RESUMEN
Lipopolysaccharide (LPS) resides in the outer membrane of Gram-negative bacteria where it is responsible for barrier function1,2. LPS can cause death as a result of septic shock, and its lipid A core is the target of polymyxin antibiotics3,4. Despite the clinical importance of polymyxins and the emergence of multidrug resistant strains5, our understanding of the bacterial factors that regulate LPS biogenesis is incomplete. Here we characterize the inner membrane protein PbgA and report that its depletion attenuates the virulence of Escherichia coli by reducing levels of LPS and outer membrane integrity. In contrast to previous claims that PbgA functions as a cardiolipin transporter6-9, our structural analyses and physiological studies identify a lipid A-binding motif along the periplasmic leaflet of the inner membrane. Synthetic PbgA-derived peptides selectively bind to LPS in vitro and inhibit the growth of diverse Gram-negative bacteria, including polymyxin-resistant strains. Proteomic, genetic and pharmacological experiments uncover a model in which direct periplasmic sensing of LPS by PbgA coordinates the biosynthesis of lipid A by regulating the stability of LpxC, a key cytoplasmic biosynthetic enzyme10-12. In summary, we find that PbgA has an unexpected but essential role in the regulation of LPS biogenesis, presents a new structural basis for the selective recognition of lipids, and provides opportunities for future antibiotic discovery.
Asunto(s)
Membrana Celular/química , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/metabolismo , Escherichia coli/química , Escherichia coli/patogenicidad , Lipopolisacáridos/química , Lipopolisacáridos/metabolismo , Amidohidrolasas/química , Amidohidrolasas/metabolismo , Secuencias de Aminoácidos , Membrana Externa Bacteriana/química , Membrana Externa Bacteriana/metabolismo , Sitios de Unión , Membrana Celular/metabolismo , Estabilidad de Enzimas , Escherichia coli/citología , Escherichia coli/efectos de los fármacos , Genes Esenciales , Hidrolasas/química , Hidrolasas/metabolismo , Lípido A/química , Lípido A/metabolismo , Lipopolisacáridos/biosíntesis , Pruebas de Sensibilidad Microbiana , Viabilidad Microbiana/efectos de los fármacos , Modelos Moleculares , Fragmentos de Péptidos/química , Fragmentos de Péptidos/metabolismo , Fragmentos de Péptidos/farmacología , Periplasma/química , Periplasma/metabolismo , Unión Proteica , VirulenciaRESUMEN
Acinetobacter baumannii is a clinically important, predominantly health care-associated gram-negative bacterium with high rates of emerging resistance worldwide. Given the urgent need for novel antibacterial therapies against A. baumannii, we focused on inhibiting lipoprotein biosynthesis, a pathway that is essential for envelope biogenesis in gram-negative bacteria. The natural product globomycin, which inhibits the essential type II signal peptidase prolipoprotein signal peptidase (LspA), is ineffective against wild-type A. baumannii clinical isolates due to its poor penetration through the outer membrane. Here, we describe a globomycin analog, G5132, that is more potent against wild-type and clinical A. baumannii isolates. Mutations leading to G5132 resistance in A. baumannii map to the signal peptide of a single hypothetical gene, which we confirm encodes an alanine-rich lipoprotein and have renamed lirL (prolipoprotein signal peptidase inhibitor resistance lipoprotein). LirL is a highly abundant lipoprotein primarily localized to the inner membrane. Deletion of lirL leads to G5132 resistance, inefficient cell division, increased sensitivity to serum, and attenuated virulence. Signal peptide mutations that confer resistance to G5132 lead to the accumulation of diacylglyceryl-modified LirL prolipoprotein in untreated cells without significant loss in cell viability, suggesting that these mutations overcome a block in lipoprotein biosynthetic flux by decreasing LirL prolipoprotein substrate sensitivity to processing by LspA. This study characterizes a lipoprotein that plays a critical role in resistance to LspA inhibitors and validates lipoprotein biosynthesis as a antibacterial target in A. baumannii.
