Current Pharmaceutical Design - Volume 11, Issue 21, 2005

Local anesthetic drugs interfere with excitation and conduction by action potentials in the nervous system and in the heart by blockade of the voltage-gated Na channel. Drug affinity varies with gating state of the channel. The drugs show low affinity at slow excitation rates, but high affinity when the channels are opened and inactivated during action potentials at high frequency, as they are during pain or during a cardiac arrhythmia. The drugs are thought to access their binding site in the inner pore by passage through the membrane and entry through the inner pore vestibule. There have been three major developments in the last decade that greatly increase our understanding of their mechanism of action. Firstly, amino acid residues critical to drug binding have been located by mutagenesis, and it is possible to develop a molecular model of the drug binding site. Secondly, a path for drug access directly from the outside has been characterized in the cardiac isoform of the channel. Thirdly, the hypothesis that high affinity binding stabilizes the fast inactivated conformation of the channel has been challenged. Rather, the drug may stabilize a slow inactivated state and immobilize the voltage sensor in domain III in its activated outward position. The combination of mutational study of the cloned Na channels and patch clamp offers the opportunity to understand the detailed molecular mechanism of drug action and to resolve drug structure-function.

TRPV1 is a channel expressed highly in small sensory neurons. TRPV1 is a ligand-gated, cation channel that is activated by heat, acid and capsaicin, a principal ingredient in hot peppers. Because of its possible role as a polymodal molecular detector, TRPV1 is studied most extensively. In mice lacking TRPV1, thermal hyperalgesia induced by inflammation is reduced, suggesting a role for mediating inflammatory pain. Activity of TRPV1 is modulated by actions of various kinases such as protein kinase A and C. Furthermore, phosphorylation by Ca2+-calmodulin-dependent kinase II is required for its ligand binding. TRPV1 is activated by various endogenous lipids, such as anandamide, N-arachidonoyldopamine, and various metabolic products of lipoxygenases. 12-hydroperoxyeicosatetraenoic acid, an immediate metabolic product of 12-lipoxygenase, activates TRPV1 and shares 3-dimensional structural similarity with capsaicin. Because lipoxygenase products can activate TRPV1 in sensory neurons, upstream signals to lipoxygenase/TRPV1 pathway have been questioned. Indeed, bradykinin, a potent pain-causing substance, is now known to activate TRPV1 via lipoxygenase pathway. However, we cannot overlook the sensitizing effect of bradykinin via the phospholipase C or protein kinase C pathway. Interestingly, histamine, a pruritogenic substance, also appears to use the lipoxygenase/TRPV1 pathway in order to excite sensory neurons. Because of its role in the mediation of nociception, antagonists of TRPV1 are targeted for development of potential analgesics. In the present review, theoretical background of organic synthesis of SC0030, a potent antagonist of TRPV1 is presented.

ATP-sensitive K+ channels, termed KATP channels, provide a link between cellular metabolism and membrane electrical activity in a variety of tissues. Channel isoforms have been identified and are targets for compounds that both stimulate and inhibit their activity resulting in membrane hyperpolarization and depolarization, respectively. Examples include relaxation of vascular smooth muscle and stimulation of insulin secretion. This article reviews the cloning, molecular biology, and structure of KATP channels, with particular focus on the SUR1/KIR6.2 neuroendocrine channels that are important for the regulation of insulin secretion. We integrate the extensive pharmacologic structure-activityrelationship data on these channels, which defines a bipartite drug binding pocket in the SUR (sulfonylurea receptor), with recent structure-function studies that identify domains of SUR and KIR6.2, the channel pore, which are critical for channel assembly, for gating, and for the ligand-receptor interactions that modulate channel activity. The atomic structure of a sulfonylurea in a protein pocket is used to develop insight into the recognition of these compounds. A homology model of KATP channels, based on VC-MsbA, another member of the ABC protein family, is described and used to position amino acids important for the action of channel openers and blockers within the core of SUR. The model has a central chamber which could serve as a multifaceted binding pocket.

Searching the DNA database has led to the identification of a class of K+ channels now referred to as two-pore or tandem-pore domain K+ (K2P) channels. The K2P channel is structurally unique in that each subunit possesses two poreforming domains and four transmembrane segments. In mammals, sixteen K2P channel genes have been identified, and their mRNA transcripts are expressed in many different cell types and tissues. K2P channels have properties of background or leak K+ channels, and therefore play a crucial role in setting the resting membrane potential and regulating cell excitability. Some K2P channels are activated by certain physical and chemical factors such as lipids, volatile anesthetics, heat, oxygen, protons and membrane tension. Some K2P channels are targets of agonists that bind receptors coupled to different types of G proteins, and are probably involved in a variety of neurotransmitter and peptide hormone-mediated signal transduction processes. Such diverse properties of K2P channels suggest that they are involved in many different physiological and pathophysiological processes. Therefore, K2P channels could become potentially important therapeutic targets for the treatment of various pathological conditions.

