Wednesday, October 17

7:30 am Registration Opens

POTASSIUM CHANNELS AND DISEASE

2:10 pm Chairperson’s Remarks 
Jesus “Tito” Gonzalez, Ph.D., Senior Director, Biology, Vertex Pharmaceuticals

2:15 KCa3.1: An Emerging Target for Several Major Diseases of Unmet Medical Needs
Chuan-Chu Chou, Ph.D., Fellow, Inflammation, Schering-Plough Research Institute
KCa3.1 is a calcium-activated intermediate-conductance potassium ion channel. The channel is expressed in several secretory organs but not detected in excitable tissues. While its physiological roles appear to be redundant, the channel was found to play key roles in the development of several diseases including autoimmune disorder, cardiovascular inflammation, and certain types of cancer. Although the mechanistic details of KCa3.1 in the diseases are still under investigation, with the availability of highly selective blockers, the channel has been validated as an attractive target of pharmacological intervention in several classic models of these diseases. 

2:45 Kv1.3 Channels: Therapeutic Target for T-cell-mediated Autoimmune Diseases
Christine Beeton, Ph.D., Physiology and Biophysics, Center for Immunology, University of California, Irvine
T-lymphocyte mediated autoimmune diseases afflict millions of people worldwide. Autoreactive memory T-lymphocytes are implicated in the pathogenesis of these diseases. We show that disease-relevant autoreactive T cells in patients with type-1 diabetes mellitus (T1DM), multiple sclerosis (MS) or rheumatoid arthritis (RA) share a characteristic: they are effector memory T (TEM) cells with elevated levels of the voltage-gated Kv1.3 K+ channel. In contrast, disease-irrelevant autoreactive T cells are Kv1.3low naïve or central memory (TCM) cells. We have generated ShK-186, an analog of a sea anemone peptide that selectively blocks Kv1.3 channels. Kv1.3 blockade suppresses IL2 and IFN production by TEM cells at picomolar concentrations, but is less effective in suppressing TNF production. Naïve/TCM cells become resistant to Kv1.3 blockade during activation by up-regulating the Ca2+-activated KCa3.1 K+ channel. Kv1.3 blockade prevents a TEM-mediated delayed type hypersensitivity reaction in rats and prevents and treats disease in rat models of MS and RA without toxic side-effects. Live imaging of TEM cells at the site of inflammation during a DTH reaction showed that ShK-186 inhibits DTH by blocking the motility of TEM cells. Kv1.3 blockade might have use for the therapy of autoimmune disorders by selectively targeting TEM cells while leaving the bulk of the immune system unimpaired.

3:15 Potassium Channels of the Pancreatic Beta-cell and Their Regulation of Insulin Secretion
James Herrington, Ph.D., Ion Channels, Merck Research Laboratories 
In pancreatic beta-cells, several different potassium channels are thought to be involved in the regulation of insulin secretion. The most extensively studied of these channels is the ATP-sensitive potassium channel, KATP, which is known to control the plasma membrane resting potential. Increased ATP levels in response to glucose results in closure of KATP channels, depolarization of the plasma membrane to produce action potentials, increased cytosolic free calcium, and stimulation of insulin secretion. The action potentials are, in turn, modulated by potassium channels activated by the rise in calcium (KCa) or by the depolarization of the plasma membrane (Kv). Delayed-rectifier potassium currents (IDR) through Kv channels are thought to contribute to action potential repolarization to thereby modulate insulin secretion. The voltage-gated potassium channel, Kv2.1, is expressed in beta-cells, and the biophysical characteristics of heterologously expressed channels are similar to those of IDR in rodent beta-cells. A novel peptidyl inhibitor of Kv2.1/Kv2.2 channels, Guangxitoxin-1 (GxTX-1, IC50 of ~1 nM), has been purified, sequenced and characterized, and a synthetic version has been used to probe the contribution of these channels to beta-cell physiology. In mouse beta-cells, GxTX-1 inhibits 90% of IDR and, as for Kv2.1, shifts the voltage-dependence of channel activation to more depolarized potentials, a characteristic of gating-modifier peptides. GxTX-1 broadens the beta-cell action potential, enhances glucose-stimulated intracellular cal-cium oscillations, and enhances insulin secretion from mouse pancreatic islets and dispersed islet cells in a glucose-dependent manner. This peptide has no effect on the KATP-independent pathway. These data point to a mecha-nism for specific enhancement of glucose-dependent insulin secretion by applying blockers of the beta-cell IDR, which may provide advantages over currently used therapies for the treatment of type 2 diabetes.

