Activation of Neuronal Voltage-Gated Potassium Kv7/KCNQ/M-Current by a Novel Channel Opener SCR2682 for Alleviation of Chronic Pain.

Treatment of chronic pain remains an unmet medical need. The neuronal voltage-gated potassium Kv7/KCNQ/M channel has been implicated as a therapeutic target for chronic pain. However, whether pharmacological activation of the Kv7 channel can alleviate pain remains elusive. In this study, we show that selective activation of native M-currents by a novel channel opener SCR2682 reduces repetitive firings of dorsal root ganglia (DRG) sensory neurons. Intraperitoneal administration of SCR2682 relieves mechanical allodynia and thermal hyperalgesia in rat models of pain induced by complete Freund's adjuvant (CFA) or spared nerve injury (SNI) in a dose-dependent manner without affecting locomotor activity. The antinociceptive efficacy of SCR2682 can be reversed by the channel-specific blocker XE991. Furthermore, SCR2682 increases Kv7.2/KCNQ2 mRNA and protein expression in DRG neurons from rats in the SNI model of neuropathic pain. Taken together, pharmacological activation of neuronal Kv7 channels by opener SCR2682 can alleviate pain in rats, thus possessing therapeutic potential for chronic pain or hyperexcitability-related neurologic disorders. SIGNIFICANCE STATEMENT: A novel voltage-gated potassium Kv7 channel opener SCR2682 inhibits action potential firings in dorsal root ganglia sensory neurons and exhibits efficacy in antinociception, thus possessing a developmental potential for treatment of chronic pain or epilepsy.

Excitatory GABAergic Action and Increased Vasopressin Synthesis in Hypothalamic Magnocellular Neurosecretory Cells Underlie the High Plasma Level of Vasopressin in Diabetic Rats.

Diabetes mellitus (DM) is associated with increased plasma levels of arginine-vasopressin (AVP), which may aggravate hyperglycemia and nephropathy. However, the mechanisms by which DM may cause the increased AVP levels are not known. Electrophysiological recordings in supraoptic nucleus (SON) slices from streptozotocin (STZ)-induced DM rats and vehicle-treated control rats revealed that γ-aminobutyric acid (GABA) functions generally as an excitatory neurotransmitter in the AVP neurons of STZ rats, whereas it usually evokes inhibitory responses in the cells of control animals. Furthermore, Western blotting analyses of Cl- transporters in the SON tissues indicated that Na+-K+-2Cl- cotransporter isotype 1 (a Cl- importer) was upregulated and K+-Cl- cotransporter isotype 2 (KCC2; a Cl- extruder) was downregulated in STZ rats. Treatment with CLP290 (a KCC2 activator) significantly lowered blood AVP and glucose levels in STZ rats. Last, investigation that used rats expressing an AVP-enhanced green fluorescent protein fusion gene showed that AVP synthesis in AVP neurons was much more intense in STZ rats than in control rats. We conclude that altered Cl- homeostasis that makes GABA excitatory and enhanced AVP synthesis are important changes in AVP neurons that would increase AVP secretion in DM. Our data suggest that Cl- transporters in AVP neurons are potential targets of antidiabetes treatments.

Identification of Potent Chloride Intracellular Channel Protein 1 Inhibitors from Traditional Chinese Medicine through Structure-Based Virtual Screening and Molecular Dynamics Analysis.

Chloride intracellular channel 1 (CLIC1) is involved in the development of most aggressive human tumors, including gastric, colon, lung, liver, and glioblastoma cancers. It has become an attractive new therapeutic target for several types of cancer. In this work, we aim to identify natural products as potent CLIC1 inhibitors from Traditional Chinese Medicine (TCM) database using structure-based virtual screening and molecular dynamics (MD) simulation. First, structure-based docking was employed to screen the refined TCM database and the top 500 TCM compounds were obtained and reranked by X-Score. Then, 30 potent hits were achieved from the top 500 TCM compounds using cluster and ligand-protein interaction analysis. Finally, MD simulation was employed to validate the stability of interactions between each hit and CLIC1 protein from docking simulation, and Molecular Mechanics/Generalized Born Surface Area (MM-GBSA) analysis was used to refine the virtual hits. Six TCM compounds with top MM-GBSA scores and ideal-binding models were confirmed as the final hits. Our study provides information about the interaction between TCM compounds and CLIC1 protein, which may be helpful for further experimental investigations. In addition, the top 6 natural products structural scaffolds could serve as building blocks in designing drug-like molecules for CLIC1 inhibition.

