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Development of a Novel Dual CCR5-Dependent and CXCR4-Dependent Cell-Cell Fusion Assay System with Inducible gp160 Expression
Changhua Ji
Roche Palo Alto, 3411 Hillview Avenue, Palo Alto, CA 94304 Changhua.ji{at}roche.com
Jun Zhang
Nick Cammack
Surya Sankuratri
In the current study, a novel coreceptor-specific cell-cell fusion (CCF) assay system is reported. The system possesses the following features: dual CCR5-dependent and CXCR4-dependent CCF assays, all stable cell lines, inducible expression of gp160 to minimize cytotoxicity, robust luciferase reporter, and 384-well format. These assays have been validated using various known HIV entry inhibitors targeting various stages of the HIV entry/fusion process, including fusion inhibitors, gp120 inhibitors, CCR5 antagonists, CCR5 antibodies, and CXCR4 antagonists. IC 50data generated from this assay system were well correlated to that from the antiviral assays. The effects of DMSOon this assay systemwere assessed, and a 2-to 3-fold increase in luciferase activitywas observed in the presence of 0.05% to2% DMSO. Although cell-cell fusion efficiencywas enhanced, no changes in drug response kinetics for entry inhibitors were found in the presence of 0.1% or 0.5% DMSO. This assay system has been successfully used for the identification and characterization of thousands of CCR5 inhibitors.
Key Words: HIV • cell-cell fusion • DMSO • luciferase reporter • entry inhibitor
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This version was published on February
1, 2006
Journal of Biomolecular Screening, Vol. 11, No. 1,
65-74 (2006)
DOI: 10.1177/1087057105282959
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