Hyperbranched Polylysine-Based mRNA Delivery Carrier Optimization and Evaluation

doi:10.19894/j.issn.1000-0518.250219

Chinese Journal of Applied Chemistry ›› 2025, Vol. 42 ›› Issue (10): 1335-1348.DOI: 10.19894/j.issn.1000-0518.250219

• Full Papers • Previous Articles Next Articles

Hyperbranched Polylysine-Based mRNA Delivery Carrier Optimization and Evaluation

Yi-Bo QI¹^,², Kun-Cheng LYU¹^,², Yi-Wei WANG¹^,², Wan-Tong SONG¹^,²(), Sheng-Xiang JI¹^,²(), Xue-Si CHEN¹^,²()

^1.State Key Laboratory of Polymer Science and Technology，Changchun Institute of Applied Chemistry，Chinese Academy of Sciences，Changchun 130022，China
^2.School of Applied Chemistry and Engineering，University of Science and Technology of China，Hefei 230026，China

Received:2025-05-28 Accepted:2025-08-19 Published:2025-10-01 Online:2025-10-29
Contact: Wan-Tong SONG,Sheng-Xiang JI,Xue-Si CHEN
About author:xschen@ciac.ac.cn
sji@ciac.ac.cn
wtsong@ciac.ac.cn
Supported by:
the National Key Research and Development Program of China(2024YFA1212200);the National Natural Science Foundation of China(22222509);Jilin Provincial Key Laboratory of Biomedical Polymer Materials for International Cooperation(YDZJ202402077CXJD)

1. .pdf(199KB)

Abstract

Abstract:

Hyperbranched polylysine （HPL） is a synthetic polymer material that is simple to prepare and inexpensive， with excellent biocompatibility. However， its transfection efficiency is not good when used as an mRNA delivery carrier. In this study， a HPL material library was established through a simple and systematic chemical modification strategy. Through amino-epoxide ring-opening reactions， Michael addition reactions， and amino-phospholane ring-opening reactions， small molecules containing hydrophobic groups such as 1，2-epoxides， acrylates， and alkylated dioxyphosphoranes were introduced into the three-dimensional backbone of HPL， and mRNA delivery carriers with high transfection efficiency were screened. Experimental results showed that HPL modified with propylene oxide （E3）， HPL-E3， was the best material in the HPL-En series， with a 1.8-fold increase compared to unmodified HPL； HPL modified with 3-（perfluorobutyl）-1，2-epoxide （F9）， HPL-F9， was the best material in the HPL-Fx series， with a 272.4-fold increase compared to unmodified HPL； HPL modified with dodecyl acrylate （A12）， HPL-A12， was the best material in the HPL-Ay series， with a 3246-fold increase compared to unmodified HPL； and HPL modified with 2-octyloxy-2-oxy-1，3，2-dioxaphospholane （P8）， HPL-P8， was the best material in the HPL-Pm series， with a 75.0-fold increase compared to unmodified HPL. This minimalist one-step synthesis strategy can introduce hydrophobic chain segments of appropriate lengths to enhance cell uptake， and can also convert primary amines on HPL to secondary or tertiary amines， thereby enhancing their endosomal escape capability， providing a new method for the construction of mRNA delivery carriers.

Key words: Hyperbranched polylysine, mRNA delivery, Epoxy compound, Acrylate ester, Alkylated dioxyphosphorinane

CLC Number:

O633

Yi-Bo QI, Kun-Cheng LYU, Yi-Wei WANG, Wan-Tong SONG, Sheng-Xiang JI, Xue-Si CHEN. Hyperbranched Polylysine-Based mRNA Delivery Carrier Optimization and Evaluation[J]. Chinese Journal of Applied Chemistry, 2025, 42(10): 1335-1348.

