTOR up-regulation in astrocytoma with an emphasis on its deleterious effects on the proliferation of Glioblastoma Multiforme. The manuscript reports consistent evidence indicating the occurrence of mTOR up-regulation both in experimental and human astrocytoma. The grading of human astrocytoma is discussed in relationship with mTOR up-regulation. In the second part of the manuscript, the biochemical pathways under the influence of mTOR are translated to cell phenotypes which are generated by mTOR up-regulation and reverted by its inhibition. A special section is dedicated to the prominent role of autophagy in mediating the effects of mTOR in glioblastoma. In detail, autophagy inhibition produced by mTOR up-regulation determines the fate of cancer stem cells. On the other hand, biochemical findings disclose the remarkable effects of autophagy activators as powerful inducers of cell differentiation with a strong prevalence towards neuronal phenotypes. Thus, mTOR modulation acts on the neurobiology of glioblastoma just like it operates in vivo at the level of brain stem cell niches by altering autophagy-dependent cell differentiation. In the light of such a critical role of autophagy we analyzed the ubiquitin proteasome system. The merging between autophagy and proteasome generates a novel organelle, named autophagoproteasome which is strongly induced by mTOR inhibitors in glioblastoma cells. Remarkably, when mTOR is maximally inhibited the proteasome component selectively moves within autophagy vacuoles, thus making the proteasome activity dependent on the entry within autophagy compartment.

  16. MTOR inhibition attenuates DNA damage and apoptosis through autophagy-mediated suppression of CREB1.

    PubMed

    Wang, Ying; Hu, Zhongdong; Liu, Zhibo; Chen, Rongrong; Peng, Haiyong; Guo, Jing; Chen, Xinxin; Zhang, Hongbing

    2013-12-01

    Hyperactivation of mechanistic target of rapamycin (MTOR) is a common feature of human cancers, and MTOR inhibitors, such as rapamycin, are thus becoming therapeutics in targeting certain cancers. However, rapamycin has also been found to compromise the efficacy of chemotherapeutics to cells with hyperactive MTOR. Here, we show that loss of TSC2 or PTEN enhanced etoposide-induced DNA damage and apoptosis, which was blunted by suppression of MTOR with either rapamycin or RNA interference. cAMP response element-binding protein 1 (CREB1), a nuclear transcription factor that regulates genes involved in survival and death, was positively regulated by MTOR in mouse embryonic fibroblasts (MEFs) and cancer cell lines. Silencing Creb1 expression with siRNA protected MTOR-hyperactive cells from DNA damage-induced apoptosis. Furthermore, loss of TSC2 or PTEN impaired either etoposide or nutrient starvation-induced autophagy, which in turn, leads to CREB1 hyperactivation. We further elucidated an inverse correlation between autophagy activity and CREB1 activity in the kidney tumor tissue obtained from a TSC patient and the mouse livers with hepatocyte-specific knockout of PTEN. CREB1 induced DNA damage and subsequent apoptosis in response to etoposide in autophagy-defective cells. Reactivation of CREB1 or inhibition of autophagy not only improved the efficacy of rapamycin but also alleviated MTOR inhibition-mediated chemoresistance. Therefore, autophagy suppression of CREB1 may underlie the MTOR inhibition-mediated chemoresistance. We suggest that inhibition of MTOR in combination with CREB1 activation may be used in the treatment of cancer caused by an abnormal PI3K-PTEN-AKT-TSC1/2-MTOR signaling pathway. CREB1 activators should potentiate the efficacy of chemotherapeutics in treatment of these cancers.

  17. mTOR Regulates Cellular Iron Homeostasis through Tristetraprolin

    PubMed Central

    Bayeva, Marina; Khechaduri, Arineh; Puig, Sergi; Chang, Hsiang-Chun; Patial, Sonika; Blackshear, Perry J.; Ardehali, Hossein

    2013-01-01

    SUMMARY Iron is an essential cofactor with unique redox properties. Iron regulatory proteins 1 and 2 (IRP1/2) have been established as important regulators of cellular iron homeostasis, but little is known about the role of other pathways in this process. Here we report that the mammalian target of rapamycin (mTOR) regulates iron homeostasis by modulating transferrin receptor 1 (TfR1) stability and altering cellular iron flux. Mechanistic studies identify tristetraprolin (TTP), a protein involved in anti-inflammatory response, as the downstream target of mTOR that binds to and enhances degradation of TfR1 mRNA. We also show that TTP is strongly induced by iron chelation, promotes downregulation of iron-requiring genes in both mammalian and yeast cells, and modulates survival in low-iron states. Taken together, our data uncover a link between metabolic, inflammatory, and iron regulatory pathways, and point towards the existence of a yeast-like TTP-mediated iron conservation program in mammals. PMID:23102618

  18. Ammonia Induces Autophagy through Dopamine Receptor D3 and MTOR.

    PubMed

    Li, Zhiyuan; Ji, Xinmiao; Wang, Wenchao; Liu, Juanjuan; Liang, Xiaofei; Wu, Hong; Liu, Jing; Eggert, Ulrike S; Liu, Qingsong; Zhang, Xin

    2016-01-01

    Hyperammonemia is frequently seen in tumor microenvironments as well as in liver diseases where it can lead to severe brain damage or death. Ammonia induces autophagy, a mechanism that tumor cells may use to protect themselves from external stresses. However, how cells sense ammonia has been unclear. Here we show that culture medium alone containing Glutamine can generate milimolar of ammonia at 37 degrees in the absence of cells. In addition, we reveal that ammonia acts through the G protein-coupled receptor DRD3 (Dopamine receptor D3) to induce autophagy. At the same time, ammonia induces DRD3 degradation, which involves PIK3C3/VPS34-dependent pathways. Ammonia inhibits MTOR (mechanistic target of Rapamycin) activity and localization in cells, which is mediated by DRD3. Therefore, ammonia has dual roles in autophagy: one to induce autophagy through DRD3 and MTOR, the other to increase autophagosomal pH to inhibit autophagic flux. Our study not only adds a new sensing and output pathway for DRD3 that bridges ammonia sensing and autophagy induction, but also provides potential mechanisms for the clinical consequences of hyperammonemia in brain damage, neurodegenerative diseases and tumors.

  19. Meta-analysis of long-term clinical outcomes of everolimus-eluting stents.

    PubMed

    Toyota, Toshiaki; Shiomi, Hiroki; Morimoto, Takeshi; Kimura, Takeshi

    2015-07-15

    The superiority of everolimus-eluting stents (EES) over sirolimus-eluting stents (SES) for long-term clinical outcomes has not been yet firmly established. We conducted a systematic review and a meta-analysis of randomized controlled trials (RCTs) comparing EES directly with SES using the longest available follow-up data. We searched PubMed, the Cochrane database, and ClinicalTrials.gov for RCTs comparing outcomes between EES and SES and identified 13,434 randomly assigned patients from 14 RCTs. EES was associated with significantly lower risks than SES for definite stent thrombosis (ST), definite/probable ST, target-lesion revascularization (TLR), and major adverse cardiac events (MACE). The risks for all-cause death and myocardial infarction were similar between EES and SES. By the stratified analysis according to the timing after stent implantation, the favorable trend of EES relative to SES for ST, TLR, and MACE was consistently observed both within and beyond 1 year. The lower risk of EES relative to SES for MACE beyond 1 year was statistically significant (pooled odds ratio 0.77, 95% confidence interval 0.61 to 0.96, p = 0.02). In conclusion, the current meta-analysis of 14 RCTs directly comparing EES with SES suggested that EES provided improvement in both safety and efficacy; EES compared with SES was associated with significantly lower risk for definite ST, definite/probable ST, TLR, and MACE. The direction and magnitude of the effect beyond 1 year were comparable with those observed within 1 year.

  20. Response to everolimus is seen in TSC-associated SEGAs and angiomyolipomas independent of mutation type and site in TSC1 and TSC2.

    PubMed

    Kwiatkowski, David J; Palmer, Michael R; Jozwiak, Sergiusz; Bissler, John; Franz, David; Segal, Scott; Chen, David; Sampson, Julian R

    2015-12-01

    Tuberous sclerosis complex is an autosomal dominant disorder that occurs owing to inactivating mutations in either TSC1 or TSC2. Tuberous sclerosis complex-related tumors in the brain, such as subependymal giant cell astrocytoma, and in the kidney, such as angiomyolipoma, can cause significant morbidity and mortality. Recently, randomized clinical trials (EXIST-1 and EXIST-2) of everolimus for each of these tuberous sclerosis complex-associated tumors demonstrated the benefit of this drug, which blocks activated mammalian target of rapamycin complex 1. Here we report on the spectrum of mutations seen in patients treated during these trials and the association between mutation and response. TSC2 mutations were predominant among patients in both trials and were present in nearly all subjects with angiomyolipoma in whom a mutation was identified (97%), whereas TSC1 mutations were rare in those subjects (3%). The spectrum of mutations seen in each gene was similar to those previously reported. In both trials, there was no apparent association between mutation type or location within each gene and response to everolimus. Everolimus responses were also seen at a similar frequency for the 16-18% of patients in each trial in whom no mutation in either gene was identified. These observations confirm the strong association between TSC2 mutation and angiomyolipoma burden seen in previous studies, and they indicate that everolimus response occurs regardless of mutation type or location or when no mutation in TSC1 or TSC2 has been identified.

  1. Longevity Pathways (mTOR, SIRT, Insulin/IGF-1) as Key Modulatory Targets on Aging and Neurodegeneration.

    PubMed

    Mazucanti, Caio Henrique; Cabral-Costa, João Victor; Vasconcelos, Andrea Rodrigues; Andreotti, Diana Zukas; Scavone, Cristoforo; Kawamoto, Elisa Mitiko

    2015-01-01

    Recent data from epidemiologic studies have shown that the majority of the public health costs are related to age-related disorders, and most of these diseases can lead to neuronal death. The specific signaling mechanisms underpinning neurodegeneration and aging are incompletely understood. Much work has been directed to the search for the etiology of neurodegeneration and aging and to new therapeutic strategies, including not only drugs but also non-pharmacological approaches, such as physical exercise and low-calorie dietary intake. The most important processes in aging-associated conditions, including neurodegeneration, include the mammalian (or mechanistic target of rapamycin (mTOR, sirtuin (SIRT and insulin/insulin growth factor 1 signaling (IIS pathways. These longevity pathways are involved in an array of different processes, including metabolism, cognition, stress response and brain plasticity. In this review we focus on the current advances involving the mTOR, SIRT and IIS longevity pathways during the course of healthy aging processes and neurodegenerative diseases, bringing new insights in the form of a better understanding of the signaling mechanisms underpinning neurodegeneration and how these differ from physiological normal aging processes. This also provides new targets for the therapeutic management and/or prevention of these devastating age-related disorders.

  2. A Phase II Study of BEZ235 in Patients with Everolimus-resistant, Advanced Pancreatic Neuroendocrine Tumours

    PubMed Central

    FAZIO, NICOLA; BUZZONI, ROBERTO; BAUDIN, ERIC; ANTONUZZO, LORENZO; HUBNER, RICHARD A.; LAHNER, HARALD; DE HERDER, WOUTER W.; RADERER, MARKUS; TEULÉ, ALEXANDRE; CAPDEVILA, JAUME; LIBUTTI, STEVEN K.; KULKE, MATTHEW H.; SHAH, MANISHA; DEY, DEBARSHI; TURRI, SABINE; AIMONE, PAOLA; MASSACESI, CRISTIAN; VERSLYPE, CHRIS

    2016-01-01

    Background This was a two-stage, phase II trial of the dual phosphatidylinositol 3-kinase/mammalian target of rapamycin inhibitor BEZ235 in patients with everolimus-resistant pancreatic neuroendocrine tumours (pNETs) (NCT01658436). Patients and Methods In stage 1, 11 patients received 400 mg BEZ235 orally twice daily (bid). Due to tolerability concerns, a further 20 patients received BEZ235 300 mg bid. Stage 2 would be triggered by a 16-week progression-free survival (PFS) rate of ≥60% in stage 1. Results As of 30 June, 2014, 29/31 patients had discontinued treatment. Treatment-related grade 3/4 adverse events were reported in eight (72.7%) patients at 400 mg and eight (40.0%) patients at 300 mg, including hyperglycaemia, diarrhoea, nausea, and vomiting. The estimated 16-week PFS rate was 51.6% (90% confidence interval=35.7–67.3%). Conclusion BEZ235 was poorly tolerated by patients with everolimus-resistant pNETs at 400 and 300 mg bid doses. Although evidence of disease stability was observed, the study did not proceed to stage 2. PMID:26851029

  3. Germline and somatic mutations in the MTOR gene in focal cortical dysplasia and epilepsy

    PubMed Central

    Møller, Rikke S.; Weckhuysen, Sarah; Chipaux, Mathilde; Marsan, Elise; Taly, Valerie; Bebin, E. Martina; Hiatt, Susan M.; Prokop, Jeremy W.; Bowling, Kevin M.; Mei, Davide; Conti, Valerio; de la Grange, Pierre; Ferrand-Sorbets, Sarah; Dorfmüller, Georg; Lambrecq, Virginie; Larsen, Line H.G.; Leguern, Eric; Guerrini, Renzo; Rubboli, Guido; Cooper, Gregory M.

    2016-01-01

    Objective: To assess the prevalence of somatic MTOR mutations in focal cortical dysplasia (FCD) and of germline MTOR mutations in a broad range of epilepsies. Methods: We collected 20 blood-brain paired samples from patients with FCD and searched for somatic variants using deep-targeted gene panel sequencing. Germline mutations in MTOR were assessed in a French research cohort of 93 probands with focal epilepsies and in a diagnostic Danish cohort of 245 patients with a broad range of epilepsies. Data sharing among collaborators allowed us to ascertain additional germline variants in MTOR. Results: We detected recurrent somatic variants (p.Ser2215Phe, p.Ser2215Tyr, and p.Leu1460Pro) in the MTOR gene in 37% of participants with FCD II and showed histologic evidence for activation of the mTORC1 signaling cascade in brain tissue. We further identified 5 novel de novo germline missense MTOR variants in 6 individuals with a variable phenotype from focal, and less frequently generalized, epilepsies without brain malformations, to macrocephaly, with or without moderate intellectual disability. In addition, an inherited variant was found in a mother–daughter pair with nonlesional autosomal dominant nocturnal frontal lobe epilepsy. Conclusions: Our data illustrate the increasingly important role of somatic mutations of the MTOR gene in FCD and germline mutations in the pathogenesis of focal epilepsy syndromes with and without brain malformation or macrocephaly. PMID:27830187

  4. Regulation of the glutamate transporter EAAT3 by mammalian target of rapamycin mTOR.

    PubMed

    Almilaji, Ahmad; Pakladok, Tatsiana; Guo, Anne; Munoz, Carlos; Föller, Michael; Lang, Florian

    2012-05-04

    The serine/threonine kinase mammalian target of rapamycin (mTOR) is stimulated by insulin, growth factors and nutrients and confers survival of several cell types. The kinase has previously been shown to stimulate amino acid uptake. In neurons, the cellular uptake of glutamate by the excitatory amino-acid transporters (EAATs) decreases excitation and thus confers protection against excitotoxicity. In epithelia, EAAT3 accomplishes transepithelial glutamate and aspartate transport. The present study explored, whether mTOR regulates EAAT3 (SLC1A1). To this end, cRNA encoding EAAT3 was injected into Xenopus oocytes with or without cRNA encoding mTOR and the glutamate induced current (I(glu)), a measure of glutamate transport, determined by dual electrode voltage clamp. Moreover, EAAT3 protein abundance was determined utilizing chemiluminescence. As a result, I(glu) was observed in Xenopus oocytes expressing EAAT3 but not in water injected oocytes. Coexpression of mTOR significantly increased I(glu), an effect reversed by rapamycin (100 nM). mTOR coexpression increased EAAT3 protein abundance in the cell membrane. The decay of I(glu) following inhibition of carrier insertion with brefeldin A in oocytes coexpressing EAAT3 with mTOR was similar in the presence and absence of rapamycin (100 nM). In conclusion, mTOR is a novel powerful regulator of EAAT3 and may thus contribute to protection against neuroexcitotoxicity.

  5. The Role of Phospholipase D in Modulating the MTOR Signaling Pathway in Polycystic Kidney Disease

    PubMed Central

    Liu, Yang; Käch, Andres; Ziegler, Urs; Ong, Albert C. M.; Wallace, Darren P.; Arcaro, Alexandre; Serra, Andreas L.

    2013-01-01

    The mammalian target of rapamycin (mTOR) signaling pathway is aberrantly activated in polycystic kidney disease (PKD). Emerging evidence suggests that phospholipase D (PLD) and its product phosphatidic acid (PA) regulate mTOR activity. In this study, we assessed in vitro the regulatory function of PLD and PA on the mTOR signaling pathway in PKD. We found that the basal level of PLD activity was elevated in PKD cells. Targeting PLD by small molecule inhibitors reduced cell proliferation and blocked mTOR signaling, whereas exogenous PA stimulated mTOR signaling and abolished the inhibitory effect of PLD on PKD cell proliferation. We also show that blocking PLD activity enhanced the sensitivity of PKD cells to rapamycin and that combining PLD inhibitors and rapamycin synergistically inhibited PKD cell proliferation. Furthermore, we demonstrate that targeting mTOR did not induce autophagy, whereas targeting PLD induced autophagosome formation. Taken together, our findings suggest that deregulated mTOR pathway activation is mediated partly by increased PLD signaling in PKD cells. Targeting PLD isoforms with pharmacological inhibitors may represent a new therapeutic strategy in PKD. PMID:24009738

  6. Activation of mTOR: a culprit of Alzheimer’s disease?

    PubMed Central

    Cai, Zhiyou; Chen, Guanghui; He, Wenbo; Xiao, Ming; Yan, Liang-Jun

    2015-01-01

    Alzheimer’s disease (AD) is characterized by cognitive impairment in clinical presentation, and by β-amyloid (Aβ) production and the hyper-phosphorylation of tau in basic research. More highlights demonstrate that the activation of the mammalian target of rapamycin (mTOR) enhances Aβ generation and deposition by modulating amyloid precursor protein (APP) metabolism and upregulating β- and γ-secretases. mTOR, an inhibitor of autophagy, decreases Aβ clearance by scissoring autophagy function. mTOR regulates Aβ generation or Aβ clearance by regulating several key signaling pathways, including phosphoinositide 3-kinase (PI3-K)/protein kinase B (Akt), glycogen synthase kinase 3 [GSK-3], AMP-activated protein kinase (AMPK), and insulin/insulin-like growth factor 1 (IGF-1). The activation of mTOR is also a contributor to aberrant hyperphosphorylated tau. Rapamycin, the inhibitor of mTOR, may mitigate cognitive impairment and inhibit the pathologies associated with amyloid plaques and neurofibrillary tangles by promoting autophagy. Furthermore, the upstream and downstream components of mTOR signaling are involved in the pathogenesis and progression of AD. Hence, inhibiting the activation of mTOR may be an important therapeutic target for AD. PMID:25914534

  7. mTOR Hyperactivation in down syndrome hippocampus appears early during development.

    PubMed

    Iyer, Anand M; van Scheppingen, Jackelien; Milenkovic, Ivan; Anink, Jasper J; Adle-Biassette, Homa; Kovacs, Gabor G; Aronica, Eleonora

    2014-07-01

    The mammalian target of rapamycin (mTOR) signaling pathway is a key developmental pathway involved in mechanisms underlying cellular aging and neurodegeneration. We hypothesized that its deregulation may occur during early brain development in patients with Down syndrome (DS). The expression patterns and cellular distribution of components of mTOR signaling (phosphorylated S6, phosphorylated S6 kinase, phosphorylated eukaryotic initiation factor 4E binding protein 1, and phosphorylated mTOR) were investigated in developing hippocampi from controls and patients with DS and from adults with DS and Alzheimer disease-associated pathology using immunocytochemistry. In control hippocampi, only phosphorylated S6 was detected prenatally (19-41 gestational weeks); it became undetectable 2 months postnatally. Increased expression of phosphorylated S6, phosphorylated S6 kinase, phosphorylated eukaryotic initiation factor 4E binding protein 1, and phosphorylated mTOR was observed in DS hippocampus compared with controls. Phosphorylated S6 and phosphorylated S6 kinase were detected prenatally and persisted throughout postnatal development. Prominent expression of mTOR components was observed in pyramidal neurons with granulovacuolar degeneration and in neurons containing neurofibrillary tangles in the hippocampi of DS subjects with Alzheimer disease pathology. These findings suggest that a dysregulated mTOR pathway may contribute to both early hippocampal developmental abnormalities and hippocampal functional impairment developing before neurodegeneration. Moreover, the expression patterns of mTOR components in adult DS hippocampus support its association with Alzheimer disease-related histopathologic changes.

  8. Caffeine affects the biological responses of human hematopoietic cells of myeloid lineage via downregulation of the mTOR pathway and xanthine oxidase activity

    PubMed Central

    Abooali, Maryam; Yasinska, Inna M.; Casely-Hayford, Maxwell A.; Berger, Steffen M.; Fasler-Kan, Elizaveta; Sumbayev, Vadim V.

    2015-01-01

    Correction of human myeloid cell function is crucial for the prevention of inflammatory and allergic reactions as well as leukaemia progression. Caffeine, a naturally occurring food component, is known to display anti-inflammatory effects which have previously been ascribed largely to its inhibitory actions on phosphodiesterase. However, more recent studies suggest an additional role in affecting the activity of the mammalian target of rapamycin (mTOR), a master regulator of myeloid cell translational pathways, although detailed molecular events underlying its mode of action have not been elucidated. Here, we report the cellular uptake of caffeine, without metabolisation, by healthy and malignant hematopoietic myeloid cells including monocytes, basophils and primary acute myeloid leukaemia mononuclear blasts. Unmodified caffeine downregulated mTOR signalling, which affected glycolysis and the release of pro-inflammatory/pro-angiogenic cytokines as well as other inflammatory mediators. In monocytes, the effects of caffeine were potentiated by its ability to inhibit xanthine oxidase, an enzyme which plays a central role in human purine catabolism by generating uric acid. In basophils, caffeine also increased intracellular cyclic adenosine monophosphate (cAMP) levels which further enhanced its inhibitory action on mTOR. These results demonstrate an important mode of pharmacological action of caffeine with potentially wide-ranging therapeutic impact for treating non-infectious disorders of the human immune system, where it could be applied directly to inflammatory cells. PMID:26384306

  9. Caffeine affects the biological responses of human hematopoietic cells of myeloid lineage via downregulation of the mTOR pathway and xanthine oxidase activity.

    PubMed

    Gibbs, Bernhard F; Gonçalves Silva, Isabel; Prokhorov, Alexandr; Abooali, Maryam; Yasinska, Inna M; Casely-Hayford, Maxwell A; Berger, Steffen M; Fasler-Kan, Elizaveta; Sumbayev, Vadim V

    2015-10-06

    Correction of human myeloid cell function is crucial for the prevention of inflammatory and allergic reactions as well as leukaemia progression. Caffeine, a naturally occurring food component, is known to display anti-inflammatory effects which have previously been ascribed largely to its inhibitory actions on phosphodiesterase. However, more recent studies suggest an additional role in affecting the activity of the mammalian target of rapamycin (mTOR), a master regulator of myeloid cell translational pathways, although detailed molecular events underlying its mode of action have not been elucidated. Here, we report the cellular uptake of caffeine, without metabolisation, by healthy and malignant hematopoietic myeloid cells including monocytes, basophils and primary acute myeloid leukaemia mononuclear blasts. Unmodified caffeine downregulated mTOR signalling, which affected glycolysis and the release of pro-inflammatory/pro-angiogenic cytokines as well as other inflammatory mediators. In monocytes, the effects of caffeine were potentiated by its ability to inhibit xanthine oxidase, an enzyme which plays a central role in human purine catabolism by generating uric acid. In basophils, caffeine also increased intracellular cyclic adenosine monophosphate (cAMP) levels which further enhanced its inhibitory action on mTOR. These results demonstrate an important mode of pharmacological action of caffeine with potentially wide-ranging therapeutic impact for treating non-infectious disorders of the human immune system, where it could be applied directly to inflammatory cells.

  10. Biodegradable-Polymer Biolimus-Eluting Stents versus Durable-Polymer Everolimus-Eluting Stents at One-Year Follow-Up: A Registry-Based Cohort Study.

    PubMed

    Parsa, Ehsan; Saroukhani, Sepideh; Majlessi, Fereshteh; Poorhosseini, Hamidreza; Lofti-Tokaldany, Masoumeh; Jalali, Arash; Salarifar, Mojtaba; Nematipour, Ebrahim; Alidoosti, Mohammad; Aghajani, Hassan; Amirzadegan, Alireza; Kassaian, Seyed Ebrahim

    2016-04-01

    We compared outcomes of percutaneous coronary intervention patients who received biodegradable-polymer biolimus-eluting stents with those who received durable-polymer everolimus-eluting stents. At Tehran Heart Center, we performed a retrospective analysis of the data from January 2007 through December 2011 on 3,270 consecutive patients with coronary artery disease who underwent percutaneous coronary intervention with the biodegradable-polymer biolimus-eluting stent or the durable-polymer everolimus-eluting stent. We excluded patients with histories of coronary artery bypass grafting or percutaneous coronary intervention, acute ST-segment-elevation myocardial infarction, or the implantation of 2 different stent types. Patients were monitored for 12 months. The primary endpoint was a major adverse cardiac event, defined as a composite of death, nonfatal myocardial infarction, and target-vessel and target-lesion revascularization. Durable-polymer everolimus-eluting stents were implanted in 2,648 (81%) and biodegradable-polymer biolimus-eluting stents in 622 (19%) of the study population. There was no significant difference between the 2 groups (2.7% vs 2.7%; P=0.984) in the incidence of major adverse cardiac events. The cumulative adjusted probability of major adverse cardiac events in the biodegradable-polymer biolimus-eluting stent group did not differ from that of such events in the durable-polymer everolimus-eluting stent group (hazard ratio=0.768; 95% confidence interval, 0.421-1.44; P=0.388). We conclude that in our patients the biodegradable-polymer biolimus-eluting stent was as effective and safe, during the 12-month follow-up period, as was the durable-polymer everolimus-eluting stent.

  11. Docosahexaenoic Acid Sensitizes Leukemia Lymphocytes to Barasertib and Everolimus by ROS-dependent Mechanism Without Affecting the Level of ROS and Viability of Normal Lymphocytes.

    PubMed

    Zhelev, Zhivko; Ivanova, Donika; Lazarova, Desislava; Aoki, Ichio; Bakalova, Rumiana; Saga, Tsuneo

    2016-04-01

    The aim of the present study was: (i) to investigate the possibility of sensitizing leukemia lymphocytes to anticancer drugs using docosahexaenoic acid (DHA); (ii) to find combinations with synergistic cytotoxic effect on leukemia lymphocytes, without or with only very low cytotoxicity towards normal lymphocytes; (iii) and to clarify the role of reactive oxygen species (ROS) in the induction of apoptosis and cytotoxicity by such combinations. The study covered 15 anticancer drugs, conventional and new-generation. Well-expressed synergistic cytotoxic effects were observed after treatment of leukemia lymphocytes (Jurkat) with DHA in combination with: barasertib, lonafarnib, everolimus, and palbociclib. We selected two synergistic combinations, DHA with everolimus or barasertib, and investigated their effects on viability of normal lymphocytes, as well as on the production of ROS and induction of apoptosis in both cell lines (leukemia and normal). At the selected concentrations, DHA, everolimus and barasertib (applied separately) were cytotoxic towards leukemia lymphocytes, but not normal lymphocytes. In leukemia cells, the cytotoxicity of combinations was accompanied by strong induction of apoptosis and production of ROS. In normal lymphocytes, drugs alone and in combination with DHA did not affect the level of ROS and did not induce apoptosis. To our knowledge, the present study is the first to report synergistic ROS-dependent cytotoxicity between DHA and new-generation anticancer drugs, such as everolimus and barasertib, that is cancer cell-specific (particularly for acute lymphoblastic leukemia cells Jurkat). These combinations are harmless to normal lymphocytes and do not induce abnormal production of ROS in these cells. The data suggest that DHA could be used as a supplementary component in anticancer chemotherapy, allowing therapeutic doses of everolimus and barasertib to be reduced, minimizing their side-effects.

  12. The Synergistic Effect of Everolimus and Chloroquine on Endothelial Cell Number Reduction Is Paralleled by Increased Apoptosis and Reduced Autophagy Occurrence

    PubMed Central

    Grimaldi, Anna; Balestrieri, Maria Luisa; D'Onofrio, Nunzia; Di Domenico, Gilda; Nocera, Cosimo; Lamberti, Monica; Tonini, Giuseppe; Zoccoli, Alice; Santini, Daniele; Caraglia, Michele; Pantano, Francesco

    2013-01-01

    Endothelial Progenitor Cells (EPCs), a minor subpopulation of the mononuclear cell fraction in peripheral blood, play a critical role in cancer development as they contribute to angiogenesis-mediated pathological neovascularization. In response to tumor cytokines, including VEGF, EPCs mobilize from the bone marrow into the peripheral circulation and move to the tumor bed where they incorporate into sprouting neovessels. In the present study, we evaluated the effects of everolimus (Afinitor, Novartis), a rapamycin analogue, alone or in combination with chloroquine, a 4-alkylamino substituted quinoline family member, one of the autophagy inhibitors, on EPCs biological functions. We found that either everolimus or chloroquine induce growth inhibition on EPCs in a dose-dependent manner after 72 h from the beginning of incubation. The combined administration of the two drugs to EPC was synergistic in inducing growth inhibition; in details, the maximal pharmacological synergism between everolimus and chloroquine in inducing growth inhibition on EPCs cells was recorded when chloroquine was administered 24 h before everolimus. Moreover, we have studied the mechanisms of cell death induced by the two agents alone or in combination on EPCs and we have found that the synergistic effect of combination on EPC growth inhibition was paralleled by increased apoptosis induction and reduced autophagy. These effects occurred together with biochemical features that are typical of reduced autophagic death such as increased co-immunoprecipitation between Beclin 1 and Bcl-2. Chloroquine antagonized the inhibition of the activity of Akt→4EBP1 axis mediated by everolimus and at the same time it blocked the feed-back activation of Erk-1/2 induced by RAD in EPCs. These data suggest a new strategy in order to block angiogenesis in tumours in which this process plays a key role in both the sustainment and spreading of cancer cells. PMID:24244540

  13. [Signaling pathways mTOR and AKT in epilepsy].

    PubMed

    Romero-Leguizamon, C R; Ramirez-Latorre, J A; Mora-Munoz, L; Guerrero-Naranjo, A

    2016-07-01

    Introduccion. La via de señalizacion AKT/mTOR es un eje central en la regulacion celular, especialmente en las enfermedades neurologicas. En la epilepsia, se ha evidenciado su alteracion dentro de su proceso fisiopatologico. Sin embargo, aun no se han descrito todos los mecanismos de estas rutas de señalizacion, las cuales podrian abrir la puerta hacia nuevas investigaciones y estrategias terapeuticas, que finalmente permitan desarrollar tratamientos efectivos en enfermedades neurologicas como la epilepsia. Objetivo. Revisar las asociaciones existentes entre las rutas de señalizacion intracelular de mTOR y AKT en la fisiopatologia de la epilepsia. Desarrollo. La epilepsia es una enfermedad neurologica con un alto impacto epidemiologico en el mundo, por lo cual es de sumo interes la investigacion de los componentes fisiopatologicos que puedan generar nuevos tratamientos farmacologicos. En esta busqueda se han involucrado diferentes rutas de señalizacion intracelular en neuronas, como determinantes epileptogenos. Los avances en esta materia han permitido incluso la implementacion de nuevas estrategias terapeuticas exitosas y que abren el camino hacia nuevas investigaciones. Conclusiones. Mejorar los conocimientos respecto al papel fisiopatologico de la via de señalizacion mTOR/AKT en la epilepsia permite plantear nuevas investigaciones que ofrezcan nuevas alternativas terapeuticas para el tratamiento de la enfermedad. El uso de inhibidores de mTOR ha surgido en los ultimos años como una alternativa eficaz en el tratamiento de algunos tipos de epilepsias, pero es evidente la necesidad de seguir en la busqueda de nuevas terapias farmacologicas involucradas en estas vias de señalizacion.

  14. Twelve-month efficacy and safety of the conversion to everolimus in maintenance heart transplant recipients

    PubMed Central

    Manito, Nicolás; Delgado, Juan F; Crespo-Leiro, María G; Arizón, José María; Segovia, Javier; González-Vílchez, Francisco; Mirabet, Sònia; Lage, Ernesto; Pascual-Figal, Domingo; Díaz, Beatriz; Palomo, Jesús; Rábago, Gregorio; Sanz, Marisa; Blasco, Teresa; Roig, Eulàlia

    2015-01-01

    AIM: To determine the clinical reasons for conversion to everolimus (EVL) and long-term outcomes in heart transplant (HT) recipients. METHODS: A retrospective 12-mo study has been carried out in 14 Spanish centres to assess the efficacy and safety of conversion to EVL in maintenance HT recipients. RESULTS: Two hundred and twenty-two patients were included (mean age: 53 ± 10.5 years; mean time from HT: 8.1 ± 4.5 years). The most common reasons for conversion were nephrotoxicity (30%), chronic allograft vasculopathy (20%) and neoplasms (17%). The doses and mean levels of EVL at baseline (conversion to EVL) and after one year were 1.3 ± 0.3 and 1.2 ± 0.6 mg/d and 6.4 ± 3.4 and 5.6 ± 2.5 ng/mL, respectively. The percentage of patients receiving calcineurin inhibitors (CNIs) at baseline and on the final visit was 95% and 65%, respectively. The doses and mean levels of CNIs decreased between baseline and month 12 from 142.2 ± 51.6 to 98.0 ± 39.4 mg/d (P < 0.001) and from 126.1 ± 50.9 to 89.2 ± 47.7 ng/mL (P < 0.001), respectively, for cyclosporine, and from 2.9 ± 1.8 to 2.6 ± 1.9 mg/d and from 8.3 ± 4.0 to 6.5 ± 2.7 ng/mL (P = 0.011) for tacrolimus. In the subgroup of patients converted because of nephrotoxicity, creatinine clearance increased from 34.9 ± 10.1 to 40.4 ± 14.4 mL/min (P < 0.001). There were 37 episodes of acute rejection in 24 patients (11%). The most frequent adverse events were oedemas (12%), infections (9%) and gastrointestinal problems (6%). EVL was suspended in 44 patients (20%). Since the database was closed at the end of the study, no further follow-up data is available. CONCLUSION: Conversion to EVL in maintenance HT recipients allowed minimisation or suspension of the CNIs, with improved kidney function in the patients with nephrotoxicity, after 12 mo. PMID:26722659

  15. Resveratrol as a novel treatment for diseases with mTOR pathway hyperactivation.

    PubMed

    Alayev, Anya; Berger, Sara Malka; Holz, Marina K

    2015-08-01

    The mechanistic target of rapamycin complex 1 (mTORC1) signaling pathway is hyperactivated in a variety of cancers and tumor syndromes. Therefore, mTORC1 inhibitors are being actively investigated for treatment of neoplasms. The concern with the monotherapy use of mTORC1 inhibitors, such as rapamycin, is that they cause upregulation of autophagy, a cell survival mechanism, and suppress the negative feedback loop to the oncogene Akt. In turn, Akt promotes cell survival, causing the therapy to be partially effective, but relapse occurs upon cessation of treatment. In this review, we describe the current literature on resveratrol as well as our work, which uses rapamycin in combination with resveratrol. We found that this combination treatment efficiently blocked upregulation of autophagy and restored inhibition of Akt in different cancer and tumor models. Interestingly, the combination of rapamycin and resveratrol selectively promoted apoptosis of cells with mTOR pathway hyperactivation. Moreover, this combination prevented tumor growth and lung metastasis when tested in mouse models. Finally, mass spectrometry-based identification of cellular targets of resveratrol provided mechanistic insight into the mode of action of resveratrol. The addition of resveratrol to rapamycin treatment may be a promising option for selective and targeted therapy for diseases with mTORC1 hyperactivation.

  16. Akt and mTOR in B Cell Activation and Differentiation.

    PubMed

    Limon, Jose J; Fruman, David A

    2012-01-01

    Activation of phosphoinositide 3-kinase (PI3K) is required for B cell proliferation and survival. PI3K signaling also controls key aspects of B cell differentiation. Upon engagement of the B cell receptor (BCR), PI3K activation promotes Ca(2+) mobilization and activation of NFκB-dependent transcription, events which are essential for B cell proliferation. PI3K also initiates a distinct signaling pathway involving the Akt and mTOR serine/threonine kinases. It has been generally assumed that activation of Akt and mTOR downstream of PI3K is essential for B cell function. However, Akt and mTOR have complex roles in B cell fate decisions and suppression of this pathway can enhance certain B cell responses while repressing others. In this review we will discuss genetic and pharmacological studies of Akt and mTOR function in normal B cells, and in malignancies of B cell origin.

  17. Adapting the Stress Response: Viral Subversion of the mTOR Signaling Pathway

    PubMed Central

    Le Sage, Valerie; Cinti, Alessandro; Amorim, Raquel; Mouland, Andrew J.

    2016-01-01

    The mammalian target of rapamycin (mTOR) is a central regulator of gene expression, translation and various metabolic processes. Multiple extracellular (growth factors) and intracellular (energy status) molecular signals as well as a variety of stressors are integrated into the mTOR pathway. Viral infection is a significant stress that can activate, reduce or even suppress the mTOR signaling pathway. Consequently, viruses have evolved a plethora of different mechanisms to attack and co-opt the mTOR pathway in order to make the host cell a hospitable environment for replication. A more comprehensive knowledge of different viral interactions may provide fruitful targets for new antiviral drugs. PMID:27231932

  18. Antidepressant action of ketamine via mTOR is mediated by inhibition of nitrergic Rheb degradation

    PubMed Central

    Harraz, Maged M.; Tyagi, Richa; Cortés, Pedro; Snyder, Solomon H.

    2016-01-01

    As traditional antidepressants act only after weeks/months, the discovery that ketamine, an antagonist of glutamate/NMDA receptors, elicits antidepressant actions in hours has been transformative. Its mechanism of action has been elusive, though enhanced mTOR signaling is a major feature. We report a novel signaling pathway wherein NMDA receptor activation stimulates generation of nitric oxide (NO), which S-nitrosylates glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Nitrosylated GAPDH complexes with the ubiquitin-E3-ligase Siah1 and Rheb, a small G protein that activates mTOR. Siah1 degrades Rheb leading to reduced mTOR signaling, while ketamine, conversely, stabilizes Rheb which enhances mTOR signaling. Drugs selectively targeting components of this pathway may offer novel approaches to the treatment of depression. PMID:26782056

  19. DEPTOR-related mTOR suppression is involved in metformin's anti-cancer action in human liver cancer cells

    SciTech Connect

    Obara, Akio; Fujita, Yoshihito; Abudukadier, Abulizi; Fukushima, Toru; Oguri, Yasuo; Ogura, Masahito; Harashima, Shin-ichi; Hosokawa, Masaya; Inagaki, Nobuya

    2015-05-15

    Metformin, one of the most commonly used drugs for patients with type 2 diabetes, recently has received much attention regarding its anti-cancer action. It is thought that the suppression of mTOR signaling is involved in metformin's anti-cancer action. Although liver cancer is one of the most responsive types of cancer for reduction of incidence by metformin, the molecular mechanism of the suppression of mTOR in liver remains unknown. In this study, we investigated the mechanism of the suppressing effect of metformin on mTOR signaling and cell proliferation using human liver cancer cells. Metformin suppressed phosphorylation of p70-S6 kinase, and ribosome protein S6, downstream targets of mTOR, and suppressed cell proliferation. We found that DEPTOR, an endogenous substrate of mTOR suppression, is involved in the suppressing effect of metformin on mTOR signaling and cell proliferation in human liver cancer cells. Metformin increases the protein levels of DEPTOR, intensifies binding to mTOR, and exerts a suppressing effect on mTOR signaling. This increasing effect of DEPTOR by metformin is regulated by the proteasome degradation system; the suppressing effect of metformin on mTOR signaling and cell proliferation is in a DEPTOR-dependent manner. Furthermore, metformin exerts a suppressing effect on proteasome activity, DEPTOR-related mTOR signaling, and cell proliferation in an AMPK-dependent manner. We conclude that DEPTOR-related mTOR suppression is involved in metformin's anti-cancer action in liver, and could be a novel target for anti-cancer therapy. - Highlights: • We elucidated a novel pathway of metformin's anti-cancer action in HCC cells. • DEPTOR is involved in the suppressing effect of metformin on mTOR signaling. • Metformin increases DEPTOR protein levels via suppression of proteasome activity. • DEPTOR-related mTOR suppression is involved in metformin's anti-cancer action.

