ABSTRACT
Voltage-gated sodium channels are known to be expressed in neurons and other excitable cells. Recently, voltage-gated sodium channels have been found to be expressed in human prostate cancer cells. α-Hydroxy-α-phenylamides are a new class of small molecules that have demonstrated potent inhibition of voltage-gated sodium channels. The hydroxyamide motif, an isostere of a hydantoin ring, provides an active scaffold from which several potent racemic sodium channel blockers have been derived. With little known about chiral preferences, the development of chiral syntheses to obtain each pure enantiomer for evaluation as sodium channel blockers is important. Using Seebach and Frater's chiral template, cyclocondensation of (R)-3-chloromandelic acid with pivaldehyde furnished both the cis- and trans-2,5-disubsituted dioxolanones. Using this chiral template, we synthesized both enantiomers of 2-(3-chlorophenyl)-2-hydroxynonanamide, and evaluated their ability to functionally inhibit hNa(v) isoforms, human prostate cancer cells and xenograft. Enantiomers of lead demonstrated significant ability to reduce prostate cancer in vivo.
Subject(s)
Amides/chemistry , Amides/therapeutic use , Antineoplastic Agents/chemistry , Antineoplastic Agents/therapeutic use , Prostatic Neoplasms/drug therapy , Sodium Channel Blockers/chemistry , Sodium Channel Blockers/therapeutic use , Amides/chemical synthesis , Animals , Antineoplastic Agents/chemical synthesis , Cell Line , Cell Line, Tumor , Chemistry Techniques, Synthetic/methods , Humans , Ion Channel Gating/drug effects , Isomerism , Male , Mice , Mice, Inbred BALB C , Mice, Nude , Models, Molecular , Prostate/drug effects , Prostate/metabolism , Prostatic Neoplasms/metabolism , Sodium Channel Blockers/chemical synthesis , Sodium Channels/chemistry , Sodium Channels/metabolismABSTRACT
A series of novel isoxazole voltage gated sodium channel blockers have been synthesized and evaluated. Substitutions on the benzylic position of benzamide were investigated to determine their effect on Na(v)1.7 inhibitory potency. The spirocyclobutyl substitution had the most significant enhancement on Na(v)1.7 inhibitory activity.
Subject(s)
Isoxazoles/therapeutic use , Pain/drug therapy , Sodium Channel Blockers/therapeutic use , Sodium Channels/metabolism , Animals , Cell Line , Chronic Disease , Humans , Isoxazoles/chemistry , Isoxazoles/pharmacology , Pain/immunology , Rats , Sodium Channel Blockers/chemistry , Sodium Channel Blockers/pharmacology , Spinal Nerves/drug effects , Structure-Activity RelationshipABSTRACT
Analogs of the previously reported voltage gated sodium channel blocker CDA54 were prepared in which one of the amide functions was replaced with aromatic and non-aromatic heterocycles. Replacement of the amide with an aromatic heterocycle resulted in significant loss of sodium channel blocking activity, while non-aromatic heterocycle replacements were well tolerated.
Subject(s)
Isoxazoles/chemistry , Isoxazoles/pharmacology , Sodium Channel Blockers/chemistry , Sodium Channel Blockers/pharmacology , Animals , Isoxazoles/therapeutic use , Models, Molecular , Molecular Structure , Pain/drug therapy , Rats , Rats, Sprague-Dawley , Sodium Channel Blockers/therapeutic use , Spinal Nerves/drug effects , Structure-Activity RelationshipABSTRACT
A series of imidazopyridines were evaluated as potential sodium channel blockers for the treatment of neuropathic pain. Several members were identified with good hNa(v)1.7 potency and excellent rat pharmacokinetic profiles. Compound 4 had good efficacy (52% and 41% reversal of allodynia at 2 and 4h post-dose, respectively) in the Chung rat spinal nerve ligation (SNL) model of neuropathic pain when dosed orally at 10mg/kg.
Subject(s)
Pyridines/chemistry , Pyridines/pharmacology , Sodium Channel Blockers/chemistry , Sodium Channel Blockers/pharmacology , Sodium Channels/metabolism , Analgesics/chemistry , Analgesics/pharmacology , Animals , Inflammation/drug therapy , Molecular Structure , NAV1.7 Voltage-Gated Sodium Channel , Pain/drug therapy , Rats , Sodium Channel Blockers/pharmacokinetics , Structure-Activity RelationshipABSTRACT
A series of 3-amino-1,5-benzodiazepinones were synthesized and evaluated as potential sodium channel blockers in a functional, membrane potential-based assay. One member of this series displayed subnanomolar, state-dependent sodium channel block, and was orally efficacious in a mouse model of epilepsy.
