RESUMEN
Protease inhibitors (PIs) remain an important component of antiretroviral therapy for the treatment of HIV-1 infection due to their high genetic barrier to resistance development. Nevertheless, the two most commonly prescribed HIV PIs, atazanavir and darunavir, still require co-administration with a pharmacokinetic boosting agent to maintain sufficient drug plasma levels which can lead to undesirable drug-drug interactions. Herein, we describe GS-9770, a novel investigational non-peptidomimetic HIV PI with unboosted once-daily oral dosing potential due to improvements in its metabolic stability and its pharmacokinetic properties in preclinical animal species. This compound demonstrates potent inhibitory activity and high on-target selectivity for recombinant HIV-1 protease versus other aspartic proteases tested. In cell culture, GS-9770 inhibits Gag polyprotein cleavage and shows nanomolar anti-HIV-1 potency in primary human cells permissive to HIV-1 infection and against a broad range of HIV subtypes. GS-9770 demonstrates an improved resistance profile against a panel of patient-derived HIV-1 isolates with resistance to atazanavir and darunavir. In resistance selection experiments, GS-9770 prevented the emergence of breakthrough HIV-1 variants at all fixed drug concentrations tested and required multiple protease substitutions to enable outgrowth of virus exposed to escalating concentrations of GS-9770. This compound also remained fully active against viruses resistant to drugs from other antiviral classes and showed no in vitro antagonism when combined pairwise with drugs from other antiretroviral classes. Collectively, these preclinical data identify GS-9770 as a potent, non-peptidomimetic once-daily oral HIV PI with potential to overcome the persistent requirement for pharmacological boosting with this class of antiretroviral agents.
Asunto(s)
Infecciones por VIH , Inhibidores de la Proteasa del VIH , VIH-1 , Humanos , Inhibidores de la Proteasa del VIH/farmacología , Inhibidores de la Proteasa del VIH/uso terapéutico , Darunavir/farmacología , Darunavir/uso terapéutico , Sulfato de Atazanavir/farmacología , Sulfato de Atazanavir/uso terapéutico , Farmacorresistencia Viral , VIH-1/genética , Antirretrovirales/uso terapéutico , Infecciones por VIH/tratamiento farmacológico , Proteasa del VIH/genética , Proteasa del VIH/metabolismoRESUMEN
The bromodomain and extra-terminal (BET) family of proteins, consisting of the bromodomains containing protein 2 (BRD2), BRD3, BRD4, and the testis-specific BRDT, are key epigenetic regulators of gene transcription and has emerged as an attractive target for anticancer therapy. Herein, we describe the discovery of a novel potent BET bromodomain inhibitor, using a systematic structure-based approach focused on improving potency, metabolic stability, and permeability. The optimized dimethylisoxazole aryl-benzimidazole inhibitor exhibited high potency towards BRD4 and related BET proteins in biochemical and cell-based assays and inhibited tumor growth in two proof-of-concept preclinical animal models.
Asunto(s)
Bencimidazoles/farmacología , Descubrimiento de Drogas , Isoxazoles/farmacología , Mieloma Múltiple/tratamiento farmacológico , Factores de Transcripción/antagonistas & inhibidores , Administración Oral , Animales , Bencimidazoles/química , Bencimidazoles/metabolismo , Disponibilidad Biológica , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Humanos , Isoxazoles/administración & dosificación , Isoxazoles/química , Isoxazoles/metabolismo , Ratones , Estructura Molecular , Mieloma Múltiple/metabolismo , Mieloma Múltiple/patología , Neoplasias Experimentales/tratamiento farmacológico , Neoplasias Experimentales/metabolismo , Neoplasias Experimentales/patología , Dominios Proteicos/efectos de los fármacos , Relación Estructura-Actividad , Factores de Transcripción/metabolismoRESUMEN
tert-Butoxy-(4-phenyl-quinolin-3-yl)-acetic acids (tBPQA) are a new class of HIV-1 integrase (IN) inhibitors that are structurally distinct from IN strand transfer inhibitors but analogous to LEDGINs. LEDGINs are a class of potent antiviral compounds that interacts with the lens epithelium-derived growth factor (LEDGF) binding pocket on IN and were identified through competition binding against LEDGF. LEDGF tethers IN to the host chromatin and enables targeted integration of viral DNA. The prevailing understanding of the antiviral mechanism of LEDGINs is that they inhibit LEDGF binding to IN, which prevents targeted integration of HIV-1. We showed that in addition to the properties already known for LEDGINs, the binding of tBPQAs to the IN dimer interface inhibits IN enzymatic activity in a LEDGF-independent manner. Using the analysis of two long terminal repeat junctions in HIV-infected cells, we showed that the inhibition by tBPQAs occurs at or prior to the viral DNA 3'-processing step. Biochemical studies revealed that this inhibition operates by compound-induced conformational changes in the IN dimer that prevent proper assembly of IN onto viral DNA. For the first time, tBPQAs were demonstrated to be allosteric inhibitors of HIV-1 IN displaying a dual mode of action: inhibition of IN-viral DNA assembly and inhibition of IN-LEDGF interaction.
