RESUMO
Metabolite identification is an integral part of both preclinical and clinical drug discovery and development. Synthesis of drug metabolites is often required to support definitive identification, preclinical safety studies and clinical trials. Here we describe the use of microbial biotransformation as a tool to produce drug metabolites, complementing traditional chemical synthesis and other biosynthetic methods such as hepatocytes, liver microsomes and recombinant human drug metabolizing enzymes. A workflow is discussed whereby microbial strains are initially screened for their ability to form the putative metabolites of interest, followed by a scale-up to afford quantities sufficient to perform definitive identification and further studies. Examples of the microbial synthesis of several difficult-to-synthesize hydroxylated metabolites and three difficult-to-synthesize glucuronidated metabolites are described, and the use of microbial biotransformation in drug discovery and development is discussed.
Assuntos
Bactérias/metabolismo , Preparações Farmacêuticas/metabolismo , Biotransformação , Humanos , Metaboloma , Oxirredução , Preparações Farmacêuticas/químicaRESUMO
The current standard of care for treating Alzheimer's disease is acetylcholinesterase inhibitors, which nonselectively increase cholinergic signaling by indirectly enhancing activity of nicotinic and muscarinic receptors. These drugs improve cognitive function in patients, but also produce unwanted side effects that limit their efficacy. In an effort to selectively improve cognition and avoid the cholinergic side effects associated with the standard of care, various efforts have been aimed at developing selective M1 muscarinic receptor activators. In this work, we describe the preclinical and clinical pharmacodynamic effects of the M1 muscarinic receptor-positive allosteric modulator, MK-7622. MK-7622 attenuated the cognitive-impairing effects of the muscarinic receptor antagonist scopolamine and altered quantitative electroencephalography (qEEG) in both rhesus macaque and human. For both scopolamine reversal and qEEG, the effective exposures were similar between species. However, across species the minimum effective exposures to attenuate the scopolamine impairment were lower than for qEEG. Additionally, there were differences in the spectral power changes produced by MK-7622 in rhesus versus human. In sum, these results are the first to demonstrate translation of preclinical cognition and target modulation to clinical effects in humans for a selective M1 muscarinic receptor-positive allosteric modulator.
Assuntos
Quinazolinas/farmacologia , Receptor Muscarínico M1/metabolismo , Regulação Alostérica/efeitos dos fármacos , Animais , Cognição/efeitos dos fármacos , Relação Dose-Resposta a Droga , Eletroencefalografia/efeitos dos fármacos , Humanos , Macaca mulatta , Masculino , Quinazolinas/farmacocinética , Pesquisa Translacional BiomédicaRESUMO
A series of methoxynaphthalene amides were prepared and evaluated as alternatives to quinolizidinone amide M1 positive allosteric modulators. A methoxy group was optimal for M1 activity and addressed key P-gp issues present in the aforementioned quinolizidinone amide series.
Assuntos
Amidas/química , Naftalenos/química , Quinolizidinas/química , Receptor Muscarínico M1/metabolismo , Regulação Alostérica , Amidas/síntese química , Amidas/metabolismo , Animais , Células CHO , Cricetinae , Cricetulus , Camundongos , Ligação Proteica , Receptor Muscarínico M1/química , Relação Estrutura-AtividadeRESUMO
Acinetobacter baumannii, a commonly multidrug-resistant Gram-negative bacterium responsible for large numbers of bloodstream and lung infections worldwide, is increasingly difficult to treat and constitutes a growing threat to human health. Structurally novel antibacterial chemical matter that can evade existing resistance mechanisms is essential for addressing this critical medical need. Herein, we describe our efforts to inhibit the essential A. baumannii lipooligosaccharide (LOS) ATP-binding cassette (ABC) transporter MsbA. An unexpected impurity from a phenotypic screening was optimized as a series of dimeric compounds, culminating with 1 (cerastecin D), which exhibited antibacterial activity in the presence of human serum and a pharmacokinetic profile sufficient to achieve efficacy against A. baumannii in murine septicemia and lung infection models.
