Interaction of an anticancer drug, gefitinib with human serum albumin: insights from fluorescence spectroscopy and computational modeling analysis

Abstract

Binding of gefitinib (GEF), a promising anticancer drug to human serum albumin (HSA), the major transport protein in blood circulation was investigated using fluorescence, UV-vis absorption and circular dichroism (CD) spectroscopy as well as computational modeling. Fluorescence quenching of HSA upon GEF addition was found to be a static quenching process, as revealed from the decreasing trend of the Stern–Volmer quenching constant with increasing temperature as well as UV-vis absorption spectral results. Fluorescence quenching titration results demonstrated moderate binding affinity with the binding constant, K_a value as 1.70 × 10⁴ M⁻¹ between GEF and HSA at 15 °C. Thermodynamic data (ΔH = −7.74 kJ mol⁻¹ and ΔS = +54.06 J mol⁻¹ K⁻¹) suggested participation of both hydrophobic interactions and hydrogen bonds in stabilizing the GEF–HSA complex, which was further supported by computational modeling results. The far-UV and the near-UV CD spectra showed secondary and tertiary structural changes in HSA, whereas three-dimensional fluorescence spectral results indicated microenvironmental perturbations around protein fluorophores upon GEF binding. Binding of GEF to HSA offered significant protection to the protein against thermal destabilization. Competitive site-marker displacement results along with computational modeling analysis suggested a preferred location of the GEF binding site as site III, located in subdomain IB of HSA. Some common metal ions have been found to interfere with GEF–HSA interaction.