Functional epitope mapping of insulin-like growth factor I (IGF-I) by anti-IGF-I monoclonal antibodies.
Based on a collection of monoclonal antibodies (mAb) against insulin-like growth factor I (IGF-I), we have defined the IGF-I epitopes involved in the interaction with IGF-binding proteins (IGFBP) and IGF-I receptors. We have also characterized the ability of these antibodies to block 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 constants ranging from less than 10(6) to 10(10) M-1 and varying crossreactivity with IGF-II, including 2 mAb with higher affinity for IGF-II than for IGF-I. None crossreact with insulin or any other growth factor tested. Using both enzyme immunoassays and real-time biospecific interaction analysis, we have identified 8 epitopic clusters related to the primary structure of IGF-I, according to mAb reactivity to synthetic 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 IGFBP-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 murine pro-B cell line Ba/F3, and BALB/c 3T3 fibroblasts overexpressing the human receptor. Finally, IGF-I-promoted survival can be blocked with mAb specific for target epitopes, and their potential use in tumor cell growth control is discussed.
Mañes
S
S
Department of Immunology and Oncology, Universidad Autonoma de Madrid, Spain. smanes@samba.cnb.uam.es
Kremer
L
L
Albar
J P
JP
Mark
C
C
Llopis
R
R
Martínez
C
C
D000818:Animals; D000911:Antibodies, Monoclonal; D018604:Epitope Mapping; D006801:Humans; D018969:Insulin-Like Growth Factor Binding Proteins / Q000378:metabolism; D007334:Insulin-Like Growth Factor I / Q000276:immunology* / Q000378:metabolism; D051379:Mice; D017451:Receptors, Somatomedin / Q000276:immunology; D011994:Recombinant Proteins; D014407:Tumor Cells, Cultured
D000911:Antibodies, Monoclonal; D018969:Insulin-Like Growth Factor Binding Proteins; D017451:Receptors, Somatomedin; D011994:Recombinant Proteins; D007334:Insulin-Like Growth Factor I