RESUMO
Respiratory syncytial virus (RSV) and human metapneumovirus (HMPV) are related RNA viruses responsible for severe respiratory infections and resulting disease in infants, elderly, and immunocompromised adults1-3. Therapeutic small molecule inhibitors that bind to the RSV polymerase and inhibit viral replication are being developed, but their binding sites and molecular mechanisms of action remain largely unknown4. Here we report a conserved allosteric inhibitory site identified on the L polymerase proteins of RSV and HMPV that can be targeted by a dual-specificity, non-nucleoside inhibitor, termed MRK-1. Cryo-EM structures of the inhibitor in complexes with truncated RSV and full-length HMPV polymerase proteins provide a structural understanding of how MRK-1 is active against both viruses. Functional analyses indicate that MRK-1 inhibits conformational changes necessary for the polymerase to engage in RNA synthesis initiation and to transition into an elongation mode. Competition studies reveal that the MRK-1 binding pocket is distinct from that of a capping inhibitor with an overlapping resistance profile, suggesting that the polymerase conformation bound by MRK-1 may be distinct from that involved in mRNA capping. These findings should facilitate optimization of dual RSV and HMPV replication inhibitors and provide insights into the molecular mechanisms underlying their polymerase activities.
Assuntos
Metapneumovirus , Vírus Sincicial Respiratório Humano , Infecções Respiratórias , Lactente , Adulto , Humanos , Idoso , Metapneumovirus/genética , Metapneumovirus/metabolismo , RNA Polimerase Dependente de RNA/genética , RNA Polimerase Dependente de RNA/metabolismo , RNA MensageiroRESUMO
Increasing the efficiency of the drug discovery process is a challenge faced by drug hunters everywhere. One strategy medicinal chemists employ to meet this challenge is learning from knowledge sources within and beyond their organization. In this Perspective, we discuss the evolution of mechanisms for medicinal chemistry knowledge capture and sharing at Merck & Co. over the past 15 years. We describe our approach to knowledge management and report on the multiple enduring and complementary teams and initiatives we have created to capture and share knowledge within a geographically diverse medicinal chemistry community. In addition, this Perspective will share the benefits we have observed and also reflect on what has allowed our efforts to be both successful and sustainable.
Assuntos
Química Farmacêutica , Descoberta de DrogasRESUMO
Although current antiretroviral therapy can control HIV-1 replication and prevent disease progression, it is not curative. Identifying mechanisms that can lead to eradication of persistent viral reservoirs in people living with HIV-1 (PLWH) remains an outstanding challenge to achieving cure. Utilizing a phenotypic screen, we identified a novel chemical class capable of killing HIV-1 infected peripheral blood mononuclear cells. Tool compounds ICeD-1 and ICeD-2 ("inducer of cell death-1 and 2"), optimized for potency and selectivity from screening hits, were used to deconvolute the mechanism of action using a combination of chemoproteomic, biochemical, pharmacological, and genetic approaches. We determined that these compounds function by modulating dipeptidyl peptidase 9 (DPP9) and activating the caspase recruitment domain family member 8 (CARD8) inflammasome. Efficacy of ICeD-1 and ICeD-2 was dependent on HIV-1 protease activity and synergistic with efavirenz, which promotes premature activation of HIV-1 protease at high concentrations in infected cells. This in vitro synergy lowers the efficacious cell kill concentration of efavirenz to a clinically relevant dose at concentrations of ICeD-1 or ICeD-2 that do not result in complete DPP9 inhibition. These results suggest engagement of the pyroptotic pathway as a potential approach to eliminate HIV-1 infected cells.
