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

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

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

scholarly journals mTOR Inhibition: From Aging to Autism and Beyond

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

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

scholarly journals Transcriptional Regulation of BK Virus by Nuclear Factor of Activated T Cells

2009 ◽  
Vol 84 (4) ◽  
pp. 1722-1730 ◽  
Author(s):  
Joslynn A. Jordan ◽  
Kate Manley ◽  
Aisling S. Dugan ◽  
Bethany A. O'Hara ◽  
Walter J. Atwood
Keyword(s):  
T Cells ◽  
Promoter Activity ◽  
Mutational Analysis ◽  
Adult Population ◽  
Bk Virus ◽  
Human Polyomavirus ◽  
Luciferase Reporter ◽  
Nuclear Factor ◽  
Activated T Cells ◽  
Transplant Patients

ABSTRACT The human polyomavirus BK virus (BKV) is a common virus for which 80 to 90% of the adult population is seropositive. BKV reactivation in immunosuppressed patients or renal transplant patients is the primary cause of polyomavirus-associated nephropathy (PVN). Using the Dunlop strain of BKV, we found that nuclear factor of activated T cells (NFAT) plays an important regulatory role in BKV infection. Luciferase reporter assays and chromatin immunoprecipitation assays demonstrated that NFAT4 bound to the viral promoter and regulated viral transcription and infection. The mutational analysis of the NFAT binding sites demonstrated complex functional interactions between NFAT, c-fos, c-jun, and the p65 subunit of NF-κB that together influence promoter activity and viral growth. These data indicate that NFAT is required for BKV infection and is involved in a complex regulatory network that both positively and negatively influences promoter activity and viral infection.

scholarly journals Analysis of 15 novel full-length BK virus sequences from three individuals: evidence of a high intra-strain genetic diversity

2004 ◽  
Vol 85 (9) ◽  
pp. 2651-2663 ◽  
Author(s):  
Yiping Chen ◽  
Paul M. Sharp ◽  
Mary Fowkes ◽  
Olivier Kocher ◽  
Jeffrey T. Joseph ◽  
...  
Keyword(s):  
Striated Muscle ◽  
Jc Virus ◽  
Regulatory Region ◽  
Bk Virus ◽  
Full Length ◽  
Human Polyomavirus ◽  
Coding Region ◽  
Nucleotide Substitutions ◽  
Source Of Infection

To determine the variability of BK virus (BKV) in vivo, the sequences of nine full-length molecular clones from the striated muscle and heart DNA of a patient with BKV-associated capillary leak syndrome (BKVCAP), as well as three clones each from the urine of one human immunodeficiency virus type 2-positive (BKVHI) and one healthy control subject (BKVHC), were analysed. The regulatory region of all clones corresponded to the archetypal regulatory region usually found in urine isolates. Analysis of the predicted conformation of BKVCAP proteins did not suggest any structural differences on the surface of the viral particles compared with BKVHI and BKVHC clones. No amino acid changes common to most BKVCAP clones could be identified that have not already been reported in non-vasculotropic strains. However, the coding region of each clone had unique nucleotide substitutions, and intra-host variability was greater among BKVCAP clones, with a mean difference of 0·29 % per site compared with 0·16 % for BKVHI and 0·14 % for BKVHC. The clones from each strain formed monophyletic clades, suggesting a single source of infection for each subject. The most divergent BKVCAP clones differed at 0·55 % of sites, implying a rate of nucleotide substitution of approximately 5×10−5 substitutions per site per year, which is two orders of magnitude faster than estimated for the other human polyomavirus, JC virus.

scholarly journals Mammalian target of rapamycin inhibitors rapamycin and RAD001 (everolimus) induce anti-proliferative effects in GH-secreting pituitary tumor cells in vitro

2009 ◽  
Vol 16 (3) ◽  
pp. 1017-1027 ◽  
Author(s):  
Alexander Gorshtein ◽  
Hadara Rubinfeld ◽  
Efrat Kendler ◽  
Marily Theodoropoulou ◽  
Vesna Cerovac ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Cyclin D1 ◽  
Pituitary Adenomas ◽  
Pituitary Tumor ◽  
Mammalian Target Of Rapamycin ◽  
Pituitary Tumors ◽  
Mtor Inhibitors ◽  
Target Of Rapamycin ◽  
Gh3 Cells ◽  
Mtor Inhibition

