Dual mTORC1/mTORC2 inhibition as a Host-Directed Therapeutic Target in Pathologically Distinct Mouse Models of Tuberculosis

Efforts to develop more effective and shorter-course therapies for tuberculosis have included a focus on host-directed therapy (HDT). The goal of HDT is to modulate the host response to infection, thereby improving immune defenses to reduce the duration of antibacterial therapy and/or the amount of lung damage. As a mediator of innate and adaptive immune responses involved in eliminating intracellular pathogens, autophagy is a potential target for HDT in tuberculosis. Because Mycobacterium tuberculosis modulates mammalian target of rapamycin (mTOR) signaling to impede autophagy, pharmacologic mTOR inhibition could provide effective HDT. mTOR exists within two distinct multiprotein complexes, mTOR complex-1 (mTORC1) and mTOR complex-2 (mTORC2). Rapamycin and its analogs only partially inhibit mTORC1. We hypothesized that novel mTOR kinase inhibitors blocking both complexes would have expanded therapeutic potential. We compared the effects of two mTOR inhibitors: rapamycin and the orally available mTOR kinase domain inhibitor CC214-2, which blocks both mTORC1 and mTORC2, as adjunctive therapies against murine TB, when added to the first-line regimen (RHZE) or the novel bedaquiline-pretomanid-linezolid (BPaL) regimen. Neither mTOR inhibitor affected lung CFU counts after 4-8 weeks of treatment when combined with BPaL or RHZE. However, addition of CC214-2 to BPaL and RHZE was associated with significantly fewer relapses in C3HeB/FeJ compared to addition of rapamycin and, in RHZE-treated mice, resulted in fewer relapses compared to RHZE alone. Therefore, CC214-2 and related mTOR kinase inhibitors may be more effective candidates for HDT than rapamycin analogs and may have the potential to shorten the duration of TB treatment.

Download Full-text

Dual mTORC1/mTORC2 inhibition as a Host-Directed Therapeutic Target in Pathologically Distinct Mouse Models of Tuberculosis

10.1101/2021.02.10.430715 ◽

2021 ◽

Author(s):

Rokeya Tasneen ◽

Deborah S. Mortensen ◽

Paul J. Converse ◽

Michael E. Urbanowski ◽

Anna Upton ◽

...

Keyword(s):

Kinase Inhibitors ◽

Therapeutic Potential ◽

Mammalian Target Of Rapamycin ◽

Mtor Inhibitors ◽

Kinase Domain ◽

Lung Damage ◽

Mtor Inhibition ◽

Mtor Kinase ◽

Tb Treatment ◽

Mtor Complex 1

AbstractEfforts to develop more effective and shorter-course therapies for tuberculosis have included a focus on host-directed therapy (HDT). The goal of HDT is to modulate the host response to infection, thereby improving immune defenses to reduce the duration of antibacterial therapy and/or the amount of lung damage. As a mediator of innate and adaptive immune responses involved in eliminating intracellular pathogens, autophagy is a potential target for HDT in tuberculosis. Because Mycobacterium tuberculosis modulates mammalian target of rapamycin (mTOR) signaling to impede autophagy, pharmacologic mTOR inhibition could provide effective HDT. mTOR exists within two distinct multiprotein complexes, mTOR complex-1 (mTORC1) and mTOR complex-2 (mTORC2). Rapamycin and its analogs only partially inhibit mTORC1. We hypothesized that novel mTOR kinase inhibitors blocking both complexes would have expanded therapeutic potential. We compared the effects of two mTOR inhibitors: rapamycin and the orally available mTOR kinase domain inhibitor CC214-2, which blocks both mTORC1 and mTORC2, as adjunctive therapies against murine TB, when added to the first-line regimen (RHZE) or the novel bedaquiline-pretomanid-linezolid (BPaL) regimen. Neither mTOR inhibitor affected lung CFU counts after 4-8 weeks of treatment when combined with BPaL or RHZE. However, addition of CC214-2 to BPaL and RHZE was associated with significantly fewer relapses in C3HeB/FeJ compared to addition of rapamycin and, in RHZE-treated mice, resulted in fewer relapses compared to RHZE alone. Therefore, CC214-2 and related mTOR kinase inhibitors may be more effective candidates for HDT than rapamycin analogs and may have the potential to shorten the duration of TB treatment.

Download Full-text

mTOR-targeted cancer therapy: great target but disappointing clinical outcomes, why?

