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This corrects the article DOI: 10.1038/ncomms16081.
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The identification and prioritization of chemically tractable therapeutic targets is a significant challenge in the discovery of new medicines. We have developed a novel method that rapidly screens multiple proteins in parallel using DNA-encoded library technology (ELT). Initial efforts were focused on the efficient discovery of antibacterial leads against 119 targets from Acinetobacter baumannii and Staphylococcus aureus. The success of this effort led to the hypothesis that the relative number of ELT binders alone could be used to assess the ligandability of large sets of proteins. This concept was further explored by screening 42 targets from Mycobacterium tuberculosis. Active chemical series for six targets from our initial effort as well as three chemotypes for DHFR from M. tuberculosis are reported. The findings demonstrate that parallel ELT selections can be used to assess ligandability and highlight opportunities for successful lead and tool discovery.
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Acinetobacter baumannii/efectos de los fármacos , Antibacterianos/farmacología , Descubrimiento de Drogas/métodos , Biblioteca de Genes , Mycobacterium tuberculosis/efectos de los fármacos , Bibliotecas de Moléculas Pequeñas , Staphylococcus aureus/efectos de los fármacos , Acinetobacter baumannii/metabolismo , Evaluación Preclínica de Medicamentos , Terapia Molecular Dirigida , Mycobacterium tuberculosis/metabolismo , Staphylococcus aureus/metabolismoRESUMEN
RIP1 regulates necroptosis and inflammation and may play an important role in contributing to a variety of human pathologies, including immune-mediated inflammatory diseases. Small-molecule inhibitors of RIP1 kinase that are suitable for advancement into the clinic have yet to be described. Herein, we report our lead optimization of a benzoxazepinone hit from a DNA-encoded library and the discovery and profile of clinical candidate GSK2982772 (compound 5), currently in phase 2a clinical studies for psoriasis, rheumatoid arthritis, and ulcerative colitis. Compound 5 potently binds to RIP1 with exquisite kinase specificity and has excellent activity in blocking many TNF-dependent cellular responses. Highlighting its potential as a novel anti-inflammatory agent, the inhibitor was also able to reduce spontaneous production of cytokines from human ulcerative colitis explants. The highly favorable physicochemical and ADMET properties of 5, combined with high potency, led to a predicted low oral dose in humans.
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Antiinflamatorios/química , Antiinflamatorios/farmacología , Colitis Ulcerosa/tratamiento farmacológico , Inflamación/tratamiento farmacológico , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacología , Proteína Serina-Treonina Quinasas de Interacción con Receptores/antagonistas & inhibidores , Animales , Benzazepinas/química , Benzazepinas/farmacología , Colitis Ulcerosa/inmunología , Citocinas/inmunología , Perros , Haplorrinos , Humanos , Inflamación/inmunología , Ratones , Simulación del Acoplamiento Molecular , Conejos , Ratas , Proteína Serina-Treonina Quinasas de Interacción con Receptores/inmunología , Porcinos , Porcinos Enanos , Factor de Necrosis Tumoral alfa/inmunologíaRESUMEN
The recent discovery of the role of receptor interacting protein 1 (RIP1) kinase in tumor necrosis factor (TNF)-mediated inflammation has led to its emergence as a highly promising target for the treatment of multiple inflammatory diseases. We screened RIP1 against GSK's DNA-encoded small-molecule libraries and identified a novel highly potent benzoxazepinone inhibitor series. We demonstrate that this template possesses complete monokinase selectivity for RIP1 plus unique species selectivity for primate versus nonprimate RIP1. We elucidate the conformation of RIP1 bound to this benzoxazepinone inhibitor driving its high kinase selectivity and design specific mutations in murine RIP1 to restore potency to levels similar to primate RIP1. This series differentiates itself from known RIP1 inhibitors in combining high potency and kinase selectivity with good pharmacokinetic profiles in rodents. The favorable developability profile of this benzoxazepinone template, as exemplified by compound 14 (GSK'481), makes it an excellent starting point for further optimization into a RIP1 clinical candidate.