The effect of mammalian target of rapamycin (mTOR) inhibitors on pituitary tumors is unknown. Akt overexpression was demonstrated in pituitary adenomas, which may render them sensitive to the anti-proliferative effects of these drugs. The objective of the study was to evaluate the anti-proliferative efficacy of the mTOR inhibitor, rapamycin, and its orally bioavailable analog RAD001 on the GH-secreting pituitary tumor GH3 and MtT/S cells and in human GH-secreting pituitary adenomas (GH-omas) in primary cell cultures. Treatment with rapamycin or RAD001 significantly decreased the number of viable cells and cell proliferation in a dose- and time-dependent manner. This was reflected by decreased phosphorylation levels of the downstream mTOR target p70S6K. Rapamycin treatment of GH3 cells induced G0/G1 cell cycle arrest. In other tumor cell types, this was attributed to a decrease in cyclin D1 levels. However, rapamycin did not affect cyclin D1 protein levels in GH3 cells. By contrast, it decreased cyclin D3 and p21/CIP, which stabilizes cyclin D/cyclin-dependent kinase 4 (cdk4) complexes. Rapamycin inhibited FCS-induced retinoblastoma phosphorylation and subsequent E2F-transcriptional activity. In response to decreased E2F activity, the expression of the E2F-regulated genes cyclin E and cdk2 was reduced. Our results showed that mTOR inhibitors potently inhibit pituitary cell proliferation, suggesting that mTOR inhibition may be a promising anti-proliferative therapy for pituitary adenomas. This therapeutic manipulation may have beneficial effects particularly for patients harboring invasive pituitary tumors resistant to current treatments.

scholarly journals Selective Deletion of the Mechanistic Target of Rapamycin From the Renal Collecting Duct Principal Cell in Mice Down-Regulates the Epithelial Sodium Channel

2022 ◽  
Vol 12 ◽  
Author(s):  
Bruce Chen ◽  
Maurice B. Fluitt ◽  
Aaron L. Brown ◽  
Samantha Scott ◽  
Anirudh Gadicherla ◽  
...  
Keyword(s):  
Sodium Channel ◽  
Collecting Duct ◽  
Epithelial Sodium Channel ◽  
Principal Cell ◽  
Target Of Rapamycin ◽  
Mtor Inhibition ◽  
Sodium Homeostasis ◽  
Mechanistic Target Of Rapamycin ◽  
Data Support ◽  
Body Sodium

The mechanistic target of rapamycin (mTOR), a serine-threonine-specific kinase, is a cellular energy sensor, integrating growth factor and nutrient signaling. In the collecting duct (CD) of the kidney, the epithelial sodium channel (ENaC) essential in the determination of final urine Na+ losses, has been demonstrated to be upregulated by mTOR, using cell culture and mTOR inhibition in ex vivo preparations. We tested whether CD-principal cell (PC) targeted deletion of mTOR using Cre-lox recombination would affect whole-body sodium homeostasis, blood pressure, and ENaC regulation in mice. Male and female CD-PC mTOR knockout (KO) mice and wild-type (WT) littermates (Cre-negative) were generated using aquaporin-2 (AQP2) promoter to drive Cre-recombinase. Under basal conditions, KO mice showed a reduced (∼30%) natriuretic response to benzamil (ENaC) antagonist, suggesting reduced in vivo ENaC activity. WT and KO mice were fed normal sodium (NS, 0.45% Na+) or a very low Na+ (LS, <0.02%) diet for 7-days. Switching from NS to LS resulted in significantly higher urine sodium losses (relative to WT) in the KO with adaptation occurring by day 2. Blood pressures were modestly (∼5–10 mm Hg) but significantly lower in KO mice under both diets. Western blotting showed KO mice had 20–40% reduced protein levels of all three subunits of ENaC under LS or NS diet. Immunohistochemistry (IHC) of kidney showed enhanced apical-vs.-cellular localization of all three subunits with LS, but a reduction in this ratio for γ-ENaC in the KO. Furthermore, the KO kidneys showed increased ubiquitination of α-ENaC and reduced phosphorylation of the serum and glucocorticoid regulated kinase, type 1 [serum glucocorticoid regulated kinase (SGK1)] on serine 422 (mTOR phosphorylation site). Taken together this suggests enhanced degradation as a consequence of reduced mTOR kinase activity and downstream upregulation of ubiquitination may have accounted for the reduction at least in α-ENaC. Overall, our data support a role for mTOR in ENaC activity likely via regulation of SGK1, ubiquitination, ENaC channel turnover and apical membrane residency. These data support a role for mTOR in the collecting duct in the maintenance of body sodium homeostasis.