TARGETING NEUROPSYCHIATRIC DISORDERS

3:45 Exploring Positive Allosteric Modulators of the Alpha7 Nicotinic Receptor
John Dunlop, Ph.D., Director, Neuropharmacology & Neurophysiology, Discovery Neuroscience, Wyeth Research
Alpha7 nicotinic receptors are highly expressed in the brain, in particular in the hippocampus and cortex, and have been implicated as a therapeutic target in schizophrenia and Alzheimer’s disease. Drug discovery efforts have resulted in the identification of a number of alpha7 nicotinic receptor agonists and partial agonists with demonstrated activity in preclinical models predictive of cognitive enhancing and antipsychotic-like activity. A more recent approach to targeting the alpha7 nicotinic receptor has been with positive allosteric modulators (PAMs). Interestingly, we find that alpha7 nicotinic receptor PAMs are mechanistically distinguishable based on their electrophysiological properties. At the present time the influence of these mechanistic differences on the in vivo properties of these agents is relatively unexplored.

4:15 Networking Refreshment Break in the Exhibit Hall

4:50 Technology Watch
Bringing Potassium Flux Assays to a Higher (Throughput) Level: Fluorescent FluxOR™ Detection Meets BacMam Ion Channel Delivery
Daniel W. Beacham, Ph.D., Scientist, Cell Physiology Group Molecular Probes, Invitrogen Labeling and Detection Technologies, INVITROGEN
Combining the sensitive FluxOR™ assay with BacMam delivery of the hERG potassium channel, researchers can measure potassium flux in a high-throughput, homogenous assay. Data shown will describe the measurement of hERG expressed in U2-OS cells that have been prepared fresh or thawed from frozen stocks for same-day convenience and maximum consistency.

5:05 Technology Watch (Sponsorship Available)

5:20 Search for New Drug Targets for Episodic Neurological Disorders through Mutation Screen of 150 Human Ion Channel Genes
Ronald G. Lafreniere, Ph.D., General Manager & Director of Research, Emerillon Therapeutics Inc.
Mutations in ion channel genes have been found to predispose to many episodic syndromes such as epilepsy and migraine. In an effort to identify additional predisposing genes involved in these and other episodic disor-ders, we have completed a high throughput mutation screen of 150 human brain-expressed ion channel genes in a population of 368 patients with epilepsy, migraine, Tourette syndrome, bipolar disorder or essential tremor. Using a combination of denaturing high performance liquid chromatography (dHPLC) and sequencing technologies, we have identified ~2000 variants in these 150 ion channel genes. Approximately two thirds of these variants are novel, being absent from publicly available SNP databases. We have devised a strategy to prioritize each variant as per potential effect on gene or protein function. High scoring variants are then geno-typed in additional patient and control populations, or tested for co-segregation in affected pedigrees, or for association in large patient and control cohorts. These genetic approaches are combined to identify and validate clinically relevant drug targets for neurological disorders. These validated drug targets are then further validated biochemically to identify appropriate therapeutic strategies. Such novel targets can be used to improve molecular diagnoses and develop novel therapeutics for these disorders.

5:50 Acid-sensing Ion Channels as Novel Therapeutic Targets for Neurological Disorders
Zhigang Xiong, M.D., Ph.D., Director, Neurophysiology, Robert S. Dow Neurobiology Laboratories, Legacy Clinical Research Center 
Ischemic stroke is a leading cause of death and long-term disability in the United States. Unfortunately, there is no effective therapeutic intervention other than the use of thrombolytics, which has a limited therapeutic time window and a potential side effect of intracranial hemorrhage. The absence of neuroprotective therapy is particularly apparent following the failure of multiple clinical trials using the glutamate antagonists as therapeutic agents. Understanding the detailed biochemical changes and the cellular mechanisms involved in brain ischemia and other neurological disorders is critical for establishing new and effective therapeutic strategy. Dramati-cally decreased tissue pH, or acidosis, is a common feature of acute neurological disorders, and has been suggested to play a role in neuronal injury. However, the detailed cellular and molecular mechanisms underlying acid induced neuronal injury remain elusive. The recent finding that acidosis activates a distinct family of cation channels, the acid-sensing ion channels (ASICs), has dramatically changed the view of acid signaling and provided a potential therapeutic strategy for neurological disorders. In CNS neurons, lowering extracellular pH to the level commonly seen in ischemic brain activates ASIC currents, with resultant membrane depolariza-tion and increased [Ca2+]i. Incubation of neurons with acidic solutions reproduces Ca2+-dependent neuronal injury. The acid-induced currents, membrane depolarization, [Ca2+]i increase, and neuronal injury are inhibited by the blockade of ASIC1a, and by ASIC1a gene knockout. Moreover, ASIC1a blockade has up to 5 hours of neuroprotective time window. Thus, Ca2+-permeable ASICs may represent novel therapeutic targets for neuro-logical disorders.

