RESUMO
The gene KCNT1 encodes the sodium-activated potassium channel KNa1.1 (Slack, Slo2.2). Variants in the KCNT1 gene induce a gain-of-function (GoF) phenotype in ionic currents and cause a spectrum of intractable neurological disorders in infants and children, including epilepsy of infancy with migrating focal seizures (EIMFS) and autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE). Effective treatment options for KCNT1-related disease are absent, and novel therapies are urgently required. We describe the development of a novel class of oxadiazole KNa1.1 inhibitors, leading to the discovery of compound 31 that reduced seizures and interictal spikes in a mouse model of KCNT1 GoF.
RESUMO
We aimed to develop radioligands for PET imaging of brain phosphodiesterase subtype 4D (PDE4D), a potential target for developing cognition enhancing or antidepressive drugs. Exploration of several chemical series gave four leads with high PDE4D inhibitory potency and selectivity, optimal lipophilicity, and good brain uptake. These leads featured alkoxypyridinyl cores. They were successfully labeled with carbon-11 (t1/2 = 20.4 min) for evaluation with PET in monkey. Whereas two of these radioligands did not provide PDE4D-specific signal in monkey brain, two others, [11C]T1660 and [11C]T1650, provided sizable specific signal, as judged by pharmacological challenge using rolipram or a selective PDE4D inhibitor (BPN14770) and subsequent biomathematical analysis. Specific binding was highest in prefrontal cortex, temporal cortex, and hippocampus, regions that are important for cognitive function. [11C]T1650 was progressed to evaluation in humans with PET, but the output measure of brain enzyme density (VT) increased with scan duration. This instability over time suggests that radiometabolite(s) were accumulating in the brain. BPN14770 blocked PDE4D uptake in human brain after a single dose, but the percentage occupancy was difficult to estimate because of the unreliability of measuring VT. Overall, these results show that imaging of PDE4D in primate brain is feasible but that further radioligand refinement is needed, most likely to avoid problematic radiometabolites.
Assuntos
Encéfalo , Tomografia por Emissão de Pósitrons , Encéfalo/diagnóstico por imagem , Encéfalo/metabolismo , Radioisótopos de Carbono , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/metabolismo , Compostos Radiofarmacêuticos , Rolipram/farmacologiaRESUMO
The discovery and selection of a highly potent and selective NaV1.7 inhibitor PF-06456384, designed specifically for intravenous infusion, is disclosed. Extensive in vitro pharmacology and ADME profiling followed by in vivo preclinical PK and efficacy model data are discussed. A proposed protein-ligand binding mode for this compound is also provided to rationalise the high levels of potency and selectivity over inhibition of related sodium channels. To further support the proposed binding mode, potent conjugates are described which illustrate the potential for development of chemical probes to enable further target evaluation.
Assuntos
Canal de Sódio Disparado por Voltagem NAV1.7/química , Piperidinas/química , Piridinas/química , Sulfonamidas/química , Bloqueadores do Canal de Sódio Disparado por Voltagem/química , Animais , Sítios de Ligação , Cães , Meia-Vida , Hepatócitos/metabolismo , Humanos , Infusões Intravenosas , Concentração Inibidora 50 , Camundongos , Microssomos Hepáticos/metabolismo , Simulação de Acoplamento Molecular , Canal de Sódio Disparado por Voltagem NAV1.7/metabolismo , Dor/tratamento farmacológico , Dor/patologia , Piperidinas/farmacocinética , Piperidinas/uso terapêutico , Ligação Proteica , Estrutura Terciária de Proteína , Piridinas/farmacocinética , Piridinas/uso terapêutico , Ratos , Relação Estrutura-Atividade , Sulfonamidas/farmacocinética , Sulfonamidas/uso terapêutico , Tiadiazóis , Bloqueadores do Canal de Sódio Disparado por Voltagem/farmacocinética , Bloqueadores do Canal de Sódio Disparado por Voltagem/uso terapêuticoRESUMO
A series of acidic diaryl ether heterocyclic sulfonamides that are potent and subtype selective NaV1.7 inhibitors is described. Optimization of early lead matter focused on removal of structural alerts, improving metabolic stability and reducing cytochrome P450 inhibition driven drug-drug interaction concerns to deliver the desired balance of preclinical in vitro properties. Concerns over nonmetabolic routes of clearance, variable clearance in preclinical species, and subsequent low confidence human pharmacokinetic predictions led to the decision to conduct a human microdose study to determine clinical pharmacokinetics. The design strategies and results from preclinical PK and clinical human microdose PK data are described leading to the discovery of the first subtype selective NaV1.7 inhibitor clinical candidate PF-05089771 (34) which binds to a site in the voltage sensing domain.
