ABSTRACT
New antibiotics are urgently needed to address increasing rates of multidrug resistant infections. Seventy-six diversely functionalized compounds, comprising five structural scaffolds, were synthesized and tested for their ability to inhibit microbial growth. Twenty-six compounds showed activity in the primary phenotypic screen at the Community for Open Antimicrobial Drug Discovery (CO-ADD). Follow-up testing of active molecules confirmed that two unnatural dipeptides inhibit the growth of Cryptococcus neoformans with a minimum inhibitory concentration (MIC) ≤ 8 µg/mL. Syntheses were carried out by undergraduate students at five schools implementing Distributed Drug Discovery (D3) programs. This report showcases that a collaborative research and educational process is a powerful approach to discover new molecules inhibiting microbial growth. Educational gains for students engaged in this project are highlighted in parallel to the research advances. Aspects of D3 that contribute to its success, including an emphasis on reproducibility of procedures, are discussed to underscore the power of this approach to solve important research problems and to inform other coupled chemical biology research and teaching endeavors.
Subject(s)
Anti-Infective Agents/pharmacology , Education/organization & administration , Interinstitutional Relations , Organizational Affiliation , Humans , Microbial Sensitivity Tests , Reproducibility of ResultsABSTRACT
Kynurenic acid (KYNA) plays a significant role in maintaining normal brain function, and abnormalities in KYNA levels have been associated with various central nervous system disorders. Confirmation of its causality in human diseases requires safe and effective modulation of central KYNA levels in the clinic. The kynurenine aminotransferases (KAT) II enzyme represents an attractive target for pharmacologic modulation of central KYNA levels; however, KAT II and KYNA turnover kinetics, which could contribute to the duration of pharmacologic effect, have not been reported. In this study, the kinetics of central KYNA-lowering effect in rats and nonhuman primates (NHPs, Cynomolgus macaques) was investigated using multiple KAT II irreversible inhibitors as pharmacologic probes. Mechanistic pharmacokinetic-pharmacodynamic analysis of in vivo responses to irreversible inhibition quantitatively revealed that 1) KAT II turnover is relatively slow [16-76 hours' half-life (t1/2)], whereas KYNA is cleared more rapidly from the brain (<1 hour t1/2) in both rats and NHPs, 2) KAT II turnover is slower in NHPs than in rats (76 hours vs. 16 hours t1/2, respectively), and 3) the percent contribution of KAT II to KYNA formation is constant (â¼80%) across rats and NHPs. Additionally, modeling results enabled establishment of in vitro-in vivo correlation for both enzyme turnover rates and drug potencies. In summary, quantitative translational analysis confirmed the feasibility of central KYNA modulation in humans. Model-based analysis, where system-specific properties and drug-specific properties are mechanistically separated from in vivo responses, enabled quantitative understanding of the KAT II-KYNA pathway, as well as assisted development of promising candidates to test KYNA hypothesis in humans.
Subject(s)
Brain/metabolism , Enzyme Inhibitors/administration & dosage , Kynurenic Acid/analysis , Transaminases/metabolism , Animals , Brain Chemistry/drug effects , Cells, Cultured , Chromatography, Liquid , Enzyme Inhibitors/pharmacology , Female , Half-Life , Humans , Macaca fascicularis , Male , Pyrazoles/administration & dosage , Pyrazoles/pharmacology , Rats , Tandem Mass Spectrometry , Transaminases/antagonists & inhibitorsABSTRACT
In 2014, a published report of the high-throughput screen of>42,000 kinase inhibitors from GlaxoSmithKline against T. brucei identified 797 potent and selective hits. From this rich data set, we selected NEU-0001101 (1) for hit-to-lead optimization. Through our preliminary compound synthesis and SAR studies, we have confirmed the previously reported activity of 1 in a T. brucei cell proliferation assay and have identified alternative groups to replace the pyridyl ring in 1. Pyrazole 24 achieves improvements in both potency and lipophilicity relative to 1, while also showing good in vitro metabolic stability. The SAR developed on 24 provides new directions for further optimization of this novel scaffold for anti-trypanosomal drug discovery.
