The interaction of sorafenib and regorafenib with membranes is modulated by their lipid composition

Abstract

Sorafenib and regorafenib are small-molecule kinase inhibitors approved for the treatment of locally recurrent or metastatic, progressive, differentiated thyroid carcinoma, renal cell carcinoma, and hepatocellular carcinoma (sorafenib) and of colorectal cancer (regorafenib). As of now, the mechanisms, which are responsible for their antitumor activities, are not completely understood. Given the lipophilic nature of the molecules, it can be hypothesized that the pharmacological impact is mediated by the interaction with cellular membranes as it is true for many pharmacologically active molecules. However, an interaction of sorafenib or regorafenib with lipid membranes has not yet been investigated in detail. Here, we characterized the interaction of both drugs with lipid membranes by applying a variety of biophysical approaches including nuclear magnetic resonance, electron spin resonance, and fluorescence spectroscopy. We found that sorafenib and regorafenib bind to lipid membranes by inserting into the lipid-water interface of the bilayer. This membrane embedding causes a disturbance of bilayer structure leading to an increased permeability of the membrane for polar molecules. One approach shows that the extent of the effects depends on the membrane lipid composition underlining a particular role of phosphatidylcholine and cholesterol. Our data for the first time characterize the impact of sorafenib and regorafenib on the lipid membrane structure and dynamics, which may contribute to a better understanding of their effectiveness in the treatment of malignancies as well as of their side effects.

Impact of Selected Small-Molecule Kinase Inhibitors on Lipid Membranes

“…The plot of these cross-relaxation rates over the molecular groups of POPC along the membrane normal can be interpreted as a distribution function of the inhibitor protons across the normal membrane. Previous works performed similar approaches for many other small molecules including kinase inhibitors [ 2 , 4 , 14 , 15 ]. From these data, conclusions about the membrane position and orientation of the respective molecule can be drawn.…”

“…Two assays were applied in order to measure the impact of drugs on membrane structure, determining the influence of kinase inhibitors on the permeation of anions, i.e., dithionite and ascorbate, across membranes. The assays employ the property of dithionite and ascorbate to rapidly reduce and irreversibly quench the fluorescence of the NBD group and the ESR signal intensity of spin-labeled lipids, respectively [ 15 , 55 , 56 , 57 ]. When the labeled lipids are incorporated into unilamellar lipid vesicles, the anions quench the signal intensities of the respective labeled lipids in the outer membrane leaflet, which is observed by a rapid initial decrease of signal intensity.…”

Membrane Interaction of Ibuprofen with Cholesterol-Containing Lipid Membranes

“…Subsequently, fluorescence decreases slowly caused by a slow permeation of dithionite, reacting with the labeled lipids in the inner membrane leaflet. The assay was performed as described [ 26 , 27 ]. From the reduction kinetics (see Supplementary Figure S4 ), the rate constants for the slow decrease (k p ), i.e., permeation of dithionite were determined.…”

“…The measurement of ascorbate permeation is similar to the dithionite assay, but here, the reduction of spin-labeled lipids (2 mol%) in liposomes is measured upon addition of ascorbate. The assay was performed as described in [ 27 , 28 ]. From the reduction kinetics, the rate constant k p of the slower component was determined as a parameter for ascorbate permeation.…”

“…LUVs were prepared in HBS additionally containing 50 mM 6-carboxyfluorescein [ 27 ]. At this large concentration, CF shows a low fluorescence due to self-quenching.…”