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1.
Nat Struct Mol Biol ; 28(10): 835-846, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34625748

RESUMO

Many regulatory PPP1R subunits join few catalytic PP1c subunits to mediate phosphoserine and phosphothreonine dephosphorylation in metazoans. Regulatory subunits engage the surface of PP1c, locally affecting flexible access of the phosphopeptide to the active site. However, catalytic efficiency of holophosphatases towards their phosphoprotein substrates remains unexplained. Here we present a cryo-EM structure of the tripartite PP1c-PPP1R15A-G-actin holophosphatase that terminates signaling in the mammalian integrated stress response (ISR) in the pre-dephosphorylation complex with its substrate, translation initiation factor 2α (eIF2α). G-actin, whose essential role in eIF2α dephosphorylation is supported crystallographically, biochemically and genetically, aligns the catalytic and regulatory subunits, creating a composite surface that engages the N-terminal domain of eIF2α to position the distant phosphoserine-51 at the active site. Substrate residues that mediate affinity for the holophosphatase also make critical contacts with eIF2α kinases. Thus, a convergent process of higher-order substrate recognition specifies functionally antagonistic phosphorylation and dephosphorylation in the ISR.

2.
Nat Commun ; 12(1): 5004, 2021 08 18.
Artigo em Inglês | MEDLINE | ID: mdl-34408154

RESUMO

The endoplasmic reticulum (ER) Hsp70 chaperone BiP is regulated by AMPylation, a reversible inactivating post-translational modification. Both BiP AMPylation and deAMPylation are catalysed by a single ER-localised enzyme, FICD. Here we present crystallographic and solution structures of a deAMPylation Michaelis complex formed between mammalian AMPylated BiP and FICD. The latter, via its tetratricopeptide repeat domain, binds a surface that is specific to ATP-state Hsp70 chaperones, explaining the exquisite selectivity of FICD for BiP's ATP-bound conformation both when AMPylating and deAMPylating Thr518. The eukaryotic deAMPylation mechanism thus revealed, rationalises the role of the conserved Fic domain Glu234 as a gatekeeper residue that both inhibits AMPylation and facilitates hydrolytic deAMPylation catalysed by dimeric FICD. These findings point to a monomerisation-induced increase in Glu234 flexibility as the basis of an oligomeric state-dependent switch between FICD's antagonistic activities, despite a similar mode of engagement of its two substrates - unmodified and AMPylated BiP.


Assuntos
Monofosfato de Adenosina/metabolismo , Proteínas de Choque Térmico/química , Proteínas de Choque Térmico/metabolismo , Proteínas de Membrana/química , Proteínas de Membrana/metabolismo , Nucleotidiltransferases/química , Nucleotidiltransferases/metabolismo , Monofosfato de Adenosina/química , Trifosfato de Adenosina/química , Trifosfato de Adenosina/metabolismo , Motivos de Aminoácidos , Biocatálise , Dimerização , Proteínas de Choque Térmico/genética , Humanos , Proteínas de Membrana/genética , Nucleotidiltransferases/genética , Processamento de Proteína Pós-Traducional
3.
Cell Rep ; 35(7): 109144, 2021 May 18.
Artigo em Inglês | MEDLINE | ID: mdl-34010647

RESUMO

Circulating polymers of α1-antitrypsin (α1AT) are neutrophil chemo-attractants and contribute to inflammation, yet cellular factors affecting their secretion remain obscure. We report on a genome-wide CRISPR-Cas9 screen for genes affecting trafficking of polymerogenic α1ATH334D. A CRISPR enrichment approach based on recovery of single guide RNA (sgRNA) sequences from phenotypically selected fixed cells reveals that cells with high-polymer content are enriched in sgRNAs targeting genes involved in "cargo loading into COPII-coated vesicles," where "COPII" is coat protein II, including the cargo receptors lectin mannose binding1 (LMAN1) and surfeit protein locus 4 (SURF4). LMAN1- and SURF4-disrupted cells display a secretion defect extending beyond α1AT monomers to polymers. Polymer secretion is especially dependent on SURF4 and correlates with a SURF4-α1ATH334D physical interaction and with their co-localization at the endoplasmic reticulum (ER). These findings indicate that ER cargo receptors co-ordinate progression of α1AT out of the ER and modulate the accumulation of polymeric α1AT not only by controlling the concentration of precursor monomers but also by promoting secretion of polymers.