Apigenin in combination with Akt inhibition significantly enhances thyrotropin-stimulated radioiodide accumulation in thyroid cells.

BACKGROUND: Selectively increased radioiodine accumulation in thyroid cells by thyrotropin (TSH) allows targeted treatment of thyroid cancer. However, the extent of TSH-stimulated radioiodine accumulation in some thyroid tumors is not sufficient to confer therapeutic efficacy. Hence, it is of clinical importance to identify novel strategies to selectively further enhance TSH-stimulated thyroidal radioiodine accumulation. METHODS: PCCl3 rat thyroid cells, PCCl3 cells overexpressing BRAF(V600E), or primary cultured tumor cells from a thyroid cancer mouse model, under TSH stimulation were treated with various reagents for 24 hours. Cells were then subjected to radioactive iodide uptake, kinetics, efflux assays, and protein extraction followed by Western blotting against selected antibodies. RESULTS: We previously reported that Akt inhibition increased radioiodine accumulation in thyroid cells under chronic TSH stimulation. Here, we identified Apigenin, a plant-derived flavonoid, as a reagent to further enhance the iodide influx rate increased by Akt inhibition in thyroid cells under acute TSH stimulation. Akt inhibition is permissive for Apigenin's action, as Apigenin alone had little effect. This action of Apigenin requires p38 MAPK activity but not PKC-δ. The increase in radioiodide accumulation by Apigenin with Akt inhibition was also observed in thyroid cells expressing BRAF(V600E) and in primary cultured thyroid tumor cells from TRß(PV/PV) mice. CONCLUSION: Taken together, Apigenin may serve as a dietary supplement in combination with Akt inhibitors to enhance therapeutic efficacy of radioiodine for thyroid cancer.

ABC transporters in multidrug resistance and pharmacokinetics, and strategies for drug development.

Multidrug resistance (MDR) is a serious problem that hampers the success of cancer pharmacotherapy. A common mechanism is the overexpression of ATP-binding cassette (ABC) efflux transporters in cancer cells such as P-glycoprotein (P-gp/ABCB1), multidrug resistance-associated protein 1 (MRP1/ABCC1) and breast cancer resistance protein (BCRP/ABCG2) that limit the exposure to anticancer drugs. One way to overcome MDR is to develop ABC efflux transporter inhibitors to sensitize cancer cells to chemotherapeutic drugs. The complete clinical trials thus far have showen that those tested chemosensitizers only add limited or no benefits to cancer patients. Some MDR modulators are merely toxic, and others induce unwanted drug-drug interactions. Actually, many ABC transporters are also expressed abundantly in the gastrointestinal tract, liver, kidney, brain and other normal tissues, and they largely determine drug absorption, distribution and excretion, and affect the overall pharmacokinetic properties of drugs in humans. In addition, ABC transporters such as P-gp, MRP1 and BCRP co-expressed in tumors show a broad and overlapped specificity for substrates and MDR modulators. Thus reliable preclinical assays and models are required for the assessment of transporter-mediated flux and potential effects on pharmacokinetics in drug development. In this review, we provide an overview of the role of ABC efflux transporters in MDR and pharmacokinetics. Preclinical assays for the assessment of drug transport and development of MDR modulators are also discussed.

Phα1ß toxin prevents capsaicin-induced nociceptive behavior and mechanical hypersensitivity without acting on TRPV1 channels.