Asunto(s)
Acinetobacter baumannii , Antibacterianos , Ácido Aspártico Endopeptidasas , Proteínas Bacterianas , Farmacorresistencia Bacteriana , Furanos , Eliminación de Gen , Lipoproteínas , Inhibidores de Proteasas , Piridinas , Acinetobacter baumannii/efectos de los fármacos , Acinetobacter baumannii/enzimología , Acinetobacter baumannii/genética , Antibacterianos/farmacología , Ácido Aspártico Endopeptidasas/genética , Proteínas Bacterianas/genética , Farmacorresistencia Bacteriana/genética , Furanos/farmacología , Lipoproteínas/biosíntesis , Lipoproteínas/genética , Péptidos/farmacología , Inhibidores de Proteasas/farmacología , Señales de Clasificación de Proteína/genética , Piridinas/farmacologíaRESUMEN
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.
Asunto(s)
Antibacterianos/farmacología , Escherichia coli/efectos de los fármacos , Péptidos/farmacología , Antibacterianos/síntesis química , Antibacterianos/química , Relación Dosis-Respuesta a Droga , Pruebas de Sensibilidad Microbiana , Estructura Molecular , Péptidos/síntesis química , Péptidos/química , Relación Estructura-ActividadRESUMEN
Disruption of interleukin-13 (IL-13) signaling with large molecule antibody therapies has shown promise in diseases of allergic inflammation. Given that IL-13 recruits several members of the Janus Kinase family (JAK1, JAK2, and TYK2) to its receptor complex, JAK inhibition may offer an alternate small molecule approach to disrupting IL-13 signaling. Herein we demonstrate that JAK1 is likely the isoform most important to IL-13 signaling. Structure-based design was then used to improve the JAK1 potency of a series of previously reported JAK2 inhibitors. The ability to impede IL-13 signaling was thereby significantly improved, with the best compounds exhibiting single digit nM IC50's in cell-based assays dependent upon IL-13 signaling. Appropriate substitution was further found to influence inhibition of a key off-target, LRRK2. Finally, the most potent compounds were found to be metabolically labile, which makes them ideal scaffolds for further development as topical agents for IL-13 mediated diseases of the lungs and skin (for example asthma and atopic dermatitis, respectively).
Asunto(s)
Dermatitis Atópica/genética , Interleucina-13/metabolismo , Janus Quinasa 1/antagonistas & inhibidores , Janus Quinasa 2/antagonistas & inhibidores , Humanos , Transducción de SeñalRESUMEN
The treatment of epidermal growth factor receptor (EGFR)-driven non-small cell lung cancers with the T790M resistance mutation remains a significant unmet medical need. We report the identification of 4-aminoindazolyl-dihydrofuro[3,4-d]pyrimidines as non-covalent inhibitors of EGFR, with excellent activity against the T790M resistance double mutants and initial single activating mutants. Using an optimization strategy focused on structure-based design and improving PK properties through metabolite identification, we obtained advanced leads with high oral exposure.
Asunto(s)
Receptores ErbB/antagonistas & inhibidores , Furanos/farmacología , Indazoles/farmacología , Inhibidores de Proteínas Quinasas/farmacología , Pirimidinas/farmacología , Acrilamidas/farmacología , Compuestos de Anilina/farmacología , Animales , Sitios de Unión , Cristalografía por Rayos X , Perros , Receptores ErbB/química , Clorhidrato de Erlotinib/farmacología , Furanos/síntesis química , Furanos/química , Furanos/farmacocinética , Hepatocitos/metabolismo , Ensayos Analíticos de Alto Rendimiento , Humanos , Enlace de Hidrógeno , Indazoles/síntesis química , Indazoles/química , Indazoles/farmacocinética , Ratones , Microsomas Hepáticos/metabolismo , Mutación Puntual , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacocinética , Pirimidinas/síntesis química , Pirimidinas/química , Pirimidinas/farmacocinética , RatasRESUMEN
Multidrug-resistant bacteria are spreading at alarming rates, and despite extensive efforts, no new antibiotic class with activity against Gram-negative bacteria has been approved in over 50 years. LepB inhibitors (LepBi) based on the arylomycin class of natural products are a novel class of antibiotics and function by inhibiting the bacterial type I signal peptidase (SPase) in Gram-negative bacteria. One critical aspect of LepBi development involves optimization of the membrane-anchored lipophilic portion of the molecule. We therefore developed an approach that assesses the effect of this portion on the complicated equilibria of plasma protein binding, crossing the outer membrane of Gram-negative bacteria and anchoring in the bacterial inner membrane to facilitate SPase binding. Our findings provide important insights into the development of antibacterial agents where the target is associated with the inner membrane of Gram-negative bacteria.