Asunto(s)
Acetatos/farmacología , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Cromatina/metabolismo , Inhibidores de Integrasa VIH/farmacología , Integrasa de VIH/metabolismo , VIH-1/enzimología , Quinolinas/farmacología , Factores de Transcripción/metabolismo , Integración Viral/efectos de los fármacos , Acetatos/química , Proteínas Adaptadoras Transductoras de Señales/genética , Línea Celular , Cromatina/genética , ADN Viral/genética , ADN Viral/metabolismo , Infecciones por VIH/tratamiento farmacológico , Infecciones por VIH/enzimología , Infecciones por VIH/genética , Integrasa de VIH/química , Integrasa de VIH/genética , Inhibidores de Integrasa VIH/química , VIH-1/genética , Humanos , Quinolinas/química , Factores de Transcripción/genética , Integración Viral/fisiologíaRESUMEN
A series of nevirapine-based analogues containing the phosphonate functionality were prepared and evaluated in vitro against HIV RT. The effect of the phosphonate was evaluated against the wild type and Y181C HIV replication. An in vivo PK study was performed on a select analogue.
Asunto(s)
Infecciones por VIH/tratamiento farmacológico , Nevirapina/análogos & derivados , Nevirapina/farmacología , Organofosfonatos/síntesis química , Organofosfonatos/farmacología , Inhibidores de la Transcriptasa Inversa/síntesis química , Inhibidores de la Transcriptasa Inversa/farmacología , VIH-1/efectos de los fármacos , Humanos , Organofosfonatos/química , Inhibidores de la Transcriptasa Inversa/química , Relación Estructura-ActividadRESUMEN
Non-nucleoside reverse transcriptase inhibitors (NNRTIs) represent cornerstones of current regimens for treatment of human immunodeficiency virus type 1 (HIV-1) infections. However, NNRTIs usually suffer from low aqueous solubility and the emergence of resistant viral strains. In the present work, novel bicyclic NNRTIs derived from etravirine (ETV) and rilpivirine (RPV), bearing modified purine, tetrahydropteridine, and pyrimidodiazepine cores, were designed and prepared. Compounds 2, 4, and 6 carrying the acrylonitrile moiety displayed single-digit nanomolar activities against the wild-type (WT) virus (EC50 = 2.5, 2.7, and 3.0 nM, respectively), where the low nanomolar activity was retained against HXB2 (EC50 = 2.2-2.8 nM) and the K103N and Y181C mutated strains (fold change, 1.2-6.7×). Most importantly, compound 2 exhibited significantly improved phosphate-buffered saline solubility (10.4 µM) compared to ETV and RPV (âª1 µM). Additionally, the binding modes of compounds 2, 4, and 6 to the reverse transcriptase were studied by X-ray crystallography.