  20. Mechanisms regulating neuronal excitability and seizure development following mTOR pathway hyperactivation

    PubMed Central

    LaSarge, Candi L.; Danzer, Steve C.

    2014-01-01

    The phosphatidylinositol-3-kinase/phosphatase and tensin homolog (PTEN)-mammalian target of rapamycin (mTOR) pathway regulates a variety of neuronal functions, including cell proliferation, survival, growth, and plasticity. Dysregulation of the pathway is implicated in the development of both genetic and acquired epilepsies. Indeed, several causal mutations have been identified in patients with epilepsy, the most prominent of these being mutations in PTEN and tuberous sclerosis complexes 1 and 2 (TSC1, TSC2). These genes act as negative regulators of mTOR signaling, and mutations lead to hyperactivation of the pathway. Animal models deleting PTEN, TSC1, and TSC2 consistently produce epilepsy phenotypes, demonstrating that increased mTOR signaling can provoke neuronal hyperexcitability. Given the broad range of changes induced by altered mTOR signaling, however, the mechanisms underlying seizure development in these animals remain uncertain. In transgenic mice, cell populations with hyperactive mTOR have many structural abnormalities that support recurrent circuit formation, including somatic and dendritic hypertrophy, aberrant basal dendrites, and enlargement of axon tracts. At the functional level, mTOR hyperactivation is commonly, but not always, associated with enhanced synaptic transmission and plasticity. Moreover, these populations of abnormal neurons can affect the larger network, inducing secondary changes that may explain paradoxical findings reported between cell and network functioning in different models or at different developmental time points. Here, we review the animal literature examining the link between mTOR hyperactivation and epileptogenesis, emphasizing the impact of enhanced mTOR signaling on neuronal form and function. PMID:24672426

  1. MTOR-Driven Metabolic Reprogramming Regulates Legionella pneumophila Intracellular Niche Homeostasis

    PubMed Central

    Abshire, Camille F.; Roy, Craig R.

    2016-01-01

    Vacuolar bacterial pathogens are sheltered within unique membrane-bound organelles that expand over time to support bacterial replication. These compartments sequester bacterial molecules away from host cytosolic immunosurveillance pathways that induce antimicrobial responses. The mechanisms by which the human pulmonary pathogen Legionella pneumophila maintains niche homeostasis are poorly understood. We uncovered that the Legionella-containing vacuole (LCV) required a sustained supply of host lipids during expansion. Lipids shortage resulted in LCV rupture and initiation of a host cell death response, whereas excess of host lipids increased LCVs size and housing capacity. We found that lipids uptake from serum and de novo lipogenesis are distinct redundant supply mechanisms for membrane biogenesis in Legionella-infected macrophages. During infection, the metabolic checkpoint kinase Mechanistic Target of Rapamycin (MTOR) controlled lipogenesis through the Serum Response Element Binding Protein 1 and 2 (SREBP1/2) transcription factors. In Legionella-infected macrophages a host-driven response that required the Toll-like receptors (TLRs) adaptor protein Myeloid differentiation primary response gene 88 (Myd88) dampened MTOR signaling which in turn destabilized LCVs under serum starvation. Inactivation of the host MTOR-suppression pathway revealed that L. pneumophila sustained MTOR signaling throughout its intracellular infection cycle by a process that required the upstream regulator Phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K) and one or more Dot/Icm effector proteins. Legionella-sustained MTOR signaling facilitated LCV expansion and inhibition of the PI3K-MTOR-SREPB1/2 axis through pharmacological or genetic interference or by activation of the host MTOR-suppression response destabilized expanding LCVs, which in turn triggered cell death of infected macrophages. Our work identified a host metabolic requirement for LCV homeostasis and demonstrated that L

  2. Mechanistically distinct cancer-associated mTOR activation clusters predict sensitivity to rapamycin

    PubMed Central

    Xu, Jianing; Pham, Can G.; Albanese, Steven K.; Dong, Yiyu; Lee, Chung-Han; Yao, Zhan; Han, Song; Chen, David; Parton, Daniel L.; Chodera, John D.; Rosen, Neal; Cheng, Emily H.; Hsieh, James J.

    2016-01-01

    Genomic studies have linked mTORC1 pathway–activating mutations with exceptional response to treatment with allosteric inhibitors of mTORC1 called rapalogs. Rapalogs are approved for selected cancer types, including kidney and breast cancers. Here, we used sequencing data from 22 human kidney cancer cases to identify the activating mechanisms conferred by mTOR mutations observed in human cancers and advance precision therapeutics. mTOR mutations that clustered in focal adhesion kinase targeting domain (FAT) and kinase domains enhanced mTORC1 kinase activity, decreased nutrient reliance, and increased cell size. We identified 3 distinct mechanisms of hyperactivation, including reduced binding to DEP domain–containing MTOR-interacting protein (DEPTOR), resistance to regulatory associated protein of mTOR–mediated (RAPTOR-mediated) suppression, and altered kinase kinetics. Of the 28 mTOR double mutants, activating mutations could be divided into 6 complementation groups, resulting in synergistic Rag- and Ras homolog enriched in brain–independent (RHEB-independent) mTORC1 activation. mTOR mutants were resistant to DNA damage–inducible transcript 1–mediated (REDD1-mediated) inhibition, confirming that activating mutations can bypass the negative feedback pathway formed between HIF1 and mTORC1 in the absence of von Hippel–Lindau (VHL) tumor suppressor expression. Moreover, VHL-deficient cells that expressed activating mTOR mutants grew tumors that were sensitive to rapamycin treatment. These data may explain the high incidence of mTOR mutations observed in clear cell kidney cancer, where VHL loss and HIF activation is pathognomonic. Our study provides mechanistic and therapeutic insights concerning mTOR mutations in human diseases. PMID:27482884

  3. Mammalian target of rapamycin (mTor) mediates tau protein dyshomeostasis: implication for Alzheimer disease.

    PubMed

    Tang, Zhi; Bereczki, Erika; Zhang, Haiyan; Wang, Shan; Li, Chunxia; Ji, Xinying; Branca, Rui M; Lehtiö, Janne; Guan, Zhizhong; Filipcik, Peter; Xu, Shaohua; Winblad, Bengt; Pei, Jin-Jing

    2013-05-31

    Previous evidence from post-mortem Alzheimer disease (AD) brains and drug (especially rapamycin)-oriented in vitro and in vivo models implicated an aberrant accumulation of the mammalian target of rapamycin (mTor) in tangle-bearing neurons in AD brains and its role in the formation of abnormally hyperphosphorylated tau. Compelling evidence indicated that the sequential molecular events such as the synthesis and phosphorylation of tau can be regulated through p70 S6 kinase, the well characterized immediate downstream target of mTor. In the present study, we further identified that the active form of mTor per se accumulates in tangle-bearing neurons, particularly those at early stages in AD brains. By using mass spectrometry and Western blotting, we identified three phosphoepitopes of tau directly phosphorylated by mTor. We have developed a variety of stable cell lines with genetic modification of mTor activity using SH-SY5Y neuroblastoma cells as background. In these cellular systems, we not only confirmed the tau phosphorylation sites found in vitro but also found that mTor mediates the synthesis and aggregation of tau, resulting in compromised microtubule stability. Changes of mTor activity cause fluctuation of the level of a battery of tau kinases such as protein kinase A, v-Akt murine thymoma viral oncogene homolog-1, glycogen synthase kinase 3β, cyclin-dependent kinase 5, and tau protein phosphatase 2A. These results implicate mTor in promoting an imbalance of tau homeostasis, a condition required for neurons to maintain physiological function.

  4. TNFAIP3 promotes survival of CD4 T cells by restricting MTOR and promoting autophagy.

    PubMed

    Matsuzawa, Yu; Oshima, Shigeru; Takahara, Masahiro; Maeyashiki, Chiaki; Nemoto, Yasuhiro; Kobayashi, Masanori; Nibe, Yoichi; Nozaki, Kengo; Nagaishi, Takashi; Okamoto, Ryuichi; Tsuchiya, Kiichiro; Nakamura, Tetsuya; Ma, Averil; Watanabe, Mamoru

    2015-01-01

    Autophagy plays important roles in metabolism, differentiation, and survival in T cells. TNFAIP3/A20 is a ubiquitin-editing enzyme that is thought to be a negative regulator of autophagy in cell lines. However, the role of TNFAIP3 in autophagy remains unclear. To determine whether TNFAIP3 regulates autophagy in CD4 T cells, we first analyzed Tnfaip3-deficient naïve CD4 T cells in vitro. We demonstrated that Tnfaip3-deficient CD4 T cells exhibited reduced MAP1LC3/LC3 (microtubule-associated protein 1 light chain 3) puncta formation, increased mitochondrial content, and exaggerated reactive oxygen species (ROS) production. These results indicate that TNFAIP3 promotes autophagy after T cell receptor (TCR) stimulation in CD4 T cells. We then investigated the mechanism by which TNFAIP3 promotes autophagy signaling. We found that TNFAIP3 bound to the MTOR (mechanistic target of rapamycin) complex and that Tnfaip3-deficient cells displayed enhanced ubiquitination of the MTOR complex and MTOR activity. To confirm the effects of enhanced MTOR activity in Tnfaip3-deficient cells, we analyzed cell survival following treatment with Torin1, an MTOR inhibitor. Tnfaip3-deficient CD4 T cells exhibited fewer cell numbers than the control cells in vitro and in vivo. In addition, the impaired survival of Tnfaip3-deficient cells was ameliorated with Torin1 treatment in vitro and in vivo. The effect of Torin1 was abolished by Atg5 deficiency. Thus, enhanced MTOR activity regulates the survival of Tnfaip3-deficient CD4 T cells. Taken together, our findings illustrate that TNFAIP3 restricts MTOR signaling and promotes autophagy, providing new insight into the manner in which MTOR and autophagy regulate survival in CD4 T cells.

  5. Emerging Role of MicroRNAs in mTOR Signaling.

    PubMed

    Zhang, Yanjie; Huang, Bo; Wang, Hui-Yun; Chang, Augustus; Zheng, X F Steven

    2017-02-25

    Mechanistic target of rapamycin (mTOR) is a conserved serine/threonine kinase that plays a critical role in the control of cellular growth and metabolism. Hyperactivation of mTOR pathway is common in human cancers, driving uncontrolled proliferation. MicroRNA (miRNA) is a class of short noncoding RNAs that regulate the expression of a wide variety of genes. Deregulation of miRNAs is a hallmark of cancer. Recent studies have revealed interplays between miRNAs and the mTOR pathway during cancer development. Such interactions appear to provide a fine-tuning of various cellular functions and contribute qualitatively to the behavior of cancer. Here we provide an overview of current knowledge regarding the reciprocal relationship between miRNAs and mTOR pathway: regulation of mTOR signaling by miRNAs and control of miRNA biogenesis by mTOR. Further research in this area may prove important for the diagnosis and therapy of human cancer.

  6. Stem cell-like transcriptional reprogramming mediates metastatic resistance to mTOR inhibition.

    PubMed

    Mateo, F; Arenas, E J; Aguilar, H; Serra-Musach, J; de Garibay, G Ruiz; Boni, J; Maicas, M; Du, S; Iorio, F; Herranz-Ors, C; Islam, A; Prado, X; Llorente, A; Petit, A; Vidal, A; Català, I; Soler, T; Venturas, G; Rojo-Sebastian, A; Serra, H; Cuadras, D; Blanco, I; Lozano, J; Canals, F; Sieuwerts, A M; de Weerd, V; Look, M P; Puertas, S; García, N; Perkins, A S; Bonifaci, N; Skowron, M; Gómez-Baldó, L; Hernández, V; Martínez-Aranda, A; Martínez-Iniesta, M; Serrat, X; Cerón, J; Brunet, J; Barretina, M P; Gil, M; Falo, C; Fernández, A; Morilla, I; Pernas, S; Plà, M J; Andreu, X; Seguí, M A; Ballester, R; Castellà, E; Nellist, M; Morales, S; Valls, J; Velasco, A; Matias-Guiu, X; Figueras, A; Sánchez-Mut, J V; Sánchez-Céspedes, M; Cordero, A; Gómez-Miragaya, J; Palomero, L; Gómez, A; Gajewski, T F; Cohen, E E W; Jesiotr, M; Bodnar, L; Quintela-Fandino, M; López-Bigas, N; Valdés-Mas, R; Puente, X S; Viñals, F; Casanovas, O; Graupera, M; Hernández-Losa, J; Ramón Y Cajal, S; García-Alonso, L; Saez-Rodriguez, J; Esteller, M; Sierra, A; Martín-Martín, N; Matheu, A; Carracedo, A; González-Suárez, E; Nanjundan, M; Cortés, J; Lázaro, C; Odero, M D; Martens, J W M; Moreno-Bueno, G; Barcellos-Hoff, M H; Villanueva, A; Gomis, R R; Pujana, M A

    2016-12-19

    Inhibitors of the mechanistic target of rapamycin (mTOR) are currently used to treat advanced metastatic breast cancer. However, whether an aggressive phenotype is sustained through adaptation or resistance to mTOR inhibition remains unknown. Here, complementary studies in human tumors, cancer models and cell lines reveal transcriptional reprogramming that supports metastasis in response to mTOR inhibition. This cancer feature is driven by EVI1 and SOX9. EVI1 functionally cooperates with and positively regulates SOX9, and promotes the transcriptional upregulation of key mTOR pathway components (REHB and RAPTOR) and of lung metastasis mediators (FSCN1 and SPARC). The expression of EVI1 and SOX9 is associated with stem cell-like and metastasis signatures, and their depletion impairs the metastatic potential of breast cancer cells. These results establish the mechanistic link between resistance to mTOR inhibition and cancer metastatic potential, thus enhancing our understanding of mTOR targeting failure.Oncogene advance online publication, 19 December 2016; doi:10.1038/onc.2016.427.

  7. Hyperactive mTOR pathway promotes lymphoproliferation and abnormal differentiation in autoimmune lymphoproliferative syndrome.

    PubMed

    Völkl, Simon; Rensing-Ehl, Anne; Allgäuer, Andrea; Schreiner, Elisabeth; Lorenz, Myriam Ricarda; Rohr, Jan; Klemann, Christian; Fuchs, Ilka; Schuster, Volker; von Bueren, André O; Naumann-Bartsch, Nora; Gambineri, Eleonora; Siepermann, Kathrin; Kobbe, Robin; Nathrath, Michaela; Arkwright, Peter D; Miano, Maurizio; Stachel, Klaus-Daniel; Metzler, Markus; Schwarz, Klaus; Kremer, Anita N; Speckmann, Carsten; Ehl, Stephan; Mackensen, Andreas

    2016-07-14

    Autoimmune lymphoproliferative syndrome (ALPS) is a human disorder characterized by defective Fas signaling, resulting in chronic benign lymphoproliferation and accumulation of TCRαβ(+) CD4(-) CD8(-) double-negative T (DNT) cells. Although their phenotype resembles that of terminally differentiated or exhausted T cells, lack of KLRG1, high eomesodermin, and marginal T-bet expression point instead to a long-lived memory state with potent proliferative capacity. Here we show that despite their terminally differentiated phenotype, human ALPS DNT cells exhibit substantial mitotic activity in vivo. Notably, hyperproliferation of ALPS DNT cells is associated with increased basal and activation-induced phosphorylation of serine-threonine kinases Akt and mechanistic target of rapamycin (mTOR). The mTOR inhibitor rapamycin abrogated survival and proliferation of ALPS DNT cells, but not of CD4(+) or CD8(+) T cells in vitro. In vivo, mTOR inhibition reduced proliferation and abnormal differentiation by DNT cells. Importantly, increased mitotic activity and hyperactive mTOR signaling was also observed in recently defined CD4(+) or CD8(+) precursor DNT cells, and mTOR inhibition specifically reduced these cells in vivo, indicating abnormal programming of Fas-deficient T cells before the DNT stage. Thus, our results identify the mTOR pathway as a major regulator of lymphoproliferation and aberrant differentiation in ALPS.

  8. Ghrelin regulates GLP-1 production through mTOR signaling in L cells.

    PubMed

    Xu, Geyang; Hong, Xiaosi; Tang, Hong; Jiang, Sushi; Liu, Fenting; Shen, Zhemin; Li, Ziru; Zhang, Weizhen

    2015-11-15

    Glucagon-like peptide (GLP-1), an intestinal incretin produced in L-cells and released in response to meal intake, functions to promote insulin secretion and to decrease plasma glucose. Ghrelin is an orexigenic hormone critical for glucose homeostasis. The molecular mechanism by which ghrelin alters GLP-1 production remains largely unknown. Here we showed that ghrelin attenuates GLP-1 production through mTOR signaling. In GHSR1a null mice, ileal mTOR signaling, proglucagon and circulating GLP-1 were significantly increased. Antagonism of the GHSR1a by D-Lys-3-GHRP-6 increased GLP-1 synthesis and release in STC-1 cells. Treatment of STC-1 cells with ghrelin decreased the production of GLP-1. This effect was associated with a significant inhibition of mTOR signaling. Overexpression of ghrelin inhibited proglucagon promoter activity and GLP-1 production. Inhibition of mTOR activity by mTOR siRNA blocked D-Lys-3-GHRP-6 induced GLP-1 production in STC-1 cells. Our results suggest that mTOR signaling mediates the inhibitory effect of ghrelin on GLP-1 production.

  9. Targeting mTOR in RET mutant medullary and differentiated thyroid cancer cells

    PubMed Central

    Gild, Matti L; Landa, Iñigo; Ryder, Mabel; Ghossein, Ronald A; Knauf, Jeffrey A; Fagin, James A

    2015-01-01

    Inhibitors of RET, a tyrosine kinase receptor encoded by a gene that is frequently mutated in medullary thyroid cancer, have emerged as promising novel therapies for the disease. Rapalogs and other mammalian target of rapamycin (mTOR) inhibitors are effective agents in patients with gastroenteropancreatic neuroendocrine tumors, which share lineage properties with medullary thyroid carcinomas. The objective of this study was to investigate the contribution of mTOR activity to RET-induced signaling and cell growth and to establish whether growth suppression is enhanced by co-targeting RET and mTOR kinase activities. Treatment of the RET mutant cell lines TT, TPC-1, and MZ-CRC-1 with AST487, a RET kinase inhibitor, suppressed growth and showed profound and sustained inhibition of mTOR signaling, which was recapitulated by siRNA-mediated RET knockdown. Inhibition of mTOR with INK128, a dual mTORC1 and mTORC2 kinase inhibitor, also resulted in marked growth suppression to levels similar to those seen with RET blockade. Moreover, combined treatment with AST487 and INK128 at low concentrations suppressed growth and induced apoptosis. These data establish mTOR as a key mediator of RET-mediated cell growth in thyroid cancer cells and provide a rationale for combinatorial treatments in thyroid cancers with oncogenic RET mutations. PMID:23828865

  10. Critical role for hypothalamic mTOR activity in energy balance

    PubMed Central

    Mori, Hiroyuki; Inoki, Ken; Münzberg, Heike; Opland, Darren; Faouzi, Miro; Villanueva, Eneida C.; Ikenoue, Tsuneo; Kwiatkowski, David; MacDougald, Ormond A; Myers, Martin G.; Guan, Kun-Liang

    2009-01-01

    Summary The mammalian target of Rapamycin (mTOR) promotes anabolic cellular processes in response to growth factors and metabolic cues. The TSC1 and TSC2 tumor suppressors are major upstream inhibitory regulators of mTOR signaling. Mice with Rip2/Cre-mediated deletion of Tsc1 (Rip-Tsc1cKO mice) developed hyperphagia and obesity, suggesting that hypothalamic disruption (for which Rip2/Cre is well known) of Tsc1 may dysregulate feeding circuits via mTOR activation. Indeed, Rip-Tsc1cKO mice displayed increased mTOR signaling and enlarged neuron cell size in a number of hypothalamic populations, including Pomc neurons. Furthermore, Tsc1 deletion with Pomc/Cre (Pomc-Tsc1cKO mice) resulted in dysregulation of Pomc neurons and hyperphagic obesity. Treatment with the mTOR inhibitor, rapamycin, ameliorated the hyperphagia, obesity, and the altered Pomc neuronal morphology in developing or adult Pomc-Tsc1cKO mice, and cessation of treatment reinstated these phenotypes. Thus, ongoing mTOR activation in Pomc neurons blocks the catabolic function of these neurons to promote nutrient intake and increased adiposity. PMID:19356717

  11. mTOR activation is critical for betulin treatment in renal cell carcinoma cells.

    PubMed

    Cheng, Wenlong; Ji, Shiqi; Zhang, Haijian; Han, Zhixing; Liu, Qingjun; Wang, Jianwen; Ping, Hao

    2017-01-22

    Betulin, a natural product isolated from the bark of the birch trees, exhibits multiple anticancer effects. Activation of mTOR signaling pathway has been found in numerous cancers, including renal cell carcinoma (RCC). Here, we attempted to study whether mTOR signaling was essential for betulin to treat RCC. Based on cell survival and colony formation assays, we found that mTOR hyperactive RCC cell line 786-O cells were more sensitive to betulin treatment compared with mTOR-inactive Caki-2 cells. Knockdown of TSC2 in Caki-2 cells had similar results to 786-O cells, and mTOR silencing in 786-O cells rescued the inhibitory effect of betulin, indicating that betulin inhibited RCC cell proliferation in an mTOR-dependent manner. Furthermore, betulin treatment decreases the levels of glucose consumption and lactate production in 786-O cells, while minimal effects were observed in Caki-2 cells. In addition, betulin significantly inhibited the expression of PKM2 and HK2 in 786-O cells. Finally, knockdown of PKM2 or HK2 in 786-O reversed the anti-proliferative effects of betulin, and overexpression of PKM2 or HK2 in Caki-2 cells enhanced the sensitivity to betulin treatment. Taken together, these findings demonstrated the critical role of mTOR activation in RCC cells to betulin treatment, suggesting that betulin might be valuable for targeted therapies in RCC patients with mTOR activation.

  12. Photic regulation of the mTOR signaling pathway in the suprachiasmatic circadian clock.

    PubMed

    Cao, Ruifeng; Lee, Boyoung; Cho, Hee-Yeon; Saklayen, Sanjida; Obrietan, Karl

    2008-07-01

    Here we analyzed the light-responsiveness of the mammalian target of rapamycin (mTOR) cascade, a key regulator of inducible translation, in the suprachiasmatic nuclei (SCN), the locus of the master circadian clock. Brief light exposure during the subjective night, but not during the subjective day, triggered rapid phosphorylation (a marker of catalytic activity) of the mTOR translation effectors p70 S6K, ribosomal S6 protein (S6) and 4E-BP1. In the absence of photic stimulation, marked S6 and 4E-BP1 phosphorylation was detected, indicating tonic mTOR activity in the SCN. Light stimulated the colocalized activation of p70 S6K and extracellular signal-regulated protein kinase (ERK), and pharmacological disruption of ERK signaling abolished light-induced mTOR activity, revealing that the MAPK cascade is an essential intermediate that couples light to mTOR. Together these data identify a light-responsive mTOR cascade in the SCN, and thus, raise the possibility that inducible translation contributes to the clock entrainment process.

  13. Modulation of mTOR signaling as a strategy for the treatment of Pompe disease.

    PubMed

    Lim, Jeong-A; Li, Lishu; Shirihai, Orian S; Trudeau, Kyle M; Puertollano, Rosa; Raben, Nina

    2017-03-01

    Mechanistic target of rapamycin (mTOR) coordinates biosynthetic and catabolic processes in response to multiple extracellular and intracellular signals including growth factors and nutrients. This serine/threonine kinase has long been known as a critical regulator of muscle mass. The recent finding that the decision regarding its activation/inactivation takes place at the lysosome undeniably brings mTOR into the field of lysosomal storage diseases. In this study, we have examined the involvement of the mTOR pathway in the pathophysiology of a severe muscle wasting condition, Pompe disease, caused by excessive accumulation of lysosomal glycogen. Here, we report the dysregulation of mTOR signaling in the diseased muscle cells, and we focus on potential sites for therapeutic intervention. Reactivation of mTOR in the whole muscle of Pompe mice by TSC knockdown resulted in the reversal of atrophy and a striking removal of autophagic buildup. Of particular interest, we found that the aberrant mTOR signaling can be reversed by arginine. This finding can be translated into the clinic and may become a paradigm for targeted therapy in lysosomal, metabolic, and neuromuscular diseases.

  14. Mammalian Target of Rapamycin (mTOR) Activation Increases Axonal Growth Capacity of Injured Peripheral Nerves*

    PubMed Central

    Abe, Namiko; Borson, Steven H.; Gambello, Michael J.; Wang, Fan; Cavalli, Valeria

    2010-01-01

    Unlike neurons in the central nervous system (CNS), injured neurons in the peripheral nervous system (PNS) can regenerate their axons and reinnervate their targets. However, functional recovery in the PNS often remains suboptimal, especially in cases of severe damage. The lack of regenerative ability of CNS neurons has been linked to down-regulation of the mTOR (mammalian target of rapamycin) pathway. We report here that PNS dorsal root ganglial neurons (DRGs) activate mTOR following damage and that this activity enhances axonal growth capacity. Furthermore, genetic up-regulation of mTOR activity by deletion of tuberous sclerosis complex 2 (TSC2) in DRGs is sufficient to enhance axonal growth capacity in vitro and in vivo. We further show that mTOR activity is linked to the expression of GAP-43, a crucial component of axonal outgrowth. However, although TSC2 deletion in DRGs facilitates axonal regrowth, it leads to defects in target innervation. Thus, whereas manipulation of mTOR activity could provide new strategies to stimulate nerve regeneration in the PNS, fine control of mTOR activity is required for proper target innervation. PMID:20615870

  15. Targeting molecules to medicine with mTOR, autophagy and neurodegenerative disorders.

    PubMed

    Maiese, Kenneth

    2016-11-01

    Neurodegenerative disorders are significantly increasing in incidence as the age of the global population continues to climb with improved life expectancy. At present, more than 30 million individuals throughout the world are impacted by acute and chronic neurodegenerative disorders with limited treatment strategies. The mechanistic target of rapamycin (mTOR), also known as the mammalian target of rapamycin, is a 289 kDa serine/threonine protein kinase that offers exciting possibilities for novel treatment strategies for a host of neurodegenerative diseases that include Alzheimer's disease, Parkinson's disease, Huntington's disease, epilepsy, stroke and trauma. mTOR governs the programmed cell death pathways of apoptosis and autophagy that can determine neuronal stem cell development, precursor cell differentiation, cell senescence, cell survival and ultimate cell fate. Coupled to the cellular biology of mTOR are a number of considerations for the development of novel treatments involving the fine control of mTOR signalling, tumourigenesis, complexity of the apoptosis and autophagy relationship, functional outcome in the nervous system, and the intimately linked pathways of growth factors, phosphoinositide 3-kinase (PI 3-K), protein kinase B (Akt), AMP activated protein kinase (AMPK), silent mating type information regulation two homologue one (Saccharomyces cerevisiae) (SIRT1) and others. Effective clinical translation of the cellular signalling mechanisms of mTOR offers provocative avenues for new drug development in the nervous system tempered only by the need to elucidate further the intricacies of the mTOR pathway.

  16. mTOR plays critical roles in pancreatic cancer stem cells through specific and stemness-related functions

    NASA Astrophysics Data System (ADS)

    Matsubara, Shyuichiro; Ding, Qiang; Miyazaki, Yumi; Kuwahata, Taisaku; Tsukasa, Koichiro; Takao, Sonshin

    2013-11-01

    Pancreatic cancer is characterized by near-universal mutations in KRAS. The mammalian target of rapamycin (mTOR), which functions downstream of RAS, has divergent effects on stem cells. In the present study, we investigated the significance of the mTOR pathway in maintaining the properties of pancreatic cancer stem cells. The mTOR inhibitor, rapamycin, reduced the viability of CD133+ pancreatic cancer cells and sphere formation which is an index of self-renewal of stem-like cells, indicating that the mTOR pathway functions to maintain cancer stem-like cells. Further, rapamycin had different effects on CD133+ cells compared to cyclopamine which is an inhibitor of the Hedgehog pathway. Thus, the mTOR pathway has a distinct role although both pathways maintain pancreatic cancer stem cells. Therefore, mTOR might be a promising target to eliminate pancreatic cancer stem cells.

  17. The role of phospholipase D and phosphatidic acid in the mechanical activation of mTOR signaling in skeletal muscle.

    PubMed

    Hornberger, T A; Chu, W K; Mak, Y W; Hsiung, J W; Huang, S A; Chien, S

    2006-03-21

    Signaling by the mammalian target of rapamycin (mTOR) has been reported to be necessary for mechanical load-induced growth of skeletal muscle. The mechanisms involved in the mechanical activation of mTOR signaling are not known, but several studies indicate that a unique [phosphotidylinositol-3-kinase (PI3K)- and nutrient-independent] mechanism is involved. In this study, we have demonstrated that a regulatory pathway for mTOR signaling that involves phospholipase D (PLD) and the lipid second messenger phosphatidic acid (PA) plays a critical role in the mechanical activation of mTOR signaling. First, an elevation in PA concentration was sufficient for the activation of mTOR signaling. Second, the isozymes of PLD (PLD1 and PLD2) are localized to the z-band in skeletal muscle (a critical site of mechanical force transmission). Third, mechanical stimulation of skeletal muscle with intermittent passive stretch ex vivo induced PLD activation, PA accumulation, and mTOR signaling. Finally, pharmacological inhibition of PLD blocked the mechanically induced increase in PA and the activation of mTOR signaling. Combined, these results indicate that mechanical stimuli activate mTOR signaling through a PLD-dependent increase in PA. Furthermore, we showed that mTOR signaling was partially resistant to rapamycin in muscles subjected to mechanical stimulation. Because rapamycin and PA compete for binding to the FRB domain on mTOR, these results suggest that mechanical stimuli activate mTOR signaling through an enhanced binding of PA to the FRB domain on mTOR.

  18. mTOR links oncogenic signaling to tumor cell metabolism.

    PubMed

    Yecies, Jessica L; Manning, Brendan D

    2011-03-01

    As a key regulator of cell growth and proliferation, the mammalian target of rapamycin (mTOR) complex 1 (mTORC1) has been the subject of intense investigation for its role in tumor development and progression. This research has revealed a signaling network of oncogenes and tumor suppressors lying upstream of mTORC1, and oncogenic perturbations to this network result in the aberrant activation of this kinase complex in the majority of human cancers. However, the molecular events downstream of mTORC1 contributing to tumor cell growth and proliferation are just coming to light. In addition to its better-known functions in promoting protein synthesis and suppressing autophagy, mTORC1 has emerged as a key regulator of cellular metabolism. Recent studies have found that mTORC1 activation is sufficient to stimulate an increase in glucose uptake, glycolysis, and de novo lipid biosynthesis, which are considered metabolic hallmarks of cancer, as well as the pentose phosphate pathway. Here, we focus on the molecular mechanisms of metabolic regulation by mTORC1 and the potential consequences for anabolic tumor growth and therapeutic strategies.

  19. Influence of cyclosporine and everolimus on the main mycophenolate mofetil pharmacokinetic parameters

    PubMed Central

    Noreikaitė, Aurelija; Saint-Marcoux, Franck; Marquet, Pierre; Kaduševičius, Edmundas; Stankevičius, Edgaras

    2017-01-01

    Abstract The objective of the present study was to assess the effect of cyclosporine (CsA) on the pharmacokinetic parameters of mycophenolic acid (MPA), an active mycophenolate mofetil (MMF) metabolite, and to compare with the effect of everolimus (EVR). Anonymized medical records of 404 kidney recipients were reviewed. The main MPA pharmacokinetic parameters (AUC(0–12) and Cmax) were evaluated. The patients treated with a higher mean dose of CsA displayed higher MPA AUC(0–12) exposure in the low-dose MMF group (1000 mg/day) (40.50 ± 10.97 vs 28.08 ± 11.03 h mg/L; rs = 0.497, P < 0.05), medium-dose MMF group (2000 mg/day) (43.00 ± 6.27 vs 28.85 ± 11.08 h mg/L; rs = 0.437, P < 0.01), and high-dose MMF group (3000 mg/day) (56.75 ± 16.78 vs 36.20 ± 3.70 h mg/L; rs = 0.608, P < 0.05). A positive correlation was also observed between the mean CsA dose and the MPA Cmax in the low-dose MMF group (Cmax 22.83 ± 10.82 vs 12.08 ± 5.59 mg/L; rs = 0.507, P < 0.05) and in the medium-dose MMF group (22.77 ± 8.86 vs 13.00 ± 6.82 mg/L; rs = 0.414, P < 0.01). The comparative analysis between 2 treatment arms (MMF + CsA and MMF + EVR) showed that MPA AUC(0–12) exposure was by 43% higher in the patients treated with a medium dose of MMF and EVR than in the patients treated with a medium dose of MMF and CsA. The data of the present study suggest a possible CsA versus EVR influence on MMF pharmacokinetics. Study results show that CsA has an impact on the main MPA pharmacokinetic parameters (AUC(0–12) and Cmax) in a CsA dose-related manner, while EVR mildly influence or does not affect MPA pharmacokinetic parameters. Low-dose CsA (lower than 180 mg/day) reduces MPA AUC(0–12) exposure under the therapeutic window and may lead to ineffective therapy, while a high-dose CsA (>240 mg/day) is related to greater than 10 mg/L MPA Cmax and increases the

  20. Influence of cyclosporine and everolimus on the main mycophenolate mofetil pharmacokinetic parameters: Cross-sectional study.

    PubMed

    Noreikaitė, Aurelija; Saint-Marcoux, Franck; Marquet, Pierre; Kaduševičius, Edmundas; Stankevičius, Edgaras

    2017-03-01

    The objective of the present study was to assess the effect of cyclosporine (CsA) on the pharmacokinetic parameters of mycophenolic acid (MPA), an active mycophenolate mofetil (MMF) metabolite, and to compare with the effect of everolimus (EVR).Anonymized medical records of 404 kidney recipients were reviewed. The main MPA pharmacokinetic parameters (AUC(0-12) and Cmax) were evaluated.The patients treated with a higher mean dose of CsA displayed higher MPA AUC(0-12) exposure in the low-dose MMF group (1000 mg/day) (40.50 ± 10.97 vs 28.08 ± 11.03 h mg/L; rs = 0.497, P < 0.05), medium-dose MMF group (2000 mg/day) (43.00 ± 6.27 vs 28.85 ± 11.08 h mg/L; rs = 0.437, P < 0.01), and high-dose MMF group (3000 mg/day) (56.75 ± 16.78 vs 36.20 ± 3.70 h mg/L; rs = 0.608, P < 0.05).A positive correlation was also observed between the mean CsA dose and the MPA Cmax in the low-dose MMF group (Cmax 22.83 ± 10.82 vs 12.08 ± 5.59 mg/L; rs = 0.507, P < 0.05) and in the medium-dose MMF group (22.77 ± 8.86 vs 13.00 ± 6.82 mg/L; rs = 0.414, P < 0.01).The comparative analysis between 2 treatment arms (MMF + CsA and MMF + EVR) showed that MPA AUC(0-12) exposure was by 43% higher in the patients treated with a medium dose of MMF and EVR than in the patients treated with a medium dose of MMF and CsA.The data of the present study suggest a possible CsA versus EVR influence on MMF pharmacokinetics. Study results show that CsA has an impact on the main MPA pharmacokinetic parameters (AUC(0-12) and Cmax) in a CsA dose-related manner, while EVR mildly influence or does not affect MPA pharmacokinetic parameters. Low-dose CsA (lower than 180 mg/day) reduces MPA AUC(0-12) exposure under the therapeutic window and may lead to ineffective therapy, while a high-dose CsA (>240 mg/day) is related to greater than 10 mg/L MPA Cmax and increases the likelihood of adverse events.

  1. Oral everolimus treatment in a preterm infant with multifocal inoperable cardiac rhabdomyoma associated with tuberous sclerosis complex and a structural heart defect.

    PubMed

    Mohamed, Ibrahim; Ethier, Guillaume; Goyer, Isabelle; Major, Philippe; Dahdah, Nagib

    2014-11-26

    Rhabdomyoma (RHM) is a benign cardiac tumour usually associated with tuberous sclerosis complex (TSC). Most RHMs are asymptomatic and regress spontaneously during the first years of life. Haemodynamically significant RHMs are classically treated with surgical excision. We present a case of a premature infant, born to a mother having TSC, with a prenatal diagnosis of pulmonary valve atresia and a large ventricular septal defect. Multiple cardiac RHMs were also present, including a large tumour affecting the right ventricular filling. Owing to the prematurity and low birth weight, the infant was inoperable. In this report, we describe our approach to pharmacologically reduce the RHM size using oral everolimus in preparation for a two-ventricle surgical repair of the structural cardiac defect. We also specifically describe the dose of everolimus that was used in this case to achieve therapeutic serum levels, which was seven times lower than the conventional dose applicable for older infants.

  2. Proangiogenic stimulation of bone marrow endothelium engages mTOR and is inhibited by simultaneous blockade of mTOR and NF-κB

    PubMed Central

    Costa, Lara F.; Balcells, Mercedes; Edelman, Elazer R.; Nadler, Lee M.; Cardoso, Angelo A.