4.
Front Neuroendocrinol ; 62: 100914, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-33845041

RESUMO

Parkinson's disease and diabetes mellitus are two chronic disorders associated with aging that are becoming increasingly prevalent worldwide. Parkinson is a multifactorial progressive condition with no available disease modifying treatments at the moment. Over the last few years there is growing interest in the relationship between diabetes (and impaired insulin signaling) and neurodegenerative diseases, as well as the possible benefit of antidiabetic treatments as neuroprotectors, even in non-diabetic patients. Insulin regulates essential functions in the brain such as neuronal survival, autophagy of toxic proteins, synaptic plasticity, neurogenesis, oxidative stress and neuroinflammation. We review the existing epidemiological, experimental and clinical evidence that supports the interplay between insulin and neurodegeneration in Parkinson's disease, as well as the role of antidiabetic treatments in this disease.

5.
BMC Cancer ; 21(1): 64, 2021 Jan 14.
Artigo em Inglês | MEDLINE | ID: mdl-33446148

RESUMO

BACKGROUND: Outcomes are poorer in metastatic colorectal cancer (mCRC) patients with BRAF V600E mutations than those without it, but the effect of these mutations on treatment response is unclear. This real-world study assessed the effects of antiangiogenic-based treatment and systemic inflammatory factors on outcomes in patients with BRAF V600-mutated mCRC. METHODS: This real-world, multicenter, retrospective, observational study included patients with BRAF V600-mutated mCRC treated in eight hospitals in Spain. The primary endpoints were overall survival (OS) and progression-free survival (PFS); overall response rate (ORR) and disease control rate (DCR) were also assessed. The effect of first- and second-line treatment type on OS, PFS, ORR, and DCR were evaluated, plus the impact of systemic inflammatory markers on these outcomes. A systemic inflammation score (SIS) of 1-3 was assigned based on one point each for platelet-lymphocyte ratio (PLR) ≥200, neutrophil-lymphocyte ratio (NLR) ≥3, and serum albumin < 3.6 g/dL. RESULTS: Of 72 patients, data from 64 were analyzed. After a median of 69.1 months, median OS was 11.9 months and median first-line PFS was 4.4 months. First-line treatment was triplet chemotherapy-antiangiogenic (12.5%), doublet chemotherapy-antiangiogenic (47.2%), doublet chemotherapy-anti-EGFR (11.1%), or doublet chemotherapy (18.1%). Although first-line treatment showed no significant effect on OS, antiangiogenic-based regimens were associated with prolonged median PFS versus non-antiangiogenic regimens. Negative predictors of survival with antiangiogenic-based treatment were NLR, serum albumin, and SIS 1-3, but not PLR. Patients with SIS 1-3 showed significantly prolonged PFS with antiangiogenic-based treatment versus non-antiangiogenic-based treatment, while those with SIS=0 showed no PFS benefit. CONCLUSIONS: Antiangiogenic-based regimens, SIS, NLR, and albumin were predictors of survival in patients with mCRC, while SIS, NLR and serum albumin may predict response to antiangiogenic-based chemotherapy. TRIAL REGISTRATION: GIT-BRAF-2017-01.


Assuntos
Inibidores da Angiogênese/uso terapêutico , Biomarcadores Tumorais/análise , Neoplasias Colorretais/patologia , Inflamação/patologia , Mutação , Proteínas Proto-Oncogênicas B-raf/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Plaquetas/patologia , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/genética , Neoplasias Colorretais/imunologia , Feminino , Seguimentos , Humanos , Linfócitos/patologia , Masculino , Pessoa de Meia-Idade , Metástase Neoplásica , Neutrófilos/patologia , Prognóstico , Estudos Retrospectivos , Espanha , Taxa de Sobrevida
6.
Mol Cell ; 81(1): 88-103.e6, 2021 01 07.
Artigo em Inglês | MEDLINE | ID: mdl-33220178