Figures/Tables 10

Fig.1 1H NMR spectrum of HPL （D2O）

Fig.2 GPC chromatogram of HPL

Table 1 Reaction conditions for the synthesis of HPL derivatives

Molecular species	Reaction solvent	Equivalent	Temperature/℃	Reaction time/h
Non-fluorinated epoxides	1 mL DMSO	2	60	24
Fluorinated epoxides	1 mL DMSO	0.5	60	24
Acrylates	0.8 mL DMSO+0.2 mL DMF	2	60	24
Alkylated dioxyphosphoranes	1 mL DMSO	2	60	24

Fig.3 HPL-En transfection efficiency of Fluc mRNA in 293T cells （m（polymer）∶m（mRNA）=20∶1）

Fig.4 HPL-Fx transfection efficiency of Fluc mRNA in 293T cells （m（polymer）∶m（mRNA）=20∶1）

Fig.5 HPL-F9 transfection efficiency of Fluc mRNA in 293T cells at different polymer/mRNA mass ratios

Fig.6 HPL-Ay transfection efficiency of Fluc mRNA in 293T cells （m（polymer）∶m（mRNA）=20∶1）

Fig.7 HPL-A8， HPL-A12 and HPL-A14 transfection efficiency of Fluc mRNA in 293T cells at different polymer/mRNA mass ratios

Fig.8 HPL-Pm transfection efficiency of Fluc mRNA in 293T cells （m（polymer）∶m（mRNA）=20∶1）

Fig.9 HPL-P8 and HPL-P10 transfection efficiency of Fluc mRNA in 293T cells at different polymer/mRNA mass ratios

