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1.
Cell ; 158(5): 1094-1109, 2014 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-25171410

RESUMO

It is increasingly appreciated that oncogenic transformation alters cellular metabolism to facilitate cell proliferation, but less is known about the metabolic changes that promote cancer cell aggressiveness. Here, we analyzed metabolic gene expression in cancer cell lines and found that a set of high-grade carcinoma lines expressing mesenchymal markers share a unique 44 gene signature, designated the "mesenchymal metabolic signature" (MMS). A FACS-based shRNA screen identified several MMS genes as essential for the epithelial-mesenchymal transition (EMT), but not for cell proliferation. Dihydropyrimidine dehydrogenase (DPYD), a pyrimidine-degrading enzyme, was highly expressed upon EMT induction and was necessary for cells to acquire mesenchymal characteristics in vitro and for tumorigenic cells to extravasate into the mouse lung. This role of DPYD was mediated through its catalytic activity and enzymatic products, the dihydropyrimidines. Thus, we identify metabolic processes essential for the EMT, a program associated with the acquisition of metastatic and aggressive cancer cell traits.


Assuntos
Transição Epitelial-Mesenquimal , Pirimidinas/metabolismo , Animais , Carcinoma/metabolismo , Linhagem Celular Tumoral , Di-Hidrouracila Desidrogenase (NADP)/genética , Citometria de Fluxo , Perfilação da Expressão Gênica , Humanos , Mesoderma/citologia , Mesoderma/metabolismo , Camundongos , RNA Interferente Pequeno/metabolismo
2.
Nature ; 517(7534): 302-10, 2015 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-25592535

RESUMO

The ability to sense and respond to fluctuations in environmental nutrient levels is a requisite for life. Nutrient scarcity is a selective pressure that has shaped the evolution of most cellular processes. Different pathways that detect intracellular and extracellular levels of sugars, amino acids, lipids and surrogate metabolites are integrated and coordinated at the organismal level through hormonal signals. During food abundance, nutrient-sensing pathways engage anabolism and storage, whereas scarcity triggers homeostatic mechanisms, such as the mobilization of internal stores through autophagy. Nutrient-sensing pathways are commonly deregulated in human metabolic diseases.


Assuntos
Aminoácidos/metabolismo , Glucose/metabolismo , Metabolismo dos Lipídeos , Animais , Autofagia , Homeostase , Humanos , Doenças Metabólicas/metabolismo
3.
Mol Cell ; 45(6): 719-30, 2012 Mar 30.
Artigo em Inglês | MEDLINE | ID: mdl-22342344

RESUMO

The IκB kinase (IKK) pathway is an essential mediator of inflammatory, oncogenic, and cell stress pathways. Recently IKK was shown to be essential for autophagy induction in mammalian cells independent of its ability to regulate NF-κB, but the mechanism by which this occurs is unclear. Here we demonstrate that the p85 regulatory subunit of PI3K is an IKK substrate, phosphorylated at S690 in vitro and in vivo in response to cellular starvation. Cells expressing p85 S690A or inhibited for IKK activity exhibit increased Akt activity following cell starvation, demonstrating that p85 phosphorylation is required for starvation-induced PI3K feedback inhibition. S690 is in a conserved region of the p85 cSH2 domain, and IKK-mediated phosphorylation of this site results in decreased affinity for tyrosine-phosphorylated proteins and decreased PI3K membrane localization. Finally, leucine deprivation is shown to be necessary and sufficient for starvation-induced, IKK-mediated p85 phosphorylation and PI3K feedback inhibition.


Assuntos
Quinase I-kappa B/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Inanição/metabolismo , Sequência de Aminoácidos , Aminoácidos/metabolismo , Animais , Linhagem Celular , Classe Ia de Fosfatidilinositol 3-Quinase/metabolismo , Sequência Conservada , Retroalimentação Fisiológica , Fibroblastos/metabolismo , Humanos , Quinase I-kappa B/genética , Leucina/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Fosforilação , Fosfotirosina/metabolismo , Domínios de Homologia de src
4.
Mol Cancer Ther ; : OF1-OF12, 2024 Jun 10.
Artigo em Inglês | MEDLINE | ID: mdl-38853438

