A system for simultaneous evaluation of the effect of signal transduction inhibitors on interleukin-2 synthesis and cell viability in a human T cell line.

Abstract

The signal transduction pathways involved in interleukin-2 (IL-2) synthesis have become a focus of interest for pharmacological intervention. Regulation of synthesis of IL-2 requires costimulation of two cell surface receptors, the T cell receptor and an appropriate costimulatory receptor. Antibodies such as anti-CD3 to stimulate the T cell receptor and anti-CD28 can be used to induce this costimulation. Previous methodologies employed antibody coupled to polystyrene microtiter plates or solution phase stimulation. Here, a method using magnetic beads to present anti-CD3 and anti-CD28, to the Jurkat T cell line was developed. Conditions were also developed for simultaneous analysis of the effect of test compounds on cell viability. The IL-2 synthesis methods and cell viability methods have been used to evaluate signal transduction inhibitors. This method was then employed to test the effects of various signal transduction inhibitors on IL-2 synthesis. Agents which increase cAMP, cyclosporin and inhibitors of calmodulin, tyrosine kinases/phosphatases, protein kinase C, and PC-PL-C decrease IL-2 synthesis at concentrations which do not affect cell viability. These results found with this novel system of stimulation with antibody to CD3 and CD28 compare favorably with activities reported in the literature thus validating this rapid facile system for evaluation of the effect of signal transduction inhibitors in IL-2 synthesis.