RESUMO
Oncogenic KRAS mutations were identified decades ago, yet the selective inhibition of specific KRAS mutant proteins represents an ongoing challenge. Recent progress has been made in targeting certain P-loop mutant proteins, in particular KRAS G12C, for which the covalent inhibition of the GDP state via the Switch II pocket is now a clinically validated strategy. Inhibition of other KRAS mutant proteins such as KRAS G13D, on the other hand, still requires clinical validation. The remoteness of the D13 residue relative to the Switch II pocket in combination with the solvent exposure and conformational flexibility of the D13 side chain, as well as the difficulties of targeting carboxylate residues covalently, renders this specific protein particularly challenging to target selectively. In this report, we describe the design and evaluation of potent and KRAS G13D-selective reversible inhibitors. Subnanomolar binding to the GDP state Switch II pocket and biochemical selectivity over WT KRAS are achieved by leveraging a salt bridge with D13.
RESUMO
The dramatic increase in the prevalence of multi-drug resistant Gram-negative bacterial infections and the simultaneous lack of new classes of antibiotics is projected to result in approximately 10 million deaths per year by 2050. We report on efforts to target the Gram-negative ATP-binding cassette (ABC) transporter MsbA, an essential inner membrane protein that transports lipopolysaccharide from the inner leaflet to the periplasmic face of the inner membrane. We demonstrate the improvement of a high throughput screening hit into compounds with on-target single digit micromolar (µM) minimum inhibitory concentrations against wild-type uropathogenic Escherichia coli, Klebsiella pneumoniae, and Enterobacter cloacae. A 2.98 Å resolution X-ray crystal structure of MsbA complexed with an inhibitor revealed a novel mechanism for inhibition of an ABC transporter. The identification of a fully encapsulated membrane binding site in Gram-negative bacteria led to unique physicochemical property requirements for wild-type activity.
Assuntos
Escherichia coli , Lipopolissacarídeos , Transportadores de Cassetes de Ligação de ATP , Proteínas de Bactérias/metabolismo , Escherichia coli/metabolismo , Klebsiella pneumoniae/metabolismo , Lipopolissacarídeos/metabolismo , Lipopolissacarídeos/farmacologiaRESUMO
Structure-based design was utilized to optimize 6,6-diaryl substituted dihydropyrone and hydroxylactam to obtain inhibitors of lactate dehydrogenase (LDH) with low nanomolar biochemical and single-digit micromolar cellular potencies. Surprisingly the replacement of a phenyl with a pyridyl moiety in the chemical structure revealed a new binding mode for the inhibitors with subtle conformational change of the LDHA active site. This led to the identification of a potent, cell-active hydroxylactam inhibitor exhibiting an in vivo pharmacokinetic profile suitable for mouse tumor xenograft study.
Assuntos
Inibidores Enzimáticos/farmacologia , L-Lactato Desidrogenase/antagonistas & inibidores , Lactamas/farmacologia , Animais , Linhagem Celular , Cães , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/química , Humanos , L-Lactato Desidrogenase/metabolismo , Lactamas/química , Camundongos , Microssomos Hepáticos/química , Microssomos Hepáticos/metabolismo , Estrutura Molecular , Relação Estrutura-AtividadeRESUMO
Fulvestrant is an FDA-approved drug with a dual mechanism of action (MOA), acting as a full antagonist and degrader of the estrogen receptor protein. A significant limitation of fulvestrant is the dosing regimen required for efficacy. Due to its high lipophilicity and poor pharmacokinetic profile, fulvestrant needs to be administered through intramuscular injections which leads to injection site soreness. This route of administration also limits the dose and target occupancy in patients. We envisioned a best-in-class molecule that would function with the same dual MOA as fulvestrant, but with improved physicochemical properties and would be orally bioavailable. Herein we report our progress toward that goal, resulting in a new lead GNE-502 which addressed some of the liabilities of our previously reported lead molecule GNE-149.
