Binding and signalling properties of a growth hormone enhancing monoclonalantibody

We have used a sequential, qualitative biosensor based assay to demonstrate that OA15, a monoclonal antibody which enhances in vivo the activity of bo vine growth hormone (bGH) does not disrupt the interaction between bGH and its cognate receptor (as represented by recombinant bovine GH binding prote in -rbGHBP). We have confirmed this using a classical cell-based radio-rece ptor assay with the GH-responsive mouse pre-adipocyte cell line 3T3-F442A. The fact that OA15 binding to bGH still allows hormone to interact with its receptor, allows us to test the hypothesis that there is any amplification of signalling events following hormone-MAb treatment of 3T3-F442A cells. W e have used as a reporter of GH activity the rapid stimulation of JAK-2 tyr osine phosphorylation which is a critical first step in GH signalling event s. We demonstrate that binding of rbGH by OA15 attenuates hormone stimulati on of JAK-2 tyrosine phosphorylation. We conclude that although OA15 does n ot disrupt GH-GH receptor (GHR) interactions it does interfere with subsequ ent GH activity at the molecular and cellular level. We further speculate t herefore that the biological enhancing activity of this antibody is most li kely due to an in vivo effect as presentation of antibody-hormone complexes to a GH-target cell inhibits hormone activity.