scholarly journals Restriction of Human Polyomavirus BK Virus DNA Replication in Murine Cells and Extracts

2009 ◽  
Vol 83 (11) ◽  
pp. 5708-5717 ◽  
Author(s):  
Cathal Mahon ◽  
Bo Liang ◽  
Irina Tikhanovich ◽  
Johanna R. Abend ◽  
Michael J. Imperiale ◽  
...  
Keyword(s):  
Dna Replication ◽  
Topoisomerase I ◽  
Simian Virus 40 ◽  
T Antigen ◽  
Bk Virus ◽  
Human Polyomavirus ◽  
Expression Vectors ◽  
Viral Dna ◽  
Viral Dna Replication ◽  
Cell Extracts

ABSTRACT BK virus (BKV) causes persistent and asymptomatic infections in most humans and is the etiologic agent of polyomavirus-associated nephropathy (PVAN) and other pathologies. Unfortunately, there are no animal models with which to study activation of BKV replication in the human kidney and the accompanying PVAN. Here we report studies of the restriction of BKV replication in murine cells and extracts and the cause(s) of this restriction. Upon infection of murine cells, BKV expressed large T antigen (TAg), but viral DNA replication and progeny were not detected. Transfection of murine cells with BKV TAg expression vectors also caused TAg expression without accompanying DNA replication. Analysis of the replication of DNAs containing chimeric BKV and murine polyomavirus origins revealed the importance of BKV core origin sequences and TAg for DNA replication. A sensitive assay was developed with purified BKV TAg that supported TAg-dependent BKV DNA replication with human but not with murine cell extracts. Addition of human replication proteins, DNA polymerase α-primase, replication protein A, or topoisomerase I to the murine extracts with BKV TAg did not rescue viral DNA replication. Notably, addition of murine extracts to human extracts inhibited BKV TAg-dependent DNA replication at a step prior to or during unwinding of the viral origin. These findings and differences in replication specificity between BKV TAg and the TAgs of simian virus 40 (SV40) and JC virus (JCV) and their respective origins implicate features of the BKV TAg and origin distinct from SV40 and JCV in restriction of BKV replication in murine cells.

scholarly journals Fructose leads to hepatic steatosis in zebrafish that is reversed by mechanistic target of rapamycin (mTOR) inhibition

Hepatology ◽  
2014 ◽  
Vol 60 (5) ◽  
pp. 1581-1592 ◽  
Author(s):  
Valerie Sapp ◽  
Leah Gaffney ◽  
Steven F. EauClaire ◽  
Randolph P. Matthews
Keyword(s):  
Hepatic Steatosis ◽  
Target Of Rapamycin ◽  
Mtor Inhibition ◽  
Mechanistic Target Of Rapamycin

scholarly journals Mechanistic Target of Rapamycin Inhibitors in Renal Cell Carcinoma: Potential, Limitations, and Perspectives

2021 ◽  
Vol 9 ◽  
Author(s):  
Seraina Faes ◽  
Nicolas Demartines ◽  
Olivier Dormond
Keyword(s):  
Renal Cell Carcinoma ◽  
Cell Proliferation ◽  
Cell Carcinoma ◽  
Renal Cell ◽  
Mtor Inhibitors ◽  
Target Of Rapamycin ◽  
Cancer Cell Proliferation ◽  
Hypoxia Inducible Factors ◽  
Mtor Signaling Pathway ◽  
Mechanistic Target Of Rapamycin

Several elements highlight the importance of the mechanistic target of rapamycin (mTOR) in the biology of renal cell carcinoma (RCC). mTOR signaling pathway is indeed frequently activated in RCC, inducing cancer cell proliferation and survival. In addition, mTOR promotes tumor angiogenesis and regulates the expression of hypoxia-inducible factors that play an important role in a subset of RCC. Despite mTOR protumorigenic effects, mTOR inhibitors have failed to provide long-lasting anticancer benefits in RCC patients, highlighting the need to readdress their role in the treatment of RCC. This review aims to present the rationale and limitations of targeting mTOR in RCC. Future roles of mTOR inhibitors in the treatment of RCC are also discussed, in particular in the context of immunotherapies.

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