( Sylviane Komesli...)Use of FlashPlate Technology for In Vitro Measurement of 125I-Labeled TGF-1 Binding on Chimeric Extracellular Domain of Type II Transforming Growth Factor Receptor

Sylviane Komesli, Martine Page, and Patrick Dutartre
Dpartement dImmunologie, Laboratoires Fournier S.A., France

Abstract

TGF- type I and type II receptors form ligand dependent heteromeric signaling complexes, in which transforming growth factor- receptor type II (TRII) tends to act as the primary receptor. In the present study, we investigated the feasibility of using the FlashPlate platform to assay a chimeric soluble type II receptor fused with the Fc regions of human immunoglobulin (TRIIs-Fc), in order to screen for potent agonists and antagonists of TGF.

Introduction

Transforming growth factors 1, 2 and 3 (TGF-s) are multifunctional cytokines involved in the regulation of cell proliferation, differentiation and extracellular matrix production. In mammalian cells, responses to TGF- are mediated by types I and II cell surface receptors (TRI and TRII respectively) which are expressed in most cell types and tissues. TGF- binds directly to TRII, allowing this receptor to associate with and phosphorylate TRI, which then propagates the signal through activation of Smad2 and Smad4 heteromeric complexes¹.

At this time, there are no known, clinically useful TGF- agonists or antagonists. A better understanding of the mechanism of activation of the TGF- receptor complexes may be useful, therefore, as a step towards the development of TGF- antagonist or agonist drugs. We have investigated the possibility of using the soluble extracellular domain of TRII in a nonseparation microplate receptor binding assay.

To facilitate this approach and increase the probability of success, we constructed a chimeric soluble receptor, by fusing the extracellular domain of TRII to the Fc regions of human immunoglobulin (TRIIs-Fc). Through the Fc region, the chimeric receptor, TRIIs-Fc, expressed in a transiently tran
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