RESUMEN
Small molecule inhibitors that target the phosphatidylinositol 3-kinase (PI3K) signaling pathway have received significant interest for the treatment of cancers. The class I isoform PI3Kα is most commonly associated with solid tumors via gene amplification or activating mutations. However, inhibitors demonstrating both PI3K isoform and mutant specificity have remained elusive. Herein, we describe the optimization and characterization of a series of benzoxazepin-oxazolidinone ATP-competitive inhibitors of PI3Kα which also induce the selective degradation of the mutant p110α protein, the catalytic subunit of PI3Kα. Structure-based design informed isoform-specific interactions within the binding site, leading to potent inhibitors with greater than 300-fold selectivity over the other Class I PI3K isoforms. Further optimization of pharmacokinetic properties led to excellent in vivo exposure and efficacy and the identification of clinical candidate GDC-0077 (inavolisib, 32), which is now under evaluation in a Phase III clinical trial as a treatment for patients with PIK3CA-mutant breast cancer.
Asunto(s)
Neoplasias de la Mama , Fosfatidilinositol 3-Quinasas , Humanos , Femenino , Inhibidores de las Quinasa Fosfoinosítidos-3/farmacología , Inhibidores de las Quinasa Fosfoinosítidos-3/uso terapéutico , Fosfatidilinositol 3-Quinasas/metabolismo , Fosfatidilinositol 3-Quinasa Clase I/uso terapéutico , Neoplasias de la Mama/tratamiento farmacológico , Línea Celular Tumoral , MutaciónRESUMEN
PIK3CA is one of the most frequently mutated oncogenes; the p110a protein it encodes plays a central role in tumor cell proliferation. Small-molecule inhibitors targeting the PI3K p110a catalytic subunit have entered clinical trials, with early-phase GDC-0077 studies showing antitumor activity and a manageable safety profile in patients with PIK3CA-mutant breast cancer. However, preclinical studies have shown that PI3K pathway inhibition releases negative feedback and activates receptor tyrosine kinase signaling, reengaging the pathway and attenuating drug activity. Here we discover that GDC-0077 and taselisib more potently inhibit mutant PI3K pathway signaling and cell viability through unique HER2-dependent mutant p110a degradation. Both are more effective than other PI3K inhibitors at maintaining prolonged pathway suppression. This study establishes a new strategy for identifying inhibitors that specifically target mutant tumors by selective degradation of the mutant oncoprotein and provide a strong rationale for pursuing PI3Kα degraders in patients with HER2-positive breast cancer. SIGNIFICANCE: The PI3K inhibitors GDC-0077 and taselisib have a unique mechanism of action; both inhibitors lead to degradation of mutant p110a protein. The inhibitors that have the ability to trigger specific degradation of mutant p110a without significant change in wild-type p110a protein may result in improved therapeutic index in PIK3CA-mutant tumors.See related commentary by Vanhaesebroeck et al., p. 20.This article is highlighted in the In This Issue feature, p. 1.