Asunto(s)
Fármacos Anti-VIH , Infecciones por VIH , VIH-1 , Humanos , Fármacos Anti-VIH/química , VIH-1/metabolismo , Inhibidores de la Transcriptasa Inversa , Transcriptasa Inversa del VIH/metabolismo , Infecciones por VIH/tratamiento farmacológico , Rilpivirina/uso terapéutico , Diseño de FármacosRESUMEN
GS-9148 is an investigational phosphonate nucleotide analogue inhibitor of reverse transcriptase (RT) (NtRTI) of human immunodeficiency virus type 1 (HIV-1). This compound is an adenosine derivative with a 2',3'-dihydrofuran ring structure that contains a 2'-fluoro group. The resistance profile of GS-9148 is unique in that the inhibitor can select for the very rare Q151L mutation in HIV-1 RT as a pathway to resistance. Q151L is not stably selected by any of the approved nucleoside or nucleotide analogues; however, it may be a transient intermediate that leads to the related Q151M mutation, which confers resistance to multiple compounds that belong to this class of RT inhibitors. Here, we employed pre-steady-state kinetics to study the impact of Q151L on substrate and inhibitor binding and the catalytic rate of incorporation. Most importantly, we found that the Q151L mutant is unable to incorporate GS-9148 under single-turnover conditions. Interference experiments showed that the presence of GS-9148-diphosphate, i.e., the active form of the inhibitor, does not reduce the efficiency of incorporation for the natural counterpart. We therefore conclude that Q151L severely compromises binding of GS-9148-diphosphate to RT. This effect is highly specific, since we also demonstrate that another NtRTI, tenofovir, is incorporated with selectivity similar to that seen with wild-type RT. Incorporation assays with other related compounds and models based on the RT/DNA/GS-9148-diphosphate crystal structure suggest that the 2'-fluoro group of GS-9148 may cause steric hindrance with the side chain of the Q151L mutant.
Asunto(s)
Farmacorresistencia Viral , Guanosina/análogos & derivados , Transcriptasa Inversa del VIH/genética , VIH-1/efectos de los fármacos , Mutación , Inhibidores de la Transcriptasa Inversa/farmacología , Secuencia de Bases , Guanosina/farmacología , VIH-1/enzimología , Modelos Moleculares , Datos de Secuencia Molecular , Relación Estructura-ActividadRESUMEN
HIV-1 RNase H breaks down the intermediate RNA-DNA hybrids during reverse transcription, requiring two divalent metal ions for activity. Pyrimidinol carboxylic acid and N-hydroxy quinazolinedione inhibitors were designed to coordinate the two metal ions in the active site of RNase H. High-resolution (1.4 Å to 2.1 Å) crystal structures were determined with the isolated RNase H domain and reverse transcriptase (RT), which permit accurate assessment of the metal and water environment at the active site. The geometry of the metal coordination suggests that the inhibitors mimic a substrate state prior to phosphodiester catalysis. Surface plasmon resonance studies confirm metal-dependent binding to RNase H and demonstrate that the inhibitors do not bind at the polymerase active site of RT. Additional evaluation of the RNase H site reveals an open protein surface with few additional interactions to optimize active-site inhibitors.
Asunto(s)
Fármacos Anti-VIH/farmacología , VIH-1/efectos de los fármacos , Pirimidinas/farmacología , Quinazolinonas/farmacología , Ribonucleasa H/antagonistas & inhibidores , Secuencia de Aminoácidos , Cristalización , Transcriptasa Inversa del VIH/antagonistas & inhibidores , Transcriptasa Inversa del VIH/química , VIH-1/enzimología , Conformación Molecular , Datos de Secuencia Molecular , Pirimidinas/química , Quinazolinonas/química , Ribonucleasa H/química , Relación Estructura-ActividadRESUMEN
The interaction between lens epithelium-derived growth factor/transcriptional co-activator p75 (LEDGF) and human immunodeficiency virus type 1 (HIV-1) integrase (IN) is essential for HIV-1 replication. Homogeneous time-resolved fluorescence resonance energy transfer assays were developed to characterize HIV-1 integrase dimerization and the interaction between LEDGF and IN dimers. Using these assays in an equilibrium end point dose-response format with mathematical modeling, we determined the dissociation constants of IN dimers (K(dimer) = 67.8 pm) and of LEDGF from IN dimers (K(d) = 10.9 nm). When used in a kinetic format, the assays allowed the determination of the on- and off-rate constants for these same interactions. Integrase dimerization had a k(on) of 0.1247 nm(-1) x min(-1) and a k(off) of 0.0080 min(-1) resulting in a K(dimer) of 64.5 pm. LEDGF binding to IN dimers had a k(on) of 0.0285 nm(-1).min(-1) and a k(off) of 0.2340 min(-1) resulting in a K(d) of 8.2 nm. These binding assays can also be used in an equilibrium end point competition format. In this format, the IN catalytic core domain produced a K(i) of 15.2 nm while competing for integrase dimerization, confirming the very tight interaction of IN with itself. In the same format, LEDGF produced a K(i) value of 35 nm when competing for LEDGF binding to IN dimers. In summary, this study describes a methodology combining homogeneous time-resolved fluorescence resonance energy transfer and mathematical modeling to derive the affinities between IN monomers and between LEDGF and IN dimers. This study revealed the significantly tighter nature of the IN-IN dimer compared with the IN-LEDGF interaction.
