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1.
J Chem Inf Model ; 60(11): 5513-5528, 2020 11 23.
Artículo en Inglés | MEDLINE | ID: mdl-32786224

RESUMEN

Bacterial glycosyltransferases of the GT51 family are key enzymes in bacterial cell wall synthesis. Inhibiting cell wall synthesis is a very effective approach for development of antibiotics, as this can lead to either bacteriostatic or bactericidal effects. Even though the existence of this family has been known for over 50 years, only one potent inhibitor exists, which is an analog of the lipid IV product and derived from a natural product. Drug development focused on bacterial transglycosylase has been hampered due to little being know about its structure and reaction mechanism. In this study, Staphylococcus aureus monoglycosyltransferase was investigated at an atomistic level using computational methods. Classical molecular dynamics simulations were used to reveal information about the large-scale dynamics of the enzyme-substrate complex and the importance of magnesium in structure and function of the protein, while mixed mode quantum mechanics/molecular mechanics calculations unveiled a novel hypothesis for the reaction mechanism. From these results, we present a new model for the binding mode of lipid II and the reaction mechanism of the GT51 glycosyltransferases. A metal-bound hydroxide catalyzed reaction mechanism yields an estimated free energy barrier of 16.1 ± 1.0 kcal/mol, which is in line with experimental values. The importance of divalent cations is also further discussed. These findings could significantly aid targeted drug design, particularly the efficient development of transition state analogues as potential inhibitors for the GT51 glycosyltransferases.


Asunto(s)
Proteínas Bacterianas/química , Glicosiltransferasas , Staphylococcus aureus , Antibacterianos , Glicosiltransferasas/química , Simulación de Dinámica Molecular , Teoría Cuántica , Staphylococcus aureus/enzimología
2.
J Chem Inf Model ; 60(10): 4881-4893, 2020 10 26.
Artículo en Inglés | MEDLINE | ID: mdl-32820916

RESUMEN

The fragment docking program solvation energy for exhaustive docking (SEED) is evaluated on 15 different protein targets, with a focus on enrichment and the hit rate. It is shown that SEED allows for consistent computational enrichment of fragment libraries, independent of the effective hit rate. Depending on the actual target protein, true positive rates ranging up to 27% are observed at a cutoff value corresponding to the experimental hit rate. The impact of variations in docking protocols and energy filters is discussed in detail. Remaining issues, limitations, and use cases of SEED are also discussed. Our results show that fragment library selection or enhancement for a particular target is likely to benefit from docking with SEED, suggesting that SEED is a useful resource for fragment screening campaigns. A workflow is presented for the use of the program in virtual screening, including filtering and postprocessing to optimize hit rates.


Asunto(s)
Proteínas , Ligandos , Unión Proteica , Proteínas/metabolismo
3.
Int J Mol Sci ; 20(9)2019 May 07.
Artículo en Inglés | MEDLINE | ID: mdl-31067645

RESUMEN

The caseinolytic protease proteolytic subunit (ClpP) is a serine protease playing an important role in proteostasis of eukaryotic organelles and prokaryotic cells. Alteration of ClpP function has been proved to affect the virulence and infectivity of a number of pathogens. Increased bacterial resistance to antibiotics has become a global problem and new classes of antibiotics with novel mechanisms of action are needed. In this regard, ClpP has emerged as an attractive and potentially viable option to tackle pathogen fitness without suffering cross-resistance to established antibiotic classes and, when not an essential target, without causing an evolutionary selection pressure. This opens a greater window of opportunity for the host immune system to clear the infection by itself or by co-administration with commonly prescribed antibiotics. A comprehensive overview of the function, regulation and structure of ClpP across the different organisms is given. Discussion about mechanism of action of this protease in bacterial pathogenesis and human diseases are outlined, focusing on the compounds developed in order to target the activation or inhibition of ClpP.