Frontiers of Medicine ◽

10.1007/s11684-020-0812-7 ◽

2020 ◽

Cited By ~ 1

Author(s):

Shi-Yong Sun

Keyword(s):

Cancer Therapy ◽

Kinase Inhibitors ◽

Mammalian Target Of Rapamycin ◽

Mtor Inhibitors ◽

Targeted Cancer Therapy ◽

Biological Functions ◽

New Findings ◽

Survival Signaling ◽

Mtor Complex 1

Abstract The mammalian target of rapamycin (mTOR) critically regulates several essential biological functions, such as cell growth, metabolism, survival, and immune response by forming two important complexes, namely, mTOR complex 1 (mTORC1) and complex 2 (mTORC2). mTOR signaling is often dysregulated in cancers and has been considered an attractive cancer therapeutic target. Great efforts have been made to develop efficacious mTOR inhibitors, particularly mTOR kinase inhibitors, which suppress mTORC1 and mTORC2; however, major success has not been achieved. With the strong scientific rationale, the intriguing question is why cancers are insensitive or not responsive to mTOR-targeted cancer therapy in clinics. Beyond early findings on induced activation of PI3K/Akt, MEK/ERK, and Mnk/eIF4E survival signaling pathways that compromise the efficacy of rapalog-based cancer therapy, recent findings on the essential role of GSK3 in mediating cancer cell response to mTOR inhibitors and mTORC1 inhibition-induced upregulation of PD-L1 in cancer cells may provide some explanations. These new findings may also offer us the opportunity to rationally utilize mTOR inhibitors in cancer therapy. Further elucidation of the biology of complicated mTOR networks may bring us the hope to develop effective therapeutic strategies with mTOR inhibitors against cancer.

Download Full-text

Conserved sequence motifs and the structure of the mTOR kinase domain

Biochemical Society Transactions ◽

10.1042/bst20130113 ◽

2013 ◽

Vol 41 (4) ◽

pp. 889-895 ◽

Cited By ~ 8

Author(s):

Evelyn Sauer ◽

Stefan Imseng ◽

Timm Maier ◽

Michael N. Hall

Keyword(s):

Crystal Structure ◽

Catalytic Domain ◽

Mammalian Target Of Rapamycin ◽

Mtor Inhibitors ◽

Homology Model ◽

Kinase Domain ◽

Sequence Motifs ◽

Serine Threonine Kinase ◽

Conserved Sequence ◽

Mtor Kinase

The atypical serine/threonine kinase mTOR (mammalian target of rapamycin) is a central regulator of cell growth and metabolism. mTOR is part of two multisubunit signalling complexes, mTORC1 and mTORC2. Although many aspects of mTOR signalling are understood, the lack of high-resolution structures impairs a detailed understanding of complex assembly, function and regulation. The structure of the kinase domain is of special interest for the development of mTOR inhibitors as anti-cancer agents. A homology model of the mTOR kinase domain was derived from the structure of PI3Ks (phosphoinositide 3-kinases). More recently, the crystal structure of the catalytic domain of human mTOR was determined, providing long-awaited structural insight into the architecture of mTOR. Interestingly, the homology model predicted several aspects of the crystal structure. In the present paper, we revisit the homology model in the context of the now available crystal structure of the mTOR kinase domain.

Download Full-text

Adaptation to chronic mTOR inhibition in cancer and in aging

Biochemical Society Transactions ◽

10.1042/bst20130080 ◽

2013 ◽

Vol 41 (4) ◽

pp. 956-961 ◽

Cited By ~ 11

Author(s):

Rebecca Gilley ◽

Kathryn Balmanno ◽

Claire L. Cope ◽

Simon J. Cook

Keyword(s):

Drug Target ◽

Kinase Inhibitors ◽

Mtor Inhibitors ◽

Allosteric Inhibitors ◽

Mtor Inhibition ◽

Cell Responses ◽

Downstream Effector ◽

Signalling Network ◽

Effector Pathways ◽

Mtor Complex 1

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

Download Full-text

Rapamycin-insensitive mTORC1 activity controls eIF4E:4E-BP1 binding

F1000Research ◽

10.12688/f1000research.1-4.v1 ◽

2012 ◽

Vol 1 ◽

pp. 4 ◽

Cited By ~ 26

Author(s):

Mark Livingstone ◽

Michael Bidinosti

Keyword(s):