Asunto(s)
Antineoplásicos , Neoplasias de la Mama , Fosfatidilinositol 3-Quinasa Clase I , Imidazoles , Oxazepinas , Inhibidores de las Quinasa Fosfoinosítidos-3 , Receptor ErbB-2 , Femenino , Humanos , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/genética , Línea Celular Tumoral/efectos de los fármacos , Fosfatidilinositol 3-Quinasa Clase I/genética , Imidazoles/farmacología , Imidazoles/uso terapéutico , Oxazepinas/farmacología , Oxazepinas/uso terapéutico , Inhibidores de las Quinasa Fosfoinosítidos-3/farmacología , Inhibidores de las Quinasa Fosfoinosítidos-3/uso terapéutico , Receptor ErbB-2/genéticaRESUMEN
This letter describes the structure-activity relationship (SAR) of the 'right-wing' α-amino acid residue of potent tetrahydroisoquinoline (THIQ)-derived LFA-1/ICAM-1 antagonists. Novel (S)-substituted heteroaryl-bearing α-amino acids have been identified as replacements of the 'right-wing' (S)-2,3-diaminopropanoic acid (DAP) moiety. Improvement of potency in the Hut-78 assay in the presence of 10% human serum has also been achieved.
Asunto(s)
Aminoácidos/química , Molécula 1 de Adhesión Intercelular/química , Antígeno-1 Asociado a Función de Linfocito/química , Tetrahidroisoquinolinas/química , Animales , Molécula 1 de Adhesión Intercelular/metabolismo , Antígeno-1 Asociado a Función de Linfocito/metabolismo , Masculino , Ratas , Ratas Sprague-Dawley , Relación Estructura-Actividad , Tetrahidroisoquinolinas/síntesis química , Tetrahidroisoquinolinas/farmacocinética , beta-Alanina/análogos & derivados , beta-Alanina/químicaRESUMEN
This letter describes the discovery of a novel series of tetrahydroisoquinoline (THIQ)-derived small molecules that potently inhibit both human T-cell migration and super-antigen induced T-cell activation through disruption of the binding of integrin LFA-1 to its receptor, ICAM-1. In addition to excellent in vitro potency, 6q shows good pharmacokinetic properties and its ethyl ester (6t) demonstrates good oral bioavailability in both mouse and rat. Either intravenous administration of 6q or oral administration of its ethyl ester (6t) produced a significant reduction of neutrophil migration in a thioglycollate-induced murine peritonitis model.
Asunto(s)
Molécula 1 de Adhesión Intercelular/efectos de los fármacos , Antígeno-1 Asociado a Función de Linfocito/efectos de los fármacos , Tetrahidroisoquinolinas/farmacología , Animales , Disponibilidad Biológica , Descubrimiento de Drogas , Humanos , Tetrahidroisoquinolinas/administración & dosificación , Tetrahidroisoquinolinas/farmacocinéticaRESUMEN
The discovery and development of targeted protein degraders have become important areas of research in the field of medicinal chemistry. Inducing degradation of a target protein presents several advantages relative to simple inhibition including a potential for extended duration of action and more profound pharmacology. While engineered heterodimeric molecules have recently been a major focus within industry and academia, this Perspective highlights examples of targeted protein degradation observed for smaller, monomeric molecules. Methods and tools for evaluating protein degradation as well as a discussion of physical properties of monomeric vs engineered heterodimeric degraders are presented.
