H7T4-4 is an ionizable lipid designed for mRNA delivery via lipid nanoparticles (LNPs). It features a cyclic amine headgroup (derived from cyclen tetrahydrochloride) and four C14 hydrophobic alkyl tails, synthesized through a Michael addition reaction between cyclen and 1,2-epoxytetradecane. With a high transition temperature (Tm = 58.6°C) due to strong intermolecular interactions from its cyclic headgroup and multi-tail structure, H7T4-4 alone forms rigid aggregates incompatible with mRNA encapsulation. However, when blended with low-Tm helper lipids (e.g., DOPE, Tm = -16°C), the system’s overall Tm decreases, enabling stable LNP formation. Optimized formulations (20% H7T4-4, 41% DOPE, 38% cholesterol, 1% DMG-PEG) exhibit efficient mRNA encapsulation (>90%) and transfection. Structural analyses (SAXS, cryo-TEM) confirm monodisperse LNPs with lamellar/hexagonal phases. In vivo, H7T4-4 LNPs show tumor-targeted and intranasal mRNA delivery with reduced off-target accumulation compared to SM-102-based LNPs. This rational design highlights Tm-guided helper lipid selection to overcome rigidity challenges in ionizable lipids.