RESUMO

Advances in linker payload technology and target selection have been at the forefront of recent improvements in antibody-drug conjugate (ADC) design, leading to several approvals over the last decade. In contrast, the potential of novel ADC technologies to enhance payload delivery to tumors is relatively underexplored. We demonstrate that incorporation of pH-dependent binding in the antibody component of a c-mesenchymal-epithelial transition (MET)-targeting ADC (MYTX-011) can overcome the requirement for high c-MET expression on tumors, an innovation that has the potential to benefit a broader population of patients with lower c-MET levels. MYTX-011 drove fourfold higher net internalization than a non-pH-engineered parent ADC in non-small cell lung cancer (NSCLC) cells and showed increased cytotoxicity against a panel of cell lines from various solid tumors. A single dose of MYTX-011 showed at least threefold higher efficacy than a benchmark ADC in mouse xenograft models of NSCLC ranging from low to high c-MET expression. Moreover, MYTX-011 showed improved pharmacokinetics over parent and benchmark ADCs. In a repeat dose toxicology study, MYTX-011 exhibited a toxicity profile similar to other monomethyl auristatin E-based ADCs. These results highlight the potential of MYTX-011 for treating a broader range of patients with NSCLC with c-MET expression than other c-MET-targeting ADCs. A first-in-human study is ongoing to determine the safety, tolerability, and preliminary efficacy of MYTX-011 in patients with NSCLC (NCT05652868).

5.
Mol Cancer Ther ; 2024 Apr 30.
Artigo em Inglês | MEDLINE | ID: mdl-38684230

RESUMO

Advances in linker payload technology and target selection have been at the forefront of recent improvements in antibody-drug conjugate (ADC) design, leading to several approvals over the last decade. In contrast, the potential of novel ADC technologies to enhance payload delivery to tumors is relatively underexplored. We demonstrate that incorporation of pH-dependent binding in the antibody component of a cMET targeting ADC (MYTX-011) can overcome the requirement for high cMET expression on tumors, an innovation that has the potential to benefit a broader population of patients with lower cMET levels. MYTX-011 drove four-fold higher net internalization than a non-pH engineered parent ADC in non-small cell lung cancer (NSCLC) cells and showed increased cytotoxicity against a panel of cell lines from various solid tumors. A single dose of MYTX-011 showed at least three-fold higher efficacy than a benchmark ADC in mouse xenograft models of NSCLC ranging from low to high cMET expression. Moreover, MYTX-011 showed improved pharmacokinetics over parent and benchmark ADCs. In a repeat dose toxicology study, MYTX-011 exhibited a toxicity profile similar to other MMAE-based ADCs. These results highlight the potential of MYTX-011 for treating a broader range of NSCLC patients with cMET expression than other cMET targeting ADCs. A first in human study is ongoing to determine the safety, tolerability, and preliminary efficacy of MYTX-011 in patients with NSCLC (NCT05652868).

6.
Science ; 377(6601): 47-56, 2022 07.
Artigo em Inglês | MEDLINE | ID: mdl-35771919

RESUMO

The mechanistic target of rapamycin complex 1 (mTORC1) kinase controls growth in response to nutrients, including the amino acid leucine. In cultured cells, mTORC1 senses leucine through the leucine-binding Sestrin proteins, but the physiological functions and distribution of Sestrin-mediated leucine sensing in mammals are unknown. We find that mice lacking Sestrin1 and Sestrin2 cannot inhibit mTORC1 upon dietary leucine deprivation and suffer a rapid loss of white adipose tissue (WAT) and muscle. The WAT loss is driven by aberrant mTORC1 activity and fibroblast growth factor 21 (FGF21) production in the liver. Sestrin expression in the liver lobule is zonated, accounting for zone-specific regulation of mTORC1 activity and FGF21 induction by leucine. These results establish the mammalian Sestrins as physiological leucine sensors and reveal a spatial organization to nutrient sensing by the mTORC1 pathway.