Assuntos
Infecções por HIV , HIV-1 , Alcinos , Benzoxazinas , Proteínas Adaptadoras de Sinalização CARD/metabolismo , Ciclopropanos , Dipeptidil Peptidases e Tripeptidil Peptidases/metabolismo , Infecções por HIV/tratamento farmacológico , HIV-1/metabolismo , Humanos , Inflamassomos/metabolismo , Leucócitos Mononucleares , Proteínas de Neoplasias/metabolismoRESUMO
A novel series of histone deacetylase (HDAC) inhibitors lacking a zinc-binding moiety has been developed and described herein. HDAC isozyme profiling and kinetic studies indicate that these inhibitors display a selectivity preference for HDACs 1, 2, 3, 10, and 11 via a rapid equilibrium mechanism, and crystal structures with HDAC2 confirm that these inhibitors do not interact with the catalytic zinc. The compounds are nonmutagenic and devoid of electrophilic and mutagenic structural elements and exhibit off-target profiles that are promising for further optimization. The efficacy of this new class in biochemical and cell-based assays is comparable to the marketed HDAC inhibitors belinostat and vorinostat. These results demonstrate that the long-standing pharmacophore model of HDAC inhibitors requiring a metal binding motif should be revised and offers a distinct class of HDAC inhibitors.
RESUMO
Incorporation of hydroxycycloalkyl fused pyridone carboxylic acids in lieu of quinolone carboxylic acids enhance free fraction without increased susceptibility to P-glycoprotein transport.
Assuntos
Ácidos Carboxílicos/farmacologia , Piridonas/química , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Regulação Alostérica , Animais , Ácidos Carboxílicos/química , Camundongos , Transporte Proteico , RatosRESUMO
A series of triarylethanolamine inhibitors of the Kv1.5 potassium channel have been prepared and evaluated for their effects in vitro and in vivo. The structure-activity relationship (SAR) studies described herein led to the development of potent, selective and orally active inhibitors of Kv1.5.
Assuntos
Etanolaminas/farmacologia , Bloqueadores dos Canais de Potássio/farmacologia , Canais de Potássio de Abertura Dependente da Tensão da Membrana/antagonistas & inibidores , Animais , Relação Dose-Resposta a Droga , Descoberta de Drogas , Avaliação Pré-Clínica de Medicamentos , Etanolaminas/química , Humanos , Bloqueadores dos Canais de Potássio/química , Relação Estrutura-AtividadeRESUMO
A full account of the enantioselective total syntheses of (+)-lyconadin A (1) and (-)-lyconadin B (2) is presented. Central to this venture was recognition and deployment of a key strategy-level intramolecular aldol/conjugate addition cascade that led, in a single operation, to two new carbon-carbon sigma-bonds, three new stereogenic centers, and two new rings, albeit with the incorrect stereogenicity at C(12) for the lyconadins. Correction of the C(12) stereogenicity was achieved via innovative use of a protonated intramolecular aminal. An aminoiodo olefin cyclization, in conjunction with alpha-pyridinone and 3,4-dihydropyridinone annulation protocols, permitted completion of the syntheses of (+)-lyconadin A (1) and (-)-lyconadin B (2), respectively.
Assuntos
Alcaloides/síntese química , Compostos Policíclicos/síntese química , Aldeídos/química , Alcaloides/química , Hidrazonas/química , Iodetos/química , Estrutura Molecular , Compostos Policíclicos/química , EstereoisomerismoRESUMO
Drug discovery efforts have focused recently on atrial-selective targets, including the Kv1.5 channel, which underlies the ultrarapid delayed rectifier current, I(Kur), to develop novel treatments for atrial fibrillation (AF). Two structurally distinct compounds, a triarylethanolamine TAEA and an isoquinolinone 3-[(dimethylamino)-methyl]-6-methoxy-2-methyl-4-phenylisoquinolin-1(2H)-one (ISQ-1), blocked I(Kur) in Chinese hamster ovary cells expressing human Kv1.5 with IC(50) values of 238 and 324 nM, respectively. In anesthetized dogs, i.v. infusions of TAEA and ISQ-1 elicited comparable 16% increases in atrial refractory period, with no effect on ventricular