    2006-01-01

    Most bone marrow (BM) malignancies develop in association with an angiogenic phenotype and increased numbers of endothelial cells. The molecular mechanisms involved in the modulation and recruitment of BM endothelium are largely unknown and may provide novel therapeutic targets for neoplastic diseases. We observed that angiogenic stimulation of BM endothelial cells activates mTOR and engages its downstream pathways 4E-BP1 and S6K1, which are inhibited by the mTOR-specific blockers rapamycin and CCI-779. Both mTOR blockers significantly inhibit growth factor- and leukemia-induced proliferation of BM endothelium by inducing G0/G1 cell-cycle arrest. This effect is associated with down-regulation of cyclin D1 and cdk2 phosphorylation, and up-regulation of the cdk inhibitors p27kip1 and p21cip1. Under conditions that reproduce the biomechanical fluidic environment of the BM, CCI-779 is equally effective in inhibiting BM endothelial-cell proliferation. Finally, simultaneous blockade of mTOR and NF-κB pathways synergize to significantly inhibit or abrogate the proliferative responses of BM endothelial cells to mitogenic stimuli. This study identifies mTOR as an important pathway for the proangiogenic stimulation of BM endothelium. Modulation of this pathway may serve as a valid therapeutic intervention in BM malignancies evolving in association with an angiogenic phenotype. PMID:16141350

  3. GADD34 Keeps the mTOR Pathway Inactivated in Endoplasmic Reticulum Stress Related Autophagy

    PubMed Central

    Holczer, Marianna; Bánhegyi, Gábor; Kapuy, Orsolya

    2016-01-01

    The balance of protein synthesis and proteolysis (i.e. proteostasis) is maintained by a complex regulatory network in which mTOR (mechanistic target of rapamycin serine/threonine kinase) pathway and unfolded protein response are prominent positive and negative actors. The interplay between the two systems has been revealed; however the mechanistic details of this crosstalk are largely unknown. The aim of the present study was to investigate the elements of crosstalk during endoplasmic reticulum stress and to verify the key role of GADD34 in the connection with the mTOR pathway. Here, we demonstrate that a transient activation of autophagy is present in endoplasmic reticulum stress provoked by thapsigargin or tunicamycin, which is turned into apoptotic cell death. The transient phase can be characterized by the elevation of the autophagic marker LC3II/I, by mTOR inactivation, AMP-activated protein kinase activation and increased GADD34 level. The switch from autophagy to apoptosis is accompanied with the appearance of apoptotic markers, mTOR reactivation, AMP-activated protein kinase inactivation and a decrease in GADD34. Inhibition of autophagy by 3-methyladenine shortens the transient phase, while inhibition of mTOR by rapamycin or resveratrol prolongs it. Inhibition of GADD34 by guanabenz or transfection of the cells with siGADD34 results in down-regulation of autophagy-dependent survival and a quick activation of mTOR, followed by apoptotic cell death. The negative effect of GADD34 inhibition is diminished when guanabenz or siGADD34 treatment is combined with rapamycin or resveratrol addition. These data confirm that GADD34 constitutes a mechanistic link between endoplasmic reticulum stress and mTOR inactivation, therefore promotes cell survival during endoplasmic reticulum stress. PMID:27992581

  4. Potential impact of mTOR inhibitors on cervical squamous cell carcinoma: A systematic review

    PubMed Central

    Assad, Daniele Xavier; Elias, Silvia Taveira; Melo, Andréia Cristina; Ferreira, Carlos Gil; De Luca Canto, Graziela; Guerra, Eliete Neves Silva

    2016-01-01

    The aim of the present systematic review was to analyze the potential impact of mammalian target of rapamycin (mTOR) inhibitors on the treatment of cervical squamous cell carcinoma (CSCC). A systematic literature search was conducted in PubMed, PMC, Scopus, Cochrane Library, LILACS, Web of Science, Google Scholar and ScienceDirect on January 19, 2015, without time and language restrictions. Studies that evaluated women of any age with CSCC and who received mTOR inhibitors alone or in association with other treatments were considered. Randomized and non-randomized clinical trials were included, and the Preferred Reporting Items for Systematic Reviews and Meta-Analyses checklist was followed. Selected studies were methodologically appraised according to the Grades of Recommendation, Assessment, Development and Evaluation method to assess the quality of evidence. Of 642 identified citations, 43 studies were fully reviewed; however, only 3 studies met the inclusion criteria and were used for qualitative analysis. Of these, two studies were phase 1 and one was a phase 2 clinical trial. The studies included were not conclusive with regard to the association between mTOR inhibitor treatment and cervical cancer. The main analysis of secondary endpoints revealed that individuals treated with other drugs in association with mTOR inhibitors achieved partial responses (15.4–33.3%) or stable disease (17.6–28%). Treatment with mTOR inhibitors in general was well tolerated in patients with metastatic disease. The predominant toxicities were grade 1 and 2. The phase 1 trials included in this review demonstrated that mTOR inhibitor treatments are feasible and safe. However, the currently available evidence is insufficient to determine the effect of mTOR inhibitors on CSCC, and further investigation in high-quality, randomized clinical trials is required. PMID:27895779

  5. mTOR disruption causes intestinal epithelial cell defects and intestinal atrophy postinjury in mice

    PubMed Central

    Sampson, Leesa L.; Davis, Ashley K.; Grogg, Matthew W.; Zheng, Yi

    2016-01-01

    Intestinal stem cells (ISCs) drive small intestinal epithelial homeostasis and regeneration. Mechanistic target of rapamycin (mTOR) regulates stem and progenitor cell metabolism and is frequently dysregulated in human disease, but its physiologic functions in the mammalian small intestinal epithelium remain poorly defined. We disrupted the genes mTOR, Rptor, Rictor, or both Rptor and Rictor in mouse ISCs, progenitors, and differentiated intestinal epithelial cells (IECs) using Villin-Cre. Mutant tissues and wild-type or heterozygous littermate controls were analyzed by histologic immunostaining, immunoblots, and proliferation assays. A total of 10 Gy irradiation was used to injure the intestinal epithelium and induce subsequent crypt regeneration. We report that mTOR supports absorptive enterocytes and secretory Paneth and goblet cell function while negatively regulating chromogranin A-positive enteroendocrine cell number. Through additional Rptor, Rictor, and Rptor/Rictor mutant mouse models, we identify mechanistic target of rapamycin complex 1 as the major IEC regulatory pathway, but mechanistic target of rapamycin complex 2 also contributes to ileal villus maintenance and goblet cell size. Homeostatic adult small intestinal crypt cell proliferation, survival, and canonical wingless-int (WNT) activity are not mTOR dependent, but Olfm4+ ISC/progenitor population maintenance and crypt regeneration postinjury require mTOR. Overall, we conclude that mTOR regulates multiple IEC lineages and promotes stem and progenitor cell activity during intestinal epithelium repair postinjury.—Sampson, L. L., Davis, A. K., Grogg, M. W., Zheng, Y. mTOR disruption causes intestinal epithelial cell defects and intestinal atrophy postinjury in mice. PMID:26631481

  6. Clinical activity of everolimus in relapsed/refractory marginal zone B-cell lymphomas: results of a phase II study of the International Extranodal Lymphoma Study Group.

    PubMed

    Conconi, Annarita; Raderer, Markus; Franceschetti, Silvia; Devizzi, Liliana; Ferreri, Andrés J M; Magagnoli, Massimo; Arcaini, Luca; Zinzani, Pier Luigi; Martinelli, Giovanni; Vitolo, Umberto; Kiesewetter, Barbara; Porro, Elena; Stathis, Anastasios; Gaidano, Gianluca; Cavalli, Franco; Zucca, Emanuele

    2014-07-01

    The International Extranodal Lymphoma Study Group coordinated a phase II trial to evaluate the activity and safety of everolimus in marginal zone lymphomas (MZLs). Thirty patients with relapsed/refractory MZLs received everolimus for six cycles or until dose-limiting toxicity or progression. Median age was 71 years (range, 51-88 years). Twenty patients had extranodal, six splenic, four nodal MZL. Twenty-four patients had stage III-IV. Median number of prior therapies was two (range 1-5). Seventeen patients had early treatment discontinuation, in most cases due to toxicity. Median number of cycles was 4.5 (range, 1-16). Among the 24 assessable patients, the overall response rate (ORR) was 25% (95% confidence interval: 10-47). Grade 3-4 adverse events were neutropenia and thrombocytopenia (17% of patients, each), infections (17%), mucositis and odontogenic infections (13%) and lung toxicity (3%). The median response duration was 6.8 months (range, 1.4-11.1+). After a median follow-up of 14.5 months, five deaths were reported: four deaths were due to lymphoma, one was due to toxicity. In an intent-to-treat analysis, the projected median progression-free survival was 14 months. The moderate antitumour activity of everolimus in relapsed/refractory MZLs and the observed toxicity limit its therapeutical applicability in these indolent entities. Lower doses of the drug and, perhaps, different strategies including combination with additional agents need to be explored.

  7. Use of a small particle solid-core packing for improved efficiency and rapid measurement of sirolimus and everolimus by LC-MS/MS.

    PubMed

    Morgan, Phillip; Nwafor, Magnus; Tredger, Mike

    2016-06-01

    Measurement of whole blood sirolimus and everolimus is required in order to optimize patient treatment following solid organ transplant. Assay by LC-MS/MS is increasingly preferred; however efficient use of the instrument and short turnaround times are crucial. Use of a 1.6 µm solid-core packing HPLC column (Cortecs) gave significant increases in efficiency, sensitivity and throughput compared with an existing method, following simple protein precipitation of small-volume (20 μL) whole blood samples. Sirolimus, everolimus and the stable isotopic internal standard ((13) C2 D4 - everolimus) eluted at around 0.8 min, and total analytical run time was 2.2 min, saving almost 4 min per sample compared with an existing method. Within-assay imprecision (CV) was 3.3-8.5%, and between-assay imprecision was 2.2-10.8%. Retrospective assay of external quality assurance samples and comparison of patient samples assayed in parallel showed only small differences (between +6.8 and -1.9%) in results using the Cortecs column when compared with the existing method. No significant interferences or ion suppression were observed. Copyright © 2015 John Wiley & Sons, Ltd.

  8. Inhibition of mTOR kinase via rapamycin blocks persistent predator stress-induced hyperarousal.

    PubMed

    Fifield, Kathleen; Hebert, Mark; Angel, Rebecca; Adamec, Robert; Blundell, Jacqueline

    2013-11-01

    Traumatic, stressful life events are thought to trigger acquired anxiety disorders such as post-traumatic stress disorder (PTSD). Recent data suggests that the mammalian target of rapamycin (mTOR) plays a key role in the formation of traumatic memories. The predator stress paradigm allows us to determine whether mTOR mediates the formation of both context-dependent (associative) and context-independent (non-associative) fear memories. Predator stress involves an acute, unprotected exposure of a rat to a cat which causes long-lasting non-associative fear memories manifested as generalized hyperarousal and increased anxiety-like behavior. Here, we show that rapamycin, an mTOR inhibitor, attenuates predator stress-induced hyperarousal, lasting at least three weeks. In addition, rapamycin blocks a subset of anxiety-like behaviors as measured in the elevated plus maze and hole board. Furthermore, when re-exposed to the predator stress context, rapamycin-treated stressed rats showed increased activity compared to vehicle controls suggesting that rapamycin blocks predator stress-induced associative fear memory. Taken together with past research, our results indicate that mTOR regulation of protein translation is required for the formation of both associative and non-associative fear memories. Overall, these data suggest that mTOR activation may contribute to the development of acquired anxiety disorders such as PTSD.

  9. Distinct amino acid-sensing mTOR pathways regulate skeletal myogenesis.

    PubMed

    Yoon, Mee-Sup; Chen, Jie

    2013-12-01

    Signaling through the mammalian target of rapamycin (mTOR) in response to amino acid availability controls many cellular and developmental processes. mTOR is a master regulator of myogenic differentiation, but the pathways mediating amino acid signals in this process are not known. Here we examine the Rag GTPases and the class III phosphoinositide 3-kinase (PI3K) Vps34, two mediators of amino acid signals upstream of mTOR complex 1 (mTORC1) in cell growth regulation, for their potential involvement in myogenesis. We find that, although both Rag and Vps34 mediate amino acid activation of mTORC1 in C2C12 myoblasts, they have opposing functions in myogenic differentiation. Knockdown of RagA/B enhances, whereas overexpression of active RagB/C mutants impairs, differentiation, and this inhibitory function of Rag is mediated by mTORC1 suppression of the IRS1-PI3K-Akt pathway. On the other hand, Vps34 is required for myogenic differentiation. Amino acids activate a Vps34-phospholipase D1 (PLD1) pathway that controls the production of insulin-like growth factor II, an autocrine inducer of differentiation, through the Igf2 muscle enhancer. The product of PLD, phosphatidic acid, activates the enhancer in a rapamycin-sensitive but mTOR kinase-independent manner. Our results uncover amino acid-sensing mechanisms controlling the homeostasis of myogenesis and underline the versatility and context dependence of mTOR signaling.

  10. Role of HHV-8 and mTOR pathway in post-transplant Kaposi sarcoma staging.

    PubMed

    Hernández-Sierra, Astrid; Rovira, Jordi; Petit, Anna; Moya-Rull, Daniel; Mazuecos, María Auxiliadora; Sánchez-Fructuoso, Ana Isabel; Errasti, Pedro; Idoate, Miguel Ángel; Cruzado, Josep María; Vidal, August; Diekmann, Fritz; Oppenheimer, Federico; Campistol, Josep M; Revuelta, Ignacio

    2016-09-01

    Kaposi's sarcoma (KS) is one of the most frequent transplant related tumors. Several pathways are involved; however, the impact of the molecular phenotype associated to the tumor stage and the behavior-depending resultant therapy is still unknown. The aim of our study was to analyze the role of HHV-8 and mTOR pathway in tumor stages of skin KS after renal transplantation. Twelve renal transplant recipients with cutaneous KS from five transplant centers (1980-2007) under reduction of immunosuppression or conversion to mTOR inhibitor were included. The expression of HHV-8, PTEN, TGFβ, VEGF, phospho-mTOR, and phospho-P70S6K in tumoral tissue was analyzed. KS lesions were classified as patch, plaque, and nodule state. HHV-8 infection was found in all tissue samples. KS lesions showed high activation of VEGF, p-mTOR and p-P70S6K, low PTEN, and null TGFβ expression. The only pathway activated in a staging-dependent manner was mTOR with higher p-mTOR and p-P70S6K expression in nodule versus patch stage. KS lesions disappeared after 5.24 months in all converted patients without any recurrence in 14.05 years of mean follow-up. The activation of mTOR pathway according to KS stages supports the rational of the mTOR inhibitor in post-transplant Kaposi.

  11. mTOR has distinct functions in generating versus sustaining humoral immunity

    PubMed Central

    Jones, Derek D.; Gaudette, Brian T.; Wilmore, Joel R.; Chernova, Irene; Bortnick, Alexandra; Weiss, Brendan M.; Allman, David

    2016-01-01

    Little is known about the role of mTOR signaling in plasma cell differentiation and function. Furthermore, for reasons not understood, mTOR inhibition reverses antibody-associated disease in a murine model of systemic lupus erythematosus. Here, we have demonstrated that induced B lineage–specific deletion of the gene encoding RAPTOR, an essential signaling adaptor for rapamycin-sensitive mTOR complex 1 (mTORC1), abrogated the generation of antibody-secreting plasma cells in mice. Acute treatment with rapamycin recapitulated the effects of RAPTOR deficiency, and both strategies led to the ablation of newly formed plasma cells in the spleen and bone marrow while also obliterating preexisting germinal centers. Surprisingly, although perturbing mTOR activity caused a profound decline in serum antibodies that were specific for exogenous antigen or DNA, frequencies of long-lived bone marrow plasma cells were unaffected. Instead, mTORC1 inhibition led to decreased expression of immunoglobulin-binding protein (BiP) and other factors needed for robust protein synthesis. Consequently, blockade of antibody synthesis was rapidly reversed after termination of rapamycin treatment. We conclude that mTOR signaling plays critical but diverse roles in early and late phases of antibody responses and plasma cell differentiation. PMID:27760048

  12. Prolonged autophagy by MTOR inhibitor leads radioresistant cancer cells into senescence.

    PubMed

    Nam, Hae Yun; Han, Myung Woul; Chang, Hyo Won; Kim, Sang Yoon; Kim, Seong Who

    2013-10-01

    Radiotherapy is one of the well-established therapeutic modalities for cancer treatment. However, the emergence of cells refractory to radiation is a major obstacle to successful treatment with radiotherapy. Many reports suggest that inhibitors targeting the mechanistic target of rapamycin (MTOR) can sensitize cancer cells to the effect of radiation, although by which mechanism MTOR inhibitors enhance the efficacy of radiation toward cancer cells remains to be elucidated. Our studies indicate that a potent and persistent activation of autophagy via inhibition of the MTOR pathway, even in cancer cells where autophagy is occurring, can trigger premature senescence, cellular proliferation arrest. Combined treatment of MTOR inhibitor and radiation induce heterochromatin formation, an irreversible growth arrest and an increase of senescence-associated GLB1 (β-galactosidase) activity, which appear to result from a constant activation of TP53 and a restoration in the activity of retinoblastoma 1 protein (RB1)-E2F1. Thus, this study provides evidence that promoting cellular senescence via inhibition of the MTOR pathway may serve as an avenue to augment radiosensitivity in cancer cells that initiate an autophagy-survival mode to radiotherapy.

  13. MHY1485 activates mTOR and protects osteoblasts from dexamethasone.

    PubMed

    Zhao, Sai; Chen, Caiyun; Wang, Shouguo; Ji, Feng; Xie, Yue

    2016-12-09

    Dexamethasone (Dex) exerts cytotoxic effects to cultured osteoblasts. The potential effect of MHY1485, a small-molecular mammalian target of rapamycin (mTOR) activator, against the process was studied here. In both osteoblastic MC3T3-E1 cells and primary murine osteoblasts, treatment with MHY1485 significantly ameliorated Dex-induced cell death and apoptosis. mTOR inhibition, through mTOR kinase inhibitor OSI-027 or mTOR shRNAs, abolished MHY1485-mediated osteoblast cytoprotection against Dex. Intriguingly, activation of mTOR complex (mTORC1), but not mTORC2, is required for MHY1485's anti-Dex activity. mTORC1 inhibitors (rapamycin and RAD001) or Raptor knockdown almost reversed MHY1485-induced osteoblast cytoprotection. mTORC2 inhibition, via shRNA knockdown of Rictor, failed to affect MHY1485's activity in MC3T3-E1 cells. Further studies showed that MHY1485 treatment in MC3T3-E1 cells and primary murine osteoblasts significantly inhibited Dex-induced mitochondrial death pathway activation, the latter was tested by mitochondrial depolarization, cyclophilin D-ANT-1 association and cytochrome C cytosol release. Together, these results suggest that MHY1485 activates mTORC1 signaling to protect osteoblasts from Dex.

  14. Che-1-induced inhibition of mTOR pathway enables stress-induced autophagy

    PubMed Central

    Desantis, Agata; Bruno, Tiziana; Catena, Valeria; De Nicola, Francesca; Goeman, Frauke; Iezzi, Simona; Sorino, Cristina; Ponzoni, Maurilio; Bossi, Gianluca; Federico, Vincenzo; La Rosa, Francesca; Ricciardi, Maria Rosaria; Lesma, Elena; De Meo, Paolo D'Onorio; Castrignanò, Tiziana; Petrucci, Maria Teresa; Pisani, Francesco; Chesi, Marta; Bergsagel, P Leif; Floridi, Aristide; Tonon, Giovanni; Passananti, Claudio; Blandino, Giovanni; Fanciulli, Maurizio

    2015-01-01

    Mammalian target of rapamycin (mTOR) is a key protein kinase that regulates cell growth, metabolism, and autophagy to maintain cellular homeostasis. Its activity is inhibited by adverse conditions, including nutrient limitation, hypoxia, and DNA damage. In this study, we demonstrate that Che-1, a RNA polymerase II-binding protein activated by the DNA damage response, inhibits mTOR activity in response to stress conditions. We found that, under stress, Che-1 induces the expression of two important mTOR inhibitors, Redd1 and Deptor, and that this activity is required for sustaining stress-induced autophagy. Strikingly, Che-1 expression correlates with the progression of multiple myeloma and is required for cell growth and survival, a malignancy characterized by high autophagy response. PMID:25770584

  15. MicroRNA-96 promotes myocardial hypertrophy by targeting mTOR.

    PubMed

    Sun, Xuemei; Zhang, Chunlai

    2015-01-01

    As a main cause of cardiac hypertrophy, myocardial hypertrophy includes the proliferation and enlongation of myocardial cell, resulting in abnormally cardiac enlargement. However, the pathogenesis and the molecular mechanism that regulate gene expression of myocardial hypertrophy remain incompletely understood. MiRNAs were deemed as an important molecules involved in a variety of pathological processes. MiR-96 has been reported being associated with the tumor proliferation, but whether miR-96 is involved in cardiac hypertrophy remains uncertain. In this study, we have confirmed that, as the myocardial hypertrophy gene, mTOR was a target gene of miR-96, who would promote the occurrence of myocardial hypertrophy. Thus, we got the conclusion that miR-96 could promote myocardial hypertrophy by inhibiting mTOR, miR-96 and mTOR were negatively correlated.

  16. Inhibition of the mechanistic target of rapamycin (mTOR) - Rapamycin and beyond

    PubMed Central

    Lamming, Dudley W.

    2016-01-01

    Rapamycin is an FDA-approved immunosuppressant and anti-cancer agent discovered in the soil of Easter Island in the early 1970s. Rapamycin is a potent and selective inhibitor of the mTOR (mechanistic Target Of Rapamycin) protein kinase, which acts as a central integrator of nutrient signaling pathways. During the last decade, genetic and pharmaceutical inhibition of mTOR pathway signaling has been found to promote longevity in yeast, worms, flies and mice. In this chapter, we will discuss the molecular biology underlying the effects of rapamycin and its physiological effects; evidence for rapamycin as an anti-aging compound; mechanisms by which rapamycin may extend lifespan; and the potential limitations of rapamycin as an anti-aging molecule. Finally, we will discuss possible strategies that may allow us to inhibit mTOR signaling safely while minimizing side effects, and reap the health, social and economic benefits from slowing the aging process. PMID:27048303

  17. Rapamycin-Resistant mTOR Activity Is Required for Sensory Axon Regeneration Induced by a Conditioning Lesion

    PubMed Central

    Lu, Na; Ding, Yue; Chan, Leung Ting; Wang, Xu; Gao, Xin; Jiang, Songshan

    2016-01-01

    Abstract Neuronal mammalian target of rapamycin (mTOR) activity is a critical determinant of the intrinsic regenerative ability of mature neurons in the adult central nervous system (CNS). However, whether its action also applies to peripheral nervous system (PNS) neurons after injury remains elusive. To address this issue unambiguously, we used genetic approaches to determine the role of mTOR signaling in sensory axon regeneration in mice. We showed that deleting mTOR in dorsal root ganglion (DRG) neurons suppressed the axon regeneration induced by conditioning lesions. To establish whether the impact of mTOR on axon regeneration results from functions of mTOR complex 1 (mTORC1) or 2 (mTORC2), two distinct kinase complexes, we ablated either Raptor or Rictor in DRG neurons. We found that suppressing mTORC1 signaling dramatically decreased the conditioning lesion effect. In addition, an injury to the peripheral branch boosts mTOR activity in DRG neurons that cannot be completely inhibited by rapamycin, a widely used mTOR-specific inhibitor. Unexpectedly, examining several conditioning lesion–induced pro-regenerative pathways revealed that Raptor deletion but not rapamycin suppressed Stat3 activity in neurons. Therefore, our results demonstrate that crosstalk between mTOR and Stat3 signaling mediates the conditioning lesion effect and provide genetic evidence that rapamycin-resistant mTOR activity contributes to the intrinsic axon growth capacity in adult sensory neurons after injury. PMID:28101526

  18. Hypothalamic roles of mTOR complex I: Integration of nutrient and hormone signals to regulate energy homeostasis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Mammalian or mechanistic target of rapamycin (mTOR) senses nutrient, energy, and hormone signals to regulate metabolism and energy homeostasis. mTOR activity in the hypothalamus, which is associated with changes in energy status, plays a critical role in the regulation of food intake and body weight...

  19. A carbazole alkaloid deactivates mTOR through the suppression of rictor and that induces apoptosis in lung cancer cells.

    PubMed

    Chatterjee, Priyajit; Seal, Soma; Mukherjee, Sandip; Kundu, Rakesh; Bhuyan, Mantu; Barua, Nabin C; Baruah, Pranab K; Babu, Santi Prasad Sinha; Bhattacharya, Samir

    2015-07-01

    Non-small cell lung cancer (NSCLC) is known to be a difficult cancer to treat because of its poor prognosis, limited option for surgery, and resistance to chemo or radiotherapy. In this study, we have demonstrated that suppression of rictor expression in A549 and H1299 NSCLC cells by mahanine, a carbazole alkaloid, disrupted constitutive activation of mTOR and Akt. Mahanine suppression of rictor gene expression and consequent attenuation of its protein expression affected the inhibition of mTOR (Ser-2481) and Akt (Ser-473) phosphorylation. Since mahanine treatment revealed this new insight of rictor-mTOR relationship, we examined an association between mTOR activation with rictor expression. Interestingly, in rictor knockdown (KD) NSCLC cells, mTOR activation was significantly impaired. Transfection of rictor over-expression vector into the NSCLC cells reversed this situation. In fact, both rictor KD and mahanine treated cells showed considerably depleted phospho-mTOR level. These results indicate that rictor is required to maintain constitutive activation of mTOR in lung cancer cells. When mTOR kinase activity in rictor KD cells was examined with Akt as substrate, a significant reduction of Akt phosphorylation indicated impairment of mTOR kinase potentiality. Disruption of mTOR and Akt activation caused drastic mortality of NSCLC cancer cells through apoptosis. Hence, our study reveals a new dimension in mTOR-rictor relationship, where rictor stands to be a suitable therapeutic target for lung cancer.

  20. Age related increase in mTOR activity contributes to the pathological changes in ovarian surface epithelium

    PubMed Central

    Bajwa, Preety; Nagendra, Prathima B.; Nielsen, Sarah; Sahoo, Subhransu S.; Bielanowicz, Amanda; Lombard, Janine M.; Wilkinson, Erby J.; Miller, Richard A.; Tanwar, Pradeep S.

    2016-01-01

    Ovarian cancer is a disease of older women. However, the molecular mechanisms of ovarian aging and their contribution to the pathogenesis of ovarian cancer are currently unclear. mTOR signalling is a major regulator of aging as suppression of this pathway extends lifespan in model organisms. Overactive mTOR signalling is present in up to 80% of ovarian cancer samples and is associated with poor prognosis. This study examined the role of mTOR signalling in age-associated changes in ovarian surface epithelium (OSE). Histological examination of ovaries from both aged mice and women revealed OSE cell hyperplasia, papillary growth and inclusion cysts. These pathological lesions expressed bonafide markers of ovarian cancer precursor lesions, Pax8 and Stathmin 1, and were presented with elevated mTOR signalling. To understand whether overactive mTOR signalling is responsible for the development of these pathological changes, we analysed ovaries of the Pten trangenic mice and found significant reduction in OSE lesions compared to controls. Furthermore, pharmacological suppression of mTOR signalling significantly decreased OSE hyperplasia in aged mice. Treatment with mTOR inhibitors reduced human ovarian cancer cell viability, proliferation and colony forming ability. Collectively, we have established the role of mTOR signalling in age-related OSE pathologies and initiation of ovarian cancer. PMID:27036037

  1. Comparative idiosyncrasies in life extension by reduced mTOR signalling and its distinctiveness from dietary restriction.

    PubMed

    Garratt, Michael; Nakagawa, Shinichi; Simons, Mirre J P

    2016-08-01

    Reduced mechanistic target of rapamycin (mTOR) signalling extends lifespan in yeast, nematodes, fruit flies and mice, highlighting a physiological pathway that could modulate aging in evolutionarily divergent organisms. This signalling system is also hypothesized to play a central role in lifespan extension via dietary restriction. By collating data from 48 available published studies examining lifespan with reduced mTOR signalling, we show that reduced mTOR signalling provides similar increases in median lifespan across species, with genetic mTOR manipulations consistently providing greater life extension than pharmacological treatment with rapamycin. In contrast to the consistency in changes in median lifespan, however, the demographic causes for life extension are highly species specific. Reduced mTOR signalling extends lifespan in nematodes by strongly reducing the degree to which mortality rates increase with age (aging rate). By contrast, life extension in mice and yeast occurs largely by pushing back the onset of aging, but not altering the shape of the mortality curve once aging starts. Importantly, in mice, the altered pattern of mortality induced by reduced mTOR signalling is different to that induced by dietary restriction, which reduces the rate of aging. Effects of mTOR signalling were also sex dependent, but only within mice, and not within flies, thus again species specific. An alleviation of age-associated mortality is not a shared feature of reduced mTOR signalling across model organisms and does not replicate the established age-related survival benefits of dietary restriction.

  2. B cell-specific deficiencies in mTOR limit humoral immune responses.

    PubMed

    Zhang, Shuling; Pruitt, Margaret; Tran, Dena; Du Bois, Wendy; Zhang, Ke; Patel, Rushi; Hoover, Shelley; Simpson, R Mark; Simmons, John; Gary, Joy; Snapper, Clifford M; Casellas, Rafael; Mock, Beverly A

    2013-08-15

    Generation of high-affinity Abs in response to Ags/infectious agents is essential for developing long-lasting immune responses. B cell maturation and Ab responses to Ag stimulation require Ig somatic hypermutation (SHM) and class-switch recombination (CSR) for high-affinity responses. Upon immunization with either the model Ag 4-hydroxy-3-nitrophenylacetyl hapten (NP) conjugated to chicken γ globulin lysine (NP-CGG) or heat-killed Streptococcus pneumoniae capsular type 14 protein (Pn14), knock-in (KI) mice hypomorphic for mTOR function had a decreased ability to form germinal centers, develop high-affinity anti-NP-specific or anti-Pn14-specific Abs, and perform SHM/CSR. Hypomorphic mTOR mice also had a high mortality (40%) compared with wild-type (WT) (0%) littermates and had lower pneumococcal surface protein A-specific Ab titers when immunized and challenged with live S. pneumoniae infection. Mice with mTOR deleted in their B cell lineage (knockout [KO]) also produced fewer splenic germinal centers and decreased high-affinity Ab responses to NP-CGG than did their WT littermates. CSR rates were lower in mTOR KI and KO mice, and pharmacologic inhibition of mTOR in WT B cells resulted in decreased rates of ex vivo CSR. RNA and protein levels of activation-induced cytidine deaminase (AID), a protein essential for SHM and CSR, were lower in B cells from both KI and B cell-specific KO mice, concomitant with increases in phosphorylated AKT and FOXO1. Rescue experiments increasing AID expression in KI B cells restored CSR levels to those in WT B cells. Thus, mTOR plays an important immunoregulatory role in the germinal center, at least partially through AID signaling, in generating high-affinity Abs.

  3. Cerebral mTOR signal and pro-inflammatory cytokines in Alzheimer’s disease rats

    PubMed Central

    Wang, Xu; Li, Guang-Jian; Hu, Hai-Xia; Ma, Chi; Ma, Di-Hui; Liu, Xiao-Liang

    2016-01-01

    Abstract As a part of Alzheimer’s disease (AD) development the mammalian target of rapamycin (mTOR) has been reported to play a crucial role in regulating cognition and can be used as a neuronal marker. Neuro-inflammation is also a cause of the pathophysiological process in AD. Thus, we examined the protein expression levels of mTOR and its downstream pathways as well as pro-inflammatory cytokines (PICs) in the brain of AD rats. We further examined the effects of blocking mTOR on PICs, namely IL-1β, IL-6 and TNF-α. Our results showed that the protein expression of p-mTOR, mTOR-mediated phosphorylation of 4E-binding protein 4 (4E-BP1) and p70 ribosomal S6 protein kinase 1 (S6K1) pathways were amplified in the hippocampus of AD rats compared with controls. Blocking mTOR by using rapamycin selectively enhanced activities of IL-6 and TNF-α signaling pathways, which was accompanied with an increase of Caspase-3, indicating cellular apoptosis and worsened learning performance. In conclusion, our data for the first time revealed specific signaling pathways engaged in the development of AD, including a regulatory role by the activation of mTOR in PIC mechanisms. Stimulation of mTOR is likely to play a beneficial role in modulating neurological deficits in AD.Targeting one or more of these signaling molecules may present with new opportunities for treatment and clinical management of AD PMID:28123835

  4. Phosphatidic acid enhances mTOR signaling and resistance exercise induced hypertrophy

    PubMed Central

    2014-01-01

    Introduction The lipid messenger phosphatidic acid (PA) plays a critical role in the stimulation of mTOR signaling. However, the mechanism by which PA stimulates mTOR is currently unknown. Therefore, the purpose of this study was to compare the effects of various PA precursors and phospholipids on their ability to stimulate mTOR signaling and its ability to augment resistance training-induced changes in body composition and performance. Methods In phase one, C2C12 myoblasts cells were stimulated with different phospholipids and phospholipid precursors derived from soy and egg sources. The ratio of phosphorylated p70 (P-p70-389) to total p70 was then used as readout for mTOR signaling. In phase two, resistance trained subjects (n = 28, 21 ± 3 years, 77 ± 4 kg, 176 ± 9 cm) consumed either 750 mg PA daily or placebo and each took part in an 8 week periodized resistance training program. Results In phase one, soy-phosphatidylserine, soy-Lyso-PA, egg-PA, and soy-PA stimulated mTOR signaling, and the effects of soy-PA (+636%) were significantly greater than egg-PA (+221%). In phase two, PA significantly increased lean body mass (+2.4 kg), cross sectional area (+1.0 cm), and leg press strength (+51.9 kg) over placebo. Conclusion PA significantly activates mTOR and significantly improved responses in skeletal muscle hypertrophy, lean body mass, and maximal strength to resistance exercise. PMID:24959196

  5. Characterization of Torin2, an ATP-competitive inhibitor of mTOR, ATM and ATR

    PubMed Central

    Liu, Qingsong; Xu, Chunxiao; Kirubakaran, Sivapriya; Zhang, Xin; Hur, Wooyoung; Liu, Yan; Kwiatkowski, Nicholas P.; Wang, Jinhua; Westover, Kenneth D.; Gao, Peng; Ercan, Dalia; Niepel, Mario; Thoreen, Carson C.; Kang, Seong A.; Patricelli, Matthew P.; Wang, Yuchuan; Tupper, Tanya; Altabef, Abigail; Kawamura, Hidemasa; Held, Kathryn D.; Chou, Danny M.; Elledge, Stephen J.; Janne, Pasi A.; Wong, Kwok-Kin; Sabatini, David M.; Gray, Nathanael S.

    2013-01-01

    mTOR is a highly conserved serine/threonine protein kinase that serves as a central regulator of cell growth, survival and autophagy. Deregulation of the PI3K/Akt/mTOR signaling pathway occurs commonly in cancer and numerous inhibitors targeting the ATP-binding site of these kinases are currently undergoing clinical evaluation. Here we report the characterization of Torin2, a second generation ATP-competitive inhibitor that is potent and selective for mTOR with a superior pharmacokinetic profile to previous inhibitors. Torin2 inhibited mTORC1-dependent T389 phosphorylation on S6K (RPS6KB1) with an EC50 of 250 pM with approximately 800-fold selectivity for cellular mTOR versus PI3K. Torin2 also exhibited potent biochemical and cellular activity against PIKK family kinases including ATM (EC50 28 nM), ATR (EC50 35 nM) and DNA-PK (EC50 118 nM) (PRKDC), the inhibition of which sensitized cells to Irradiation. Similar to the earlier generation compound Torin1 and in contrast to other reported mTOR inhibitors, Torin2 inhibited mTOR kinase and mTORC1 signaling activities in a sustained manner suggestive of a slow dissociation from the kinase. Cancer cell treatment with Torin2 for 24 hours resulted in a prolonged block in negative feedback and consequent T308 phosphorylation on Akt. These effects were associated with strong growth inhibition in vitro. Single agent treatment with Torin2 in vivo did not yield significant efficacy against KRAS-driven lung tumors, but the combination of Torin2 with MEK inhibitor AZD6244 yielded a significant growth inhibition. Taken together, our findings establish Torin2 as a strong candidate for clinical evaluation in a broad number of oncological settings where mTOR signaling has a pathogenic role. PMID:23436801

  6. B Cell-specific Deficiencies in mTOR Limit Humoral Immune Responses1

    PubMed Central

    Zhang, Shuling; Pruitt, Margaret; Tran, Dena; Bois, Wendy Du; Zhang, Ke; Patel, Rushi; Hoover, Shelley; Simpson, R. Mark; Simmons, John; Gary, Joy; Snapper, Clifford M.; Casellas, Rafael; Mock, Beverly A.

    2013-01-01

    Generation of high-affinity antibodies in response to antigens/infectious agents is essential for developing long-lasting immune responses. B cell maturation and antibody responses to antigen stimulation require immunoglobulin (Ig) somatic hypermutation (SHM) and class-switch recombination (CSR) for high-affinity responses. Upon immunization with either the model antigen NP-CGG or heat-killed Pn14 derived from Streptococcus pneumoniae, knock-in (KI) mice hypomorphic for mTOR function had decreased ability to form germinal centers, develop high-affinity anti-NP or –Pn14 specific antibodies, and perform SHM/CSR. Hypomorphic mTOR mice also had a high mortality rate (40%) compared to WT (0%) littermates and had lower PspA specific antibody titers when immunized and challenged with live S. pneumoniae infection. Mice with mTOR deleted in their B cell lineage (KO) also produced fewer splenic germinal centers and decreased high-affinity antibody responses to NP-CGG than their WT littermates. CSR rates were lower in mTOR KI and KO mice, and pharmacologic inhibition of mTOR in WT B cells resulted in decreased rates of ex vivo CSR. RNA and protein levels of activation-induced cytidine deaminase (AID), a protein essential for SHM and CSR, were lower in B cells from both KI and B-cell specific KO mice, concomitant with increases in phosphorylated AKT and FOXO1. Rescue experiments increasing AID expression in KI B cells restored CSR levels to those in wild-type B cells. Thus, mTOR plays an important immunoregulatory role in the germinal center, at least partially through AID signaling, in generating high affinity antibodies. PMID:23858034

  7. Dietary flavonoid fisetin: A novel dual inhibitor of PI3K/Akt and mTOR for prostate cancer management

    PubMed Central

    Adhami, Vaqar Mustafa; Syed, Deeba; Khan, Naghma; Mukhtar, Hasan

    2013-01-01

    Epidemiologic and case control population based studies over the past few decades have identified diet as an important determinant of cancer risk. This evidence has kindled an interest into research on bioactive food components and has till date resulted in the identification of many compounds with cancer preventive and therapeutic potential. Among such compounds has been fisetin (3,7,3’,4’-tetrahydroxyflavone), a flavonol and a member of the flavonoid polyphenols that also include quercetin, myricetin and kaempferol. Fisetin is commonly found in many fruits and vegetables such as apples, persimmons, grapes, kiwis, strawberries, onions and cucumbers. We evaluated the effects of fisetin against melanoma and cancers of the prostate, pancreas and the lungs. Using prostate and lung adenocarcinoma cells, we observed that fisetin acts as a dual inhibitor of the PI3K/Akt and the mTOR pathways. This is a significant finding considering the fact that mTOR is phosphorylated and its activation is more frequent in tumors with overexpression of PI3K/Akt. Dual inhibitors of PI3K/Akt and mTOR signaling have been suggested as valuable agents for treating such cancers. Here, we summarize our findings on the dietary flavonoid fisetin and its effects on cancer with particular focus on prostate cancer. Our observations and findings from other laboratories suggest that fisetin could be a useful chemotherapeutic agent that could be used either alone or as an adjuvant with conventional chemotherapeutic drugs for the management of prostate and other cancers. PMID:22842629

  8. mTOR ensures increased release and reduced uptake of the organic osmolyte taurine under hypoosmotic conditions in mouse fibroblasts.

    PubMed

    Lambert, Ian Henry; Jensen, Jane Vendelbo; Pedersen, Per Amstrup

    2014-06-01

    Mammalian target of rapamycin (mTOR) is a serine/threonine kinase that modulates translation in response to growth factors and alterations in nutrient availability following hypoxia and DNA damage. Here we demonstrate that mTOR activity in Ehrlich Lettré ascites (ELA) cells is transiently increased within minutes following osmotic cell swelling and that inhibition of phosphatidylinositol-3-phosphatase (PTEN) counteracts the upstream phosphatidylinositol kinase and potentiates mTOR activity. PTEN inhibition concomitantly potentiates swelling-induced taurine release via the volume-sensitive transporter for organic osmolytes and anion channels (VSOAC) and enhances swelling-induced inhibition of taurine uptake via the taurine-specific transporter (TauT). Chronic osmotic stress, i.e., exposure to hypotonic or hypertonic media for 24 h, reduces and increases mTOR activity in ELA cells, respectively. Using rapamycin, we demonstrate that mTOR inhibition is accompanied by reduction in TauT activity and increase in VSOAC activity in cells expressing high (NIH3T3 fibroblasts) or low (ELA) amounts of mTOR protein. The effect of mTOR inhibition on TauT activity reflects reduced TauT mRNA, TauT protein abundance, and an overall reduction in protein synthesis, whereas the effect on VSOAC is mimicked by catalase inhibition and correlates with reduced catalase mRNA abundance. Hence, mTOR activity favors loss of taurine following hypoosmotic cell swelling, i.e., release via VSOAC and uptake via TauT during acute hypotonic exposure is potentiated and reduced, respectively, by phosphorylation involving mTOR and/or the kinases upstream to mTOR. Decrease in TauT activity during chronic hypotonic exposure, on the other hand, involves reduction in expression/activity of TauT and enzymes in antioxidative defense.

