Interaction of 5-fluoro-5′-deoxyuridine with human serum albumin under physiological and non-physiological condition: A biophysical investigation

Uridine analogs 5′dFUrd (a cytotoxic metabolite of a prodrug capecitabine that enzymatically converted into 5′dFUrd) commonly used in the treatment of advanced human cancers, especially gastrointestinal tract, ovary, colorectal, breast cancers etc. Drugs/metabolites are transported in the blood by transporter proteins like human serum albumin (HSA). Here we investigate the interaction of 5′dFUrd to HSA by spectroscopic and calorimetric techniques at physiological (pH 7.4) and non-physiological (pH 9.0) conditions. The binding constant (Kb), enthalpy change (ΔH°), entropy change (ΔS°) and Gibbs free energy change (ΔG°) were also calculated under both conditions. The secondary structure of HSA showed greater alteration in helicity at physiological pH. ITC measurement reveals that HSA have high binding affinity at physiological pH as compares to non-physiological conditions. The thermostability of HSA alone as well of the HSA–drug complex was found to be higher at physiological pH. The binding study was also explored through molecular docking studies which revealed that 5′dFUrd was bound to subdomain IIA of Sudlowʹs site I through multiple mode of interaction. These results suggest that 5′dFUrd have high binding affinity at physiological condition or “N” isoform so lower drug concentrations are required in compare to non-physiological or “B” isoform of HSA to completely occupied the binding site of the protein.