Subject(s)
Anticonvulsants/pharmacology , Benzodiazepinones/pharmacology , Epilepsy/drug therapy , Ether-A-Go-Go Potassium Channels/antagonists & inhibitors , Sodium Channel Blockers/pharmacology , Animals , Anticonvulsants/chemical synthesis , Anticonvulsants/pharmacokinetics , Benzodiazepinones/chemical synthesis , Benzodiazepinones/pharmacokinetics , Electrophysiology , Electroshock , Epilepsy/metabolism , Ether-A-Go-Go Potassium Channels/metabolism , Fluorescence Resonance Energy Transfer , Humans , Mice , Molecular Structure , Rats , Sodium Channel Blockers/chemical synthesis , Sodium Channel Blockers/pharmacokineticsABSTRACT
Voltage-gated sodium channels (Nav1) transmit pain signals from peripheral nociceptive neurons, and blockers of these channels have been shown to ameliorate a number of pain conditions. Because these drugs can have adverse effects that limit their efficacy, more potent and selective Nav1 inhibitors are being pursued. Recent human genetic data have provided strong evidence for the involvement of the peripheral nerve sodium channel subtype, Nav1.7, in the signaling of nociceptive information, highlighting the importance of identifying selective Nav1.7 blockers for the treatment of chronic pain. Using a high-throughput functional assay, novel Nav1.7 blockers, namely, the 1-benzazepin-2-one series, have recently been identified. Further characterization of these agents indicates that, in addition to high-affinity inhibition of Nav1.7 channels, selectivity against the Nav1.5 and Nav1.8 subtypes can also be achieved within this structural class. The most potent, nonselective member of this class of Nav1.7 blockers has been radiolabeled with tritium. [3H]BNZA binds with high affinity to rat brain synaptosomal membranes (Kd = 1.5 nM) and to membranes prepared from HEK293 cells stably transfected with hNav1.5 (Kd = 0.97 nM). In addition, and for the first time, high-affinity binding of a radioligand to hNav1.7 channels (Kd = 1.6 nM) was achieved with [3H]BNZA, providing an additional means for identifying selective Nav1.7 channel inhibitors. Taken together, these data suggest that members of the novel 1-benzazepin-2-one structural class of Nav1 blockers can display selectivity toward the peripheral nerve Nav1.7 channel subtype, and with appropriate pharmacokinetic and drug metabolism properties, these compounds could be developed as analgesic agents.
Subject(s)
Benzazepines/chemistry , Sodium Channel Blockers/chemistry , Sodium Channels/physiology , Animals , Benzazepines/metabolism , Benzazepines/pharmacology , Binding, Competitive , Cell Line , Cell Membrane/metabolism , Electrophysiology , Humans , Membrane Potentials/drug effects , Molecular Structure , NAV1.7 Voltage-Gated Sodium Channel , NAV1.8 Voltage-Gated Sodium Channel , Radioligand Assay , Rats , Sodium Channel Blockers/metabolism , Sodium Channel Blockers/pharmacology , Sodium Channels/drug effects , Sodium Channels/metabolism , Synaptosomes/metabolismABSTRACT
A series of benzazepinones were synthesized and evaluated as hNa(v)1.7 sodium channel blockers. Several compounds from this series displayed good oral bioavailability and exposure and were efficacious in a rat model of neuropathic pain.
Subject(s)
Benzodiazepinones/chemical synthesis , Benzodiazepinones/therapeutic use , Neuralgia/drug therapy , Sodium Channel Blockers/chemical synthesis , Sodium Channel Blockers/therapeutic use , Sodium Channels/drug effects , Animals , Benzodiazepinones/pharmacokinetics , Biological Availability , Disease Models, Animal , Dogs , Drug Evaluation, Preclinical , Molecular Structure , NAV1.7 Voltage-Gated Sodium Channel , Rats , Sodium Channel Blockers/pharmacokinetics , Sodium Channels/chemistryABSTRACT
A series of benzodiazepines and benzazepinones were synthesized and evaluated as potential sodium channel blockers in a functional, membrane potential-based assay. One member of the benzazepinone series, compound 47, displayed potent, state-dependent block of hNa(v)1.7, and was orally efficacious in a rat model of neuropathic pain.