Subject(s)
Drug Design , Pyrazoles/chemical synthesis , Quinazolinones/chemistry , Trypanocidal Agents/chemical synthesis , Cell Line , Cell Proliferation/drug effects , Humans , Pyrazoles/pharmacology , Pyrazoles/therapeutic use , Quinazolinones/chemical synthesis , Quinazolinones/pharmacology , Quinazolinones/therapeutic use , Structure-Activity Relationship , Trypanocidal Agents/pharmacology , Trypanocidal Agents/therapeutic use , Trypanosoma brucei brucei/drug effects , Trypanosoma brucei brucei/growth & development , Trypanosomiasis, African/drug therapyABSTRACT
The kynurenine pathway is responsible for the metabolism of more than 95% of dietary tryptophan (TRP) and produces numerous bioactive metabolites. Recent studies have focused on three enzymes in this pathway: indoleamine dioxygenase (IDO1), kynurenine monooxygenase (KMO), and kynurenine aminotransferase II (KAT II). IDO1 inhibitors are currently in clinical trials for the treatment of cancer, and these agents may also have therapeutic utility in neurological disorders, including multiple sclerosis. KMO inhibitors are being investigated as potential treatments for neurodegenerative diseases, such as Huntington's and Alzheimer's diseases. KAT II inhibitors have been proposed in new therapeutic approaches toward psychiatric and cognitive disorders, including cognitive impairment associated with schizophrenia. Numerous medicinal chemistry studies are currently aimed at the design of novel, potent, and selective inhibitors for each of these enzymes. The emerging opportunities and significant challenges associated with pharmacological modulation of these enzymes will be explored in this review.
Subject(s)
Kynurenine/metabolism , Signal Transduction/drug effects , Animals , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/pharmacology , Central Nervous System Diseases/drug therapy , Enzyme Inhibitors/pharmacology , Enzyme Inhibitors/therapeutic use , Humans , Indoleamine-Pyrrole 2,3,-Dioxygenase/antagonists & inhibitors , Kynurenine 3-Monooxygenase/antagonists & inhibitors , Transaminases/antagonists & inhibitorsABSTRACT
The elevation of kynurenic acid (KYNA) observed in schizophrenic patients may contribute to core symptoms arising from glutamate hypofunction, including cognitive impairments. Although increased KYNA levels reduce excitatory neurotransmission, KYNA has been proposed to act as an endogenous antagonist at the glycine site of the glutamate NMDA receptor (NMDAR) and as a negative allosteric modulator at the α7 nicotinic acetylcholine receptor. Levels of KYNA are elevated in CSF and the postmortem brain of schizophrenia patients, and these elevated levels of KYNA could contribute to NMDAR hypofunction and the cognitive deficits and negative symptoms associated with this disease. However, the impact of endogenously produced KYNA on brain function and behavior is less well understood due to a paucity of pharmacological tools. To address this issue, we identified PF-04859989, a brain-penetrable inhibitor of kynurenine aminotransferase II (KAT II), the enzyme responsible for most brain KYNA synthesis. In rats, systemic administration of PF-04859989 dose-dependently reduced brain KYNA to as little as 28% of basal levels, and prevented amphetamine- and ketamine-induced disruption of auditory gating and improved performance in a sustained attention task. It also prevented ketamine-induced disruption of performance in a working memory task and a spatial memory task in rodents and nonhuman primates, respectively. Together, these findings support the hypotheses that endogenous KYNA impacts cognitive function and that inhibition of KAT II, and consequent lowering of endogenous brain KYNA levels, improves cognitive performance under conditions considered relevant for schizophrenia.