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ADN/química , Isoxazoles/farmacología , Oxazepinas/farmacología , Inhibidores de Proteínas Quinasas/farmacología , Proteína Serina-Treonina Quinasas de Interacción con Receptores/antagonistas & inhibidores , Bibliotecas de Moléculas Pequeñas/farmacología , Animales , Línea Celular Tumoral , Cristalografía por Rayos X , Relación Dosis-Respuesta a Droga , Células HT29 , Humanos , Isoxazoles/síntesis química , Isoxazoles/química , Ratones , Modelos Moleculares , Estructura Molecular , Oxazepinas/síntesis química , Oxazepinas/química , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/química , Proteína Serina-Treonina Quinasas de Interacción con Receptores/metabolismo , Bibliotecas de Moléculas Pequeñas/síntesis química , Bibliotecas de Moléculas Pequeñas/química , Relación Estructura-Actividad , Células U937RESUMEN
In the search of PI3K p110α wild type and H1047R mutant selective small molecule leads, an encoded library technology (ELT) campaign against the desired target proteins was performed which led to the discovery of a selective chemotype for PI3K isoforms from a three-cycle DNA encoded library. An X-ray crystal structure of a representative inhibitor from this chemotype demonstrated a unique binding mode in the p110α protein.
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Nod-like receptors (NLRs) are cytoplasmic pattern recognition receptors that are promising targets for the development of anti-inflammatory therapeutics. Drug discovery efforts targeting NLRs have been hampered by their inherent tendency to form aggregates making protein generation and the development of screening assays very challenging. Herein we report the results of an HTS screen of NLR family member NLRP1 (NLR family, pyrin domain-containing 1) which was achieved through the large scale generation of recombinant GST-His-Thrombin-NLRP1 protein. The screen led to the identification of a diverse set of ATP competitive inhibitors with micromolar potencies. Activity of these hits was confirmed in a FP binding assay, and two homology models were employed to predict the possible binding mode of the leading series and facilitate further lead-optimization. These results highlight a promising strategy for the identification of inhibitors of NLR family members which are rapidly emerging as key drivers of inflammation in human disease.
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Proteínas Adaptadoras Transductoras de Señales/antagonistas & inhibidores , Adenosina Trifosfato/química , Proteínas Reguladoras de la Apoptosis/antagonistas & inhibidores , Inflamasomas/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Adenosina Trifosfato/metabolismo , Proteínas Reguladoras de la Apoptosis/genética , Proteínas Reguladoras de la Apoptosis/metabolismo , Sitios de Unión , Unión Competitiva , Ensayos Analíticos de Alto Rendimiento , Humanos , Simulación del Acoplamiento Molecular , Proteínas NLR , Unión Proteica , Estructura Terciaria de Proteína , Pirazoles/química , Pirazoles/metabolismo , Proteínas Recombinantes de Fusión/biosíntesis , Proteínas Recombinantes de Fusión/química , Proteínas Recombinantes de Fusión/aislamiento & purificación , Relación Estructura-ActividadRESUMEN
Potent inhibitors of RIP1 kinase from three distinct series, 1-aminoisoquinolines, pyrrolo[2,3-b]pyridines, and furo[2,3-d]pyrimidines, all of the type II class recognizing a DLG-out inactive conformation, were identified from screening of our in-house kinase focused sets. An exemplar from the furo[2,3-d]pyrimidine series showed a dose proportional response in protection from hypothermia in a mouse model of TNFα induced lethal shock.
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ATP citrate lyase (ACL) catalyzes an ATP-dependent biosynthetic reaction which produces acetyl-coenzyme A and oxaloacetate from citrate and coenzyme A (CoA). Studies were performed with recombinant human ACL to ascertain the nature of the catalytic phosphorylation that initiates the ACL reaction and the identity of the active site residues involved. Inactivation of ACL by treatment with diethylpyrocarbonate suggested the catalytic role of an active site histidine (i.e., His760), which was proposed to form a phosphohistidine species during catalysis. The pH-dependence of the pre-steady-state phosphorylation of ACL with [γ-(33)P]-ATP revealed an ionizable group with a pK(a) value of ~7.5, which must be unprotonated for the catalytic phosphorylation of ACL to occur. Mutagenesis of His760 to an alanine results in inactivation of the biosynthetic reaction of ACL, in good agreement with the involvement of a catalytic histidine. The nature of the formation of the phospho-ACL was further investigated by positional isotope exchange using [γ-(18)O(4)]-ATP. The ß,γ-bridge to nonbridge positional isotope exchange rate of [γ-(18)O(4)]-ATP achieved its maximal rate of 14 s(-1) in the absence of citrate and CoA. This rate decreased to 5 s(-1) when citrate was added, and was found to be 10 s(-1) when both citrate and CoA were present. The rapid positional isotope exchange rates indicated the presence of one or more catalytically relevant, highly reversible phosphorylated intermediates. Steady-state measurements in the absence of citrate and CoA showed that MgADP was produced by both wild type and H760A forms of ACL, with rates at three magnitudes lower than that of k(cat) for the full biosynthetic reaction. The ATPase activity of ACL, along with the small yet significant positional isotope exchange rate observed in H760A mutant ACL (~150 fold less than wild type), collectively suggested the presence of a second, albeit unproductive, phosphoryl transfer in ACL. Mathematical analysis and computational simulation suggested that the desorption of MgADP at a rate of ~7 s(-1) was the rate-limiting step in the biosynthesis of AcCoA and oxaloacetate.