Assuntos
Canal de Sódio Disparado por Voltagem NAV1.7/metabolismo , Éteres Fenílicos/farmacologia , Sulfonamidas/farmacologia , Bloqueadores do Canal de Sódio Disparado por Voltagem/farmacologia , Linhagem Celular , Citocromo P-450 CYP2C9/metabolismo , Inibidores do Citocromo P-450 CYP2C9/síntese química , Inibidores do Citocromo P-450 CYP2C9/química , Inibidores do Citocromo P-450 CYP2C9/farmacocinética , Inibidores do Citocromo P-450 CYP2C9/farmacologia , Citocromo P-450 CYP3A/metabolismo , Inibidores do Citocromo P-450 CYP3A/síntese química , Inibidores do Citocromo P-450 CYP3A/química , Inibidores do Citocromo P-450 CYP3A/farmacocinética , Inibidores do Citocromo P-450 CYP3A/farmacologia , Desenho de Fármacos , Humanos , Microssomos Hepáticos/metabolismo , Canal de Sódio Disparado por Voltagem NAV1.7/química , Éteres Fenílicos/síntese química , Éteres Fenílicos/química , Éteres Fenílicos/farmacocinética , Relação Estrutura-Atividade , Sulfonamidas/síntese química , Sulfonamidas/química , Sulfonamidas/farmacocinética , Bloqueadores do Canal de Sódio Disparado por Voltagem/síntese química , Bloqueadores do Canal de Sódio Disparado por Voltagem/química , Bloqueadores do Canal de Sódio Disparado por Voltagem/farmacocinéticaRESUMO
A series of TRPA1 antagonists is described which has as its core structure an indazole moiety. The physical properties and in vitro DMPK profiles are discussed. Good in vivo exposure was obtained with several analogs, allowing efficacy to be assessed in rodent models of inflammatory pain. Two compounds showed significant activity in these models when administered either systemically or topically. Protein chimeras were constructed to indicate compounds from the series bound in the S5 region of the channel, and a computational docking model was used to propose a binding mode for example compounds.
RESUMO
Human genetic studies show that the voltage gated sodium channel 1.7 (Nav1.7) is a key molecular determinant of pain sensation. However, defining the Nav1.7 contribution to nociceptive signalling has been hampered by a lack of selective inhibitors. Here we report two potent and selective arylsulfonamide Nav1.7 inhibitors; PF-05198007 and PF-05089771, which we have used to directly interrogate Nav1.7's role in nociceptor physiology. We report that Nav1.7 is the predominant functional TTX-sensitive Nav in mouse and human nociceptors and contributes to the initiation and the upstroke phase of the nociceptor action potential. Moreover, we confirm a role for Nav1.7 in influencing synaptic transmission in the dorsal horn of the spinal cord as well as peripheral neuropeptide release in the skin. These findings demonstrate multiple contributions of Nav1.7 to nociceptor signalling and shed new light on the relative functional contribution of this channel to peripheral and central noxious signal transmission.
Assuntos
Axônios/fisiologia , Canal de Sódio Disparado por Voltagem NAV1.7/efeitos dos fármacos , Terminações Pré-Sinápticas/fisiologia , Potenciais de Ação , Animais , Gânglios Espinais/efeitos dos fármacos , Gânglios Espinais/fisiologia , Células HEK293 , Humanos , Masculino , Camundongos , Canal de Sódio Disparado por Voltagem NAV1.7/fisiologia , Técnicas de Patch-Clamp , Éteres Fenílicos/farmacologia , Sulfonamidas/farmacologiaRESUMO
Voltage-gated sodium (NaV) channels are a family of transmembrane ion channel proteins. They function by forming a gated, water-filled pore to help establish and control cell membrane potential via control of the flow of ions between the intracellular and the extracellular environments. Blockade of NaVs has been successfully accomplished in the clinic to enable control of pathological firing patterns that occur in a diverse range of conditions such as chronic pain, epilepsy, and cardiac arrhythmias. First generation sodium channel modulator drugs, despite low inherent subtype selectivity, preferentially act on over-excited cells which reduces undesirable side effects in the clinic. However, the limited therapeutic indices observed with the first generation demanded a new generation of sodium channel inhibitors. The structure, function and the state of the art in sodium channel modulator drug discovery are discussed in this chapter.