Subject(s)
Brain/metabolism , Cognition/physiology , Kynurenic Acid/metabolism , Schizophrenia/cerebrospinal fluid , Schizophrenia/pathology , Animals , Attention/drug effects , Attention/physiology , Enzyme Inhibitors/pharmacology , Evoked Potentials, Auditory/drug effects , Evoked Potentials, Auditory/physiology , Female , Hippocampus/cytology , Humans , Macaca mulatta , Male , Maze Learning/drug effects , Maze Learning/physiology , Memory, Short-Term/drug effects , Memory, Short-Term/physiology , Neurons/drug effects , Neurons/physiology , Pyrazoles/pharmacology , Rats , Rats, Long-Evans , Rats, Sprague-Dawley , WakefulnessABSTRACT
The structure-based design, synthesis, and biological evaluation of a new pyrazole series of irreversible KAT II inhibitors are described herein. The modification of the inhibitor scaffold of 1 and 2 from a dihydroquinolinone core to a tetrahydropyrazolopyridinone core led to discovery of a new series of potent KAT II inhibitors with excellent physicochemical properties. Compound 20 is the most potent and lipophilically efficient of these new pyrazole analogs, with a k(inact)/K(i) value of 112,000 M(-1)s(-1) and lipophilic efficiency (LipE) of 8.53. The X-ray crystal structure of 20 with KAT II demonstrates key features that contribute to this remarkable potency and binding efficiency.
Subject(s)
Drug Design , Enzyme Inhibitors/pharmacology , Pyrazoles/pharmacology , Transaminases/antagonists & inhibitors , Catalytic Domain/drug effects , Crystallography, X-Ray , Dose-Response Relationship, Drug , Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/chemistry , Humans , Hydrophobic and Hydrophilic Interactions/drug effects , Models, Molecular , Molecular Structure , Pyrazoles/chemical synthesis , Pyrazoles/chemistry , Structure-Activity Relationship , Transaminases/metabolismABSTRACT
A series of aryl hydroxamates recently have been disclosed as irreversible inhibitors of kynurenine amino transferase II (KAT II), an enzyme that may play a role in schizophrenia and other psychiatric and neurological disorders. The utilization of structure-activity relationships (SAR) in conjunction with X-ray crystallography led to the discovery of hydroxamate 4, a disubstituted analogue that has a significant potency enhancement due to a novel interaction with KAT II. The use of k inact/K i to assess potency was critical for understanding the SAR in this series and for identifying compounds with improved pharmacodynamic profiles.
ABSTRACT
Evidence suggests that compounds possessing both norepinephrine reuptake inhibition and 5-HT(1A) partial agonism (NRI/5-HT(1A)) activities may have a greater efficacy in treating neuropsychiatric disorders than compounds possessing either activity alone. The objectives of the present study were first to characterize the pharmacokinetic/pharmacodynamic (PK/PD) relationship of the plasma concentrations of atomoxetine (NRI) and buspirone (5-HT(1A) partial agonist), administered alone and in combination, on the prefrontal cortex dopamine levels in rats, and second to use the model developed to characterize the PK/PD relationship of novel NRI/5-HT(1A) compounds, PF-04269339 and PF-03529936, in a NRI/5-HT(1A) drug discovery program. Maximal dopamine elevation was twofold higher after administration of atomoxetine and buspirone in combination, PF-04269339, or PF-03529936 than after administration of atomoxetine or buspirone alone. A mechanism-based extended indirect response model characterized the time profiles of the prefrontal cortex dopamine response to atomoxetine and buspirone, administered alone or in combination. After fixing three mechanism-specific pharmacodynamic parameters (I (max) and γ2 for NRI and γ1 for 5-HT(1A)) based on the model for atomoxetine and/or buspirone, the model fitted the exposure-response profiles of PF-04269339 and PF-03529936 well. Good in vitro-to-in vivo correlation was demonstrated with the compound-specific pharmacodynamic parameters (IC(50) for NRI and SC(50) and S (max) for 5-HT(1A)) across the compounds. In summary, a piecewise modeling approach was used successfully for the characterization of the PK/PD relationship of novel NRI/5-HT(1A) compounds on prefrontal cortex dopamine levels in rats. The application and value of the mechanism-based modeling in the dual pharmacology drug discovery program are also discussed.
