RESUMEN
Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) ion channel are established as the primary causative factor in the devastating lung disease cystic fibrosis (CF). More recently, cigarette smoke exposure has been shown to be associated with dysfunctional airway epithelial ion transport, suggesting a role for CFTR in the pathogenesis of chronic obstructive pulmonary disease (COPD). Here, the identification and characterization of a high throughput screening hit 6 as a potentiator of mutant human F508del and wild-type CFTR channels is reported. The design, synthesis, and biological evaluation of compounds 7-33 to establish structure-activity relationships of the scaffold are described, leading to the identification of clinical development compound icenticaftor (QBW251) 33, which has subsequently progressed to deliver two positive clinical proofs of concept in patients with CF and COPD and is now being further developed as a novel therapeutic approach for COPD patients.
Asunto(s)
Aminopiridinas/química , Regulador de Conductancia de Transmembrana de Fibrosis Quística/metabolismo , Administración Oral , Aminopiridinas/metabolismo , Aminopiridinas/uso terapéutico , Animales , Fibrosis Quística/tratamiento farmacológico , Regulador de Conductancia de Transmembrana de Fibrosis Quística/antagonistas & inhibidores , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Modelos Animales de Enfermedad , Evaluación Preclínica de Medicamentos , Eliminación de Gen , Semivida , Humanos , Unión Proteica , Enfermedad Pulmonar Obstructiva Crónica/tratamiento farmacológico , Ratas , Ratas Sprague-Dawley , Solubilidad , Relación Estructura-ActividadRESUMEN
Spinal muscular atrophy (SMA) is a debilitating neuromuscular disease caused by low levels of functional survival motor neuron protein (SMN) resulting from a deletion or loss of function mutation of the survival motor neuron 1 (SMN1) gene. Branaplam (1) elevates levels of full-length SMN protein in vivo by modulating the splicing of the related gene SMN2 to enhance the exon-7 inclusion and increase levels of the SMN. The intramolecular hydrogen bond present in the 2-hydroxyphenyl pyridazine core of 1 enforces a planar conformation of the biaryl system and is critical for the compound activity. Scaffold morphing revealed that the pyridazine could be replaced by a 1,3,4-thiadiazole, which provided additional opportunities for a conformational constraint of the biaryl through intramolecular 1,5-sulfur-oxygen (S···O) or 1,5-sulfur-halogen (S···X) noncovalent interactions. Compound 26, which incorporates a 2-fluorophenyl thiadiazole motif, demonstrated a greater than 50% increase in production of full-length SMN protein in a mouse model of SMA.
Asunto(s)
Diseño de Fármacos , Empalme del ARN , Tiadiazoles/química , Animales , Semivida , Halógenos/química , Humanos , Masculino , Ratones , Conformación Molecular , Neuronas Motoras/metabolismo , Atrofia Muscular Espinal/genética , Atrofia Muscular Espinal/patología , Oxígeno/química , Piridazinas/química , Empalme del ARN/efectos de los fármacos , Ratas , Ratas Sprague-Dawley , Relación Estructura-Actividad , Azufre/química , Proteína 1 para la Supervivencia de la Neurona Motora/genética , Proteína 1 para la Supervivencia de la Neurona Motora/metabolismo , Proteína 2 para la Supervivencia de la Neurona Motora/genética , Proteína 2 para la Supervivencia de la Neurona Motora/metabolismo , Tiadiazoles/metabolismo , Tiadiazoles/farmacologíaRESUMEN
Spinal muscular atrophy (SMA), a rare neuromuscular disorder, is the leading genetic cause of death in infants and toddlers. SMA is caused by the deletion or a loss of function mutation of the survival motor neuron 1 (SMN1) gene. In humans, a second closely related gene SMN2 exists; however it codes for a less stable SMN protein. In recent years, significant progress has been made toward disease modifying treatments for SMA by modulating SMN2 pre-mRNA splicing. Herein, we describe the discovery of LMI070/branaplam, a small molecule that stabilizes the interaction between the spliceosome and SMN2 pre-mRNA. Branaplam (1) originated from a high-throughput phenotypic screening hit, pyridazine 2, and evolved via multiparameter lead optimization. In a severe mouse SMA model, branaplam treatment increased full-length SMN RNA and protein levels, and extended survival. Currently, branaplam is in clinical studies for SMA.
