EVIDENCE FOR MULTIPLE BINDING SITES FOR SEVERAL SPECIES OF HU IFN-ALPHA

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The antiproliferative and competitive binding activities of several purified species of Hu lymphoblastoid Interferon-alpha were compared to that of recombinant Interferon-alpha 2b on Daudi and AU937 cells. We found that the antiproliferative activity of the interferon species on each cell line has a broad range and that Daudi cells were more sensitive than AU937 cells. The 50% inhibition of cell growth ranged from .003 ng/ml to greater than 10 ng/ml on Daudi cells and 1.2 ng/ml to >100 ng/ml on AU937 cells. Scatchard analyses demonstrated that the IFN-alpha 2b binding site number is greater on Daudi cells (12,700 binding sites/cell) than AU937 cells (3,300 binding sites/cell); however, their receptor affinities were similar. Interestingly, there appears to be three species (n,o,p) which exhibit the highest antiproliferative activities on both cell lines but do not compete for the Interferon-alpha-2b binding site. These data suggest that there may be more than one receptor or multiple receptor components involved in the biological actions of these molecules. In order to further study different receptors among species O IFN-alpha2b and other species, we have radiolabeled species O and performed competitive binding experiments. We also performed cross-linking to studies with IFN-alpha2b and species O to compare the molecular properties of the binding sites.