RESUMO

The small molecule ISRIB antagonizes the activation of the integrated stress response (ISR) by phosphorylated translation initiation factor 2, eIF2(αP). ISRIB and eIF2(αP) bind distinct sites in their common target, eIF2B, a guanine nucleotide exchange factor for eIF2. We have found that ISRIB-mediated acceleration of eIF2B's nucleotide exchange activity in vitro is observed preferentially in the presence of eIF2(αP) and is attenuated by mutations that desensitize eIF2B to the inhibitory effect of eIF2(αP). ISRIB's efficacy as an ISR inhibitor in cells also depends on presence of eIF2(αP). Cryoelectron microscopy (cryo-EM) showed that engagement of both eIF2B regulatory sites by two eIF2(αP) molecules remodels both the ISRIB-binding pocket and the pockets that would engage eIF2α during active nucleotide exchange, thereby discouraging both binding events. In vitro, eIF2(αP) and ISRIB reciprocally opposed each other's binding to eIF2B. These findings point to antagonistic allostery in ISRIB action on eIF2B, culminating in inhibition of the ISR.


Assuntos
Acetamidas/química , Cicloexilaminas/química , Fator de Iniciação 2B em Eucariotos/química , Fator de Iniciação 2 em Eucariotos/química , Regulação Alostérica , Animais , Sítios de Ligação , Células CHO , Cricetulus , Microscopia Crioeletrônica , Fator de Iniciação 2 em Eucariotos/genética , Fator de Iniciação 2 em Eucariotos/metabolismo , Fator de Iniciação 2B em Eucariotos/genética , Fator de Iniciação 2B em Eucariotos/metabolismo , Células HeLa , Humanos , Fosforilação
7.
J Geriatr Oncol ; 12(3): 410-415, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33357975

RESUMO

BACKGROUND: Nivolumab is an anti PD1 immunotherapy drug approved for advanced Non-Small Cell Lung Cancer (NSCLC) patients who previously received at least one prior line of treatment. Older patients are often not represented in clinical trials and drugs with acceptable safety profiles are necessary. We aim to report the efficacy and safety profile of Nivolumab in the real-world older subgroup of the Galician lung cancer group study. PATIENTS AND METHODS: We retrospectively reviewed 188 advanced NSCLC patients treated with at least one prior therapy. We collected data from patients who were ≥70 years old treated with Nivolumab in second or subsequent lines. Patient characteristics, treatment efficacy (overall survival, progression-free survival, and response rate), and safety profile were reported. RESULTS: Thirty-eight patients aged ≥70 years were included in the subgroup analysis. The median age was 74.5 years, a high percentage of patients were males (95%), most had a Performance Status of 1 (79%) and only 13% were non-smokers. The predominant histology was adenocarcinoma (53%), and 18% of patients received 2 or more lines. The median Progression-Free Survival was 7.53 months (CI 4.3-17.3, p = 0.15) and the median Overall Survival was 14.85 months (CI 10.5-20.7, p = 0.44). The objective response rate was 42%. No new adverse events were reported in comparison to a global population. CONCLUSIONS: The efficacy and safety profile of Nivolumab in advanced NSCLC patients treated with at least one prior therapy and age ≥70 years old can be overlapped to a global population. Further prospective trials are needed to define and confirm these results.


Assuntos
Antineoplásicos Imunológicos , Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Idoso , Antineoplásicos Imunológicos/efeitos adversos , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Masculino , Nivolumabe/efeitos adversos , Estudos Retrospectivos
8.
Elife ; 92020 12 09.
Artigo em Inglês | MEDLINE | ID: mdl-33295873

RESUMO

The metazoan endoplasmic reticulum (ER) serves both as a hub for maturation of secreted proteins and as an intracellular calcium storage compartment, facilitating calcium-release-dependent cellular processes. ER calcium depletion robustly activates the unfolded protein response (UPR). However, it is unclear how fluctuations in ER calcium impact organellar proteostasis. Here, we report that calcium selectively affects the dynamics of the abundant metazoan ER Hsp70 chaperone BiP, by enhancing its affinity for ADP. In the calcium-replete ER, ADP rebinding to post-ATP hydrolysis BiP-substrate complexes competes with ATP binding during both spontaneous and co-chaperone-assisted nucleotide exchange, favouring substrate retention. Conversely, in the calcium-depleted ER, relative acceleration of ADP-to-ATP exchange favours substrate release. These findings explain the rapid dissociation of certain substrates from BiP observed in the calcium-depleted ER and suggest a mechanism for tuning ER quality control and coupling UPR activity to signals that mobilise ER calcium in secretory cells.