Protein Complexes ◽

Mammalian Target Of Rapamycin ◽

Phosphorylation Site ◽

Mtor Inhibitors ◽

Kinase Domain ◽

Genetic Dissection ◽

Post Translational Modification ◽

Mtor Kinase ◽

Phosphorylation Event ◽

Mtor Protein

The recent development of mammalian target of rapamycin (mTOR) kinase domain inhibitors and genetic dissection of rapamycin-sensitive and -insensitive mTOR protein complexes (mTORC1 and mTORC2) have revealed that phosphorylation of the mTOR substrate 4E-BP1 on amino acids Thr37 and/or Thr46 represents a rapamycin-insensitive activity of mTORC1. Despite numerous previous reports utilizing serine (Ser)-to-alanine (Ala) and threonine (Thr)-to-Ala phosphorylation site mutants of 4E-BP1 to assess which post-translational modification(s) directly regulate binding to eIF4E, an ambiguous understanding persists. This manuscript demonstrates that the initial, rapamycin-insensitive phosphorylation event at Thr46 is sufficient to prevent eIF4E:4E-BP1 binding. This finding is relevant, particularly as mTOR kinase domain inhibitors continue to be assessed for clinical efficacy, since it clarifies a difference between the action of these second-generation mTOR inhibitors and those of rapamycin analogues.

Download Full-text

Pancreatic endocrine disorders and multiple endocrine neoplasia

Oxford Textbook of Medicine ◽

10.1093/med/9780198746690.003.0258 ◽

2020 ◽

pp. 2449-2463

Author(s):

B. Khoo ◽

T.M. Tan ◽

S.R. Bloom

Keyword(s):

Kinase Inhibitors ◽

Neuroendocrine Tumours ◽

Mammalian Target Of Rapamycin ◽

Peptide Receptor Radionuclide Therapy ◽

Mtor Inhibitors ◽

Somatostatin Analogues ◽

Endocrine Disorders ◽

Pancreatic Neuroendocrine Tumours ◽

Islet Cell Tumours ◽

Cancer Syndromes

Pancreatic neuroendocrine tumours (islet-cell tumours) are rare and usually sporadic, but they may be associated with complex familial endocrine cancer syndromes. Recognized types of pancreatic neuroendocrine tumours are those that are non-functioning (often advanced at diagnosis and presenting with mass effects due to the absence of symptoms attributable to hormone hypersecretion), insulinoma (the most frequent type), and others including gastrinoma, VIPoma, and glucagonoma. The following should be considered in addition to the symptomatic treatments: surgical resection—the only curative treatment, but not possible in many cases; tyrosine kinase inhibitors which inhibit specific kinases involved in tumour cell proliferation, growth, and angiogenesis; mammalian Target of Rapamycin (mTOR) inhibitors; peptide-receptor radionuclide therapy (radiolabelled somatostatin analogues).

Download Full-text

mTOR inhibitors lower an intrinsic barrier to virus infection mediated by IFITM3

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1811892115 ◽

2018 ◽

Vol 115 (43) ◽

pp. E10069-E10078 ◽

Cited By ~ 23

Author(s):

Guoli Shi ◽

Stosh Ozog ◽

Bruce E. Torbett ◽

Alex A. Compton

Keyword(s):

Influenza A ◽

Lentiviral Vector ◽

Sendai Virus ◽

Mammalian Target Of Rapamycin ◽

Mtor Inhibitors ◽

Endosomal Trafficking ◽

Mtor Inhibition ◽

Rapamycin Treatment ◽

Endosomal Sorting ◽

Basal Expression

Rapamycin and its derivatives are specific inhibitors of mammalian target of rapamycin (mTOR) kinase and, as a result, are well-established immunosuppressants and antitumorigenic agents. Additionally, this class of drug promotes gene delivery by facilitating lentiviral vector entry into cells, revealing its potential to improve gene therapy efforts. However, the precise mechanism was unknown. Here, we report that mTOR inhibitor treatment results in down-regulation of the IFN-induced transmembrane (IFITM) proteins. IFITM proteins, especially IFITM3, are potent inhibitors of virus–cell fusion and are broadly active against a range of pathogenic viruses. We found that the effect of rapamycin treatment on lentiviral transduction is diminished upon IFITM silencing or knockout in primary and transformed cells, and the extent of transduction enhancement depends on basal expression of IFITM proteins, with a major contribution from IFITM3. The effect of rapamycin treatment on IFITM3 manifests at the level of protein, but not mRNA, and is selective, as many other endosome-associated transmembrane proteins are unaffected. Rapamycin-mediated degradation of IFITM3 requires endosomal trafficking, ubiquitination, endosomal sorting complex required for transport (ESCRT) machinery, and lysosomal acidification. Since IFITM proteins exhibit broad antiviral activity, we show that mTOR inhibition also promotes infection by another IFITM-sensitive virus, Influenza A virus, but not infection by Sendai virus, which is IFITM-resistant. Our results identify the molecular basis by which mTOR inhibitors enhance virus entry into cells and reveal a previously unrecognized immunosuppressive feature of these clinically important drugs. In addition, this study uncovers a functional convergence between the mTOR pathway and IFITM proteins at endolysosomal membranes.