Asunto(s)
Integrasa de VIH/química , Integrasa de VIH/metabolismo , Péptidos y Proteínas de Señalización Intercelular/química , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Algoritmos , Secuencia de Aminoácidos , Unión Competitiva , Transferencia Resonante de Energía de Fluorescencia , Integrasa de VIH/genética , Humanos , Péptidos y Proteínas de Señalización Intercelular/genética , Cinética , Modelos Biológicos , Modelos Químicos , Datos de Secuencia Molecular , Unión Proteica , Dominios y Motivos de Interacción de Proteínas , Mapeo de Interacción de Proteínas , Multimerización de ProteínaRESUMEN
A series of N1-heterocyclic pyrimidinediones were extensively evaluated as HIV-1 non-nucleoside reverse transcriptase inhibitors (NNRTIs). Inhibitor 1 is active against NNRTI-resistant viruses including RT mutant K103N. The co-crystal structure of inhibitor 1 with HIV-1 RT revealed that H-bonds are formed with K101 and K103. Efforts to improve the suboptimal pharmacokinetic profile of 1 resulted in the discovery of compound 13, which represents the lead compound in this series with improved pharmacokinetics and similar potency as inhibitor 1.
Asunto(s)
Fármacos Anti-VIH/química , Transcriptasa Inversa del VIH/antagonistas & inhibidores , Compuestos Heterocíclicos/química , Pirimidinonas/química , Inhibidores de la Transcriptasa Inversa/química , Timina/análogos & derivados , Animales , Fármacos Anti-VIH/síntesis química , Fármacos Anti-VIH/farmacocinética , Sitios de Unión , Cristalografía por Rayos X , Perros , Transcriptasa Inversa del VIH/metabolismo , Humanos , Enlace de Hidrógeno , Microsomas/metabolismo , Proteínas Mutantes/antagonistas & inhibidores , Proteínas Mutantes/metabolismo , Pirimidinonas/síntesis química , Pirimidinonas/farmacocinética , Inhibidores de la Transcriptasa Inversa/síntesis química , Inhibidores de la Transcriptasa Inversa/farmacocinética , Relación Estructura-Actividad , Timina/síntesis química , Timina/química , Timina/farmacocinéticaRESUMEN
A series of N1-alkyl pyrimidinediones were designed, synthesized and evaluated as HIV-1 non-nucleoside reverse transcriptase inhibitors (NNRTIs). Our efforts identified compound 10b, which represents the lead compound in this series with pharmacokinetics and antiviral potency that may support once-daily dosing.
Asunto(s)
Fármacos Anti-VIH/química , Transcriptasa Inversa del VIH/química , Pirimidinonas/química , Inhibidores de la Transcriptasa Inversa/química , Timina/análogos & derivados , Animales , Fármacos Anti-VIH/síntesis química , Fármacos Anti-VIH/farmacocinética , Sitios de Unión , Cristalografía por Rayos X , Perros , Transcriptasa Inversa del VIH/metabolismo , Humanos , Microsomas/metabolismo , Pirimidinonas/síntesis química , Pirimidinonas/farmacocinética , Inhibidores de la Transcriptasa Inversa/síntesis química , Inhibidores de la Transcriptasa Inversa/farmacocinética , Relación Estructura-Actividad , Timina/síntesis química , Timina/química , Timina/farmacocinéticaRESUMEN
Spleen tyrosine kinase (SYK) is a critical regulator of signaling in a variety of immune cell types such as B-cells, monocytes, and macrophages. Accordingly, there have been numerous efforts to identify compounds that selectively inhibit SYK as a means to treat autoimmune and inflammatory diseases. We previously disclosed GS-9973 (entospletinib) as a selective SYK inhibitor that is under clinical evaluation in hematological malignancies. However, a BID dosing regimen and drug interaction with proton pump inhibitors (PPI) prevented development of entospletinib in inflammatory diseases. Herein, we report the discovery of a second-generation SYK inhibitor, GS-9876 (lanraplenib), which has human pharmacokinetic properties suitable for once-daily administration and is devoid of any interactions with PPI. Lanraplenib is currently under clinical evaluation in multiple autoimmune indications.