Assuntos
Analgésicos/farmacologia , Antiarrítmicos/farmacologia , Anticonvulsivantes/farmacologia , Descoberta de Drogas/métodos , Bloqueadores dos Canais de Sódio/farmacologia , Canais de Sódio Disparados por Voltagem/metabolismo , Sequência de Aminoácidos , Analgésicos/química , Animais , Antiarrítmicos/química , Anticonvulsivantes/química , Humanos , Dados de Sequência Molecular , Bloqueadores dos Canais de Sódio/químicaRESUMO
Voltage-gated sodium channels (Navs) are an important family of transmembrane ion channel proteins and Nav drug discovery is an exciting field. Pharmaceutical investment in Navs for pain therapeutics has expanded exponentially due to genetic data such as SCN10A mutations and an improved ability to establish an effective screen sequence for example IonWorks Barracuda®, Synchropatch® and Qube®. Moreover, emerging clinical data (AZD-3161, XEN402, CNV1014802, PF-05089771, PF-04531083) combined with recent breakthroughs in Nav structural biology pave the way for a future of fruitful prospective Nav drug discovery.
Assuntos
Dor/tratamento farmacológico , Bloqueadores dos Canais de Sódio/farmacologia , Bloqueadores dos Canais de Sódio/uso terapêutico , Canais de Sódio Disparados por Voltagem/metabolismo , Animais , Descoberta de Drogas , Humanos , Bloqueadores dos Canais de Sódio/química , Canais de Sódio Disparados por Voltagem/químicaRESUMO
Voltage-gated sodium (Nav) channels play a fundamental role in the generation and propagation of electrical impulses in excitable cells. Here we describe two unique structurally related nanomolar potent small molecule Nav channel inhibitors that exhibit up to 1,000-fold selectivity for human Nav1.3/Nav1.1 (ICA-121431, IC50, 19 nM) or Nav1.7 (PF-04856264, IC50, 28 nM) vs. other TTX-sensitive or resistant (i.e., Nav1.5) sodium channels. Using both chimeras and single point mutations, we demonstrate that this unique class of sodium channel inhibitor interacts with the S1-S4 voltage sensor segment of homologous Domain 4. Amino acid residues in the "extracellular" facing regions of the S2 and S3 transmembrane segments of Nav1.3 and Nav1.7 seem to be major determinants of Nav subtype selectivity and to confer differences in species sensitivity to these inhibitors. The unique interaction region on the Domain 4 voltage sensor segment is distinct from the structural domains forming the channel pore, as well as previously characterized interaction sites for other small molecule inhibitors, including local anesthetics and TTX. However, this interaction region does include at least one amino acid residue [E1559 (Nav1.3)/D1586 (Nav1.7)] that is important for Site 3 α-scorpion and anemone polypeptide toxin modulators of Nav channel inactivation. The present study provides a potential framework for identifying subtype selective small molecule sodium channel inhibitors targeting interaction sites away from the pore region.
Assuntos
Acetamidas/farmacologia , Fenômenos Eletrofisiológicos/fisiologia , Canal de Sódio Disparado por Voltagem NAV1.3/metabolismo , Tiazóis/farmacologia , Bloqueadores do Canal de Sódio Disparado por Voltagem/farmacologia , Motivos de Aminoácidos/genética , Sítios de Ligação/genética , Células HEK293 , Humanos , Concentração Inibidora 50 , Dados de Sequência Molecular , Canal de Sódio Disparado por Voltagem NAV1.3/genética , Técnicas de Patch-Clamp , Alinhamento de SequênciaRESUMO
Na(v)1.8 (also known as PN3) is a tetrodotoxin-resistant (TTx-r) voltage-gated sodium channel (VGSC) that is highly expressed on small diameter sensory neurons. It has been implicated in the pathophysiology of inflammatory and neuropathic pain, and we envisioned that selective blockade of Na(v)1.8 would be analgesic, while reducing adverse events typically associated with non-selective VGSC blocking therapeutic agents. Herein, we describe the preparation and characterization of a series of 6-aryl-2-pyrazinecarboxamides, which are potent blockers of the human Na(v)1.8 channel and also block TTx-r sodium currents in rat dorsal root ganglia (DRG) neurons. Selected derivatives display selectivity versus human Na(v)1.2. We further demonstrate that an example from this series is orally bioavailable and produces antinociceptive activity in vivo in a rodent model of neuropathic pain following oral administration.