Asunto(s)
Antibacterianos/farmacología , Proteínas Bacterianas/metabolismo , Depsipéptidos/farmacología , Endopeptidasa Clp/metabolismo , Antibacterianos/química , Bacterias/efectos de los fármacos , Bacterias/enzimología , Proteínas Bacterianas/agonistas , Proteínas Bacterianas/química , Depsipéptidos/química , Diseño de Fármacos , Endopeptidasa Clp/química
4.
J Chem Inf Model ; 58(11): 2193-2202, 2018 11 26.
Artículo en Inglés | MEDLINE | ID: mdl-30336018

RESUMEN

Simulations of membrane proteins have been rising in popularity in the past decade. Advancements in technology and force fields made it possible to simulate behavior of membrane proteins. Membrane protein simulations can now be used as supporting evidence for experimental findings, for elucidating protein mechanisms, and validating protein crystal structures. Unrelated to experimental data, these simulations can also serve to investigate larger scale processes like protein sorting, protein-membrane interactions, and more. In this review, the history as well as the state-of-the-art methodologies in membrane protein simulations will be summarized. An emphasis will be put on how to set up the system and on the current models for the different components of the simulation system. An overview of the available tools for membrane protein simulation will be given, and current limitations and prospects will also be discussed.


Asunto(s)
Membrana Dobles de Lípidos/química , Proteínas de la Membrana/química , Simulación de Dinámica Molecular , Animales , Bases de Datos de Proteínas , Humanos , Fosfolípidos/química , Programas Informáticos
5.
Clin Chem Lab Med ; 53(8): 1197-204, 2015 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-25720076

RESUMEN

BACKGROUND: Manufacturers and laboratories might benefit from using a modern integrated tool for quality management/assurance. The tool should not be confounded by commutability issues and focus on the intrinsic analytical quality and comparability of assays as performed in routine laboratories. In addition, it should enable monitoring of long-term stability of performance, with the possibility to quasi "real-time" remedial action. Therefore, we developed the "Empower" project. METHODS: The project comprises four pillars: (i) master comparisons with panels of frozen single-donation samples, (ii) monitoring of patient percentiles and (iii) internal quality control data, and (iv) conceptual and statistical education about analytical quality. In the pillars described here (i and ii), state-of-the-art as well as biologically derived specifications are used. RESULTS: In the 2014 master comparisons survey, 125 laboratories forming 8 peer groups participated. It showed not only good intrinsic analytical quality of assays but also assay biases/non-comparability. Although laboratory performance was mostly satisfactory, sometimes huge between-laboratory differences were observed. In patient percentile monitoring, currently, 100 laboratories participate with 182 devices. Particularly, laboratories with a high daily throughput and low patient population variation show a stable moving median in time with good between-instrument concordance. Shifts/drifts due to lot changes are sometimes revealed. There is evidence that outpatient medians mirror the calibration set-points shown in the master comparisons. CONCLUSIONS: The Empower project gives manufacturers and laboratories a realistic view on assay quality/comparability as well as stability of performance and/or the reasons for increased variation. Therefore, it is a modern tool for quality management/assurance toward improved patient care.


Asunto(s)
Análisis Químico de la Sangre/normas , Servicios de Laboratorio Clínico/normas , Manejo de Especímenes/normas , Congelación , Humanos , Control de Calidad
6.
Front Chem ; 11: 1160164, 2023.
Artículo en Inglés | MEDLINE | ID: mdl-37090247

RESUMEN

Receptor-Interacting serine/threonine-Protein Kinase 1 (RIPK1) emerged as an important driver of inflammation and, consequently, inflammatory pathologies. The enzymatic activity of RIPK1 is known to indirectly promote inflammation by triggering cell death, in the form of apoptosis, necroptosis and pyroptosis. Small molecule Receptor-Interacting serine/threonine-Protein Kinase 1 inhibitors have therefore recently entered clinical trials for the treatment of a subset of inflammatory pathologies. We previously identified GSK2656157 (GSK'157), a supposedly specific inhibitor of protein kinase R (PKR)-like ER kinase (PERK), as a much more potent type II Receptor-Interacting serine/threonine-Protein Kinase 1 inhibitor. We now performed further structural optimisation on the GSK'157 scaffold in order to develop a novel class of more selective Receptor-Interacting serine/threonine-Protein Kinase 1 inhibitors. Based on a structure-activity relationship (SAR) reported in the literature, we anticipated that introducing a substituent on the para-position of the pyridinyl ring would decrease the interaction with PERK. Herein, we report a series of novel GSK'157 analogues with different para-substituents with increased selectivity for Receptor-Interacting serine/threonine-Protein Kinase 1. The optimisation led to UAMC-3861 as the best compound of this series in terms of activity and selectivity for Receptor-Interacting serine/threonine-Protein Kinase 1 over PERK. The most selective compounds were screened in vitro for their ability to inhibit RIPK1-dependent apoptosis and necroptosis. With this work, we successfully synthesised a novel series of potent and selective type II Receptor-Interacting serine/threonine-Protein Kinase 1 inhibitors based on the GSK'157 scaffold.