RESUMEN
We describe the discovery of three structurally differentiated potent and selective MTH1 inhibitors and their subsequent use to investigate MTH1 as an oncology target, culminating in target (in)validation. Tetrahydronaphthyridine 5 was rapidly identified as a highly potent MTH1 inhibitor (IC50 = 0.043 nM). Cocrystallization of 5 with MTH1 revealed the ligand in a Φ-cis-N-(pyridin-2-yl)acetamide conformation enabling a key intramolecular hydrogen bond and polar interactions with residues Gly34 and Asp120. Modification of literature compound TH287 with O- and N-linked aryl and alkyl aryl substituents led to the discovery of potent pyrimidine-2,4,6-triamine 25 (IC50 = 0.49 nM). Triazolopyridine 32 emerged as a highly selective lead compound with a suitable in vitro profile and desirable pharmacokinetic properties in rat. Elucidation of the DNA damage response, cell viability, and intracellular concentrations of oxo-NTPs (oxidized nucleoside triphosphates) as a function of MTH1 knockdown and/or small molecule inhibition was studied. Based on our findings, we were unable to provide evidence to further pursue MTH1 as an oncology target.
RESUMEN
BACKGROUND: Antiretroviral therapy that targets HIV type-1 (HIV-1) reverse transcriptase (RT) can be linked to mutations in the thumb-connection (amino acids [AA] 241-426) and RNase H (AA 427-560) domains, which could affect drug resistance. METHODS: Genotypical and statistical analyses were performed on HIV-1 RT from 100 antiretroviral treatment-naive and 248 antiretroviral treatment-experienced patients, the majority of whom were infected with HIV-1 subtype B. The RT region was analysed in three parts: the polymerase (AA 1-240), thumb-connection (AA 241-426) and RNase H (AA 427-560) domains. RESULTS: The polymerase domain had statistically significant changes between the two groups at 24 AA positions that are known resistance sites. Within the thumb-connection domain, R284 and N348 had statistically significant changes between the groups (P=0.007 and P< or =0.001, respectively). In treatment-experienced patients, 17.3% had R284K, whereas 24.5% had N348I substitutions. Both R284 and N348 were 100% conserved in treatment-naive patients. Within the RNase H domain, only K451 showed a statistically significant change (P=0.001), with K451R present in 11% of treatment-experienced patients but remaining 100% conserved among treatment-naive patients. CONCLUSIONS: RT mutations at three positions outside of the polymerase region were associated with antiretroviral therapy: R284K, N348I and K451R. Both R284K and K451R interact with the phosphate backbone of the template or primer in HIV-1 RT crystal structures and could potentially influence the positioning of the primer strand, thus affecting polymerization, the efficiency of nucleoside reverse transcriptase inhibitor excision and/or RNase H activity.