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ATP Citrato (pro-S)-Liasa/química , ATP Citrato (pro-S)-Liasa/farmacocinética , ATP Citrato (pro-S)-Liasa/genética , Acetilcoenzima A/biosíntesis , Biocatálisis , Dominio Catalítico/genética , Secuencia Conservada , Medición de Intercambio de Deuterio , Histidina/química , Histidina/genética , Histidina/metabolismo , Humanos , Mutación , Ácido Oxaloacético/metabolismo , FosforilaciónRESUMEN
Steady-state kinetic analysis of focal adhesion kinase-1 (FAK1) was performed using radiometric measurement of phosphorylation of a synthetic peptide substrate (Ac-RRRRRRSETDDYAEIID-NH(2), FAK-tide) which corresponds to the sequence of an autophosphorylation site in FAK1. Initial velocity studies were consistent with a sequential kinetic mechanism, for which apparent kinetic values k(cat) (0.052 +/- 0.001 s(-1)), K(MgATP) (1.2 +/- 0.1 microM), K(iMgATP) (1.3 +/- 0.2 microM), K(FAK-tide) (5.6 +/- 0.4 microM), and K(iFAK-tide) (6.1 +/- 1.1 microM) were obtained. Product and dead-end inhibition data indicated that enzymatic phosphorylation of FAK-tide by FAK1 was best described by a random bi bi kinetic mechanism, for which both E-MgADP-FAK-tide and E-MgATP-P-FAK-tide dead-end complexes form. FAK1 catalyzed the betagamma-bridge:beta-nonbridge positional oxygen exchange of [gamma-(18)O(4)]ATP in the presence of 1 mM [gamma-(18)O(4)]ATP and 1.5 mM FAK-tide with a progressive time course which was commensurate with catalysis, resulting in a rate of exchange to catalysis of k(x)/k(cat) = 0.14 +/- 0.01. These results indicate that phosphoryl transfer is reversible and that a slow kinetic step follows formation of the E-MgADP-P-FAK-tide complex. Further kinetic studies performed in the presence of the microscopic viscosogen sucrose revealed that solvent viscosity had no effect on k(cat)/K(FAK-tide), while k(cat) and k(cat)/K(MgATP) were both decreased linearly at increasing solvent viscosity. Crystallographic characterization of inactive versus AMP-PNP-liganded structures of FAK1 showed that a large conformational motion of the activation loop upon ATP binding may be an essential step during catalysis and would explain the viscosity effect observed on k(cat)/K(m) for MgATP but not on k(cat)/K(m) for FAK-tide. From the positional isotope exchange, viscosity, and structural data it may be concluded that enzyme turnover (k(cat)) is rate-limited by both reversible phosphoryl group transfer (k(forward) approximately 0.2 s(-1) and k(reverse) approximately 0.04 s(-1)) and a slow step (k(conf) approximately 0.1 s(-1)) which is probably the opening of the activation loop after phosphoryl group transfer but preceding product release.