Phα1ß toxin is a peptide purified from the venom of the armed spider Phoneutria nigriventer, with markedly antinociceptive action in models of acute and persistent pain in rats. Similarly to ziconotide, its analgesic action is related to inhibition of high voltage activated calcium channels with more selectivity for N-type. In this study we evaluated the effect of Phα1ß when injected peripherally or intrathecally in a rat model of spontaneous pain induced by capsaicin. We also investigated the effect of Phα1ß on Ca²âº transients in cultured dorsal root ganglia (DRG) neurons and HEK293 cells expressing the TRPV1 receptor. Intraplantar or intrathecal administered Phα1ß reduced both nocifensive behavior and mechanical hypersensitivity induced by capsaicin similarly to that observed with SB366791, a specific TRPV1 antagonist. Peripheral nifedipine and mibefradil did also decrease nociceptive behavior induced by intraplantar capsaicin. In contrast, ω-conotoxin MVIIA (a selective N-type Ca²âº channel blocker) was effective only when administered intrathecally. Phα1ß, MVIIA and SB366791 inhibited, with similar potency, the capsaicin-induced Ca²âº transients in DRG neurons. The simultaneous administration of Phα1ß and SB366791 inhibited the capsaicin-induced Ca²âº transients that were additive suggesting that they act through different targets. Moreover, Phα1ß did not inhibit capsaicin-activated currents in patch-clamp recordings of HEK293 cells that expressed TRPV1 receptors. Our results show that Phα1ß may be effective as a therapeutic strategy for pain and this effect is not related to the inhibition of TRPV1 receptors.

Ion channel modulators and urinary tract function.

The membrane potential fulfils an important role in initiating smooth muscle contraction, through its depolarization and the subsequent influx of Ca(2+) through voltage-gated Ca(2+) channels. Changes in membrane potential can also coordinate contraction across great distances, utilizing the speed of electrical current flow through gap junctions. Hence, regulating membrane potential can greatly influence smooth muscle function. In this chapter, we will consider the influence of ion channels, as dynamic gatekeepers of membrane permeability, on urogenital function. Through their ability to act as key regulators of both the resting membrane potential and its dynamic changes, they provide important pharmacological targets for influencing urogenital function.Urogenital smooth muscle and urothelia contain a diverse range of molecularly and functionally distinct K(+) channels, which are key to regulating the resting membrane and for re-establishing the normal membrane potential following both active and passive changes. The voltage-gated Ca(2+) channels are key to initiating contraction and causing rapid depolarization, supplemented in some smooth muscles by rapid Na(+) conductances. The Cl(-) channels, often assumed to be passive, can actively change the membrane potential, and hence, cellular function, because Cl(-) is not usually at its equilibrium potential. The useful ways in which these ion channels can be targeted therapeutically in the ureter, bladder and urethra are discussed, focussing particularly on treatments for ureteric obstruction and detrusor overactivity. Current treatments for many urinary tract disorders, particularly the overactive bladder, are complicated by side effects. While ion channels have traditionally been considered as poor therapeutic targets by the pharmaceutical industry, our increasing knowledge of the molecular diversity of K(+) and Cl(-) channels gives new hope for more narrowly focused drug targeting, while the exciting discoveries of active currents in interstitial cells give us a new set of cellular targets for drugs.

[New antiarrhythmic drugs for treatment of atrial fibrillation]. / Uudet rytmihäiriölääkkeet eteisvärinän hoidossa.

Pharmacotherapy of atrial fibrillation (AF) is demanding, because currently available antiarrhythmic drugs have low efficacy and many side effects. In drug development, the focus has been on amiodarone-like multichannel blockers, atrial-specific ion channel blockers, and novel non-channel agents targeting atrial remodelling. Dronedarone, an amiodarone analogue without iodine, was recently approved for treatment of AF. It is less effective than amiodarone, but serious adverse events are rare. Vernakalant, an atrial-selective drug with low proarrhythmic risk, is effective in cardioversion and it may also prevent AF recurrences. So-called upstream therapy with angiotensin converting enzyme and angiotensin receptor inhibitors, statins and omega-3 fatty acids needs further clinical validation.

Inhibition of TRPV1 for the treatment of sensitive skin.

