Protein-Containing Lipid Bilayers Intercalated with Size-Matched Mesoporous Silica Thin Films.

Proteins are key components in a multitude of biological processes, of which the functions carried out by transmembrane (membrane-spanning) proteins are especially demanding for investigations. This is because this class of protein needs to be incorporated into a lipid bilayer representing its native environment, and in addition, many experimental conditions also require a solid support for stabilization and analytical purposes. The solid support substrate may, however, limit the protein functionality due to protein-material interactions and a lack of physical space. We have in this work tailored the pore size and pore ordering of a mesoporous silica thin film to match the native cell-membrane arrangement of the transmembrane protein human aquaporin 4 (hAQP4). Using neutron reflectivity (NR), we provide evidence of how substrate pores host the bulky water-soluble domain of hAQP4, which is shown to extend 7.2 nm into the pores of the substrate. Complementary surface analytical tools, including quartz crystal microbalance with dissipation monitoring (QCM-D) and fluorescence microscopy, revealed successful protein-containing supported lipid bilayer (pSLB) formation on mesoporous silica substrates, whereas pSLB formation was hampered on nonporous silica. Additionally, electron microscopy (TEM and SEM), light scattering (DLS and stopped-flow), and small-angle X-ray scattering (SAXS) were employed to provide a comprehensive characterization of this novel hybrid organic-inorganic interface, the tailoring of which is likely to be generally applicable to improve the function and stability of a broad range of membrane proteins containing water-soluble domains.

Amphotericin B and monoacyl-phosphatidylcholine form a stable amorphous complex.

Amphotericin B (AmB) is a "life-saving" medicine for the treatment of invasive fungal infections and visceral leishmaniasis. To date, all marketed AmB formulations require parenteral administration, which causes high rates of acute infusion-related side effects and dose-dependent nephrotoxicity. The development of an oral AmB formulation will entail numerous advantages including increased patient compliance, eliminated infusion-related toxicities and reduced nephrotoxicity. Unfortunately, the gastrointestinal absorption of AmB is negligible due to its extremely low solubility in both aqueous and lipid solvents, and its poor gastrointestinal permeability. Drug-phospholipid complexation is an emerging strategy for oral delivery of poorly soluble drugs. In this study, monoacyl-phosphatidylcholine (MAPC) was complexed with AmB forming an AmB-MAPC complex (APC), to enhance the dissolution rate and aqueous solubility of AmB, in order to enable oral delivery of AmB. X-ray powder diffraction demonstrated that AmB was transformed to its amorphous form following complexation with MAPC, i.e. in the APC. Fourier-transform infrared spectroscopy suggested molecular interactions between AmB and MAPC. Dynamic light scattering indicated formation of colloidal structures after aqueous dispersion of APC; Cryogenic transmission electron microscopy showed that APC formed small round, "rod-like" and "worm-like" micellar structures and Small-angle neutron scattering provided three-dimensional micellar structures formed by APC upon aqueous dispersion, which indicated that AmB was inserted into the micellar mono-layer membrane formed by MAPC. Additionally, APC showed an increased dissolution rate and a higher amount of AmB solubilized in fasted state simulated intestinal fluid, compared to AmB/MAPC physical mixtures and crystalline AmB. In conclusion, an APC exhibiting amorphous properties was developed, the APC showed improved dissolution rate and increased apparent aqueous solubility compared to AmB, indicating that the application of APC could be a promising strategy to enable the oral delivery of AmB.