Spectroscopic studies of simultaneous binding of cyclophosphamide and imatinib mesylate to human holo-transferrin

The interactions of proteins with drugs are very important from a pharmacological point of view. Holo-transferrin is a blood-plasma glycoprotein whose main function is iron-binding and the transport of other ligands. Additionally, the protein is only transferrin-form recognized by TfR1 and TfR2 receptors at the surface of rapidly proliferating malignant cells. Imatinib mesylate is a tyrosine-kinase inhibitor mainly used in the treatment of blood cancers, frequently in multidrug therapy with cyclophosphamide. In this study the effect of cyclophosphamide on the interaction of imatinib mesylate with human holo-transferrin has been investigated. Using spectroscopic techniques such as fluorescence, circular dichroism, ultraviolet-visible and electrophoretic light scattering additive parameters, system stability and the effect of the ligands on the protein conformation at varying pH values have been defined. Calculated quenching constants are in the order of 2 × 10⁴ M^-1 and the type of interaction depends on the reaction medium. Under physiological conditions binding constant is 1.329 × 10⁶ M^-1 whereas in an environment similar to that of cancer cells the constant is significantly lower, K_a = 6.060 × 10⁴ M^-1. N values are approximate to 1 in all cases. Moreover, some changes are observed in the α-helical structure of the protein after interaction with the drugs and the presence of cyclophosphamide slightly stabilizes the protein secondary structure. All collected data proves the effect of cyclophosphamide on the interaction between imatinib mesylate and human holo-transferrin. It is of great clinical interest due to anticancer, multidrug therapies including both imatinib mesylate and cyclophosphamide.