During the past years, the topic sensitive skin became one of the most important fields in dermatology. The tremendous interest is based on several studies showing that about 50% of the population declares to have sensitive skin. The human thermoreceptor hTRPV1 was previously identified to contribute to this skin condition while facilitating neurogenic inflammation leading to hyperalgesia. Furthermore, skin sensitivity towards capsaicin, a natural activator of TRPV1, was shown to correlate with sensitive skin. In a screening campaign based on recombinant HEK293-cells stably transfected with hTRPV1, the selective antagonist trans-4-tert-butylcyclohexanol was identified. This antagonist is able to inhibit capsaicin-induced hTRPV1 activation with an IC(50) value of 34 ± 5 µm tested in HEK293-cells as well as in electrophysiological recordings performed in oocytes expressing hTRPV1. Strikingly, in a clinical study with 30 women using topical treatment with o/w emulsions containing 31.6 ppm capsaicin, we were able to show that 0.4% of this inhibitor significantly reduces capsaicin-induced burning (P < 0.0001) in vivo. Thus trans-4-tert-butylcyclohexanol has the potential as a novel bioactive for the treatment of sensitive skin.

Future treatment strategies for neuropathic pain.

The prevalence of people suffering from chronic pain is extremely high and pain affects millions of people worldwide. As such, persistent pain represents a major health problem and an unmet clinical need. The reason for the high incidence of chronic pain patients is in a large part due to a paucity of effective pain control. An important reason for poor pain control is undoubtedly a deficit in our understanding of the underlying causes of chronic pain and as a consequence our arsenal of analgesic therapies is limited. However, there is considerable hope for the development of new classes of analgesic drugs by targeting novel processes contributing to clinically relevant pain. In this chapter we highlight a number of molecular species which are potential therapeutic targets for future neuropathic pain treatments. In particular, the roles of voltage-gated ion channels, neuroinflammation, protein kinases and neurotrophins are discussed in relation to the generation of neuropathic pain and how by targeting these molecules it may be possible to provide better pain control than is currently available.

Review of the If selective channel inhibitor ivabradine in the treatment of chronic stable angina.

Coronary heart disease is the major cause of morbidity and mortality in industrialized countries, and its prevalence is predicted to grow as the population ages. Current drugs for chronic stable angina (such as beta-blockers, calcium-channel blockers, long- and short-acting nitrates, and potassium-channel activators) are often effective, either as monotherapy or in combination, but side effects and contraindications may limit their use. The "I(f)" (for "funny") channel, discovered in 1979, is expressed mainly in the membrane of pacemaker cells present in the sinus node, the atrioventricular node, the ventricular conduction pathways, and ventricular myocytes. By determining the slope of diastolic depolarization, which in turn controls action potential frequency, it is a key determinant of heart rate and so provides a new therapeutic target for controlling angina symptoms. A new antiangina drug, ivabradine, has been developed and licensed for clinical use. It exclusively reduces the heart rate by selectively blocking the I(f) channel of the sino-atrial node. As clinical trials have shown it to be remarkably well-tolerated, ivabradine offers an alternative for patients who cannot take, or are intolerant of, beta blockade. This review provides an insight into this new agent, its historical background, mechanism of action, and pathophysiologic basis, and provides up-to-date evidence-based information on its optimum use in stable angina.

BK channel modulators: a comprehensive overview.

The large Ca(2+)-activated K(+) channel (BK channel) reflects per excellence the dilemma of the molecular target driven drug discovery process. Significant experimental evidence suggests that the BK channels play a pivotal and specific role in many pathophysiological conditions supporting the notion that the channel represents an innovative and promising drug target. However, after more than ten years of intense research effort both in academia and industry, scientists have yet to witness the approval of a single BK channel modulator for clinical use. On the contrary, three BK openers that were progressed to clinical development have recently been discontinued (NS8, BMS204352 and TA1702) and, at the present time, only one drug candidate targeting BK channels (andolast) remains in the early phases of clinical development. Since biological studies keep strengthening the concept of BK channels as a potentially attractive target, the design and synthesis of potent and selective BK modulators continue based on novel chemical ideas. A comprehensive overview of BK channel modulators is therefore timely and important to the current medicinal chemist for review, summary, and classification of the multitude of chemical entities claimed to be BK-modulating agents. Such chemical entities are, herein, classified by both origin and chemical structure in 1) Endogenous BK channel modulators and structural analogues 2) Naturally-occurring BK channel inhibitors and blockers 3) Synthetic BK channel inhibitors and blockers 4) Marketed and/or investigational drugs with BK-modulating side properties and structural analogues 5) Naturally-occurring BK channel openers and structural analogues 6) Synthetic BK channel openers. This review is intended to provide readers with current opinion on the BK channel as a drug target, the chemical structures of BK channel modulators, the structural and chemical features involved in the BK channel modulating activity and, where and when possible, with highlights of structure-activity relationships.

