Development of a Cationic Polymeric Micellar Structure with Endosomal Escape Capability Enables Enhanced Intramuscular Transfection of MRNA-LNPs

Overview

Journal Vaccines (Basel)

Date 2025 Jan 24

PMID 39852804

Authors

Siyuan Deng

Han Shao

Hongtao Shang

Lingjin Pang

Xiaomeng Chen

Jingyi Cao

Yi Wang

Zhao Zhao

Affiliations

Soon will be listed here.

Abstract

The endosomal escape of lipid nanoparticles (LNPs) is crucial for efficient mRNA-based therapeutics. Here, we present a cationic polymeric micelle (cPM) as a safe and potent co-delivery system with enhanced endosomal escape capabilities. We synthesized a cationic and ampholytic di-block copolymer, poly (poly (ethylene glycol) methacrylate--hexyl methacrylate)--poly(butyl methacrylate--dimethylaminoethyl methacrylate--propyl acrylate) (p(PEGMA--HMA)--p(BMA--DMAEMA--PAA)), via reversible addition-fragmentation chain transfer polymerization. The cPMs were then formulated using the synthesized polymer by the dispersion-diffusion method and characterized by dynamic light scattering (DLS) and cryo-transmission electron microscopy (CryoTEM). The membrane-destabilization activity of the cPMs was evaluated by a hemolysis assay. We performed an in vivo functional assay of firefly luciferase (Fluc) mRNA using two of the most commonly studied LNPs, SM102 LNP and Dlin-MC3-DMA LNPs. With a particle size of 61.31 ± 0.68 nm and a zeta potential of 37.76 ± 2.18 mV, the cPMs exhibited a 2-3 times higher firefly luciferase signal at the injection site compared to the control groups without cPMs following intramuscular injection in mice, indicating the high potential of cPMs to enhance the endosomal escape efficiency of mRNA-LNPs. The developed cPM, with enhanced endosomal escape capabilities, presents a promising strategy to improve the expression efficiency of delivered mRNAs. This approach offers a novel alternative strategy with no modifications to the inherent properties of mRNA-LNPs, preventing any unforeseeable changes in formulation characteristics. Consequently, this polymer-based nanomaterial holds immense potential for clinical applications in mRNA-based vaccines.

References

Convertine A, Benoit D, Duvall C, Hoffman A, Stayton P . Development of a novel endosomolytic diblock copolymer for siRNA delivery. J Control Release. 2008; 133(3):221-9. PMC: 3110267. DOI: 10.1016/j.jconrel.2008.10.004. View

Hald Albertsen C, Kulkarni J, Witzigmann D, Lind M, Petersson K, Simonsen J . The role of lipid components in lipid nanoparticles for vaccines and gene therapy. Adv Drug Deliv Rev. 2022; 188:114416. PMC: 9250827. DOI: 10.1016/j.addr.2022.114416. View

Gilleron J, Querbes W, Zeigerer A, Borodovsky A, Marsico G, Schubert U . Image-based analysis of lipid nanoparticle-mediated siRNA delivery, intracellular trafficking and endosomal escape. Nat Biotechnol. 2013; 31(7):638-46. DOI: 10.1038/nbt.2612. View

Obst K, Yealland G, Balzus B, Miceli E, Dimde M, Weise C . Protein Corona Formation on Colloidal Polymeric Nanoparticles and Polymeric Nanogels: Impact on Cellular Uptake, Toxicity, Immunogenicity, and Drug Release Properties. Biomacromolecules. 2017; 18(6):1762-1771. DOI: 10.1021/acs.biomac.7b00158. View

Baden L, El Sahly H, Essink B, Kotloff K, Frey S, Novak R . Efficacy and Safety of the mRNA-1273 SARS-CoV-2 Vaccine. N Engl J Med. 2020; 384(5):403-416. PMC: 7787219. DOI: 10.1056/NEJMoa2035389. View

Rybak S, Murphy R . Primary cell cultures from murine kidney and heart differ in endosomal pH. J Cell Physiol. 1998; 176(1):216-22. DOI: 10.1002/(SICI)1097-4652(199807)176:1<216::AID-JCP23>3.0.CO;2-3. View

Nelson C, Kintzing J, Hanna A, Shannon J, Gupta M, Duvall C . Balancing cationic and hydrophobic content of PEGylated siRNA polyplexes enhances endosome escape, stability, blood circulation time, and bioactivity in vivo. ACS Nano. 2013; 7(10):8870-80. PMC: 3857137. DOI: 10.1021/nn403325f. View