Subject(s)
Heterocyclic Compounds, 3-Ring/chemistry , Heterocyclic Compounds, 3-Ring/therapeutic use , Pain/drug therapy , Sodium Channel Blockers/classification , Sodium Channel Blockers/pharmacology , Sodium Channels/metabolism , Animals , Heterocyclic Compounds, 3-Ring/administration & dosage , Heterocyclic Compounds, 3-Ring/pharmacology , Molecular Structure , NAV1.7 Voltage-Gated Sodium Channel , Rats , Sodium Channel Blockers/chemistry , Sodium Channel Blockers/therapeutic use , Structure-Activity RelationshipABSTRACT
Novel cyclopentane-based 3-phenyl-1-hydroxypropyl compounds were evaluated for inhibitory activity against the peripheral nerve sodium channel Na(V)1.7 and off-target activity against the cardiac potassium channel hERG. The stereochemistry of the hydroxyl group and substitution on the phenyl rings with either fluorinated O-alkyl or alkyl groups were found to be critical for conferring potency against Na(V)1.7. A benchmark compound from this series displayed efficacy in rat models of inflammatory and neuropathic pain.
Subject(s)
Cyclopentanes/chemistry , Cyclopentanes/pharmacology , Sodium Channel Blockers/chemical synthesis , Sodium Channel Blockers/pharmacology , Sodium Channels/metabolism , Animals , Cyclopentanes/chemical synthesis , Cyclopentanes/pharmacokinetics , Ether-A-Go-Go Potassium Channels/metabolism , Humans , Inhibitory Concentration 50 , Molecular Structure , Rats , Sodium Channel Blockers/chemistry , Sodium Channel Blockers/pharmacokinetics , Structure-Activity RelationshipABSTRACT
A new series of voltage-gated sodium channel blockers with potential for treatment of chronic pain is reported. Systematic structure-activity relationship studies, starting with compound 1, led to identification of potent analogs that displayed use-dependent block of sodium channels, were efficacious in pain models in vivo, and most importantly, were devoid of activity against the cardiac potassium channel hERG.
Subject(s)
Pain/drug therapy , Sodium Channel Blockers/therapeutic use , Chronic Disease , Humans , Molecular Conformation , Molecular Probes , Sodium Channel Blockers/chemistryABSTRACT
The discovery of novel therapeutic agents that act on voltage-gated sodium channels requires the establishment of high-capacity screening assays that can reliably measure the activity of these proteins. Fluorescence resonance energy transfer (FRET) technology using membrane potential-sensitive dyes has been shown to provide a readout of voltage-gated sodium channel activity in stably transfected cell lines. Due to the inherent rapid inactivation of sodium channels, these assays require the presence of a channel activator to prolong channel opening. Because sodium channel activators and test compounds may share related binding sites on the protein, the assay protocol is critical for the proper identification of channel inhibitors. In this study, high throughput, functional assays for the voltage-gated sodium channels, hNa(V)1.5 and hNa(V)1.7, are described. In these assays, channels stably expressed in HEK cells are preincubated with test compound in physiological medium and then exposed to a sodium channel activator that slows channel inactivation. Sodium ion movement through open channels causes membrane depolarization that can be measured with a FRET dye membrane potential-sensing system, providing a large and reproducible signal. Unlike previous assays, the signal obtained in the agonist initiation assay is sensitive to all sodium channel modulators that were tested and can be used in high throughput mode, as well as in support of Medicinal Chemistry efforts for lead optimization.