Asunto(s)
Transcriptasa Inversa del VIH/genética , VIH-1 , Mutación/efectos de los fármacos , Inhibidores de la Transcriptasa Inversa/uso terapéutico , Adenina/análogos & derivados , Adenina/uso terapéutico , Alquinos , Sustitución de Aminoácidos/efectos de los fármacos , Benzoxazinas/uso terapéutico , Ciclopropanos , Farmacorresistencia Viral Múltiple , Variación Genética/efectos de los fármacos , Infecciones por VIH/tratamiento farmacológico , Infecciones por VIH/virología , Transcriptasa Inversa del VIH/química , VIH-1/efectos de los fármacos , VIH-1/genética , Humanos , Lamivudine/uso terapéutico , Organofosfonatos/uso terapéutico , Estructura Terciaria de Proteína/genética , Ribonucleasa H del Virus de la Inmunodeficiencia Humana/genética , Estavudina/uso terapéutico , TenofovirRESUMEN
Emtricitabine (FTC) and lamivudine (3TC), containing an oxathiolane ring with unnatural (-)-stereochemistry, are widely used nucleoside reverse transcriptase inhibitors (NRTIs) in anti-HIV therapy. Treatment with FTC or 3TC primarily selects for the HIV-1 RT M184V/I resistance mutations. Here we provide a comprehensive kinetic and structural basis for inhibiting HIV-1 RT by (-)-FTC-TP and (-)-3TC-TP and drug resistance by M184V. (-)-FTC-TP and (-)-3TC-TP have higher binding affinities (1/Kd) for wild-type RT but slower incorporation rates than dCTP. HIV-1 RT ternary crystal structures with (-)-FTC-TP and (-)-3TC-TP corroborate kinetic results demonstrating that their oxathiolane sulfur orients toward the DNA primer 3'-terminus and their triphosphate exists in two different binding conformations. M184V RT displays greater (>200-fold) Kd for the L-nucleotides and moderately higher (>9-fold) Kd for the D-isomers compared to dCTP. The M184V RT structure illustrates how the mutation repositions the oxathiolane of (-)-FTC-TP and shifts its triphosphate into a non-productive conformation.
Asunto(s)
Farmacorresistencia Viral , Transcriptasa Inversa del VIH/química , Transcriptasa Inversa del VIH/genética , VIH-1/efectos de los fármacos , VIH-1/genética , Nucleótidos/química , Inhibidores de la Transcriptasa Inversa/química , Alelos , Sustitución de Aminoácidos , Bases de Datos Genéticas , Infecciones por VIH/tratamiento farmacológico , Infecciones por VIH/virología , Transcriptasa Inversa del VIH/antagonistas & inhibidores , VIH-1/enzimología , Humanos , Pruebas de Sensibilidad Microbiana , Mutación , Nucleótidos/farmacología , Inhibidores de la Transcriptasa Inversa/farmacologíaRESUMEN
The thermophilic chemolithotroph, Aquifex aeolicus, expresses a gene product that exhibits both ATP sulfurylase and adenosine-5'-phosphosulfate (APS) kinase activities. These enzymes are usually segregated on two separate proteins in most bacteria, fungi, and plants. The domain arrangement in the Aquifex enzyme is reminiscent of the fungal ATP sulfurylase, which contains a C-terminal domain that is homologous to APS kinase yet displays no kinase activity. Rather, in the fungal enzyme, the motif serves as a sulfurylase regulatory domain that binds the allosteric effector 3'-phosphoadenosine-5'-phosphosulfate (PAPS), the product of true APS kinase. Therefore, the Aquifex enzyme may represent an ancestral homolog of a primitive bifunctional enzyme, from which the fungal ATP sulfurylase may have evolved. In heterotrophic sulfur-assimilating organisms such as fungi, ATP sulfurylase catalyzes the first committed step in sulfate assimilation to produce APS, which is subsequently metabolized to generate all sulfur-containing biomolecules. In contrast, ATP sulfurylase in sulfur chemolithotrophs catalyzes the reverse reaction to produce ATP and sulfate from APS and pyrophosphate. Here, the 2.3 A resolution X-ray crystal structure of Aquifex ATP sulfurylase-APS kinase bifunctional enzyme is presented. The protein dimerizes through its APS kinase domain and contains ADP bound in all four active sites. Comparison of the Aquifex ATP sulfurylase active site with those from sulfate assimilators reveals similar dispositions of the bound nucleotide and nearby residues. This suggests that minor perturbations are responsible for optimizing the kinetic properties for the physiologically relevant direction. The APS kinase active-site lid adopts two distinct conformations, where one conformation is distorted by crystal contacts. Additionally, a disulfide bond is observed in one ATP-binding P-loop of the APS kinase active site. This linkage accounts for the low kinase activity of the enzyme under oxidizing conditions. The thermal stability of the Aquifex enzyme can be explained by the 43% decreased cavity volume found within the protein core.