  9. CC-223, a Potent and Selective Inhibitor of mTOR Kinase: In Vitro and In Vivo Characterization.

    PubMed

    Mortensen, Deborah S; Fultz, Kimberly E; Xu, Shuichan; Xu, Weiming; Packard, Garrick; Khambatta, Godrej; Gamez, James C; Leisten, Jim; Zhao, Jingjing; Apuy, Julius; Ghoreishi, Kamran; Hickman, Matt; Narla, Rama Krishna; Bissonette, Rene; Richardson, Samantha; Peng, Sophie X; Perrin-Ninkovic, Sophie; Tran, Tam; Shi, Tao; Yang, Wen Qing; Tong, Zeen; Cathers, Brian E; Moghaddam, Mehran F; Canan, Stacie S; Worland, Peter; Sankar, Sabita; Raymon, Heather K

    2015-06-01

    mTOR is a serine/threonine kinase that regulates cell growth, metabolism, proliferation, and survival. mTOR complex-1 (mTORC1) and mTOR complex-2 (mTORC2) are critical mediators of the PI3K-AKT pathway, which is frequently mutated in many cancers, leading to hyperactivation of mTOR signaling. Although rapamycin analogues, allosteric inhibitors that target only the mTORC1 complex, have shown some clinical activity, it is hypothesized that mTOR kinase inhibitors, blocking both mTORC1 and mTORC2 signaling, will have expanded therapeutic potential. Here, we describe the preclinical characterization of CC-223. CC-223 is a potent, selective, and orally bioavailable inhibitor of mTOR kinase, demonstrating inhibition of mTORC1 (pS6RP and p4EBP1) and mTORC2 [pAKT(S473)] in cellular systems. Growth inhibitory activity was demonstrated in hematologic and solid tumor cell lines. mTOR kinase inhibition in cells, by CC-223, resulted in more complete inhibition of the mTOR pathway biomarkers and improved antiproliferative activity as compared with rapamycin. Growth inhibitory activity and apoptosis was demonstrated in a panel of hematologic cancer cell lines. Correlative analysis revealed that IRF4 expression level associates with resistance, whereas mTOR pathway activation seems to associate with sensitivity. Treatment with CC-223 afforded in vivo tumor biomarker inhibition in tumor-bearing mice, after a single oral dose. CC-223 exhibited dose-dependent tumor growth inhibition in multiple solid tumor xenografts. Significant inhibition of mTOR pathway markers pS6RP and pAKT in CC-223-treated tumors suggests that the observed antitumor activity of CC-223 was mediated through inhibition of both mTORC1 and mTORC2. CC-223 is currently in phase I clinical trials.

  10. mTOR regulates skeletal muscle regeneration in vivo through kinase-dependent and kinase-independent mechanisms.

    PubMed

    Ge, Yejing; Wu, Ai-Luen; Warnes, Christine; Liu, Jianming; Zhang, Chongben; Kawasome, Hideki; Terada, Naohiro; Boppart, Marni D; Schoenherr, Christopher J; Chen, Jie

    2009-12-01

    Rapamycin-sensitive signaling is required for skeletal muscle differentiation and remodeling. In cultured myoblasts, the mammalian target of rapamycin (mTOR) has been reported to regulate differentiation at different stages through distinct mechanisms, including one that is independent of mTOR kinase activity. However, the kinase-independent function of mTOR remains controversial, and no in vivo studies have examined those mTOR myogenic mechanisms previously identified in vitro. In this study, we find that rapamycin impairs injury-induced muscle regeneration. To validate the role of mTOR with genetic evidence and to probe the mechanism of mTOR function, we have generated and characterized transgenic mice expressing two mutants of mTOR under the control of human skeletal actin (HSA) promoter: rapamycin-resistant (RR) and RR/kinase-inactive (RR/KI). Our results show that muscle regeneration in rapamycin-administered mice is restored by RR-mTOR expression. In the RR/KI-mTOR mice, nascent myofiber formation during the early phase of regeneration proceeds in the presence of rapamycin, but growth of the regenerating myofibers is blocked by rapamycin. Igf2 mRNA levels increase drastically during early regeneration, which is sensitive to rapamycin in wild-type muscles but partially resistant to rapamycin in both RR- and RR/KI-mTOR muscles, consistent with mTOR regulation of Igf2 expression in a kinase-independent manner. Furthermore, systemic ablation of S6K1, a target of mTOR kinase, results in impaired muscle growth but normal nascent myofiber formation during regeneration. Therefore, mTOR regulates muscle regeneration through kinase-independent and kinase-dependent mechanisms at the stages of nascent myofiber formation and myofiber growth, respectively.

  11. Long-Term Follow-Up of Patients after Percutaneous Coronary Intervention with Everolimus-Eluting Bioresorbable Vascular Scaffold

    PubMed Central

    Meneguz-Moreno, Rafael Alexandre; Costa Junior, José de Ribamar; Moscoso, Freddy Antônio Britto; Staico, Rodolfo; Tanajura, Luiz Fernando Leite; Centemero, Marinella Patrizia; Chaves, Auréa Jacob; Abizaid, Andrea Claudia Leão de Sousa; Sousa, Amanda Guerra de Moraes Rego e; Abizaid, Alexandre Antonio Cunha

    2017-01-01

    Background Bioresorbable vascular scaffolds (BVS) were developed to improve the long-term results of percutaneous coronary intervention, restoring vasomotion. Objectives To report very late follow-up of everolimus-eluting Absorb BVS (Abbott Vascular, Santa Clara, USA) in our center. Methods Observational retrospective study, in a single Brazilian center, from August 2011 to October 2013, including 49 patients submitted to Absorb BVS implantation. Safety and efficacy outcomes were analyzed in the in-hospital and very late follow-up phases (> 2 years). Results All 49 patients underwent a minimum follow-up of 2.5 years and a maximum of 4.6 years. Mean age was 56.8 ± 7.6 years, 71.4% of the patients were men, and 26.5% were diabetic. Regarding clinical presentation, the majority (94%) had stable angina or silent ischemia. Device success was achieved in 100% of cases with 96% overall procedure success rate. Major adverse cardiovascular events rate was 4% at 30 days, 8.2% at 1 year, and 12.2% at 2 years, and there were no more events until 4.6 years. There were 2 cases of thrombosis (1 subacute and 1 late). Conclusions In this preliminary analysis, Absorb BVS showed to be a safe and effective device in the very late follow-up. Establishing the efficacy and safety profiles of these devices in more complex scenarios is necessary. PMID:28076449

  12. First Report of a Successful Pregnancy in an Everolimus-Treated Heart-Transplanted Patient: Neonatal Disappearance of Immunosuppressive Drugs.

    PubMed

    Fiocchi, R; D'Elia, E; Vittori, C; Sebastiani, R; Strobelt, N; Eleftheriou, G; Introna, M; Freddi, C; Crippa, A

    2016-04-01

    The use of everolimus (EVL) as primary immunosuppression is steadily increasing in heart transplantation (HTx) patients. Limited data currently exist in kidney transplantation, but there is no report of EVL use during pregnancy after HTx and its pharmacokinetics in the newborn. We report a case of an unplanned pregnancy discovered at 21 weeks of gestation in a female HTx patient aged 40 years treated with EVL and cyclosporine (CyA). Because pregnancy was advanced, immunosuppression therapy was left unchanged. At 36 weeks, a healthy infant was delivered. At birth, CyA blood levels were lower in the neonate, but EVL concentrations in maternal and neonatal umbilical blood were similar. Amniotic fluid concentrations were undetectable for both drugs. In the newborn, EVL was measurable at 5 days after birth, whereas CyA disappeared within 2 days. Cord blood displayed a normal count of B and T cells and CD4, CD8 and natural killer cell populations. At birth, both mother and newborn displayed the same blood levels of EVL; therefore, a filter effect of the placenta may be hypothesized for CyA but not for EVL. No immediate complications were observed with this pregnancy.

  13. One-Year Outcomes After Everolimus-Eluting Stents Implantation in Ostial Lesions of Left Anterior Descending Coronary Arteries

    PubMed Central

    Golmohamadi, Zahra; Sokhanvar, Sepideh; Aslanabadi, Naser; Ghaffari, Samad; Sohrabi, Bahram

    2013-01-01

    Background In recent years, stents are increasingly used in variety of coronary lesions. Ostial lesion of left anterior descending coronary artery (LAD) however remains a challenge area because of the invariable involvement of distal left main coronary artery (LMCA). This study was designed to evaluate the clinical and angiographic outcomes of everolimus-eluting stent (EES) implantation for ostial LAD. Methods EESs were implanted in 45 consecutive patients with ostial LAD stenoses. For complete lesion coverage, stent positing was extended into the distal LMCA in 6 patients (13.3%) with intermediated LMCA narrowing. We assess MACE during one-year follow-up. Results In-hospital success rate was 100%; neither cardiac death nor stent thrombosis in our patients, but two patients had myocardial infarction in non-related coronary artery during follow-up. Two patients had angiographic restenosis and underwent TLR. The cumulative MACE-free survival rate was 95.6% at one year. Conclusion EES was in ostial LAD lesions with complete lesion coverage achieving high procedural success rate and acceptable clinical outcomes during one-year follow-up period.

  14. mTOR and differential activation of mitochondria orchestrate neutrophil chemotaxis

    PubMed Central

    Bao, Yi; Ledderose, Carola; Graf, Amelie F.; Brix, Bianca; Birsak, Theresa; Lee, Albert; Zhang, Jingping

    2015-01-01

    Neutrophils use chemotaxis to locate invading bacteria. Adenosine triphosphate (ATP) release and autocrine purinergic signaling via P2Y2 receptors at the front and A2a receptors at the back of cells regulate chemotaxis. Here, we examined the intracellular mechanisms that control these opposing signaling mechanisms. We found that mitochondria deliver ATP that stimulates P2Y2 receptors in response to chemotactic cues, and that P2Y2 receptors promote mTOR signaling, which augments mitochondrial activity near the front of cells. Blocking mTOR signaling with rapamycin or PP242 or mitochondrial ATP production (e.g., with CCCP) reduced mitochondrial Ca2+ uptake and membrane potential, and impaired cellular ATP release and neutrophil chemotaxis. Autocrine stimulation of A2a receptors causes cyclic adenosine monophosphate accumulation at the back of cells, which inhibits mTOR signaling and mitochondrial activity, resulting in uropod retraction. We conclude that mitochondrial, purinergic, and mTOR signaling regulates neutrophil chemotaxis and may be a pharmacological target in inflammatory diseases. PMID:26416965

  15. Combined inhibition of MEK and mTOR has a synergic effect on angiosarcoma tumorgrafts

    PubMed Central

    ANDERSEN, NICHOLAS J.; BOGUSLAWSKI, ELISSA B.; KUK, CYNTHIA Y.; CHAMBERS, CHRISTOPHER M.; DUESBERY, NICHOLAS S.

    2015-01-01

    Angiosarcoma (AS) is a rare neoplasm of endothelial origin that has limited treatment options and poor five-year survival. Using tumorgraft models, we previously showed that AS is sensitive to small-molecule inhibitors that target mitogen-activated/extracellular-signal-regulated protein kinase kinases 1 and 2 (MEK). The objective of this study was to identify drugs that combine with MEK inhibitors to more effectively inhibit AS growth. We examined the in vitro synergy between the MEK inhibitor PD0325901 and inhibitors of eleven common cancer pathways in melanoma cell lines and canine angiosarcoma cell isolates. Combination indices were calculated using the Chou-Talalay method. Optimized combination therapies were evaluated in vivo for toxicity and efficacy using canine angiosarcoma tumorgrafts. Among the drugs we tested, rapamycin stood out because it showed strong synergy with PD0325901 at nanomolar concentrations. We observed that angiosarcomas are insensitive to mTOR inhibition. However, treatment with nanomolar levels of mTOR inhibitor renders these cells as sensitive to MEK inhibition as a melanoma cell line with mutant BRAF. Similar results were observed in B-Raf wild-type melanoma cells as well as in vivo, where treatment of canine AS tumorgrafts with MEK and mTOR inhibitors was more effective than monotherapy. Our data show that a low dose of an mTOR inhibitor can dramatically enhance angiosarcoma and melanoma response to MEK inhibition, potentially widening the field of applications for MEK-targeted therapy. PMID:25955301

  16. mTOR and differential activation of mitochondria orchestrate neutrophil chemotaxis.

    PubMed

    Bao, Yi; Ledderose, Carola; Graf, Amelie F; Brix, Bianca; Birsak, Theresa; Lee, Albert; Zhang, Jingping; Junger, Wolfgang G

    2015-09-28

    Neutrophils use chemotaxis to locate invading bacteria. Adenosine triphosphate (ATP) release and autocrine purinergic signaling via P2Y2 receptors at the front and A2a receptors at the back of cells regulate chemotaxis. Here, we examined the intracellular mechanisms that control these opposing signaling mechanisms. We found that mitochondria deliver ATP that stimulates P2Y2 receptors in response to chemotactic cues, and that P2Y2 receptors promote mTOR signaling, which augments mitochondrial activity near the front of cells. Blocking mTOR signaling with rapamycin or PP242 or mitochondrial ATP production (e.g., with CCCP) reduced mitochondrial Ca(2+) uptake and membrane potential, and impaired cellular ATP release and neutrophil chemotaxis. Autocrine stimulation of A2a receptors causes cyclic adenosine monophosphate accumulation at the back of cells, which inhibits mTOR signaling and mitochondrial activity, resulting in uropod retraction. We conclude that mitochondrial, purinergic, and mTOR signaling regulates neutrophil chemotaxis and may be a pharmacological target in inflammatory diseases.

  17. IKKbeta suppression of TSC1 function links the mTOR pathway with insulin resistance.

    PubMed

    Lee, Dung-Fang; Kuo, Hsu-Ping; Chen, Chun-Te; Wei, Yongkun; Chou, Chao-Kai; Hung, Jen-Yu; Yen, Chia-Jui; Hung, Mien-Chie

    2008-11-01

    The proinflammatory cytokine TNFalpha is one of the factors that links obesity-derived chronic inflammation with insulin resistance. Activation of mTOR signaling pathway has been found to suppress insulin sensitivity through serine phosphorylation and the inhibition of IRS1 by mTOR and its downstream effector, S6K1. It remains elusive that whether the mTOR pathway has a role in TNFalpha-mediated insulin resistance. In the present study, we demonstrated that TNFalpha-IKKbeta-mediated inactivation of TSC1 resulted in increasing phosphorylation of IRS1 serine 307 and serine 636/639, impaired insulin-induced glucose uptake, tyrosine phosphorylation of IRS1, and the association between IRS1 and PI3K p85. Furthermore, a higher expression of pIKKbeta (S181), pTSC1(S511), and pS6(S240/244) was found in livers obtained from both C57BL/6J mice on a high-fat diet and B6.V-Lepob/J mice. Collectively, dysregulation of the TSC1/ TSC2/mTOR signaling pathway by IKKbeta is a common molecular switch for both cancer pathogenesis and diet- and obesity-induced insulin resistance.

  18. mTOR regulates MAPKAPK2 translation to control the senescence-associated secretory phenotype

    PubMed Central

    Herranz, Nicolás; Gallage, Suchira; Mellone, Massimiliano; Wuestefeld, Torsten; Klotz, Sabrina; Hanley, Christopher J.; Raguz, Selina; Acosta, Juan Carlos; Innes, Andrew J; Banito, Ana; Georgilis, Athena; Montoya, Alex; Wolter, Katharina; Dharmalingam, Gopuraja; Faull, Peter; Carroll, Thomas; Martínez-Barbera, Juan Pedro; Cutillas, Pedro; Reisinger, Florian; Heikenwalder, Mathias; Miller, Richard A.; Withers, Dominic; Zender, Lars; Thomas, Gareth J.; Gil, Jesús

    2015-01-01

    Senescent cells secrete a combination of factors collectively known as the senescence-associated secretory phenotype (SASP). The SASP reinforces senescence and activates an immune surveillance response but it can also display pro-tumorigenic properties and contribute to age-related pathologies. In a drug screen to find novel SASP regulators, we uncovered the mTOR inhibitor rapamycin as a potent SASP suppressor. Here we report a mechanism by which mTOR controls the SASP by differentially regulating the translation of the MK2/MAPKAPK2 kinase through 4EBP1. In turn, MAPKAPK2 phosphorylates the RNA binding protein ZFP36L1 during senescence, inhibiting its ability to degrade the transcripts of numerous SASP components. Consequently, mTOR inhibition or constitutive activation of ZFP36L1 impairs the non-cell-autonomous effects of senescent cells both in tumour-suppressive and promoting-promoting contexts. Altogether, our results place regulation of the SASP as a key mechanism by which mTOR could influence cancer, age-related diseases and immune responses. PMID:26280535

  19. mTOR Complex 1 Plays Critical Roles in Hematopoiesis and Pten-Loss-Evoked Leukemogenesis

    PubMed Central

    Kalaitzidis, Demetrios; Sykes, Stephen M.; Wang, Zhu; Punt, Natalie; Tang, Yuefeng; Ragu, Christine; Sinha, Amit U.; Lane, Steven W.; Souza, Amanda L.; Clish, Clary B.; Anastasiou, Dimitrios; Gilliland, D. Gary; Scadden, David T.; Guertin, David A.; Armstrong, Scott A.

    2013-01-01

    SUMMARY The mammalian target of rapamycin (mTOR)-pathway serves as a key sensor of cellular-energetic state, and functions to maintain tissue homeostasis. Hyperactivation of the mTOR pathway impairs hematopoietic stem cell (HSC) function and is associated with leukemogenesis. However, the roles of the unique mTOR complexes (mTORCs) in hematopoeisis and leukemogenesis have not been adequately elucidated. We deleted the mTORC1 component, Raptor (regulatory-associated protein of mTOR), in mouse HSC and its loss causes a non-lethal phenotype characterized by pancytopenia, splenomegaly, and the accumulation of monocytoid cells. Furthermore, Raptor is required for HSC regeneration, and plays largely non-redundant roles with Rictor (rapamycin-insensitive companion of mTOR), in these processes. Ablation of Raptor also significantly extends survival of mice in models of leukemogenesis evoked by Pten deficiency. These data delineate critical roles for mTORC1 in hematopoietic function and leukemogenesis, and inform clinical strategies based on chronic mTORC1 inhibition. PMID:22958934

  20. Potential of GPCRs to modulate MAPK and mTOR pathways in Alzheimer's disease.

    PubMed

    Franco, Rafael; Martínez-Pinilla, Eva; Navarro, Gemma; Zamarbide, Marta

    2017-02-09

    Despite efforts to understand the mechanism of neuronal cell death, finding effective therapies for neurodegenerative diseases is still a challenge. Cognitive deficits are often associated with neurodegenerative diseases. Remarkably, in the absence of consensus biomarkers, diagnosis of diseases such as Alzheimer's still relies on cognitive tests. Unfortunately, all efforts to translate findings in animal models to the patients have been unsuccessful. Alzheimer's disease may be addressed from two different points of view, neuroprotection or cognitive enhancement. Based on recent data, the mammalian target of rapamycin (mTOR) pathway arises as a versatile player whose modulation may impact on mechanisms of both neuroprotection and cognition. Whereas direct targeting of mTOR does not seem to constitute a convenient approach in drug discovery, its indirect modulation by other signaling pathways seems promising. In fact, G-protein-coupled receptors (GPCRs) remain the most common 'druggable' targets and as such pharmacological manipulation of GPCRs with selective ligands may modulate phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K), mitogen-activated protein (MAP) kinase and mTOR signaling pathways. Thus, GPCRs become important targets for potential drug treatments in different neurodegenerative disorders including, but not limited to, Alzheimer's disease. GPCR-mediated modulation of mTOR may take advantage of different GPCRs coupled to different G-dependent and G-independent signal transduction routes, of functional selectivity and/or of biased agonism. Signals mediated by GPCRs may act as coincidence detectors to achieve different benefits in different stages of the neurodegenerative disease.

  1. Foxp3+ T cells inhibit antitumor immune memory modulated by mTOR inhibition.

    PubMed

    Wang, Yanping; Sparwasser, Tim; Figlin, Robert; Kim, Hyung L

    2014-04-15

    Inhibition of mTOR signaling enhances antitumor memory lymphocytes. However, pharmacologic mTOR inhibition also enhances regulatory T-cell (Treg) activity. To counter this effect, Treg control was added to mTOR inhibition in preclinical models. Tregs were controlled with CD4-depleting antibodies because CD4 depletion has high translational potential and already has a well-established safety profile in patients. The antitumor activity of the combination therapy was CD8 dependent and controlled growth of syngeneic tumors even when an adoptive immunotherapy was not used. Lymphocytes resulting from the combination therapy could be transferred into naïve mice to inhibit aggressive growth of lung metastases. The combination therapy enhanced CD8 memory formation as determined by memory markers and functional studies of immune recall. Removal of FoxP3-expressing T lymphocytes was the mechanism underlying immunologic memory formation following CD4 depletion. This was confirmed using transgenic DEREG (depletion of regulatory T cells) mice to specifically remove Foxp3(+) T cells. It was further confirmed with reciprocal studies where stimulation of immunologic memory because of CD4 depletion was completely neutralized by adoptively transferring tumor-specific Foxp3(+) T cells. Also contributing to tumor control, Tregs that eventually recovered following CD4 depletion were less immunosuppressive. These results provide a rationale for further study of mTOR inhibition and CD4 depletion in patients.

  2. A systems study reveals concurrent activation of AMPK and mTOR by amino acids

    PubMed Central

    Pezze, Piero Dalle; Ruf, Stefanie; Sonntag, Annika G.; Langelaar-Makkinje, Miriam; Hall, Philip; Heberle, Alexander M.; Navas, Patricia Razquin; van Eunen, Karen; Tölle, Regine C.; Schwarz, Jennifer J.; Wiese, Heike; Warscheid, Bettina; Deitersen, Jana; Stork, Björn; Fäßler, Erik; Schäuble, Sascha; Hahn, Udo; Horvatovich, Peter; Shanley, Daryl P.; Thedieck, Kathrin

    2016-01-01

    Amino acids (aa) are not only building blocks for proteins, but also signalling molecules, with the mammalian target of rapamycin complex 1 (mTORC1) acting as a key mediator. However, little is known about whether aa, independently of mTORC1, activate other kinases of the mTOR signalling network. To delineate aa-stimulated mTOR network dynamics, we here combine a computational–experimental approach with text mining-enhanced quantitative proteomics. We report that AMP-activated protein kinase (AMPK), phosphatidylinositide 3-kinase (PI3K) and mTOR complex 2 (mTORC2) are acutely activated by aa-readdition in an mTORC1-independent manner. AMPK activation by aa is mediated by Ca2+/calmodulin-dependent protein kinase kinase β (CaMKKβ). In response, AMPK impinges on the autophagy regulators Unc-51-like kinase-1 (ULK1) and c-Jun. AMPK is widely recognized as an mTORC1 antagonist that is activated by starvation. We find that aa acutely activate AMPK concurrently with mTOR. We show that AMPK under aa sufficiency acts to sustain autophagy. This may be required to maintain protein homoeostasis and deliver metabolite intermediates for biosynthetic processes. PMID:27869123

  3. Inhibition of mTOR pathway attenuates migration and invasion of gallbladder cancer via EMT inhibition.

    PubMed

    Zong, Huajie; Yin, Baobing; Zhou, Huading; Cai, Duan; Ma, Baojin; Xiang, Yang

    2014-07-01

    Gallbladder cancer (GBC) is an aggressive disease in which epithelial-mesenchymal transition (EMT) plays a critical role. Whether inhibition of mTOR effects via EMT reversal in GBC remains unclear. Using genetic and pharmacologic inhibitions of mTOR, we investigated the changes of EMT levels in GBC cells. Expressions of EMT related genes were also studied. Migration and invasion assays were carried out and in vivo tumour metastasis mouse models were established. Circulating tumour DNA was quantified. We used EMT index (ratio of Vimentin/Ecadherin expression) to profile EMT levels. We found that inhibition of mTOR using shRNAs and rapamycin inhibited EMT in GBC-SD gallbladder cancer cells. Inhibition of mTOR inhibited EMT in GBC-SD cells in TGF-β-dependent manner, which was contributed majorly by mTORC2 inhibition. Rapamycin decreased invasiveness and migration of GBC-SD cells in vitro and in vivo. We have in the current study shown that rapamycin diminishes the ability of invasion and migration of GBC via inhibition of TGF-β-dependent EMT. Our findings contribute to the understanding of the carcinogenesis of GBC.

  4. Trichomonas vaginalis metalloproteinase induces mTOR cleavage of SiHa cells.

    PubMed

    Quan, Juan-Hua; Choi, In-Wook; Yang, Jung-Bo; Zhou, Wei; Cha, Guang-Ho; Zhou, Yu; Ryu, Jae-Sook; Lee, Young-Ha

    2014-12-01

    Trichomonas vaginalis secretes a number of proteases which are suspected to be the cause of pathogenesis; however, little is understood how they manipulate host cells. The mammalian target of rapamycin (mTOR) regulates cell growth, cell proliferation, cell motility, cell survival, protein synthesis, and transcription. We detected various types of metalloproteinases including GP63 protein from T. vaginalis trophozoites, and T. vaginalis GP63 metalloproteinase was confirmed by sequencing and western blot. When SiHa cells were stimulated with live T. vaginalis, T. vaginalis excretory-secretory products (ESP) or T. vaginalis lysate, live T. vaginalis and T. vaginalis ESP induced the mTOR cleavage in both time- and parasite load-dependent manner, but T. vaginalis lysate did not. Pretreatment of T. vaginalis with a metalloproteinase inhibitor, 1,10-phenanthroline, completely disappeared the mTOR cleavage in SiHa cells. Collectively, T. vaginalis metallopeptidase induces host cell mTOR cleavage, which may be related to survival of the parasite.

  5. Identification of Palmitoleic Acid Controlled by mTOR Signaling as a Biomarker of Polymyositis.

    PubMed

    Yin, Geng; Wang, Ying; Cen, Xiao-Min; Yang, Yuan; Yang, Min; Xie, Qi-Bing

    2017-01-01

    Polymyositis (PM) is a chronic disease characterized by muscle pain, weakness, and increase in muscle-related enzymes, accompanied with inflammations in lymphocytes. However, it is not well understood how the molecular alternations in lymphocytes contribute to the development of polymyositis. The mechanistic target of rapamycin (mTOR) signaling is the central regulator of metabolism and inflammation in mammalian cells. Based on previous studies, we proposed that mTOR signaling may control inflammatory reactions via lipid metabolism. In this study, we aim to figure out the role of mTOR signaling in the development of polymyositis and identify novel biomarkers for the detection and therapy of polymyositis. After screening and validation, we found that palmitoleic acid, a monounsaturated fatty acid, is highly regulated by mTOR signaling. Inhibition of mTORC1 activity decreases palmitoleic acid level. Moreover, mTORC1 regulates the level of palmitoleic acid by controlling its de novo synthesis. Importantly, increased palmitoleic acid has been proven to be a marker of polymyositis. Our work identifies palmitoleic acid in peripheral blood mononuclear cells (PBMC) as a biomarker of polymyositis and offers new targets to the clinical therapy.

  6. Identification of Palmitoleic Acid Controlled by mTOR Signaling as a Biomarker of Polymyositis

    PubMed Central

    Wang, Ying; Cen, Xiao-min; Yang, Yuan; Yang, Min

    2017-01-01

    Polymyositis (PM) is a chronic disease characterized by muscle pain, weakness, and increase in muscle-related enzymes, accompanied with inflammations in lymphocytes. However, it is not well understood how the molecular alternations in lymphocytes contribute to the development of polymyositis. The mechanistic target of rapamycin (mTOR) signaling is the central regulator of metabolism and inflammation in mammalian cells. Based on previous studies, we proposed that mTOR signaling may control inflammatory reactions via lipid metabolism. In this study, we aim to figure out the role of mTOR signaling in the development of polymyositis and identify novel biomarkers for the detection and therapy of polymyositis. After screening and validation, we found that palmitoleic acid, a monounsaturated fatty acid, is highly regulated by mTOR signaling. Inhibition of mTORC1 activity decreases palmitoleic acid level. Moreover, mTORC1 regulates the level of palmitoleic acid by controlling its de novo synthesis. Importantly, increased palmitoleic acid has been proven to be a marker of polymyositis. Our work identifies palmitoleic acid in peripheral blood mononuclear cells (PBMC) as a biomarker of polymyositis and offers new targets to the clinical therapy. PMID:28194428

  7. Coordinate Autophagy and mTOR Pathway Inhibition Enhances Cell Death in Melanoma

    PubMed Central

    Xie, Xiaoqi; White, Eileen P.; Mehnert, Janice M.

    2013-01-01

    The phosphatidylinositol 3-kinase/AKT/mammalian target of rapamycin (PI3K/AKT/mTOR) pathway promotes melanoma tumor growth and survival while suppressing autophagy, a catabolic process through which cells collect and recycle cellular components to sustain energy homeostasis in starvation. Conversely, inhibitors of the PI3K/AKT/mTOR pathway, in particular the mTOR inhibitor temsirolimus (CCI-779), induce autophagy, which can promote tumor survival and thus, these agents potentially limit their own efficacy. We hypothesized that inhibition of autophagy in combination with mTOR inhibition would block this tumor survival mechanism and hence improve the cytotoxicity of mTOR inhibitors in melanoma. Here we found that melanoma cell lines of multiple genotypes exhibit high basal levels of autophagy. Knockdown of expression of the essential autophagy gene product ATG7 resulted in cell death, indicating that survival of melanoma cells is autophagy-dependent. We also found that the lysosomotropic agent and autophagy inhibitor hydroxychloroquine (HCQ) synergizes with CCI-779 and led to melanoma cell death via apoptosis. Combination treatment with CCI-779 and HCQ suppressed melanoma growth and induced cell death both in 3-dimensional (3D) spheroid cultures and in tumor xenografts. These data suggest that coordinate inhibition of the mTOR and autophagy pathways promotes apoptosis and could be a new therapeutic paradigm for the treatment of melanoma. PMID:23383069

  8. mTOR masters monocytic myeloid-derived suppressor cells in mice with allografts or tumors.

    PubMed

    Wu, Tingting; Zhao, Yang; Wang, Hao; Li, Yang; Shao, Lijuan; Wang, Ruoyu; Lu, Jun; Yang, Zhongzhou; Wang, Junjie; Zhao, Yong

    2016-02-01

    CD11b(+) Gr1(+) myeloid-derived suppressor cells (MDSCs) play critical roles in controlling the processes of tumors, infections, autoimmunity and graft rejection. Immunosuppressive drug rapamycin (RPM), targeting on the key cellular metabolism molecule mTOR, is currently used in clinics to treat patients with allo-grafts, autoimmune diseases and tumors. However, the effect of RPM on MDSCs has not been studied. RPM significantly decreases the cell number and the immunosuppressive ability on T cells of CD11b(+) Ly6C(high) monocytic MDSCs (M-MDSCs) in both allo-grafts-transplanted and tumor-bearing mice respectively. Mice with a myeloid-specific deletion of mTOR have poor M-MDSCs after grafting with allo-skin tissue or a tumor. Grafting of allo-skin or tumors significantly activates glycolysis pathways in myeloid precursor cells in bone marrow, which is inhibited by RPM or mTOR deletion. 2-deoxyglucose (2-DG), an inhibitor of the glycolytic pathway, inhibits M-MDSC differentiation from precursors, while enhancing glycolysis by metformin significantly rescues the RPM-caused deficiency of M-MDSCs. Therefore, we offer evidence supporting that mTOR is an intrinsic factor essential for the differentiation and immunosuppressive function of M-MDSCs and that these metabolism-relevant medicines may impact MDSCs-mediated immunosuppression or immune tolerance induction, which is of considerable clinical importance in treating graft rejection, autoimmune diseases and cancers.

  9. Evidence of molecular links between PKR and mTOR signalling pathways in Abeta neurotoxicity: role of p53, Redd1 and TSC2.

    PubMed

    Morel, Milena; Couturier, Julien; Pontcharraud, Raymond; Gil, Roger; Fauconneau, Bernard; Paccalin, Marc; Page, Guylène

    2009-10-01

    The control of translation is disturbed in Alzheimer's disease (AD). This study analysed the crosslink between the up regulation of double-stranded RNA-dependent-protein kinase (PKR) and the down regulation of mammalian target of rapamycin (mTOR) signalling pathways via p53, the protein Regulated in the Development and DNA damage response 1 (Redd1) and the tuberous sclerosis complex (TSC2) factors in two beta-amyloid peptide (Abeta) neurotoxicity models. In SH-SY5Y cells, Abeta42 induced an increase of P(T451)-PKR and of the ratio p66/(p66+p53) in nuclei and a physical interaction between these proteins. Redd1 gene levels increased and P(T1462)-TSC2 decreased. These disturbances were earlier in rat primary neurons with nuclear co-localization of Redd1 and PKR. The PKR gene silencing in SH-SY5Y cells prevented these alterations. p53, Redd1 and TSC2 could represent the molecular links between PKR and mTOR in Abeta neurotoxicity. PKR could be a critical target in a therapeutic program of AD.

  10. Ampelopsin attenuates brain aging of D-gal-induced rats through miR-34a-mediated SIRT1/mTOR signal pathway

    PubMed Central

    Chen, Xianbing; Li, Jie; Yang, Xiaoqi; Fan, Jingjing; Yang, Yi; Chen, Ning

    2016-01-01

    The underlying molecular mechanisms for aging-related neurodegenerative diseases such as Alzheimer's disease (AD) are not fully understood. Currently, growing evidences have revealed that microRNAs (miRNAs) are involved in aging and aging-related diseases. The up-regulation of miR-34a has been reported to be associated with aging-related diseases, and thus it should be a promising therapeutic target. Ampelopsin, also called dihydromyricetin (DHM), a natural flavonoid from Chinese herb Ampelopsis grossedentata, has been reported to possess multiple pharmacological functions including anti-inflammatory, anti-oxidative and anti-cancer functions. Meanwhile, it has also gained tremendous attention against neurodegenerative diseases as an anti-aging compound. In the present study, the model rats with D-gal-induced brain aging revealed an obvious expression of miR-34a; in contrast, it could be significantly suppressed upon DHM treatment. In addition, target genes associated with miR-34a in the presence of DHM treatment were also explored. DHM supplementation inhibited D-gal-induced apoptosis and rescued impaired autophagy of neurons in hippocampus tissue. Moreover, DHM activated autophagy through up-regulated SIRT1 and down-regulated mTOR signal pathways due to the down-regulated miR-34a. In conclusion, DHM can execute the prevention and treatment of D-gal-induced brain aging by miR-34a-mediated SIRT1-mTOR signal pathway. PMID:27780933

  11. Hyperactive mTOR signals in the proopiomelanocortin-expressing hippocampal neurons cause age-dependent epilepsy and premature death in mice

    PubMed Central

    Matsushita, Yuki; Sakai, Yasunari; Shimmura, Mitsunori; Shigeto, Hiroshi; Nishio, Miki; Akamine, Satoshi; Sanefuji, Masafumi; Ishizaki, Yoshito; Torisu, Hiroyuki; Nakabeppu, Yusaku; Suzuki, Akira; Takada, Hidetoshi; Hara, Toshiro

    2016-01-01

    Epilepsy is a frequent comorbidity in patients with focal cortical dysplasia (FCD). Recent studies utilizing massive sequencing data identified subsets of genes that are associated with epilepsy and FCD. AKT and mTOR-related signals have been recently implicated in the pathogenic processes of epilepsy and FCD. To clarify the functional roles of the AKT-mTOR pathway in the hippocampal neurons, we generated conditional knockout mice harboring the deletion of Pten (Pten-cKO) in Proopiomelanocortin-expressing neurons. The Pten-cKO mice developed normally until 8 weeks of age, then presented generalized seizures at 8–10 weeks of age. Video-monitored electroencephalograms detected paroxysmal discharges emerging from the cerebral cortex and hippocampus. These mice showed progressive hypertrophy of the dentate gyrus (DG) with increased expressions of excitatory synaptic markers (Psd95, Shank3 and Homer). In contrast, the expression of inhibitory neurons (Gad67) was decreased at 6–8 weeks of age. Immunofluorescence studies revealed the abnormal sprouting of mossy fibers in the DG of the Pten-cKO mice prior to the onset of seizures. The treatment of these mice with an mTOR inhibitor rapamycin successfully prevented the development of seizures and reversed these molecular phenotypes. These data indicate that the mTOR pathway regulates hippocampal excitability in the postnatal brain. PMID:26961412

  12. Screening mTOR siRNA based on bioinformatics and detecting the transcription by the gold nanoparticle beacon

    NASA Astrophysics Data System (ADS)

    Tian, Caiping; Ma, Yi; Li, Siwen; Gu, Yueqing

    2014-09-01

    Mammalian target of rapamycin (mTOR) as a key protein in PI3K-AKT-mTOR signaling pathway ,plays an important role in the tumor growth. The small interfering RNA (siRNA) of mTOR would decrease the expression of mTOR protein. In this study, we screened the mTOR siRNA sequence using MATLAB software and ascertained it based on BLAST. Then we imported it with the aid of Lipofectamine2000 into MCF-7 cancer cells where mTOR is over expression .And then we used a special hairpin deoxyribonucleic acid (DNA) for combining with the human mTOR mRNA to functionalize gold nanoparticles, which served as a molecule beacon for detecting human mTOR mRNA transcription. Laser scanning confocal microscope and Flow Cytometry data showed that the quenching efficiency was up to 90%,which are consistent with the RT-PCR measurement and Western. Compared to the previous approaches, this beacon has advantages of higher target to background ratio of detection. The strategy reported in this study is a promising approach for the intracellular measurement of the result of siRNA or protein expression in living cells, and has great potential in the study of drug screening and discovery.

  13. Oocyte-dependent activation of MTOR in cumulus cells controls the development and survival of cumulus-oocyte complexes.

    PubMed

    Guo, Jing; Shi, Lanying; Gong, Xuhong; Jiang, Mengjie; Yin, Yaoxue; Zhang, Xiaoyun; Yin, Hong; Li, Hui; Emori, Chihiro; Sugiura, Koji; Eppig, John J; Su, You-Qiang

    2016-08-15

    Communication between oocytes and their companion somatic cells promotes the healthy development of ovarian follicles, which is crucial for producing oocytes that can be fertilized and are competent to support embryogenesis. However, how oocyte-derived signaling regulates these essential processes remains largely undefined. Here, we demonstrate that oocyte-derived paracrine factors, particularly GDF9 and GDF9-BMP15 heterodimer, promote the development and survival of cumulus-cell-oocyte complexes (COCs), partly by suppressing the expression of Ddit4l, a negative regulator of MTOR, and enabling the activation of MTOR signaling in cumulus cells. Cumulus cells expressed less Ddit4l mRNA and protein than mural granulosa cells, which is in striking contrast to the expression of phosphorylated RPS6 (a major downstream effector of MTOR). Knockdown of Ddit4l activated MTOR signaling in cumulus cells, whereas inhibition of MTOR in COCs compromised oocyte developmental competence and cumulus cell survival, with the latter likely to be attributable to specific changes in a subset of transcripts in the transcriptome of COCs. Therefore, oocyte suppression of Ddit4l expression allows for MTOR activation in cumulus cells, and this oocyte-dependent activation of MTOR signaling in cumulus cells controls the development and survival of COCs.

  14. mTOR signaling in aging and neurodegeneration: At the crossroad between metabolism dysfunction and impairment of autophagy.

