Profiling the Interaction between Human Serum Albumin and Clinically Relevant HIV Reverse Transcriptase Inhibitors.

Tenofovir (TFV) is the active form of the prodrugs tenofovir disoproxil fumarate (TDF) and tenofovir alafenamide (TAF), both clinically prescribed as HIV reverse transcriptase inhibitors. The biophysical interactions between these compounds and human serum albumin (HSA), the primary carrier of exogenous compounds in the human bloodstream, have not yet been thoroughly characterized. Thus, the present study reports the interaction profile between HSA and TFV, TDF, and TAF via UV-Vis, steady-state, and time-resolved fluorescence techniques combined with isothermal titration calorimetry (ITC) and in silico calculations. A spontaneous interaction in the ground state, which does not perturb the microenvironment close to the Trp-214 residue, is classified as weak. In the case of HSA/TFV and HSA/TDF, the binding is both enthalpically and entropically driven, while for HSA/TAF, the binding is only entropically dominated. The binding constant (Ka) and thermodynamic parameters obtained via ITC assays agree with those obtained using steady-state fluorescence quenching measurements, reinforcing the reliability of the data. The small internal cavity known as site I is probably the main binding pocket for TFV due to the low steric volume of the drug. In contrast, most external sites (II and III) can better accommodate TAF due to the high steric volume of this prodrug. The cross-docking approach corroborated experimental drug-displacement assays, indicating that the binding affinity of TFV and TAF might be impacted by the presence of different compounds bound to albumin. Overall, the weak binding capacity of albumin to TFV, TDF, and TAF is one of the main factors for the low residence time of these antiretrovirals in the human bloodstream; however, positive cooperativity for TAF and TDF was detected in the presence of some drugs, which might improve their residence time (pharmacokinetic profile).

Interaction between a water-soluble anionic porphyrin and human serum albumin unexpectedly stimulates the aggregation of the photosensitizer at the surface of the albumin.

The 5,10,15,20-tetrakis(2,6-difluoro-3-sulfophenyl)porphyrin (TDFPPS4) was reported as a potential photosensitizer for photodynamic therapy. The capacity of the photosensitizers to be carried in the human bloodstream is predominantly determined by its extension of binding, binding location, and binding mechanism to human serum albumin (HSA), influencing its biodistribution and ultimately its photodynamic therapy efficacy in vivo. Thus, the present work reports a biophysical characterization on the interaction between the anionic porphyrin TDFPPS4 and HSA by UV-visible absorption, circular dichroism, steady-state, time-resolved, and synchronous fluorescence techniques under physiological conditions, combined with molecular docking calculations and molecular dynamics simulations. The interaction HSA:TDFPPS4 is spontaneous (ΔG° < 0), strong, and enthalpically driven (ΔH° = -70.1 ± 3.3 kJ mol-1) into subdomain IIA (site I). Curiously, despite the porphyrin binding into an internal pocket, about 50 % of TDFPPS4 structure is still accessible to the solvent, making aggregation in the bloodstream possible. In silico calculations were reinforced by spectroscopic data indicating porphyrin aggregation between bound and unbound porphyrins. This results in an adverse scenario for anionic porphyrins to achieve their therapeutical potential as photosensitizers and control of effective dosages. Finally, a trend of anionic porphyrins to have a combination of quenching mechanisms (static and dynamic) was noticed.