Asunto(s)
Encéfalo/efectos de los fármacos , Canal de Potasio ERG1/metabolismo , Atrofia Muscular Espinal/tratamiento farmacológico , Piridazinas/química , Administración Oral , Animales , Encéfalo/metabolismo , Línea Celular , Cristalografía por Rayos X , Relación Dosis-Respuesta a Droga , Evaluación Preclínica de Medicamentos/métodos , Canal de Potasio ERG1/antagonistas & inhibidores , Humanos , Ratones Endogámicos C57BL , Neuronas Motoras/efectos de los fármacos , Atrofia Muscular Espinal/genética , Piridazinas/farmacología , Relación Estructura-Actividad Cuantitativa , Empalme del ARN , Ratas Sprague-Dawley , Proteína 1 para la Supervivencia de la Neurona Motora/genética , Proteína 1 para la Supervivencia de la Neurona Motora/metabolismo , Proteína 2 para la Supervivencia de la Neurona Motora/genéticaRESUMEN
Spinal muscular atrophy (SMA), which results from the loss of expression of the survival of motor neuron-1 (SMN1) gene, represents the most common genetic cause of pediatric mortality. A duplicate copy (SMN2) is inefficiently spliced, producing a truncated and unstable protein. We describe herein a potent, orally active, small-molecule enhancer of SMN2 splicing that elevates full-length SMN protein and extends survival in a severe SMA mouse model. We demonstrate that the molecular mechanism of action is via stabilization of the transient double-strand RNA structure formed by the SMN2 pre-mRNA and U1 small nuclear ribonucleic protein (snRNP) complex. The binding affinity of U1 snRNP to the 5' splice site is increased in a sequence-selective manner, discrete from constitutive recognition. This new mechanism demonstrates the feasibility of small molecule-mediated, sequence-selective splice modulation and the potential for leveraging this strategy in other splicing diseases.
Asunto(s)
Empalme Alternativo , Atrofia Muscular Espinal/tratamiento farmacológico , ARN Bicatenario/agonistas , Ribonucleoproteína Nuclear Pequeña U1/agonistas , Bibliotecas de Moléculas Pequeñas/farmacología , Proteína 2 para la Supervivencia de la Neurona Motora/metabolismo , Animales , Sitios de Unión , Modelos Animales de Enfermedad , Femenino , Expresión Génica , Humanos , Ratones , Ratones Transgénicos , Modelos Moleculares , Atrofia Muscular Espinal/metabolismo , Atrofia Muscular Espinal/mortalidad , Atrofia Muscular Espinal/patología , Unión Proteica/efectos de los fármacos , Estabilidad Proteica/efectos de los fármacos , Proteolisis , Precursores del ARN/agonistas , Precursores del ARN/química , Precursores del ARN/metabolismo , ARN Bicatenario/química , ARN Bicatenario/metabolismo , Ribonucleoproteína Nuclear Pequeña U1/química , Ribonucleoproteína Nuclear Pequeña U1/metabolismo , Bibliotecas de Moléculas Pequeñas/síntesis química , Bibliotecas de Moléculas Pequeñas/metabolismo , Análisis de Supervivencia , Proteína 2 para la Supervivencia de la Neurona Motora/química , Proteína 2 para la Supervivencia de la Neurona Motora/genéticaRESUMEN
Toll-like receptor (TLR) signaling is a key component of innate immunity. Aberrant TLR activation leads to immune disorders via dysregulation of cytokine production, such as IL-12/IL-23. Herein, we identify and characterize PIKfyve, a lipid kinase, as a critical player in TLR signaling using apilimod as an affinity tool. Apilimod is a potent small molecular inhibitor of IL-12/IL-23 with an unknown target and has been evaluated in clinical trials for patients with Crohn's disease or rheumatoid arthritis. Using a chemical genetic approach, we show that it binds to PIKfyve and blocks its phosphotransferase activity, leading to selective inhibition of IL-12/IL-23p40. Pharmacological or genetic inactivation of PIKfyve is necessary and sufficient for suppression of IL-12/IL-23p40 expression. Thus, we have uncovered a phosphoinositide-mediated regulatory mechanism that controls TLR signaling.