    PubMed

    Perluigi, Marzia; Di Domenico, Fabio; Butterfield, D Allan

    2015-12-01

    Compelling evidence indicates that the mammalian target of rapamycin (mTOR) signaling pathway is involved in cellular senescence, organismal aging and age-dependent diseases. mTOR is a conserved serine/threonine kinase that is known to be part of two different protein complexes: mTORC1 and mTORC2, which differ in some components and in upstream and downstream signalling. In multicellular organisms, mTOR regulates cell growth and metabolism in response to nutrients, growth factors and cellular energy conditions. Growing studies highlight that disturbance in mTOR signalling in the brain affects multiple pathways including glucose metabolism, energy production, mitochondrial function, cell growth and autophagy. All these events are key players in age-related cognitive decline such as development of Alzheimer disease (AD). The current review discusses the main regulatory roles of mTOR signalling in the brain, in particular focusing on autophagy, glucose metabolism and mitochondrial functions. Targeting mTOR in the CNS can offer new prospective for drug discovery; however further studies are needed for a comprehensive understanding of mTOR, which lies at the crossroads of multiple signals involved in AD etiology and pathogenesis.

  15. Fluoxetine regulates mTOR signalling in a region-dependent manner in depression-like mice.

    PubMed

    Liu, Xiao-Long; Luo, Liu; Mu, Rong-Hao; Liu, Bin-Bin; Geng, Di; Liu, Qing; Yi, Li-Tao

    2015-11-02

    Previous studies have demonstrated that the mammalian target of rapamycin (mTOR) signaling pathway has an important role in ketamine-induced, rapid antidepressant effects despite the acute administration of fluoxetine not affecting mTOR phosphorylation in the brain. However, the effects of long-term fluoxetine treatment on mTOR modulation have not been assessed to date. In the present study, we examined whether fluoxetine, a type of commonly used antidepressant agent, alters mTOR signaling following chronic administration in different brain regions, including the frontal cortex, hippocampus, amygdala and hypothalamus. We also investigated whether fluoxetine enhanced synaptic protein levels in these regions via the activation of the mTOR signaling pathway and its downstream regulators, p70S6K and 4E-BP-1. The results indicated that chronic fluoxetine treatment attenuated the chronic, unpredictable, mild stress (CUMS)-induced mTOR phosphorylation reduction in the hippocampus and amygdala of mice but not in the frontal cortex or the hypothalamus. Moreover, the CUMS-decreased PSD-95 and synapsin I levels were reversed by fluoxetine, and these effects were blocked by rapamycin only in the hippocampus. In conclusion, our findings suggest that chronic treatment with fluoxetine can induce synaptic protein expression by activating the mTOR signaling pathway in a region-dependent manner and mainly in the hippocampus.

  16. FOXO3-mTOR Metabolic Cooperation in the Regulation of Erythroid Cell Maturation and Homeostasis

    PubMed Central

    Zhang, Xin; Campreciós, Genís; Rimmelé, Pauline; Liang, Raymond; Yalcin, Safak; Mungamuri, Sathish Kumar; Barminko, Jeffrey; D’Escamard, Valentina; Baron, Margaret H.; Brugnara, Carlo; Papatsenko, Dmitri; Rivella, Stefano; Ghaffari, Saghi

    2014-01-01

    Ineffective erythropoiesis is observed in many erythroid disorders including β-thalassemia and anemia of chronic disease in which increased production of erythroblasts that fail to mature exacerbate the underlying anemias. As loss of the transcription factor FOXO3 results in erythroblast abnormalities similar to the ones observed in ineffective erythropoiesis, we investigated the underlying mechanisms of the defective Foxo3−/− erythroblast cell cycle and maturation. Here we show that loss of Foxo3 results in overactivation of the JAK2/AKT/mTOR signaling pathway in primary bone marrow erythroblasts partly mediated by redox modulation. We further show that hyperactivation of mTOR signaling interferes with cell cycle progression in Foxo3 mutant erythroblasts. Importantly, inhibition of mTOR signaling, in vivo or in vitro enhances significantly Foxo3 mutant erythroid cell maturation. Similarly, in vivo inhibition of mTOR remarkably improves erythroid cell maturation and anemia in a model of β-thalassemia. Finally we show that FOXO3 and mTOR are likely part of a larger metabolic network in erythroblasts as together they control the expression of an array of metabolic genes some of which are implicated in erythroid disorders. These combined findings indicate that a metabolism-mediated regulatory network centered by FOXO3 and mTOR control the balanced production and maturation of erythroid cells. They also highlight physiological interactions between these proteins in regulating erythroblast energy. Our results indicate that alteration in the function of this network might be implicated in the pathogenesis of ineffective erythropoiesis. PMID:24966026

  17. Glycerol-3-phosphate acyltransferase-1 upregulation by O-GlcNAcylation of Sp1 protects against hypoxia-induced mouse embryonic stem cell apoptosis via mTOR activation

    PubMed Central

    Lee, H J; Ryu, J M; Jung, Y H; Lee, K H; Kim, D I; Han, H J

    2016-01-01

    Oxygen signaling is critical for stem cell regulation, and oxidative stress-induced stem cell apoptosis decreases the efficiency of stem cell therapy. Hypoxia activates O-linked β-N-acetyl glucosaminylation (O-GlcNAcylation) of stem cells, which contributes to regulation of cellular metabolism, as well as cell fate. Our study investigated the role of O-GlcNAcylation via glucosamine in the protection of hypoxia-induced apoptosis of mouse embryonic stem cells (mESCs). Hypoxia increased mESCs apoptosis in a time-dependent manner. Moreover, hypoxia also slightly increased the O-GlcNAc level. Glucosamine treatment further enhanced the O-GlcNAc level and prevented hypoxia-induced mESC apoptosis, which was suppressed by O-GlcNAc transferase inhibitors. In addition, hypoxia regulated several lipid metabolic enzymes, whereas glucosamine increased expression of glycerol-3-phosphate acyltransferase-1 (GPAT1), a lipid metabolic enzyme producing lysophosphatidic acid (LPA). In addition, glucosamine-increased O-GlcNAcylation of Sp1, which subsequently leads to Sp1 nuclear translocation and GPAT1 expression. Silencing of GPAT1 by gpat1 siRNA transfection reduced glucosamine-mediated anti-apoptosis in mESCs and reduced mammalian target of rapamycin (mTOR) phosphorylation. Indeed, LPA prevented mESCs from undergoing hypoxia-induced apoptosis and increased phosphorylation of mTOR and its substrates (S6K1 and 4EBP1). Moreover, mTOR inactivation by rapamycin (mTOR inhibitor) increased pro-apoptotic proteins expressions and mESC apoptosis. Furthermore, transplantation of non-targeting siRNA and glucosamine-treated mESCs increased cell survival and inhibited flap necrosis in mouse skin flap model. Conversely, silencing of GPAT1 expression reversed those glucosamine effects. In conclusion, enhancing O-GlcNAcylation of Sp1 by glucosamine stimulates GPAT1 expression, which leads to inhibition of hypoxia-induced mESC apoptosis via mTOR activation. PMID:27010859

  18. Outcomes in Ethnic Minority Renal Transplant Recipients Receiving Everolimus versus Mycophenolate: Comparative Risk Assessment Results From a Pooled Analysis

    PubMed Central

    Melancon, Keith; Mulgaonkar, Shamkant P.; Delcoro, Carlos; Wiland, Anne; McCague, Kevin; Shihab, Fuad S.

    2013-01-01

    Background Everolimus (EVR) has demonstrated good efficacy after renal transplantation. Racial disparities in clinical outcomes after de novo renal transplantation are well documented; whether the efficacy of EVR varies based on recipient ethnicity is unknown. We conducted a comparative risk assessment of EVR by ethnicity. Methods Data on 2004 renal transplant recipients from three EVR studies were pooled to identify the impact of ethnicity on efficacy outcomes across EVR dosing groups and control groups. Ethnic groups compared were African Americans, non-U.S. blacks, Asians, Hispanics, and Caucasians. EVR groups received either 1.5 or 3 mg per day, with either standard-dose cyclosporine or reduced-dose cyclosporine. Control groups received mycophenolic acid (MPA) with standard-dose cyclosporine. Composite efficacy failure endpoint was graft loss, death, biopsy-proven acute rejection, or lost to follow-up. Adjusted odds ratios were calculated using a logistic regression model. Results The proportion of renal transplant recipients who met the composite endpoint was African Americans (46%), non-U.S. black (35%), Caucasian (31%), Hispanic (28%), and Asian (25%). The odds of meeting the composite endpoint were significantly (P=0.0001) greater for African Americans versus Caucasians but did not differ among the other ethnic groups (ethnic groups were only compared with Caucasians). EVR and MPA were associated with similar efficacy among each of the ethnic groups. Conclusion In this pooled data analysis in more than 2000 renal transplant recipients, EVR versus MPA resulted in similar composite endpoint incidence events across ethnicities. Consistent with previously published data, African Americans had poorer clinical outcomes. EVR is efficacious regardless of ethnicity. PMID:24345868

  19. Longitudinal growth on an everolimus- versus an MMF-based steroid-free immunosuppressive regimen in paediatric renal transplant recipients.

    PubMed

    Billing, Heiko; Burmeister, Greta; Plotnicki, Lukasz; Ahlenstiel, Thurid; Fichtner, Alexander; Sander, Anja; Höcker, Britta; Tönshoff, Burkhard; Pape, Lars

    2013-09-01

    Concerns have been raised that mammalian target of rapamycin inhibitors in pediatric transplant recipients might interfere with longitudinal bone growth by inhibition of growth factor signaling and growth plate chondrocyte proliferation. We therefore undertook a prospective nested, case-control study on longitudinal growth over 2 years in steroid-free pediatric renal transplant recipients. Fourteen patients on a steroid-free maintenance immunosuppressive regimen consisting of low-dose everolimus (EVR) in conjunction with low-dose cyclosporine (CsA) were compared to a matched cohort of 14 steroid-free patients on a standard dose mycophenolate mofetil (MMF) regimen in conjunction with a standard dose calcineurin inhibitor (CNI). The mean change in height standard deviation (SD) score in the first study year was 0.31 ± 0.71 SD score in the EVR group compared to 0.31 ± 0.64 SD score in the MMF group (P = 0.20). For the entire study period of 2 years, the change in height SD score in the EVR group was 0.43 ± 0.81 SDS compared to 0.75 ± 0.85 SDS in the MMF group (P = 0.32). The percentage of prepubertal patients experiencing catch-up growth, defined as an increase in height SD score ≥0.5 in 2 years, was similar in the EVR group (5/8, 65%) and the MMF group (6/8, 75%; P = 1.00). Longitudinal growth over 2 years in steroid-free pediatric patients on low-dose EVR and CsA is not different to that of a matched steroid-free control group on an immunosuppressive regimen with standard-dose CNI and MMF. Hence, low-dose EVR does not appear to negatively impact short-term growth in pediatric renal transplant recipients.

  20. eIF4GI links nutrient sensing by mTOR to cell proliferation and inhibition of autophagy.

    PubMed

    Ramírez-Valle, Francisco; Braunstein, Steve; Zavadil, Jiri; Formenti, Silvia C; Schneider, Robert J

    2008-04-21

    Translation initiation factors have complex functions in cells that are not yet understood. We show that depletion of initiation factor eIF4GI only modestly reduces overall protein synthesis in cells, but phenocopies nutrient starvation or inhibition of protein kinase mTOR, a key nutrient sensor. eIF4GI depletion impairs cell proliferation, bioenergetics, and mitochondrial activity, thereby promoting autophagy. Translation of mRNAs involved in cell growth, proliferation, and bioenergetics were selectively inhibited by reduction of eIF4GI, as was the mRNA encoding Skp2 that inhibits p27, whereas catabolic pathway factors were increased. Depletion or overexpression of other eIF4G family members did not recapitulate these results. The majority of mRNAs that were translationally impaired with eIF4GI depletion were excluded from polyribosomes due to the presence of multiple upstream open reading frames and low mRNA abundance. These results suggest that the high levels of eIF4GI observed in many breast cancers might act to specifically increase proliferation, prevent autophagy, and release tumor cells from control by nutrient sensing.

  1. MTOR regulates the pro-tumorigenic senescence-associated secretory phenotype by promoting IL1A translation

    PubMed Central

    Laberge, Remi-Martin; Sun, Yu; Orjalo, Arturo V.; Patil, Christopher K.; Freund, Adam; Zhou, Lili; Curran, Samuel C.; Davalos, Albert R.; Wilson-Edell, Kathleen A.; Liu, Su; Limbad, Chandani; Demaria, Marco; Li, Patrick; Hubbard, Gene B.; Ikeno, Yuji; Javors, Martin; Desprez, Pierre-Yves; Benz, Christopher C.; Kapahi, Pankaj; Nelson, Peter S.; Campisi, Judith

    2015-01-01

    The TOR (target of rapamycin) kinase limits longevity by poorly understood mechanisms. Rapamycin suppresses the mammalian TORC1 complex, which regulates translation, and extends lifespan in diverse species, including mice. We show that rapamycin selectively blunts the pro-inflammatory phenotype of senescent cells. Cellular senescence suppresses cancer by preventing cell proliferation. However, as senescent cells accumulate with age, the senescence-associated secretory phenotype (SASP) can disrupt tissues and contribute to age-related pathologies, including cancer. MTOR inhibition suppressed the secretion of inflammatory cytokines by senescent cells. Rapamycin reduced IL6 and other cytokine mRNA levels, but selectively suppressed translation of the membrane-bound cytokine IL1A. Reduced IL1A diminished NF-κB transcriptional activity, which controls much of the SASP; exogenous IL1A restored IL6 secretion to rapamycin-treated cells. Importantly, rapamycin suppressed the ability of senescent fibroblasts to stimulate prostate tumour growth in mice. Thus, rapamycin might ameliorate age-related pathologies, including late-life cancer, by suppressing senescence-associated inflammation. PMID:26147250

  2. MTOR regulates the pro-tumorigenic senescence-associated secretory phenotype by promoting IL1A translation.

    PubMed

    Laberge, Remi-Martin; Sun, Yu; Orjalo, Arturo V; Patil, Christopher K; Freund, Adam; Zhou, Lili; Curran, Samuel C; Davalos, Albert R; Wilson-Edell, Kathleen A; Liu, Su; Limbad, Chandani; Demaria, Marco; Li, Patrick; Hubbard, Gene B; Ikeno, Yuji; Javors, Martin; Desprez, Pierre-Yves; Benz, Christopher C; Kapahi, Pankaj; Nelson, Peter S; Campisi, Judith

    2015-08-01

    The TOR (target of rapamycin) kinase limits longevity by poorly understood mechanisms. Rapamycin suppresses the mammalian TORC1 complex, which regulates translation, and extends lifespan in diverse species, including mice. We show that rapamycin selectively blunts the pro-inflammatory phenotype of senescent cells. Cellular senescence suppresses cancer by preventing cell proliferation. However, as senescent cells accumulate with age, the senescence-associated secretory phenotype (SASP) can disrupt tissues and contribute to age-related pathologies, including cancer. MTOR inhibition suppressed the secretion of inflammatory cytokines by senescent cells. Rapamycin reduced IL6 and other cytokine mRNA levels, but selectively suppressed translation of the membrane-bound cytokine IL1A. Reduced IL1A diminished NF-κB transcriptional activity, which controls much of the SASP; exogenous IL1A restored IL6 secretion to rapamycin-treated cells. Importantly, rapamycin suppressed the ability of senescent fibroblasts to stimulate prostate tumour growth in mice. Thus, rapamycin might ameliorate age-related pathologies, including late-life cancer, by suppressing senescence-associated inflammation.

  3. [An Elderly Patient with Metastatic Breast Cancer Who Developed Severe Adverse Events such as Stomatitis and Interstitial Pneumonia after Everolimus plus Exemestane Treatment].

    PubMed

    Sakiyama, Kana; Yoshida, Takashi; Goto, Yoshinari; Kimura, Morihiko

    2016-06-01

    An 80-year-old woman was diagnosed with right breast cancer with clinical Stage IIIA 6 years previously. She underwent mastectomy and axillary lymph node dissection. The pathological diagnosis was invasive micropapillary carcinoma with lymph node involvement. Immunohistochemically, the tumor was positive for estrogen receptor and progesterone receptor, and negative for HER2. Postoperatively, the patient was treated with adjuvant chemotherapy consisting of cyclophosphamide, epirubicin, 5-fluorouracil, and paclitaxel, followed by endocrine therapy with letrozole. Four years after surgery, she experienced a recurrence of breast cancer in the thoracic wall, and was treated with exemestane, toremifene, and fulvestrant for 1 year and 5 months. However, she developed carcinomatous pleurisy and was treated with eribulin. This last treatment was ineffective. Subsequently, she received combination therapy with everolimus and exemestane. Although the pleural effusion reduced markedly after 5 weeks, stomatitis, diarrhea, melena, and interstitial pneumonia occurred as adverse events. The symptoms improved after drug discontinuation and steroid therapy. The combination therapy with everolimus and exemestane is a prospective therapy for hormone-resistant recurrent breast cancer, but the management of adverse events is very important.

  4. Essential role of D1R in the regulation of mTOR complex1 signaling induced by cocaine.

    PubMed

    Sutton, Laurie P; Caron, Marc G

    2015-12-01

    The mammalian target of rapamycin (mTOR) is a serine/threonine kinase that is involved in neuronal adaptions that underlie cocaine-induced sensitization and reward. mTOR exists in two functionally distinct multi-component complexes known as mTORC1 and mTORC2. In this study, we show that increased mTORC1 activity induced by cocaine is mediated by the dopamine D1 receptor (D1R). Specifically, cocaine treatment increased the phosphorylation on residues Thr2446 and Ser2481 but not on Ser2448 in the nucleus accumbens (NAc) and that this increase in phosphorylated mTOR levels was also apparent when complexed with its binding partner Raptor. Furthermore, the increase in phosphorylated mTOR levels, as well as phosphorylated 4E-BP1 and S6K, downstream targets of mTORC1 were blocked with SCH23390 treatment. Similar results were also observed in the dopamine-transporter knockout mice as the increase in phosphorylated mTOR Thr2446 and Ser2481 was blocked by SCH23390 but not with raclopride. To further validate D1R role in mTORC1 signaling, decrease in phosphorylated mTOR levels were observed in D1R knockout mice, whereas administration of SKF81297 elevated phosphorylated mTOR in the NAc. Lastly deletion of mTOR or Raptor in D1R expressing neurons reduced cocaine-induced locomotor activity. Together, our data supports a mechanism whereby mTORC1 signaling is activated by cocaine administration through the stimulation of D1R.

  5. mTOR direct interactions with Rheb-GTPase and raptor: sub-cellular localization using fluorescence lifetime imaging

    PubMed Central

    2013-01-01

    Background The mammalian target of rapamycin (mTOR) signalling pathway has a key role in cellular regulation and several diseases. While it is thought that Rheb GTPase regulates mTOR, acting immediately upstream, while raptor is immediately downstream of mTOR, direct interactions have yet to be verified in living cells, furthermore the localisation of Rheb has been reported to have only a cytoplasmic cellular localization. Results In this study a cytoplasmic as well as a significant sub-cellular nuclear mTOR localization was shown , utilizing green and red fluorescent protein (GFP and DsRed) fusion and highly sensitive single photon counting fluorescence lifetime imaging microscopy (FLIM) of live cells. The interaction of the mTORC1 components Rheb, mTOR and raptor, tagged with EGFP/DsRed was determined using fluorescence energy transfer-FLIM. The excited-state lifetime of EGFP-mTOR of ~2400 ps was reduced by energy transfer to ~2200 ps in the cytoplasm and to 2000 ps in the nucleus when co-expressed with DsRed-Rheb, similar results being obtained for co-expressed EGFP-mTOR and DsRed-raptor. The localization and distribution of mTOR was modified by amino acid withdrawal and re-addition but not by rapamycin. Conclusions The results illustrate the power of GFP-technology combined with FRET-FLIM imaging in the study of the interaction of signalling components in living cells, here providing evidence for a direct physical interaction between mTOR and Rheb and between mTOR and raptor in living cells for the first time. PMID:23311891

  6. Negative regulation of HLA-DR expression on endothelial cells by anti-blood group A/B antibody ligation and mTOR inhibition.

    PubMed

    Iwasaki, Kenta; Miwa, Yuko; Uchida, Kazuharu; Kodera, Yasuhiro; Kobayashi, Takaaki

    2017-02-01

    Donor-specific antibody (DSA), particularly against HLA class II, is a major cause of chronic antibody-mediated rejection (CAMR) after transplantation, although ABO-incompatible kidney transplantation has recently demonstrated favorable graft outcomes. The condition of no injury even in the presence of anti-donor antibody has been referred to as "accommodation", which would be one of the key factors for successful long-term graft survival. The purpose of this study was to analyze the beneficial effect of anti-blood group A/B antibody ligation on endothelial cells against HLA-DR antibody-mediated, complement-dependent cytotoxicity (CDC). Blood group A/B-expressing endothelial cells EA.hy926 or Human Umbilical Vein Endothelia Cells (HUVEC) were incubated with IFNγ in the presence or absence of anti-blood group A/B antibody or mTOR inhibitor (mTOR-i) for 48h. The effects on signaling pathway, HLA expression, complement regulatory factors, and CDC were investigated. Expression of HLA-DR on EA.hy926 or HUVEC were successfully elicited by IFNγ treatment, although little or no expression was observed in quiescent cells. Pre-incubation with anti-blood group A/B antibody had resistance to HLA-DR antibody-mediated CDC against IFNγ-treated cells in a concentration-dependent manner. This finding was ascribed to decreased expression of HLA-DR by post-translational regulation and increased expression of CD55/59, which was related to ERK and mTOR pathway inhibition. mTOR-i also inhibited HLA-DR expression by itself. Furthermore, the combination of mTOR-I and anti-blood group A/B ligation had an additive effect in preventing HLA-DR antibody-mediated CDC. Anti-blood group A/B antibody might play a preventive role in CAMR. Inhibition of the ERK and mTOR pathways may contribute to the development of a novel treatment in the maintenance period after transplantation.

  7. Involvement of PI3K/Akt/GSK-3β and mTOR in the antidepressant-like effect of atorvastatin in mice.

    PubMed

    Ludka, Fabiana Kalyne; Constantino, Leandra Celso; Dal-Cim, Tharine; Binder, Luisa Bandeira; Zomkowski, Andréa; Rodrigues, Ana Lúcia S; Tasca, Carla Inês

    2016-11-01

    Atorvastatin is a cholesterol-lowering statin that has been shown to exert several pleiotropic effects in the nervous system as a neuroprotective and antidepressant-like agent. Antidepressant-like effect of atorvastatin in mice is mediated by glutamatergic and serotoninergic receptors, although the precise intracellular signaling pathways involved are unknown. PI3K/Akt/GSK-3β/mTOR signaling pathway has been associated to neurobiology of depression and seems to be modulated by some pharmacological antidepressant strategies. The present study investigated the participation of the PI3K/Akt/GSK-3β/mTOR signaling pathway in the antidepressant-like effect of an acute atorvastatin treatment in mice. Atorvastatin sub-effective (0.01 mg/kg) or effective (0.1 mg/kg) doses in the tail suspension test (TST) was administered orally alone or in combination with PI3K, GSK-3β or mTOR inhibitors. The administration of PI3K inhibitor, LY294002 (10 nmol/site, i.c.v) completely prevented the antidepressant-like effect of atorvastatin (0.1 mg/kg, p.o.). The participation of GSK-3β in the antidepressant-like effect of atorvastatin was demonstrated by co-administration of a sub-effective dose of atorvastatin (0.01 mg/kg, p.o.) with AR-A014418 (0.01 μg/site, i.c.v., a selective GSK-3β inhibitor) or with lithium chloride (10 mg/kg, p.o., a non-selective GSK-3β inhibitor). The mTOR inhibitor, rapamycin (0.2 nmol/site, i.c.v.) was also able to prevent atorvastatin (0.1 mg/kg, p.o.) antidepressant-like effect. These behavioral findings were supported by neurochemical observations, as atorvastatin treatment increased the immunocontent of the phosphorylated isoforms of Akt, GSK-3β and mTOR in the hippocampus of mice. Taken together, our results suggest an involvement of the PI3K/Akt/GSK-3β/mTOR signaling pathway in the antidepressant-like effect of atorvastatin in mice.

  8. Judicious Toggling of mTOR Activity to Combat Insulin Resistance and Cancer: Current Evidence and Perspectives

    PubMed Central

    Ong, Pei Shi; Wang, Louis Z.; Dai, Xiaoyun; Tseng, Sheng Hsuan; Loo, Shang Jun; Sethi, Gautam

    2016-01-01

    The mechanistic target of rapamycin (mTOR), via its two distinct multiprotein complexes, mTORC1, and mTORC2, plays a central role in the regulation of cellular growth, metabolism, and migration. A dysregulation of the mTOR pathway has in turn been implicated in several pathological conditions including insulin resistance and cancer. Overactivation of mTORC1 and disruption of mTORC2 function have been reported to induce insulin resistance. On the other hand, aberrant mTORC1 and mTORC2 signaling via either genetic alterations or increased expression of proteins regulating mTOR and its downstream targets have contributed to cancer development. These underlined the attractiveness of mTOR as a therapeutic target to overcome both insulin resistance and cancer. This review summarizes the evidence supporting the notion of intermittent, low dose rapamycin for treating insulin resistance. It further highlights recent data on the continuous use of high dose rapamycin analogs and related second generation mTOR inhibitors for cancer eradication, for overcoming chemoresistance and for tumor stem cell suppression. Within these contexts, the potential challenges associated with the use of mTOR inhibitors are also discussed. PMID:27826244

  9. mTOR kinase is needed for the development and stabilization of dendritic arbors in newly born olfactory bulb neurons

    PubMed Central

    Skalecka, Agnieszka; Liszewska, Ewa; Bilinski, Robert; Gkogkas, Christos; Khoutorsky, Arkady; Malik, Anna R.; Sonenberg, Nahum

    2016-01-01

    ABSTRACT Neurogenesis is the process of neuron generation, which occurs not only during embryonic development but also in restricted niches postnatally. One such region is called the subventricular zone (SVZ), which gives rise to new neurons in the olfactory bulb (OB). Neurons that are born postnatally migrate through more complex territories and integrate into fully functional circuits. Therefore, differences in the differentiation of embryonic and postnatally born neurons may exist. Dendritogenesis is an important process for the proper formation of future neuronal circuits. Dendritogenesis in embryonic neurons cultured in vitro was shown to depend on the mammalian target of rapamycin (mTOR). Still unknown, however, is whether mTOR could regulate the dendritic arbor morphology of SVZ‐derived postnatal OB neurons under physiological conditions in vivo. The present study used in vitro cultured and differentiated SVZ‐derived neural progenitors and found that both mTOR complex 1 and mTOR complex 2 were required for the dendritogenesis of SVZ‐derived neurons. Furthermore, using a combination of in vivo electroporation of neural stem cells in the SVZ and genetic and pharmacological inhibition of mTOR, it was found that mTOR was crucial for the growth of basal and apical dendrites in postnatally born OB neurons under physiological conditions and contributed to the stabilization of their basal dendrites. © 2016 Wiley Periodicals, Inc. Develop Neurobiol 76: 1308–1327, 2016 PMID:27008592

  10. BMAL1-dependent regulation of the mTOR signaling pathway delays aging.

    PubMed

    Khapre, Rohini V; Kondratova, Anna A; Patel, Sonal; Dubrovsky, Yuliya; Wrobel, Michelle; Antoch, Marina P; Kondratov, Roman V

    2014-01-01

    The circadian clock, an internal time-keeping system, has been linked with control of aging, but molecular mechanisms of regulation are not known. BMAL1 is a transcriptional factor and core component of the circadian clock; BMAL1 deficiency is associated with premature aging and reduced lifespan. Here we report that activity of mammalian Target of Rapamycin Complex 1 (mTORC1) is increased upon BMAL1 deficiency both in vivo and in cell culture. Increased mTOR signaling is associated with accelerated aging; in accordance with that, treatment with the mTORC1 inhibitor rapamycin increased lifespan of Bmal1-/- mice by 50%. Our data suggest that BMAL1 is a negative regulator of mTORC1 signaling. We propose that the circadian clock controls the activity of the mTOR pathway through BMAL1-dependent mechanisms and this regulation is important for control of aging and metabolism.

  11. Rapalogs and mTOR inhibitors as anti-aging therapeutics

    PubMed Central

    Lamming, Dudley W.; Ye, Lan; Sabatini, David M.; Baur, Joseph A.

    2013-01-01

    Rapamycin, an inhibitor of mechanistic target of rapamycin (mTOR), has the strongest experimental support to date as a potential anti-aging therapeutic in mammals. Unlike many other compounds that have been claimed to influence longevity, rapamycin has been repeatedly tested in long-lived, genetically heterogeneous mice, in which it extends both mean and maximum life spans. However, the mechanism that accounts for these effects is far from clear, and a growing list of side effects make it doubtful that rapamycin would ultimately be beneficial in humans. This Review discusses the prospects for developing newer, safer anti-aging therapies based on analogs of rapamycin (termed rapalogs) or other approaches targeting mTOR signaling. PMID:23454761

  12. Rapalogs and mTOR inhibitors as anti-aging therapeutics.

    PubMed

    Lamming, Dudley W; Ye, Lan; Sabatini, David M; Baur, Joseph A

    2013-03-01

    Rapamycin, an inhibitor of mechanistic target of rapamycin (mTOR), has the strongest experimental support to date as a potential anti-aging therapeutic in mammals. Unlike many other compounds that have been claimed to influence longevity, rapamycin has been repeatedly tested in long-lived, genetically heterogeneous mice, in which it extends both mean and maximum life spans. However, the mechanism that accounts for these effects is far from clear, and a growing list of side effects make it doubtful that rapamycin would ultimately be beneficial in humans. This Review discusses the prospects for developing newer, safer anti-aging therapies based on analogs of rapamycin (termed rapalogs) or other approaches targeting mTOR signaling.

  13. A pilot study of JI-101, an inhibitor of VEGFR-2, PDGFR-β, and EphB4 receptors, in combination with everolimus and as a single agent in an ovarian cancer expansion cohort.

    PubMed

    Werner, Theresa L; Wade, Mark L; Agarwal, Neeraj; Boucher, Kenneth; Patel, Jesal; Luebke, Aaron; Sharma, Sunil

    2015-12-01

    JI-101 is an oral multi-kinase inhibitor that targets vascular endothelial growth factor receptor type 2 (VEGFR-2), platelet derived growth factor receptor β (PDGFR-β), and ephrin type-B receptor 4 (EphB4). None of the currently approved angiogenesis inhibitors have been reported to inhibit EphB4, and therefore, JI-101 has a novel mechanism of action. We conducted a pilot trial to assess the pharmacokinetics (PK), tolerability, and efficacy of JI-101 in combination with everolimus in advanced cancers, and pharmacodynamics (PD), tolerability, and efficacy of JI-101 in ovarian cancer. This was the first clinical study assessing anti-tumor activity of JI-101 in a combinatorial regimen. In the PK cohort, four patients received single agent 10 mg everolimus on day 1, 10 mg everolimus and 200 mg JI-101 combination on day 8, and single agent 200 mg JI-101 on day 15. In the PD cohort, eleven patients received single agent JI-101 at 200 mg twice daily for 28 day treatment cycles. JI-101 was well tolerated as a single agent and in combination with everolimus. No serious adverse events were observed. Common adverse events were hypertension, nausea, and abdominal pain. JI-101 increased exposure of everolimus by approximately 22%, suggestive of drug-drug interaction. The majority of patients had stable disease at their first set of restaging scans (two months), although no patients demonstrated a response to the drug per RECIST criteria. The novel mechanism of action of JI-101 is promising in ovarian cancer treatment and further prospective studies of this agent may be pursued in a less refractory patient population or in combination with cytotoxic chemotherapy.

  14. Muscle wasting associated with the long-term use of mTOR inhibitors

    PubMed Central

    Gyawali, Bishal; Shimokata, Tomoya; Honda, Kazunori; Kondoh, Chihiro; Hayashi, Naomi; Yoshino, Yasushi; Sassa, Naoto; Nakano, Yasuyuki; Gotoh, Momokazu; Ando, Yuichi

    2016-01-01

    Some targeted therapies alter muscle mass due to interference with pathways of muscle metabolism. The effects of mammalian target of ra pamycin (mTOR) inhibitors on muscle mass have yet to be fully elucidated. In the present study, the computerized tomography (CT) scans of patients receiving mTOR inhibitors for at least 6 months taken at baseline and post-therapy were retrospectively retrieved, and body composition analyses were performed using the software, sliceOmatic version 5.0 (TomoVision, Inc., Magog, QC, Canada). The difference in body composition parameters was evaluated for significance. The time to treatment (TTF) failure was also compared between the sarcopenic and non-sarcopenic patients at the baseline. Of the 75 patients studied, 20 matched the inclusion criteria (including 16 males). The mean duration between the CT scans was 14.4±2.0 months. A total of 12 (60%) patients were sarcopenic at the baseline, whereas three more (75% in total) became sarcopenic following treatment. The use of mTOR inhibitors significantly decreased the skeletal muscle area (P=0.011) and lean body mass (P=0.007), although it had no effect on adipose tissue (P=0.163) or body weight (P=0.262). The rate of skeletal muscle wasting was 2.6 cm2/m2, or 2.3 kg in 6 months. The TTF did not differ between sarcopenic and non-sarcopenic patients, and was not significantly associated with any other parameter. To the best of our knowledge, this is the first study to demonstrate that the long-term use of mTOR inhibitors induces a marked loss of muscle mass. Due to the predictive and prognostic role of sarcopenia in cancer patients, these findings may have important clinical implications. PMID:27900103

  15. Putrescine stimulates the mTOR signaling pathway and protein synthesis in porcine trophectoderm cells.

    PubMed

    Kong, Xiangfeng; Wang, Xiaoqiu; Yin, Yulong; Li, Xilong; Gao, Haijun; Bazer, Fuller W; Wu, Guoyao

    2014-11-01

    Insufficient placental growth is a major factor contributing to intrauterine growth retardation in mammals. There is growing evidence that putrescine produced from arginine (Arg) and proline via ornithine decarboxylase is a key regulator of angiogenesis, embryogenesis, as well as placental and fetal growth. However, the underlying mechanisms are largely unknown. The present study tested the hypothesis that putrescine stimulates protein synthesis by activating the mechanistic target of rapamycin (mTOR) signaling pathway in porcine trophectoderm cell line 2 cells. The cells were cultured for 2 to 4 days in customized Arg-free Dulbecco modified Eagle Ham medium containing 0, 10, 25, or 50 μM putrescine or 100 μM Arg. Cell proliferation, protein synthesis, and degradation, as well as the abundance of total and phosphorylated mTOR, ribosomal protein S6 kinase 1, and eukaryotic initiation factor 4E-binding protein-1 (4EBP1), were determined. Our results indicate that putrescine promotes cell proliferation and protein synthesis in a dose- and time-dependent manner, which was inhibited by difluoro-methylornithine (an inhibitor of ornithine decarboxylase). Moreover, supplementation of culture medium with putrescine increased the abundance of phosphorylated mTOR and its downstream targets, 4EBP1 and p70 S6K1 proteins. Collectively, these findings reveal a novel and important role for putrescine in regulating the mTOR signaling pathway in porcine placental cells. We suggest that dietary supplementation with or intravenous administration of putrescine may provide a new and effective strategy to improve survival and growth of embryos/fetuses in mammals.

  16. Akt- or MEK-mediated mTOR inhibition suppresses Nf1 optic glioma growth

    PubMed Central

    Kaul, Aparna; Toonen, Joseph A.; Cimino, Patrick J.; Gianino, Scott M.; Gutmann, David H.

    2015-01-01

    Background Children with neurofibromatosis type 1 (NF1) develop optic pathway gliomas, which result from impaired NF1 protein regulation of Ras activity. One obstacle to the implementation of biologically targeted therapies is an incomplete understanding of the individual contributions of the downstream Ras effectors (mitogen-activated protein kinase kinase [MEK], Akt) to optic glioma maintenance. This study was designed to address the importance of MEK and Akt signaling to Nf1 optic glioma growth. Methods Primary neonatal mouse astrocyte cultures were employed to determine the consequence of phosphatidylinositol-3 kinase (PI3K)/Akt and MEK inhibition on Nf1-deficient astrocyte growth. Nf1 optic glioma–bearing mice were used to assess the effect of Akt and MEK inhibition on tumor volume, proliferation, and retinal ganglion cell dysfunction. Results Both MEK and Akt were hyperactivated in Nf1-deficient astrocytes in vitro and in Nf1 murine optic gliomas in vivo. Pharmacologic PI3K or Akt inhibition reduced Nf1-deficient astrocyte proliferation to wild-type levels, while PI3K inhibition decreased Nf1 optic glioma volume and proliferation. Akt inhibition of Nf1-deficient astrocyte and optic glioma growth reflected Akt-dependent activation of mammalian target of rapamycin (mTOR). Sustained MEK pharmacologic blockade also attenuated Nf1-deficient astrocytes as well as Nf1 optic glioma volume and proliferation. Importantly, these MEK inhibitory effects resulted from p90RSK-mediated, Akt-independent mTOR activation. Finally, both PI3K and MEK inhibition reduced optic glioma–associated retinal ganglion cell loss and nerve fiber layer thinning. Conclusion These findings establish that the convergence of 2 distinct Ras effector pathways on mTOR signaling maintains Nf1 mouse optic glioma growth, supporting the evaluation of pharmacologic inhibitors that target mTOR function in future human NF1–optic pathway glioma clinical trials. PMID:25534823

  17. Stoichiometry and assembly of mTOR complexes revealed by single-molecule pulldown

    PubMed Central

    Jain, Ankur; Arauz, Edwin; Aggarwal, Vasudha; Ikon, Nikita; Chen, Jie; Ha, Taekjip

    2014-01-01

    The mammalian target of rapamycin (mTOR) kinase is a master regulator of cellular, developmental, and metabolic processes. Deregulation of mTOR signaling is implicated in numerous human diseases including cancer and diabetes. mTOR functions as part of either of the two multisubunit complexes, mTORC1 and mTORC2, but molecular details about the assembly and oligomerization of mTORCs are currently lacking. We use the single-molecule pulldown (SiMPull) assay that combines principles of conventional pulldown assays with single-molecule fluorescence microscopy to investigate the stoichiometry and assembly of mTORCs. After validating our approach with mTORC1, confirming a dimeric assembly as previously reported, we show that all major components of mTORC2 exist in two copies per complex, indicating that mTORC2 assembles as a homodimer. Interestingly, each mTORC component, when free from the complexes, is present as a monomer and no single subunit serves as the dimerizing component. Instead, our data suggest that dimerization of mTORCs is the result of multiple subunits forming a composite surface. SiMPull also allowed us to distinguish complex disassembly from stoichiometry changes. Physiological conditions that abrogate mTOR signaling such as nutrient deprivation or energy stress did not alter the stoichiometry of mTORCs. On the other hand, rapamycin treatment leads to transient appearance of monomeric mTORC1 before complete disruption of the mTOR–raptor interaction, whereas mTORC2 stoichiometry is unaffected. These insights into assembly of mTORCs may guide future mechanistic studies and exploration of therapeutic potential. PMID:25453101

  18. Stoichiometry and assembly of mTOR complexes revealed by single-molecule pulldown.

    PubMed

    Jain, Ankur; Arauz, Edwin; Aggarwal, Vasudha; Ikon, Nikita; Chen, Jie; Ha, Taekjip

    2014-12-16

    The mammalian target of rapamycin (mTOR) kinase is a master regulator of cellular, developmental, and metabolic processes. Deregulation of mTOR signaling is implicated in numerous human diseases including cancer and diabetes. mTOR functions as part of either of the two multisubunit complexes, mTORC1 and mTORC2, but molecular details about the assembly and oligomerization of mTORCs are currently lacking. We use the single-molecule pulldown (SiMPull) assay that combines principles of conventional pulldown assays with single-molecule fluorescence microscopy to investigate the stoichiometry and assembly of mTORCs. After validating our approach with mTORC1, confirming a dimeric assembly as previously reported, we show that all major components of mTORC2 exist in two copies per complex, indicating that mTORC2 assembles as a homodimer. Interestingly, each mTORC component, when free from the complexes, is present as a monomer and no single subunit serves as the dimerizing component. Instead, our data suggest that dimerization of mTORCs is the result of multiple subunits forming a composite surface. SiMPull also allowed us to distinguish complex disassembly from stoichiometry changes. Physiological conditions that abrogate mTOR signaling such as nutrient deprivation or energy stress did not alter the stoichiometry of mTORCs. On the other hand, rapamycin treatment leads to transient appearance of monomeric mTORC1 before complete disruption of the mTOR-raptor interaction, whereas mTORC2 stoichiometry is unaffected. These insights into assembly of mTORCs may guide future mechanistic studies and exploration of therapeutic potential.

