Differential Inhibition of Inducible T Cell Cytokine Secretion by Potent Iron Chelators

Abstract

The authors used the PatchXpress ® 7000A system to measure compound activity at the hERG channel using procedures that mimicked the "gold-standard" conventional whole-cell patch clamp. A set of 70 compounds, including hERG antagonists with potencies spanning 3 orders of magnitude, were tested on hERG302-HEK cells using protocols aimed at either identifying compound activity at a single concentration or obtaining compound potency from a cumulative concentration dependence paradigm. After exposure to compounds and subsequent washout of the wells to determine reversibility of the block, blockade by a reference compound served as a quality control. Electrical parameters and voltage dependence were similar to those obtained using a conventional whole-cell patch clamp. Rank order of compound potency was also comparable to that determined by conventional methods. One exception was flunarizine, a particularly lipophilic compound. The PatchXpress ® accurately identified the activity of 29 moderately potent antagonists, which only weakly displace radiolabeled astemizole and are false negatives in the binding assay. Finally, no false hits were observed from a collection of relatively inactive compounds. High-quality data acquisition by PatchXpress ® should help accelerate secondary screening for ion channel modulators and the drug discovery process. (Journal of Biomolecular Screening 2005:168-181)

Role of hERG potassium channel assays in drug development

“…Typically, hERG potencies measured by automated and manual electrophysiology are in good agreement; however, some compounds are known to appear less potent when tested by automated electrophysiology. [23][24][25][26] Discrepancies typically arise from hydrophobic compounds being adsorbed to the surface of the plate used to hold the compound or to the patch plate itself. Glass-coated compound plates can help to minimize the nonspecific binding to the compound plate but come at a significant cost.…”

Automated Electrophysiology in Drug Discovery

“…The quality of hERG data obtained on these platforms is high, and most known hERG blockers display potencies similar to those observed in conventional voltage clamp experiments [94,95]. However, potency shifts are observed with very hydrophobic compounds, requiring multiple additions to achieve the proper steady-state concentration of free compound [96].…”

A Place for High-Throughput Electrophysiology in Cardiac Safety: Screening hERG Cell Lines and Novel Compounds with the Ion Works HT™ System

“…Some correlations to hydrophobicity and solubility can be made with the chemical structure and classes of compounds, and assessing these characteristics may become an integral part of an ion channel screen to assist in predicting a shifted IC 50 value with an automated system. [20][21][22] Typically with standard patch-clamp hERG experiments, the voltage pulse protocol is applied to the cell for several minutes while exposed to compound (≥ 5 min). The ability to continuously apply the voltage pulse protocol can facilitate obtaining steadystate block.…”