Locating the binding sites of antitumor drug tamoxifen and its metabolites with DNA

• The binding sites of tamoxifen and its metabolites are located with DNA duplex.
• Drug–DNA bindings are via both hydrophobic and hydrophilic contacts.
• 4-Hydroxytamoxifen forms more stable complexes than tamoxifen and endoxifen.
• Drug–DNA interaction induces biopolymer aggregation.

We located the binding sites of antitumor drugs tamoxifen, 4-hydroxytamoxifen and endoxifen with calf-thymus DNA. FTIR, CD, UV–vis and fluorescence spectroscopic methods as well as molecular modeling were used to characterize the drug binding sites, binding constant and the effect of drug binding on DNA stability and conformation. Structural analysis showed that tamoxifen and its metabolites bind DNA via hydrophobic and hydrophilic interactions with overall binding constants of Ktam-DNA = 3.5 (±0.2) × 104 M−1, K4-hydroxytam-DNA = 3.3 (±0.4) × 104 M−1 and Kendox-DNA = 2.8 (±0.8) × 104 M−1. The number of binding sites occupied by drug is 1 (tamoxifen), 0.8 (4-hydroxitamoxifen) and 1.2 (endoxifen). Docking showed the participation of several nucleobases in drug–DNA complexes with the free binding energy of −3.85 (tamoxifen), −4.18 (4-hydroxtamoxifen) and −3.74 kcal/mol (endoxifen). The order of binding is 4-hydroxy-tamoxen > tamoxifen > endoxifen. Drug binding did not alter DNA conformation from B-family structure, while major biopolymer aggregation occurred at high drug concentrations. The drug binding mode is correlated with the mechanism of action of antitumor activity of tamoxifen and its metabolites.

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