Interactions of Cubosomes with Cell Membrane Lipids: Revealed through Cubic Membrane Property Analysis

Leveraged for their large drug payloads and fusogenic properties, cubosomes are promising platforms for therapeutic applications. However, research regarding the properties of the cubic membranes and their interactions with cell membranes is still lacking. The study of these properties is an important factor to consider in the design process of cubosomes due to their influence on the cubosome structure and capability for drug release. To address this research gap, we demonstrate the effectiveness of the utilization of membrane-bound fluorescence probes to examine the properties of cubosomes and their interaction behavior. First, we demonstrated the cubosomal membrane changes with increasing concentration of DOPC using steady-state and time-resolved fluorescence of Laurdan and DPH probes. The steady-state generalized polarization of Laurdan (GP₃₄₀) and anisotropy of DPH (r) showed that the well-hydrated and fluidic membranes transitioned into less hydrated and fluidic states, due to the transition to the lamellar phase. Finally, we verified the results in the first part by examining the interactions of MO cubosomes and DOPC liposomes using time-resolved small-angle X-ray scattering (TR-SAXS) and the changes to the GP₃₄₀ values of Laurdan probed cubosomes over time. The cubic-to-lamellar phase transition was confirmed from the TR-SAXS spectra. By using the GP₃₄₀ of the MO/DOPC compositions in the first part as a calibration curve, the kinetics of apparent lipid exchange (k_app) were obtained, and the apparent amount of DOPC in the cubosomes over the course of the interaction could be estimated. The approach developed in this work showcases the applications of fluorescence probes such as estimation of apparent lipid exchange kinetics that could be generalized to other types of carriers. In addition, the results obtained from this work provide a deeper understanding of changes in the cubic membrane and structure in relation to the interaction between the drug carrier and model cell membranes. Such insights may prove to be valuable for the design of cubosomes as therapeutic carriers.