Subject(s)
Adrenergic Uptake Inhibitors/pharmacokinetics , Dopamine/metabolism , Drug Partial Agonism , Models, Biological , Norepinephrine/antagonists & inhibitors , Prefrontal Cortex/metabolism , Serotonin 5-HT1 Receptor Agonists/pharmacokinetics , Animals , Atomoxetine Hydrochloride , Buspirone/pharmacokinetics , Dose-Response Relationship, Drug , Humans , Male , Norepinephrine/metabolism , Prefrontal Cortex/drug effects , Propylamines/pharmacokinetics , Rats , Rats, Sprague-DawleyABSTRACT
Kynurenine aminotransferase (KAT) II has been identified as a potential new target for the treatment of cognitive impairment associated with schizophrenia and other psychiatric disorders. Following a high-throughput screen, cyclic hydroxamic acid PF-04859989 was identified as a potent and selective inhibitor of human and rat KAT II. An X-ray crystal structure and (13)C NMR studies of PF-04859989 bound to KAT II have demonstrated that this compound forms a covalent adduct with the enzyme cofactor, pyridoxal phosphate (PLP), in the active site. In vivo pharmacokinetic and efficacy studies in rat show that PF-04859989 is a brain-penetrant, irreversible inhibitor and is capable of reducing brain kynurenic acid by 50% at a dose of 10 mg/kg (sc). Preliminary structure-activity relationship investigations have been completed and have identified the positions on this scaffold best suited to modification for further optimization of this novel series of KAT II inhibitors.
ABSTRACT
We describe a generalized approach to stereocontrolled synthesis of substituted cyclic hydroxamic acids (3-amino-1-hydroxy-3,4-dihydroquinolinones) by selective reduction of substituted 2-nitrophenylalanine substrates. Compounds in this series have antibacterial properties and have also recently been reported as KAT II inhibitors. The key nitrophenyl alanine intermediates are prepared enantioselectively in excellent yield by phase transfer catalyzed alkylation of the corresponding nitrobenzyl bromides. The scope and limitations of the reductive cyclization transformation have been explored with attention to the effects of substitution pattern and electronics on reaction efficiency and byproduct formation. In addition, a novel activated trifluoroethyl ester cyclization strategy has been developed as an alternate approach to the most sterically demanding systems in this series.
Subject(s)
Hydroxamic Acids/chemistry , Hydroxamic Acids/chemical synthesis , Nitro Compounds/chemistry , Cyclization , Esters , Oxidation-Reduction , Phenylalanine/analogs & derivatives , Phenylalanine/chemical synthesis , Phenylalanine/chemistry , Sodium Acetate/chemistry , Substrate Specificity , Tin Compounds/chemistryABSTRACT
Compounds with combined norepinephrine reuptake inhibitor (NRI) and serotonin 1A (5-HT(1A)) partial agonist pharmacology may offer a new therapeutic approach for treating symptoms of neuropsychiatric disorders including ADHD, depression, and anxiety. Herein we describe the design and optimization of novel chemical matter that exhibits favorable dual NRI and 5-HT(1A) partial agonist activity. Lead compounds in this series were found to be devoid of activity at the dopamine transporter and were shown to be brain penetrant with high receptor occupancy.
Subject(s)
Adrenergic Uptake Inhibitors/chemistry , Adrenergic Uptake Inhibitors/pharmacology , Norepinephrine/metabolism , Receptor, Serotonin, 5-HT1A/metabolism , Serotonin 5-HT1 Receptor Agonists/chemistry , Serotonin 5-HT1 Receptor Agonists/pharmacology , Adrenergic Uptake Inhibitors/pharmacokinetics , Animals , Anxiety/drug therapy , Attention Deficit Disorder with Hyperactivity/drug therapy , Azetidines/chemistry , Azetidines/pharmacokinetics , Azetidines/pharmacology , Brain/metabolism , Depressive Disorder/drug therapy , Dogs , Humans , Pyrrolidines/chemistry , Pyrrolidines/pharmacokinetics , Pyrrolidines/pharmacology , Serotonin 5-HT1 Receptor Agonists/pharmacokineticsABSTRACT
Preclinical studies suggest that compounds with dual norepinephrine reuptake inhibitor (NRI) and 5-HT(1A) partial agonist properties may provide an important new therapeutic approach to ADHD, depression, and anxiety. Reported herein is the discovery of a novel chemical series with a favorable NRI and 5-HT(1A) partial agonist pharmacological profile as well as excellent selectivity for the norepinephrine transporter over the dopamine transporter.