Assuntos
Cálcio/deficiência , Retículo Endoplasmático/metabolismo , Proteínas de Choque Térmico/metabolismo , Proteostase , Difosfato de Adenosina/metabolismo , Adenosina Trifosfatases/metabolismo , Animais , Células CHO , Cálcio/metabolismo , Cricetulus , Cristalografia por Raios X , Drosophila , Escherichia coli , Citometria de Fluxo , Proteínas de Choque Térmico HSP70/metabolismo , Imunoprecipitação , Resposta a Proteínas não Dobradas
10.
Case Rep Oncol ; 13(1): 69-75, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32110222

RESUMO

In a patient who had been diagnosed in 2006 with appendiceal adenocarcinoma with peritoneal metastases after an incomplete surgery, palliative chemotherapy was administered. First-line treatment with 5-fluorouracil, leucovorin and oxaliplatin (FOLFOX) and second-line treatment including 5-fluorouracil, leucovorin and irinotecan (FOLFIRI) plus panitumumab showed inefficiency in controlling disease progression. Third-line chemotherapy combining capecitabine plus bevacizumab was started, achieving good control of the tumour growth and a minor response in the second computed tomography scan. We decided to maintain the treatment, although forced bevacizumab "breaks" were necessary due to unexpected adverse events, with the patient suffering disease progression every time bevacizumab was stopped and reaching minor response again once the antiangiogenic treatment was reintroduced. During more than 10 years after starting third-line treatment, the patient maintained good performance status and disease stability with this "up and down" management until January 2019, when a neurological adverse event during bevacizumab infusion drove us to abandon it definitely.

12.
Nature ; 578(7795): 444-448, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31875646

RESUMO

Metformin, the world's most prescribed anti-diabetic drug, is also effective in preventing type 2 diabetes in people at high risk1,2. More than 60% of this effect is attributable to the ability of metformin to lower body weight in a sustained manner3. The molecular mechanisms by which metformin lowers body weight are unknown. Here we show-in two independent randomized controlled clinical trials-that metformin increases circulating levels of the peptide hormone growth/differentiation factor 15 (GDF15), which has been shown to reduce food intake and lower body weight through a brain-stem-restricted receptor. In wild-type mice, oral metformin increased circulating GDF15, with GDF15 expression increasing predominantly in the distal intestine and the kidney. Metformin prevented weight gain in response to a high-fat diet in wild-type mice but not in mice lacking GDF15 or its receptor GDNF family receptor α-like (GFRAL). In obese mice on a high-fat diet, the effects of metformin to reduce body weight were reversed by a GFRAL-antagonist antibody. Metformin had effects on both energy intake and energy expenditure that were dependent on GDF15, but retained its ability to lower circulating glucose levels in the absence of GDF15 activity. In summary, metformin elevates circulating levels of GDF15, which is necessary to obtain its beneficial effects on energy balance and body weight, major contributors to its action as a chemopreventive agent.


Assuntos
Peso Corporal/efeitos dos fármacos , Metabolismo Energético/efeitos dos fármacos , Fator 15 de Diferenciação de Crescimento/metabolismo , Metformina/farmacologia , Administração Oral , Adulto , Idoso , Animais , Glicemia/análise , Glicemia/metabolismo , Dieta Hiperlipídica , Método Duplo-Cego , Ingestão de Energia/efeitos dos fármacos , Enterócitos/citologia , Enterócitos/efeitos dos fármacos , Feminino , Receptores de Fator Neurotrófico Derivado de Linhagem de Célula Glial/antagonistas & inibidores , Receptores de Fator Neurotrófico Derivado de Linhagem de Célula Glial/deficiência , Receptores de Fator Neurotrófico Derivado de Linhagem de Célula Glial/genética , Fator 15 de Diferenciação de Crescimento/sangue , Fator 15 de Diferenciação de Crescimento/deficiência , Fator 15 de Diferenciação de Crescimento/genética , Homeostase/efeitos dos fármacos , Humanos , Intestinos/citologia , Intestinos/efeitos dos fármacos , Masculino , Metformina/administração & dosagem , Camundongos , Camundongos Obesos , Pessoa de Meia-Idade , Perda de Peso/efeitos dos fármacos
13.
Diabetes ; 69(3): 477-483, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31882561