Download Full-text

New Frontiers in Mucositis

American Society of Clinical Oncology Educational Book ◽

10.14694/edbook_am.2012.32.46 ◽

2012 ◽

pp. 545-551 ◽

Cited By ~ 9

Author(s):

Douglas E. Peterson ◽

Dorothy M. Keefe ◽

Stephen T. Sonis

Keyword(s):

Kinase Inhibitors ◽

Management Strategies ◽

Mammalian Target Of Rapamycin ◽

Cancer Therapeutics ◽

Mtor Inhibitors ◽

Mucosal Injury ◽

Mucosal Damage ◽

Nucleotide Polymorphisms ◽

Oncology Patient ◽

Novel Technologies

Overview: Mucositis is among the most debilitating side effects of radiotherapy, chemotherapy, and targeted anticancer therapy. Research continues to escalate regarding key issues such as etiopathology, incidence and severity across different mucosae, relationships between mucosal and nonmucosal toxicities, and risk factors. This approach is being translated into enhanced management strategies. Recent technology advances provide an important foundation for this continuum. For example, evolution of applied genomics is fostering development of new algorithms to rapidly screen genomewide single-nucleotide polymorphisms (SNPs) for patient-associated risk prediction. This modeling will permit individual tailoring of the most effective, least toxic treatment in the future. The evolution of novel cancer therapeutics is changing the mucositis toxicity profile. These agents can be associated with unique mechanisms of mucosal damage. Additional research is needed to optimally manage toxicity caused by agents such as mammalian target of rapamycin (mTOR) inhibitors and tyrosine kinase inhibitors, without reducing antitumor effect. There has similarly been heightened attention across the health professions regarding clinical practice guidelines for mucositis management in the years following the first published guidelines in 2004. New opportunities exist to more effectively interface this collective guideline portfolio by capitalizing upon novel technologies such as an Internet-based Wiki platform. Substantive progress thus continues across many domains associated with mucosal injury in oncology patients. In addition to enhancing oncology patient care, these advances are being integrated into high-impact educational and scientific venues including the National Cancer Institute Physician Data Query (PDQ) portfolio as well as a new Gordon Research Conference on mucosal health and disease scheduled for June 2013.

Download Full-text

DEPTOR Is a Regulator of Response to mTOR Kinase Inhibitors in Multiple Myeloma

Blood ◽

10.1182/blood.v118.21.2916.2916 ◽

2011 ◽

Vol 118 (21) ◽

pp. 2916-2916

Author(s):

Diana Cirstea ◽

Teru Hideshima ◽

Loredana Santo ◽

Homare Eda ◽

Miriam Canavese ◽

...

Keyword(s):