Subject(s)
Adrenergic Uptake Inhibitors/chemical synthesis , Drug Design , Norepinephrine Plasma Membrane Transport Proteins/antagonists & inhibitors , Pyridines/chemical synthesis , Serotonin 5-HT1 Receptor Agonists , Serotonin Receptor Agonists/chemical synthesis , Adrenergic Uptake Inhibitors/metabolism , Adrenergic Uptake Inhibitors/pharmacology , Cell Line , Crystallography, X-Ray , Drug Evaluation, Preclinical/methods , Humans , Microsomes, Liver/drug effects , Microsomes, Liver/metabolism , Norepinephrine Plasma Membrane Transport Proteins/metabolism , Phenols/chemical synthesis , Phenols/metabolism , Phenols/pharmacology , Pyridines/metabolism , Pyridines/pharmacology , Receptor, Serotonin, 5-HT1A/metabolism , Serotonin Receptor Agonists/metabolism , Serotonin Receptor Agonists/pharmacologyABSTRACT
Compounds that are both norepinephrine reuptake inhibitors (NRI) and 5-HT1(A) partial agonists may have the potential to treat neuropsychiatric disorders including attention deficit hyperactivity disorder (ADHD) and depression. Targeted screening of NRI-active compounds for binding to the 5-HT(1A) receptor provided a series of thiomorpholinone hits with this dual activity profile. Several iterations of design, synthesis, and testing led to substituted piperidine diphenyl ethers which are potent NRIs with 5-HT1(A) partial agonist properties. In addition, optimization of these molecules provided compounds which exhibit selectivity for NRI over the dopamine (DAT) and serotonin (SERT) reuptake transporters. Monoamine and 5-HT(1A) in vitro functional activities for select compounds from the developed piperidine diphenyl ether series are also presented.
Subject(s)
Drug Discovery , Ethers/pharmacology , Neurotransmitter Uptake Inhibitors/pharmacology , Piperazines/pharmacology , Piperidines/pharmacology , Serotonin 5-HT1 Receptor Agonists , Dopamine/metabolism , Ethers/chemical synthesis , Ethers/chemistry , Molecular Structure , Neurotransmitter Uptake Inhibitors/chemical synthesis , Neurotransmitter Uptake Inhibitors/chemistry , Piperazines/chemical synthesis , Piperazines/chemistry , Piperidines/chemical synthesis , Piperidines/chemistry , Serotonin Plasma Membrane Transport Proteins/metabolism , Stereoisomerism , Structure-Activity RelationshipABSTRACT
Aminopyrimidine 2 (4-(1-(2-(1H-indol-3-yl)ethyl)piperidin-3-yl)-N-cyclopropylpyrimidin-2-amine) emerged from a high throughput screen as a novel 5-HT(1A) agonist. This compound showed moderate potency for 5-HT(1A) in binding and functional assays, as well as moderate metabolic stability. Implementation of a strategy for improving metabolic stability by lowering the lipophilicity (cLogD) led to identification of methyl ether 31 (4-(1-(2-(1H-indol-3-yl)ethyl)piperidin-3-yl)-N-(2-methoxyethyl)pyrimidin-2-amine) as a substantially improved compound within the series.