Asunto(s)
Interleucina-12/antagonistas & inhibidores , Interleucina-23/antagonistas & inhibidores , Morfolinas/farmacología , Fosfatidilinositol 3-Quinasas/metabolismo , Inhibidores de las Quinasa Fosfoinosítidos-3 , Transducción de Señal/efectos de los fármacos , Receptores Toll-Like/metabolismo , Triazinas/farmacología , Animales , Línea Celular , Citocinas/metabolismo , Inhibidores Enzimáticos/metabolismo , Inhibidores Enzimáticos/farmacología , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Hidrazonas , Ratones , Morfolinas/metabolismo , Unión Proteica , Pirimidinas , Especificidad por Sustrato , Triazinas/metabolismoRESUMEN
Tankyrase 1 and 2 have been shown to be redundant, druggable nodes in the Wnt pathway. As such, there has been intense interest in developing agents suitable for modulating the Wnt pathway in vivo by targeting this enzyme pair. By utilizing a combination of structure-based design and LipE-based structure efficiency relationships, the core of XAV939 was optimized into a more stable, more efficient, but less potent dihydropyran motif 7. This core was combined with elements of screening hits 2, 19, and 33 and resulted in highly potent, selective tankyrase inhibitors that are novel three pocket binders. NVP-TNKS656 (43) was identified as an orally active antagonist of Wnt pathway activity in the MMTV-Wnt1 mouse xenograft model. With an enthalpy-driven thermodynamic signature of binding, highly favorable physicochemical properties, and high lipophilic efficiency, NVP-TNKS656 is a novel tankyrase inhibitor that is well suited for further in vivo validation studies.
Asunto(s)
Acetamidas/farmacología , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Pirimidinonas/farmacología , Tanquirasas/antagonistas & inhibidores , Acetamidas/administración & dosificación , Acetamidas/química , Administración Oral , Animales , Área Bajo la Curva , Disponibilidad Biológica , Inhibidores Enzimáticos/administración & dosificación , Ratones , Modelos Moleculares , Pirimidinonas/administración & dosificación , Pirimidinonas/química , Relación Estructura-ActividadRESUMEN
The crystal structures of tankyrase 1 (TNKS1) in complex with two small-molecule inhibitors, PJ34 and XAV939, both at 2.0 Å resolution, are reported. The structure of TNKS1 in complex with PJ34 reveals two molecules of PJ34 bound in the NAD(+) donor pocket. One molecule is in the nicotinamide portion of the pocket, as previously observed in other PARP structures, while the second molecule is bound in the adenosine portion of the pocket. Additionally, unlike the unliganded crystallization system, the TNKS1-PJ34 crystallization system has the NAD(+) donor site accessible to bulk solvent in the crystal, which allows displacement soaking. The TNKS1-PJ34 crystallization system was used to determine the structure of TNKS1 in complex with XAV939. These structures provide a basis for the start of a structure-based drug-design campaign for TNKS1.