Asunto(s)
Bacterias Gramnegativas Quimiolitotróficas/enzimología , Fosfotransferasas (Aceptor de Grupo Alcohol)/química , Sulfato Adenililtransferasa/química , Sitios de Unión , Cristalografía por Rayos X , Estabilidad de Enzimas , Calor , Ligandos , Estructura Cuaternaria de Proteína , Estructura Secundaria de Proteína , Estructura Terciaria de Proteína , Subunidades de Proteína/química , Homología Estructural de ProteínaRESUMEN
A series of 3,4,5-trisubstituted 1,2,4-4H triazole derivatives was synthesized and investigated for HIV-1 reverse transcriptase inhibition. An X-ray structure with HIV-1 RT secured the binding mode and allowed the key interactions with the enzyme to be identified.
Asunto(s)
Fármacos Anti-VIH/síntesis química , Fármacos Anti-VIH/farmacología , Transcriptasa Inversa del VIH/antagonistas & inhibidores , VIH-1/efectos de los fármacos , Inhibidores de la Transcriptasa Inversa/síntesis química , Inhibidores de la Transcriptasa Inversa/farmacología , Triazoles/síntesis química , Triazoles/farmacología , Fármacos Anti-VIH/química , Técnicas Químicas Combinatorias , Cristalografía por Rayos X , Conformación Molecular , Estructura Molecular , Inhibidores de la Transcriptasa Inversa/química , Relación Estructura-Actividad , Triazoles/químicaRESUMEN
Oxidative stress is an underlying component of acute and chronic kidney disease. Apoptosis signal-regulating kinase 1 (ASK1) is a widely expressed redox-sensitive serine threonine kinase that activates p38 and c-Jun N-terminal kinase (JNK) mitogen-activated protein kinase kinases, and induces apoptotic, inflammatory, and fibrotic signaling in settings of oxidative stress. We describe the discovery and characterization of a potent and selective small-molecule inhibitor of ASK1, GS-444217, and demonstrate the therapeutic potential of ASK1 inhibition to reduce kidney injury and fibrosis. Activation of the ASK1 pathway in glomerular and tubular compartments was confirmed in renal biopsies from patients with diabetic kidney disease (DKD) and was decreased by GS-444217 in several rodent models of kidney injury and fibrosis that collectively represented the hallmarks of DKD pathology. Treatment with GS-444217 reduced progressive inflammation and fibrosis in the kidney and halted glomerular filtration rate decline. Combination of GS-444217 with enalapril, an angiotensin-converting enzyme inhibitor, led to a greater reduction in proteinuria and regression of glomerulosclerosis. These results identify ASK1 as an important target for renal disease and support the clinical development of an ASK1 inhibitor for the treatment of DKD.
Asunto(s)
Nefropatías Diabéticas/enzimología , Fibroblastos/enzimología , Glomérulos Renales/enzimología , MAP Quinasa Quinasa Quinasa 5/metabolismo , Sistema de Señalización de MAP Quinasas , Animales , Nefropatías Diabéticas/tratamiento farmacológico , Nefropatías Diabéticas/genética , Nefropatías Diabéticas/patología , Modelos Animales de Enfermedad , Femenino , Fibroblastos/patología , Fibrosis , Humanos , Glomérulos Renales/patología , MAP Quinasa Quinasa Quinasa 5/antagonistas & inhibidores , MAP Quinasa Quinasa Quinasa 5/genética , Masculino , Ratones , Ratones Noqueados , Inhibidores de Proteínas Quinasas/farmacología , Distribución Aleatoria , Ratas Sprague-DawleyRESUMEN
To elucidate the structure-geometry-activity relationship in diarylpyrimidine family (DAPYs) containing carbonyl linker between the central pyrimidine core and phenyl type B-arm, a series of (2,6-difluorophenyl)(2-(phenylamino)pyrimidin-4-yl)methanones was designed, prepared and tested for their anti-HIV-1 activity. The carbonyl linker bearing B phenyl arm was successfully attached at both C-2 and C-4 positions of the central pyrimidine ring using a new synthetic approach. Further modifications of target compounds are present at C-5 position of the pyrimidine ring. In vitro anti-HIV-1 activity study performed on a series of 22 compounds confirmed the crucial importance of both conformational rigidity between phenyl B arm and the pyrimidine core linked through the carbonyl bridge, as well as presence of fluoro substituents in ortho-positions of phenyl B moiety. The most potent derivative of the series, compound 17, having almost perpendicular angle within the two planes made from the B aromatic arm and the pyrimidine ring, exhibited low nanomolar anti-HIV-1 activity (EC50 = 4 nM) with no significant toxicity (CC50 > 57.1 µM).
