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1.
Front Oncol ; 13: 1250195, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37965471

RESUMO

The core activity of population-based cancer registries (PBCRs) is to gather information from all new cancer cases in a defined geographic area, in order to measure the magnitude of cancer burden and to provide a basis for cancer research. The Joint Research Centre-European Network of Cancer Registries Quality Check Software (JRC-ENCR QCS) is a Java standalone desktop application, under development since 2015, created to support PBCRs in the validation of the collected data. The JRC-ENCR QCS performs internal consistency checks on the cancer registry dataset, to detect impossible or unlikely codes or combination of codes, and is thereby an important tool to support the validation efforts by registries and improve data quality and European-wide harmonisation. The software package also includes the JRC CSV Data layout converter, a complementary tool for transforming PBCR incidence files into a format compatible with the JRC-ENCR QCS. This paper gives an overview of the JRC-ENCR QCS, describing the role of the software in processing data files submitted by PBCRs contributing to the European Cancer Information System (ECIS) as well as its functionalities. The development of the JRC-ENCR QCS is an evolving process, with regular updates implementing new and revised European and International recommendations and classifications.

2.
J Biomed Semantics ; 13(1): 7, 2022 02 22.
Artigo em Inglês | MEDLINE | ID: mdl-35193690

RESUMO

BACKGROUND: Population-based cancer registries are a critical reference source for the surveillance and control of cancer. Cancer registries work extensively with the internationally recognised TNM classification system used to stage solid tumours, but the system is complex and compounded by the different TNM editions in concurrent use. TNM ontologies exist but the design requirements are different for the needs of the clinical and cancer-registry domains. Two TNM ontologies developed specifically for cancer registries were designed for different purposes and have limitations for serving wider application. A unified ontology is proposed to serve the various cancer registry TNM-related tasks and reduce the multiplication effects of different ontologies serving specific tasks. The ontology is comprehensive of the rules for TNM edition 7 as required by cancer registries and designed on a modular basis to allow extension to other TNM editions. RESULTS: A unified ontology was developed building on the experience and design of the existing ontologies. It follows a modular approach allowing plug in of components dependent upon any particular TNM edition. A Java front-end was developed to interface with the ontology via the Web Ontology Language application programme interface and enables batch validation or classification of cancer registry records. The programme also allows the means of automated error correction in some instances. Initial tests verified the design concept by correctly inferring TNM stage and successfully handling the TNM-related validation checks on a number of cancer case records, with a performance similar to that of an existing ontology dedicated to the task. CONCLUSIONS: The unified ontology provides a multi-purpose tool for TNM-related tasks in a cancer registry and is scalable for different editions of TNM. It offers a convenient way of quickly checking validity of cancer case stage information and for batch processing of multi-record data via a dedicated front-end programme. The ontology is adaptable to many uses, either as a standalone TNM module or as a component in applications of wider focus. It provides a first step towards a single, unified TNM ontology for cancer registries.


Assuntos
Ontologias Biológicas , Neoplasias , Humanos , Idioma , Estadiamento de Neoplasias , Neoplasias/patologia , Sistema de Registros
3.
Curr Opin Cell Biol ; 20(4): 470-5, 2008 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-18456488

RESUMO

Animal cells have evolved different mechanisms to communicate with one another. In 2004, a new route of cell-to-cell communication mediated by tunneling nanotubes (TNT) was reported. These membranous cell bridges form de novo between cells and mediate the intercellular transfer of organelles, plasma membrane components and cytoplasmic molecules. The characterization of TNT-like bridges from several cell types revealed variations in the cytoskeletal composition as well as in the modality by which they interconnect cells, suggesting that different subclasses may exist. Furthermore, the growing number of cell types for which TNT-like structures were detected, supports the view that they represent a general mechanism for functional connectivity between cells, which could have important implications under physiological conditions.


Assuntos
Comunicação Celular/fisiologia , Membrana Celular/fisiologia , Nanotubos , Animais , Transporte Biológico , Humanos
4.
Biochemistry ; 44(12): 4886-96, 2005 Mar 29.
Artigo em Inglês | MEDLINE | ID: mdl-15779915

RESUMO

Recombinant human thymidine kinase 2 (hTK2) expressed in Escherichia coli has been found to bind tightly a substoichiometric amount of deoxyribonucleoside triphosphates (dTTP > dCTP >> dATP), known to be strong feedback inhibitors of the enzyme. Incubation of hTK2 with the substrate dThd was able to release the dNTPs from the active site during purification from E. coli and thus allowed the kinetic characterization of the noninhibited enzyme, with the tetrameric hTK2 showing slightly higher activity than the most abundant dimeric form. The unliganded hTK2 revealed a lower structural stability than the inhibitor-bound enzyme forms, being more prone to aggregation, thermal denaturation, and limited proteolysis. Moreover, intrinsic tryptophan fluorescence (ITF), far-UV circular dichroism (CD), and limited proteolysis have revealed that hTK2 undergoes distinct conformational changes upon binding different substrates and inhibitors, which are known to occur in the nucleoside monophosphate kinase family. The CD-monitored thermal denaturation of hTK2 dimer/tetramer revealed an irreversible process that can be satisfactorily described by the two-state irreversible denaturation model. On the basis of this model, the parameters of the Arrhenius equation were calculated, providing evidence for a significant structural stabilization of the enzyme upon ligand binding (dCyd < MgdCTP < dThd < dCTP < dTTP < MgdTTP), whereas MgATP further destabilizes the enzyme. Finally, surface plasmon resonance (SPR) was used to study in real time the reversible binding of substrates and inhibitors to the immobilized enzyme. The binding affinities for the inhibitors were found to be 1-2 orders of magnitude higher than for the corresponding substrates, both by SPR and ITF analysis.