6:20 End of Conference Day

Thursday, October 18

7:30-3:40 pm Registration Open

CRAC – AN EMERGING TARGET

8:35 Chairperson’s Remarks
Martin Gosling, Ph.D., Lab Head, Ion Channel Pharmacology, Novartis Institutes for BioMedical Research

8:40 CRAC Channel Inhibition A Novel Strategy for Immunomodulation
Gonul Velicelebi, Ph.D., Founder, President and CEO, CalciMedica

9:10 Targeting the CRAC Channel for Drug Discovery: Biophysical and Pharmacological Considerations
Murali Prakriya, Ph.D., Assistant Professor, Dept. of Molecular Pharmacology and Biological Chemistry, Northwestern University Feinberg School of Medicine
The activation of Ca²⁺ entry through store-operated CRAC channels mediates several essential functions in the immune system including T-cell activation by antigen and release of inflammatory mediators from mast cells. Recent work using genetic approaches have identified Orai1 and STIM1 as the pore-forming subunit of the CRAC channel and the ER Ca²⁺ sensor, respectively. This talk will summarize the key biophysical and pharmacological properties of CRAC channels and highlight opportunities provided by the discovery of its molecular components for drug discovery.

TECHNOLOGIES FOR ION CHANNEL DRUG DISCOVERY

9:40 Screening Ion Channel Targets with Therapeutic Potential in Respiratory Disease: Application of Automated Electrophysiology 
Martin Gosling, Ph.D.
Enhancing mucociliary clearance offers therapeutic potential to both cystic fibrosis and chronic obstructive pulmonary disease (COPD) patients, whose airways are congested with poorly hydrated viscous mucus. Poten-tiation of the activity of the cystic fibrosis transmembrane regulator (CFTR) or inhibition of the epithelial sodium channel (ENaC) are proposed strategies to accelerate or restore mucociliary clearance. We have developed a series of cell-based high and medium-throughput screening assays using either a fluorescent membrane potential dye or automated planar patch clamp electrophysiology capable of identifying compounds that modulate either CFTR or ENaC - the development and performance of these assays will be described.

10:10 Networking Coffee Break in the Exhibit Hall

10:55 Novel Applications of Planar Array Electrophysiology in Ion Channel Drug Discovery 
Derek J. Trezise, Ph.D., Director, Biological Reagents & Assay Development, GlaxoSmithKline
Planar array patch clamp electrophysiology systems such as IonWorks Quattro and Sophion Q-patch are now embedded assay methodologies in most ion channel drug discovery laboratories. For screening voltage-gated ion channels targets the quantal improvements in speed and capacity afforded by this technology have proved truly enabling. In contrast, ligand-gated ion channels (LGICs) have received relatively little attention. For the highest throughout system, the IonWorks Quattro, the theoretical limitations of the inability to simultaneously add compounds and measure the ionic current have discouraged workers from using this approach. Here, data will be described from experiments using GABAA and hSK/IK channels as test systems to evaluate the true extent of these limitations for screening. With detailed quantitative pharmacological analysis of agonists, an-tagonists, pore blockers and positive and negative modulators we were able to confirm the majority of previously published data from conventional patch clamp experiments. Our findings indicate that with caution planar array electrophysiology is a valuable approach for studying certain LGICs and thus provides another useful tool in the armoury of the drug discovery pharmacologist. 

11:25 Probing Voltage-gated Channels in Neurons
Jesus “Tito” Gonzalez, Ph.D., Senior Director, Biology, Vertex Pharmaceuticals
Voltage-gated ion channels regulate many physiological functions and are targets for numerous drugs. Two important challenges for developing new drug candidates are to understand the biological function of individual channel subtypes and to test candidate compounds in primary cells. Neuronal populations are particularly difficult because of functional and channel heterogeneity; isolation and culturing constraints, and compound assay methods. Here we discuss the use of a subtype-selective peptide toxin and fluorescence imaging methods in peripheral neurons to address these issues.

