Search

Irvalec© (PM02734, Elisidepsin) is a marine-derived cytotoxic depsipeptide that is currently undergoing phase II clinical studies in non-small cell lung cancer. In vitro treatment of tumor cells with Irvalec© induces necrotic cell death, a process associated with rapid loss of membrane integrity and subsequent cell permeabilization. These results suggested an effect of the drug, either on an ion channel or in the plasma membrane. In order to explore this hypothesis patch-clamp experiments were performed in different tumor cell lines (HeLa, A549 and HCT116). Treated cells underwent rapid and dramatic morphological changes, including cell blebbing, severe swelling, plasma membrane permeabilization and cell lysis. Apart from the numerous small blebs, membranes from damaged cells also re-organized to form enormous bubbles surrounded by cell membrane. Using electrophysiological techniques, it was shown that Irvalec© induced an important increase in membrane conductance. The compound permeabilized the plasma membrane to ions, even when the cells were not pulsed, causing important changes in the holding current. It has been described that zinc attenuates the drastic effects of some membrane disrupting agents. Hence, to test if zinc exerted some protective effect against Irvalec© effects, cells were treated with this drug in the presence or absence of zinc salts and its membrane permeability was analyzed by using electrophysiology techniques, measuring the variations in the ion currents induced by the drug. Interestingly, in cells treated with zinc (10 mM), a decrease in the membrane permeability induced by Irvalec© was observed. Altogether, these results suggest that Irvalec© induces a rapid membrane permeabilization that lead to a necrotic cell death.

Irvalec is a marine-derived antitumor agent currently undergoing phase II clinical trials. In vitro, Irvalec induces a rapid loss of membrane integrity in tumor cells, accompanied of a significant Ca(2+) influx, perturbations of membrane conductivity, severe swelling and the formation of giant membranous vesicles. All these effects are not observed in Irvalec-resistant cells, or are significantly delayed by pretreating the cells with Zn(2+). Using fluorescent derivatives of Irvalec it was demonstrated that the compound rapidly interacts with the plasma membrane of tumor cells promoting lipid bilayer restructuration. Also, FRET experiments demonstrated that Irvalec molecules localize in the cell membrane close enough to each other as to suggest that the compound could self-organize, forming supramolecular structures that likely trigger cell death by necrosis through the disruption of membrane integrity.

Kv1.3 plays a crucial role in the activation and proliferation of T-lymphocytes and macrophages. While Kv1.3 is responsible for the voltage-dependent potassium current in T-cells, in macrophages this K+ current is generated by the association of Kv1.3 and Kv1.5. Patients with autoimmune diseases show a high number of effector memory T cells that are characterized by a high expression of Kv1.3 and Kv1.3 antagonists ameliorate autoimmune disorders in vivo. Diclofenac is a non-steroidal anti-inflammatory drug (NSAID) used in patients who suffer from painful autoimmune diseases such as rheumatoid arthritis. In this study, we show that diclofenac impairs immune response via a mechanism that involves Kv1.3. While diclofenac inhibited Kv1.3 expression in activated macrophages and T-lymphocytes, Kv1.5 remained unaffected. Diclofenac also decreased iNOS levels in Raw 264.7 cells, impairing their activation in response to lipopolysaccharide (LPS). LPS-induced macrophage migration and IL-2 production in stimulated Jurkat T-cells were also blocked by pharmacological doses of diclofenac. These effects were mimicked by Margatoxin, a specific Kv1.3 inhibitor, and Charybdotoxin, which blocks both Kv1.3 and Ca2+-activated K+ channels (KCa3.1). Because Kv1.3 is a very good target for autoimmune therapies, the effects of diclofenac on Kv1.3 are of high pharmacological relevance. © 2010 Elsevier Inc.