Asunto(s)
Inhibidores Enzimáticos/química , Compuestos Heterocíclicos con 3 Anillos/química , Fenantrenos/química , Tanquirasas/química , Cristalografía por Rayos X , Humanos , Modelos Moleculares , Dominios y Motivos de Interacción de Proteínas , Tanquirasas/antagonistas & inhibidoresRESUMEN
The Wnt signaling pathway is critical to the regulation of key cellular processes. When deregulated, it has been shown to play a crucial role in the growth and progression of multiple human cancers. The identification of small molecule modulators of Wnt signaling has proven challenging, largely due to the relative paucity of druggable nodes in this pathway. Several recent publications have identified small molecule inhibitors of the Wnt pathway, and tankyrase (TNKS) inhibition has been demonstrated to antagonize Wnt signaling via axin stabilization. Herein, we report the early hit assessment of a series of compounds previously reported to antagonize Wnt signaling. We report the biophysical, computational characterization, structure-activity relationship, and physicochemical properties of a novel series of [1,2,4]triazol-3-ylsulfanylmethyl)-3-phenyl-[1,2,4]oxadiazole inhibitors of TNKS1 and 2. Furthermore, a cocrystal structure of compound 24 complexed to TNKS1 demonstrates an alternate binding mode for PARP family member proteins that does not involve interactions with the nicotinamide binding pocket.
Asunto(s)
Adenosina/metabolismo , Modelos Moleculares , Oxadiazoles/síntesis química , Sulfuros/síntesis química , Tanquirasas/antagonistas & inhibidores , Triazoles/síntesis química , Vía de Señalización Wnt/efectos de los fármacos , Adenosina/química , Sitios de Unión , Cristalografía por Rayos X , Células HEK293 , Humanos , Oxadiazoles/química , Oxadiazoles/farmacología , Conformación Proteica , Relación Estructura-Actividad , Sulfuros/química , Sulfuros/farmacología , Triazoles/química , Triazoles/farmacologíaRESUMEN
Identification of novel, validated targets remains a top priority in modern drug discovery. Chemical genetics represents a powerful approach to the discovery of new targets. Unlike the traditional target-based screen that relies on a predefined, sometimes poorly validated target, a chemical genetics-based phenotypic screen probes the entire molecular signaling pathway in an efficient and unbiased manner for the most drug-sensitive node. The most significant obstacle associated with this approach is identification of the efficacy targets of small-molecule probes. The huge potential of chemical genetics cannot be realized without the establishment of reliable mechanisms for target identification. In this article, we describe each essential element of the chemical genetics process, discuss common challenges that the field is facing, and critically review various biochemical and genetics approaches recently developed for target deconvolution. We also attempt to summarize lessons that we have collectively learned and provide a practical perspective to facilitate the advancement of chemical genetics.
Asunto(s)
Sistemas de Liberación de Medicamentos , Descubrimiento de Drogas/métodos , Perfilación de la Expresión Génica/métodos , Farmacogenética/métodos , Animales , Biología Computacional/métodos , Diseño de Fármacos , Humanos , Fenotipo , Proteómica/métodos , Transducción de Señal , Transcripción GenéticaRESUMEN
The application of chemical proteomics to new target discovery can lead to a rapid understanding of disease mechanism and new therapeutic methods. Successful application includes a thorough understanding of SAR and the validation of target relevance using multiple genetic and biochemical methods. This feature review highlights several successful applications of chemical proteomics and outlines the strategy and approaches that lead to the discovery of novel therapeutic targets.
Asunto(s)
Descubrimiento de Drogas , Terapia Molecular Dirigida , Farmacología/métodos , Proteómica/métodos , Animales , Evaluación Preclínica de Medicamentos/métodos , Industria Farmacéutica/métodos , Humanos , Transducción de Señal/efectos de los fármacos , Relación Estructura-ActividadRESUMEN
The stability of the Wnt pathway transcription factor beta-catenin is tightly regulated by the multi-subunit destruction complex. Deregulated Wnt pathway activity has been implicated in many cancers, making this pathway an attractive target for anticancer therapies. However, the development of targeted Wnt pathway inhibitors has been hampered by the limited number of pathway components that are amenable to small molecule inhibition. Here, we used a chemical genetic screen to identify a small molecule, XAV939, which selectively inhibits beta-catenin-mediated transcription. XAV939 stimulates beta-catenin degradation by stabilizing axin, the concentration-limiting component of the destruction complex. Using a quantitative chemical proteomic approach, we discovered that XAV939 stabilizes axin by inhibiting the poly-ADP-ribosylating enzymes tankyrase 1 and tankyrase 2. Both tankyrase isoforms interact with a highly conserved domain of axin and stimulate its degradation through the ubiquitin-proteasome pathway. Thus, our study provides new mechanistic insights into the regulation of axin protein homeostasis and presents new avenues for targeted Wnt pathway therapies.
