Design and development of drug delivery nanocarriers based on liquid-liquid phase separation, improved stability,...

Liquid-liquid phase separation (LLPS) of nuclear pore complex (NPC) with nuclear transport proteins (NTPs) via intrinsically disordered regions (IDRs) plays a crucial role in the nucleocytoplasmic transport. The development of efficient targeted delivery systems based on LLPS has attracted widespread attention. Here, we developed nanocarriers of casein peptides, a natural intrinsically disordered proteins (IDPs), modified with fatty acids of different alkyl chains (C10-C18) and decorated by shellac for highly effective drug delivery and cancer therapy. The curcumin (Cur)-loading nanocarriers (CSLNCs) showed excellent stability and dispersity in the natural environment over 30 days, with Cur encapsulation efficiency and loading capacity of ~90 % and ~57 %. Electron microscope (EM) indicated an aggregated homogeneous elliptical shape of CSLNCs(C10) and the morphology of CSLNCs(C18) transited to a distributed cubic shape. CSLNCs(C10, C12, C14 and C18) exhibited cytotoxicity against human lung adenocarcinoma NCI-H1975 cells with an IC₅₀ value of 17.5 μM, 17.3 μM, 10.2 μM and 19.3 μM after 24 h of incubation, respectively. CSLNCs were also found to inhibit the cell wound healing with a migration rate of 12.72 %, 10.93 %, 4.28 % and 13.62 %, respectively. CSLNCs especially increased the percentage of late apoptotic cells. As indications of confocal microscopy, the fluorescence intensities of NCI-H1975 cells were enhanced with a cytosolic distribution and noticeably florescence in the nucleus after 0.5 h of incubation CSLNCs. CSLNCs treated cells adopted a rounded morphology with a dramatic reduction in fluorescence intensity after 1 h of incubation. Among CSLNCs, CSLNCs(C14) improved considerably the cytotoxicity activity and intercellular localization in the nucleus. The cell-penetration ability was also confirmed by the binding of CSLNCs in a model bicelles membrane system composed of DMPC and DHPC investigated by ¹H NMR. It was proposed that CSLNCs with cell-penetrating and nuclear targeting performance may regulate the LLPS of nuclear pore complex and thus improved its nuclear penetration and cytotoxic activity.