Work

Cell Treatment and Lysis in 96-Well Filter-Bottom Plates for Screening Bcr-Abl Activity and Inhibition in Whole-Cell Extracts

Year: 2010

Type: article

Abstract: Although conventional high-throughput screens performed in vitro with purified protein kinases are powerful tools to discover new kinase inhibitors, they are far from ideal for determining efficacy in vivo. As a complementary approach, cell-based, target-driven secondary screens may help predict in vivo compound potency and specificity as well as evaluate bioavailability and toxicity. Here the authors report a simple protocol for treating K562 Bcr-Abl-expressing cells with small-molecule kinase inhibitors in 96-well filter-bottom plates followed by in-plate cell lysis. The lysates were assayed via a solid-phase kinase assay, allowing determination of apparent IC(50) for known Bcr-Abl inhibitors as well as facilitating the screening of a small kinase inhibitor library. This approach may have further applications in generating lysates for analyzing kinase activity and inhibition in other nonadherent suspension cell lines.

Source: SLAS DISCOVERY

DOI: 10.1177/1087057110363307

Authors: Michael R. Mand, Ding Wu, Darren R. Veach, Stephen J. Kron

Institutions: University of Chicago, The University of Texas at Austin, Memorial Sloan Kettering Cancer Center

Language: English

Topic: Chronic Myeloid Leukemia Treatments

Subfield: Hematology

Field: Medicine

Domain: Health Sciences

SDG: Good health and well-being

Open Access status: hybrid

Estimated APC paid: $2,070