Binding of antitumor tamoxifen and its metabolites 4-hydroxytamoxifen and endoxifen to human serum albumin

Tamoxifen is extensively metabolized, and several metabolites have been detected in human serum. The aim of this study was to examine the interaction of human serum albumin (HSA) with tamoxifen and its metabolites 4-hydroxytamoxifen and endoxifen at physiological conditions, using constant protein concentration and various drug contents. FTIR, UV–Visible, CD and fluorescence spectroscopic methods as well as molecular modeling were used to analyse drug binding mode, the binding constant and the effects of drug complexation on HSA stability and conformation. Structural analysis showed that tamoxifen and its metabolites bound HSA via both hydrophobic and hydrophilic interactions with overall binding constants of Ktam = 1.8 (±0.2) × 104 M−1, K4-hydroxytam = 1.8 (±0.4) × 104 M−1 and Kendox = 2.0 (±0.5) × 104 M−1. The number of bound drugs per protein is 1.2 (tamoxifen), 1.7 (4-hydroxitamoxifen) and 1.0 (endoxifen). Structural modeling showed the participation of several amino acid residues in drug–HSA complexation, with extended H-bonding network. HSA conformation was altered by tamoxifen and its metabolites with a major reduction of α-helix and an increase in β-sheet, random coil and turn structures, indicating a partial protein unfolding. Our results suggest that serum albumins can act as carrier proteins for tamoxifen and its metabolites in delivering them to target tissues.

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► The binding sites of anticancer tamoxifen and its metabolites are located on HSA.
► Endoxifen formed stronger complexes than tamoxifen and 4-hydroxytamoxifen.
► The complexation of tamoxifen and its metabolites induced protein unfolding.
► HSA can be used to transport tamoxifen and its metabolites in vitro.