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Quinasa 1 de Adhesión Focal/química , Quinasa 1 de Adhesión Focal/metabolismo , Péptidos/metabolismo , Adenosina Trifosfato/metabolismo , Adenilil Imidodifosfato/química , Adenilil Imidodifosfato/metabolismo , Secuencia de Aminoácidos , Cristalografía por Rayos X , Humanos , Cinética , Modelos Biológicos , Modelos Moleculares , Datos de Secuencia Molecular , Péptidos/química , Fosforilación , Unión ProteicaRESUMEN
Phosphoinositide 3-kinases have been targeted for therapeutic research because they are key components of a cell signaling cascade controlling proliferation, growth, and survival. Direct activation of the PI3Kalpha pathway contributes to the development and progression of solid tumors in breast, endometrial, colon, ovarian, and gastric cancers. In the context of a drug discovery effort, the availability of a robust crystallographic system is a means to understand the subtle differences between ATP competitive inhibitor interactions with the active site and their selectivity against other PI3Kinase enzymes. To generate a suitable recombinant design for this purpose, a p85alpha-p110alpha fusion system was developed which enabled the expression and purification of a stoichiometrically homogeneous, constitutively active enzyme for structure determination with potent ATP competitive inhibitors (Raha et al., in preparation) [56]. This approach has yielded preparations with activity and inhibition characteristics comparable to those of the full-length PI3Kalpha from which X-ray diffracting crystals were grown with inhibitors bound in the active site.
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Fosfatidilinositol 3-Quinasas Clase II/metabolismo , Cristalografía por Rayos X , Inhibidores Enzimáticos/farmacología , Adenosina Trifosfato/metabolismo , Animales , Fusión Artificial Génica , Baculoviridae/metabolismo , Sitios de Unión , Células Cultivadas , Fosfatidilinositol 3-Quinasas Clase II/química , Fosfatidilinositol 3-Quinasas Clase II/genética , Fosfatidilinositol 3-Quinasa Clase Ia/genética , Diseño de Fármacos , Concentración 50 Inhibidora , Modelos Moleculares , Proteínas Recombinantes de Fusión/química , Proteínas Recombinantes de Fusión/aislamiento & purificación , Proteínas Recombinantes de Fusión/metabolismo , Spodoptera/citología , Spodoptera/metabolismo , Difracción de Rayos XRESUMEN
Differential activation of the phosphoinositide 3-kinase (PI3K)/AKT pathway has been linked to cancer. Activation occurs through gene amplification and activating mutations. High-frequency mutations in the gene encoding the p110alpha catalytic subunit of PI3K (PIK3CA) have been observed in a variety of tumors including colon, brain, breast, ovarian, and gastric. Inhibition of PI3K kinase activity may provide a specific way to treat multiple types of human cancer. A scintillation proximity assay (SPA) was developed to detect phosphatidylinositol 3-kinase catalytic activity. Using this assay format, steady-state kinetic parameters were compared for the PI3K class IA enzymes p110alpha, p110beta, and p110delta, each coexpressed with the regulatory subunit p85alpha or splice variant p55alpha. Inhibition by the natural product wortmannin and LY294002 was detected with potencies consistent with alternate assay formats. Other biochemical assay formats have been described for phosphoinositide 3-kinases but each has its unique limitations. The simple, inexpensive, sensitive high-throughput nature of the SPA format has advanced our knowledge of isoform-specific enzymology and will facilitate the discovery of novel PI3K inhibitors.
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Fosfatidilinositol 3-Quinasas/metabolismo , Subunidades de Proteína/metabolismo , Conteo por Cintilación/métodos , Biocatálisis/efectos de los fármacos , Productos Biológicos/farmacología , Relación Dosis-Respuesta a Droga , Evaluación Preclínica de Medicamentos , Inhibidores Enzimáticos/farmacología , Humanos , Concentración de Iones de Hidrógeno , Isoenzimas/antagonistas & inhibidores , Isoenzimas/metabolismo , Microesferas , Inhibidores de las Quinasa Fosfoinosítidos-3 , Subunidades de Proteína/antagonistas & inhibidores , VolumetríaRESUMEN
The PIK3CA gene, encoding the p110alpha catalytic subunit of Class IA PI3Ks (phosphoinositide 3-kinases), is frequently mutated in many human tumours. The three most common tumour-derived alleles of p110alpha, H1047R, E542K and E545K, were shown to potently activate PI3K signalling in human epithelial cells. In the present study, we examine the biochemical activity of the recombinantly purified PI3K oncogenic mutants. The kinetic characterizations of the wt (wild-type) and the three 'hot spot' PI3K mutants show that the mutants all have approx. 2-fold increase in lipid kinase activities. Interestingly, the phosphorylated IRS-1 (insulin receptor substrate-1) protein shows activation of the lipid kinase activity for the wt and H1047R but not E542K and E545K PI3Kalpha, suggesting that these mutations represent different mechanisms of lipid kinase activation and hence transforming activity in cancer cells.