Idiopathic epilepsies are genetically determined diseases of the central nervous system characterized by typical epileptic seizures and EEG abnormalities but not associated with structural brain lesions. In recent years, an increasing number of mutations associated with idiopathic epilepsy syndromes were identified in genes encoding subunits of voltageor ligand-gated ion channels. These encouraging results provide a plausible pathophysiological concept, since ion channels form the basis for neuronal excitability and are the major targets for anticonvulsive pharmacotherapy. The first epilepsy genes were identified for rare autosomal dominant syndromes within large pedigrees. Recently, a few mutations were also found for the frequent classical forms of idiopathic generalized epilepsies (IGE), for example absence or juvenile myoclonic epilepsy. The mutations can affect ion channels which on one hand have been known since several decades to be crucial for neuronal function, such as the voltage-gated sodium channel or the GABAA receptor, or on the other hand were newly identified within the last decade as KCNQ potassium channels or the ClC-2 chloride channel. Functional studies characterizing the molecular defects of the mutant channels point to a central role of GABAergic synaptic inhibition in the pathophysiology of IGE. Furthermore, newly discovered genes may be suitable as novel targets for pharmacotherapy such as KCNQ channels for the anticonvulsant drug retigabine. Altogether, these genetic and pathophysiological investigations will enhance our knowledge about the understanding of epileptogenesis and can help to improve anticonvulsive therapy.

The passage of ions to form and maintain electrochemical gradients is a key element for regulating cellular activities and is dependent on specific channel proteins or complexes. Certain ion channels have been the targets of pharmaceuticals that have had impact on a variety of cardiovascular and neurological diseases. Chloride channels regulate the movement of a major cellular anion, and in so doing they in part determine cell membrane potential, modify transepithelial transport, and maintain intracellular pH and cell volume. There are multiple families of chloride channel proteins, and respiratory, neuromuscular, and renal dysfunction may result from mutations in specific family members. Interest in chloride channels related to cancer first arose when the multidrug resistance protein (MDR/ P-glycoprotein) was linked to volume-activated chloride channel activity in cancer cells from patients undergoing chemotherapy. More recently, CLC, CLIC, and CLCA intracellular chloride channels have been recognized for their contributions in modifying cell cycle, apoptosis, cell adhesion, and cell motility. Moreover, advances in structural biology and high-throughput screening provide a platform to identify chemical compounds that modulate the activities of intracellular chloride channels thereby influencing chloride ion transport and altering cell behavior. This review will focus on several chloride channel families that may contribute to the cancer phenotype and suggest how they may serve as novel targets for primary cancer therapy.

The molecular identification of novel ion channels expressed in the immune context provides very attractive potential targets for new therapeutic drugs designed to modulate immune system function. Several members of the recently discovered TRPM family of cation channels are expressed in immune cells. The TRPM channels show an astonishing diversity regarding their gating mechanisms and permeability profiles. First functional studies demonstrate that this diversity correlates with the range of potential biological processes TRPM channels might be involved in. Even though this field of research is still in its infancy, new insights into the regulation of ion homeostasis in immune cells start to emerge. An overview of the current knowledge about the TRPM family members in general, and in the immune context in particular, will be given throughout this review.

The MEK/MAPK signaling module is a key integration point along signal transduction cascades that regulate cell growth, survival, and differentiation, and is aberrantly activated in many human tumors. In tumor cells, constitutive MAPK activation affords increased proliferation and resistance to apoptotic stimuli, including classical cytotoxic drugs. In most instances, however, MAPK inhibition has cytostatic rather than cytotoxic effects, which may explain the lack of objective responses observed in early clinical trials of MEK inhibitors. Nevertheless, amenability of the MAPK pathway to pharmacodynamic evaluation and negligible clinical toxicity make MEK inhibitors an ideal platform to build pharmacological combinations with synergistic antitumor activity. In AML, the MEK/MAPK pathway is constitutively activated in the majority of cases (75%), conferring a uniformly poor prognosis; in preclinical models of AML, MEK blockade profoundly inhibits cell growth and proliferation and downregulates the expression of several anti-apoptotic players, thereby lowering the apoptotic threshold. Apoptosis induction, however, requires concentrations of MEK inhibitors much higher than those required to inhibit proliferation. Nevertheless, MEK blockade efficiently and selectively sensitizes leukemic cells to sub-optimal doses of other apoptotic stimuli, including classical cytotoxics (nucleoside analogs, microtubule-targeted drugs, γ-irradiation), biologicals (retinoids, interferons, arsenic trioxide), and, most interestingly, other signal transduction/apoptosis modulators (UCN-01, STI571, Bcl-2 antagonists). In most instances, these MEK inhibition-based combinations result in a striking pro-apoptotic synergism in preclinical models. Here we briefly discuss evidence suggesting that MAPK pathway inhibition could play a prominent role in the development of integrated therapeutic strategies aimed at synergistic anti-leukemic effects.

It is well documented that sensory transmission, including pain, receives endogenous inhibitory modulatory influences at dorsal horn of the spinal cord. Recent results, from behavioral to molecular studies, demonstrate that injury caused plastic changes in forebrain areas. In addition to encoding pain, these supraspinal areas may also affect pain transmission in the spinal cord level by activating “top-down” descending facilitatory systems. In this review, I provide review of evidence related to these new progresses, from human brain imaging to work from genetically mutant mice.