References 41

[1]	MIYAZAKI T， UCHIDA S， NAGATOISHI S， et al. Polymeric nanocarriers with controlled chain flexibility boost mRNA delivery in vivo through enhanced structural fastening［J］. Adv Healthc Mater， 2020， 9（16）： e2000538.
[2]	SHI J Q， YIN H Y， SUN Q M， et al. Rational design of polymeric mRNA delivery vectors to achieve excellent room-temperature storage stability and delivery efficiency［J］. Chem Mater， 2024， 36（11）： 5422-5435.
[3]	ZHANG H P， MENG C Y， YI X W， et al. Fluorinated lipid nanoparticles for enhancing mRNA delivery efficiency［J］. ACS Nano， 2024， 18（11）： 7825-7836.
[4]	樊渝川，殷涵，李钰，等. mRNA疫苗与脂质纳米颗粒递送载体的研究进展［J］. 科学通报， 2024， 69（33）： 4813-4823.
	FAN Y C， YIN H， LI Y， et al. Progress on mRNA vaccines and lipid nanoparticles［J］. Chin Sci Bull， 2024， 69（33）： 4813-4823.
[5]	张苗苗，李港，侯泰霖，等. 基于纳米技术的mRNA递送系统的研究进展［J］. 科学通报， 2024， 69（33）： 4858-4873.
	ZHANG M M， LI G， HOU T L， et al. Advancements in nanotechnology-enabled mRNA delivery systems［J］. Chin Sci Bull， 2024， 69（33）： 4858-4873.
[6]	LIU X， YANG Y L， HAN M M， et al. Guanylated hyperbranched polylysines with high in vitro and in vivo antifungal activity［J］. Adv Healthc Mater， 2022， 11（18）： e2201091.
[7]	SCHOLL M， NGUYEN T Q， BRUCHMANN B， et al. The thermal polymerization of amino acids revisited； synthesis and structural characterization of hyperbranched polymers from L-lysine［J］. J Polym Sci Pol Chem， 2007， 45（23）： 5494-5508.
[8]	HAJJ K A， MELAMED J R， CHAUDHARY N， et al. A potent branched-tail lipid nanoparticle enables multiplexed mRNA delivery and gene editing in vivo［J］. Nano Lett， 2020， 20（7）： 5167-5175.
[9]	FAN C Y， WANG S W， CHUNG C， et al. Synthesis of a dendritic cell-targeted self-assembled polymeric nanoparticle for selective delivery of mRNA vaccines to elicit enhanced immune responses［J］. Chem Sci， 2024， 15（29）： 11626-11632.
[10]	LAI Q Y， LI W L， HU D D， et al. Hepatic stellate cell-targeted chemo-gene therapy for liver fibrosis using fluorinated peptide-lipid hybrid nanoparticles［J］. J Control Release， 2024， 376： 601-617.
[11]	LIU S， WANG X， YU X L， et al. Zwitterionic phospholipidation of cationic polymers facilitates systemic mRNA delivery to spleen and lymph nodes［J］. J Am Chem Soc， 2021， 143（50）： 21321-21330.
[12]	JARZEBINSKA A， PASEWALD T， LAMBRECHT J， et al. A single methylene group in oligoalkylamine-based cationic polymers and lipids promotes enhanced mRNA delivery［J］. Angew Chem Int Ed， 2016， 55（33）： 9591-9595.
[13]	WANG L Y， LI Y C， JIANG P G， et al. Enhanced mRNA delivery via incorporating hydrophobic amines into lipid nanoparticles［J］. Colloids Surf B Biointerfaces， 2025， 249： 114528.
[14]	YONG H Y， LIN L X， LI Z L， et al. Tailoring highly branched poly（beta-amino ester）s for efficient and organ-selective mRNA delivery［J］. Nano Lett， 2024， 24（30）： 9368-9376.
[15]	ZHAO H Q， MA S， QI Y B， et al. A polyamino acid-based phosphatidyl polymer library for in vivo mRNA delivery with spleen targeting ability［J］. Mater Horiz， 2024， 11（11）： 2739-2748.
[16]	LONG J R， YU C X， ZHANG H L， et al. Novel ionizable lipid nanoparticles for SARS-CoV-2 Omicron mRNA delivery［J］. Adv Healthc Mater， 2023， 12（13）： e2202590.
[17]	GUO X Y， YANG Z Y， FANG H P， et al. Modulating the oxidation degree of linear polyethyleneimine for preparation of highly efficient and low-cytotoxicity degradable gene delivery carriers［J］. Chin J Polym Sci， 2024， 42（11）： 1699-1709.
[18]	KOWALSKI P S， PALMIERO C U， HUANG Y X， et al. Ionizable amino-polyesters synthesized via ring opening polymerization of tertiary amino-alcohols for tissue selective mRNA delivery［J］. Adv Mater， 2018： e1801151.
[19]	KACZMAREK J C， PATEL A K， KAUFFMAN K J， et al. Polymer-lipid nanoparticles for systemic delivery of mRNA to the lungs［J］. Angew Chem Int Ed， 2016， 55（44）： 13808-13812.
[20]	KIM H L， SARAVANAKUMAR G， LEE S， et al. Poly（beta-amino ester） polymer library with monomer variation for mRNA delivery［J］. Biomaterials， 2025， 314： 122896.
[21]	XUE L L， ZHAO G， GONG N Q， et al. Combinatorial design of siloxane-incorporated lipid nanoparticles augments intracellular processing for tissue-specific mRNA therapeutic delivery［J］. Nat Nanotechnol， 2025， 20（1）： 132-143.