Subject(s)
Indoles/chemical synthesis , Indoles/pharmacology , Microsomes, Liver/drug effects , Pyrimidines/chemical synthesis , Pyrimidines/pharmacology , Receptor, Serotonin, 5-HT1A/drug effects , Serotonin Receptor Agonists/chemical synthesis , Serotonin Receptor Agonists/pharmacology , Buspirone/pharmacology , Combinatorial Chemistry Techniques , Drug Design , Humans , Indoles/chemistry , Microsomes, Liver/metabolism , Molecular Structure , Pyrimidines/chemistry , Serotonin Receptor Agonists/chemistry , Structure-Activity RelationshipABSTRACT
A 1,2,3,4-tetrahydro-9a,4a-(iminoethano)-9H-carbazole (4) is a central structural feature of the Strychnos alkaloid minfiensine (1) and akuammiline alkaloids such as vincorine (5) and echitamine (6). A cascade catalytic asymmetric Heck-iminium cyclization was developed that rapidly provides 3,4-dihydro-9a,4a-(iminoethano)-9H-carbazoles in high enantiomeric purity. Two sequences were developed for advancing 3,4-dihydro-9a,4a-(iminoethano)-9H-carbazole 27 to (+)-minfiensine. In our first-generation approach, a reductive Heck cyclization was employed to form the fifth ring of (+)-minfiensine. In a second more concise total synthesis, an intramolecular palladium-catalyzed ketone enolate vinyl iodide coupling was employed to construct the final ring of (+)-minfiensine. This second-generation total synthesis of enantiopure (+)-minfiensine was accomplished in 6.5% overall yield and 15 steps from 1,2-cyclohexanedione and anisidine 13. A distinctive feature of this sequence is the use of palladium-catalyzed reactions to form all carbon-carbon bonds in the transformation of these simple precursors to (+)-minfiensine.
Subject(s)
Alkaloids/chemical synthesis , Carbazoles/chemical synthesis , Imines/chemistry , Strychnos/chemistry , Alkaloids/chemistry , Carbazoles/chemistry , Crystallography, X-Ray , Cyclization , Cycloparaffins/chemistry , Epoxy Compounds/chemistry , Ions/chemistry , Models, Molecular , Molecular Structure , StereoisomerismABSTRACT
A catalytic asymmetric method for the chemical synthesis of alkaloids containing the 1,2,3,4-tetrahydro-9a,4a-(iminoethano)-9H-carbazole (1) moiety is reported and verified by the enantioselective total synthesis of (+)-minfiensine (4). The central step in this total synthesis is the sequential catalytic asymmetric Heck-N-acyliminium ion cyclization of dienyl carbamate triflate 10, prepared in six steps from 1,2-cyclohexanedione, to give enantiopure 3,4-dihydro-9a,4a-(iminoethano)-9H-carbazole (12) in 75% yield. Iminoethano-9H-carbazole 12 is transformed in six steps to dienyl iodide 17, which undergoes diastereoselective intramolecular Heck cyclization to form pentacyclic intermediate 18. In eight additional steps, this latter intermediate is transformed to (+)-minfiensine (4).
Subject(s)
Alkaloids/chemical synthesis , Carbazoles/chemical synthesis , Cyclization , Imines/chemistry , Stereoisomerism , Strychnos/chemistryABSTRACT
A practical sequence involving three consecutive palladium(0)-catalyzed reactions has been developed for synthesizing 3-alkyl-3-aryloxindoles in high enantiopurity. The Heck cyclization precursors 10 and 11a-k are generated in one step by chemoselective Stille cross-coupling of 2'-triflato-(Z)-2-stannyl-2-butenanilide 9 with aryl or heteroaryl iodides. The pivotal catalytic asymmetric Heck cyclization step of this sequence takes place in high yield and with high enantioselectivity (71-98% ee) with the Pd-BINAP catalyst derived from Pd(OAc)(2) to construct oxindoles containing a diaryl-substituted all-carbon quaternary carbon center. A wide variety of aryl and heteroaryl substituents, including ones of considerable steric bulk, can be introduced at C3 of oxindoles in this way (Table 4). The only limitations encountered to date are aryl substituents containing ortho nitro or basic amine functionalities and the bulky N-alkyl-7-oxindolyl group. Asymmetric Heck cyclization of butenalide 22 having an o-(N-acetyl-N-benzylamino)phenyl substituent at C2 provided a approximately 1:1 mixture of amide atropisomers 23 and 24 in high yield and high enantioselectivity. These atropisomers are formed directly upon Heck cyclization of 22 at 80 degrees C, as they interconvert thermally to only a small extent at this temperature.