Assuntos
Dieta , Leucina , Fígado , Alvo Mecanístico do Complexo 1 de Rapamicina , Sestrinas , Tecido Adiposo Branco/enzimologia , Animais , Leucina/metabolismo , Fígado/metabolismo , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Camundongos , Sestrinas/metabolismo , Transdução de Sinais
7.
Nat Commun ; 12(1): 5282, 2021 09 06.
Artigo em Inglês | MEDLINE | ID: mdl-34489418

RESUMO

Homeostasis is one of the fundamental concepts in physiology. Despite remarkable progress in our molecular understanding of amino acid transport, metabolism and signaling, it remains unclear by what mechanisms cytosolic amino acid concentrations are maintained. We propose that amino acid transporters are the primary determinants of intracellular amino acid levels. We show that a cell's endowment with amino acid transporters can be deconvoluted experimentally and used this data to computationally simulate amino acid translocation across the plasma membrane. Transport simulation generates cytosolic amino acid concentrations that are close to those observed in vitro. Perturbations of the system are replicated in silico and can be applied to systems where only transcriptomic data are available. This work explains amino acid homeostasis at the systems-level, through a combination of secondary active transporters, functionally acting as loaders, harmonizers and controller transporters to generate a stable equilibrium of all amino acid concentrations.


Assuntos
Sistemas de Transporte de Aminoácidos/metabolismo , Aminoácidos/metabolismo , Homeostase/genética , Modelos Estatísticos , Neuroglia/metabolismo , Células A549 , Sistemas de Transporte de Aminoácidos/química , Sistemas de Transporte de Aminoácidos/classificação , Sistemas de Transporte de Aminoácidos/genética , Animais , Transporte Biológico , Linhagem Celular Tumoral , Membrana Celular/metabolismo , Simulação por Computador , Expressão Gênica , Humanos , Cinética , Metabolômica/métodos , Neuroglia/citologia , Oócitos/citologia , Oócitos/metabolismo , Xenopus laevis
8.
Clin Transl Gastroenterol ; 11(8): e00222, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32955200

RESUMO

INTRODUCTION: AXA1665 is a novel investigational amino acid (AA) composition specifically designed to impact AA imbalance, ammoniagenesis, and dysregulated anabolic activity associated with cirrhosis. METHODS: This 2-part study examined AXA1665 effects on safety, tolerability, and hepatic/muscle physiology in subjects with Child-Pugh A and B cirrhosis. Part 1 established plasma ammonia and AA concentration baselines with a standardized protein supplement. Part 2 included two 15-day domiciled periods separated by a 14-day washout. In period 1, subjects were randomly distributed to 2 groups: AXA1665 14.7 g t.i.d. (group 1) or control t.i.d. (group 2). In period 2, subjects from group 1 crossed over to control and those in group 2 crossed over to AXA1665 4.9 g t.i.d. All subjects were maintained on standard of care (standardized meals; 30-minute daily, supervised, mandatory physical activity; and daily late-evening snack). RESULTS: In parts 1 and 2, 23 and 17 participants were enrolled, respectively. Dose-dependent increases were observed in plasma concentrations of AXA1665-constituent AAs. Fasted branched-chain AA-to-aromatic AA and valine-to-phenylalanine ratios were both increased (AXA1665 14.7 g t.i.d. control-adjusted change: 44.3% ± 2.7% and 47.2% ± 3.9%, respectively; P < 0.0001). Despite provision of additional nitrogen, mean fasted plasma ammonia concentration at day 15 numerically decreased (-21.1% in AXA1665 14.7 g t.i.d. vs -3.8% in control; P > 0.05). AXA1665 14.7 g t.i.d. produced a leaner body composition and significantly decreased Liver Frailty Index at day 15 vs control (-0.70 ± 0.15 vs -0.14 ± 0.17; P < 0.05). AXA1665 was safe and well tolerated. DISCUSSION: AXA1665 has potential to mitigate core metabolic derangements associated with cirrhosis.


