A multimodal approach integrating spectroscopy, deep learning guided molecular docking, and molecular dynamics...

Binding affinity is a critical parameter that can influence the state of the drug in vivo and help to define the formulation strategy. The current study implements a multimodal approach to analyse the binding affinity between human serum albumin (HSA) and pioglitazone. Ultraviolet (UV) absorbance and fluorescence spectrometry analyses were performed on different combinations of HSA and pioglitazone complexes, and the absorbance and fluorescence intensities were mapped to calculate the binding constant. DynamicBind, a distinct deep-learning artificial intelligence tool, was implemented to perform in silico docking studies using a non-conventional approach. Furthermore, molecular dynamics simulation was also performed to generate root mean square deviation, radius of gyration, and root mean square fluctuation values, followed by principal component analysis, probability distribution function, and free energy landscape analysis. The simulation output was analysed to interpret the binding affinity and associated conformation of the protein-active pharmaceutical ingredient (API) complex. The binding constant calculated through UV analysis was 1.1 × 10⁴ M^-1. Fluorescence spectroscopic analysis derived a value of 1.7 × 10⁵ M^-1. At the same time, DynamicBind predicted the cLDDT score for the top predicted model to be 0.634, and a binding affinity value of greater than 5, indicating a relatively moderate binding between pioglitazone and HSA. The results from molecular dynamics simulations further complemented our earlier observations, indicating non-covalent binding interactions and a stable protein-API complex, which is desirable for developing a formulation using HSA as a carrier polymer. This orthogonal approach also provided critical information on the fate of the API and possible considerations that needed to be made during the design of the formulation process, highlighting the need for similar approaches that could provide multifaceted advantages and help in optimising R&D costs and timelines.