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A novel multi-parameter assay to dissect the pharmacological effects of different modes of integrin ?L?2 inhibition in whole blood.

ABSTRACT: BACKGROUND AND PURPOSE:The integrin ?L?2 plays central roles in leukocyte adhesion and T cell activation, rendering ?L?2 an attractive therapeutic target. Compounds with different modes of ?L?2 inhibition are in development, currently. Consequently, there is a foreseeable need for bedside assays, which allow assessment of the different effects of diverse types of ?L?2 inhibitors in the peripheral blood of treated patients. EXPERIMENTAL APPROACH:Here, we describe a flow cytometry-based technology that simultaneously quantitates ?L?2 conformational change upon inhibitor binding, ?L?2 expression and T cell activation at the single-cell level in human blood. Two classes of allosteric low MW inhibitors, designated ? I and ?/? I allosteric ?L?2 inhibitors, were investigated. The first application revealed intriguing inhibitor class-specific profiles. KEY RESULTS:Half-maximal inhibition of T cell activation was associated with 80% epitope loss induced by ? I allosteric inhibitors and with 40% epitope gain induced by ?/? I allosteric inhibitors. This differential establishes that inhibitor-induced ?L?2 epitope changes do not directly predict the effect on T cell activation. Moreover, we show here for the first time that ?/? I allosteric inhibitors, in contrast to ? I allosteric inhibitors, provoked partial downmodulation of ?L?2, revealing a novel property of this inhibitor class. CONCLUSIONS AND IMPLICATIONS:The multi-parameter whole blood ?L?2 assay described here may enable therapeutic monitoring of ?L?2 inhibitors in patients' blood. The assay dissects differential effect profiles of different classes of ?L?2 inhibitors.

SUBMITTER: Welzenbach K 

PROVIDER: S-EPMC4621979 | BioStudies | 2015-01-01

REPOSITORIES: biostudies

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