RESUMEN
Vaccinia-related kinase 1 (VRK1) and the δ and ε isoforms of casein kinase 1 (CK1) are linked to various disease-relevant pathways. However, the lack of tool compounds for these kinases has significantly hampered our understanding of their cellular functions and therapeutic potential. Here, we describe the structure-based development of potent inhibitors of VRK1, a kinase highly expressed in various tumor types and crucial for cell proliferation and genome integrity. Kinome-wide profiling revealed that our compounds also inhibit CK1δ and CK1ε. We demonstrate that dihydropteridinones 35 and 36 mimic the cellular outcomes of VRK1 depletion. Complementary studies with existing CK1δ and CK1ε inhibitors suggest that these kinases may play overlapping roles in cell proliferation and genome instability. Together, our findings highlight the potential of VRK1 inhibition in treating p53-deficient tumors and possibly enhancing the efficacy of existing cancer therapies that target DNA stability or cell division.
Asunto(s)
Inhibidores de Proteínas Quinasas , Proteínas Serina-Treonina Quinasas , Pteridinas , Humanos , Inhibidores de Proteínas Quinasas/farmacología , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/síntesis química , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Serina-Treonina Quinasas/metabolismo , Pteridinas/farmacología , Pteridinas/química , Pteridinas/síntesis química , Péptidos y Proteínas de Señalización Intracelular/antagonistas & inhibidores , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Proliferación Celular/efectos de los fármacos , Relación Estructura-Actividad , Quinasa Idelta de la Caseína/antagonistas & inhibidores , Quinasa Idelta de la Caseína/metabolismo , Caseína Cinasa 1 épsilon/antagonistas & inhibidores , Caseína Cinasa 1 épsilon/metabolismo , Línea Celular TumoralRESUMEN
The discovery of potent and selective inhibitors for understudied kinases can provide relevant pharmacological tools to illuminate their biological functions. DYRK1A and DYRK1B are protein kinases linked to chronic human diseases. Current DYRK1A/DYRK1B inhibitors also antagonize the function of related protein kinases, such as CDC2-like kinases (CLK1, CLK2, CLK4) and DYRK2. Here, we reveal narrow spectrum dual inhibitors of DYRK1A and DYRK1B based on a benzothiophene scaffold. Compound optimization exploited structural differences in the ATP-binding sites of the DYRK1 kinases and resulted in the discovery of 3n, a potent and cell-permeable DYRK1A/DYRK1B inhibitor. This compound has a different scaffold and a narrower off-target profile compared to current DYRK1A/DYRK1B inhibitors. We expect the benzothiophene derivatives described here to aid establishing DYRK1A/DYRK1B cellular functions and their role in human pathologies.
Asunto(s)
Proteínas Serina-Treonina Quinasas , Proteínas Tirosina Quinasas , Humanos , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Quinasas , Proteínas Tirosina Quinasas/metabolismo , TiofenosRESUMEN
Vaccinia-related kinases 1 and 2 (VRK1 and VRK2) are human Ser/Thr protein kinases associated with increased cell division and neurological disorders. Nevertheless, the cellular functions of these proteins are not fully understood. Despite their therapeutic potential, there are no potent and specific inhibitors available for VRK1 or VRK2. We report here the discovery and elaboration of an aminopyridine scaffold as a basis for VRK1 and VRK2 inhibitors. The most potent compound for VRK1 (26) displayed an IC50 value of 150 nM and was fairly selective in a panel of 48 human kinases (selectivity score S(50%) of 0.04). Differences in compound binding mode and substituent preferences between the two VRKs were identified by the structure-activity relationship combined with the crystallographic analysis of key compounds. We expect our results to serve as a starting point for the design of more specific and potent inhibitors against each of the two VRKs.
RESUMEN
Studies have linked the serine-threonine kinase MAP4K4 to the regulation of a number of biological processes and/or diseases, including diabetes, cancer, inflammation, and angiogenesis. With a majority of the members of our lead series (e.g., 1) suffering from time-dependent inhibition (TDI) of CYP3A4, we sought design avenues that would eliminate this risk. One such approach arose from the observation that carboxylic acid-based intermediates employed in our discovery efforts retained high MAP4K4 inhibitory potency and were devoid of the TDI risk. The medicinal chemistry effort that led to the discovery of this central nervous system-impaired inhibitor together with its preclinical safety profile is described.
