Drug discovery technology for ion channels

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  • Although technologies for ion channelscreening have not kept pace with other HTS assays in the past, emergingtechnologies promise to shift this para-digm in the near future. This was thefocus of the 2nd annual Drug Discoveryfor Ion Channels satellite meeting of the Biophysical Society (22 February 2002,San Francisco, CA, USA), sponsored byAxon Instruments. It represented an op-portunity for screening technology com-panies to report their progress made in2001, to academic and pharmaceuticalor biotechnology investigators interestedin ion channel drug discovery.

    Alan Finkel of Axon Instruments(Union City, CA, USA) set the stage forthe meeting by remarking on the explo-sion of information about ion channelfunction that has occurred over the past20 years with the advent of the patchclamp techniques and more recently, theability to study ion channel functionusing high-throughput fluorescent screen-ing techniques such as the FluorescentImaging Plate Reader (FLIPR; MolecularDevices, Sunnyvale, CA, USA) and theVoltage Ion Plate Reader (VIPR; AuroraBioscience, San Diego, CA, USA).

    However, the excitement at this yearsmeeting was the fast-paced progress in the automated patch-clamp field that has occurred in the past year.Specifically, up until 2002, planar patch-clamp technologies had yet to be vali-dated because no research group hadpresented whole-cell recordings fromsingle mammalian cells at the meeting in2001. By contrast, several groups pre-sented whole-cell data examples at thisyears meeting.

    Pharmaceutical perspectiveEarly discussions focused on how someof the current technologies are imple-mented in the pharmaceutical industrytoday. Owen McManus from Merck(Rahway, NJ, USA) shared his perspectiveon ion-channel technologies used indrug discovery. Specifically, he describedthe use of classical ligand-binding tech-niques, as well as rubidium efflux (whichis useful for assessing K+ channel func-tion). In addition, he described fluor-escent assays on both FLIPR and VIPRplatforms using Fluo-4, a Ca2+ indicatorfor studying vanilloid receptor 1 (VR1).He commented that the patch-clamptechnique is undoubtedly the best tech-nique for ion channel function because it has the required flexibility and tem-poral resolution. Furthermore, he statedthat for high-throughput patch-clamp-ing to be successful, it will need to have the ability to generate robust, high-quality data using established cell linesat reasonable cost with efficient data handling.

    Automated electrophysiologyAxon Instruments launched their auto-mated two-electrode voltage clamp(TEVC) system (OpusXpress) for Xenopusoocytes at the meeting with CathySmith-Maxwell highlighting the featuresof the system. OpusXpress records fromeight oocytes in parallel and can run unattended for a total of 24 automatedsolution changes per oocyte. In additionto Axon Instruments, Michael Fejtl of Multi-Channel Systems (Reutlingen,Germany) presented information abouttheir commercially available TEVC system

    called Roboocyte. The Roboocyte mea-sures ion channel currents from channels expressed in oocytes using a singlerecording head containing the voltagerecording, current injection and groundelectrodes, as well as the perfusion inletand outlet. The Roboocyte is capable ofrunning unattended and will automati-cally inject mRNA or cDNA into theoocytes before recording. In contrast to these TEVC systems, Morton Bech ofSophion Bioscience (Ballerup, Denmark)presented information about their Apatchi-1, an automated patch-clamp system ca-pable of generating whole cell data. Thissystem uses a conventional microscope,amplifier, and patch-clamp pipettes.

    Planar patch-clamp technologiesThe hot topic at the meeting was planarpatch-clamp, a new technology in whichthe conventional glass patch-clamppipette is replaced by a hole in a flat substrate that can be made of variousmaterials including glass, polydimethyl-siloxane (PDMS), or silicon. The mem-brane seal is formed where the cell con-tacts the substrate rather then at the tipof a conventional pipette. Positioning of the cell over the hole is achieved by various means, including electrophore-sis, suction and physical positioning. Thepresentation of data using the planarpatch-clamp technologies began with JiaXu of Aviva Biosciences (San Diego, CA,USA). He presented whole-cell sodiumand potassium currents recorded fromneuroblastoma and Chinese hamsterovary (CHO) cells, respectively, as well ascurrents from rat basophilic leukaemia(RBL-1) cells. The substrate being used

    DDT Vol. 7, No. 12 June 2002 updateconference

    1359-6446/02/$ see front matter 2002 Elsevier Science Ltd. All rights reserved. PII: S1359-6446(02)02329-2 651www.drugdiscoverytoday.com

    Drug discovery technology for ionchannelsWilhelm G. Lachnit and James L. Costantin, Molecular Devices, 1311 Orleans Drive, Sunnyvale, CA, USA 94089, tel: +1 408 548 6016, fax: +1 408 548 6430, e-mail: wilhelm_lachnit@moldev.com

  • update conference DDT Vol. 7, No. 12 June 2002

    652 www.drugdiscoverytoday.com

    by Aviva is a modified silicon chip with theedges of the hole smoothed by thermo-oxidation, which greatly increases thesuccess rate for seal formation. Aviva report that most cells form seals within15 sec and the mean values for seals are~10 G for RBL-1 cells and 4G for CHOcells.

    Chris Mathes of Axon Instruments pre-sented whole-cell potassium currents(Kv1.1) measured in neuroblastoma cellsusing 2 M apertures in PDMS. Axon is currently developing a high-through-put planar patch-clamp machine(PatchXpress) with 16 headstages inparallel, capable of both voltage andcurrent clamp, which they expect tolaunch at the end of 2002. Jan Behrendsof Nanion Technologies (Munich,Germany) presented whole-cell calciumand potassium currents measured fromneuroblastoma and CHO cells, respec-tively. Nanion Technologies uses aborosilicate or quartz microstructured

    substrate with a varied aperture size de-pending on the cell type. Alfred Stettfrom Cytocentrics (Reutlingen, Germany)presented success rates of 97% for cellpositioning and 68% for gigaseal forma-tion using a novel substrate that containsa larger hole (~40 M) for cell position-ing surrounding a smaller hole (16 M)used for seal formation and recording.Some spontaneous whole-cell access(16%) and additional whole-cell accessusing suction was also reported.

    IonWorks platforms that are currentlyunder development at Molecular DevicesCorporation (Sunnyvale, CA) were described by Wilhelm Lachnit. TheIonWorks APC system is an assay de-velopment and safety profiling systemthat offers flexibility and sensitivity torecord whole-cell currents or single channel events from cells, whereas theIonWorks HT system is a high-through-put system capable of generating datafrom 1000 to 3000 patches per day in

    whole-cell mode. Lachnit presentedwhole-cell potassium currents (Kv1.5 andHERG) expressed in CHO and human em-bryonic kidney (HEK) cells generated onthese platforms, which will be availablelater this year.

    Overall, the explosion of planar patch-clamp data over the past year does muchmore than validate proof-of-concept.The success of multiple groups usingwidely varied substrates suggests thatthe planar patch-clamp technology isfeasible and will probably become com-mercially available in the near future.However, which substrate works best forwhich applications is yet to be deter-mined. The next 1218 months shouldprove the value of these novel technol-ogies by providing an enormous oppor-tunity for pharmaceutical companies to screen pharmaceutical compound libraries directly at the electrophysio-logical level and, therefore, effectivelyexploit ion channel targets.

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