Assuntos
Neuralgia/tratamento farmacológico , Pirazinas/química , Bloqueadores dos Canais de Sódio/química , Canais de Sódio/química , Administração Oral , Animais , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Gânglios Espinais/citologia , Humanos , Microssomos/metabolismo , Canal de Sódio Disparado por Voltagem NAV1.8 , Neurônios/metabolismo , Pirazinas/farmacocinética , Pirazinas/uso terapêutico , Ratos , Bloqueadores dos Canais de Sódio/farmacocinética , Bloqueadores dos Canais de Sódio/uso terapêutico , Canais de Sódio/metabolismo , Relação Estrutura-AtividadeRESUMO
A series of aryl-substituted nicotinamide derivatives with selective inhibitory activity against the Na(v)1.8 sodium channel is reported. Replacement of the furan nucleus and homologation of the anilide linker in subtype-selective blocker A-803467 (1) provided potent, selective derivatives with improved aqueous solubility and oral bioavailability. Representative compounds from this series displayed efficacy in rat models of inflammatory and neuropathic pain.
Assuntos
Niacinamida/farmacologia , Bloqueadores dos Canais de Sódio/farmacologia , Administração Oral , Animais , Disponibilidade Biológica , Niacinamida/química , Niacinamida/farmacocinética , Ratos , Bloqueadores dos Canais de Sódio/administração & dosagem , Bloqueadores dos Canais de Sódio/química , Bloqueadores dos Canais de Sódio/farmacocinética , Relação Estrutura-AtividadeRESUMO
Activation of sodium channels is essential to action potential generation and propagation. Recent genetic and pharmacological evidence indicates that activation of Na(v)1.8 channels contributes to chronic pain. Herein, we describe the identification of a novel series of structurally related pyridine derivatives as potent Na(v)1.8 channel blockers. A-887826 exemplifies this series and potently (IC(50)=11nM) blocked recombinant human Na(v)1.8 channels. A-887826 was approximately 3 fold less potent to block Na(v)1.2, approximately 10 fold less potent to block tetrodotoxin-sensitive sodium (TTX-S Na(+)) currents and was >30 fold less potent to block Na(V)1.5 channels. A-887826 potently blocked tetrodotoxin-resistant sodium (TTX-R Na(+)) currents (IC(50)=8nM) from small diameter rat dorsal root ganglion (DRG) neurons in a voltage-dependent fashion. A-887826 effectively suppressed evoked action potential firing when DRG neurons were held at depolarized potentials and reversibly suppressed spontaneous firing in small diameter DRG neurons from complete Freund's adjuvant inflamed rats. Following oral administration, A-887826 significantly attenuated tactile allodynia in a rat neuropathic pain model. Further characterization of TTX-R current block in rat DRG neurons demonstrated that A-887826 (100nM) shifted the mid-point of voltage-dependent inactivation of TTX-R currents by approximately 4mV without affecting voltage-dependent activation and did not exhibit frequency-dependent inhibition. The present data demonstrate that A-887826 is a structurally novel and potent Na(v)1.8 blocker that inhibits rat DRG TTX-R currents in a voltage-, but not frequency-dependent fashion. The ability of this structurally novel Na(v)1.8 blocker to effectively reduce tactile allodynia in neuropathic rats further supports the role of Na(v)1.8 sodium channels in pathological pain states.
Assuntos
Hiperalgesia/tratamento farmacológico , Limiar da Dor/efeitos dos fármacos , Bloqueadores dos Canais de Sódio/farmacologia , Bloqueadores dos Canais de Sódio/uso terapêutico , Canais de Sódio/metabolismo , Animais , Biofísica , Células Cultivadas , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Estimulação Elétrica/métodos , Gânglios Espinais/citologia , Humanos , Hiperalgesia/etiologia , Masculino , Potenciais da Membrana/efeitos dos fármacos , Morfolinas/química , Morfolinas/farmacologia , Morfolinas/uso terapêutico , Canal de Sódio Disparado por Voltagem NAV1.8 , Neuralgia/complicações , Neuralgia/etiologia , Niacinamida/análogos & derivados , Niacinamida/química , Niacinamida/farmacologia , Niacinamida/uso terapêutico , Técnicas de Patch-Clamp/métodos , Ratos , Ratos Sprague-Dawley , Células Receptoras Sensoriais/efeitos dos fármacos , Bloqueadores dos Canais de Sódio/química , Canais de Sódio/efeitos dos fármacos , Traumatismos da Medula Espinal/complicações , Tetrodotoxina/farmacologia , Transfecção/métodosRESUMO
The synthesis and pharmacological characterization of a novel furan-based class of voltage-gated sodium channel blockers is reported. Compounds were evaluated for their ability to block the tetrodotoxin-resistant sodium channel Na(v)1.8 (PN3) as well as the Na(v)1.2 and Na(v)1.5 subtypes. Benchmark compounds from this series possessed enhanced potency, oral bioavailability, and robust efficacy in a rodent model of neuropathic pain, together with improved CNS and cardiovascular safety profiles compared to the clinically used sodium channel blockers mexiletine and lamotrigine.