Asunto(s)
Aminopiridinas/síntesis química , Aminopiridinas/farmacología , Péptidos y Proteínas de Señalización Intracelular/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Aminopiridinas/efectos adversos , Animales , Disponibilidad Biológica , Ácidos Carboxílicos/química , Inhibidores del Citocromo P-450 CYP3A/síntesis química , Inhibidores del Citocromo P-450 CYP3A/farmacología , Descubrimiento de Drogas , Semivida , Lipopolisacáridos/farmacología , Masculino , Ratones , Ratones Endogámicos C57BL , Modelos Moleculares , Conformación Molecular , Inhibidores de Proteínas Quinasas/efectos adversos , Ratas , Ratas Wistar , Relación Estructura-Actividad , Factor de Necrosis Tumoral alfa/sangreRESUMEN
The human genome encodes two active Vaccinia-related protein kinases (VRK), VRK1 and VRK2. These proteins have been implicated in a number of cellular processes and linked to a variety of tumors. However, understanding the cellular role of VRKs and establishing their potential use as targets for therapeutic intervention has been limited by the lack of tool compounds that can specifically modulate the activity of these kinases in cells. Here we identified BI-D1870, a dihydropteridine inhibitor of RSK kinases, as a promising starting point for the development of chemical probes targeting the active VRKs. We solved co-crystal structures of both VRK1 and VRK2 bound to BI-D1870 and of VRK1 bound to two broad-spectrum inhibitors. These structures revealed that both VRKs can adopt a P-loop folded conformation, which is stabilized by different mechanisms on each protein. Based on these structures, we suggest modifications to the dihydropteridine scaffold that can be explored to produce potent and specific inhibitors towards VRK1 and VRK2.
Asunto(s)
Antineoplásicos/química , Péptidos y Proteínas de Señalización Intracelular/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/química , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Pteridinas/química , Secuencia de Aminoácidos , Antineoplásicos/farmacología , Sitios de Unión , Clonación Molecular , Cristalografía por Rayos X , Escherichia coli/genética , Escherichia coli/metabolismo , Expresión Génica , Vectores Genéticos/química , Vectores Genéticos/metabolismo , Genoma Humano , Humanos , Péptidos y Proteínas de Señalización Intracelular/química , Péptidos y Proteínas de Señalización Intracelular/genética , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Cinética , Modelos Moleculares , Unión Proteica , Conformación Proteica en Hélice alfa , Conformación Proteica en Lámina beta , Pliegue de Proteína , Dominios y Motivos de Interacción de Proteínas , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Serina-Treonina Quinasas/química , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Pteridinas/farmacología , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Relación Estructura-Actividad , Virus Vaccinia/genética , Virus Vaccinia/metabolismoRESUMEN
Recent studies in adipose tissue, pancreas, muscle, and macrophages suggest that MAP4K4, a serine/threonine protein kinase may be a viable target for antidiabetic drugs. As part of the evaluation of MAP4K4 as a novel antidiabetic target, a tool compound, 16 (PF-6260933) and a lead 17 possessing excellent kinome selectivity and suitable properties were delivered to establish proof of concept in vivo. The medicinal chemistry effort that led to the discovery of these lead compounds is described herein together with in vivo pharmacokinetic properties and activity in a model of insulin resistance.
RESUMEN
Achiral [3.2.1] bridged heterocycles containing a bridging amide can undergo enantioselective hydroarylation reactions under rhodium(I) catalysis. These reactions proceed in high yield and enantioselectivity in most cases, under mild reaction conditions and using commercially available Josiphos ligands. The phosphine ligand structure and the protecting group on the nitrogen both have significant effects on the selectivity and yield of the reactions.
Asunto(s)
Compuestos Bicíclicos Heterocíclicos con Puentes/química , Compuestos Bicíclicos Heterocíclicos con Puentes/síntesis química , Compuestos Heterocíclicos de Anillo en Puente/química , Compuestos Heterocíclicos de Anillo en Puente/síntesis química , Amidas/química , Catálisis , Estructura Molecular , Rodio/química , EstereoisomerismoRESUMEN
A series of GPR119 agonists based on a 2,6-diazatricyclo[3.3.1.1â¼3,7â¼]decane ring system is described. Also provided is a detailed account of the development of a multigram scale synthesis of the diazatricyclic ring system, which was achieved using a Hofmann-Löffler-Freytag reaction as the key step. The basis for the use of this complex framework lies in an attempt to constrain one end of the molecule in the "agonist conformation" as was previously described for 3-oxa-7-aza-bicyclo[3.3.1]nonanes. Optimization of carbamate analogues of the diazatricylic compounds led to the identification of 32i as a potent agonist of the GPR119 receptor with low unbound human liver microsomal clearance. The use of an agonist response weighted ligand lipophilic efficiency (LLE) termed AgLLE is discussed along with the issues of applying efficiency measures to agonist programs. Ultimately, solubility limited absorption and poor exposure reduced further interest in these molecules.
