Referenced in: none

Automatically associated channels: Kv10.1

Title: Evaluation and Optimization of Compound Solubilization and Delivery Methods in a Two-Tiered Ion Channel Lead Optimization Triage.

Authors: Adam W Hendricson, Liz Gallagher, Michele Matchett, Meredith Ferrante, Steve Spence, Tony Paiva, Wilson Shou, Svetlana Tertyshnikova, Mike Krambis, Deborah Post-Munson, Litao Zhang, Ron Knox

Journal, date & volume: , 2011 Nov 15 , ,

PubMed link: http://www.ncbi.nlm.nih.gov/pubmed/22085018

Abstract
Low-volume dispensing of neat dimethyl sulfoxide (DMSO) into plate-based assays conserves compound, assay reagents, and intermediate dilution plate cost and, as we demonstrate here, significantly improves structure-activity relationship resolution. Acoustic dispensing of DMSO solutions into standard volume 384W plates yielded inconsistent results in studies with 2 cell lines because of apparent effects on the integrity of the cell monolayer (increased intracellular Ca⁺⁺ levels as indicated by elevated basal dye fluorescence after acoustic transfer). PocketTip-mediated transfer was successful at increasing apparent potency on a more consistent basis. Notably, the correlation coefficient among fluorescence imaging plate reader (FLIPR):electrophysiology (EP) across a representative ~125 compound collection was increased ~5× via conversion to a PocketTip direct dispensation, indicating a triage assay more predictive of activity in the decisional patch-clamp assay. Very importantly, the EP-benchmarked false-negative rate as measured by compounds with FLIPR EC₅₀ more than the highest concentration tested fell from >11% to 5% assay-wide, and the relative FLIPR:EP rank-order fidelity increased from 55% to 78%. Elimination of the aqueous intermediate step provided additional benefits, including reduced assay cost, decreased cycle time, and reduced wet compound consumption rate. Direct DMSO dispensing has broad applicability to cell-based functional assays of multiple varieties, especially in cases where limit solubility in assay buffer is a recognized impediment to maximizing interassay connectivity.