Assuntos
Aminoácidos de Cadeia Ramificada/administração & dosagem , Drogas em Investigação/administração & dosagem , Cirrose Hepática/tratamento farmacológico , Adulto , Idoso , Aminoácidos de Cadeia Ramificada/efeitos adversos , Aminoácidos de Cadeia Ramificada/sangue , Aminoácidos de Cadeia Ramificada/metabolismo , Amônia/sangue , Amônia/metabolismo , Estudos Cross-Over , Drogas em Investigação/efeitos adversos , Feminino , Humanos , Fígado/metabolismo , Cirrose Hepática/sangue , Cirrose Hepática/diagnóstico , Cirrose Hepática/metabolismo , Masculino , Pessoa de Meia-Idade , Índice de Gravidade de Doença , Soluções , Resultado do Tratamento
9.
Cell Rep ; 12(9): 1445-55, 2015 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-26299971

RESUMO

mTORC1 controls key processes that regulate cell growth, including mRNA translation, ribosome biogenesis, and autophagy. Environmental amino acids activate mTORC1 by promoting its recruitment to the cytosolic surface of the lysosome, where its kinase is activated downstream of growth factor signaling. mTORC1 is brought to the lysosome by the Rag GTPases, which are tethered to the lysosomal membrane by Ragulator, a lysosome-bound scaffold. Here, we identify c17orf59 as a Ragulator-interacting protein that regulates mTORC1 activity through its interaction with Ragulator at the lysosome. The binding of c17orf59 to Ragulator prevents Ragulator interaction with the Rag GTPases, both in cells and in vitro, and decreases Rag GTPase lysosomal localization. Disruption of the Rag-Ragulator interaction by c17orf59 impairs mTORC1 activation by amino acids by preventing mTOR from reaching the lysosome. By disrupting the Rag-Ragulator interaction to inhibit mTORC1, c17orf59 expression may represent another mechanism to modulate nutrient sensing by mTORC1.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Lisossomos/metabolismo , Proteínas Monoméricas de Ligação ao GTP/metabolismo , Complexos Multiproteicos/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Células HEK293 , Células HeLa , Humanos , Alvo Mecanístico do Complexo 1 de Rapamicina , Proteínas Monoméricas de Ligação ao GTP/genética , Ligação Proteica
10.
Mol Cell Biol ; 28(16): 5061-70, 2008 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-18541671

RESUMO

Glutamate is a critical neurotransmitter of the central nervous system (CNS) and also an important regulator of cell survival and proliferation. The binding of glutamate to metabotropic glutamate receptors induces signal transduction cascades that lead to gene-specific transcription. The transcription factor NF-kappaB, which regulates cell proliferation and survival, is activated by glutamate; however, the glutamate receptor-induced signaling pathways that lead to this activation are not clearly defined. Here we investigate the glutamate-induced activation of NF-kappaB in glial cells of the CNS, including primary astrocytes. We show that glutamate induces phosphorylation, nuclear accumulation, DNA binding, and transcriptional activation function of glial p65. The glutamate-induced activation of NF-kappaB requires calcium-dependent IkappaB kinase alpha (IKKalpha) and IKKbeta activation and induces p65-IkappaBalpha dissociation in the absence of IkappaBalpha phosphorylation or degradation. Moreover, glutamate-induced IKK preferentially targets the phosphorylation of p65 but not IkappaBalpha. Finally, we show that the ability of glutamate to activate NF-kappaB requires cross-coupled signaling with the epidermal growth factor receptor. Our results provide insight into a glutamate-induced regulatory pathway distinct from that described for cytokine-induced NF-kappaB activation and have important implications with regard to both normal glial cell physiology and pathogenesis.


Assuntos
Receptores ErbB/metabolismo , Receptores de Glutamato/metabolismo , Transdução de Sinais , Fator de Transcrição RelA/metabolismo , Animais , Sinalização do Cálcio/efeitos dos fármacos , Linhagem Celular , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Proliferação de Células/efeitos dos fármacos , Ácido Glutâmico/farmacologia , Glicina/análogos & derivados , Glicina/farmacologia , Humanos , Quinase I-kappa B/metabolismo , Proteínas I-kappa B/metabolismo , Espaço Intracelular/efeitos dos fármacos , Espaço Intracelular/metabolismo , Camundongos , Neuroglia/citologia , Neuroglia/efeitos dos fármacos , Neuroglia/metabolismo , Fenilacetatos/farmacologia , Fosforilação/efeitos dos fármacos , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Receptor de Glutamato Metabotrópico 5 , Receptores de Glutamato Metabotrópico/metabolismo , Transdução de Sinais/efeitos dos fármacos , Ativação Transcricional/efeitos dos fármacos
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