Asunto(s)
Proteínas Represoras/metabolismo , Transducción de Señal/efectos de los fármacos , Tanquirasas/antagonistas & inhibidores , Proteínas Wnt/antagonistas & inhibidores , Proteína Axina , División Celular/efectos de los fármacos , Línea Celular , Línea Celular Tumoral , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/metabolismo , Compuestos Heterocíclicos con 3 Anillos/farmacología , Humanos , Complejo de la Endopetidasa Proteasomal/metabolismo , Unión Proteica , Proteómica , Proteínas Represoras/química , Tanquirasas/metabolismo , Transcripción Genética/efectos de los fármacos , Ubiquitina/metabolismo , Ubiquitinación , Proteínas Wnt/metabolismo , beta Catenina/antagonistas & inhibidores , beta Catenina/metabolismoRESUMEN
A new approach to (+)-cacospongionolide was developed to access conformationally restricted variants of the natural product. The flexible aliphatic region between the decalin and side chain portion of the natural product was replaced with alkenyl and alkynyl linkers to probe the influence of structural rigidity in the inhibition of secretary phospholipase A2 (sPLA2). It was found that when the aliphatic section is replaced with a Z-olefin or an alkyne, sPLA2 inhibitory activity suffered relative to the natural product; however, an E-olefin-containing analogue led to an enhanced activity. These results suggest that preferred sPLA2 binding conformation of the natural product is similar to the geometry of the E-olefin-containing analogue.
Asunto(s)
4-Butirolactona/análogos & derivados , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/síntesis química , Fosfolipasas A/antagonistas & inhibidores , Poríferos/química , Piranos/química , Piranos/síntesis química , 4-Butirolactona/síntesis química , 4-Butirolactona/química , 4-Butirolactona/farmacología , Animales , Venenos de Abeja/enzimología , Inhibidores Enzimáticos/farmacología , Furaldehído/química , Humanos , Indicadores y Reactivos , Espectroscopía de Resonancia Magnética , Modelos Moleculares , Conformación Molecular , Fosfolipasas A2 , Piranos/farmacología , Proteínas Recombinantes/químicaRESUMEN
The total syntheses of the antiinflammatory marine sponge metabolites (+)-cacospongionolide B and E are described. The pivotal steps in the synthetic route include a three-step sequence that couples the two main regions of the natural product, as well as generates the side chain dihydropyran ring. The activity of the synthetic analogues against bee venom phospholipase A2 suggests that the cacospongionolides have enantiospecific interactions with the enzyme that may be independent of the gamma-hydroxybutenolide moiety.
Asunto(s)
4-Butirolactona/análogos & derivados , 4-Butirolactona/síntesis química , Inhibidores Enzimáticos/síntesis química , Fosfolipasas A/antagonistas & inhibidores , Piranos/síntesis química , 4-Butirolactona/farmacología , Animales , Inhibidores Enzimáticos/farmacología , Estructura Molecular , Fosfolipasas A2 , Poríferos/química , Piranos/farmacología , Estereoisomerismo , Relación Estructura-ActividadRESUMEN
The first total synthesis of the antiinflammatory marine sponge metabolite (+)-cacospongionolide B has been accomplished in 12 linear steps. The pivotal transformations include a three-step sequence coupling the two main regions of the natural product as well as generating the side chain dihydropyran ring. The activity of the synthetic analogues against bee venom phospholipase A(2) suggests that cacospongionolide B has an enantiospecific interaction with the enzyme that is independent of the gamma-hydroxybutenolide moiety.