Asunto(s)
Descubrimiento de Drogas/métodos , Proteolisis/efectos de los fármacos , Bibliotecas de Moléculas Pequeñas , Sitios de Unión , Línea Celular Tumoral , Humanos , Estructura Molecular , Unión Proteica , Proteínas/antagonistas & inhibidores , Bibliotecas de Moléculas Pequeñas/química , Bibliotecas de Moléculas Pequeñas/farmacología , Ubiquitina-Proteína LigasasRESUMEN
Clinical development of antibiotics with novel mechanisms of action to kill pathogenic bacteria is challenging, in part, due to the inevitable emergence of resistance. A phenomenon of potential clinical importance that is broadly overlooked in preclinical development is heteroresistance, an often-unstable phenotype in which subpopulations of bacterial cells show decreased antibiotic susceptibility relative to the dominant population. Here, we describe a new globomycin analog, G0790, with potent activity against the Escherichia coli type II signal peptidase LspA and uncover two novel resistance mechanisms to G0790 in the clinical uropathogenic E. coli strain CFT073. Building on the previous finding that complete deletion of Lpp, the major Gram-negative outer membrane lipoprotein, leads to globomycin resistance, we also find that an unexpectedly modest decrease in Lpp levels mediated by insertion-based disruption of regulatory elements is sufficient to confer G0790 resistance and increase sensitivity to serum killing. In addition, we describe a heteroresistance phenotype mediated by genomic amplifications of lspA that result in increased LspA levels sufficient to overcome inhibition by G0790 in culture. These genomic amplifications are highly unstable and are lost after as few as two subcultures in the absence of G0790, which places amplification-containing resistant strains at high risk of being misclassified as susceptible by routine antimicrobial susceptibility testing. In summary, our study uncovers two vastly different mechanisms of resistance to LspA inhibitors in E. coli and emphasizes the importance of considering the potential impact of unstable and heterogenous phenotypes when developing antibiotics for clinical use.IMPORTANCE Despite increasing evidence suggesting that antibiotic heteroresistance can lead to treatment failure, the significance of this phenomena in the clinic is not well understood, because many clinical antibiotic susceptibility testing approaches lack the resolution needed to reliably classify heteroresistant strains. Here we present G0790, a new globomycin analog and potent inhibitor of the Escherichia coli type II signal peptidase LspA. We demonstrate that in addition to previously known mechanisms of resistance to LspA inhibitors, unstable genomic amplifications containing lspA can lead to modest yet biologically significant increases in LspA protein levels that confer a heteroresistance phenotype.
Asunto(s)
Antibacterianos/farmacología , Ácido Aspártico Endopeptidasas/antagonistas & inhibidores , Proteínas Bacterianas/antagonistas & inhibidores , Farmacorresistencia Bacteriana/genética , Lipoproteínas/metabolismo , Escherichia coli Uropatógena/efectos de los fármacos , Escherichia coli Uropatógena/enzimología , Animales , Ácido Aspártico Endopeptidasas/genética , Proteínas Bacterianas/genética , Enterobacteriaceae/clasificación , Enterobacteriaceae/efectos de los fármacos , Escherichia coli/efectos de los fármacos , Escherichia coli/enzimología , Escherichia coli/genética , Infecciones por Escherichia coli/microbiología , Femenino , Humanos , Ratones , Ratones Endogámicos C57BL , Péptidos/química , Péptidos/farmacología , Escherichia coli Uropatógena/genética , Escherichia coli Uropatógena/patogenicidadRESUMEN
This Letter describes the discovery and key structure-activity relationship (SAR) of a series of 2-aminobenzimidazoles as potent Aurora kinase inhibitors. 2-Aminobenzimidazole serves as a bioisostere of the biaryl urea residue of SNS-314 (1c), which is a potent Aurora kinase inhibitor and entered clinical testing in patients with solid tumors. Compared to SNS-314, this series of compounds offers better aqueous solubility while retaining comparable in vitro potency in biochemical and cell-based assays; in particular, 6m has also demonstrated a comparable mouse iv PK profile to SNS-314.
Asunto(s)
Antineoplásicos/química , Bencimidazoles/química , Inhibidores de Proteínas Quinasas/química , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Animales , Antineoplásicos/síntesis química , Antineoplásicos/farmacocinética , Aurora Quinasas , Bencimidazoles/síntesis química , Bencimidazoles/farmacocinética , Línea Celular Tumoral , Humanos , Ratones , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/farmacocinética , Proteínas Serina-Treonina Quinasas/metabolismo , Relación Estructura-ActividadRESUMEN
A series of 2-amino-isoxazolopyridines was designed and synthesized as Polo-like kinase (Plk) inhibitors. Key SAR and crystallographic data are discussed. More advanced analogues inhibit Plk1 with good enzymatic activity and modest cell-based activity. Differential selectivity among the three Plk isoforms is observed.
