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1.
BMC Biol ; 18(1): 143, 2020 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-33059680

RESUMO

BACKGROUND: ADP-ribosylation is a ubiquitous post-translational modification that involves both mono- and poly-ADP-ribosylation. ARTD10, also known as PARP10, mediates mono-ADP-ribosylation (MARylation) of substrate proteins. A previous screen identified protein kinase C delta (PKCδ) as a potential ARTD10 substrate, among several other kinases. The voltage-gated K+ channel Kv1.1 constitutes one of the dominant Kv channels in neurons of the central nervous system and the inactivation properties of Kv1.1 are modulated by PKC. In this study, we addressed the role of ARTD10-PKCδ as a regulator of Kv1.1. RESULTS: We found that ARTD10 inhibited PKCδ, which increased Kv1.1 current amplitude and the proportion of the inactivating current component in HeLa cells, indicating that ARTD10 regulates Kv1.1 in living cells. An inhibitor of ARTD10, OUL35, significantly decreased peak amplitude together with the proportion of the inactivating current component of Kv1.1-containing channels in primary hippocampal neurons, demonstrating that the ARTD10-PKCδ signaling cascade regulates native Kv1.1. Moreover, we show that the pharmacological blockade of ARTD10 increases excitability of hippocampal neurons. CONCLUSIONS: Our results, for the first time, suggest that MARylation by ARTD10 controls neuronal excitability.


Assuntos
Canal de Potássio Kv1.1/genética , Poli(ADP-Ribose) Polimerases/genética , Proteína Quinase C-delta/genética , Processamento de Proteína Pós-Traducional , Proteínas Proto-Oncogênicas/genética , Transdução de Sinais , Animais , Células HEK293 , Células HeLa , Humanos , Canal de Potássio Kv1.1/metabolismo , Camundongos , Poli(ADP-Ribose) Polimerases/metabolismo , Proteína Quinase C-delta/metabolismo , Proteínas Proto-Oncogênicas/metabolismo
2.
Hepatol Commun ; 8(4)2024 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-38517202

RESUMO

BACKGROUND: Bile salts of hepatic and microbial origin mediate interorgan cross talk in the gut-liver axis. Here, we assessed whether the newly discovered class of microbial bile salt conjugates (MBSCs) activate the main host bile salt receptors (Takeda G protein-coupled receptor 5 [TGR5] and farnesoid X receptor [FXR]) and enter the human systemic and enterohepatic circulation. METHODS: N-amidates of (chenodeoxy) cholic acid and leucine, tyrosine, and phenylalanine were synthesized. Receptor activation was studied in cell-free and cell-based assays. MBSCs were quantified in mesenteric and portal blood and bile of patients undergoing pancreatic surgery. RESULTS: MBSCs were activating ligands of TGR5 as evidenced by recruitment of Gsα protein, activation of a cAMP-driven reporter, and diminution of lipopolysaccharide-induced cytokine release from macrophages. Intestine-enriched and liver-enriched FXR isoforms were both activated by MBSCs, provided that a bile salt importer was present. The affinity of MBSCs for TGR5 and FXR was not superior to host-derived bile salt conjugates. Individual MBSCs were generally not detected (ie, < 2.5 nmol/L) in human mesenteric or portal blood, but Leu-variant and Phe-variant were readily measurable in bile, where MBSCs comprised up to 213 ppm of biliary bile salts. CONCLUSIONS: MBSCs activate the cell surface receptor TGR5 and the transcription factor FXR and are substrates for intestinal (apical sodium-dependent bile acid transporter) and hepatic (Na+ taurocholate co-transporting protein) transporters. Their entry into the human circulation is, however, nonsubstantial. Given low systemic levels and a surplus of other equipotent bile salt species, the studied MBSCs are unlikely to have an impact on enterohepatic TGR5/FXR signaling in humans. The origin and function of biliary MBSCs remain to be determined.


Assuntos
Ácidos e Sais Biliares , Receptores Citoplasmáticos e Nucleares , Receptores Acoplados a Proteínas G , Humanos , Bile/química , Ácidos e Sais Biliares/farmacologia , Ácidos e Sais Biliares/metabolismo , Fígado/metabolismo , Receptores Citoplasmáticos e Nucleares/metabolismo , Fatores de Transcrição , Receptores Acoplados a Proteínas G/metabolismo
3.
Nature ; 449(7164): 898-901, 2007 Oct 18.
Artigo em Inglês | MEDLINE | ID: mdl-17882164