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Oncogenes , Fosfatidilinositol 3-Quinasas/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Adenosina Trifosfato/metabolismo , Alelos , Dominio Catalítico , Fosfatidilinositol 3-Quinasa Clase I , Activación Enzimática , Humanos , Proteínas Sustrato del Receptor de Insulina , Cinética , Mutación , Fosfatidilinositol 3-Quinasas/metabolismo , Fosforilación , Proteínas Recombinantes/genética , Proteínas Recombinantes/aislamiento & purificación , Proteínas Recombinantes/metabolismo , Células Tumorales CultivadasRESUMEN
Serum and glucocorticoid-regulated kinase 1 (SGK1) is a serine/threonine protein kinase of the AGC family which participates in the control of epithelial ion transport and is implicated in proliferation and apoptosis. We report here the 1.9 A crystal structure of the catalytic domain of inactive human SGK1 in complex with AMP-PNP. SGK1 exists as a dimer formed by two intermolecular disulfide bonds between Cys258 in the activation loop and Cys193. Although most of the SGK1 structure closely resembles the common protein kinase fold, the structure around the active site is unique when compared to most protein kinases. The alphaC helix is not present in this inactive form of SGK1 crystal structure; instead, the segment corresponding to the C helix forms a beta-strand that is stabilized by the N-terminal segment of the activation loop through a short antiparallel beta-sheet. Since the differences from other kinases occur around the ATP binding site, this structure can provide valuable insight into the design of selective and highly potent ATP-competitive inhibitors of SGK1 kinase.
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Adenilil Imidodifosfato/química , Proteínas Inmediatas-Precoces/química , Proteínas Serina-Treonina Quinasas/química , Adenosina Trifosfato/metabolismo , Adenilil Imidodifosfato/metabolismo , Secuencia de Aminoácidos , Sitios de Unión , Dominio Catalítico , Cristalografía por Rayos X , Dimerización , Humanos , Interacciones Hidrofóbicas e Hidrofílicas , Proteínas Inmediatas-Precoces/aislamiento & purificación , Proteínas Inmediatas-Precoces/metabolismo , Modelos Moleculares , Datos de Secuencia Molecular , Estructura Molecular , Conformación Proteica , Proteínas Serina-Treonina Quinasas/aislamiento & purificación , Proteínas Serina-Treonina Quinasas/metabolismo , Estructura Secundaria de Proteína , Estructura Terciaria de Proteína , Alineación de SecuenciaRESUMEN
The mitotic kinesin KSP (kinesin spindle protein, or Eg5) has an essential role in centrosome separation and formation of the bipolar mitotic spindle. Its exclusive involvement in the mitotic spindle of proliferating cells presents an opportunity for developing new anticancer agents with reduced side effects relative to antimitotics that target tubulin. Ispinesib is an allosteric small-molecule KSP inhibitor in phase 2 clinical trials. Mutations that attenuate ispinesib binding to KSP have been identified, which highlights the need for inhibitors that target different binding sites. We describe a new class of selective KSP inhibitors that are active against ispinesib-resistant forms of KSP. These ATP-competitive KSP inhibitors do not bind in the nucleotide binding pocket. Cumulative data from generation of resistant cells, site-directed mutagenesis and photo-affinity labeling suggest that they compete with ATP binding via a novel allosteric mechanism.