  19. Regulation of mTOR activity in Snell dwarf and GH receptor gene-disrupted mice.

    PubMed

    Dominick, Graham; Berryman, Darlene E; List, Edward O; Kopchick, John J; Li, Xinna; Miller, Richard A; Garcia, Gonzalo G

    2015-02-01

    The involvement of mammalian target of rapamycin (mTOR) in lifespan control in invertebrates, calorie-restricted rodents, and extension of mouse lifespan by rapamycin have prompted speculation that diminished mTOR function may contribute to mammalian longevity in several settings. We show here that mTOR complex-1 (mTORC1) activity is indeed lower in liver, muscle, heart, and kidney tissue of Snell dwarf and global GH receptor (GHR) gene-disrupted mice (GHR-/-), consistent with previous studies. Surprisingly, activity of mTORC2 is higher in fasted Snell and GHR-/- than in littermate controls in all 4 tissues tested. Resupply of food enhanced mTORC1 activity in both controls and long-lived mutant mice but diminished mTORC2 activity only in the long-lived mice. Mice in which GHR has been disrupted only in the liver do not show extended lifespan and also fail to show the decline in mTORC1 and increase in mTORC2 seen in mice with global loss of GHR. The data suggest that the antiaging effects in the Snell dwarf and GHR-/- mice are accompanied by both a decline in mTORC1 in multiple organs and an increase in fasting levels of mTORC2. Neither the lifespan nor mTOR effects appear to be mediated by direct GH effects on liver or by the decline in plasma IGF-I, a shared trait in both global and liver-specific GHR-/- mice. Our data suggest that a more complex pattern of hormonal effects and intertissue interactions may be responsible for regulating both lifespan and mTORC2 function in these mouse models of delayed aging.

  20. mTOR kinase inhibitors promote antibody class switching via mTORC2 inhibition.

    PubMed

    Limon, Jose J; So, Lomon; Jellbauer, Stefan; Chiu, Honyin; Corado, Juana; Sykes, Stephen M; Raffatellu, Manuela; Fruman, David A

    2014-11-25

    The mammalian target of rapamycin (mTOR) is a kinase that functions in two distinct complexes, mTORC1 and mTORC2. In peripheral B cells, complete deletion of mTOR suppresses germinal center B-cell responses, including class switching and somatic hypermutation. The allosteric mTORC1 inhibitor rapamycin blocks proliferation and differentiation, but lower doses can promote protective IgM responses. To elucidate the complexity of mTOR signaling in B cells further, we used ATP-competitive mTOR kinase inhibitors (TOR-KIs), which inhibit both mTORC1 and mTORC2. Although TOR-KIs are in clinical development for cancer, their effects on mature lymphocytes are largely unknown. We show that high concentrations of TOR-KIs suppress B-cell proliferation and differentiation, yet lower concentrations that preserve proliferation increase the fraction of B cells undergoing class switching in vitro. Transient treatment of mice with the TOR-KI compound AZD8055 increased titers of class-switched high-affinity antibodies to a hapten-protein conjugate. Mechanistic investigation identified opposing roles for mTORC1 and mTORC2 in B-cell differentiation and showed that TOR-KIs enhance class switching in a manner dependent on forkhead box, subgroup O (FoxO) transcription factors. These observations emphasize the distinct actions of TOR-KIs compared with rapamycin and suggest that TOR-KIs might be useful to enhance production of class-switched antibodies following vaccination.

  1. PML nuclear bodies contribute to the basal expression of the mTOR inhibitor DDIT4

    PubMed Central

    Salsman, Jayme; Stathakis, Alex; Parker, Ellen; Chung, Dudley; Anthes, Livia E.; Koskowich, Kara L.; Lahsaee, Sara; Gaston, Daniel; Kukurba, Kimberly R.; Smith, Kevin S.; Chute, Ian C.; Léger, Daniel; Frost, Laura D.; Montgomery, Stephen B.; Lewis, Stephen M.; Eskiw, Christopher; Dellaire, Graham

    2017-01-01

    The promyelocytic leukemia (PML) protein is an essential component of PML nuclear bodies (PML NBs) frequently lost in cancer. PML NBs coordinate chromosomal regions via modification of nuclear proteins that in turn may regulate genes in the vicinity of these bodies. However, few PML NB-associated genes have been identified. PML and PML NBs can also regulate mTOR and cell fate decisions in response to cellular stresses. We now demonstrate that PML depletion in U2OS cells or TERT-immortalized normal human diploid fibroblasts results in decreased expression of the mTOR inhibitor DDIT4 (REDD1). DNA and RNA immuno-FISH reveal that PML NBs are closely associated with actively transcribed DDIT4 loci, implicating these bodies in regulation of basal DDIT4 expression. Although PML silencing did reduce the sensitivity of U2OS cells to metabolic stress induced by metformin, PML loss did not inhibit the upregulation of DDIT4 in response to metformin, hypoxia-like (CoCl2) or genotoxic stress. Analysis of publicly available cancer data also revealed a significant correlation between PML and DDIT4 expression in several cancer types (e.g. lung, breast, prostate). Thus, these findings uncover a novel mechanism by which PML loss may contribute to mTOR activation and cancer progression via dysregulation of basal DDIT4 gene expression. PMID:28332630

  2. mTOR target NDRG1 confers MGMT-dependent resistance to alkylating chemotherapy.

    PubMed

    Weiler, Markus; Blaes, Jonas; Pusch, Stefan; Sahm, Felix; Czabanka, Marcus; Luger, Sebastian; Bunse, Lukas; Solecki, Gergely; Eichwald, Viktoria; Jugold, Manfred; Hodecker, Sibylle; Osswald, Matthias; Meisner, Christoph; Hielscher, Thomas; Rübmann, Petra; Pfenning, Philipp-Niklas; Ronellenfitsch, Michael; Kempf, Tore; Schnölzer, Martina; Abdollahi, Amir; Lang, Florian; Bendszus, Martin; von Deimling, Andreas; Winkler, Frank; Weller, Michael; Vajkoczy, Peter; Platten, Michael; Wick, Wolfgang

    2014-01-07

    A hypoxic microenvironment induces resistance to alkylating agents by activating targets in the mammalian target of rapamycin (mTOR) pathway. The molecular mechanisms involved in this mTOR-mediated hypoxia-induced chemoresistance, however, are unclear. Here we identify the mTOR target N-myc downstream regulated gene 1 (NDRG1) as a key determinant of resistance toward alkylating chemotherapy, driven by hypoxia but also by therapeutic measures such as irradiation, corticosteroids, and chronic exposure to alkylating agents via distinct molecular routes involving hypoxia-inducible factor (HIF)-1alpha, p53, and the mTOR complex 2 (mTORC2)/serum glucocorticoid-induced protein kinase 1 (SGK1) pathway. Resistance toward alkylating chemotherapy but not radiotherapy was dependent on NDRG1 expression and activity. In posttreatment tumor tissue of patients with malignant gliomas, NDRG1 was induced and predictive of poor response to alkylating chemotherapy. On a molecular level, NDRG1 bound and stabilized methyltransferases, chiefly O(6)-methylguanine-DNA methyltransferase (MGMT), a key enzyme for resistance to alkylating agents in glioblastoma patients. In patients with glioblastoma, MGMT promoter methylation in tumor tissue was not more predictive for response to alkylating chemotherapy in patients who received concomitant corticosteroids.

  3. Reciprocal regulation of cilia and autophagy via the MTOR and proteasome pathways.

    PubMed

    Wang, Shixuan; Livingston, Man J; Su, Yunchao; Dong, Zheng

    2015-04-03

    Primary cilium is an organelle that plays significant roles in a number of cellular functions ranging from cell mechanosensation, proliferation, and differentiation to apoptosis. Autophagy is an evolutionarily conserved cellular function in biology and indispensable for cellular homeostasis. Both cilia and autophagy have been linked to different types of genetic and acquired human diseases. Their interaction has been suggested very recently, but the underlying mechanisms are still not fully understood. We examined autophagy in cells with suppressed cilia and measured cilium length in autophagy-activated or -suppressed cells. It was found that autophagy was repressed in cells with short cilia. Further investigation showed that MTOR activation was enhanced in cilia-suppressed cells and the MTOR inhibitor rapamycin could largely reverse autophagy suppression. In human kidney proximal tubular cells (HK2), autophagy induction was associated with cilium elongation. Conversely, autophagy inhibition by 3-methyladenine (3-MA) and chloroquine (CQ) as well as bafilomycin A1 (Baf) led to short cilia. Cilia were also shorter in cultured atg5-knockout (KO) cells and in atg7-KO kidney proximal tubular cells in mice. MG132, an inhibitor of the proteasome, could significantly restore cilium length in atg5-KO cells, being concomitant with the proteasome activity. Together, the results suggest that cilia and autophagy regulate reciprocally through the MTOR signaling pathway and ubiquitin-proteasome system.

  4. From mTOR to cognition: molecular and cellular mechanisms of cognitive impairments in tuberous sclerosis

    PubMed Central

    Ehninger, D.; de Vries, P. J.; Silva, A. J.

    2010-01-01

    Background Tuberous sclerosis (TSC) is a multi-system disorder caused by heterozygous mutations in the TSC1 or TSC2 gene and is often associated with neuropsychiatric symptoms, including intellectual disability, specific neuropsychological deficits, autism, other behavioural disorders and epilepsy. Method Here, we review evidence from animal models of TSC for the role of specific molecular and cellular processes in the pathogenesis of cognitive, developmental and epilepsy-related manifestations seen in the disorder. Results Recent evidence shows that, in animal models, disinhibited mTOR (mammalian target of rapamycin) signalling substantially contributes to neuropsychiatric phenotypes, including cognitive deficits and seizures. We discuss potential pathogenetic mechanisms involved in the cognitive phenotypes of TSC and present implications regarding mTOR inhibitor-based treatments for TSC-related neuropsychiatric features. Conclusions Results suggest that reversing the underlying molecular deficits of TSC with rapamycin or other mTOR inhibitors could result in clinically significant improvements of cognitive function and neurological symptoms, even if treatments are started in adulthood. PMID:19694899

  5. Baicalein inhibits lipid accumulation by regulating early adipogenesis and m-TOR signaling.

    PubMed

    Seo, Min-Jung; Choi, Hyeon-Son; Jeon, Hui-Jeon; Woo, Mi-Seon; Lee, Boo-Yong

    2014-05-01

    Baicalein is a type of flavonoid that originates from Scutellaria baicalensis. In this study, we examined how baicalein inhibits lipid accumulation during adipogenesis in 3T3-L1 cells. Our data show that baicalein inhibited lipid accumulation during adipogenesis in a dose-dependent manner. Baicalein inhibition was limited to the early adipogenic stage. Cell cycle analysis showed that baicalein induced cell cycle arrest in the G0/G1 phase through cyclin downregulation. In addition, baicalein suppressed the mRNA expression of early adipogenic factors leading to downregulation of late adipogenic factors at mRNA and protein levels. Inhibition of adipogenic factors by baicalein was correlated with downregulation of lipid synthetic enzymes. Additionally, baicalein negatively regulated the m-TOR signaling pathway involved in lipid accumulation during adipogenesis, thus inhibiting phosphorylation of m-TOR and p70S6K. In a zebrafish study, baicalein significantly reduced lipid accumulation in Nile Red staining. Consistent with a report using cell lines, mRNA expression of adipogenic factors was decreased in a dose-dependent manner by baicalein. This result reflects a reduction in total triglyceride levels based on a triglyceride assay. Our data suggest that baicalein inhibits lipid accumulation by controlling the cell cycle and m-TOR signaling in 3T3-L1 cells, and its anti-adipogenic effect was found in a zebrafish model.

  6. mTOR is critical for intestinal T-cell homeostasis and resistance to Citrobacter rodentium

    PubMed Central

    Lin, Xingguang; Yang, Jialong; Wang, Jinli; Huang, Hongxiang; Wang, Hong-Xia; Chen, Pengcheng; Wang, Shang; Pan, Yun; Qiu, Yu-Rong; Taylor, Gregory A.; Vallance, Bruce A.; Gao, Jimin; Zhong, Xiao-Ping

    2016-01-01

    T-cells play an important role in promoting mucosal immunity against pathogens, but the mechanistic basis for their homeostasis in the intestine is still poorly understood. We report here that T-cell-specific deletion of mTOR results in dramatically decreased CD4 and CD8 T-cell numbers in the lamina propria of both small and large intestines under both steady-state and inflammatory conditions. These defects result in defective host resistance against a murine enteropathogen, Citrobacter rodentium, leading to the death of the animals. We further demonstrated that mTOR deficiency reduces the generation of gut-homing effector T-cells in both mesenteric lymph nodes and Peyer’s patches without obviously affecting expression of gut-homing molecules on those effector T-cells. Using mice with T-cell-specific ablation of Raptor/mTORC1 or Rictor/mTORC2, we revealed that both mTORC1 and, to a lesser extent, mTORC2 contribute to both CD4 and CD8 T-cell accumulation in the gastrointestinal (GI) tract. Additionally, mTORC1 but not mTORC2 plays an important role regulating the proliferative renewal of both CD4 and CD8 T-cells in the intestines. Our data thus reveal that mTOR is crucial for T-cell accumulation in the GI tract and for establishing local adaptive immunity against pathogens. PMID:27731345

  7. mTOR regulate EMT through RhoA and Rac1 pathway in prostate cancer.

    PubMed

    Chen, XianGuo; Cheng, HaiYan; Pan, TengFei; Liu, Yi; Su, Yang; Ren, CuiPing; Huang, DaKe; Zha, XiaoJun; Liang, ChaoZhao

    2015-10-01

    Recently, an increasing number of studies have suggested that mTOR plays a critical role in the regulation of tumor cell motility, invasion and cancer metastasis. However, little is known about the signaling mechanisms in regulating epithelial-mesenchymal transition (EMT) of prostate cancer. In this study, we found that the expression levels of Raptor and Rictor in prostate cancer tissues were elevated, which may suggest that Raptor and Rictor signaling pathways are associated with prostate cancer progression and metastasis. Inhibition of mTORC1 or mTORC2 by knock down of Raptor or Rictor, respectively, migration and invasion of prostate cancer were attenuated. Furthermore, EMT, a characterized by the changed expression levels of various markers like E-cadherin, β-catenin, N-cadherin, and vimentin emergend following inhibition of Raptor or Rictor. Finally, the small GTPases (RhoA and Rac1) which were crucial regulatory proteins in cell migration and invasion were inactivited after downregulating Raptor and Rictor. These results suggest that mTOR regulate EMT at least in part by down regulation of RhoA and Rac1 signaling pathways. Our findings provide novel very attractive target strategies that the inhibition of mTOR signaling pathways may retard prostate cancer migration and invasion at early stages.

  8. PML nuclear bodies contribute to the basal expression of the mTOR inhibitor DDIT4.

    PubMed

    Salsman, Jayme; Stathakis, Alex; Parker, Ellen; Chung, Dudley; Anthes, Livia E; Koskowich, Kara L; Lahsaee, Sara; Gaston, Daniel; Kukurba, Kimberly R; Smith, Kevin S; Chute, Ian C; Léger, Daniel; Frost, Laura D; Montgomery, Stephen B; Lewis, Stephen M; Eskiw, Christopher; Dellaire, Graham

    2017-03-23

    The promyelocytic leukemia (PML) protein is an essential component of PML nuclear bodies (PML NBs) frequently lost in cancer. PML NBs coordinate chromosomal regions via modification of nuclear proteins that in turn may regulate genes in the vicinity of these bodies. However, few PML NB-associated genes have been identified. PML and PML NBs can also regulate mTOR and cell fate decisions in response to cellular stresses. We now demonstrate that PML depletion in U2OS cells or TERT-immortalized normal human diploid fibroblasts results in decreased expression of the mTOR inhibitor DDIT4 (REDD1). DNA and RNA immuno-FISH reveal that PML NBs are closely associated with actively transcribed DDIT4 loci, implicating these bodies in regulation of basal DDIT4 expression. Although PML silencing did reduce the sensitivity of U2OS cells to metabolic stress induced by metformin, PML loss did not inhibit the upregulation of DDIT4 in response to metformin, hypoxia-like (CoCl2) or genotoxic stress. Analysis of publicly available cancer data also revealed a significant correlation between PML and DDIT4 expression in several cancer types (e.g. lung, breast, prostate). Thus, these findings uncover a novel mechanism by which PML loss may contribute to mTOR activation and cancer progression via dysregulation of basal DDIT4 gene expression.

  9. mTOR Signaling and Entrainment of the Mammalian Circadian Clock.

    PubMed

    Cao, Ruifeng; Obrietan, Karl

    2010-01-01

    The biochemistry, physiology and behavior of nearly all organisms are influenced by an inherent circadian (24 hr) clock timing mechanism. For mammals, the linchpin of this biological timing process is located in the suprachiasmatic nuclei (SCN) of the hypothalamus. One key feature of the SCN clock is that it is tightly entrained to lighting cues, thus ensuring that the clock is synchronized to the ever-changing seasonal light cycle. Within the field of circadian biology, there has been intense interest in understanding the intracellular signaling events that drive this process. To this end, our recent studies have revealed a role for an evolutionarily conserved translational control kinase, the mammalian target of rapamycin (mTOR), in the SCN clock entrainment process. Here we provide an overview of mechanisms of inducible mTOR activation in the SCN, and describe the effects of mTOR on clock protein synthesis and behavioral rhythmicity. Given that dysregulation of SCN timing has been associated with an array of clinical conditions (e.g., hypertension, obesity, diabetes, depression), new insights into the molecular mechanisms that regulate clock timing may provide new therapeutic treatments for circadian rhythm-associated disorders.

  10. FK866-induced NAMPT inhibition activates AMPK and downregulates mTOR signaling in hepatocarcinoma cells

    SciTech Connect

    Schuster, Susanne; Penke, Melanie; Gorski, Theresa; Gebhardt, Rolf; Weiss, Thomas S.; Kiess, Wieland; Garten, Antje

    2015-03-06

    Background: Nicotinamide phosphoribosyltransferase (NAMPT) is the key enzyme of the NAD salvage pathway starting from nicotinamide. Cancer cells have an increased demand for NAD due to their high proliferation and DNA repair rate. Consequently, NAMPT is considered as a putative target for anti-cancer therapies. There is evidence that AMP-activated protein kinase (AMPK) and mammalian target of rapamycin (mTOR) become dysregulated during the development of hepatocellular carcinoma (HCC). Here, we investigated the effects of NAMPT inhibition by its specific inhibitor FK866 on the viability of hepatocarcinoma cells and analyzed the effects of FK866 on the nutrient sensor AMPK and mTOR complex1 (mTORC1) signaling. Results: FK866 markedly decreased NAMPT activity and NAD content in hepatocarcinoma cells (Huh7 cells, Hep3B cells) and led to delayed ATP reduction which was associated with increased cell death. These effects could be abrogated by administration of nicotinamide mononucleotide (NMN), the enzyme product of NAMPT. Our results demonstrated a dysregulation of the AMPK/mTOR pathway in hepatocarcinoma cells compared to non-cancerous hepatocytes with a higher expression of mTOR and a lower AMPKα activation in hepatocarcinoma cells. We found that NAMPT inhibition by FK866 significantly activated AMPKα and inhibited the activation of mTOR and its downstream targets p70S6 kinase and 4E-BP1 in hepatocarcinoma cells. Non-cancerous hepatocytes were less sensitive to FK866 and did not show changes in AMPK/mTOR signaling after FK866 treatment. Conclusion: Taken together, these findings reveal an important role of the NAMPT-mediated NAD salvage pathway in the energy homeostasis of hepatocarcinoma cells and suggest NAMPT inhibition as a potential treatment option for HCC. - Highlights: • FK866 increases cell death in p53-deficient hepatocarcinoma cells. • AMPK/mTOR signaling is dysregulated in hepatocarcinoma cells. • FK866-induced NAMPT inhibition activates AMPK

  11. Inhibitory Effect of mTOR Activator MHY1485 on Autophagy: Suppression of Lysosomal Fusion

    PubMed Central

    Choi, Yeon Ja; Park, Yun Jung; Park, Ji Young; Jeong, Hyoung Oh; Kim, Dae Hyun; Ha, Young Mi; Kim, Ji Min; Song, Yu Min; Heo, Hyoung-Sam; Yu, Byung Pal; Chun, Pusoon; Moon, Hyung Ryong; Chung, Hae Young

    2012-01-01

    Autophagy is a major degradative process responsible for the disposal of cytoplasmic proteins and dysfunctional organelles via the lysosomal pathway. During the autophagic process, cells form double-membraned vesicles called autophagosomes that sequester disposable materials in the cytoplasm and finally fuse with lysosomes. In the present study, we investigated the inhibition of autophagy by a synthesized compound, MHY1485, in a culture system by using Ac2F rat hepatocytes. Autophagic flux was measured to evaluate the autophagic activity. Autophagosomes were visualized in Ac2F cells transfected with AdGFP-LC3 by live-cell confocal microscopy. In addition, activity of mTOR, a major regulatory protein of autophagy, was assessed by western blot and docking simulation using AutoDock 4.2. In the result, treatment with MHY1485 suppressed the basal autophagic flux, and this inhibitory effect was clearly confirmed in cells under starvation, a strong physiological inducer of autophagy. The levels of p62 and beclin-1 did not show significant change after treatment with MHY1485. Decreased co-localization of autophagosomes and lysosomes in confocal microscopic images revealed the inhibitory effect of MHY1485 on lysosomal fusion during starvation-induced autophagy. These effects of MHY1485 led to the accumulation of LC3II and enlargement of the autophagosomes in a dose- and time- dependent manner. Furthermore, MHY1485 induced mTOR activation and correspondingly showed a higher docking score than PP242, a well-known ATP-competitive mTOR inhibitor, in docking simulation. In conclusion, MHY1485 has an inhibitory effect on the autophagic process by inhibition of fusion between autophagosomes and lysosomes leading to the accumulation of LC3II protein and enlarged autophagosomes. MHY1485 also induces mTOR activity, providing a possibility for another regulatory mechanism of autophagy by the MHY compound. The significance of this study is the finding of a novel inhibitor of autophagy

  12. Inhibitory effect of mTOR activator MHY1485 on autophagy: suppression of lysosomal fusion.

    PubMed

    Choi, Yeon Ja; Park, Yun Jung; Park, Ji Young; Jeong, Hyoung Oh; Kim, Dae Hyun; Ha, Young Mi; Kim, Ji Min; Song, Yu Min; Heo, Hyoung-Sam; Yu, Byung Pal; Chun, Pusoon; Moon, Hyung Ryong; Chung, Hae Young

    2012-01-01

    Autophagy is a major degradative process responsible for the disposal of cytoplasmic proteins and dysfunctional organelles via the lysosomal pathway. During the autophagic process, cells form double-membraned vesicles called autophagosomes that sequester disposable materials in the cytoplasm and finally fuse with lysosomes. In the present study, we investigated the inhibition of autophagy by a synthesized compound, MHY1485, in a culture system by using Ac2F rat hepatocytes. Autophagic flux was measured to evaluate the autophagic activity. Autophagosomes were visualized in Ac2F cells transfected with AdGFP-LC3 by live-cell confocal microscopy. In addition, activity of mTOR, a major regulatory protein of autophagy, was assessed by western blot and docking simulation using AutoDock 4.2. In the result, treatment with MHY1485 suppressed the basal autophagic flux, and this inhibitory effect was clearly confirmed in cells under starvation, a strong physiological inducer of autophagy. The levels of p62 and beclin-1 did not show significant change after treatment with MHY1485. Decreased co-localization of autophagosomes and lysosomes in confocal microscopic images revealed the inhibitory effect of MHY1485 on lysosomal fusion during starvation-induced autophagy. These effects of MHY1485 led to the accumulation of LC3II and enlargement of the autophagosomes in a dose- and time-dependent manner. Furthermore, MHY1485 induced mTOR activation and correspondingly showed a higher docking score than PP242, a well-known ATP-competitive mTOR inhibitor, in docking simulation. In conclusion, MHY1485 has an inhibitory effect on the autophagic process by inhibition of fusion between autophagosomes and lysosomes leading to the accumulation of LC3II protein and enlarged autophagosomes. MHY1485 also induces mTOR activity, providing a possibility for another regulatory mechanism of autophagy by the MHY compound. The significance of this study is the finding of a novel inhibitor of autophagy

  13. BIM and mTOR expression levels predict outcome to erlotinib in EGFR-mutant non-small-cell lung cancer

    PubMed Central

    Karachaliou, Niki; Codony-Servat, Jordi; Teixidó, Cristina; Pilotto, Sara; Drozdowskyj, Ana; Codony-Servat, Carles; Giménez-Capitán, Ana; Molina-Vila, Miguel Angel; Bertrán-Alamillo, Jordi; Gervais, Radj; Massuti, Bartomeu; Morán, Teresa; Majem, Margarita; Felip, Enriqueta; Carcereny, Enric; García-Campelo, Rosario; Viteri, Santiago; González-Cao, María; Morales-Espinosa, Daniela; Verlicchi, Alberto; Crisetti, Elisabetta; Chaib, Imane; Santarpia, Mariacarmela; Luis Ramírez, José; Bosch-Barrera, Joaquim; Felipe Cardona, Andrés; de Marinis, Filippo; López-Vivanco, Guillermo; Miguel Sánchez, José; Vergnenegre, Alain; Sánchez Hernández, José Javier; Sperduti, Isabella; Bria, Emilio; Rosell, Rafael

    2015-01-01

    BIM is a proapoptotic protein that initiates apoptosis triggered by EGFR tyrosine kinase inhibitors (TKI). mTOR negatively regulates apoptosis and may influence response to EGFR TKI. We examined mRNA expression of BIM and MTOR in 57 patients with EGFR-mutant NSCLC from the EURTAC trial. Risk of mortality and disease progression was lower in patients with high BIM compared with low/intermediate BIM mRNA levels. Analysis of MTOR further divided patients with high BIM expression into two groups, with those having both high BIM and MTOR experiencing shorter overall and progression-free survival to erlotinib. Validation of our results was performed in an independent cohort of 19 patients with EGFR-mutant NSCLC treated with EGFR TKIs. In EGFR-mutant lung adenocarcinoma cell lines with high BIM expression, concomitant high mTOR expression increased IC50 of gefitinib for cell proliferation. We next sought to analyse the signalling pattern in cell lines with strong activation of mTOR and its substrate P-S6. We showed that mTOR and phosphodiesterase 4D (PDE4D) strongly correlate in resistant EGFR-mutant cancer cell lines. These data suggest that the combination of EGFR TKI with mTOR or PDE4 inhibitors could be adequate therapy for EGFR-mutant NSCLC patients with high pretreatment levels of BIM and mTOR. PMID:26639561

  14. BIM and mTOR expression levels predict outcome to erlotinib in EGFR-mutant non-small-cell lung cancer.

    PubMed

    Karachaliou, Niki; Codony-Servat, Jordi; Teixidó, Cristina; Pilotto, Sara; Drozdowskyj, Ana; Codony-Servat, Carles; Giménez-Capitán, Ana; Molina-Vila, Miguel Angel; Bertrán-Alamillo, Jordi; Gervais, Radj; Massuti, Bartomeu; Morán, Teresa; Majem, Margarita; Felip, Enriqueta; Carcereny, Enric; García-Campelo, Rosario; Viteri, Santiago; González-Cao, María; Morales-Espinosa, Daniela; Verlicchi, Alberto; Crisetti, Elisabetta; Chaib, Imane; Santarpia, Mariacarmela; Luis Ramírez, José; Bosch-Barrera, Joaquim; Felipe Cardona, Andrés; de Marinis, Filippo; López-Vivanco, Guillermo; Miguel Sánchez, José; Vergnenegre, Alain; Sánchez Hernández, José Javier; Sperduti, Isabella; Bria, Emilio; Rosell, Rafael

    2015-12-07

    BIM is a proapoptotic protein that initiates apoptosis triggered by EGFR tyrosine kinase inhibitors (TKI). mTOR negatively regulates apoptosis and may influence response to EGFR TKI. We examined mRNA expression of BIM and MTOR in 57 patients with EGFR-mutant NSCLC from the EURTAC trial. Risk of mortality and disease progression was lower in patients with high BIM compared with low/intermediate BIM mRNA levels. Analysis of MTOR further divided patients with high BIM expression into two groups, with those having both high BIM and MTOR experiencing shorter overall and progression-free survival to erlotinib. Validation of our results was performed in an independent cohort of 19 patients with EGFR-mutant NSCLC treated with EGFR TKIs. In EGFR-mutant lung adenocarcinoma cell lines with high BIM expression, concomitant high mTOR expression increased IC50 of gefitinib for cell proliferation. We next sought to analyse the signalling pattern in cell lines with strong activation of mTOR and its substrate P-S6. We showed that mTOR and phosphodiesterase 4D (PDE4D) strongly correlate in resistant EGFR-mutant cancer cell lines. These data suggest that the combination of EGFR TKI with mTOR or PDE4 inhibitors could be adequate therapy for EGFR-mutant NSCLC patients with high pretreatment levels of BIM and mTOR.

  15. Fisetin induces autophagic cell death through suppression of mTOR signaling pathway in prostate cancer cells

    PubMed Central

    Suh, Yewseok; Afaq, Farrukh; Khan, Naghma; Johnson, Jeremy J.; Khusro, Fatima H.; Mukhtar, Hasan

    2010-01-01

    The mammalian target of rapamycin (mTOR) kinase is an important component of PTEN/PI3K/Akt signaling pathway, which is frequently deregulated in prostate cancer (CaP). Recent studies suggest that targeting PTEN/PI3K/Akt and mTOR signaling pathway could be an effective strategy for the treatment of hormone refractory CaP. Here, we show that the treatment of androgen-independent and PTEN-negative human CaP PC3 cells with fisetin, a dietary flavonoid, resulted in inhibition of mTOR kinase signaling pathway. Treatment of cells with fisetin inhibited mTOR activity and downregulated Raptor, Rictor, PRAS40 and GβL that resulted in loss of mTOR complexes (mTORC)1/2 formation. Fisetin also activated the mTOR repressor TSC2 through inhibition of Akt and activation of AMPK. Fisetin-mediated inhibition of mTOR resulted in hypophosphorylation of 4EBP1 and suppression of Cap-dependent translation. We also found that fisetin treatment leads to induction of autophagic-programmed cell death rather than cytoprotective autophagy as shown by small interfering RNA Beclin1-knockdown and autophagy inhibitor. Taken together, we provide evidence that fisetin functions as a dual inhibitor of mTORC1/2 signaling leading to inhibition of Cap-dependent translation and induction of autophagic cell death in PC3 cells. These results suggest that fisetin could be a useful chemotherapeutic agent in treatment of hormone refractory CaP. PMID:20530556

  16. Tuberous sclerosis complex suppression in cerebellar development and medulloblastoma: separate regulation of mTOR activity and p27Kip1 localization

    PubMed Central

    Bhatia, Bobby; Northcott, Paul A.; Hambardzumyan, Dolores; Govindarajan, Baskaran; Brat, Daniel J.; Arbiser, Jack L.; Holland, Eric C.; Taylor, Michael D.; Kenney, Anna Marie

    2009-01-01

    During development, proliferation of cerebellar granule neuron precursors (CGNPs), candidate cells-of-origin for the pediatric brain tumor medulloblastoma, requires signaling by Sonic hedgehog (Shh) and insulin-like growth factor (IGF), whose pathways are also implicated in medulloblastoma. One of the consequences of IGF signaling is inactivation of the mTOR-suppressing Tuberous Sclerosis Complex (TSC), comprised of TSC1 and TSC2, leading to increased mRNA translation. We show that mice in which TSC function is impaired display increased mTOR pathway activation, enhanced CGNP proliferation, GSK-3α/β inactivation, and cytoplasmic localization of the cyclin-dependent kinase (cdk) inhibitor p27Kip1, which has been proposed to cause its inactivation or gain of oncogenic functions. We observed the same characteristics in wild-type primary cultures of CGNPs in which TSC1 and/or TSC2 were knocked down, and in mouse medulloblastomas induced by ectopic Shh pathway activation. Moreover, Shh-induced mouse medulloblastomas manifested Akt-mediated TSC2 inactivation, and the mutant TSC2 allele synergized with aberrant Shh signaling to increase medulloblastoma incidence in mice. Driving exogenous TSC2 expression in Shh-induced medulloblastoma cells corrected p27Kip1 localization and reduced proliferation. GSK-3α/β inactivation in the tumors in vivo and in primary CGNP cultures was mTOR-dependent, whereas p27Kip1 cytoplasmic localization was regulated upstream of mTOR, by TSC2. These results indicate that a balance between Shh mitogenic signaling and TSC function regulating new protein synthesis and cdk inhibition is essential for normal development and prevention of tumor formation or expansion. PMID:19738049

  17. Structure–activity relationships study of mTOR kinase inhibition using QSAR and structure-based drug design approaches

    PubMed Central

    Lakhlili, Wiame; Yasri, Abdelaziz; Ibrahimi, Azeddine

    2016-01-01

    The discovery of clinically relevant inhibitors of mammalian target of rapamycin (mTOR) for anticancer therapy has proved to be a challenging task. The quantitative structure–activity relationship (QSAR) approach is a very useful and widespread technique for ligand-based drug design, which can be used to identify novel and potent mTOR inhibitors. In this study, we performed two-dimensional QSAR tests, and molecular docking validation tests of a series of mTOR ATP-competitive inhibitors to elucidate their structural properties associated with their activity. The QSAR tests were performed using partial least square method with a correlation coefficient of r2=0.799 and a cross-validation of q2=0.714. The chemical library screening was done by associating ligand-based to structure-based approach using the three-dimensional structure of mTOR developed by homology modeling. We were able to select 22 compounds from two databases as inhibitors of the mTOR kinase active site. We believe that the method and applications highlighted in this study will help future efforts toward the design of selective ATP-competitive inhibitors. PMID:27980424

  18. Activation of the mTOR signaling pathway in peritumoral tissues can cause glioma-associated seizures.

    PubMed

    Yuan, Yang; Xiang, Wang; Yanhui, Liu; Ruofei, Liang; Jiewen, Luo; Shu, Jiang; Qing, Mao

    2017-01-01

    Epileptic seizures, the most common symptom accompanying glioma, are closely associated with tumor growth and patient quality of life. However, the association between glioma and glioma-related epilepsy is poorly understood. In fact, findings related to the location of epileptogenicity have been inconsistent in previous studies. We investigated seizure foci in patients with glioma and the corresponding association between glioma-related epilepsy and the tumoral and peritumoral microenvironment. Clinical characteristics, extracellular electrophysiology, immunohistochemistry, and western blots were conducted on 12 patients with glioma; nine patients had histories of preoperative seizures while three did not. Samples from included patients were used to identify seizure foci and mTOR pathway status. Electrophysiological recordings were conducted on 36 samples (tumor, peritumoral, and normal brain tissues) from 12 patients. Interictal-like discharges (ILDs) were observed in seven of nine peritumoral tissues obtained from patients with glioma that had experienced perioperative seizures. No ILDs were observed in any other sample groups. Western blots and immunohistochemistry for mTOR pathway proteins (mTOR and S6k) suggested that the mTOR pathway was activated in peritumoral tissues of patients with seizure history, but inactivated in patients without seizure history. Our results suggest that mTOR pathway expression in peritumoral tissues is associated with tumor-related seizures, thus providing a potential target for therapeutics aimed at simultaneously controlling gliomas and seizures.

  19. mTOR signaling promotes stem cell activation via counterbalancing BMP-mediated suppression during hair regeneration.

    PubMed

    Deng, Zhili; Lei, Xiaohua; Zhang, Xudong; Zhang, Huishan; Liu, Shuang; Chen, Qi; Hu, Huimin; Wang, Xinyue; Ning, Lina; Cao, Yujing; Zhao, Tongbiao; Zhou, Jiaxi; Chen, Ting; Duan, Enkui

    2015-02-01

    Hair follicles (HFs) undergo cycles of degeneration (catagen), rest (telogen), and regeneration (anagen) phases. Anagen begins when the hair follicle stem cells (HFSCs) obtain sufficient activation cues to overcome suppressive signals, mainly the BMP pathway, from their niche cells. Here, we unveil that mTOR complex 1 (mTORC1) signaling is activated in HFSCs, which coincides with the HFSC activation at the telogen-to-anagen transition. By using both an inducible conditional gene targeting strategy and a pharmacological inhibition method to ablate or inhibit mTOR signaling in adult skin epithelium before anagen initiation, we demonstrate that HFs that cannot respond to mTOR signaling display significantly delayed HFSC activation and extended telogen. Unexpectedly, BMP signaling activity is dramatically prolonged in mTOR signaling-deficient HFs. Through both gain- and loss-of-function studies in vitro, we show that mTORC1 signaling negatively affects BMP signaling, which serves as a main mechanism whereby mTORC1 signaling facilitates HFSC activation. Indeed, in vivo suppression of BMP by its antagonist Noggin rescues the HFSC activation defect in mTORC1-null skin. Our findings reveal a critical role for mTOR signaling in regulating stem cell activation through counterbalancing BMP-mediated repression during hair regeneration.

  20. Discovering new mTOR inhibitors for cancer treatment through virtual screening methods and in vitro assays

    PubMed Central

    Wang, Ling; Chen, Lei; Yu, Miao; Xu, Li-Hui; Cheng, Bao; Lin, Yong-Sheng; Gu, Qiong; He, Xian-Hui; Xu, Jun

    2016-01-01

    Mammalian target of rapamycin (mTOR) is an attractive target for new anticancer drug development. We recently developed in silico models to distinguish mTOR inhibitors and non-inhibitors. In this study, we developed an integrated strategy for identifying new mTOR inhibitors using cascaded in silico screening models. With this strategy, fifteen new mTOR kinase inhibitors including four compounds with IC50 values below 10 μM were discovered. In particular, compound 17 exhibited potent anticancer activities against four tumor cell lines, including MCF-7, HeLa, MGC-803, and C6, with IC50 values of 1.90, 2.74, 3.50 and 11.05 μM. Furthermore, cellular studies and western blot analyses revealed that 17 induces cell death via apoptosis by targeting both mTORC1 and mTORC2 within cells and arrests the cell cycle of HeLa at the G1/G0-phase. Finally, multi-nanosecond explicit solvent simulations and MM/GBSA analyses were carried out to study the inhibitory mechanisms of 13, 17, and 40 for mTOR. The potent compounds presented here are worthy of further investigation. PMID:26732172

  1. Reduced AMPK-ACC and mTOR signaling in muscle from older men, and effect of resistance exercise

    PubMed Central

    Li, Mengyao; Verdijk, Lex B.; Sakamoto, Kei; Ely, Brian; van Loon, Luc J.C.; Musi, Nicolas

    2012-01-01

    AMP-activated protein kinase (AMPK) is a key energy-sensitive enzyme that controls numerous metabolic and cellular processes. Mammalian target of rapamycin (mTOR) is another energy/nutrient-sensitive kinase that controls protein synthesis and cell growth. In this study we determined whether older versus younger men have alterations in the AMPK and mTOR pathways in skeletal muscle, and examined the effect of a long term resistance type exercise training program on these signaling intermediaries. Older men had decreased AMPKα2 activity and lower phosphorylation of AMPK and its downstream signaling substrate acetyl-CoA carboxylase (ACC). mTOR phosphylation also was reduced in muscle from older men. Exercise training increased AMPKα1 activity in older men, however, AMPKα2 activity, and the phosphorylation of AMPK, ACC and mTOR, were not affected. In conclusion, older men have alterations in the AMPK-ACC and mTOR pathways in muscle. In addition, prolonged resistance type exercise training induces an isoform-selective up regulation of AMPK activity. PMID:23000302

  2. Discovering new mTOR inhibitors for cancer treatment through virtual screening methods and in vitro assays

    NASA Astrophysics Data System (ADS)

    Wang, Ling; Chen, Lei; Yu, Miao; Xu, Li-Hui; Cheng, Bao; Lin, Yong-Sheng; Gu, Qiong; He, Xian-Hui; Xu, Jun

    2016-01-01

    Mammalian target of rapamycin (mTOR) is an attractive target for new anticancer drug development. We recently developed in silico models to distinguish mTOR inhibitors and non-inhibitors. In this study, we developed an integrated strategy for identifying new mTOR inhibitors using cascaded in silico screening models. With this strategy, fifteen new mTOR kinase inhibitors including four compounds with IC50 values below 10 μM were discovered. In particular, compound 17 exhibited potent anticancer activities against four tumor cell lines, including MCF-7, HeLa, MGC-803, and C6, with IC50 values of 1.90, 2.74, 3.50 and 11.05 μM. Furthermore, cellular studies and western blot analyses revealed that 17 induces cell death via apoptosis by targeting both mTORC1 and mTORC2 within cells and arrests the cell cycle of HeLa at the G1/G0-phase. Finally, multi-nanosecond explicit solvent simulations and MM/GBSA analyses were carried out to study the inhibitory mechanisms of 13, 17, and 40 for mTOR. The potent compounds presented here are worthy of further investigation.