RESUMO

Permanent neonatal diabetes mellitus (PNDM) is caused by reduced ß-cell number or impaired ß-cell function. Understanding of the genetic basis of this disorder highlights fundamental ß-cell mechanisms. We performed trio genome sequencing for 44 patients with PNDM and their unaffected parents to identify causative de novo variants. Replication studies were performed in 188 patients diagnosed with diabetes before 2 years of age without a genetic diagnosis. EIF2B1 (encoding the eIF2B complex α subunit) was the only gene with novel de novo variants (all missense) in at least three patients. Replication studies identified two further patients with de novo EIF2B1 variants. In addition to having diabetes, four of five patients had hepatitis-like episodes in childhood. The EIF2B1 de novo mutations were found to map to the same protein surface. We propose that these variants render the eIF2B complex insensitive to eIF2 phosphorylation, which occurs under stress conditions and triggers expression of stress response genes. Failure of eIF2B to sense eIF2 phosphorylation likely leads to unregulated unfolded protein response and cell death. Our results establish de novo EIF2B1 mutations as a novel cause of permanent diabetes and liver dysfunction. These findings confirm the importance of cell stress regulation for ß-cells and highlight EIF2B1's fundamental role within this pathway.


Assuntos
Diabetes Mellitus/genética , Fator de Iniciação 2B em Eucariotos/genética , Hepatopatias/genética , Adolescente , Pré-Escolar , Simulação por Computador , Fator de Iniciação 2 em Eucariotos/metabolismo , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Lactente , Masculino , Modelos Moleculares , Mutação , Mutação de Sentido Incorreto , Recidiva , Análise de Sequência de DNA , Estresse Fisiológico
14.
Elife ; 82019 12 24.
Artigo em Inglês | MEDLINE | ID: mdl-31873072

RESUMO

Coupling of endoplasmic reticulum (ER) stress to dimerisation-dependent activation of the UPR transducer IRE1 is incompletely understood. Whilst the luminal co-chaperone ERdj4 promotes a complex between the Hsp70 BiP and IRE1's stress-sensing luminal domain (IRE1LD) that favours the latter's monomeric inactive state and loss of ERdj4 de-represses IRE1, evidence linking these cellular and in vitro observations is presently lacking. We report that enforced loading of endogenous BiP onto endogenous IRE1α repressed UPR signalling in CHO cells and deletions in the IRE1α locus that de-repressed the UPR in cells, encode flexible regions of IRE1LD that mediated BiP-induced monomerisation in vitro. Changes in the hydrogen exchange mass spectrometry profile of IRE1LD induced by ERdj4 and BiP confirmed monomerisation and were consistent with active destabilisation of the IRE1LD dimer. Together, these observations support a competition model whereby waning ER stress passively partitions ERdj4 and BiP to IRE1LD to initiate active repression of UPR signalling.


Assuntos
Estresse do Retículo Endoplasmático/genética , Endorribonucleases/química , Proteínas de Choque Térmico HSP40/química , Proteínas de Membrana/química , Chaperonas Moleculares/química , Proteínas Serina-Treonina Quinases/química , Resposta a Proteínas não Dobradas/genética , Animais , Células CHO , Cricetinae , Cricetulus , Retículo Endoplasmático/genética , Endorribonucleases/genética , Escherichia coli/genética , Proteínas de Choque Térmico HSP40/genética , Proteínas de Choque Térmico HSP70/química , Proteínas de Choque Térmico HSP70/genética , Humanos , Proteínas de Membrana/genética , Chaperonas Moleculares/genética , Chaperonas Moleculares/metabolismo , Ligação Proteica/genética , Conformação Proteica , Multimerização Proteica/genética , Proteínas Serina-Treonina Quinases/genética
15.
Elife ; 82019 11 21.
Artigo em Inglês | MEDLINE | ID: mdl-31749445