RESUMO

The short-chain hydrocarbons ethane, propane and butane are constituents of natural gas. They are usually assumed to be of thermochemical origin, but biological formation of ethane and propane has been also observed. Microbial utilization of short-chain hydrocarbons has been shown in some aerobic species but not in anaerobic species of bacteria. On the other hand, anaerobic utilization of short-chain hydrocarbons would in principle be expected because various anaerobic bacteria grow with higher homologues (> or =C(6)). Indeed, chemical analyses of hydrocarbon-rich habitats with limited or no access of oxygen indicated in situ biodegradation of short-chain hydrocarbons. Here we report the enrichment of sulphate-reducing bacteria (SRB) with such capacity from marine hydrocarbon seep areas. Propane or n-butane as the sole growth substrate led to sediment-free sulphate-reducing enrichment cultures growing at 12, 28 or 60 degrees C. With ethane, a slower enrichment with residual sediment was obtained at 12 degrees C. Isolation experiments resulted in a mesophilic pure culture (strain BuS5) that used only propane and n-butane (methane, isobutane, alcohols or carboxylic acids did not support growth). Complete hydrocarbon oxidation to CO2 and the preferential oxidation of 12C-enriched alkanes were observed with strain BuS5 and other cultures. Metabolites of propane included iso- and n-propylsuccinate, indicating a subterminal as well as an unprecedented terminal alkane activation with involvement of fumarate. According to 16S ribosomal RNA analyses, strain BuS5 affiliates with Desulfosarcina/Desulfococcus, a cluster of widespread marine SRB. An enrichment culture with propane growing at 60 degrees C was dominated by Desulfotomaculum-like SRB. Our results suggest that diverse SRB are able to thrive in seep areas and gas reservoirs on propane and butane, thus altering the gas composition and contributing to sulphide production.


Assuntos
Bactérias Anaeróbias/metabolismo , Hidrocarbonetos/química , Hidrocarbonetos/metabolismo , Sulfatos/metabolismo , Bactérias Redutoras de Enxofre/metabolismo , Anaerobiose , Bactérias Anaeróbias/classificação , Bactérias Anaeróbias/genética , Butanos/metabolismo , Etano/metabolismo , Cinética , Dados de Sequência Molecular , Oceanos e Mares , Oxirredução , Filogenia , Propano/metabolismo , RNA Ribossômico 16S , Bactérias Redutoras de Enxofre/classificação , Bactérias Redutoras de Enxofre/genética
5.
Metabolites ; 12(2)2022 Feb 13.
Artigo em Inglês | MEDLINE | ID: mdl-35208250

RESUMO

Biotransformation of host bile salts by gut microbes results in generation of secondary bile salt species that have biological and physicochemical properties that are distinct from the parent compounds. There is increased awareness that a bile salt-gut microbiome axis modulates various processes in the host, including innate and adaptive immunity, by interaction of microbial bile salt metabolites with host receptors. Omics and targeted approaches have vastly expanded the number and repertoire of secondary bile salt species. A new class of microbial bile salt metabolites was reported in 2020 and comprises bile salts that are conjugated by microbial enzymes. Amino acids other than those employed by host enzymes (glycine and taurine) are used as substrates in the formation of these microbial bile salt conjugates (MBSCs). Leucocholic acid, phenylalanocholic acid and tyrosocholic acid were the first MBSCs identified in mice and humans. The number of distinct MBSCs is now approaching 50, with variation both at the level of bile salt and amino acid employed for conjugation. Evidence is emerging that MBSC generation is a common feature of human gut bacteria, and initial links with disease states have been reported. In this review, we discuss this intriguing new class of secondary bile salts, with yet enigmatic function.

6.
Cancers (Basel) ; 14(1)2022 Jan 04.
Artigo em Inglês | MEDLINE | ID: mdl-35008392

RESUMO

PARP1 inhibitors (PARPi) are currently approved for BRCAmut metastatic breast cancer, but they have shown limited response in triple negative breast cancer (TNBC) patients. Combination of an Auger emitter with PARPis enables PARP inhibition and DNA strand break induction simultaneously. This will enhance cytotoxicity and additionally allow a theranostic approach. This study presents the radiosynthesis of the Auger emitter [125I] coupled olaparib derivative: [125I]-PARPi-01, and its therapeutic evaluation in a panel of TNBC cell lines. Specificity was tested by a blocking assay. DNA strand break induction was analysed by γH2AX immunofluorescence staining. Cell cycle analysis and apoptosis assays were studied using flow cytometry in TNBC cell lines (BRCAwt/mut). Anchorage independent growth potential was evaluated using soft agar assay. [125I]-PARPi-01 showed PARP1-specificity and higher cytotoxicity than olaparib in TNBC cell lines irrespective of BRCA their status. Cell lines harbouring DNA repair deficiency showed response to [125I]-PARPi-01 monotherapy. Combined treatment with Dox-NP further enhanced therapeutic efficiency in metastatic resistant BRCAwt cell lines. The clonogenic survival was significantly reduced after treatment with [125I]-PARPi-01 in all TNBC lines investigated. Therapeutic efficacy was further enhanced after combined treatment with chemotherapeutics. [125I]-PARPi-01 is a promising radiotherapeutic agent for low radiation dosages, and mono/combined therapies of TNBC.