7.
Eur J Med Chem ; 259: 115668, 2023 Nov 05.
Artículo en Inglés | MEDLINE | ID: mdl-37490800

RESUMEN

The taxane class of microtubule stabilizers are some of the most effective and widely used chemotherapeutics. The anticancer activity of taxanes arises from their ability to induce tubulin assembly by selectively recognizing the curved (c-) conformation in unassembled tubulin as compared to the straight (s-) conformation in assembled tubulin. We first designed and synthesized a series of 3'N-modified taxanes bearing covalent groups. Instead of discovering covalent taxanes, we found a series of non-covalent taxanes 2, in which the 3'N side chain was found to be essential for cytotoxicity due to its role in locking tubulin in the s-conformation. A representative compound bearing an acrylamide moiety (2h) exhibited increased binding affinity to the unassembled tubulin c-conformation and less cytotoxicity than paclitaxel. Further exploration of chemical space around 2h afforded a new series 3, in which derivatives such as 3l bind more tightly to both the s- and c-conformations of tubulin compared to paclitaxel, leading to more efficient promotion of tubulin polymerization and a greater persistence of in vitro efficacy against breast cancer cells after drug washout. Although 3l also had improved in vivo potency as compared to paclitaxel, it was also associated with increased systemic toxicity that required localized, intratumoral injection to observe potent and prolonged antitumor efficacy.


Asunto(s)
Paclitaxel , Tubulina (Proteína) , Tubulina (Proteína)/metabolismo , Paclitaxel/farmacología , Paclitaxel/química , Taxoides/farmacología , Taxoides/química , Microtúbulos
10.
J Med Chem ; 61(22): 10126-10140, 2018 11 21.
Artículo en Inglés | MEDLINE | ID: mdl-30354101

RESUMEN

Ferroptosis is an iron-catalyzed, nonapoptotic form of regulated necrosis that results in oxidative lipid damage in cell membranes that can be inhibited by the radical-trapping antioxidant Ferrostatin-1 (Fer-1). Novel inhibitors derived from the Fer-1 scaffold inhibited ferroptosis potently but suffered from solubility issues. In this paper, we report the synthesis of a more stable and readily soluble series of Fer-1 analogues that potently inhibit ferroptosis. The most promising compounds (37, 38, and 39) showed an improved protection compared to Fer-1 against multiorgan injury in mice. No toxicity was observed in mice after daily injection of 39 (UAMC-3203) for 4 weeks. UAMC-3203 inserts rapidly in a phospholipid bilayer in silico, which aligns with the current understanding of the mechanism of action of these compounds. In conclusion, these analogues have superior properties compared to Fer-1, show in vivo efficacy, and represent novel lead compounds with therapeutic potential in relevant ferroptosis-driven disease models.


Asunto(s)
Apoptosis/efectos de los fármacos , Ciclohexilaminas/metabolismo , Diseño de Fármacos , Fenilendiaminas/metabolismo , Animales , Línea Celular Tumoral , Membrana Celular/efectos de los fármacos , Membrana Celular/metabolismo , Humanos , Ratones , Modelos Moleculares , Conformación Molecular , Estrés Oxidativo/efectos de los fármacos , Ratas , Distribución Tisular
11.
J Med Chem ; 61(5): 1895-1920, 2018 03 08.
Artículo en Inglés | MEDLINE | ID: mdl-29437386

RESUMEN

Receptor interacting protein kinase 1 (RIPK1) plays a crucial role in tumor necrosis factor (TNF)-induced necroptosis, suggesting that this pathway might be druggable. Most inhibitors of RIPK1 are classified as either type II or type III kinase inhibitors. This opened up some interesting perspectives for the discovery of novel inhibitors that target the active site of RIPK1. Tozasertib, a type I pan-aurora kinase (AurK) inhibitor, was found to show a very high affinity for RIPK1. Because tozasertib presents the typical structural elements of a type I kinase inhibitor, the development of structural analogues of tozasertib is a good starting point for identifying novel type I RIPK1 inhibitors. In this paper, we identified interesting inhibitors of mTNF-induced necroptosis with no significant effect on AurK A and B, resulting in no nuclear abnormalities as is the case for tozasertib. Compounds 71 and 72 outperformed tozasertib in an in vivo TNF-induced systemic inflammatory response syndrome (SIRS) mouse model.