RESUMO

The eukaryotic translation initiation factor 2α (eIF2α) kinase GCN2 is activated by amino acid starvation to elicit a rectifying physiological program known as the Integrated Stress Response (ISR). A role for uncharged tRNAs as activating ligands of yeast GCN2 is supported experimentally. However, mouse GCN2 activation has recently been observed in circumstances associated with ribosome stalling with no global increase in uncharged tRNAs. We report on a mammalian CHO cell-based CRISPR-Cas9 mutagenesis screen for genes that contribute to ISR activation by amino acid starvation. Disruption of genes encoding components of the ribosome P-stalk, uL10 and P1, selectively attenuated GCN2-mediated ISR activation by amino acid starvation or interference with tRNA charging without affecting the endoplasmic reticulum unfolded protein stress-induced ISR, mediated by the related eIF2α kinase PERK. Wildtype ribosomes isolated from CHO cells, but not those with P-stalk lesions, stimulated GCN2-dependent eIF2α phosphorylation in vitro. These observations support a model whereby lack of a cognate charged tRNA exposes a latent capacity of the ribosome P-stalk to activate GCN2 in cells and help explain the emerging link between ribosome stalling and ISR activation.


Assuntos
Aminoácidos/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Ribossomos/metabolismo , Inanição/metabolismo , Animais , Células CHO , Sistemas CRISPR-Cas , Cricetulus , Retículo Endoplasmático/metabolismo , Regulação Enzimológica da Expressão Gênica , Células HeLa , Humanos , Cinética , Ligantes , Camundongos , Modelos Moleculares , Mutagênese , Fosforilação , Ligação Proteica , Conformação Proteica , Desdobramento de Proteína , Proteínas Serina-Treonina Quinases/química , Proteínas Serina-Treonina Quinases/genética , RNA de Transferência/metabolismo , Ribossomos/química , Transdução de Sinais , Transcriptoma , eIF-2 Quinase/genética , eIF-2 Quinase/metabolismo
16.
EMBO J ; 38(21): e102177, 2019 10 04.
Artigo em Inglês | MEDLINE | ID: mdl-31531998

RESUMO

AMPylation is an inactivating modification that alters the activity of the major endoplasmic reticulum (ER) chaperone BiP to match the burden of unfolded proteins. A single ER-localised Fic protein, FICD (HYPE), catalyses both AMPylation and deAMPylation of BiP. However, the basis for the switch in FICD's activity is unknown. We report on the transition of FICD from a dimeric enzyme, that deAMPylates BiP, to a monomer with potent AMPylation activity. Mutations in the dimer interface, or of residues along an inhibitory pathway linking the dimer interface to the enzyme's active site, favour BiP AMPylation in vitro and in cells. Mechanistically, monomerisation relieves a repressive effect allosterically propagated from the dimer interface to the inhibitory Glu234, thereby permitting AMPylation-competent binding of MgATP. Moreover, a reciprocal signal, propagated from the nucleotide-binding site, provides a mechanism for coupling the oligomeric state and enzymatic activity of FICD to the energy status of the ER.


Assuntos
Retículo Endoplasmático/metabolismo , Proteínas de Choque Térmico/química , Proteínas de Choque Térmico/metabolismo , Proteínas de Membrana/química , Proteínas de Membrana/metabolismo , Nucleotidiltransferases/química , Nucleotidiltransferases/metabolismo , Multimerização Proteica , Processamento de Proteína Pós-Traducional , Células HEK293 , Humanos , Conformação Proteica
17.
Front Oncol ; 9: 819, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31552167

RESUMO

In a patient who had been diagnosed of located squamous cell lung carcinoma, pneumonectomy, and adjuvant chemotherapy were performed. Brain recurrence and subsequent lung metastatic disease were uncontrolled by neurosurgery, holocranial radiotherapy, and first-line chemotherapy. In August 2015, appearance of leptomeningeal carcinomatosis triggered severe clinical deterioration and threatened the patient's life. Anti-PD1 immune checkpoint inhibitor Nivolumab was initiated in an attempt to stop tumor growth, achieving a spectacular brain and pulmonary complete response and clinical improvement, without serious adverse effects. High expression PD-L1 level (100%) was found in the pathological tissue sample. Nivolumab was maintained for more than 2 years and stopped in December 2017 after 28 months of treatment, with no disease evidence. More than 3 years after its onset, the patient maintains an outstanding PS with complete tumor response and no evidence of disease in last surveillance CT scan and brain MRI.