Assuntos
Antineoplásicos/farmacologia , Antineoplásicos/farmacocinética , Neoplasias da Mama/tratamento farmacológico , Descoberta de Drogas , Receptores de Estrogênio/metabolismo , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/química , Relação Dose-Resposta a Droga , Feminino , Humanos , Células MCF-7 , Camundongos , Estrutura Molecular , Conformação Proteica , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Herein we present an investigation into the scope and mechanism for the synthesis of cyclopentyl and heterocyclic fused pyridones from the corresponding enyne amides. In the presence of a secondary amine, cyclization proceeds smoothly to form 5,6-bicyclic pyridones in 12-90% yield. The cyclization fails with enyne amides of six-membered and larger ring systems. The ring closure reaction is catalytic in nature with respect to the secondary amine and proceeds via sequential 1,6-addition of the amine, 6-exo-trig ring closure of the iminium intermediate, and subsequent elimination of the secondary amine. Computations show reduced conjugation between the enyne and amide for six-membered and larger systems, thereby providing an explanation for the inability of such enyne amides to form fused pyridones.
Assuntos
Amidas , Piridonas , Catálise , Ciclização , Estrutura MolecularRESUMO
Breast cancer remains a leading cause of cancer death in women, representing a significant unmet medical need. Here, we disclose our discovery efforts culminating in a clinical candidate, 35 (GDC-9545 or giredestrant). 35 is an efficient and potent selective estrogen receptor degrader (SERD) and a full antagonist, which translates into better antiproliferation activity than known SERDs (1, 6, 7, and 9) across multiple cell lines. Fine-tuning the physiochemical properties enabled once daily oral dosing of 35 in preclinical species and humans. 35 exhibits low drug-drug interaction liability and demonstrates excellent in vitro and in vivo safety profiles. At low doses, 35 induces tumor regressions either as a single agent or in combination with a CDK4/6 inhibitor in an ESR1Y537S mutant PDX or a wild-type ERα tumor model. Currently, 35 is being evaluated in Phase III clinical trials.
Assuntos
Antineoplásicos/uso terapêutico , Neoplasias da Mama/tratamento farmacológico , Carbolinas/uso terapêutico , Antagonistas do Receptor de Estrogênio/uso terapêutico , Receptor alfa de Estrogênio/metabolismo , Animais , Antineoplásicos/química , Antineoplásicos/farmacocinética , Carbolinas/química , Carbolinas/farmacocinética , Cães , Antagonistas do Receptor de Estrogênio/química , Antagonistas do Receptor de Estrogênio/farmacocinética , Feminino , Humanos , Células MCF-7 , Macaca fascicularis , Camundongos , Estrutura Molecular , Ratos , Relação Estrutura-Atividade , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Discovery of novel classes of Gram-negative antibiotics with activity against multi-drug resistant infections is a critical unmet need. As an essential member of the lipoprotein biosynthetic pathway, lipoprotein signal peptidase II (LspA) is an attractive target for antibacterial drug discovery, with the natural product inhibitor globomycin offering a modestly-active starting point. Informed by structure-based design, the globomycin depsipeptide was optimized to improve activity against E. coli. Backbone modifications, together with adjustment of physicochemical properties, afforded potent compounds with good in vivo pharmacokinetic profiles. Optimized compounds such as 51 (E. coli MIC 3.1 µM) and 61 (E. coli MIC 0.78 µM) demonstrate broad spectrum activity against gram-negative pathogens and may provide opportunities for future antibiotic discovery.