11:55 Primary Screening on the hERG Channel Using Automated Patch Clamp
Mads P.G. Korsgaard, Ph.D., Senior Research Scientist, Ion Channel Pharmacology, NeuroSearch A/S
hERG (human ether-à-go-go-related gene) encodes the channel protein that is the molecular correlate of the potassium current known as I(Kr) in the heart. Compared to I(Ks) which is carried through Kv7.1/minK (KCNQ1/minK) channels, the kinetics of the hERG channel is characterized by fast activation with even faster inactivation occurring simultaneously which results in only a small current during depolarization. Upon re-polarization the channel transition from inactivated (non-conducting) to closed goes via the open channel state and gives rise to a transient tail current followed by deactivation to the closed channel. Together with KCNQ1/mink the hERG channel is important in the repolarization phase of the heart action potential and loss of function in these channels lead to prolongation of the QT interval in the ECG as a result of compro-mised repolarization. The tail current rises sharply (few ms) before is decays (100s of ms) and in physiological K+-gradients results in an outward current. Pharmacological effects can be mediated via interaction with different sites on the channel complex with effects differing from pore block, diminished/increased inactivation, accelerated activation as well as modulation of the deactivation kinetics. These potential properties make it difficult to measure modulation of the channel with indirect methods but necessitate electrophysiology in an attempt to detect all sorts of pharmacological effects. At NeuroSearch we used the QPatchHT to conduct a screening campaign for modulators of the hERG channel on an in-house library of 10,000 compounds. At a pace of app. 500 compounds (n = 4) a week we tested single concentrations (10 µM) by multiple additions to each whole-cell. On average the whole-cells each received 8 compound additions compared to a theoretical (waste-limited) maximum of 12. Potential hits were detected with an arbitrary cut-off for effect on tail current and follow-up validation experiments were conducted also on the QPatchHT by concentration response and only one compound pr. whole-cell. The stability of the system, the analysis software and the suitability and limitations of the QPatchHT for larger primary electrophysiological screening campaigns will be discussed.

12:25 pm Luncheon Workshop (Sponsorship Available) or Lunch on Your Own 

12:55 Session Break

TARGETING PAIN MANAGEMENT

1:55 Chairperson’s Remarks 
Iain Chessell, Ph.D., Head, Pain Research, GlaxoSmithKline

2:00 Novel and Translatable Mechanisms for Pain Treatment - Lessons From the P2X Receptor Family 
Iain Chessell, Ph.D.
The P2X receptor ion channel family offers multiple opportunities for drug development in the pain area. However, the diversity of localisation, mechanisms and tractability makes choice of a single molecular target difficult. We have characterised some of the mechanisms which underlie activity of individual members of the P2X family in the pain pathway, and also have utilised translational technologies to provide prioritisation between these targets. In this presentation the evidence for roles of the P2X receptor family members in pain will be reviewed along with underlying mechanisms, and the use of translational markers to provide early assesment of clinical pharmacodynamic activity will be presented. 

2:30 TRP Channels as Drug Targets 
David E. Clapham, M.D., Ph.D., HHMI, Cardiology, Children’s Hospital Boston; Neurobiology, Harvard Medical School
TRP channels are cation channels with polymodal activation properties. Mammalian TRP channel proteins are six transmembrane (6-TM) cation-permeable channels that may be grouped by homology into 6 sub-families. Advances in understanding the function of these channels, and their potentials as therapeutic targets, will be summarized.

3:00 Networking Refreshment Break in the Exhibit Hall

3:40 Identification and Characterisation of Novel Small Molecule
Inhibitors of the Kv1.x/KvβChannel Subunit Interactions for Pain Management
Roland Kozlowski, Ph.D., Chief Executive Officer, Lectus Therapeutics, Ltd.
The identification of inhibitors of the interaction between Kv1 channels and their regulatory Kv1.x/Kvβ subunits will be presented. This will include the development of Kv1.x/Kvβ small molecule inhibitors, LEPTICS® protein-protein interaction assays, high throughput screening data and hit characterisation. Secondary in vitro electrophysiological characterisation of these novel inhibitors of Kv1.x/Kvβ subunits in rat DRGs will be presented in addition to in vivo evaluation of these novel inhibitors in models of inflammatory and neuropathic hyperalgesia. 

4:10 Role of Nav1.8 in Chronic Pain States
Douglas S. Krafte, Ph.D., Vice President – Biology, Icagen Inc.
Sodium channels are important regulators of neuronal excitability and play a key role in both normal and aberrant pain signaling. We have been able to probe the role of one specific sodium channel, Nav1.8, by using a selective small molecule blocker of the channel. This compound, A-803467, blocks Nav1.8 in vitro and shows efficacy in a variety of pain models when tested in vivo. Such subtype selective molecules should allow a greater understanding of the role various sodium channels play in regulating pain sensation.

4:40 Potent and Selective TRPV1 Antagonist for Pain Management
Arthur Gomtsyan, Ph.D., Associate Research Fellow, Global Pharmaceutical Research and Development, Neuroscience Research, Abbott Laboratories 
Therapeutic potential of vanilloid receptor TRPV1, a member of the transient potential ion channel family, is well documented. This receptor is considered the principle integrator of noxious stimuli, which points to a significant role that TRPV1 may play in pain pathways. While both TRPV1 agonists and antagonists are being targeted as potential analgesics, the major focus of the drug discovery effort is on the identification of TRPV1 antagonists. We discovered a novel, potent and selective TRPV1 antagonist that possesses broad-spectrum analgesic profile in preclinical models of chronic pain and is suitable for clinical development. This presentation will describe SAR, pharmacokinetic and pharmacological properties of the featured compound.

5:10 Close of Ion Channels Conference