Asunto(s)
Necrosis/prevención & control , Piperazinas/química , Inhibidores de Proteínas Quinasas/farmacología , Animales , Aurora Quinasa A/efectos de los fármacos , Aurora Quinasa B/efectos de los fármacos , Muerte Celular/efectos de los fármacos , Ratones , Piperazinas/efectos adversos , Inhibidores de Proteínas Quinasas/química , Proteína Serina-Treonina Quinasas de Interacción con Receptores/antagonistas & inhibidores , Síndrome de Respuesta Inflamatoria Sistémica/inducido químicamente , Síndrome de Respuesta Inflamatoria Sistémica/tratamiento farmacológico , Factor de Necrosis Tumoral alfa/efectos adversos
12.
Clin Chim Acta ; 467: 8-14, 2017 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-27132242

RESUMEN

Clinicians diagnose thyroid dysfunction based on TSH and FT4 testing. However, the current lack of comparability between assays limits the optimal use of laboratory data. The International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) gave a mandate to the Committee for Standardization of Thyroid Function Tests (C-STFT) to resolve this limitation by standardization. Recently, the Committee members and their partners felt ready to set the step towards the technical recalibration. However, before implementation, they were furthered by the Food and Drugs Administration (FDA) to develop a tool to assess the sustainability of the new calibration basis. C-STFT began to use 2 online applications, i.e., the "Percentiler" and "Flagger", with the intention to assess their utility for this purpose. The tools monitor the course of instrument-specific moving medians of outpatient results (Percentiler) and flagging rates (Flagger) from data of individual laboratories grouped by instrument/assay peer. They additionally document the mid- to long-term medians, hence, are quality indicators of stability of performance of both laboratories and peers/assays. Here, the first experiences built up in the pre-standardization phase are reported. They suggest the suitability of both applications to document the sustainability of the calibration basis in the post-standardization phase.


Asunto(s)
Análisis Químico de la Sangre/normas , Pacientes Ambulatorios , Tirotropina/sangre , Tiroxina/sangre , Humanos , Estándares de Referencia , Factores de Tiempo
13.
Clin Chim Acta ; 445: 12-8, 2015 May 20.
Artículo en Inglés | MEDLINE | ID: mdl-25771106

RESUMEN

BACKGROUND: "The Percentiler" project provides quasi real-time access to patient medians across laboratories and manufacturers. This data can serve as "clearinghouse" for electronic health record applications, e.g., use of laboratory data for global health-care research. METHODS: Participants send their daily outpatient medians to the Percentiler application. After 6 to 8weeks, the laboratory receives its login information, which gives access to the user interface. Data is assessed by peer group, i.e., 10 or more laboratories using the same test system. Participation is free of charge. RESULTS: Participation is global with, to date, >120 laboratories and >250 instruments. Up to now, several reports have been produced that address i) the general features of the project, ii) peer group observations; iii) synergisms between "The Percentiler" and dedicated external quality assessment surveys. Reasons for long-term instability and bias (calibration- or lot-effects) have been observed for the individual laboratory and manufacturers. CONCLUSIONS: "The Percentiler" project has the potential to build a continuous, global evidence base on in vitro diagnostic test comparability and stability. As such, it may be beneficial for all stakeholders and, in particular, the patient. The medical laboratory is empowered for contributing to the development, implementation, and management of global health-care policies.


Asunto(s)
Técnicas de Química Analítica/normas , Laboratorios/normas , Juego de Reactivos para Diagnóstico/normas , Humanos , Cooperación Internacional , Variaciones Dependientes del Observador , Control de Calidad , Reproducibilidad de los Resultados
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