DEVELOPMENT OF GM-CSF ANTAGONIST PEPTIDES

Granulocyte/macrophage colony stimulating factor (GM-CSF) is both a he matopoietic growth factor and a cytokine implicated in inflammatory di sease. The development of GM-CSF antagonist peptides corresponding to the GM-CSF native sequence should allow their modification into higher affinity analogs, but this is hampered by the low affinity of linear peptides. To adequately evaluate such low affinity peptides, the use o f several independent assays should allow specific versus nonspecific inhibitors to be distinguished In this study; inhibition of GM-CSF-dep endent cell growth, inhibition of GM-CSF binding and immunologic cross reactivity between GM-CSF-derived peptides and native protein by neutr alizing antibodies have been used to evaluate peptide analogs with pot ential bioactivity. The GM-CSF sequence was divided into 6 peptides ra nging in size from 15-24 amino acids. Antisera were raised to these pe ptides in mice and assayed for immunologic cross-reactivity. 4/6 anti- peptide antisera bound GM-CSF on ELISA and 3/6 on immunoprecipitation. Antisera to two of the peptides (corresponding to residues 17-31 and 96-112) inhibited GM-CSF-dependent cellular proliferation in two cell lines, with one peptide derived from residues 17-31 demonstrating inhi bition of GM-CSF binding and direct biological inhibitory activity. A peptide that did not elicit native GM-CSF reactive antibodies, corresp onding to residues 54-78, was recognized by two neutralizing monoclona l antibodies. It exhibited inhibition of GM-CSF binding and direct bio logical antagonist activity. These studies implicate two sites in medi ating GM-CSF biological activity, and indicate that biological antagon ists can be developed based on these sites.