Assuntos
Antibacterianos/farmacologia , Escherichia coli/efeitos dos fármacos , Peptídeos/farmacologia , Antibacterianos/síntese química , Antibacterianos/química , Relação Dose-Resposta a Droga , Testes de Sensibilidade Microbiana , Estrutura Molecular , Peptídeos/síntese química , Peptídeos/química , Relação Estrutura-AtividadeRESUMO
Estrogen receptor alpha (ERα) is a well-validated drug target for ER-positive (ER+) breast cancer. Fulvestrant is FDA-approved to treat ER+ breast cancer and works through two mechanisms-as a full antagonist and selective estrogen receptor degrader (SERD)-but lacks oral bioavailability. Thus, we envisioned a "best-in-class" molecule with the same dual mechanisms as fulvestrant, but with significant oral exposure. Through lead optimization, we discovered a tool molecule 12 (GNE-149) with improved degradation and antiproliferative activity in both MCF7 and T47D cells. To illustrate the binding mode and key interactions of this scaffold with ERα, we obtained a cocrystal structure of 6 that showed ionic interaction of azetidine with Asp351 residue. Importantly, 12 showed favorable metabolic stability and good oral exposure. 12 exhibited antagonist effect in the uterus and demonstrated robust dose-dependent efficacy in xenograft models.
RESUMO
Phenolic groups are responsible for the high clearance and low oral bioavailability of the estrogen receptor alpha (ERα) clinical candidate GDC-0927. An exhaustive search for a backup molecule with improved pharmacokinetic (PK) properties identified several metabolically stable analogs, although in general at the expense of the desired potency and degradation efficiency. C-8 hydroxychromene 30 is the first example of a phenol-containing chromene that not only maintained excellent potency but also exhibited 10-fold higher oral exposure in rats. The improved in vivo clearance in rat was hypothesized to be the result of C-8 hydroxy group being sterically protected from glucuronide conjugation. The excellent potency underscores the possibility of replacing the presumed indispensable phenolic group at C-6 or C-7 of the chromene core. Co-crystal structures were obtained to highlight the change in key interactions and rationalize the retained potency.
Assuntos
Azetidinas/farmacologia , Receptor alfa de Estrogênio/metabolismo , Flavonoides/farmacologia , Administração Oral , Animais , Azetidinas/administração & dosagem , Azetidinas/metabolismo , Azetidinas/farmacocinética , Cristalografia por Raios X , Descoberta de Drogas , Estabilidade de Medicamentos , Flavonoides/administração & dosagem , Flavonoides/metabolismo , Flavonoides/farmacocinética , Humanos , Células MCF-7 , Microssomos Hepáticos/metabolismo , Ratos , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
There is a critical need for new antibacterial strategies to counter the growing problem of antibiotic resistance. In Gram-negative bacteria, the outer membrane (OM) provides a protective barrier against antibiotics and other environmental insults. The outer leaflet of the outer membrane is primarily composed of lipopolysaccharide (LPS). Outer membrane biogenesis presents many potentially compelling drug targets as this pathway is absent in higher eukaryotes. Most proteins involved in LPS biosynthesis and transport are essential; however, few compounds have been identified that inhibit these proteins. The inner membrane ABC transporter MsbA carries out the first essential step in the trafficking of LPS to the outer membrane. We conducted a biochemical screen for inhibitors of MsbA and identified a series of quinoline compounds that kill Escherichia coli through inhibition of its ATPase and transport activity, with no loss of activity against clinical multidrug-resistant strains. Identification of these selective inhibitors indicates that MsbA is a viable target for new antibiotics, and the compounds we identified serve as useful tools to further probe the LPS transport pathway in Gram-negative bacteria.
Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Proteínas da Membrana Bacteriana Externa/metabolismo , Proteínas de Bactérias/metabolismo , Proteínas de Escherichia coli/metabolismo , Escherichia coli/metabolismo , Lipopolissacarídeos/metabolismo , Antibacterianos/farmacologia , Transporte Biológico/efeitos dos fármacos , Transporte Biológico/fisiologia , Escherichia coli/efeitos dos fármacosRESUMO
The movement of core-lipopolysaccharide across the inner membrane of Gram-negative bacteria is catalysed by an essential ATP-binding cassette transporter, MsbA. Recent structures of MsbA and related transporters have provided insights into the molecular basis of active lipid transport; however, structural information about their pharmacological modulation remains limited. Here we report the 2.9 Å resolution structure of MsbA in complex with G907, a selective small-molecule antagonist with bactericidal activity, revealing an unprecedented mechanism of ABC transporter inhibition. G907 traps MsbA in an inward-facing, lipopolysaccharide-bound conformation by wedging into an architecturally conserved transmembrane pocket. A second allosteric mechanism of antagonism occurs through structural and functional uncoupling of the nucleotide-binding domains. This study establishes a framework for the selective modulation of ABC transporters and provides rational avenues for the design of new antibiotics and other therapeutics targeting this protein family.
Assuntos
Transportadores de Cassetes de Ligação de ATP/antagonistas & inibidores , Transportadores de Cassetes de Ligação de ATP/química , Antibacterianos/química , Antibacterianos/farmacologia , Proteínas de Bactérias/antagonistas & inibidores , Proteínas de Bactérias/química , Quinolinas/química , Quinolinas/farmacologia , Transportadores de Cassetes de Ligação de ATP/metabolismo , Regulação Alostérica/efeitos dos fármacos , Proteínas de Bactérias/metabolismo , Sítios de Ligação/efeitos dos fármacos , Cristalografia por Raios X , Relação Dose-Resposta a Droga , Escherichia coli/química , Hidrocarbonetos/química , Hidrocarbonetos/metabolismo , Lipopolissacarídeos/química , Lipopolissacarídeos/metabolismo , Lipopolissacarídeos/farmacologia , Modelos Moleculares , Domínios Proteicos/efeitos dos fármacosRESUMO
A high-throughput screening (HTS) of the Genentech/Roche library identified a novel, uncharged scaffold as a KDM5A inhibitor. Lacking insight into the binding mode, initial attempts to improve inhibitor potency failed to improve potency, and synthesis of analogs was further hampered by the presence of a C-C bond between the pyrrolidine and pyridine. Replacing this with a C-N bond significantly simplified synthesis, yielding pyrazole analog 35, of which we obtained a co-crystal structure with KDM5A. Using structure-based design approach, we identified 50 with improved biochemical, cell potency and reduced MW and lower lipophilicity (LogD) compared with the original hit. Furthermore, 50 showed lower clearance than 9 in mice. In combination with its remarkably low plasma protein binding (PPB) in mice (40%), oral dosing of 50 at 5mg/kg resulted in unbound Cmax â¼2-fold of its cell potency (PC9 H3K4Me3 0.96µM), meeting our criteria for an in vivo tool compound from a new scaffold.
Assuntos
Inibidores Enzimáticos/farmacologia , Ensaios de Triagem em Larga Escala , Pirazóis/farmacologia , Proteína 2 de Ligação ao Retinoblastoma/antagonistas & inibidores , Administração Oral , Animais , Disponibilidade Biológica , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/administração & dosagem , Inibidores Enzimáticos/química , Humanos , Camundongos , Microssomos Hepáticos/química , Microssomos Hepáticos/metabolismo , Simulação de Acoplamento Molecular , Estrutura Molecular , Pirazóis/administração & dosagem , Pirazóis/química , Ratos , Proteína 2 de Ligação ao Retinoblastoma/metabolismo , Relação Estrutura-AtividadeRESUMO
A series of trisubstituted hydroxylactams was identified as potent enzymatic and cellular inhibitors of human lactate dehydrogenase A. Utilizing structure-based design and physical property optimization, multiple inhibitors were discovered with <10 µM lactate IC50 in a MiaPaca2 cell line. Optimization of the series led to 29, a potent cell active molecule (MiaPaca2 IC50 = 0.67 µM) that also possessed good exposure when dosed orally to mice.