He C, Hu Y, Yin L, Tang C, Yin C . Effects of particle size and surface charge on cellular uptake and biodistribution of polymeric nanoparticles. Biomaterials. 2010; 31(13):3657-66. DOI: 10.1016/j.biomaterials.2010.01.065. View

Li Y, Cheng Q, Jiang Q, Huang Y, Liu H, Zhao Y . Enhanced endosomal/lysosomal escape by distearoyl phosphoethanolamine-polycarboxybetaine lipid for systemic delivery of siRNA. J Control Release. 2013; 176:104-14. DOI: 10.1016/j.jconrel.2013.12.007. View

10.

Stewart M, Lorenz A, Dahlman J, Sahay G . Challenges in carrier-mediated intracellular delivery: moving beyond endosomal barriers. Wiley Interdiscip Rev Nanomed Nanobiotechnol. 2015; 8(3):465-78. DOI: 10.1002/wnan.1377. View

11.

Kim H, Ogura S, Otabe T, Kamegawa R, Sato M, Kataoka K . Fine-Tuning of Hydrophobicity in Amphiphilic Polyaspartamide Derivatives for Rapid and Transient Expression of Messenger RNA Directed Toward Genome Engineering in Brain. ACS Cent Sci. 2019; 5(11):1866-1875. PMC: 6891845. DOI: 10.1021/acscentsci.9b00843. View

12.

Herrera M, Kim J, Eygeris Y, Jozic A, Sahay G . Illuminating endosomal escape of polymorphic lipid nanoparticles that boost mRNA delivery. Biomater Sci. 2021; 9(12):4289-4300. PMC: 8769212. DOI: 10.1039/d0bm01947j. View

13.

Jones R, Cheung C, Black F, Zia J, Stayton P, Hoffman A . Poly(2-alkylacrylic acid) polymers deliver molecules to the cytosol by pH-sensitive disruption of endosomal vesicles. Biochem J. 2003; 372(Pt 1):65-75. PMC: 1223370. DOI: 10.1042/BJ20021945. View

14.

Alexis F, Pridgen E, Molnar L, Farokhzad O . Factors affecting the clearance and biodistribution of polymeric nanoparticles. Mol Pharm. 2008; 5(4):505-15. PMC: 2663893. DOI: 10.1021/mp800051m. View

15.

Lee Y, Jeong M, Park J, Jung H, Lee H . Immunogenicity of lipid nanoparticles and its impact on the efficacy of mRNA vaccines and therapeutics. Exp Mol Med. 2023; 55(10):2085-2096. PMC: 10618257. DOI: 10.1038/s12276-023-01086-x. View

16.

Wittrup A, Ai A, Liu X, Hamar P, Trifonova R, Charisse K . Visualizing lipid-formulated siRNA release from endosomes and target gene knockdown. Nat Biotechnol. 2015; 33(8):870-6. PMC: 4663660. DOI: 10.1038/nbt.3298. View

17.

Wang Z, Zhang J, Wang Y, Zhou J, Jiao X, Han M . Overcoming Endosomal Escape Barriers in Gene Drug Delivery Using De Novo Designed pH-Responsive Peptides. ACS Nano. 2024; 18(14):10324-10340. DOI: 10.1021/acsnano.4c02400. View

18.

Prieve M, Harvie P, Monahan S, Roy D, Li A, Blevins T . Targeted mRNA Therapy for Ornithine Transcarbamylase Deficiency. Mol Ther. 2018; 26(3):801-813. PMC: 5910669. DOI: 10.1016/j.ymthe.2017.12.024. View

19.

Betker J, Tilden S, Anchordoquy T . Escaping to silence using an endosome-disrupting polymer. Mol Ther. 2021; 29(10):2893-2894. PMC: 8531147. DOI: 10.1016/j.ymthe.2021.09.006. View

20.

Homma K, Miura Y, Kobayashi M, Chintrakulchai W, Toyoda M, Ogi K . Fine tuning of the net charge alternation of polyzwitterion surfaced lipid nanoparticles to enhance cellular uptake and membrane fusion potential. Sci Technol Adv Mater. 2024; 25(1):2338785. PMC: 11028023. DOI: 10.1080/14686996.2024.2338785. View