[22]	唐国鸿，赵振，仲家慧，等. 氨基酸及其衍生物的生物基聚氨酯的制备和性能的研究进展［J］. 应用化学， 2025， 42（1）： 42-57.
	TANG G H， ZHAO Z， ZHONG J H， et al. Research progress on preparation and properties of bio-based polyurethanes from amino acid and its derivatives ［J］. Chin J Appl Chem， 2025， 42（1）： 42-57.
[23]	ZHANG X Y， SU K X， WU S Q， et al. One-component cationic lipids for systemic mRNA delivery to splenic T cells［J］. Angew Chem Int Ed， 2024， 63（26）： e202405444.
[24]	李港，侯泰霖，蒋为，等. 聚合物载体在mRNA递送领域的研究进展［J］. 药学进展， 2024， 48（6）： 437-449.
	LI G， HOU T L， JIANG W， et al. Research progress of polymer carriers in the field of mRNA delivery［J］. Prog Pharm Sci， 2024， 48（6）： 437-449.
[25]	CHEN G H， LIU X， LIU H， et al. Quaternary ammonium salt derivatives of hyperbranched polylysine with enhanced antibacterial activity against multidrug-resistant Gram-negative bacteria［J］. ACS Appl Bio Mater， 2024， 7（11）： 7444-7452.
[26]	DONG W， LI Z B， HOU T L， et al. Multicomponent synthesis of imidazole-based ionizable lipids for highly efficient and spleen-selective messenger RNA delivery［J］. J Am Chem Soc， 2024， 146（22）： 15085-15095.
[27]	KADLECOVA Z， BALDI L， HACKER D， et al. Comparative study on the in vitro cytotoxicity of linear， dendritic， and hyperbranched polylysine analogues［J］. Biomacromolecules， 2012， 13（10）： 3127-3137.
[28]	KADLECOVA Z， RAJENDRA Y， MATASCI M， et al. DNA delivery with hyperbranched polylysine： a comparative study with linear and dendritic polylysine［J］. J Control Release， 2013， 169（3）： 276-288.
[29]	NAIDU G S， YONG S B， RAMISHETTI S， et al. A combinatorial library of lipid nanoparticles for cell type-specific mRNA delivery［J］. Adv Sci， 2023， 10（19）： e2301929.
[30]	LV K， YU Z L， WANG J， et al. Discovery of ketal-ester ionizable lipid nanoparticle with reduced hepatotoxicity， enhanced spleen tropism for mRNA vaccine delivery［J］. Adv Sci， 2024， 11（45）： e2404684.
[31]	焦元昊，崔洪燕，张留伟，等. 基于原位聚合技术构建细胞内微环境响应型DNA递送系统［J］. 应用化学， 2022， 39（10）： 1510-1522.
	JIAO Y H， CUI H Y， ZHANG L W，et al. Fabrication of multifunctional gene delivery systems responsible to intracellular microenvironments through in situ polymerization［J］. Chin J Appl Chem， 2022， 39（10）： 1510-1522.
[32]	GAO Y X， ZHAO H Q， ZHAO J Y， et al. Polymer-based synthetic oncolytic virus-like nanoparticles for cancer immunotherapy［J］. Sci China Chem， 2023， 66（12）： 3576-3586.
[33]	DENG Y H， ZHANG J， SUN X M， et al. Potent gene delivery from fluorinated poly（beta-amino ester） in adhesive and suspension difficult-to-transfect cells for apoptosis and ferroptosis［J］. J Control Release， 2023， 363： 597-605.
[34]	DONG L Y， DENG X Q， LI Y， et al. Stimuli-responsive mRNA vaccines to induce robust CD8⁺ T Cell response via ROS-mediated innate immunity boosting［J］. J Am Chem Soc， 2024， 146（28）： 19218-19228.
[35]	ZHANG Y Y， HU Y Y， TIAN H Y， et al. Opportunities and challenges for mRNA delivery nanoplatforms［J］. J Phys Chem Lett， 2022， 13（5）： 1314-1322.
[36]	HAN X X， ALAMEH M G， XU Y， et al. Optimization of the activity and biodegradability of ionizable lipids for mRNA delivery via directed chemical evolution［J］. Nat Biomed Eng， 2024， 8（11）： 1412-1424.
[37]	LIU H， LIU X， CAO Y Q， et al. Engineering antibacterial activities and biocompatibility of hyperbranched lysine-based random copolymers［J］. Chin J Polym Sci， 2023， 41（3）： 345-355.
[38]	ZENG G G， HE Z P， YANG H H， et al. Cationic lipid pairs enhance liver-to-lung tropism of lipid nanoparticles for in vivo mRNA delivery［J］. ACS Appl Mater Interfaces， 2024， 16（20）： 25698-25709.
[39]	LI S R， LV M Y， MEI W K， et al. Fluorinated polyethylenimine and fluorinated choline phosphate lipids complex system for efficient mRNA delivery to deep-seated tumor tissues［J］. Biomacromolecules， 2024， 25（8）： 5251-5259.
[40]	KRANZ L M， DIKEN M， HAAS H， et al. Systemic RNA delivery to dendritic cells exploits antiviral defence for cancer immunotherapy［J］. Nature， 2016， 534（7607）： 396-401.
[41]	KADLECOVA Z， RAJENDRA Y， MATASCI M， et al. Hyperbranched polylysine： a versatile， biodegradable transfection agent for the production of recombinant proteins by transient gene expression and the transfection of primary cells［J］. Macromol Biosci， 2012， 12（6）： 794-804.