Assuntos
Analgésicos não Narcóticos/química , Analgésicos não Narcóticos/farmacologia , Furanos/química , Furanos/farmacologia , Neuralgia/tratamento farmacológico , Piperazinas/química , Piperazinas/farmacologia , Bloqueadores dos Canais de Sódio/química , Bloqueadores dos Canais de Sódio/farmacologia , Canais de Sódio/efeitos dos fármacos , Analgésicos não Narcóticos/síntese química , Animais , Canais de Potássio Éter-A-Go-Go/antagonistas & inibidores , Furanos/síntese química , Humanos , Masculino , Camundongos , Piperazinas/síntese química , Ratos , Ratos Sprague-Dawley , Bloqueadores dos Canais de Sódio/síntese química , Relação Estrutura-AtividadeRESUMO
Nav1.8 (also known as PN3) is a tetrodotoxin-resistant (TTx-r) voltage-gated sodium channel (VGSC) that is highly expressed on small diameter sensory neurons and has been implicated in the pathophysiology of inflammatory and neuropathic pain. Recent studies using an Nav1.8 antisense oligonucleotide in an animal model of chronic pain indicated that selective blockade of Nav1.8 was analgesic and could provide effective analgesia with a reduction in the adverse events associated with nonselective VGSC blocking therapeutic agents. Herein, we describe the preparation and characterization of a series of 5-substituted 2-furfuramides, which are potent, voltage-dependent blockers (IC50 < 10 nM) of the human Nav1.8 channel. Selected derivatives, such as 7 and 27, also blocked TTx-r sodium currents in rat dorsal root ganglia (DRG) neurons with comparable potency and displayed >100-fold selectivity versus human sodium (Nav1.2, Nav1.5, Nav1.7) and human ether-a-go-go (hERG) channels. Following systemic administration, compounds 7 and 27 dose-dependently reduced neuropathic and inflammatory pain in experimental rodent models.
Assuntos
Amidas/síntese química , Analgésicos/síntese química , Anti-Inflamatórios não Esteroides/síntese química , Furanos/síntese química , Bloqueadores dos Canais de Sódio/síntese química , Canais de Sódio/fisiologia , Amidas/química , Amidas/farmacologia , Analgésicos/farmacocinética , Analgésicos/farmacologia , Animais , Anti-Inflamatórios não Esteroides/farmacocinética , Anti-Inflamatórios não Esteroides/farmacologia , Linhagem Celular , Cricetinae , Cricetulus , Furanos/química , Furanos/farmacocinética , Furanos/farmacologia , Gânglios Espinais/citologia , Humanos , Técnicas In Vitro , Masculino , Camundongos , Canal de Sódio Disparado por Voltagem NAV1.8 , Proteínas do Tecido Nervoso/fisiologia , Neurônios/efeitos dos fármacos , Neurônios/fisiologia , Dor/tratamento farmacológico , Dor/etiologia , Técnicas de Patch-Clamp , Doenças do Sistema Nervoso Periférico/tratamento farmacológico , Ratos , Ratos Sprague-Dawley , Proteínas Recombinantes/antagonistas & inibidores , Bloqueadores dos Canais de Sódio/farmacocinética , Bloqueadores dos Canais de Sódio/farmacologia , Relação Estrutura-AtividadeRESUMO
Screening in a 'well-less' or lawn format provides a means to screen large compound collections against many targets in a fast, versatile and cost effective manner. The development of generic lawn format assays to screen various gene families against large compound collections should facilitate the identification of hits and tools to use in drug discovery and chemogenomic endeavours. Lawn format holds particular promise for screening GPCRs and selected enzyme families with potential use in other gene families.
Assuntos
Técnicas de Química Combinatória/métodos , Avaliação Pré-Clínica de Medicamentos/métodos , Animais , Anti-Infecciosos/síntese química , Anti-Infecciosos/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Células Cultivadas , Desenho de Fármacos , Enzimas/química , Humanos , Ligação ProteicaRESUMO
The high-throughput manual solid-phase parallel synthesis of libraries comprising thousands of discrete samples using pellicular supports (i.e. SynPhase crowns and lanterns) and a suite of novel tools and techniques is described. Key aspects of this approach include the combination of a split-split-split synthesis strategy with spatial encoding to differentiate thousands of crowns, the rapid washing and filtration of up to 48 reaction vessels in parallel, the application of an inexpensive and environmentally friendly technique to remove trifluoroacetic acid from sixteen 96-well plates in parallel, and a high-throughput method for removing cleaved crowns from reusable pin racks. Tens of thousands of discrete samples have been produced in-house using this conceptually and operationally straightforward strategy.