RESUMO
Metabolic reprogramming in tumors represents a potential therapeutic target. Herein we used shRNA depletion and a novel lactate dehydrogenase (LDHA) inhibitor, GNE-140, to probe the role of LDHA in tumor growth in vitro and in vivo. In MIA PaCa-2 human pancreatic cells, LDHA inhibition rapidly affected global metabolism, although cell death only occurred after 2 d of continuous LDHA inhibition. Pancreatic cell lines that utilize oxidative phosphorylation (OXPHOS) rather than glycolysis were inherently resistant to GNE-140, but could be resensitized to GNE-140 with the OXPHOS inhibitor phenformin. Acquired resistance to GNE-140 was driven by activation of the AMPK-mTOR-S6K signaling pathway, which led to increased OXPHOS, and inhibitors targeting this pathway could prevent resistance. Thus, combining an LDHA inhibitor with compounds targeting the mitochondrial or AMPK-S6K signaling axis may not only broaden the clinical utility of LDHA inhibitors beyond glycolytically dependent tumors but also reduce the emergence of resistance to LDHA inhibition.
Assuntos
Plasticidade Celular/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , L-Lactato Desidrogenase/antagonistas & inibidores , Piridonas/farmacologia , Tiofenos/farmacologia , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/química , Humanos , L-Lactato Desidrogenase/metabolismo , Modelos Moleculares , Estrutura Molecular , Piridonas/química , Relação Estrutura-Atividade , Tiofenos/químicaRESUMO
Features from a high throughput screening (HTS) hit and a previously reported scaffold were combined to generate 1,7-naphthyridones as novel KDM5 enzyme inhibitors with nanomolar potencies. These molecules exhibited high selectivity over the related KDM4C and KDM2B isoforms. An X-ray co-crystal structure of a representative molecule bound to KDM5A showed that these inhibitors are competitive with the co-substrate (2-oxoglutarate or 2-OG).
Assuntos
Histona Desmetilases com o Domínio Jumonji/antagonistas & inibidores , Naftiridinas/farmacologia , Proteínas Nucleares/antagonistas & inibidores , Proteínas Repressoras/antagonistas & inibidores , Proteína 2 de Ligação ao Retinoblastoma/antagonistas & inibidores , Animais , Cristalografia por Raios X , Cães , Desenho de Fármacos , Humanos , Células Madin Darby de Rim Canino , Naftiridinas/química , Relação Estrutura-AtividadeRESUMO
Starting with a lead [1,5-a]pyrimidin-7(4H)-one-containing molecule (1), we generated potent, selective and orally bioavailable KDM5 inhibitors. Using structure- and property-based approaches, we designed 48 with improved cell potency (PC9 H3K4Me3 EC50=0.34µM). Furthermore, 48 maintained suitable physiochemical properties and displayed an excellent pharmacokinetic (PK) profile in mice. When dosed orally in mice at 50mg/kg twice a day (BID), 48 showed an unbound maximal plasma concentration (Cmax) >15-fold over its cell EC50, thereby providing a robust chemical probe for studying KDM5 biological functions in vivo.
Assuntos
Pirazóis/química , Pirimidinonas/química , Proteína 2 de Ligação ao Retinoblastoma/antagonistas & inibidores , Administração Oral , Animais , Sítios de Ligação , Cristalografia por Raios X , Feminino , Meia-Vida , Histonas/metabolismo , Humanos , Fígado/metabolismo , Camundongos , Microssomos Hepáticos/metabolismo , Simulação de Dinâmica Molecular , Pirazóis/síntese química , Pirazóis/farmacocinética , Pirimidinonas/sangue , Pirimidinonas/síntese química , Pirimidinonas/farmacocinética , Ratos , Proteína 2 de Ligação ao Retinoblastoma/metabolismo , Relação Estrutura-AtividadeRESUMO
Optimization of 5-(2,6-dichlorophenyl)-3-hydroxy-2-mercaptocyclohex-2-enone using structure-based design strategies resulted in inhibitors with considerable improvement in biochemical potency against human lactate dehydrogenase A (LDHA). These potent inhibitors were typically selective for LDHA over LDHB isoform (410 fold) and other structurally related malate dehydrogenases, MDH1 and MDH2 (>500 fold). An X-ray crystal structure of enzymatically most potent molecule bound to LDHA revealed two additional interactions associated with enhanced biochemical potency.