refractory period or QTc interval. Plasma concentrations at end infusion for TAEA and ISQ-1 were 58.5 +/- 23.6 and 330.3 +/- 43.5 nM, respectively. The abilities of TAEA and ISQ-1 to terminate AF, with comparison to the rapidly activating component of delayed rectifier potassium current blocker (+)-N-[1'-(6-cyano-1,2,3,4-tetrahydro-2(R)-naphthalenyl)-3,4-dihydro-4(R)-hydroxyspiro(2H-1-benzopyran-2,4'-piperidin)-6-yl]methanesulfonamide] monohydrochloride (MK-499) and the class IC 1-[2-[2-hydroxy-3-(propylamino)-propoxy]phenyl]-3-phenyl-1-propanone (propafenone), were assessed in conscious dogs with heart failure and inducible AF (entry criterion). All test agents administered in i.v. bolus regimens terminated AF in at least half of animals tested; conversely no agent was universally effective. MK-499, ISQ-1, TAEA, and propafenone terminated AF in five of six, four of seven, four of six, and five of six animals at plasma concentrations of 32.6 +/- 18.7, 817 +/- 274, 714 +/- 622, and 816 +/- 240 nM, respectively. Directed cardiac electrophysiologic studies in anesthetized dogs using i.v. bolus (consistent with AF studies) plus infusion regimens with TAEA and ISQ-1 demonstrated significant increases in atrial refractory period (12-15%), A-H and P-A intervals, but no effects on ventricular refractory period, H-V, and HEG intervals. The demonstration of AF termination with TAEA and ISQ-1 in the dog heart failure model extends the profile of antiarrhythmic efficacy of Kv1.5 blockade.
Assuntos
Antiarrítmicos/uso terapêutico , Fibrilação Atrial/tratamento farmacológico , Insuficiência Cardíaca/tratamento farmacológico , Isoquinolinas/uso terapêutico , Canal de Potássio Kv1.5/antagonistas & inibidores , Bloqueadores dos Canais de Potássio/uso terapêutico , Piridinas/uso terapêutico , Animais , Fibrilação Atrial/fisiopatologia , Benzopiranos/uso terapêutico , Linhagem Celular , Cães , Feminino , Átrios do Coração/efeitos dos fármacos , Átrios do Coração/fisiopatologia , Humanos , Masculino , Piperidinas/uso terapêutico , Propafenona/uso terapêutico , Bloqueadores dos Canais de Sódio/uso terapêuticoRESUMO
Identification of ligands that selectively activate the M1 muscarinic signaling pathway has been sought for decades to treat a range of neurological and cognitive disorders. Herein, we describe the optimization efforts focused on addressing key physicochemical and safety properties, ultimately leading to the clinical candidate MK-7622, a highly selective positive allosteric modulator of the M1 muscarinic receptor that has entered Phase II studies in patients with Alzheimer's disease.
RESUMO
Agonist shift assays feature cross-titrations of allosteric modulators and orthosteric ligands. Information generated in agonist shift assays can include a modulator's effect on the orthosteric agonist's potency (alpha) and efficacy (beta), as well as direct agonist activity of the allosteric ligand (tauB) and the intrinsic binding affinity of the modulator to the unoccupied receptor (KB). Because of the heavy resource demand and complex data handling, these allosteric parameters are determined infrequently during the course of a drug discovery program and on a relatively small subset of compounds. Automation of agonist shift assays enables this data-rich analysis to evaluate a larger number of compounds, offering the potential to differentiate compound classes earlier and prospectively prioritize based on desired molecular pharmacology. A high-throughput calcium-imaging agonist shift assay was pursued to determine the allosteric parameters of over 1000 positive allosteric modulator (PAM) molecules for the human muscarinic acetylcholine receptor 1 (M1). Control compounds were run repeatedly to demonstrate internal consistency. Comparisons between potency measurements and the allosteric parameter results demonstrate that these different types of measurements do not necessarily correlate, highlighting the importance of fully characterizing and understanding the allosteric properties of leads.
