Binding of Doxorubicin-Conjugated Transferrin to U937 Cells

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Binding of transferrin (Trf) and its doxorubicin-conjugated forms (Conj) to U937 cells at 0 degrees C were compared using 125I-labelled Trf or Conj. The apparent binding affinity (Ka) of Conj to the surface of U937 cells was (1.9 +/- 0.4).10(8) l/mol; it is about 40% of that of Trf [(5.0 +/- 1.2).10(8) l/mol]. Binding of 125I-labelled ligands was blocked by the unlabelled ligands to the same degree, however, it was not blocked by a great excess of doxorubicin (Dox). N-ethylmaleimide caused about 10% inhibition while dithiothreitol was without effect. Dissociation of 125I-labelled ligands in the presence of different concentrations of unlabelled ligands (Trf and Conj in the all 4 variations) resulted in different R50 values (the concentration of the unlabelled ligand where 50% of the radiolabelled ligand was released). While Trf displaced Trf with an R50 value close to the binding affinity, Conj displacement by Conj occurred with much lower efficiency. The heterolog displacement experiments yielded R50 values in between the two extrema. These results suggest that 1) binding of Conj to the surface of cells is governed by the binding of the Trf part of Conj to the transferrin receptor, 2) -SH groups are not involved in the binding, and 3) a second interaction between the Conj and some constituent(s) of the plasma membrane may modify the binding of Conj in comparison to that of Trf.