7.
EJNMMI Res ; 12(1): 60, 2022 Sep 14.
Artigo em Inglês | MEDLINE | ID: mdl-36104637

RESUMO

BACKGROUND: Triple-negative breast cancer (TNBC) lacks biomarkers for targeted therapy. Auger emitters display the best therapeutic effect, if delivered directly into the nucleus proximal to DNA. The nuclear protein Poly (ADP-ribose)-Polymerase 1 (PARP1) is a suitable target against which few inhibitors (PARPi) are clinically approved for treatment of breast cancer with germline BRCA mutation (BRCAmut). In this study, a theranostic approach was investigated in a TNBC xenografted mouse model by radiolabelling a close derivative of a PARPi Olaparib (termed PARPi-01) with the Auger emitters 123/125I. METHODS: TNBC cell line MDA-MB-231 was subcutaneously implanted in female NOD/SCID mice. At a tumour size of ~ 500mm3, [123I]PARPi-01 was administered intravenously, and SPECT/CT images were obtained at 4 h or 24 h post injection (p.i). A therapy study was performed with [125I]PARPi-01 in 4 doses (10 MBq/dose, 10 days apart). Tumour growth was monitored by CT scans longitudinally once per week. Upon reaching study endpoint, tissues were harvested and stained with TUNEL assay for detection of apoptosis induction. RESULTS: SPECT/CT images showed rapid hepatobiliary tracer clearance at 4 h post injection (p.i.). Retention in thyroid at 24 h p.i. suggested tracer deiodination in vivo. The tumour and liver uptake were 0.2%ID/g and 2.5%ID/g, respectively. The tumour: blood ratio was 1.3. Endogenous therapy induced a significant delay in tumour growth (doubling time increased from 8.3 to 14.2 days), but no significant survival advantage. Significantly higher apoptosis ratio was observed in [125I]PARPi-01 treated tumour tissues. No radiotoxicity was detected in the liver and thyroid. CONCLUSION: Considering the radio-cytotoxic effect in the tumour tissue and a delay on tumour doubling time, [125I]PARPi-01 presents a potential radiotherapeutics for treatment of TNBC. Improvements to overcome the suboptimal pharmacokinetics are necessary for its potential clinical application.

8.
ChemistryOpen ; 10(10): 939-948, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34145784

RESUMO

Intracellular ADP-ribosyltransferases catalyze mono- and poly-ADP-ribosylation and affect a broad range of biological processes. The mono-ADP-ribosyltransferase PARP10 is involved in signaling and DNA repair. Previous studies identified OUL35 as a selective, cell permeable inhibitor of PARP10. We have further explored the chemical space of OUL35 by synthesizing and investigating structurally related analogs. Key synthetic steps were metal-catalyzed cross-couplings and functional group modifications. We identified 4-(4-cyanophenoxy)benzamide and 3-(4-carbamoylphenoxy)benzamide as PARP10 inhibitors with distinct selectivities. Both compounds were cell permeable and interfered with PARP10 toxicity. Moreover, both revealed some inhibition of PARP2 but not PARP1, unlike clinically used PARP inhibitors, which typically inhibit both enzymes. Using crystallography and molecular modeling the binding of the compounds to different ADP-ribosyltransferases was explored regarding selectivity. Together, these studies define additional compounds that interfere with PARP10 function and thus expand our repertoire of inhibitors to further optimize selectivity and potency.


Assuntos
ADP Ribose Transferases , Inibidores de Poli(ADP-Ribose) Polimerases , ADP Ribose Transferases/metabolismo , Modelos Moleculares , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Transdução de Sinais
10.
Bioorg Med Chem Lett ; 12(14): 1795-8, 2002 Jul 22.
Artigo em Inglês | MEDLINE | ID: mdl-12086819

RESUMO

A new chiral MeO-PEG-supported ferrocenyl oxazoline was synthesized and successfully employed in the enantioselective phenyl transfer to aldehydes. The products were obtained in high yields and with excellent enantioselectivities (up to 97% ee). Furthermore, the recovery of the ferrocene was facile, and catalyst efficiency was maintained in subsequent reactions.


Assuntos
Benzaldeídos/química , Compostos Benzidrílicos/química , Compostos Ferrosos/síntese química , Oxazolona/análogos & derivados , Oxazolona/síntese química , Catálise , Compostos Ferrosos/química , Metalocenos , Polímeros/química , Estereoisomerismo , Zinco/química
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