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Published on: August 13, 2021
Thiourea-modified poly(l-lysine) carriers with multiple hydrogen bonding interactions for safe and robust mRNA
Jiangai Long1, Sheng Ma2, Renming Wan1
1State Key Laboratory of Polymer Science and Technology, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun 130022, China; School of Applied Chemistry and Engineering, University of Science and Technology of China, Hefei 230026, China.
Abstract:
Cationic polymers have garnered considerable attention as mRNA carriers due to their structural tunability and chemical diversity. Nevertheless, their broader application remains limited by poor stability under physiological conditions and cytotoxicity associated with excessive charge density. Here, we report thiourea-modified poly(l-lysine) (PLL-T) carriers constructed by combinatorial side-chain modification with ionizable and hydrophobic isothiocyanates. The resulting thiourea linkage introduced two hydrogen bonding donor sites, enabling stronger hydrogen bonding with mRNA. Such hydrogen bonding could synergize with electrostatic and hydrophobic interactions, forming multiple cooperative interactions with mRNA. This design enhanced mRNA encapsulation, colloidal stability, and high transfection efficiency without increasing cytotoxicity. The optimized PLL-1A1B(T) variant achieved over 10,000-fold higher transfection efficiency than the parent polymer PLL, and outperformed the commercial gold-standard Lipofectamine MessengerMAX in both HEK293T and RAW264.7 cells. Compared with its amide-linked analogue PLL-1A1B(A), in which the amide linkage provided only a single hydrogen bonding donor site, PLL-1A1B(T) exhibited superior mRNA encapsulation (91 % vs. 42 %), higher storage stability, and over 50-fold increase in intramuscular transfection. Unlike PLL-1A1B(A), which induced high levels of inflammation, PLL-1A1B(T) exhibited minimal inflammation in vivo. Importantly, PLL-1A1B(T) enabled efficient delivery of mRNA cancer vaccines and elicited potent CD8+ T cell mediated antitumor immunity. A single administration of PLL-1A1B(T)@E6/E7 mRNA vaccine achieved 77 % tumor suppression rate (TSR) in the TC-1 tumor model. Moreover, in combination with anti PD-1, PLL-1A1B(T)@OVA mRNA vaccine achieved 90 % TSR in the B16-OVA tumor model. This study demonstrates hydrogen bonding-driven carriers engineering as a versatile strategy to tune nucleic acid-carrier interactions, offering a potent and safe platform for mRNA therapeutics.