Assuntos
Inibidores Enzimáticos/síntese química , L-Lactato Desidrogenase/antagonistas & inibidores , Animais , Cristalografia por Raios X , Cães , Inibidores Enzimáticos/metabolismo , Inibidores Enzimáticos/farmacologia , Humanos , L-Lactato Desidrogenase/metabolismo , Células Madin Darby de Rim CaninoRESUMO
A series of 3,6-disubstituted dihydropyrones were identified as inhibitors of human lactate dehydrogenase (LDH)-A. Structure activity relationships were explored and a series of 6,6-spiro analogs led to improvements in LDHA potency (IC50 <350 nM). An X-ray crystal structure of an improved compound bound to human LDHA was obtained and it illustrated additional opportunities to enhance the potency of these compounds, resulting in the identification of 51 (IC50=30 nM).
Assuntos
Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/farmacologia , L-Lactato Desidrogenase/antagonistas & inibidores , Pironas/síntese química , Pironas/farmacologia , Sítios de Ligação , Cristalografia por Raios X , Humanos , L-Lactato Desidrogenase/metabolismo , Modelos Moleculares , Estrutura Molecular , Relação Estrutura-AtividadeRESUMO
A novel class of 3-hydroxy-2-mercaptocyclohex-2-enone-containing inhibitors of human lactate dehydrogenase (LDH) was identified through a high-throughput screening approach. Biochemical and surface plasmon resonance experiments performed with a screening hit (LDHA IC50=1.7 µM) indicated that the compound specifically associated with human LDHA in a manner that required simultaneous binding of the NADH co-factor. Structural variation of this screening hit resulted in significant improvements in LDHA biochemical inhibition activity (best IC50=0.18 µM). Two crystal structures of optimized compounds bound to human LDHA were obtained and explained many of the observed structure-activity relationships. In addition, an optimized inhibitor exhibited good pharmacokinetic properties after oral administration to rats (F=45%).
Assuntos
Cicloexanonas/farmacologia , Inibidores Enzimáticos/farmacologia , L-Lactato Desidrogenase/antagonistas & inibidores , Compostos de Sulfidrila/farmacologia , Administração Oral , Animais , Cicloexanonas/administração & dosagem , Cicloexanonas/química , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/administração & dosagem , Inibidores Enzimáticos/química , Ensaios de Triagem em Larga Escala , Humanos , L-Lactato Desidrogenase/metabolismo , Modelos Moleculares , Estrutura Molecular , Ratos , Relação Estrutura-Atividade , Compostos de Sulfidrila/administração & dosagem , Compostos de Sulfidrila/químicaRESUMO
In the past few years, there have been many advances in the efforts to cure patients with hepatitis C virus (HCV). The ultimate goal of these efforts is to develop a combination therapy consisting of only direct-antiviral agents (DAAs). In this paper, we discuss our efforts that led to the identification of a bicyclic template with potent activity against the NS5B polymerase, a critical enzyme on the life cycle of HCV. In continuation of our exploration to improve the stilbene series, the 3,5,6,8-tetrasubstituted quinoline core was identified as replacement of the stilbene moiety. 6-Methoxy-2(1H)-pyridone was identified among several heterocyclic headgroups to have the best potency. Solubility of the template was improved by replacing a planar aryl linker with a saturated pyrrolidine. Profiling of the most promising compounds led to the identification of quinoline 41 (RG7109), which was selected for advancement to clinical development.