Asunto(s)
Compuestos Aza/síntesis química , Hidrocarburos Aromáticos con Puentes/síntesis química , Ciclodecanos/síntesis química , Receptores Acoplados a Proteínas G/agonistas , Animales , Compuestos Aza/química , Compuestos Aza/farmacología , Disponibilidad Biológica , Hidrocarburos Aromáticos con Puentes/química , Hidrocarburos Aromáticos con Puentes/farmacología , Cristalografía por Rayos X , Ciclodecanos/química , Ciclodecanos/farmacología , Perros , Diseño de Fármacos , Humanos , Masculino , Microsomas Hepáticos/metabolismo , Estructura Molecular , Pirimidinas/síntesis química , Pirimidinas/química , Pirimidinas/farmacología , Ratas , Ratas Sprague-Dawley , Receptores Acoplados a Proteínas G/química , Solubilidad , Estereoisomerismo , Relación Estructura-ActividadRESUMEN
A novel GPR119 agonist based on the 2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazole scaffold was designed through lead optimization starting from pyrazole-based GPR119 agonist 1. The design is centered on the conformational restriction of the core scaffold, while minimizing the change in spatial relationships of two key pharmacophoric elements (piperidine-carbamate and aryl sulfone).
Asunto(s)
Pirazoles/química , Receptores Acoplados a Proteínas G/agonistas , Carbamatos/química , Humanos , Piperidinas/química , Unión Proteica , Pirazoles/síntesis química , Pirazoles/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Relación Estructura-ActividadRESUMEN
Previous drug discovery efforts identified classical PYK2 kinase inhibitors such as 2 and 3 that possess selectivity for PYK2 over its intra-family isoform FAK. Efforts to identify more kinome-selective chemical matter that stabilize a DFG-out conformation of the enzyme are described herein. Two sub-series of PYK2 inhibitors, an indole carboxamide-urea and a pyrazole-urea have been identified and found to have different binding interactions with the hinge region of PYK2. These leads proved to be more selective than the original classical inhibitors.
Asunto(s)
Quinasa 2 de Adhesión Focal/antagonistas & inhibidores , Indoles/farmacología , Inhibidores de Proteínas Quinasas/farmacología , Pirazoles/farmacología , Urea/farmacología , Animales , Cristalografía por Rayos X , Relación Dosis-Respuesta a Droga , Quinasa 2 de Adhesión Focal/metabolismo , Células HEK293 , Humanos , Indoles/síntesis química , Indoles/química , Modelos Moleculares , Estructura Molecular , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/química , Pirazoles/síntesis química , Pirazoles/química , Ratas , Relación Estructura-Actividad , Urea/análogos & derivados , Urea/químicaRESUMEN
The amination of bis-allylic imidates using an Iridium(I) catalyst leads to the efficient formation of 2,6-divinyl heterocycles. Careful screening of amines, solvents, and conditions has led to the discovery of a system that favors formation of the desired cis products with synthetically useful levels of diastereoselectivity, and these results are further explained by computer based transition state energy calculations. Exposure of the heterocycles to ring-closing metathesis catalysts leads to the desired bridged heterocyclic systems.
Asunto(s)
Aminas/síntesis química , Compuestos Heterocíclicos de Anillo en Puente/síntesis química , Iridio/química , Aminación , Aminas/química , Catálisis , Compuestos Heterocíclicos de Anillo en Puente/química , Estructura Molecular , EstereoisomerismoRESUMEN
In this paper, we describe a lead transformation tool, NEAT (Novel and Electronically equivalent Aromatic Template), which can help identify novel aromatic rings that are estimated to have similar electrostatic potentials, dipoles, and hydrogen bonding capabilities to a query template; hence, they may offer similar bioactivity profiles. In this work, we built a comprehensive heteroaryl database, and precalculated high-level quantum mechanical (QM) properties, including electrostatic potential charges, hydrogen bonding ability, dipole moments, chemical reactivity, and othe properties. NEAT bioisosteric similarities are based on the electrostatic potential surface calculated by Brood, using the precalculated QM ESP charges and other QM properties. Compared with existing commercial lead transformation software, (1) NEAT is the only one that covers the comprehensive heteroaryl chemical space, and (2) NEAT offers a better characterization of novel aryl cores by using high-evel QM properties that are relevant to molecular interactions. NEAT provides unique value to medicinal chemists quickly exploring the largely uncharted aromatic chemical space, and one successful example of its application is discussed herein.
