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- Wacker, Soeren J., et al.
(författare)
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Identification of Selective Inhibitors of the Potassium Channel Kv1.1-1.2(3) by High-Throughput Virtual Screening and Automated Patch Clamp
- 2012
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Ingår i: ChemMedChem. - : Wiley. - 1860-7179 .- 1860-7187. ; 7:10, s. 1775-1783
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Tidskriftsartikel (refereegranskat)abstract
- Two voltage-dependent potassium channels, Kv1.1 (KCNA1) and Kv1.2 (KCNA2), are found to co-localize at the juxtaparanodal region of axons throughout the nervous system and are known to co-assemble in heteromultimeric channels, most likely in the form of the concatemer Kv1.11.2(3). Loss of the myelin sheath, as is observed in multiple sclerosis, uncovers the juxtaparanodal region of nodes of Ranvier in myelinated axons leading to potassium conductance, resulting in loss of nerve conduction. The selective blocking of these Kv channels is therefore a promising approach to restore nerve conduction and function. In the present study, we searched for novel inhibitors of Kv1.11.2(3) by combining a virtual screening protocol and electrophysiological measurements on a concatemer Kv1.11.2(3) stably expressed in Chinese hamster ovary K1 (CHO-K1) cells. The combined use of four popular virtual screening approaches (eHiTS, FlexX, Glide, and Autodock-Vina) led to the identification of several compounds as potential inhibitors of the Kv1.11.2(3) channel. From 89 electrophysiologically evaluated compounds, 14 novel compounds were found to inhibit the current carried by Kv1.11.2(3) channels by more than 80?% at 10 mu M. Accordingly, the IC50 values calculated from concentrationresponse curve titrations ranged from 0.6 to 6 mu M. Two of these compounds exhibited at least 30-fold higher potency in inhibition of Kv1.11.2(3) than they showed in inhibition of a set of cardiac ion channels (hERG, Nav1.5, and Cav1.2), resulting in a profile of selectivity and cardiac safety. The results presented herein provide a promising basis for the development of novel selective ion channel inhibitors, with a dramatically lower demand in terms of experimental time, effort, and cost than a sole high-throughput screening approach of large compound libraries.
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| 2. |
- Stary, Anna, et al.
(författare)
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Toward a Consensus Model of the hERG Potassium Channel
- 2010
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Ingår i: ChemMedChem. - : Wiley. - 1860-7179 .- 1860-7187. ; 5:3, s. 455-467
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Tidskriftsartikel (refereegranskat)abstract
- Malfunction of hERG potassium channels, due to inherited mutations or inhibition by drugs, can cause long QT syndrome, which can lead to life-threatening arrhythmias. A three-dimensional structure of hERG is a prerequisite to understand the molecular basis of hERG malfunction. To achieve a consensus model, we carried out an extensive analysis of hERG models based on various alignments of helix S5. We analyzed seven models using a combination of conventional geometry/packing/normality validation methods as well as molecular dynamics simulations and molecular docking. A synthetic test set with the X-ray crystal structure of K(v)1.2 with artificially shifted S5 sequences modeled into the structure served as a reference case. We docked the known hERG inhibitors (+)-cisapride, (S)-terfenadine, and MK-499 into the hERG models and simulation snapshots. None of the single analyses unambiguously identified a preferred model, but the combination of all three revealed that there is only one model that fulfils all quality criteria. This model is confirmed by a recent mutation scanning experiment (P. Ju, G. Pages, R. R Riek, P. C. Chen, A. M. Torres, R S. Bansal, S. Kuyucak, R W. Kuchel, J. I. Vandenberg, J. Biol. Chem. 2009, 284, 1000-1008).([1]) We expect the modeled structure to be useful as a basis both for computational studies of channel function and kinetics as well as the design of experiments.
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| 3. |
- Wacker, Soeren J., et al.
(författare)
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The identification of novel, high affinity AQP9 inhibitors in an intracellular binding site
- 2013
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Ingår i: Molecular Membrane Biology. - : Informa UK Limited. - 0968-7688 .- 1464-5203. ; 30:3, s. 246-260
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Tidskriftsartikel (refereegranskat)abstract
- Background: The involvement of aquaporin (AQP) water and small solute channels in the etiology of several diseases, including cancer, neuromyelitis optica and body fluid imbalance disorders, has been suggested previously. Furthermore, results obtained in a mouse model suggested that AQP9 function contributes to hyperglycemia in type-2 diabetes. In addition, the physiological role of several AQP family members remains poorly understood. Small molecule inhibitors of AQPs are therefore desirable to further study AQP physiological and pathophysiological functions. Methods: The binding of recently established AQP9 inhibitors to a homology model of AQP9 was investigated by molecular dynamics simulations and molecular docking. Putative inhibitor binding sites identified with this procedure were modified by site-directed mutagenesis. Active compounds were measured in a mammalian cell water permeability assay of mutated AQP9 isoforms and tested for changes in inhibitory effects. Controls: Three independent cell lines were established for each mutated AQP9 isoform and functionality of mutant isoforms was established. Principal findings: We have identified putative binding sites of recently established AQP9 inhibitors. This information facilitated successful identification of novel AQP9 inhibitors with low micromolar IC50 values in a cell based assay by in silico screening of a compound library targeting specifically this binding site. Significance: We have established a successful strategy for AQP small molecule inhibitor identification. AQP inhibitors may be relevant as experimental tools, to enhance our understanding of AQP function, and in the treatment of various diseases.
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