Molecular pharmacology of the glycine receptor chloride channel.

The glycine receptor (GlyR) Cl(-) channel belongs to the cysteine-loop family of ligand-gated ion channel receptors. It is best known for mediating inhibitory neurotransmission in motor and sensory reflex circuits of the spinal cord, although glycinergic synapses are also present in the brain stem, cerebellum and retina. Extrasynaptic GlyRs are widely distributed throughout the central nervous system and they are also found in sperm and macrophages. A total of 5 GlyR subunits (alpha1-4 and beta) have been identified. Embryonic receptors comprise alpha2 homomers whereas adult receptors comprise predominantly alpha beta heteromers in a 2:3 stoichiometry. Notably, the alpha3 subunit is present in synaptic GlyRs that mediate inhibitory neurotransmission onto spinal nociceptive neurons. These receptors are specifically inhibited by inflammatory mediators, implying a role for alpha3-containing GlyRs in inflammatory pain sensitisation. Because molecules that increase GlyR current may have clinical potential as muscle relaxant and peripheral analgesic drugs, this review focuses on the molecular pharmacology of GlyR potentiating substances. Of all GlyR potentiating substances identified to date, we conclude that 5HT(3)R antagonists such as tropisetron offer the most promise as therapeutic lead compounds. However, one problem is that that virtually all known GlyR potentiating compounds, including tropisetron analogues, lack specificity for the GlyR. Another is that almost nothing is known about the pharmacological properties of alpha3-containing GlyRs, which is the subtype of choice for targeting by novel antinociceptive agents. These issues need to be addressed before GlyR-specific therapeutics can be developed.

Nature's clarion call of antibacterial resistance: are we listening?

Antibiotic resistance is recognized as a major problem worldwide in the management of infectious disease, both in hospital settings and in the community. Therefore, there is an urgent need for new antibiotics, particularly those effective against multidrug-resistant bacteria such as Acinetobacter, Pseudomonas, many Enterobacteriaceae, as well as enterococci and staphylococci. There is also a growing need for new agents with activity against resistant community-acquired pathogens. Major pharmaceutical companies have either abandoned or reduced antibiotic research and development in favor of areas such as chronic illness. In general, the anti-infectives pipelines of pharmaceutical companies are more focused on the antiviral market; however, some large pharmaceutical companies have entered into collaborative discovery projects with smaller companies. Although these smaller companies face many additional challenges, several are still pursuing a wide variety of approaches.

[Clinical trials of new therapeutic pharmacology for sickle cell disease]. / Essais cliniques de nouvelles thérapeutiques pharmacologiques de la drépanocytose.

Following the generation of transgenic mouse models of sickle cell disease, pre-clinical trials have shown the beneficial effects of various potential therapeutic molecules for the acute or chronic manifestations of the disease. Several molecules are upon evaluation in phase I to phase III clinical trials. These therapeutic approaches target: 1) membrane cation transport systems and channels involved in sickle cell dehydration; 2) adherence of erythrocytes to endothelium; 3) activation of circulating and endothelial cells participating in the vasoocclusive events and local ischemia. The Gardos channel (calcium activated potassium channel KCNN4) is inhibited by the clotrimazole metabolite ICA17043, in phase III trial. The K-Cl co-transport (KCC1/3/4) activated by the depletion of erythrocyte magnesium is inhibited by Magnesium pidolate; dipyridamole inhibits ion transports upon deoxygenation. Sulfasalazyne (inhibitor of the NF-jB pathway) inhibits the abnormal activation of endothelial cells. Nitric oxide (NO) is the most potent vasodilator. It prevents the activation of leucocytes, platelets and endothelial cells in patients with sickle cell disease and vascular remodelling. The L-arginine, the NO precursor, provides could be beneficial in sickle cell patients.