18.
Nat Commun ; 10(1): 541, 2019 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-30710085

RESUMO

Despite its known role as a secreted neuroprotectant, much of the mesencephalic astrocyte-derived neurotrophic factor (MANF) is retained in the endoplasmic reticulum (ER) of producer cells. There, by unknown mechanisms, MANF plays a role in protein folding homeostasis in complex with the ER-localized Hsp70 chaperone BiP. Here we report that the SAF-A/B, Acinus, and PIAS (SAP) domain of MANF selectively associates with the nucleotide binding domain (NBD) of ADP-bound BiP. In crystal structures the SAP domain engages the cleft between NBD subdomains Ia and IIa, stabilizing the ADP-bound conformation and clashing with the interdomain linker that occupies this site in ATP-bound BiP. MANF inhibits both ADP release from BiP and ATP binding to BiP, and thereby client release. Cells lacking MANF have fewer ER stress-induced BiP-containing high molecular weight complexes. These findings suggest that MANF contributes to protein folding homeostasis as a nucleotide exchange inhibitor that stabilizes certain BiP-client complexes.


Assuntos
Retículo Endoplasmático/metabolismo , Proteínas de Choque Térmico/metabolismo , Fatores de Crescimento Neural/metabolismo , Nucleotídeos/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Células CHO , Chlorocebus aethiops , Cricetulus , Cristalografia por Raios X , Células HEK293 , Proteínas de Choque Térmico/química , Humanos , Modelos Biológicos , Fatores de Crescimento Neural/química , Ligação Proteica , Domínios Proteicos , Eletricidade Estática , Resposta a Proteínas não Dobradas
19.
Cell Metab ; 29(3): 707-718.e8, 2019 03 05.
Artigo em Inglês | MEDLINE | ID: mdl-30639358

RESUMO

GDF15 is an established biomarker of cellular stress. The fact that it signals via a specific hindbrain receptor, GFRAL, and that mice lacking GDF15 manifest diet-induced obesity suggest that GDF15 may play a physiological role in energy balance. We performed experiments in humans, mice, and cells to determine if and how nutritional perturbations modify GDF15 expression. Circulating GDF15 levels manifest very modest changes in response to moderate caloric surpluses or deficits in mice or humans, differentiating it from classical intestinally derived satiety hormones and leptin. However, GDF15 levels do increase following sustained high-fat feeding or dietary amino acid imbalance in mice. We demonstrate that GDF15 expression is regulated by the integrated stress response and is induced in selected tissues in mice in these settings. Finally, we show that pharmacological GDF15 administration to mice can trigger conditioned taste aversion, suggesting that GDF15 may induce an aversive response to nutritional stress.


Assuntos
Ingestão de Energia/fisiologia , Fator 15 de Diferenciação de Crescimento/metabolismo , Adulto , Animais , Linhagem Celular , Dieta Hiperlipídica/métodos , Fator 15 de Diferenciação de Crescimento/farmacologia , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Adulto Jovem
20.
Artigo em Inglês | MEDLINE | ID: mdl-30396883

RESUMO

The physiological consequences of the unfolded protein response (UPR) are mediated by changes in gene expression. Underlying them are rapid processes involving preexisting components. We review recent insights gained into the regulation of the endoplasmic reticulum (ER) Hsp70 chaperone BiP, whose incorporation into inactive oligomers and reversible AMPylation and de-AMPylation present a first line of response to fluctuating levels of unfolded proteins. BiP activity is tied to the regulation of the UPR transducers by a recently discovered cycle of ER-localized, J protein-mediated formation of a repressive IRE1-BiP complex, whose working we contrast to an alternative model for UPR regulation that relies on direct recognition of unfolded proteins. We conclude with a discussion of mechanisms that repress messenger RNA (mRNA) translation to limit the flux of newly synthesized proteins into the ER, a rapid adaptation that does not rely on new macromolecule biosynthesis.


Assuntos
Retículo Endoplasmático/metabolismo , Resposta a Proteínas não Dobradas , Proteínas de Choque Térmico/metabolismo , Biossíntese de Proteínas , Processamento de Proteína Pós-Traducional
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