Asunto(s)
Descubrimiento de Drogas , Hidrocarburos Aromáticos/química , Modelos Químicos , Teoría Cuántica , Humanos , Piperazinas/química , Purinas/química , Citrato de Sildenafil , Sulfonas/químicaRESUMEN
The application of conformationally dependent measures of size and polarity to characterize beyond rule-of-5 (Ro5) space for passive permeation was investigated. Specifically, radius of gyration, an alternative to molecular weight, and three-dimensional polar surface area and the generalized Born/surface area dehydration free energy, alternatives to hydrogen-bond donor and acceptor counts, were computed on models of the permeating conformations of over 35 000 molecules. The resulting guidelines for size and polarity, described by the 3D properties, should aid the design of Ro5 violators with passive permeability.
Asunto(s)
Membrana Celular/metabolismo , Modelos Químicos , Medicamentos bajo Prescripción/química , Animales , Permeabilidad de la Membrana Celular , Perros , Enlace de Hidrógeno , Interacciones Hidrofóbicas e Hidrofílicas , Cinética , Células de Riñón Canino Madin Darby , Conformación Molecular , Estructura Molecular , Peso Molecular , Electricidad Estática , TermodinámicaRESUMEN
Myeloperoxidase (MPO) is known to be inactivated and covalently modified by treatment with hydrogen peroxide and agents similar to 3-(2-ethoxypropyl)-2-thioxo-2,3-dihydro-1H-purin-6(9H)-one (1), a 254.08 Da derivative of 2-thioxanthine. Peptide mapping by liquid chromatography and mass spectrometry detected modification by 1 in a labile peptide-heme-peptide fragment of the enzyme, accompanied by a mass increase of 252.08 Da. The loss of two hydrogen atoms was consistent with mechanism-based oxidative coupling. Multistage mass spectrometry (MS(4)) of the modified fragment in an ion trap/Orbitrap spectrometer demonstrated that 1 was coupled directly to heme. Use of a 10 amu window delivered the full isotopic envelope of each precursor ion to collision-induced dissociation, preserving definitive isotopic profiles for iron-containing fragments through successive steps of multistage mass spectrometry. Iron isotope signatures and accurate mass measurements supported the structural assignments. Crystallographic analysis confirmed linkage between the methyl substituent of the heme pyrrole D ring and the sulfur atom of 1. The final orientation of 1 perpendicular to the plane of the heme ring suggested a mechanism consisting of two consecutive one-electron oxidations of 1 by MPO. Multistage mass spectrometry using stage-specific collision energies permits stepwise deconstruction of modifications of heme enzymes containing covalent links between the heme group and the polypeptide chain.
Asunto(s)
Hemo/química , Peroxidasa/química , Peroxidasa/metabolismo , Secuencia de Aminoácidos , Dominio Catalítico , Cromatografía Liquida , Cristalografía por Rayos X , Humanos , Modelos Químicos , Modelos Moleculares , Datos de Secuencia Molecular , Peso Molecular , Neutrófilos/enzimología , Oxidación-Reducción , Fragmentos de Péptidos/química , Mapeo Peptídico , Espectrometría de Masas en TándemRESUMEN
The critical issues in docking include the prediction of the correct binding pose and the accurate estimation of the corresponding binding affinity. Different docking methodologies have all been successful in reproducing the crystallographic binding modes, but struggle when predicting the corresponding binding affinities. The rescoring of docking poses using the MM-GB/SA technique has emerged as an important computational approach in structure-based lead optimization as it provides for congeneric molecules, clearly superior correlations with experimental data to those obtained with typical docking scoring functions. Although the technique has been collectively referred as MM-GB/SA, there are in fact many flavors in the literature. Here we describe the details of our MM-GB/SA scoring protocol, highlighting not only its strengths but also the limitations.