Hyperbranched Polylysine-Based mRNA Delivery Carrier Optimization and Evaluation

RichHTML

PDF

Supplement

Knowledge

Abstract

Cite this article

share this article

Figures/Tables 10

References 41

Related Articles 15

Recommended Articles

Metrics

Comments

[1]	Yu-Song LUO, Rui WANG. Copolymerization Modification and Properties of Poly（butylene succinate） by 2，2'-Bithiazole-4，4'-Dicarboxylic Acid [J]. Chinese Journal of Applied Chemistry, 2024, 41(11): 1565-1571.
[2]	Yun HONG, Xiao-Ye MA, Jing-Wei HOU, Ding-Xiao JIANG, Chuan-Qing KANG. Synthesis and Properties of Polyurethane Containing Polypeptide from Silk Fibroin [J]. Chinese Journal of Applied Chemistry, 2024, 41(10): 1425-1435.
[3]	Zhao-Yang DIAO, Li LIU, Chen-Ming LI, Huan-Li SUN. Research Progress on the Application of Nanomedicines in the Treatment of Acute Myeloid Leukemia [J]. Chinese Journal of Applied Chemistry, 2024, 41(9): 1259-1270.
[4]	Kun ZHANG, Bo ZHU, Chang-Bin DU, Yi HAN. Preparation and Radiation Shielding Mechanism of Lead-Boron-Polyethylene Composites [J]. Chinese Journal of Applied Chemistry, 2024, 41(8): 1168-1174.
[5]	Fang XIA, Zi-Ying XU, He WANG, Yu-Fang HE. Graphene Oxide Characterization Based on Bibliometrics [J]. Chinese Journal of Applied Chemistry, 2023, 40(10): 1448-1455.
[6]	Xiao-Hu LIU, Xiao-Juan LAI, Hong-Yan CAO, Ting-Ting WANG, Zhi-Qiang DANG. Synergistic Performance of Foaming Agent/Stabilizer/SiO₂ Composite Foam Retarded Acid System [J]. Chinese Journal of Applied Chemistry, 2023, 40(1): 91-99.
[7]	Tong CAO, Jun PENG, Yan FENG, Xiao-Bo LIU, Yu-Min HUANG. Research Progress of Side-Chain Sulfonated Polyarylene Ether Proton Exchange Membranes [J]. Chinese Journal of Applied Chemistry, 2022, 39(12): 1783-1802.
[8]	Yong-Xin TANG, Li-Wu NIE. Effect of Epoxy Resin Content on the Performance of Luminous Resin Permeable Concrete [J]. Chinese Journal of Applied Chemistry, 2022, 39(11): 1665-1671.
[9]	Bing-Gang CHEN, San-Rong LIU, Zi-Jiang JIANG, Xi-Fei YU. Preparation and Properties Characterization of Hydrophilic Polysiloxane and Polyvinyl Alcohol Composite as Skin Barrier Material [J]. Chinese Journal of Applied Chemistry, 2022, 39(8): 1224-1236.
[10]	Guan-Yu XIE, Mao LI. Topological Metallopolymers Synthesized by Electropolymerization and Their Photoelectric Properties [J]. Chinese Journal of Applied Chemistry, 2022, 39(8): 1246-1251.
[11]	Hong-Xia CHEN, Bao-Xia LI, Ying-Ming YAO, Peng LIU. Synthesis， Characterization and Catalytic Activity of Benzylamine⁃bridged Bis（phenolato）lanthanide Complexes [J]. Chinese Journal of Applied Chemistry, 2022, 39(5): 843-851.
[12]	. [J]. Chinese Journal of Applied Chemistry, 2022, 39(5): 855-856.
[13]	ZHANG Qiao-Feng, LI Guo, JIANG Jin-Qiang, LIU Zhao-Tie. Preparation and Luminescent Properties of Linear Multi-hydroxy Polyester [J]. Chinese Journal of Applied Chemistry, 2021, 38(8): 938-945.
[14]	WANG Xu-Yao, FANG Ying-Jun, ZHANG Ling-Zhi. Synthesis and Characterization of Cross-linked Solid Polymer Electrolyte Based on N-Cyanoethylated Polyethylenimine for Lithium-Ion Batteries [J]. Chinese Journal of Applied Chemistry, 2021, 38(8): 946-953.
[15]	WANG Yu-Cheng, QI Ying-Xia, LU Yi-Xuan, LU Yang. Preparation and Heat Storage/Release of Paraffin/Octadecanoic Acid/Octadecanol Composite [J]. Chinese Journal of Applied Chemistry, 2021, 38(8): 969-975.