Assuntos
Ensaios de Triagem em Larga Escala/métodos , Receptor Muscarínico M1/metabolismo , Acetilcolina/farmacologia , Regulação Alostérica/efeitos dos fármacos , Animais , Automação , Células CHO , Cricetinae , Cricetulus , Receptor Muscarínico M1/agonistas , Receptor Muscarínico M1/química , Reprodutibilidade dos TestesRESUMO
High-throughput screening (HTS) has enabled millions of compounds to be assessed for biological activity, but challenges remain in the prioritization of hit series. While biological, absorption, distribution, metabolism, excretion, and toxicity (ADMET), purity, and structural data are routinely used to select chemical matter for further follow-up, the scarcity of historical ADMET data for screening hits limits our understanding of early hit compounds. Herein, we describe a process that utilizes a battery of in-house quantitative structure-activity relationship (QSAR) models to generate in silico ADMET profiles for hit series to enable more complete characterizations of HTS chemical matter. These profiles allow teams to quickly assess hit series for desirable ADMET properties or suspected liabilities that may require significant optimization. Accordingly, these in silico data can direct ADMET experimentation and profoundly impact the progression of hit series. Several prospective examples are presented to substantiate the value of this approach.
Assuntos
Descoberta de Drogas/métodos , Ensaios de Triagem em Larga Escala/métodos , Preparações Farmacêuticas/química , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Animais , Simulação por Computador , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos , Humanos , Preparações Farmacêuticas/metabolismo , Farmacocinética , Farmacologia , Relação Quantitativa Estrutura-AtividadeRESUMO
Novel 3-cyanoisoquinoline Kv1.5 antagonists have been prepared and evaluated in in vitro and in vivo assays for inhibition of the Kv1.5 potassium channel and its associated cardiac potassium current, IKur. Structural modifications of isoquinolinone lead 1 afforded compounds with excellent potency, selectivity, and oral bioavailability.
Assuntos
Antiarrítmicos/síntese química , Fibrilação Atrial/tratamento farmacológico , Isoquinolinas/síntese química , Canal de Potássio Kv1.5/antagonistas & inibidores , Nitrilas/síntese química , Administração Oral , Animais , Antiarrítmicos/química , Antiarrítmicos/farmacologia , Disponibilidade Biológica , Eletrofisiologia , Coração/efeitos dos fármacos , Coração/fisiologia , Humanos , Isoquinolinas/química , Isoquinolinas/farmacologia , Canal de Potássio Kv1.5/fisiologia , Nitrilas/química , Nitrilas/farmacologia , Técnicas de Patch-Clamp , Ratos , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
A series of macrocyclic 3-aminopyrrolidinone farnesyltransferase inhibitors (FTIs) has been synthesized. Compared with previously described linear 3-aminopyrrolidinone FTIs such as compound 1, macrocycles such as 49 combined improved pharmacokinetic properties with a reduced potential for side effects. In dogs, oral bioavailability was good to excellent, and increases in plasma half-life were due to attenuated clearance. It was observed that in vivo clearance correlated with the flexibility of the molecules and this concept proved useful in the design of FTIs that exhibited low clearance, such as FTI 78. X-ray crystal structures of compounds 49 and 66 complexed with farnesyltransferase (FTase)-farnesyl diphosphate (FPP) were determined, and they provide details of the key interactions in such ternary complexes. Optimization of this 3-aminopyrrolidinone series of compounds led to significant increases in potency, providing 83 and 85, the most potent inhibitors of FTase in cells described to date.
Assuntos
Alquil e Aril Transferases/antagonistas & inibidores , Hidrocarboneto de Aril Hidroxilases , Proteínas de Transporte de Cátions , Proteínas de Ligação a DNA , Inibidores Enzimáticos/síntese química , Naftalenos/síntese química , Canais de Potássio de Abertura Dependente da Tensão da Membrana , Pirrolidinas/síntese química , Transativadores , Animais , Linhagem Celular , Cromatografia Líquida , Cristalografia por Raios X , Citocromo P-450 CYP3A , Inibidores das Enzimas do Citocromo P-450 , Cães , Canal de Potássio ERG1 , Eletrocardiografia , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacocinética , Canais de Potássio Éter-A-Go-Go , Farnesiltranstransferase , Humanos , Técnicas In Vitro , Espectroscopia de Ressonância Magnética , Espectrometria de Massas , Microssomos Hepáticos/efeitos dos fármacos , Microssomos Hepáticos/enzimologia , Modelos Moleculares , Estrutura Molecular , Naftalenos/química , Naftalenos/farmacocinética , Oxirredutases N-Desmetilantes/antagonistas & inibidores , Canais de Potássio/metabolismo , Ligação Proteica , Pirrolidinas/química , Pirrolidinas/farmacocinética , Estereoisomerismo , Relação Estrutura-Atividade , Regulador Transcricional ERGRESUMO
[reaction: see text] A one-pot, tandem reductive amination-transamidation-cyclization reaction was employed to produce substituted piperazin-2-ones in good yields. Various amino acid methyl esters and transferable acyl groups were examined to establish the reaction's scope.