Asunto(s)
Proteínas de Ciclo Celular/antagonistas & inhibidores , Diseño de Fármacos , Isoxazoles/síntesis química , Isoxazoles/farmacología , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Proto-Oncogénicas/antagonistas & inhibidores , Piridinas/síntesis química , Piridinas/farmacología , Técnicas Químicas Combinatorias , Cristalografía por Rayos X , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Isoenzimas/antagonistas & inhibidores , Isoxazoles/química , Microsomas Hepáticos/efectos de los fármacos , Conformación Molecular , Estructura Molecular , Piridinas/química , Relación Estructura-Actividad , Quinasa Tipo Polo 1RESUMEN
A series of 2-amino-pyrazolopyridines was designed and synthesized as Polo-like kinase (Plk) inhibitors based on a low micromolar hit. The SAR was developed to provide compounds exhibiting low nanomolar inhibitory activity of Plk1; the phenotype of treated cells is consistent with Plk1 inhibition. A co-crystal structure of one of these compounds with zPlk1 confirms an ATP-competitive binding mode.
Asunto(s)
Proteínas de Ciclo Celular/antagonistas & inhibidores , Química Farmacéutica/métodos , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Proto-Oncogénicas/antagonistas & inhibidores , Pirazoles/síntesis química , Piridinas/síntesis química , Adenosina Trifosfato/química , Secuencias de Aminoácidos , Ciclo Celular , Cristalografía por Rayos X , Diseño de Fármacos , Humanos , Concentración 50 Inhibidora , Modelos Químicos , Conformación Molecular , Fenotipo , Pirazoles/química , Pirazoles/farmacología , Piridinas/química , Piridinas/farmacología , Relación Estructura-Actividad , Quinasa Tipo Polo 1RESUMEN
This communication describes the discovery of a novel series of Aurora kinase inhibitors. Key SAR and critical binding elements are discussed. Some of the more advanced analogues potently inhibit cellular proliferation and induce phenotypes consistent with Aurora kinase inhibition. In particular, compound 21 (SNS-314) is a potent and selective Aurora kinase inhibitor that exhibits significant activity in pre-clinical in vivo tumor models.
Asunto(s)
Neoplasias Experimentales/tratamiento farmacológico , Compuestos de Fenilurea/química , Compuestos de Fenilurea/farmacología , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Quinazolinas/farmacología , Tiazoles/química , Tiazoles/farmacología , Animales , Antineoplásicos/química , Antineoplásicos/farmacología , Aurora Quinasas , Ensayos de Selección de Medicamentos Antitumorales , Células HCT116 , Humanos , Ratones , Trasplante de Neoplasias , Neoplasias Experimentales/enzimología , Quinazolinas/química , Relación Estructura-ActividadRESUMEN
Polo-like kinase 1 (Plk1) is a member of the Polo-like kinase family of serine/threonine kinases involved in the regulation of cell-cycle progression and cytokinesis and is an attractive target for the development of anticancer therapeutics. The catalytic domain of this enzyme shares significant primary amino-acid homology and structural similarity with another mitotic kinase, Aurora A. While screening an Aurora A library of ATP-competitive compounds, a urea-containing inhibitor with low affinity for mouse Aurora A but with submicromolar potency for human and zebrafish Plk1 (hPlk1 and zPlk1, respectively) was identified. A crystal structure of the zebrafish Plk1 kinase domain-inhibitor complex reveals that the small molecule occupies the purine pocket and extends past the catalytic lysine into the adaptive region of the active site. Analysis of the structures of this protein-inhibitor complex and of similar small molecules cocrystallized with other kinases facilitates understanding of the specificity of the inhibitor for Plk1 and documents for the first time that Plk1 can accommodate extended ATP-competitive compounds that project toward the adaptive pocket and help the enzyme order its activation segment.