Subject(s)
Coloring Agents/analysis , Fluorescence Resonance Energy Transfer/methods , Sodium Channels/analysis , Sodium Channels/physiology , Cell Line , Coloring Agents/pharmacology , Dose-Response Relationship, Drug , Humans , Membrane Potentials/drug effects , Membrane Potentials/physiology , Muscle Proteins/analysis , Muscle Proteins/physiology , NAV1.5 Voltage-Gated Sodium Channel , NAV1.7 Voltage-Gated Sodium Channel , Sodium Channel Blockers/pharmacology , Veratridine/pharmacologyABSTRACT
Sodium channel blockers are used clinically to treat a number of neuropathic pain conditions, but more potent and selective agents should improve on the therapeutic index of currently used drugs. In a high-throughput functional assay, a novel sodium channel (Na(V)) blocker, N-[[2'-(aminosulfonyl)biphenyl-4-yl]methyl]-N'-(2,2'-bithien-5-ylmethyl)succinamide (BPBTS), was discovered. BPBTS is 2 orders of magnitude more potent than anticonvulsant and antiarrhythmic sodium channel blockers currently used to treat neuropathic pain. Resembling block by these agents, block of Na(V)1.2, Na(V)1.5, and Na(V)1.7 by BPBTS was found to be voltage- and use-dependent. BPBTS appeared to bind preferentially to open and inactivated states and caused a dose-dependent hyperpolarizing shift in the steady-state availability curves for all sodium channel subtypes tested. The affinity of BPBTS for the resting and inactivated states of Na(V)1.2 was 1.2 and 0.14 microM, respectively. BPBTS blocked Na(V)1.7 and Na(V)1.2 with similar potency, whereas block of Na(V)1.5 was slightly more potent. The slow tetrodotoxin-resistant Na(+) current in small-diameter DRG neurons was also potently blocked by BPBTS. [(3)H]BPBTS bound with high affinity to a single class of sites present in rat brain synaptosomal membranes (K(d) = 6.1 nM), and in membranes derived from HEK cells stably expressing Na(V)1.5 (K(d) = 0.9 nM). BPBTS dose-dependently attenuated nociceptive behavior in the formalin test, a rat model of tonic pain. On the basis of these findings, BPBTS represents a structurally novel and potent sodium channel blocker that may be used as a template for the development of analgesic agents.
Subject(s)
Amides/therapeutic use , Biphenyl Compounds/therapeutic use , Muscle Proteins/metabolism , Pain Measurement/drug effects , Sodium Channel Blockers/therapeutic use , Sodium Channels/metabolism , Amides/chemical synthesis , Amides/metabolism , Analgesics/chemical synthesis , Analgesics/metabolism , Analgesics/therapeutic use , Animals , Binding Sites , Biphenyl Compounds/chemical synthesis , Biphenyl Compounds/metabolism , Brain/metabolism , Cell Line , Disease Models, Animal , Formaldehyde/administration & dosage , Ganglia, Spinal/drug effects , Ganglia, Spinal/metabolism , Humans , Mice , Muscle Proteins/biosynthesis , Muscle Proteins/genetics , NAV1.2 Voltage-Gated Sodium Channel , NAV1.5 Voltage-Gated Sodium Channel , NAV1.7 Voltage-Gated Sodium Channel , Nerve Tissue Proteins/antagonists & inhibitors , Nerve Tissue Proteins/biosynthesis , Nerve Tissue Proteins/genetics , Patch-Clamp Techniques , Rats , Recombinant Proteins/antagonists & inhibitors , Recombinant Proteins/biosynthesis , Sodium Channel Blockers/chemical synthesis , Sodium Channel Blockers/metabolism , Sodium Channels/biosynthesis , Sodium Channels/genetics , Succinates , Synaptosomes/metabolism , Tetrodotoxin/antagonists & inhibitors , Tetrodotoxin/chemistryABSTRACT
35S-labeled derivatives of the insecticides nodulisporic acid and ivermectin were synthesized and demonstrated to bind with high affinity to a population of receptors in Drosophila head membranes that were previously shown to be associated with a glutamate-gated chloride channel. Nodulisporic acid binding was modeled as binding to a single population of receptors. Ivermectin binding was composed of at least two kinetically distinct receptor populations, only one of which was associated with nodulisporic acid binding. The binding of these two ligands was modulated by glutamate, ivermectin, and antagonists of invertebrate gamma-aminobutyric acid (GABA)ergic receptors. Because solubilized nodulisporic acid and ivermectin receptors comigrated as 230-kDa complexes by gel filtration, antisera specific for both the Drosophila glutamate-gated chloride channel subunit GluCl alpha (DmGluCl alpha) and the GABA-gated chloride channel subunit Rdl (DmRdl) proteins were generated and used to examine the possible coassembly of these two subunits within a single receptor complex. DmGluCl alpha antibodies immunoprecipitated all of the ivermectin and nodulisporic acid receptors solubilized by detergent from Drosophila head membranes. DmRdl antibodies also immunoprecipitated all solubilized nodulisporic receptors, but only approximately 70% of the ivermectin receptors. These data suggest that both DmGluCl alpha and DmRdl are components of nodulisporic acid and ivermectin receptors, and that there also exists a distinct class of ivermectin receptors that contains the DmGluCl alpha subunit but not the DmRdl subunit. This co-association of DmGluCl alpha and DmRdl represents the first biochemical and immunological evidence of coassembly of subunits from two different subclasses of ligand-gated ion channel subunits.