  3. mTOR mediates human trophoblast invasion through regulation of matrix-remodeling enzymes and is associated with serine phosphorylation of STAT3

    SciTech Connect

    Busch, Susann; Renaud, Stephen J.; Schleussner, Ekkehard; Graham, Charles H.; Markert, Udo R.

    2009-06-10

    The intracellular signaling molecule mammalian target of rapamycin (mTOR) is essential for cell growth and proliferation. It is involved in mouse embryogenesis, murine trophoblast outgrowth and linked to tumor cell invasiveness. In order to assess the role of mTOR in human trophoblast invasion we analyzed the in vitro invasiveness of HTR-8/SVneo immortalized first-trimester trophoblast cells in conjunction with enzyme secretion upon mTOR inhibition and knockdown of mTOR protein expression. Additionally, we also tested the capability of mTOR to trigger signal transducer and activator of transcription (STAT)-3 by its phosphorylation status. Rapamycin inhibited mTOR kinase activity as demonstrated with a lower phosphorylation level of the mTOR substrate p70 S6 kinase (S6K). With the use of rapamycin and siRNA-mediated mTOR knockdown we could show that cell proliferation, invasion and secretion of matrix-metalloproteinases (MMP)-2 and -9, urokinase-like plasminogen activator (uPA) and its major physiological uPA inhibitor (PAI)-1 were inhibited. While tyrosine phosphorylation of STAT3 was unaffected by mTOR inhibition and knockdown, serine phosphorylation was diminished. We conclude that mTOR signaling is one major mechanism in a tightly regulated network of intracellular signal pathways including the JAK/STAT system to regulate invasion in human trophoblast cells by secretion of enzymes that remodel the extra-cellular matrix (ECM) such as MMP-2, -9, uPA and PAI-1. Dysregulation of mTOR may contribute to pregnancy-related pathologies caused through impaired trophoblast invasion.

  4. Alcohol intoxication following muscle contraction in mice decreases muscle protein synthesis but not mTOR signal transduction

    PubMed Central

    Steiner, Jennifer L.; Lang, Charles H.

    2014-01-01

    Background Alcohol [ethanol (EtOH)] intoxication antagonizes stimulation of muscle protein synthesis and mTOR signaling. However, whether the anabolic response can be reversed when alcohol is consumed after the stimulus is unknown. Methods A single bout of electrically stimulated muscle contractions (10 sets of 6 contractions) were induced in fasted male C57BL/6 mice 2 h prior to alcohol intoxication. EtOH was injected IP (3g/kg) and the gastrocnemius/plantaris muscle complex was collected 2 h later from the stimulated and contralateral unstimulated control leg. Results Muscle contraction increased protein synthesis 28% in control mice while EtOH abolished this stimulation-induced increase. Further, EtOH suppressed the rate of synthesis ~75% compared to control muscle irrespective of stimulation. This decrease was associated with impaired protein elongation as EtOH increased the phosphorylation of eEF2 Thr56. In contrast, stimulation-induced increases in mTORC1 (S6K1 Thr421/Ser424, S6K1 Thr389, rpS6 Ser240/244, 4E-BP1 Thr37/46) and MAPK (JNK Thr183/Tyr185, p38 Thr180/Tyr182, and rpS6S235/236) signaling were not reversed by acute EtOH. Conclusion These data suggest that EtOH-induced decreases in protein synthesis in fasted mice may be independent of mTORC1 and MAPK signaling following muscle contraction and instead due to the antagonistic actions of EtOH on mRNA translation elongation. Therefore, EtOH suppresses the contraction-induced increase in protein synthesis and over time has the potential to prevent skeletal muscle hypertrophy induced by repeated muscle contraction. PMID:25623400

  5. Methods to identify molecular expression of mTOR pathway: a rationale approach to stratify patients affected by clear cell renal cell carcinoma for more likely response to mTOR inhibitors

    PubMed Central

    Fiorini, Claudia; Massari, Francesco; Pedron, Serena; Sanavio, Sara; Ciccarese, Chiara; Porcaro, Antonio Benito; Artibani, Walter; Bertoldo, Francesco; Zampini, Claudia; Sava, Teodoro; Ficial, Miriam; Caliò, Anna; Chilosi, Marco; D’Amuri, Alessandro; Sanguedolce, Francesca; Tortora, Giampaolo; Scarpa, Aldo; Delahunt, Brett; Porta, Camillo; Martignoni, Guido; Brunelli, Matteo

    2014-01-01

    Since target therapy with mTOR inhibitors plays an important role in the current management of clear cell renal cell carcinoma (RCC), there is an increasing demand for predictive biomarkers, which may help to select patients that are most likely to benefit from personalized treatment. When dealing with formalin-fixed paraffin-embedded (FFPE) cancer tissue specimens, several techniques may be used to identify potential molecular markers, yielding different outcome in terms of accuracy. We sought to investigate and compare the capability of three main techniques to detect molecules performing an active function in mTOR pathway in RCC. Immunohistochemistry (IHC), Western blot (WB) and immunofluorescence (IF) analyses were performed on FFPE RCC tissue specimens from 16 patients by using the following mTOR pathway-related: mTOR (Ser235/236), phospho-mTOR (p-mTOR/Ser2448), phospho-p70S6k (p-p70S6k/Thr389), both monoclonal and polyclonal, phospho-S6Rb (p-S6Rb) and phospho-4EBP1 (p-4EBP1/Thr37/46). No single molecule was simultaneously revealed by all three techniques. Only p-p70S6k was detected by two methods (IHC and IF) using a monoclonal antibody. The other molecules were detected exclusively by one technique, as follows: p-mTOR and polyclonal p-p70S6K by IHC, p70S6K, p-S6Rb and p-4EBP1 by WB, and, finally, mTOR by IF. We found significant differences in detecting mTOR pathway-related active biomarkers by using three common techniques such as IHC, WB and IF on RCC samples. Such results have important implications in terms of predictive biomarker testing, and need to be related to clinical end-points such as responsiveness to targeted drugs by prospective studies. PMID:25520878

  6. Bridges between mitochondrial oxidative stress, ER stress and mTOR signaling in pancreatic β cells.

    PubMed

    Wang, Jing; Yang, Xin; Zhang, Jingjing

    2016-08-01

    Pancreatic β cell dysfunction, i.e., failure to provide insulin in concentrations sufficient to control blood sugar, is central to the etiology of all types of diabetes. Current evidence implicates mitochondrial oxidative stress and endoplasmic reticulum (ER) stress in pancreatic β cell loss and impaired insulin secretion. Oxidative and ER stress are interconnected so that misfolded proteins induce reactive oxygen species (ROS) production; likewise, oxidative stress disturbs the ER redox state thereby disrupting correct disulfide bond formation and proper protein folding. mTOR signaling regulates many metabolic processes including protein synthesis, cell growth, survival and proliferation. Oxidative stress inhibits mTORC1, which is considered an important suppressor of mitochondrial oxidative stress in β cells, and ultimately, controls cell survival. The interplay between ER stress and mTORC1 is complicated, since the unfolded protein response (UPR) activation can occur upstream or downstream of mTORC1. Persistent activation of mTORC1 initiates protein synthesis and UPR activation, while in the later phase induces ER stress. Chronic activation of ER stress inhibits Akt/mTORC1 pathway, while under particular settings, acute activation of UPR activates Akt-mTOR signaling. Thus, modulating mitochondrial oxidative stress and ER stress via mTOR signaling may be an approach that will effectively suppress obesity- or glucolipotoxicity-induced metabolic disorders such as insulin resistance and type 2 diabetes mellitus (T2DM). In this review, we focus on the regulations between mTOR signaling and mitochondrial oxidative or ER stress in pancreatic β cells.

  7. Identification of Protor as a novel Rictor-binding component of mTOR complex-2.

    PubMed

    Pearce, Laura R; Huang, Xu; Boudeau, Jérôme; Pawłowski, Rafał; Wullschleger, Stephan; Deak, Maria; Ibrahim, Adel F M; Gourlay, Robert; Magnuson, Mark A; Alessi, Dario R

    2007-08-01

    The mTOR (mammalian target of rapamycin) protein kinase is an important regulator of cell growth. Two complexes of mTOR have been identified: complex 1, consisting of mTOR-Raptor (regulatory associated protein of mTOR)-mLST8 (termed mTORC1), and complex 2, comprising mTOR-Rictor (rapamycininsensitive companion of mTOR)-mLST8-Sin1 (termed mTORC2). mTORC1 phosphorylates the p70 ribosomal S6K (S6 kinase) at its hydrophobic motif (Thr389), whereas mTORC2 phosphorylates PKB (protein kinase B) at its hydrophobic motif (Ser473). In the present study, we report that widely expressed isoforms of unstudied proteins termed Protor-1 (protein observed with Rictor-1) and Protor-2 interact with Rictor and are components of mTORC2. We demonstrate that immunoprecipitation of Protor-1 or Protor-2 results in the co-immunoprecipitation of other mTORC2 subunits, but not Raptor, a specific component of mTORC1. We show that detergents such as Triton X-100 or n-octylglucoside dissociate mTOR and mLST8 from a complex of Protor-1, Sin1 and Rictor. We also provide evidence that Rictor regulates the expression of Protor-1, and that Protor-1 is not required for the assembly of other mTORC2 subunits into a complex. Protor-1 is a novel Rictor-binding subunit of mTORC2, but further work is required to establish its role.

  8. Dose-dependent effects of mTOR inhibition on weight and mitochondrial disease in mice

    PubMed Central

    Johnson, Simon C.; Yanos, Melana E.; Bitto, Alessandro; Castanza, Anthony; Gagnidze, Arni; Gonzalez, Brenda; Gupta, Kanav; Hui, Jessica; Jarvie, Conner; Johnson, Brittany M.; Letexier, Nicolas; McCanta, Lanny; Sangesland, Maya; Tamis, Oliver; Uhde, Lauren; Van Den Ende, Alex; Rabinovitch, Peter S.; Suh, Yousin; Kaeberlein, Matt

    2015-01-01

    Rapamycin extends lifespan and attenuates age-related pathologies in mice when administered through diet at 14 parts per million (PPM). Recently, we reported that daily intraperitoneal injection of rapamycin at 8 mg/kg attenuates mitochondrial disease symptoms and progression in the Ndufs4 knockout mouse model of Leigh Syndrome. Although rapamycin is a widely used pharmaceutical agent dosage has not been rigorously examined and no dose-response profile has been established. Given these observations we sought to determine if increased doses of oral rapamycin would result in more robust impact on mTOR driven parameters. To test this hypothesis, we compared the effects of dietary rapamycin at doses ranging from 14 to 378 PPM on developmental weight in control and Ndufs4 knockout mice and on health and survival in the Ndufs4 knockout model. High dose rapamycin was well tolerated, dramatically reduced weight gain during development, and overcame gender differences. The highest oral dose, approximately 27-times the dose shown to extend murine lifespan, increased survival in Ndufs4 knockout mice similarly to daily rapamycin injection without observable adverse effects. These findings have broad implications for the effective use of rapamycin in murine studies and for the translational potential of rapamycin in the treatment of mitochondrial disease. This data, further supported by a comparison of available literature, suggests that 14 PPM dietary rapamycin is a sub-optimal dose for targeting mTOR systemically in mice. Our findings suggest that the role of mTOR in mammalian biology may be broadly underestimated when determined through treatment with rapamycin at commonly used doses. PMID:26257774

  9. Dietary and donepezil modulation of mTOR signaling and neuroinflammation in the brain

    PubMed Central

    Dasuri, Kalavathi; Zhang, Le; Kim, Sun OK Fernandez; Bruce-Keller, Annadora J.; Keller, Jeffrey N.

    2016-01-01

    Recent clinical and laboratory evidences suggest that high fat diet (HFD) induced obesity and its associated metabolic syndrome conditions promotes neuropathology in aging and age-related neurological disorders. However, the effects of high fat diet on brain pathology are poorly understood, and the effective strategies to overcome these effects remain elusive. In the current study, we examined the effects of HFD on brain pathology and further evaluated whether donepezil, an AChE inhibitor with neuroprotective functions, could suppress the ongoing HFD induced pathological changes in the brain. Our data demonstrates that HFD induced obesity results in increased neuroinflammation and increased AChE activity in the brain when compared with the mice fed on low fat diet (LFD). HFD administration to mice activated mTOR pathway resulting in increased phosphorylation of mTORser2448, AKTthr308 and S6K proteins involved in the signaling. Interestingly, donepezil administration with HFD suppressed HFD induced increases in AChE activity, and partially reversed HFD effects on microglial reactivity and the levels of mTOR signaling proteins in the brain when compared to the mice on LFD alone. However, gross levels of synaptic proteins were not altered in the brain tissues of mice fed either diet with or without donepezil. In conclusion, these results present a new insight in to the detrimental effects of HFD on brain via microglial activation and involvement of mTOR pathway, and further demonstrates the possible therapeutic role for donepezil in ameliorating the early effects of HFD that could help preserve the brain function in metabolic syndrome conditions. PMID:26554604

  10. Practical recommendations for the early use of m-TOR inhibitors (sirolimus) in renal transplantation.

    PubMed

    Campistol, Josep M; Cockwell, Paul; Diekmann, Fritz; Donati, Donato; Guirado, Luis; Herlenius, Gustaf; Mousa, Dujanah; Pratschke, Johann; San Millán, Juan Carlos Ruiz

    2009-07-01

    m-TOR inhibitors (e.g. sirolimus) are well-tolerated immunosuppressants used in renal transplantation for prophylaxis of organ rejection, and are associated with long-term graft survival. Early use of sirolimus is often advocated by clinicians, but this may be associated with a number of side-effects including impaired wound-healing, lymphoceles and delayed graft function. As transplant clinicians with experience in the use of sirolimus, we believe such side-effects can be limited by tailored clinical management. We present recommendations based on published literature and our clinical experience. Furthermore, guidance is provided on sirolimus use during surgery, both at transplantation and for subsequent operations.

  11. Genetic variants in the mTOR pathway and breast cancer risk in African American women

    PubMed Central

    Cheng, Ting-Yuan David; Ambrosone, Christine B.; Hong, Chi-Chen; Lunetta, Kathryn L.; Liu, Song; Hu, Qiang; Yao, Song; Sucheston-Campbell, Lara; Bandera, Elisa V.; Ruiz-Narváez, Edward A.; Haddad, Stephen; Troester, Melissa A.; Haiman, Christopher A.; Bensen, Jeannette T.; Olshan, Andrew F.; Palmer, Julie R.; Rosenberg, Lynn

    2016-01-01

    The phosphatidylinositol 3-kinase–AKT–mammalian target of rapamycin (mTOR) pathway has been implicated in breast carcinogenesis. However, there has been no large-scale investigation of genetic variants in the mTOR pathway and breast cancer risk. We examined 28847 single-nucleotide polymorphisms (SNPs) in 61 mTOR pathway genes in the African American Breast Cancer Epidemiology and Risk consortium of 3663 cases [1983 estrogen receptor-positive (ER+) and 1098 ER-negative (ER−)] and 4687 controls. Gene-level analyses were conducted using the adaptive rank truncated product (ARTP) test for 10773 SNPs that were not highly correlated (r 2 < 0.8), and SNP-level analyses were conducted with logistic regression. Among genes that were prioritized (nominal P < 0.05, ARTP tests), associations were observed for intronic SNPs TSC2 rs181088346 [odds ratio (OR) of each copy of variant allele = 0.77, 95% confidence interval (CI) = 0.65–0.88 for all breast cancer] and BRAF rs114729114 (OR = 1.53, 95% CI = 1.24–1.91 for all breast cancer and OR = 2.03, 95% CI = 1.50–2.76 for ER− tumors). For ER− tumors, intronic SNPs PGF rs11542848 (OR = 1.38, 95% CI = 1.15–1.66) and rs61759375 (OR = 1.34, 95% CI = 1.14–1.57) and MAPK3 rs78564187 (OR = 1.26, 95% CI = 1.11–1.43) were associated with increased risk. These SNPs were significant at a gene-wide level (Bonferroni-corrected P < 0.05). The variant allele of RPS6KB2 rs35363135, a synonymous coding SNP, was more likely to be observed in ER− than ER+ tumors (OR = 1.18, 95% CI = 1.05–1.31, gene-wide Bonferroni-corrected P = 0.06). In conclusion, specific mTOR pathway genes are potentially important to breast cancer risk and to the ER negativity in African American women. PMID:26577839

  12. mTOR: from growth signal integration to cancer, diabetes and ageing

    PubMed Central

    Zoncu, Roberto; Sabatini, David M.; Efeyan, Alejo

    2012-01-01

    Preface In all eukaryotes, the target of rapamycin (TOR) signaling pathway couples energy and nutrient abundance to the execution of cell growth and division, owing to the ability of TOR protein kinase to simultaneously sense energy, nutrients and stress, and, in metazoan, growth factors. Mammalian TOR complexes 1 and 2 (mTORC1 and mTORC2) exert their actions by regulating other important kinases, such as S6K and Akt. In the last few years, a significant advance in our understanding of the regulation and functions of mTOR has revealed its critical involvement in the onset and progression of diabetes, cancer and ageing. PMID:21157483

  13. Neuroprotective Strategy in Retinal Degeneration: Suppressing ER Stress-Induced Cell Death via Inhibition of the mTOR Signal

    PubMed Central

    Fan, Bin; Sun, Ying-Jian; Liu, Shu-Yan; Che, Lin; Li, Guang-Yu

    2017-01-01

    The retina is a specialized sensory organ, which is essential for light detection and visual formation in the human eye. Inherited retinal degenerations are a heterogeneous group of eye diseases that can eventually cause permanent vision loss. UPR (unfolded protein response) and ER (endoplasmic reticulum) stress plays an important role in the pathological mechanism of retinal degenerative diseases. mTOR (the mammalian target of rapamycin) kinase, as a signaling hub, controls many cellular processes, covering protein synthesis, RNA translation, ER stress, and apoptosis. Here, the hypothesis that inhibition of mTOR signaling suppresses ER stress-induced cell death in retinal degenerative disorders is discussed. This review surveys knowledge of the influence of mTOR signaling on ER stress arising from misfolded proteins and genetic mutations in retinal degenerative diseases and highlights potential neuroprotective strategies for treatment and therapeutic implications. PMID:28106827

  14. mTOR inhibitors block Kaposi sarcoma growth by inhibiting essential autocrine growth factors and tumor angiogenesis.

    PubMed

    Roy, Debasmita; Sin, Sang-Hoon; Lucas, Amy; Venkataramanan, Raman; Wang, Ling; Eason, Anthony; Chavakula, Veenadhari; Hilton, Isaac B; Tamburro, Kristen M; Damania, Blossom; Dittmer, Dirk P

    2013-04-01

    Kaposi sarcoma originates from endothelial cells and it is one of the most overt angiogenic tumors. In Sub-Saharan Africa, where HIV and the Kaposi sarcoma-associated herpesvirus (KSHV) are endemic, Kaposi sarcoma is the most common cancer overall, but model systems for disease study are insufficient. Here, we report the development of a novel mouse model of Kaposi sarcoma, where KSHV is retained stably and tumors are elicited rapidly. Tumor growth was sensitive to specific allosteric inhibitors (rapamycin, CCI-779, and RAD001) of the pivotal cell growth regulator mTOR. Inhibition of tumor growth was durable up to 130 days and reversible. mTOR blockade reduced VEGF secretion and formation of tumor vasculature. Together, the results show that mTOR inhibitors exert a direct anti-Kaposi sarcoma effect by inhibiting angiogenesis and paracrine effectors, suggesting their application as a new treatment modality for Kaposi sarcoma and other cancers of endothelial origin.

  15. Chronic leucine supplementation of a low protein diet increases protein synthesis in skeletal muscle and visceral tissues of neonatal pigs through mTOR signaling

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Leucine acutely stimulates protein synthesis by activating the mammalian target of rapamycin (mTOR) signaling pathway. We hypothesized that leucine supplementation of a low protein diet will enhance protein synthesis and mTOR signaling in the neonate for prolonged periods. Fasted 5-d-old pigs (n=6–8...

  16. MicroRNA-224 aggrevates tumor growth and progression by targeting mTOR in gastric cancer.

    PubMed

    Zhang, Ying; Li, Chang-Feng; Ma, Lian-Jun; Ding, Meng; Zhang, Bin

    2016-09-01

    Growing evidence suggests that microRNA plays an essential role in the development and metastasis of many tumors, including gastric cancer. Aberrant miR-224 expression has been indicated in tumor growth, the mechanism of miR-224 promoting the proliferation and metastatic ability for gastric cancer remains unclear. Accumulating evidence reports that mTOR signal pathway plays an important role in the cellular process, such as apoptosis, cell growth and proliferation. The goal of the present study was to identify whether miR-224 could inhibit the growth, migration, invasion, proliferation and metastasis of gastric cancer through targeting mTOR expression. Real-time PCR (RT-PCR) was used to quantify miR-224 expression in vitro and in vivo experiments. Luciferase reporter assays were performed to confirm the activity of mTOR pathway, and immunofluorescence staining assay was conducted to observe apoptosis and cell proliferation ability. Bioinformatics as well as cell luciferase function studies distinguished the direct modulation of miR-224 on the 3'-UTR of the mTOR, which leads to the inactivation of apoptosis signaling and the activation of cell proliferation. In addition, inhibition of miR-224 significantly reduced the expression of mTOR and improved caspase-9/3 expression while decreased cyclin D1/2 levels, attenuating gastric cancer cell proliferation. Therefore, the present study revealed the mechanistic links between miR-224 and mTOR in the pathogenesis of gastric cancer through modulation of caspase-9/3 and cyclin D1/2. In addition, targeting miR-224 could serve as a novel strategy for future gastric cancer therapy.

  17. mTOR plays an important role in cow's milk allergy-associated behavioral and immunological deficits.

    PubMed

    Wu, Jiangbo; de Theije, Caroline G M; da Silva, Sofia Lopes; van der Horst, Hilma; Reinders, Margot T M; Broersen, Laus M; Willemsen, Linette E M; Kas, Martien J H; Garssen, Johan; Kraneveld, Aletta D

    2015-10-01

    Autism spectrum disorder (ASD) is multifactorial, with both genetic as well as environmental factors working in concert to develop the autistic phenotype. Immunological disturbances in autistic individuals have been reported and a role for food allergy has been suggested in ASD. Single gene mutations in mammalian target of rapamycin (mTOR) signaling pathway are associated with the development of ASD and enhanced mTOR signaling plays a central role in directing immune responses towards allergy as well. Therefore, the mTOR pathway may be a pivotal link between the immune disturbances and behavioral deficits observed in ASD. In this study it was investigated whether the mTOR pathway plays a role in food allergy-induced behavioral and immunological deficits. Mice were orally sensitized and challenged with whey protein. Meanwhile, cow's milk allergic (CMA) mice received daily treatment of rapamycin. The validity of the CMA model was confirmed by showing increased allergic immune responses. CMA mice showed reduced social interaction and increased repetitive self-grooming behavior. Enhanced mTORC1 activity was found in the brain and ileum of CMA mice. Inhibition of mTORC1 activity by rapamycin improved the behavioral and immunological deficits of CMA mice. This effect was associated with increase of Treg associated transcription factors in the ileum of CMA mice. These findings indicate that mTOR activation may be central to both the intestinal, immunological, and psychiatric ASD-like symptoms seen in CMA mice. It remains to be investigated whether mTOR can be seen as a therapeutic target in cow's milk allergic children suffering from ASD-like symptoms.

  18. Caloric restriction protects against electrical kindling of the amygdala by inhibiting the mTOR signaling pathway

    PubMed Central

    Phillips-Farfán, Bryan V.; Rubio Osornio, María del Carmen; Custodio Ramírez, Verónica; Paz Tres, Carlos; Carvajal Aguilera, Karla G.

    2015-01-01

    Caloric restriction (CR) has been shown to possess antiepileptic properties; however its mechanism of action is poorly understood. CR might inhibit the activity of the mammalian or mechanistic target of rapamycin (mTOR) signaling cascade, which seems to participate crucially in the generation of epilepsy. Thus, we investigated the effect of CR on the mTOR pathway and whether CR modified epilepsy generation due to electrical amygdala kindling. The former was studied by analyzing the phosphorylation of adenosine monophosphate-activated protein kinase, protein kinase B and the ribosomal protein S6. The mTOR cascade is regulated by energy and by insulin levels, both of which may be changed by CR; thus we investigated if CR altered the levels of energy substrates in the blood or the level of insulin in plasma. Finally, we studied if CR modified the expression of genes that encode proteins participating in the mTOR pathway. CR increased the after-discharge threshold and tended to reduce the after-discharge duration, indicating an anti-convulsive action. CR diminished the phosphorylation of protein kinase B and ribosomal protein S6, suggesting an inhibition of the mTOR cascade. However, CR did not change glucose, β-hydroxybutyrate or insulin levels; thus the effects of CR were independent from them. Interestingly, CR also did not modify the expression of any investigated gene. The results suggest that the anti-epileptic effect of CR may be partly due to inhibition of the mTOR pathway. PMID:25814935

  19. Effects of the single and combined treatment with dopamine agonist, somatostatin analog and mTOR inhibitors in a human lung carcinoid cell line: an in vitro study.

    PubMed

    Pivonello, Claudia; Rousaki, Panagoula; Negri, Mariarosaria; Sarnataro, Maddalena; Napolitano, Maria; Marino, Federica Zito; Patalano, Roberta; De Martino, Maria Cristina; Sciammarella, Concetta; Faggiano, Antongiulio; Rocco, Gaetano; Franco, Renato; Kaltsas, Gregory A; Colao, Annamaria; Pivonello, Rosario

    2016-09-29

    Somatostatin analogues and mTOR inhibitors have been used as medical therapy in lung carcinoids with variable results. No data are available on dopamine agonists as treatment for lung carcinoids. The main aim of the current study was to evaluate the effect of the combined treatment of somatostatin analogue octreotide and the dopamine agonist cabergoline with mTOR inhibitors in an in vitro model of typical lung carcinoids: the NCI-H727 cell line. In NCI-H727 cell line, reverse transcriptase-quantitative polymerase chain reaction and immunofluorescence were assessed to characterize the expression of the somatostatin receptor 2 and 5, dopamine receptor 2 and mTOR pathway components. Fifteen typical lung carcinoids tissue samples have been used for somatostatin receptor 2, dopamine receptor 2, and the main mTOR pathway component p70S6K expression and localization by immunohistochemistry. Cell viability, fluorescence-activated cell sorting analysis and western blot have been assessed to test the pharmacological effects of octreotide, cabergoline and mTOR inhibitors, and to evaluate the activation of specific cell signaling pathways in NCI-H727 cell line. NCI-H727 cell line expressed somatostatin receptor 2, somatostatin receptor 5 and dopamine receptor 2 and all mTOR pathway components at messenger and protein levels. Somatostatin receptor 2, dopamine receptor 2, and p70S6K (non phosphorylated and phosphorylated) proteins were expressed in most typical lung carcinoids tissue samples. Octreotide and cabergoline did not reduce cell viability as single agents but, when combined with mTOR inhibitors, they potentiate mTOR inhibitors effect after long-term exposure, reducing Akt and ERK phosphorylation, mTOR escape mechanisms, and increasing the expression DNA-damage-inducible transcript 4, an mTOR suppressor. In conclusion, the single use of octreotide and cabergoline is not sufficient to block cell viability but the combined approach of these agents with mTOR inhibitors

  20. The Role and Application of Sirtuins and mTOR Signaling in the Control of Ovarian Functions

    PubMed Central

    Sirotkin, Alexander V.

    2016-01-01

    The present short review demonstrates the involvement of sirtuins (SIRTs) in the control of ovarian functions at various regulatory levels. External and endocrine factors can affect female reproduction via SIRTs-mammalian target of rapamycin (mTOR) system, which, via hormones and growth factors, can in turn regulate basic ovarian functions (proliferation, apoptosis, secretory activity of ovarian cells, their response to upstream hormonal regulators, ovarian folliculo- and oogenesis, and fecundity). SIRTs and SIRTs-related signaling molecules and drugs regulating mTOR can be used for characterization, prediction, and regulation of ovarian functions, as well as for diagnostics and treatment of ovarian disorders. PMID:27886120

  1. Wide spectrum of developmental brain disorders from megalencephaly to focal cortical dysplasia and pigmentary mosaicism caused by mutations of MTOR

    PubMed Central

    Solovieff, Nadia; Goold, Carleton; Jansen, Laura A.; Menon, Suchithra; Timms, Andrew E.; Conti, Valerio; Biag, Jonathan D.; Adams, Carissa; Boyle, Evan August; Collins, Sarah; Ishak, Gisele; Poliachik, Sandra; Girisha, Katta M.; Yeung, Kit San; Chung, Brian Hon Yin; Rahikkala, Elisa; Gunter, Sonya A.; McDaniel, Sharon S.; Macmurdo, Colleen Forsyth; Bernstein, Jonathan A.; Martin, Beth; Leary, Rebecca; Mahan, Scott; Liu, Shanming; Weaver, Molly; Doerschner, Michael; Jhangiani, Shalini; Muzny, Donna M.; Boerwinkle, Eric; Gibbs, Richard A.; Lupski, James R.; Shendure, Jay; Saneto, Russell P.; Novotny, Edward J.; Wilson, Christopher J.; Sellers, William R.; Morrissey, Michael; Hevner, Robert F.; Ojemann, Jeffrey G.; Guerrini, Renzo; Murphy, Leon O.; Winckler, Wendy; Dobyns, William B.

    2016-01-01

    Importance Focal cortical dysplasia (FCD), hemimegalencephaly (HMEG) and megalencephaly constitute a spectrum of malformations of cortical development with shared neuropathologic features. Collectively, these disorders are associated with significant childhood morbidity and mortality. FCD, in particular, represents the most frequent cause of intractable focal epilepsy in children. Objective To identify the underlying molecular etiology of FCD, HMEG, and diffuse megalencephaly. Design, Setting and Participants We performed whole exome sequencing (WES) on eight children with FCD or HMEG using standard depth (~50-60X) sequencing in peripheral samples (blood, saliva or skin) from the affected child and their parents, and deep (~150-180X) sequencing in affected brain tissue. We used both targeted sequencing and WES to screen a cohort of 93 children with molecularly unexplained diffuse or focal brain overgrowth (42 with FCD-HMEG, and 51 with diffuse megalencephaly). Histopathological and functional assays of PI3K-AKT-MTOR pathway activity in resected brain tissue and cultured neurons were performed to validate mutations. Main Outcomes and Measures Whole exome sequencing and targeted sequencing identified variants associated with this spectrum of developmental brain disorders. Results We identified low-level mosaic mutations of MTOR in brain tissue in four children with FCD type 2a with alternative allele fractions ranging from 0.012–0.086. We also identified intermediate level mosaic mutation of MTOR (p.Thr1977Ile) in three unrelated children with diffuse megalencephaly and pigmentary mosaicism in skin that resembles hypomelanosis of Ito. Finally, we identified a constitutional de novo mutation of MTOR (p.Glu1799Lys) in three unrelated children with diffuse megalencephaly and intellectual disability. Molecular and functional analysis in two children with FCD type 2a from whom multiple affected brain tissue samples were available revealed a gradient of alternate allele

  2. DUAL INHIBITION OF PI3K/AKT AND mTOR SIGNALING IN HUMAN NON-SMALL CELL LUNG CANCER CELLS BY A DIETARY FLAVONOID FISETIN

    PubMed Central

    Khan, Naghma; Afaq, Farrukh; Khusro, Fatima H.; Adhami, Vaqar Mustafa; Suh, Yewseok; Mukhtar, Hasan

    2011-01-01

    Lung cancer is one of the most commonly occurring malignancies. It has been reported that mTOR is phosphorylated in lung cancer and its activation was more frequent in tumors with over-expression of PI3K/Akt. Therefore, dual inhibitors of PI3K/Akt and mTOR signaling could be valuable agents for treating lung cancer. In the present study, we show that fisetin, a dietary tetrahydroxyflavone inhibits cell-growth with the concomitant suppression of PI3K/Akt and mTOR signaling in human non-small cell lung cancer (NSCLC) cells. Using autodock 4, we found that fisetin physically interacts with the mTOR complex at two sites. Fisetin treatment was also found to reduce the formation of A549 cell colonies in a dose-dependent manner. Treatment of cells with fisetin caused decrease in the protein expression of PI3K (p85 and p110), inhibition of phosphorylation of Akt, mTOR, p70S6K1, eIF-4E and 4E-BP1. Fisetin-treated cells also exhibited dose-dependent inhibition of the constituents of mTOR signaling complex like Rictor, Raptor, GβL and PRAS40. There was increase in the phosphorylation of AMPKα and decrease in the phosphorylation of TSC2 on treatment of cells with fisetin. We also found that treatment of cells with mTOR inhibitor rapamycin and mTOR-siRNA caused decrease in phosphorylation of mTOR and its target proteins which were further downregulated on treatment with fisetin, suggesting that these effects are mediated in part, through mTOR signaling. Our results show that fisetin suppressed PI3K/Akt and mTOR signaling in NSCLC cells and thus, could be developed as a chemotherapeutic agent against human lung cancer. PMID:21618507

  3. Global ischemia induces lysosomal-mediated degradation of mTOR and activation of autophagy in hippocampal neurons destined to die

    PubMed Central

    Hwang, Jee-Yeon; Gertner, Michael; Pontarelli, Fabrizio; Court-Vazquez, Brenda; Bennett, Michael Vander Laan; Ofengeim, Dimitry; Zukin, Ruth Suzanne

    2017-01-01

    The mammalian target of rapamycin (mTOR) is a key regulator of cell growth, autophagy, translation, and survival. Dysregulation of mTOR signaling is associated with cancer, diabetes, and autism. However, a role for mTOR signaling in neuronal death is not well delineated. Here we show that global ischemia triggers a transient increase in mTOR phosphorylation at S2448, whereas decreasing p-mTOR and functional activity in selectively vulnerable hippocampal CA1 neurons. The decrease in mTOR coincides with an increase in biochemical markers of autophagy, pS317-ULK-1, pS14-Beclin-1, and LC3-II, a decrease in the cargo adaptor p62, and an increase in autophagic flux, a functional readout of autophagy. This is significant in that autophagy, a catabolic process downstream of mTORC1, promotes the formation of autophagosomes that capture and target cytoplasmic components to lysosomes. Inhibitors of the lysosomal (but not proteasomal) pathway rescued the ischemia-induced decrease in mTOR, consistent with degradation of mTOR via the autophagy/lysosomal pathway. Administration of the mTORC1 inhibitor rapamycin or acute knockdown of mTOR promotes autophagy and attenuates ischemia-induced neuronal death, indicating an inverse causal relation between mTOR, autophagy, and neuronal death. Our findings identify a novel and previously unappreciated mechanism by which mTOR self-regulates its own levels in hippocampal neurons in a clinically relevant model of ischemic stroke. PMID:27935582

  4. Pharmacogenetic predictor of extrapyramidal symptoms induced by antipsychotics: multilocus interaction in the mTOR pathway.

    PubMed

    Mas, S; Gassó, P; Ritter, M A; Malagelada, C; Bernardo, M; Lafuente, A

    2015-01-01

    Antipsychotic (AP) treatment-emergent extrapyramidal symptoms (EPS) are acute adverse reactions of APs. The aim of the present study is to analyze gene-gene interactions in nine genes related to the mTOR pathway, in order to develop genetic predictors of the appearance of EPS. 243 subjects (78 presenting EPS: 165 not) from three cohorts participated in the present study: Cohort 1, patients treated with risperidone, (n=114); Cohort 2, patients treated with APs other than risperidone (n=102); Cohort 3, AP-naïve patients with first-episode psychosis treated with risperidone, paliperidone or amisulpride, n=27. We analyzed gene-gene interactions by multifactor dimensionality reduction assay (MDR). In Cohort 1, we identified a four-way interaction, including the rs1130214 (AKT1), rs456998 (FCHSD1), rs7211818 (Raptor) and rs1053639 (DDIT4), that correctly predicted 97 of the 114 patients (85% accuracy). We validated the predictive power of the four-way interaction in Cohort 2 and in Cohort 3 with 86% and 88% accuracy respectively. We develop and validate a powerful pharmacogenetic predictor of AP-induced EPS. For the first time, the mTOR pathway has been related to EPS susceptibility and AP response. However, validation in larger and independent populations will be necessary for optimal generalization.

  5. Cyclin B Translation Depends on mTOR Activity after Fertilization in Sea Urchin Embryos

    PubMed Central

    Boulben, Sandrine; Glippa, Virginie; Morales, Julia; Cormier, Patrick

    2016-01-01

    The cyclin B/CDK1 complex is a key regulator of mitotic entry. Using PP242, a specific ATP-competitive inhibitor of mTOR kinase, we provide evidence that the mTOR signalling pathway controls cyclin B mRNA translation following fertilization in Sphaerechinus granularis and Paracentrotus lividus. We show that PP242 inhibits the degradation of the cap-dependent translation repressor 4E-BP (eukaryotic initiation factor 4E-Binding Protein). PP242 inhibits global protein synthesis, delays cyclin B accumulation, cyclin B/CDK1 complex activation and consequently entry into the mitotic phase of the cell cycle triggered by fertilization. PP242 inhibits cyclin B mRNA recruitment into active polysomes triggered by fertilization. An amount of cyclin B mRNA present in active polysomes appears to be insensitive to PP242 treatment. Taken together, our results suggest that, following sea urchin egg fertilization, cyclin B mRNA translation is controlled by two independent mechanisms: a PP242-sensitive and an additional PP242-insentitive mechanism. PMID:26962866

  6. Autophagic lysosomal reformation depends on mTOR reactivation in H2O2-induced autophagy.

    PubMed

    Zhang, Jiqian; Zhou, Wei; Lin, Jun; Wei, Pengfei; Zhang, Yunjiao; Jin, Peipei; Chen, Ming; Man, Na; Wen, Longping

    2016-01-01

    Autophagic lysosomal reformation, a key cellular process for maintaining lysosome homeostasis in elevated autophagy, so far has only been reported for cells under certain forms of starvation. For this reason, it is controversial that whether this phenomenon is starvation-specific and its importance in lysosomal regeneration at the late stage of autophagy is often challenged. Here we show that exogenous hydrogen peroxide (H2O2) induced lysosome depletion and recovery characteristic of autophagic lysosomal reformation, and we confirmed the occurrence of autophagic lysosomal reformation after H2O2 treatment by demonstrating Rab7 dissociation from autolysosomes, recruitment of Phosphatidylinositol 4-phosphate (PI4P) and clathrin to the surface of autolysosomes, and the existence of tubular "pro-lysosome" structures extending from autolysosomes. Similar to starvation, H2O2 caused an initial deactivation and a subsequent reactivation for mTOR, and mTOR reactivation was essential for ALR. Our results provided a first non-starvation example of autophagic lysosomal reformation and provide evidence for its importance for some autophagic processes other than that of starvation.