Voltage-dependent potassium (K-v) channels play a pivotal role in the modulation of macrophage physiology. Macrophages are professional antigen-presenting cells and produce inflammatory and immunoactive substances that modulate the immune response. Blockage of K-v channels by specific antagonists decreases macrophage cytokine production and inhibits proliferation. Numerous pharmacological agents exert their effects on specific target cells by modifying the activity of their plasma membrane ion channels. Investigation of the mechanisms involved in the regulation of potassium ion conduction is, therefore, essential to the understanding of potassium channel functions in the immune response to infection and inflammation. Here, we demonstrate that the biophysical properties of voltage-dependent K+ currents are modified upon activation or immunosuppression in macrophages. This regulation is in accordance with changes in the molecular characteristics of the heterotetrameric K(v)1.3/K(v)1.5 channels, which generate the main K-v in macrophages. An increase in K+ current amplitude in lipopolysaccharide-activated macrophages is characterized by a faster C-type inactivation, a greater percentage of cumulative inactivation, and a more effective margatoxin (MgTx) inhibition than control cells. These biophysical parameters are related to an increase in K(v)1.3 subunits in the K(v)1.3/K(v)1.5 hybrid channel. In contrast, dexamethasone decreased the C-type inactivation, the cumulative inactivation, and the sensitivity to MgTx concomitantly with a decrease in K(v)1.3 expression. Neither of these treatments apparently altered the expression of K(v)1.5. Our results demonstrate that the immunomodulation of macrophages triggers molecular and biophysical consequences in K(v)1.3/K(v)1.5 hybrid channels by altering the subunit stoichiometry.

Several epidemiologic and clinical studies show that following myocardial infarction, dietary supplements of omega-3 polyunsaturated fatty acids (omega3FA) reduce sudden death. Animal data show that omega3FA have antiarrhythmic properties, but their mechanisms of action require further elucidation. The effects of omega3FA supplementation were studied in female rabbits to analyze whether their antiarrhythmic effects are due to a reduction of triangulation, reverse use-dependence, instability, and dispersion (TRIaD) of the cardiac action potential (TRIaD as a measure of proarrhythmic effects). In Langendorff-perfused hearts challenged by a selective rapidly activating delayed rectifier potassium current inhibitor that has been shown to exhibit proarrhythmic effects (dofetilide; 1 to 100 nM), omega3FA pretreatment (30 days; n=6) prolonged the plateau phase of the monophasic action potential; did not slow the terminal fast repolarization; reduced the dofetilide-induced prolongation of the action potential duration; reduced dofetilide-induced triangulation; and reduced dofetilide-induced reverse use-dependence, instability of repolarization, and dispersion. Dofetilide reduced excitability in omega3FA-pretreated hearts but not in control hearts. Whereas torsades de pointes (TdP) were observed in five out of six in control hearts, none were observed in omega3FA-pretreated hearts. Docosahexaenoic acid (DHA) inhibited the sodium current with ultrafast kinetics. Dietary omega3FA supplementation markedly reduced dofetilide-induced TRIaD and abolished dofetilide-induced TdP. Ultrafast sodium channel block by DHA may account for the antiarrhythmic protection of the dietary supplements of omega3FA against dofetilide-induced proarrhythmia observed in this animal model.

Ionic channels are membrane proteins that can be blocked by many different types of drugs such as local anesthetics, antiarrhythmics, etc. Therefore, they are considered drug targets, whose topology, at the ion channel level, has been analyzed by studying the interactions of specific ion channel blockers and site-directed mutant ion channels. Stereoselective interactions are especially interesting because they can reveal three-dimensional relationships between drugs and channels with otherwise identical biophysical and physicochemical properties. Furthermore, stereoselectivity suggests direct and specific receptor-mediated action, and identification of such stereospecific interactions may have important clinical consequences. However, ≈25% of drugs used in clinical practice are racemic mixtures, the individual enantiomers of which frequently differ in both their pharmacodynamic and pharmacokinetic profile. Furthermore, these different effects induced by one of the enantiomers of a racemic drug may contribute to the undesired effects that can be similar or different to the pharmacological effect of the racemic drug. In other cases, the enantiomers on the molecular target are opposite. In this review, we focus on the stereoselective effects of bupivacaine on different Kv channels. Bupivacaine stereoselectively blocks Kv1.5 and Kv11.1 channels, whereas non-stereoselectively it blocks Kv2.1 and Kv4.3 channels. © 2005 Bentham Science Publishers Ltd.