Assuntos
Timidina Quinase/química , Timidina Quinase/metabolismo , Dicroísmo Circular/métodos , Desoxirribonucleotídeos/metabolismo , Dimerização , Inibidores Enzimáticos/metabolismo , Humanos , Hidrólise , Isoenzimas/química , Isoenzimas/isolamento & purificação , Isoenzimas/metabolismo , Cinética , Ligantes , Conformação Proteica , Desnaturação Proteica , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Espectrometria de Fluorescência , Ressonância de Plasmônio de Superfície/métodos , Timidina Quinase/isolamento & purificação , Tripsina/química , Triptofano/química
5.
J Biol Chem ; 279(25): 26571-80, 2004 Jun 18.
Artigo em Inglês | MEDLINE | ID: mdl-15060071

RESUMO

Phenylalanine hydroxylase (PAH) is generally considered to undergo a large and reversible conformational transition upon l-Phe binding, which is closely linked to the substrate-induced catalytic activation of this hysteretic enzyme. Recently, several crystallographically solvent-exposed hinge-bending regions including residues 31-34, 111-117, 218-226, and 425-429 have been defined/predicted to be involved in the intra-protomer propagation of the substrate-triggered molecular motions generated at the active site. On this basis, single-site mutagenesis of key residues in these regions of the human PAH tetramer was performed in the present study, and their functional impact was measured by steady-state kinetics and the global conformational transition as assessed by surface plasmon resonance and intrinsic tryptophan fluorescence spectroscopy. A strong correlation (r(2) = 0.93-0.96) was observed between the l-Phe-induced global conformational transition and V(max) values for wild-type human PAH and the mutant forms K113P, N223D, N426D, and N32D, in contrast to the substitution T427P, which resulted in a tetrameric form with no kinetic cooperativity. Furthermore, the flexible intra-domain linker region (residues 31-34) seems to be involved in a more local conformational change, and the biochemical/biophysical properties of the G33A/G33V mutant forms support a key function of this residue in the positioning of the autoregulatory sequence (residues 1-30) and thus in the regulation of the solvent and substrate access to the active site. The mutant forms revealed a variably reduced global conformational stability compared with wild-type human PAH, as measured by thermal denaturation and limited proteolysis.


Assuntos
Fenilalanina Hidroxilase/química , Sítios de Ligação , Domínio Catalítico , Dicroísmo Circular , Cristalografia por Raios X , Relação Dose-Resposta a Droga , Escherichia coli/metabolismo , Humanos , Cinética , Microscopia de Fluorescência , Modelos Químicos , Modelos Moleculares , Mutagênese Sítio-Dirigida , Mutação , Ligação Proteica , Conformação Proteica , Estrutura Terciária de Proteína , Espectrometria de Fluorescência , Ressonância de Plasmônio de Superfície , Temperatura , Tripsina/química , Triptofano/química
6.
J Biol Chem ; 278(17): 15142-52, 2003 Apr 25.
Artigo em Inglês | MEDLINE | ID: mdl-12554741

RESUMO

Recombinant human phenylalanine hydroxylase (hPAH) expressed in Escherichia coli for 24 h at 28 degrees C has been found by two-dimensional electrophoresis to exist as a mixture of four to five molecular forms as a result of nonenzymatic deamidation of labile Asn residues. The multiple deamidations alter the functional properties of the enzyme including its affinity for l-phenylalanine and tetrahydrobiopterin, catalytic efficiency, and substrate inhibition and also result in enzyme forms more susceptible to limited tryptic proteolysis. Asn(32) in the regulatory domain deamidates very rapidly because of its nearest neighbor amino acid Gly(33) (Solstad, T., Carvalho, R. N., Andersen, O. A., Waidelich, D., and Flatmark, T. (2003) Eur. J. Biochem., in press). Matrix-assisted laser desorption/ionization time of flight-mass spectrometry of the tryptic peptides in the catalytic domain of a 24-h (28 degrees C) expressed enzyme has shown Asn(376) and Asn(133) to be labile residues. Site-directed mutagenesis of nine Asn residues revealed that the deamidations of Asn(32) and Asn(376) are the main determinants for the functional and regulatory differences observed between the 2- and 24-h-induced wild-type (wt) enzyme. The Asn(32) --> Asp, Asn(376) --> Asp, and the double mutant forms expressed for 2 h at 28 degrees C revealed qualitatively similar regulatory properties as the highly deamidated 24-h expressed wt-hPAH. Moreover, deamidation of Asn(32) in the wt-hPAH (24 h expression at 28 degrees C) and the Asn(32) --> Asp mutation both increase the initial rate of phosphorylation of Ser(16) by cAMP-dependent protein kinase (p < 0.005). By contrast, the substitution of Gly(33) with Ala or Val, both preventing the deamidation of Asn(32), resulted in enzyme forms that were phosphorylated at a similar rate as nondeamidated wt-hPAH, even on 24-h expression. The other Asn --> Asp substitutions (in the catalytic domain) revealed that Asn(207) and Asn(223) have an important stabilizing structural function. Finally, two recently reported phenylketonuria mutations at Asn residues in the catalytic domain were studied, i.e. Asn(167) --> Ile and Asn(207) --> Asp, and their phenotypes were characterized.


Assuntos
Asparagina/metabolismo , Estabilidade Enzimática , Fenilalanina Hidroxilase/metabolismo , Amidas/metabolismo , Sequência de Aminoácidos , Asparagina/genética , Ácido Aspártico , Domínio Catalítico , Humanos , Cinética , Mutagênese Sítio-Dirigida , Mutação de Sentido Incorreto , Fenilalanina Hidroxilase/química , Fenilalanina Hidroxilase/genética , Proteínas Recombinantes/metabolismo , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz
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