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Adenosina Trifosfato/antagonistas & inhibidores , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Proteínas Quinasas/química , Proteínas Quinasas/metabolismo , Regulación Alostérica/efectos de los fármacos , Animales , Línea Celular , Supervivencia Celular/efectos de los fármacos , Concentración 50 Inhibidora , Modelos Moleculares , Estructura Molecular , Estructura Terciaria de ProteínaRESUMEN
Granulocyte colony-stimulating factor regulates neutrophil production by binding to a specific receptor, the granulocyte colony-stimulating factor receptor, expressed on cells of the granulocytic lineage. Recombinant forms of granulocyte colony-stimulating factor are used clinically to treat neutropenias. As part of an effort to develop granulocyte colony-stimulating factor mimics with the potential for oral bioavailability, we previously identified a nonpeptidyl small molecule (SB-247464) that selectively activates murine granulocyte colony-stimulating factor signal transduction pathways and promotes neutrophil formation in vivo. To elucidate the mechanism of action of SB-247464, a series of cell-based and biochemical assays were performed. The activity of SB-247464 is strictly dependent on the presence of zinc ions. Titration microcalorimetry experiments using a soluble murine granulocyte colony-stimulating factor receptor construct show that SB-247464 binds to the extracellular domain of the receptor in a zinc ion-dependent manner. Analytical ultracentrifugation studies demonstrate that SB-247464 induces self-association of the N-terminal three-domain fragment in a manner that is consistent with dimerization. SB-247464 induces internalization of granulocyte colony-stimulating factor receptor on intact cells, consistent with a mechanism involving receptor oligomerization. These data show that small nonpeptidyl compounds are capable of selectively binding and inducing productive oligomerization of cytokine receptors.
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Receptores de Factor Estimulante de Colonias de Granulocito/química , Receptores de Factor Estimulante de Colonias de Granulocito/metabolismo , Animales , Bencimidazoles/farmacología , Células de la Médula Ósea/metabolismo , Calorimetría , Línea Celular , Dicroismo Circular , Citocinas/metabolismo , Dimerización , Relación Dosis-Respuesta a Droga , Ácido Edético/farmacología , Guanidinas/farmacología , Iones , Ligandos , Ratones , Modelos Químicos , Péptidos/química , Unión Proteica , Estructura Terciaria de Proteína , Proteínas Recombinantes de Fusión/metabolismo , Transducción de Señal , Transfección , Ultracentrifugación , ZincRESUMEN
We have recently reported the identification of four novel members of the interleukin-1 (IL-1) family which we designated as IL-1 homologue 1-4 (IL-1H1-4). These proteins exhibit significant sequence homology to other members of the IL-1 family. Of these homologues, only IL-1H4 (renamed IL-1F7b) was predicted to contain a propeptide domain and a caspase cleavage site. We now report that caspase-1 cleaves IL-1F7b at the predicted site to generate mature IL-1F7b. Caspase-4 was also able to process IL-1F7b, albeit inefficiently. Other caspases and Granzyme-B did not cleave IL-1F7b. Furthermore, adenovirus-mediated expression of IL-1F7b in HEK 293 cells led to in situ processing and secretion of mature IL-1F7b. In a screen to identify a potential receptor, both pro and mature IL-1F7b bound to the soluble IL-18 receptor alpha-Fc (IL-18Ralpha-Fc) but not to the soluble IL-1R-Fc or ST2R-Fc fusion proteins. Mature IL-1F7b bound to the IL-18Ralpha-Fc protein with higher affinity than the pro form, although the affinities for both proteins were significantly lower than that observed for IL-18. Consistent with this observation, only IL-18 and not IL-1F7b induced IFN-gamma production by KG1a cells. We also report that pro and mature IL-1F7b form homodimers with association constants of 4 microM and 5 nM, respectively, suggesting biological relevance to IL-1F7b processing. Finally, we have localized the expression of IL-1F7b protein in discrete cell populations including plasma cells and tumor cells. These data suggest that IL-1F7b may be involved in immune response, inflammatory diseases and/or cancer.
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Caspasa 1/metabolismo , Caspasas/metabolismo , Interferón gamma/biosíntesis , Interleucina-18/metabolismo , Interleucina-1/genética , Interleucina-1/metabolismo , Receptores de Interleucina/metabolismo , Secuencia de Aminoácidos , Sitios de Unión , Línea Celular , ADN Complementario , Humanos , Subunidad alfa del Receptor de Interleucina-18 , Receptores de Interleucina-18 , Proteínas Recombinantes de Fusión/metabolismoRESUMEN
Screening of our internal compound collection for inhibitors of the transforming growth factor beta1 (TGF-beta1) type I receptor (ALK5) identified several hits. Optimization of the dihydropyrroloimidazole hit 2 by introduction of a 2-pyridine and 3,4-methylenedioxyphenyl group gave 7, a selective ALK5 inhibitor. With this information, optimization of the triarylimidazole hit 8 gave the selective inhibitor 14, which inhibits TGF-beta1-induced fibronectin mRNA formation while displaying no measurable cytotoxicity in the 48 h XTT assay.