FUNCTIONAL EPITOPE MAPPING OF INSULIN-LIKE GROWTH-FACTOR-I (IGF-I) BYANTI-IGF-I MONOCLONAL-ANTIBODIES

Based on a collection of monoclonal antibodies (mAb) against insulin-l ike growth factor I(IGF-I), we have defined the IGF-I epitopes involve d in the interaction with IGF-binding proteins (IGFBP) and IGF-I recep tors. We have also characterized the ability of these antibodies to bl ock IGF-I-induced survival of the IL-3-dependent Ba/F3 cell line. More than 140 hybridomas secreting IGF-I-specific mAb were characterized, of which 28 were studied in detail. They display apparent affinity con stants ranging from less than 10(6) to 10(10) M(-1) and varying crossr eactivity with IGF-II, including 2 mAb with higher affinity for IGF-II than for IGF-I. None crossreact with insulin or any other growth fact or tested. Using both enzyme immunoassays and real-time biospecific in teraction analysis, we have identified 8 epitopic clusters related to the primary structure of IGF-I, according to mAb reactivity to synthet ic peptides, proteolytic fragments of IGF-I, and various IGF-I mutants . The mAb panel also was used to map the IGF domains implicated in the interaction with IGFBP and IGF-I receptors. An IGF-I domain has been identified that remains exposed after IGF-I binding to IGFBP-1 or to I GFBP-3, which is recognized by 6 different mAb. The mAb in this group also bind IGF-I, when complexed to the type-1 IGF receptor on the muri ne pro-B cell line Ba/F3, and BALB/c 3T3 fibroblasts overexpressing th e human receptor. Finally, IGF-I-promoted survival can be blocked with mAb specific for target epitopes, and their potential use in tumor ce ll growth control is discussed.