Asunto(s)
Proteínas de Ciclo Celular/química , Proteínas Serina-Treonina Quinasas/química , Proteínas Proto-Oncogénicas/química , Proteínas de Pez Cebra/química , Pez Cebra/metabolismo , Animales , Secuencia de Bases , Dominio Catalítico , Proteínas de Ciclo Celular/antagonistas & inhibidores , Proteínas de Ciclo Celular/genética , Cristalografía por Rayos X , Cartilla de ADN , Modelos Moleculares , Mutagénesis Sitio-Dirigida , Conformación Proteica , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Serina-Treonina Quinasas/genética , Proteínas Proto-Oncogénicas/antagonistas & inhibidores , Proteínas Proto-Oncogénicas/genética , Especificidad por Sustrato , Proteínas de Pez Cebra/antagonistas & inhibidores , Proteínas de Pez Cebra/genética , Quinasa Tipo Polo 1RESUMEN
The rapid advancement of a series of noncovalent inhibitors of T790M mutants of EGFR is discussed. The optimization of pyridone 1, a nonselective high-throughput screening hit, to potent molecules with high levels of selectivity over wtEGFR and the broader kinome is described herein.
RESUMEN
Inhibitors targeting the activating mutants of the epidermal growth factor receptor (EGFR) have found success in the treatment of EGFR mutant positive non-small-cell lung cancer. A secondary point mutation (T790M) in the inhibitor binding site has been linked to the acquired resistance against those first generation therapeutics. Herein, we describe the lead optimization of a series of reversible, pan-mutant (L858R, del746-750, T790M/L858R, and T790M/del746-750) EGFR inhibitors. By use of a noncovalent double mutant (T790M/L858R and T790M/del746-750) selective EGFR inhibitor (2) as a starting point, activities against the single mutants (L858R and del746-750) were introduced through a series of structure-guided modifications. The in vitro ADME-PK properties of the lead molecules were further optimized through a number of rational structural changes. The resulting inhibitor (21) exhibited excellent cellular activity against both the single and double mutants of EGFR, demonstrating target engagement in vivo and ADME-PK properties that are suitable for further evaluation. The reversible, noncovalent inhibitors described complement the covalent pan-mutant EGFR inhibitors that have shown encouraging results in recent clinical trials.
Asunto(s)
Antineoplásicos/química , Antineoplásicos/uso terapéutico , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Receptores ErbB/antagonistas & inhibidores , Neoplasias Pulmonares/tratamiento farmacológico , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/uso terapéutico , Animales , Antineoplásicos/farmacocinética , Antineoplásicos/farmacología , Carcinoma de Pulmón de Células no Pequeñas/genética , Carcinoma de Pulmón de Células no Pequeñas/patología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Cristalografía por Rayos X , Resistencia a Antineoplásicos , Receptores ErbB/genética , Humanos , Pulmón/efectos de los fármacos , Pulmón/metabolismo , Pulmón/patología , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patología , Ratones , Modelos Moleculares , Mutación , Inhibidores de Proteínas Quinasas/farmacocinética , Inhibidores de Proteínas Quinasas/farmacologíaRESUMEN
Because of their increased activity against activating mutants, first-generation epidermal growth factor receptor (EGFR) kinase inhibitors have had remarkable success in treating non-small-cell lung cancer (NSCLC) patients, but acquired resistance, through a secondary mutation of the gatekeeper residue, means that clinical responses only last for 8-14 months. Addressing this unmet medical need requires agents that can target both of the most common double mutants: T790M/L858R (TMLR) and T790M/del(746-750) (TMdel). Herein we describe how a noncovalent double mutant selective lead compound was optimized using a strategy focused on the structure-guided increase in potency without added lipophilicity or reduction of three-dimensional character. Following successive rounds of design and synthesis it was discovered that cis-fluoro substitution on 4-hydroxy- and 4-methoxypiperidinyl groups provided synergistic, substantial, and specific potency gain through direct interaction with the enzyme and/or effects on the proximal ligand oxygen atom. Further development of the fluorohydroxypiperidine series resulted in the identification of a pair of diastereomers that showed 50-fold enzyme and cell based selectivity for T790M mutants over wild-type EGFR (wtEGFR) in vitro and pathway knock-down in an in vivo xenograft model.