Characterization of Locally Excited and Charge‐Transfer States of the Anticancer Drug Lapatinib by Ultrafast Spectroscopy and Computational Studies

Abstract

Lapatinib (LAP) is an anticancer drug, which is metabolized to the N-a nd O-dealkylated products (N-LAP and O-LAP,r espectively). In view of the photosensitizing potential of relatedd rugs, ac omplete experimentala nd theoretical study has been performedo nL AP, N-LAP and O-LAP, both in solution and upon complexationw ith human serum albumin (HSA). In organic solvents, coplanar locally excited (LE) emissive states are generated;t hey rapidly evolve towards twisted intramolecularc harge-transfer (ICT) states. By contrast, within HSA only LE states are detected. Accordingly,f emtosecond transient absorption reveals av ery fast switching (ca. 2ps) from LE (l max = 550 nm) to ICT states (l max = 480 nm) in solution, whereas within HSA the LE species become stabilized and live much longer(up to the ns scale). Interestingly,m olecular dynamics simulation studies confirm that the coplanar orientation is preferred for LAP (or to al esser extent N-LAP) within HSA, explaining the experimental results.

Protein Binding of Lapatinib and Its N- and O-Dealkylated Metabolites Interrogated by Fluorescence, Ultrafast Spectroscopy and Molecular Dynamics Simulations

“…For this purpose, two types of transport proteins were selected: serum albumins and α 1 -acid glycoproteins from human (HSA and HAG) and bovine (BSA and BAG) origin, respectively. As previously observed, LAP and N-LAP formed aggregates in PBS solution (Wilson et al, 2015;Vayá et al, 2020); however, upon interaction with proteins they were completely solubilized. The UV absorption spectra of LAP, N-LAP and O-LAP bound to HSA, BSA, HAG and BAG did not reveal significant differences (Supplementary Figure S1).…”

“…However, in the presence of protein, which provides a more constrained environment, a clear enhancement of LAP fluorescence in addition to quenching of the protein emission (λ max ∼ 340 nm) was noticed (see Supplementary Figure S2). This effect was previously detected for the drug complexed to HSA, and was associated to singlet-singlet energy transfer (SSET) from HSA to LAP (Vayá et al, 2020). On the other hand, the spectra within each protein were unstructured and displayed their maxima at shorter wavelengths (ca.…”

“…Emission of the protein-bound LAP (or metabolites) was first investigated at λ exc 295 nm, where both the protein and the drug absorb light. In PBS, the fluorescence of LAP was weak and unstructured (λ max ∼ 475 nm) due to aggregation but most importantly due to emission from intramolecular charge transfer (ICT) states, which are favored in a twisted orientation between the furan and quinazoline rings due to the freedom in the degrees of movement of the drug in solution (Vayá et al, 2020). However, in the presence of protein, which provides a more constrained environment, a clear enhancement of LAP fluorescence in addition to quenching of the protein emission (λ max ∼ 340 nm) was noticed (see Supplementary Figure S2).…”

“…425 nm detected for LAP and N-LAP within HSA (Figures 2A,B), the emissions of the other ligand@protein complexes were much weaker and unstructured, and the maxima were found at longer wavelengths (∼450 nm). The behavior within HSA was associated to the more constrained environment provided by the protein but also to the frozen coplanar orientation adopted by the furan and quinazoline rings of LAP or N-LAP, which favors emission from LE (locally excited) states (Vayá et al, 2020). On the contrary, binding to BSA, HAG or BAG might disfavor emission from LE states due to a certain degree of freedom for twisting between both rings, resulting in a diminished and red-shifted emission of the encapsulated LAP or N-LAP.…”

“…In this framework, preliminary findings on LAP@SA complexation point to a moderate binding of the drug in the so-called site III (subdomain IB) of the protein (Shen et al, 2015;Wilson et al, 2015;Kabir et al, 2016). In addition, a recent photophysical study on the interactions between LAP, N-LAP or O-LAP and HSA (Vayá et al, 2020) has shown that within the constrained environment provided by the HSA cavities emission occurs from long-lived locally excited (LE) states, whereas in the bulk solution shorter-lived (70-90 ps) intramolecular charge transfer (ICT) states predominate. This is related to the relative conformational orientation of the furan vs. the quinazoline ring in the different media.…”