Assuntos
Piperazinas/síntese química , Aminação , Ciclização , Modelos Moleculares , Conformação Molecular , OxirreduçãoRESUMO
There is mounting evidence from preclinical and early proof-of-concept studies suggesting that selective modulation of the M1 muscarinic receptor is efficacious in cognitive models of Alzheimer's disease (AD). A number of nonselective M1 muscarinic agonists have previously shown positive effects on cognitive function in AD patients, but were limited due to cholinergic adverse events thought to be mediated by pan activation of the M2 to M5 sub-types. Thus, there is a need to identify selective activators of the M1 receptor to evaluate their potential in cognitive disorders. One strategy to confer selectivity for M1 is the identification of allosteric agonists or positive allosteric modulators, which would target an allosteric site on the M1 receptor rather than the highly conserved orthosteric acetylcholine binding site. BQCA has been identified as a highly selective carboxylic acid M1 PAM and this review focuses on an extensive lead optimization campaign undertaken on this compound.
Assuntos
Doença de Alzheimer/tratamento farmacológico , Ácidos Carboxílicos/química , Agonistas Muscarínicos/química , Quinolonas/química , Receptor Muscarínico M1/agonistas , Bibliotecas de Moléculas Pequenas/química , Regulação Alostérica , Sítio Alostérico , Doença de Alzheimer/fisiopatologia , Animais , Ácidos Carboxílicos/farmacologia , Cognição/efeitos dos fármacos , Modelos Animais de Doenças , Humanos , Camundongos , Agonistas Muscarínicos/farmacologia , Quinolonas/farmacologia , Receptor Muscarínico M1/química , Bibliotecas de Moléculas Pequenas/farmacologia , Relação Estrutura-Atividade , Especificidade por SubstratoRESUMO
INTRODUCTION: There is substantial evidence from preclinical and early proof-of-concept studies suggesting that selective modulation of the M(1) muscarinic receptor is efficacious in cognitive models of Alzheimer's disease (AD) and antipsychotic models of schizophrenia. For example, a number of nonselective M(1) muscarinic agonists have previously shown positive effects on cognitive function in AD patients, but were limited due to cholinergic adverse events thought to be mediated by pan activation of the M(2) to M(5) subtypes. Thus, there is a need to identify selective activators of the M(1) receptor to evaluate their potential in cognitive disorders. One strategy to confer selectivity for M(1) is the identification of allosteric agonists or positive allosteric modulators, which would target an allosteric site on the M(1) receptor rather than the highly conserved orthosteric acetylcholine binding site. AREAS COVERED: This review discusses the M(1) muscarinic receptor and its potential therapeutic value in the treatment of CNS disorders such as AD and schizophrenia. Specifically, novel allosteric ligands that activate or positively modulate the M(1) receptor are examined and peer-reviewed articles associated with these patents publications are also described. EXPERT OPINION: There is substantial evidence supporting activation of the M(1) receptor might be effective in treating symptoms of AD and schizophrenia, but therapeutic success has been elusive and is hypothesized to be due to the lack of selectivity among orthosteric agonists. During the past decade, allosteric modulation of GPCRs has evolved as a viable strategy toward generating subtype selective molecules. A number of novel, selective ligands in the form of allosteric agonists and positive allosteric modulators of the M(1) receptor have been identified offering the potential for clinical evaluation of M(1)-specific receptor activation.