Structural Insights on Ionizable Dlin-MC3-DMA Lipids in DOPC Layers by Combining Accurate Atomistic Force Fields, Molecular Dynamics Simulations and Neutron Reflectivity

Abstract

Ionizable lipids such as the promising Dlin-MC3-DMA (MC3) are essential for the successful design of lipid nanoparticles (LNPs) as drug delivery agents. Combining molecular dynamics simulations with experimental data such as neutron reflectivity experiments and other scattering techniques is essential to provide insights into the internal structure of LNPs, which is not fully understood to date. However, the accuracy of the simulations relies on the choice of force field parameters and high-quality experimental data is indispensable to verify the parametrization. For MC3, different parameterizations in combination with the CHARMM and the Slipids force field have recently emerged. Here, we complement the existing efforts by providing parameters for cationic and neutral MC3 compatible with the AMBER Lipid17 force field. Subsequently, we carefully assess the accuracy of the different force fields by providing a direct comparison to neutron reflectivity experiments of mixed lipid bilayers consisting of MC3 and DOPC at different pH. At low pH (cationic MC3) and at high pH (neutral MC3) the newly developed MC3 parameters in combination with AMBER Lipid17 for DOPC give good agreement with the experiments. Overall, the agreement is slightly better compared to the Park-Im parameters for MC3 in combination with the CHARMM36 force field for DOPC. The Ermilova-Swenson MC3 parameters in combination with the Slipids force field underestimate the bilayer thickness. While the distribution of cationic MC3 is very similar, the different force fields for neutral MC3 reveal distinct differences ranging from strong accumulation in the membrane center (current MC3/AMBER Lipid17 DOPC), over mild accumulation (Park-Im MC3/CHARMM36 DOPC) to surface accumulation (Ermilova-Swenson MC3/Slipids DOPC). These pronounced differences highlight the importance of accurate force field parameters and their experimental validation.

Combining molecular dynamics simulations and X-ray scattering techniques for the accurate treatment of protonation degree and packing of ionizable lipids in monolayers

“…S8), consistent observations can be made at α = 15.0 %, corresponding to pH 7.5. This behavior has been noticed earlier 43 and may provide the explanation for the small yet significant increase in A lip when increasing the pH from 5.0 to 7.5 (see above). Namely, in contrast to the naive expectation that a reduction in the density of charged lipids would decrease lateral repulsion and thus the lipids’ effective area requirement, the deep insertion of uncharged MC3 0 into the monolayer effectively increases the area requirement at high pH, where MC3 0 is more abundant.…”

“…The AMBER Lipid 17 force field 42 was used to describe the POPC lipids. For the cationic and neutral MC3 molecules we used our recently developed force fields 43 . These parameters have the advantage that they closely reproduce the structure of MC3 in lipid layers as judged by neutron reflectometry experiments.…”