Multiple Myeloma ◽

Board Of Directors ◽

Research Funding ◽

Kinase Inhibitors ◽

Mtor Inhibitors ◽

Mtor Pathway ◽

Differential Sensitivity ◽

Advisory Committees ◽

Mtor Kinase ◽

Regulatory Cascade

Abstract Abstract 2916 Inhibition of the PI3K/mTOR pathway is a promising therapeutic strategy in targeting multiple myeloma (MM) cells in the bone marrow (BM) microenvironment, which abnormally activates PI3K/mTOR signaling cascade mediating proliferation, anti-apoptosis and drug resistance. Exploring the targeting of PI3K/mTOR pathway has led to the development of different therapeutic approaches; however, mTORC1 inhibitors (i.e., temsirolimus and everolimus) have demonstrated only modest activity as single agents. In this regard, several mechanisms underlying rapamycin resistance, including mTOR/S6K1-mediated feedback loops resulting in activation of PI3K/Akt and ERK signaling, have been proposed. Importantly, recent studies have identified mTOR kinase and the mTOR-DEPTOR counter-regulatory cascade as key mediators of mTORC1 and mTORC2 multi-protein complexes, with differential sensitivity to rapamycin. Indeed, targeting DEPTOR/mTORC1/mTORC2 signaling by inhibition of mTOR kinase proved an effective strategy to overcome some of the limitations of TORC1 inhibition in MM cells, evidenced in our studies of the novel dual mTORC1 and mTORC2 selective inhibitor AZD8055. Unlike rapamycin, AZD8055 induced apoptosis and inhibited MM cell growth even when co-cultured with cytokines (i.e., IL-6, IGF1) or BMSCs, presumably through simultaneous suppression of mTORC1 and mTORC2 signaling including the rapamycin-resistant 4E-BP1 (downstream of mTORC1) and Akt as well as NDRG1 (effectors of mTORC2). We examined mRNA and protein level of DEPTOR in MM cell lines treated with AZD8055 versus rapamycin and observed no significant changes. To examine the functional significance of DEPTOR in response to mTOR inhibitors, we utilized lentiviral shRNA to knockdown DEPTOR in OPM1 MM cells. DEPTOR-knockdown cells acquired resistance to AZD8055 treatment, suggesting that DEPTOR is a key modulator of mTORC1/2 signaling. Moreover, DEPTOR knockdown triggered decrease in Akt phosphorylation (Ser473), associated with suppression of Rictor phosphorylation (Thr1135). DEPTOR co-immunoprecipitation with Rictor was also abrogated by both AZD8055 and rapamycin treatment. Taken together, our results indicate the role of DEPTOR, either alone or as an mTOR/Rictor interacting molecule, in mediating the anti-MM activity induced by mTOR kinase inhibitors in MM cells. These data therefore both provide insights into the molecular profiles that may predict sensitivity/resistance to second generation of mTOR inhibitors in MM, and may be useful to select MM patients for mTOR inhibitor therapy. Disclosures: Hideshima: Acetylon: Consultancy. Anderson:Celgene: Membership on an entity's Board of Directors or advisory committees; Millennium: Membership on an entity's Board of Directors or advisory committees; Onyx: Membership on an entity's Board of Directors or advisory committees; Merck: Membership on an entity's Board of Directors or advisory committees; Bristol Myers Squibb: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Acetylon: Membership on an entity's Board of Directors or advisory committees. Guichard:AstraZeneca, UK: Employment, Shares from AstraZeneca, UK. Raje:Celgene: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Millenium: Membership on an entity's Board of Directors or advisory committees; Astra Zeneca: Research Funding; Amgen: Membership on an entity's Board of Directors or advisory committees; Acetylon: Research Funding.

Download Full-text

Rapamycin derivatives reduce mTORC2 signaling and inhibit AKT activation in AML

Blood ◽

10.1182/blood-2006-06-030833 ◽

2006 ◽

Vol 109 (8) ◽

pp. 3509-3512 ◽

Cited By ~ 239

Author(s):

Zhihong Zeng ◽

Dos D. Sarbassov ◽

Ismael J. Samudio ◽

Karen W. L. Yee ◽

Mark F. Munsell ◽

...

Keyword(s):

Therapeutic Potential ◽

Hematologic Malignancies ◽

Mtor Inhibition ◽

Akt Activation ◽

Mtor Complex 1 ◽

Prosurvival Kinase ◽

Glut1 Mrna ◽

Made In

Abstract The mTOR complex 2 (mTORC2) containing mTOR and rictor is thought to be rapamycin insensitive and was recently shown to regulate the prosurvival kinase AKT by phosphorylation on Ser473. We investigated the molecular effects of mTOR inhibition by the rapamycin derivatives (RDs) temsirolimus (CCI-779) and everolimus (RAD001) in acute myeloid leukemia (AML) cells. Unexpectedly, RDs not only inhibited the mTOR complex 1 (mTORC1) containing mTOR and raptor with decreased p70S6K, 4EPB1 phosphorylation, and GLUT1 mRNA, but also blocked AKT activation via inhibition of mTORC2 formation. This resulted in suppression of phosphorylation of the direct AKT substrate FKHR and decreased transcription of D-cyclins in AML cells. Similar observations were made in samples from patients with hematologic malignancies who received RDs in clinical studies. Our study provides the first evidence that rapamycin derivatives inhibit AKT signaling in primary AML cells both in vitro and in vivo, and supports the therapeutic potential of mTOR inhibition strategies in leukemias.

Download Full-text