Asunto(s)
Diseño de Fármacos , Transcriptasa Inversa del VIH/antagonistas & inhibidores , VIH-1/enzimología , Pirimidinas/química , Pirimidinas/farmacología , Inhibidores de la Transcriptasa Inversa/química , Inhibidores de la Transcriptasa Inversa/farmacología , Transcriptasa Inversa del VIH/química , Transcriptasa Inversa del VIH/metabolismo , VIH-1/efectos de los fármacos , Simulación del Acoplamiento Molecular , Conformación Proteica , Pirimidinas/metabolismo , Inhibidores de la Transcriptasa Inversa/metabolismoRESUMEN
Spleen tyrosine kinase (Syk) is an attractive drug target in autoimmune, inflammatory, and oncology disease indications. The most advanced Syk inhibitor, R406, 1 (or its prodrug form fostamatinib, 2), has shown efficacy in multiple therapeutic indications, but its clinical progress has been hampered by dose-limiting adverse effects that have been attributed, at least in part, to the off-target activities of 1. It is expected that a more selective Syk inhibitor would provide a greater therapeutic window. Herein we report the discovery and optimization of a novel series of imidazo[1,2-a]pyrazine Syk inhibitors. This work culminated in the identification of GS-9973, 68, a highly selective and orally efficacious Syk inhibitor which is currently undergoing clinical evaluation for autoimmune and oncology indications.
Asunto(s)
Indazoles/farmacología , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Tirosina Quinasas/antagonistas & inhibidores , Pirazinas/farmacología , Bazo/efectos de los fármacos , Administración Oral , Animales , Células Cultivadas , Descubrimiento de Drogas , Humanos , Indazoles/administración & dosificación , Indazoles/química , Espectroscopía de Resonancia Magnética , Ratones , Inhibidores de Proteínas Quinasas/administración & dosificación , Inhibidores de Proteínas Quinasas/química , Pirazinas/administración & dosificación , Pirazinas/química , Ratas , Espectrometría de Masa por Ionización de Electrospray , Bazo/enzimología , Relación Estructura-ActividadRESUMEN
BACKGROUND: The registrational phase III clinical trials of the nonnucleoside reverse transcriptase (RT) inhibitor (NNRTI) rilpivirine (RPV) in combination with two nucleoside/nucleotide RT inhibitors (NRTIs) found a unique genotypic resistance pattern involving the NNRTI mutation E138K with the NRTI mutation M184I. Eighty percent of subjects used emtricitabine (FTC) and tenofovir disoproxil fumarate (TDF); a single tablet regimen of FTC/RPV/TDF is in development. METHODS: HIV-1 with E138K and/or M184V or I mutations were constructed and phenotyped in MT-2 cells and the PhenoSense and Antivirogram assays. Viral fitness was determined using growth competitions. Molecular models of the mutants were constructed from the RT-RPV crystal structure. RESULTS: The E138K mutant showed low-level reduced susceptibility to RPV (2.4-fold), but full susceptibility to FTC and tenofovir (TFV). Viruses with M184V or M184I showed high-level resistance to FTC and full susceptibility to RPV and TFV. Addition of M184I, but not M184V, to E138K, further decreased susceptibility to RPV and maintained FTC resistance. The E138K and M184V or I single and double mutants showed decreased replication fitness compared with wild type. M184V outcompeted M184I when compared directly and in the background of E138K. E138K + M184I was less fit than either E138K or M184I alone. Removing a salt bridge between E138/K101 is implicated in resistance to RPV. CONCLUSIONS: The higher frequency of E138K and M184I among RPV + FTC/TDF virologic failures is due to reduced susceptibility of the single mutants to RPV and FTC and the enhanced resistance to RPV for the double mutant at the cost of decreased viral fitness.