  7. Androgen receptor functions as a negative transcriptional regulator of DEPTOR, mTOR inhibitor.

    PubMed

    Kanno, Yuichiro; Zhao, Shuai; Yamashita, Naoya; Yanai, Kazuyuki; Nemoto, Kiyomitsu; Inouye, Yoshio

    2015-12-01

    It has been noticed that crosstalk between androgen receptor (AR) and mammalian target of rapamycin (mTOR) signaling pathways plays a crucial role in the proliferation of prostate cancer cells. To clarify this mechanism, we focused on DEPTOR, a naturally occurring inhibitor of mTOR. The treatment of a human AR-positive prostate cancer cell line, LNCaP, with the AR-agonist dihydrotestosterone (DHT) repressed DEPTOR mRNA expression in a time-dependent manner. This repression was abrogated by treatment with the AR-antagonist bicalutamide. Knockdown of DEPTOR mRNA by siRNA resulted in the increased phosphorylation of 70 kDa ribosomal protein S6 kinase 1 (S6K), a substrate of mTORC1, accompanied by the elevated expression of cyclin D1, a positive regulator of cell proliferation. Furthermore, the ChIP assay demonstrated that AR could bind to AR-responsible element-like region within the 4th intron of the DEPTOR gene. The amount of acetylated histone H3 (Lys9, Lys14) was reduced by the DHT treatment in this region. Taken together, these results propose that AR-dependent prostate cancer cell proliferation requires decreased DEPTOR transcription directly controlled by AR.

  8. Cyclin B Translation Depends on mTOR Activity after Fertilization in Sea Urchin Embryos.

    PubMed

    Chassé, Héloïse; Mulner-Lorillon, Odile; Boulben, Sandrine; Glippa, Virginie; Morales, Julia; Cormier, Patrick

    2016-01-01

    The cyclin B/CDK1 complex is a key regulator of mitotic entry. Using PP242, a specific ATP-competitive inhibitor of mTOR kinase, we provide evidence that the mTOR signalling pathway controls cyclin B mRNA translation following fertilization in Sphaerechinus granularis and Paracentrotus lividus. We show that PP242 inhibits the degradation of the cap-dependent translation repressor 4E-BP (eukaryotic initiation factor 4E-Binding Protein). PP242 inhibits global protein synthesis, delays cyclin B accumulation, cyclin B/CDK1 complex activation and consequently entry into the mitotic phase of the cell cycle triggered by fertilization. PP242 inhibits cyclin B mRNA recruitment into active polysomes triggered by fertilization. An amount of cyclin B mRNA present in active polysomes appears to be insensitive to PP242 treatment. Taken together, our results suggest that, following sea urchin egg fertilization, cyclin B mRNA translation is controlled by two independent mechanisms: a PP242-sensitive and an additional PP242-insentitive mechanism.

  9. Salmonella Typhimurium disrupts Sirt1/AMPK checkpoint control of mTOR to impair autophagy

    PubMed Central

    Ganesan, Raja; Hos, Nina Judith; Gutierrez, Saray; Fischer, Julia; Stepek, Joanna Magdalena; Daglidu, Evmorphia; Krönke, Martin

    2017-01-01

    During intracellular infections, autophagy significantly contributes to the elimination of pathogens, regulation of pro-inflammatory signaling, secretion of immune mediators and in coordinating the adaptive immune system. Intracellular pathogens such as S. Typhimurium have evolved mechanisms to circumvent autophagy. However, the regulatory mechanisms targeted by S. Typhimurium to modulate autophagy have not been fully resolved. Here we report that cytosolic energy loss during S. Typhimurium infection triggers transient activation of AMPK, an important checkpoint of mTOR activity and autophagy. The activation of AMPK is regulated by LKB1 in a cytosolic complex containing Sirt1 and LKB1, where Sirt1 is required for deacetylation and subsequent activation of LKB1. S. Typhimurium infection targets Sirt1, LKB1 and AMPK to lysosomes for rapid degradation resulting in the disruption of the AMPK-mediated regulation of mTOR and autophagy. The degradation of cytosolic Sirt1/LKB1/AMPK complex was not observed with two mutant strains of S. Typhimurium, ΔssrB and ΔssaV, both compromising the pathogenicity island 2 (SPI2). The results highlight virulence factor-dependent degradation of host cell proteins as a previously unrecognized strategy of S. Typhimurium to evade autophagy. PMID:28192515

  10. A Review of mTOR Pathway Inhibitors in Gynecologic Cancer

    PubMed Central

    Paulino, Eduardo; Garces, Álvaro Henrique Ingles

    2017-01-01

    The treatment of advanced gynecologic cancers remains palliative in most of cases. Although systemic treatment has entered into the era of targeted drugs the antitumor efficacies of current therapies are still limited. In this context there is a great need for more active treatment and rationally designed targeted therapies. The PI3K/AKT/mTOR is a signaling pathway in mammal cells that coordinates important cell activities. It has a critical function in the survival, growth, and proliferation of malignant cells and was object of important research in the last two decades. The mTOR pathway emerges as an attractive therapeutic target in cancer because it serves as a convergence point for many growth stimuli and, through its downstream substrates, controls cellular processes that contribute to the initiation and maintenance of cancer. Aberrant PI3K-dependent signaling occurs frequently in a wide range of tumor types, including endometrial, cervical, and ovarian cancers. The present study reviewed the available evidence regarding the potential impact of some mTOR pathway inhibitors in the treatment of gynecological cancer. Few advances in medical management have occurred in recent years in the treatment of advanced or recurrent gynecological malignancies, and a poor prognosis remains. Rationally designed molecularly targeted therapy is an emerging and important option in this setting; then more investigation in PI3K/AKT/mTOR pathway-targeted therapies is warranted. PMID:28286604

  11. Is mTOR Inhibitor Good Enough for Treatment All Tumors in TSC Patients?

    PubMed Central

    Habib, Samy L; Al-Obaidi, Noor Y; Nowacki, Maciej; Pietkun, Katarzyna; Zegarska, Barbara; Kloskowski, Tomasz; Zegarski, Wojciech; Drewa, Tomasz; Medina, Edward A.; Zhao, Zhenze; Liang, Sitai

    2016-01-01

    Tuberous sclerosis complex (TSC) is an autosomal dominant and multi-system genetic disorder in humans. TSC affects around 25,000 to 40,000 individuals in the United States and about 1 to 2 million individuals worldwide, with an estimated prevalence of one in 6,000 newborns. TSC occurs in all races and ethnic groups, and in both genders. TSC is caused by defects or mutations in two genes, TSC1 and TSC2. Loss of TSC1/TSC2 leads to dysregulation of mTOR, resulting in aberrant cell differentiation and development, and abnormal enlargement of cells. TSC is characterized by the development of benign and/or malignant tumors in several organs including renal/liver angiomyolipomas, facial angiofibroma, lymphangiomyomatosis, cardiac rhabdomyomas, retinal astrocytic, renal cell carcinoma, and brain subependymal giant cell astrocytomas (SEGA). In addition, TSC disease causes disabling neurologic disorders, including epilepsy, mental retardation and autism. Particularly problematic are the development of renal angiomyolipomas, which tend to be larger, bilateral, multifocal and present at a younger age compared with sporadic forms. In addition, SEGA block the flow of fluid within the brain, causing a buildup of fluid and pressure that leads to blurred vision and seizures. In the current review, we describe the pathology of TSC disease in key organs and summarize the use of mTOR inhibitors to treat tumors in TSC patients. PMID:27698899

  12. Metabolic clock generates nutrient anticipation rhythms in mTOR signaling.

    PubMed

    Khapre, Rohini V; Patel, Sonal A; Kondratova, Anna A; Chaudhary, Amol; Velingkaar, Nikkhil; Antoch, Marina P; Kondratov, Roman V

    2014-08-01

    The mTOR signaling pathway modulates metabolic processes with respect to nutrient availability and other growth-related cues. According to the existing paradigm, mTOR complex 1 (mTORC1) activityin vivo is induced by food and gradually decreases during fasting. We found that mTORC1 activity is controlled by an internal clock mechanism different from the known light-entrainable circadian clock. We observed 24-hr rhythms in phosphorylation of mTORC1 downstream targets, which were entrained by food, persisted during fasting and could be uncoupled from oscillating expression of the canonical circadian clock genes. Furthermore, these rhythms were present in tissues of mice with disrupted light-entrainable circadian clock. We propose tissue-specific rhythms in the expression of tor and its negative regulator deptor as the molecular mechanism of the mTORC1 activity oscillation. Our data demonstrate the existence of at least two independent molecular circadian clocks: one providing metabolic adaptation to periodic light/darkness and the other - to feeding.

  13. The oncometabolite 2-hydroxyglutarate activates the mTOR signalling pathway

    PubMed Central

    Carbonneau, Mélissa; M. Gagné, Laurence; Lalonde, Marie-Eve; Germain, Marie-Anne; Motorina, Alena; Guiot, Marie-Christine; Secco, Blandine; Vincent, Emma E.; Tumber, Anthony; Hulea, Laura; Bergeman, Jonathan; Oppermann, Udo; Jones, Russell G.; Laplante, Mathieu; Topisirovic, Ivan; Petrecca, Kevin; Huot, Marc-Étienne; Mallette, Frédérick A.

    2016-01-01

    The identification of cancer-associated mutations in the tricarboxylic acid (TCA) cycle enzymes isocitrate dehydrogenases 1 and 2 (IDH1/2) highlights the prevailing notion that aberrant metabolic function can contribute to carcinogenesis. IDH1/2 normally catalyse the oxidative decarboxylation of isocitrate into α-ketoglutarate (αKG). In gliomas and acute myeloid leukaemias, IDH1/2 mutations confer gain-of-function leading to production of the oncometabolite R-2-hydroxyglutarate (2HG) from αKG. Here we show that generation of 2HG by mutated IDH1/2 leads to the activation of mTOR by inhibiting KDM4A, an αKG-dependent enzyme of the Jumonji family of lysine demethylases. Furthermore, KDM4A associates with the DEP domain-containing mTOR-interacting protein (DEPTOR), a negative regulator of mTORC1/2. Depletion of KDM4A decreases DEPTOR protein stability. Our results provide an additional molecular mechanism for the oncogenic activity of mutant IDH1/2 by revealing an unprecedented link between TCA cycle defects and positive modulation of mTOR function downstream of the canonical PI3K/AKT/TSC1-2 pathway. PMID:27624942

  14. mTOR activity in AIDS-related diffuse large B-cell lymphoma

    PubMed Central

    Diaz-Perez, Julio A.; Preziosi, Michael; King, Charles C.; Jones, George A.; Jain, Sonia; Sun, Xiaoying; Reid, Erin G.; VandenBerg, Scott; Wang, Huan-You

    2017-01-01

    Background Patients infected with HIV have a significantly increased risk of developing non–Hodgkin lymphomas despite the widespread use of HAART. To investigate mTOR pathway activity in acquired immunodeficiency syndrome (AIDS) related diffuse large B-cell lymphoma AR-DLBCL, we used immunohistochemistry to examine the presence of the phosphorylated 70 ribosomal S6 protein-kinase (p70S6K), an extensively studied effector of mTOR Complex 1 (mTORC1) and the phosphorylated phosphatase and tensin homolog (pPTEN), a negative regulator of mTORC1 pathway. Materials and methods We evaluated tissue samples from 126 patients with AR-DLBCL. Among them, 98 samples were from tissue microarrays (TMAs) supplied by the Aids and Cancer Specimen Resource (ACSR), the remaining 28 samples were from cases diagnosed and treated at the University of California, San Diego (UCSD). The presence of p70S6K was evaluated with two antibodies directed against the combined epitopes Ser235/236 and Ser240/244, respectively; and additional monoclonal anti-bodies were used to identify pPTEN and phosphorylated proline-rich Akt substrate of 40kDa (pPRAS40). The degree of intensity and percentage of cells positive for p70S6K and pPTEN were assessed in all the samples. In addition, a subgroup of 28 patients from UCSD was studied to assess the presence of pPRAS40, an insulin-regulated activator of the mTORC1. The expression of each of these markers was correlated with clinical and histopathologic features. Results The majority of the patients evaluated were males (88%); only two cases (1.6%) were older than 65 years of age. We found high levels of both p70S6K-paired epitopes studied, 48% positivity against Ser235/236 (44% in ACSR and 64% in UCSD group), and 86% positivity against Ser240/244 (82% in ACSR and 100% in UCSD group). We observed more positive cells and stronger intensity with epitope Ser240/244 in comparison to Ser235/236 (p<0.0001). The degree of intensity and percentage of cells positive

  15. Effects of chronic overload on muscle hypertrophy and mTOR signaling in adult and aged rats

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We examined the effect of 28 days of overload on mammalian target of rapamycin (mTOR) and extracellular signal-regulated kinase (ERK) signaling in young adult (Y; 6 mo old) and aged (O; 30 mo old) Fischer 344 x Brown Norway rats subjected to bilateral synergist ablation (SA) of two-thirds of the gas...

  16. Estradiol-Induced Object Recognition Memory Consolidation Is Dependent on Activation of mTOR Signaling in the Dorsal Hippocampus

    ERIC Educational Resources Information Center

    Fortress, Ashley M.; Fan, Lu; Orr, Patrick T.; Zhao, Zaorui; Frick, Karyn M.

    2013-01-01

    The mammalian target of rapamycin (mTOR) signaling pathway is an important regulator of protein synthesis and is essential for various forms of hippocampal memory. Here, we asked whether the enhancement of object recognition memory consolidation produced by dorsal hippocampal infusion of 17[Beta]-estradiol (E[subscript 2]) is dependent on mTOR…

  17. Seizure-dependent mTOR activation in 5-HT neurons promotes autism-like behaviors in mice

    PubMed Central

    McMahon, John J.; Yu, Wilson; Yang, Jun; Feng, Haihua; Helm, Meghan; McMahon, Elizabeth; Zhu, Xinjun; Shin, Damian; Huang, Yunfei

    2014-01-01

    Epilepsy and autism spectrum disorder (ASD) are common comorbidities of one another. Despite the prevalent correlation between the two disorders, few studies have been able to elucidate a mechanistic link. We demonstrate that forebrain specific Tsc1 deletion in mice causes epilepsy and autism-like behaviors, concomitant with disruption of 5-HT neurotransmission. We find that epileptiform activity propagates to the raphe nuclei, resulting in seizure-dependent hyperactivation of mTOR in 5-HT neurons. To dissect whether mTOR hyperactivity in 5-HT neurons alone was sufficient to recapitulate an autism-like phenotype we utilized Tsc1flox/flox;Slc6a4-cre mice, in which mTOR is restrictively hyperactivated in 5-HT neurons. Tsc1flox/flox;Slc6a4-cre mice displayed alterations of the 5-HT system and autism-like behaviors, without causing epilepsy. Rapamycin treatment in these mice was sufficient to rescue the phenotype. We conclude that the spread of seizure activity to the brainstem is capable of promoting hyperactivation of mTOR in the raphe nuclei, which in turn promotes autism-like behaviors. Thus our study provides a novel mechanism describing how epilepsy can contribute to the development of autism-like behaviors, suggesting new therapeutic strategies for autism. PMID:25315683

  18. Inhibition of Histone Deacetylases (HDACs) and mTOR Signaling: Novel Strategies Toward the Treatment of Prostate Cancer

    DTIC Science & Technology

    2011-04-01

    protein and vessel density in xenograft models with constitutive mTOR activity, either through loss of PTEN (PC3 cells) or VHL (C2 cells) [19]. Myc-CaP/AS...et al. (2006) Class II histone deacetylases are associated with VHL -independent regulation of hypoxia- inducible factor 1 alpha. Cancer Res 66: 8814

  19. Biphasic activation of the mTOR pathway in the gustatory cortex is correlated with and necessary for taste learning.

    PubMed

    Belelovsky, Katya; Kaphzan, Hanoch; Elkobi, Alina; Rosenblum, Kobi

    2009-06-10

    Different forms of memories and synaptic plasticity require synthesis of new proteins at the time of acquisition or immediately after. We are interested in the role of translation regulation in the cortex, the brain structure assumed to store long-term memories. The mammalian target of rapamycin, mTOR (also known as FRAP and RAFT-1), is part of a key signal transduction mechanism known to regulate translation of specific subset of mRNAs and to affect learning and synaptic plasticity. We report here that novel taste learning induces two waves of mTOR activation in the gustatory cortex. Interestingly, the first wave can be identified both in synaptoneurosomal and cellular fractions, whereas the second wave is detected in the cellular fraction but not in the synaptic one. Inhibition of mTOR, specifically in the gustatory cortex, has two effects. First, biochemically, it modulates several known downstream proteins that control translation and reduces the expression of postsynaptic density-95 in vivo. Second, behaviorally, it attenuates long-term taste memory. The results suggest that the mTOR pathway in the cortex modulates both translation factor activity and protein expression, to enable normal taste memory consolidation.

  20. Chronic overload induced hypertrophy is associated with age-related muscle mass loss and diminished mTOR signaling

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The purpose of this study was to assess activation of the mTOR signaling pathway in young and aging rats in response to chronic muscle overload. Young (6 mo; n = 16) and older (30 mo; n = 23) male rats (F344xBN) were subjected to 4 weeks of bilateral surgical ablation (SA) of two-thirds of the gastr...

  1. Downregulation of cancer stem cell properties via mTOR signaling pathway inhibition by rapamycin in nasopharyngeal carcinoma

    PubMed Central

    YANG, CHUNGUANG; ZHANG, YUE; ZHANG, YU; ZHANG, ZIHENG; PENG, JIANHUA; LI, ZHI; HAN, LIANG; YOU, QUANJIE; CHEN, XIAOYU; RAO, XINGWANG; ZHU, YI; LIAO, ZHISU

    2015-01-01

    Rapamycin, a mammalian target of rapamycin (mTOR) signaling inhibitor, inhibits cancer cell proliferation and tumor formation, including in nasopharyngeal carcinoma (NPC), which we proved in a previous study. However, whether rapamycin affects cancer stem cells (CSCs) is unclear. In examining samples of NPCs, we found regions of CD44-positive cancer cells co-expressing the stem cell biomarker OCT4, suggesting the presence of CSCs. Following this, we used double-label immunohistochemistry to identify whether the mTOR signaling pathway was activated in CD44-positive CSCs in NPCs. We used a CCK-8 assay and western blotting to explore whether the stem cell biomarkers CD44 and SOX2 and the invasion protein MMP-2 could be suppressed by treatment with rapamycin in cultured primary NPC cells and secondary tumors in BALB/c nude mice. Interestingly, we found that rapamycin inhibited mTOR signaling in addition to simultaneously downregulating the expression of CD44, SOX2 and MMP-2 and that it affected cell growth and tumor size and weight both in vitro and in vivo. Collectively, we confirmed for the first time that CSC properties are reduced and invasion potential is restrained in response to mTOR signaling inhibition in NPC. This evidence indicates that the targeted inhibition of CSC properties may provide a novel strategy to treat cancer. PMID:26202311

  2. Alleviation of neuronal energy deficiency by mTOR inhibition as a treatment for mitochondria-related neurodegeneration

    PubMed Central

    Zheng, Xinde; Boyer, Leah; Jin, Mingji; Kim, Yongsung; Fan, Weiwei; Bardy, Cedric; Berggren, Travis; Evans, Ronald M; Gage, Fred H; Hunter, Tony

    2016-01-01

    mTOR inhibition is beneficial in neurodegenerative disease models and its effects are often attributable to the modulation of autophagy and anti-apoptosis. Here, we report a neglected but important bioenergetic effect of mTOR inhibition in neurons. mTOR inhibition by rapamycin significantly preserves neuronal ATP levels, particularly when oxidative phosphorylation is impaired, such as in neurons treated with mitochondrial inhibitors, or in neurons derived from maternally inherited Leigh syndrome (MILS) patient iPS cells with ATP synthase deficiency. Rapamycin treatment significantly improves the resistance of MILS neurons to glutamate toxicity. Surprisingly, in mitochondrially defective neurons, but not neuroprogenitor cells, ribosomal S6 and S6 kinase phosphorylation increased over time, despite activation of AMPK, which is often linked to mTOR inhibition. A rapamycin-induced decrease in protein synthesis, a major energy-consuming process, may account for its ATP-saving effect. We propose that a mild reduction in protein synthesis may have the potential to treat mitochondria-related neurodegeneration. DOI: http://dx.doi.org/10.7554/eLife.13378.001 PMID:27008180

  3. Activation of the mTOR Pathway by Oxaliplatin in the Treatment of Colorectal Cancer Liver Metastasis

    PubMed Central

    Lu, Min; Zessin, Amelia S.; Glover, Wayne

    2017-01-01

    Background Standard of care treatment for colorectal cancer liver metastasis consists of a cytotoxic chemotherapy in combination with a targeted agent. Clinical trials have guided the use of these combinatory therapies, but it remains unclear what the optimal combinations of cytotoxic chemotherapy with a targeted agent are. Methods Using a genomic based approach, gene expression profiling was obtained from tumor samples of patient with colorectal cancer liver metastasis who received an oxaliplatin based therapy. Early passaged colorectal cancer liver metastasis cell lines and patient derived xenografts of colorectal cancer liver metastasis were then treated with oxaliplatin and a mTOR inhibitor. Results Gene set enrichment analysis revealed that the mTOR pathway was activated in patients receiving oxaliplatin based therapy. Treatment of early passaged colorectal cancer lines and patient derived xenografts with oxaliplatin resulted in activation of the mTOR pathway. Combination therapy with oxaliplatin and a mTOR inhibitor resulted in a synergistic effect both in vitro and in vivo. Conclusion Our findings suggest a genomic based approach can be used to identify optimal combinations of cytotoxic chemotherapy with a targeted agent and that these observations can be validated both in vitro and in vivo using patient derived colorectal cancer cell lines and patient derived xenografts prior to clinical use. PMID:28060954

  4. Down-regulation of mTOR leads to up-regulation of osteoprotegerin in bone marrow cells

    SciTech Connect

    Mogi, Makio; Kondo, Ayami

    2009-06-19

    Osteoprotegerin (OPG)/osteoclastogenesis inhibitory factor regulates bone mass by inhibiting osteoclastic bone resorption. mTOR, which is the mammalian target of rapamycin, is a kinase and central regulator of cell growth, proliferation, and survival. By using Rapamycin, we studied whether mTOR pathway is associated with OPG protein production in the mouse bone marrow-derived stromal cell line ST2. Rapamycin markedly increased the level of soluble OPG in ST2 cells. This antibiotic treatment resulted in the suppression of phosphorylation of mTOR. Rapamycin had no effects on the proliferation, differentiation, or apoptosis of the cells. Treatment with bone morphogenetic protein-4, which can induce OPG protein in ST2 cells, also resulted in a decrease in the density of the phospho-mTOR-band, suggesting that the suppression of the phospho-mTOR pathway is necessary for OPG production in ST2 cells. Thus, suitable suppression of mTOR phosphorylation is a necessary requirement for OPG production in bone marrow stromal cells.

  5. Inhibition of 11β-Hydroxysteroid Dehydrogenase Type II Suppresses Lung Carcinogenesis by Blocking Tumor COX-2 Expression as Well as the ERK and mTOR Signaling Pathways

    PubMed Central

    Yang, Shilin; Yao, Bing; Zhang, Bixiang; Chen, Xiaoping; Pozzi, Ambra; Zhang, Ming-Zhi

    2015-01-01

    Lung cancer is by far the leading cause of cancer death. Early diagnosis and prevention remain the best approach to reduce the overall morbidity and mortality. Experimental and clinical evidence have shown that cyclooxygenase-2 (COX-2) derived prostaglandin E2 (PGE2) contributes to lung tumorigenesis. COX-2 inhibitors suppress the development and progression of lung cancer. However, increased cardiovascular risks of COX-2 inhibitors limit their use in chemoprevention of lung cancers. Glucocorticoids are endogenous and potent COX-2 inhibitors, and their local actions are down-regulated by 11β–hydroxysteroid dehydrogenase type II (11ßHSD2)-mediated metabolism. We found that 11βHSD2 expression was increased in human lung cancers and experimental lung tumors. Inhibition of 11βHSD2 activity enhanced glucocorticoid-mediated COX-2 inhibition in human lung carcinoma cells. Furthermore, 11βHSD2 inhibition suppressed lung tumor growth and invasion in association with increased tissue active glucocorticoid levels, decreased COX-2 expression, inhibition of ERK and mTOR signaling pathways, increased tumor endoplasmic reticulum stress as well as increased lifespan. Therefore, 11βHSD2 inhibition represents a novel approach for lung cancer chemoprevention and therapy by increasing tumor glucocorticoid activity, which in turn selectively blocks local COX-2 activity and/or inhibits the ERK and mTOR signaling pathways. PMID:26011146

  6. Potential anti-aging agents suppress the level of constitutive mTOR- and DNA damage- signaling.

    PubMed

    Halicka, H Dorota; Zhao, Hong; Li, Jiangwei; Lee, Yong-Syu; Hsieh, Tze-Chen; Wu, Joseph M; Darzynkiewicz, Zbigniew

    2012-12-01

    Two different mechanisms are considered to be the primary cause of aging. Cumulative DNA damage caused by reactive oxygen species (ROS), the by-products of oxidative phosphorylation, is one of these mechanisms (ROS concept). Constitutive stimulation of mitogen- and nutrient-sensing mTOR/S6 signaling is the second mechanism (TOR concept). The flow- and laser scanning- cytometric methods were developed to measure the level of the constitutive DNA damage/ROS- as well as of mTOR/S6- signaling in individual cells. Specifically, persistent activation of ATM and expression of γH2AX in untreated cells appears to report constitutive DNA damage induced by endogenous ROS. The level of phosphorylation of Ser235/236-ribosomal protein (RP), of Ser2448-mTOR and of Ser65-4EBP1, informs on constitutive signaling along the mTOR/S6 pathway. Potential gero-suppressive agents rapamycin, metformin, 2-deoxyglucose, berberine, resveratrol, vitamin D3 and aspirin, all decreased the level of constitutive DNA damage signaling as seen by the reduced expression of γH2AX in proliferating A549, TK6, WI-38 cells and in mitogenically stimulated human lymphocytes. They all also decreased the level of intracellular ROS and mitochondrial trans-membrane potential ΔΨm, the marker of mitochondrial energizing as well as reduced phosphorylation of mTOR, RP-S6 and 4EBP1. The most effective was rapamycin. Although the primary target of each on these agents may be different the data are consistent with the downstream mechanism in which the decline in mTOR/S6K signaling and translation rate is coupled with a decrease in oxidative phosphorylation, (revealed by ΔΨm) that leads to reduction of ROS and oxidative DNA damage. The decreased rate of translation induced by these agents may slow down cells hypertrophy and alleviate other features of cell aging/senescence. Reduction of oxidative DNA damage may lower predisposition to neoplastic transformation which otherwise may result from errors in repair of DNA

  7. CDK1 substitutes for mTOR kinase to activate mitotic cap-dependent protein translation

    PubMed Central

    Shuda, Masahiro; Velásquez, Celestino; Cheng, Erdong; Cordek, Daniel G.; Kwun, Hyun Jin; Chang, Yuan; Moore, Patrick S.

    2015-01-01

    Mitosis is commonly thought to be associated with reduced cap-dependent protein translation. Here we show an alternative control mechanism for maintaining cap-dependent translation during mitosis revealed by a viral oncoprotein, Merkel cell polyomavirus small T (MCV sT). We find MCV sT to be a promiscuous E3 ligase inhibitor targeting the anaphase-promoting complex, which increases cell mitogenesis. MCV sT binds through its Large T stabilization domain region to cell division cycle protein 20 (Cdc20) and, possibly, cdc20 homolog 1 (Cdh1) E3 ligase adapters. This activates cyclin-dependent kinase 1/cyclin B1 (CDK1/CYCB1) to directly hyperphosphorylate eukaryotic initiation factor 4E (eIF4E)-binding protein (4E-BP1) at authentic sites, generating a mitosis-specific, mechanistic target of rapamycin (mTOR) inhibitor-resistant δ phospho-isoform not present in G1-arrested cells. Recombinant 4E-BP1 inhibits capped mRNA reticulocyte translation, which is partially reversed by CDK1/CYCB1 phosphorylation of 4E-BP1. eIF4G binding to the eIF4E–m7GTP cap complex is resistant to mTOR inhibition during mitosis but sensitive during interphase. Flow cytometry, with and without sT, reveals an orthogonal pH3S10+ mitotic cell population having higher inactive p4E-BP1T37/T46+ saturation levels than pH3S10– interphase cells. Using a Click-iT flow cytometric assay to directly measure mitotic protein synthesis, we find that most new protein synthesis during mitosis is cap-dependent, a result confirmed using the eIF4E/4G inhibitor drug 4E1RCat. For most cell lines tested, cap-dependent translation levels were generally similar between mitotic and interphase cells, and the majority of new mitotic protein synthesis was cap-dependent. These findings suggest that mitotic cap-dependent translation is generally sustained during mitosis by CDK1 phosphorylation of 4E-BP1 even under conditions of reduced mTOR signaling. PMID:25883264

  8. CDK1 substitutes for mTOR kinase to activate mitotic cap-dependent protein translation.

    PubMed

    Shuda, Masahiro; Velásquez, Celestino; Cheng, Erdong; Cordek, Daniel G; Kwun, Hyun Jin; Chang, Yuan; Moore, Patrick S

    2015-05-12

    Mitosis is commonly thought to be associated with reduced cap-dependent protein translation. Here we show an alternative control mechanism for maintaining cap-dependent translation during mitosis revealed by a viral oncoprotein, Merkel cell polyomavirus small T (MCV sT). We find MCV sT to be a promiscuous E3 ligase inhibitor targeting the anaphase-promoting complex, which increases cell mitogenesis. MCV sT binds through its Large T stabilization domain region to cell division cycle protein 20 (Cdc20) and, possibly, cdc20 homolog 1 (Cdh1) E3 ligase adapters. This activates cyclin-dependent kinase 1/cyclin B1 (CDK1/CYCB1) to directly hyperphosphorylate eukaryotic initiation factor 4E (eIF4E)-binding protein (4E-BP1) at authentic sites, generating a mitosis-specific, mechanistic target of rapamycin (mTOR) inhibitor-resistant δ phospho-isoform not present in G1-arrested cells. Recombinant 4E-BP1 inhibits capped mRNA reticulocyte translation, which is partially reversed by CDK1/CYCB1 phosphorylation of 4E-BP1. eIF4G binding to the eIF4E-m(7)GTP cap complex is resistant to mTOR inhibition during mitosis but sensitive during interphase. Flow cytometry, with and without sT, reveals an orthogonal pH3(S10+) mitotic cell population having higher inactive p4E-BP1(T37/T46+) saturation levels than pH3(S10-) interphase cells. Using a Click-iT flow cytometric assay to directly measure mitotic protein synthesis, we find that most new protein synthesis during mitosis is cap-dependent, a result confirmed using the eIF4E/4G inhibitor drug 4E1RCat. For most cell lines tested, cap-dependent translation levels were generally similar between mitotic and interphase cells, and the majority of new mitotic protein synthesis was cap-dependent. These findings suggest that mitotic cap-dependent translation is generally sustained during mitosis by CDK1 phosphorylation of 4E-BP1 even under conditions of reduced mTOR signaling.

  9. Respiratory syncytial virus induces phosphorylation of mTOR at ser2448 in CD8 T cells from nasal washes of infected infants.

    PubMed

    de Souza, A P Duarte; de Freitas, D Nascimento; Antuntes Fernandes, K E; D'Avila da Cunha, M; Antunes Fernandes, J L; Benetti Gassen, R; Fazolo, T; Pinto, L A; Scotta, M; Mattiello, R; Pitrez, P M; Bonorino, C; Stein, R T

    2016-02-01

    Respiratory syncytial virus (RSV)-specific CD8(+) T cell responses do not protect against reinfection. Activation of mammalian target of rapamycin (mTOR) impairs memory CD8(+) T cell differentiation. Our hypothesis was that RSV inhibits the formation of CD8(+) T cells memory responses through mTOR activation. To explore this, human and mouse T cells were used. RSV induced mTOR phosphorylation at Ser2448 in CD8 T cells. mTOR activation by RSV was completely inhibited using rapamycin. RSV-infected children presented higher mTOR gene expression on nasal washes comparing to children infected with metapneumovirus and rhinovirus. In addition, RSV-infected infants presented a higher frequency of CD8(+) pmTORser2448(+) T cells in nasal washes compared to RSV-negative infants. Rapamycin treatment increased the frequency of mouse CD8 RSV-M282-90 pentamer-positive T cells and the frequency of RSV-specific memory T cells precursors. These data demonstrate that RSV is activating mTOR directly in CD8 T cells, indicating a role for mTOR during the course of RSV infection.

  10. Requirement for the eIF4E binding proteins for the synergistic down-regulation of protein synthesis by hypertonic conditions and mTOR inhibition.

    PubMed

    Clemens, Michael J; Elia, Androulla; Morley, Simon J

    2013-01-01

    The protein kinase mammalian target of rapamycin (mTOR) regulates the phosphorylation and activity of several proteins that have the potential to control translation, including p70S6 kinase and the eIF4E binding proteins 4E-BP1 and 4E-BP2. In spite of this, in exponentially growing cells overall protein synthesis is often resistant to mTOR inhibitors. We report here that sensitivity of wild-type mouse embryonic fibroblasts (MEFs) to mTOR inhibitors can be greatly increased when the cells are subjected to the physiological stress imposed by hypertonic conditions. In contrast, protein synthesis in MEFs with a double knockout of 4E-BP1 and 4E-BP2 remains resistant to mTOR inhibitors under these conditions. Phosphorylation of p70S6 kinase and protein kinase B (Akt) is blocked by the mTOR inhibitor Ku0063794 equally well in both wild-type and 4E-BP knockout cells, under both normal and hypertonic conditions. The response of protein synthesis to hypertonic stress itself does not require the 4E-BPs. These data suggest that under certain stress conditions: (i) translation has a greater requirement for mTOR activity and (ii) there is an absolute requirement for the 4E-BPs for regulation by mTOR. Importantly, dephosphorylation of p70S6 kinase and Akt is not sufficient to affect protein synthesis acutely.

  11. Requirement for lysosomal localization of mTOR for its activation differs between leucine and other amino acids.

    PubMed

    Averous, Julien; Lambert-Langlais, Sarah; Carraro, Valérie; Gourbeyre, Ophélie; Parry, Laurent; B'Chir, Wafa; Muranishi, Yuki; Jousse, Céline; Bruhat, Alain; Maurin, Anne-Catherine; Proud, Christopher G; Fafournoux, Pierre

    2014-09-01

    The mammalian target of rapamycin complex 1 (mTORC1) is a master regulator of cell growth and metabolism. It controls many cell functions by integrating nutrient availability and growth factor signals. Amino acids, and in particular leucine, are among the main positive regulators of mTORC1 signaling. The current model for the regulation of mTORC1 by amino acids involves the movement of mTOR to the lysosome mediated by the Rag-GTPases. Here, we have examined the control of mTORC1 signaling and mTOR localization by amino acids and leucine in serum-fed cells, because both serum growth factors (or, e.g., insulin) and amino acids are required for full activation of mTORC1 signaling. We demonstrate that mTORC1 activity does not closely correlate with the lysosomal localization of mTOR. In particular, leucine controls mTORC1 activity without any detectable modification of the lysosomal localization of mTOR, indicating that the signal(s) exerted by leucine is likely distinct from those exerted by other amino acids. In addition, knock-down of the Rag-GTPases attenuated the inhibitory effect of amino acid- or leucine-starvation on the phosphorylation of mTORC1 targets. Furthermore, data from cells where Rag expression has been knocked down revealed that leucine can promote mTORC1 signaling independently of the lysosomal localization of mTOR. Our data complement existing models for the regulation of mTORC1 by amino acids and provide new insights into this important topic.

  12. Delineating the mTOR kinase pathway using a dual TORC1/2 inhibitor, AZD8055, in multiple myeloma.

    PubMed

    Cirstea, Diana; Santo, Loredana; Hideshima, Teru; Eda, Homare; Mishima, Yuko; Nemani, Neeharika; Mahindra, Anuj; Yee, Andrew; Gorgun, Gullu; Hu, Yiguo; Ohguchi, Hiroto; Suzuki, Rikio; Cottini, Francesca; Guichard, Sylvie M; Anderson, Kenneth C; Raje, Noopur

    2014-11-01

    Despite promising preclinical results with mTOR kinase inhibitors in multiple myeloma, resistance to these drugs may arise via feedback activation loops. This concern is especially true for insulin-like growth factor 1 receptor (IGF1R), because IGF1R signaling is downregulated by multiple AKT and mTOR feedback mechanisms. We have tested this hypothesis in multiple myeloma using the novel selective mTOR kinase inhibitor AZD8055. We evaluated p-mTOR S(2481) as the readout for mTORC2/Akt activity in multiple myeloma cells in the context of mTOR inhibition via AZD8055 or rapamycin. We next validated AZD8055 inhibition of mTORC1 and mTORC2 functions in multiple myeloma cells alone or in culture with bone marrow stroma cells and growth factors. Unlike rapamycin, AZD8055 resulted in apoptosis of multiple myeloma cells. AZD8055 treatment, however, induced upregulation of IGF1R phosphorylation in p-Akt S(473)-expressing multiple myeloma cell lines. Furthermore, exposure of AZD8055-treated cells to IGF1 induced p-Akt S(473) and rescued multiple myeloma cells from apoptosis despite mTOR kinase inhibition and TORC2/Akt blockage. The addition of blocking IGF1R antibody resulted in reversing this effect and increased AZD8055-induced apoptosis. Our study suggests that combination treatment with AZD8055 and IGF1R-blocking agents is a promising strategy in multiple myeloma with potential IGF1R/Akt signaling-mediated survival.

  13. Systemic inhibition of mTOR kinase via rapamycin disrupts consolidation and reconsolidation of auditory fear memory.

    PubMed

    Mac Callum, Phillip E; Hebert, Mark; Adamec, Robert E; Blundell, Jacqueline

    2014-07-01

    The mammalian target of rapamycin (mTOR) kinase is a critical regulator of mRNA translation and is known to be involved in various long lasting forms of synaptic and behavioural plasticity. However, information concerning the temporal pattern of mTOR activation and susceptibility to pharmacological intervention during both consolidation and reconsolidation of long-term memory (LTM) remains scant. Male C57BL/6 mice were injected systemically with rapamycin at various time points following conditioning or retrieval in an auditory fear conditioning paradigm, and compared to vehicle (and/or anisomycin) controls for subsequent memory recall. Systemic blockade of mTOR with rapamycin immediately or 12h after training or reactivation impairs both consolidation and reconsolidation of an auditory fear memory. Further behavioural analysis revealed that the enduring effects of rapamycin on reconsolidation are dependent upon reactivation of the memory trace. Rapamycin, however, has no effect on short-term memory or the ability to retrieve an established fear memory. Collectively, our data suggest that biphasic mTOR signalling is essential for both consolidation and reconsolidation-like activities that contribute to the formation, re-stabilization, and persistence of long term auditory-fear memories, while not influencing other aspects of the memory trace. These findings also provide evidence for a cogent treatment model for reducing the emotional strength of established, traumatic memories analogous to those observed in acquired anxiety disorders such as posttraumatic stress disorder (PTSD) and specific phobias, through pharmacologic blockade of mTOR</