Asunto(s)
Biología Computacional/métodos , Proteínas/metabolismo , Programas Informáticos , Automatización , Bases de Datos de Proteínas , Unión Proteica , Conformación Proteica , Proteínas/química , TermodinámicaRESUMEN
This work addresses the link between selectivity and an unusual, folded conformation for the P-loop observed initially for MAP4K4 and subsequently for other kinases. Statistical and computational analyses of our crystal structure database demonstrate that inhibitors that induce the P-loop folded conformation tend to be more selective, especially if they take advantage of this specific conformation by interacting more favorably with a conserved Tyr or Phe residue from the P-loop.
Asunto(s)
Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Modelos Moleculares , Bases de Datos de Proteínas , Quinasas MAP Reguladas por Señal Extracelular/antagonistas & inhibidores , Quinasas MAP Reguladas por Señal Extracelular/química , Conformación Proteica , Pliegue de Proteína , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacología , Especificidad por SustratoRESUMEN
The synthesis and properties of the bridged piperidine (oxaazabicyclo) compounds 8, 9, and 11 are described. A conformational analysis of these structures is compared with the representative GPR119 ligand 1. These results and the differences in agonist pharmacology are used to formulate a conformation-based hypothesis to understand activation of the GPR119 receptor. We also show for these structures that the agonist pharmacology in rat masks the important differences in human pharmacology.
Asunto(s)
Compuestos de Azabiciclo/química , Compuestos de Azabiciclo/farmacología , Pirimidinas/química , Pirimidinas/farmacología , Receptores Acoplados a Proteínas G/agonistas , Animales , Compuestos de Azabiciclo/síntesis química , Prueba de Tolerancia a la Glucosa , Humanos , Conformación Molecular , Pirimidinas/síntesis química , Ratas , Receptores Acoplados a Proteínas G/antagonistas & inhibidores , Especificidad de la Especie , Estereoisomerismo , Relación Estructura-ActividadRESUMEN
The design and synthesis of a GPR119 agonist bearing a 2-(2,3,6-trifluorophenyl)acetamide group is described. The design capitalized on the conformational restriction found in N-ß-fluoroethylamide derivatives to help maintain good levels of potency while driving down both lipophilicity and oxidative metabolism in human liver microsomes. The chemical stability and bioactivation potential are discussed.
Asunto(s)
Acetamidas/química , Acetamidas/farmacología , Diseño de Fármacos , Receptores Acoplados a Proteínas G/agonistas , Acetamidas/síntesis química , Humanos , Concentración 50 Inhibidora , Espectroscopía de Resonancia Magnética , Estructura Molecular , Receptores Acoplados a Proteínas G/químicaRESUMEN
A variable dielectric model based on residue types for better description of protein-ligand electrostatics in MM-GBSA scoring is reported. The variable dielectric approach provides better correlation with binding data and reduces the score dynamic range, typically observed in the standard MM-GB/SA method. The latter supports the view that exaggerated enthalpic separation between weak and potent compounds due to the lack of shielding effects in the model is greatly responsible for the wide scoring spread.
RESUMEN
MM-GB/SA scoring and free energy perturbation (FEP) calculations have emerged as reliable methodologies to understand structural and energetic relationships to binding. In spite of successful applications to elucidate the structure-activity relationships for few pairs of ligands, the reality is that the performance of FEP calculations has rarely been tested for more than a handful of compounds. In this work, a series of 13 benzene sulfonamide inhibitors of carbonic anhydrase with binding free energies determined by isothermal titration calorimetry was selected as a test case. R(2) values of 0.70, 0.71, and 0.49 with the experiment were obtained with MM-GB/SA and FEP simulations run with MCPRO+ and Desmond, respectively. All methods work well, but the results obtained with Desmond are inferior to MM-GB/SA and MCPRO+. The main contrast between the methods is the level of sampling, ranging from full to restricted flexibility to single conformation for the complexes in Desmond, MCPRO+, and MM-GB/SA, respectively. The current and historical results obtained with MM-GB/SA qualify this approach as a more attractive alternative for rank-ordering; it can achieve equivalent or superior predictive accuracy and handle more structurally dissimilar ligands at a fraction of the computational cost of the rigorous free-energy methods. As for the large theoretical dynamic range for the binding energies, that seems to be a direct result of the degree of sampling in the simulations since MCPRO+ as well as MM-GB/SA are plagued by this. Van't Hoff analysis for selected pairs of ligands suggests that the wider scoring spread is not only affected by missing entropic contributions due to restricted sampling but also exaggerated enthalpic separation between the weak and potent compounds caused by diminished shielding of electrostatic interactions, thermal effects, and protein relaxation/strain.