Efficient Scaling up EV‐AAVs Production via Cellular Nanoporation for Familial Hypercholesterolaemia Therapy

作者：Yuting Yan, Yi You, Shuhong Ma, Hui Yi, Guangduo Chen, Jie Ni, Changyan Chen, Wujian Ke, Lingying Li, Rui Bai, Yuqing Ran, Wenjing Lu, Min Zhu, Yongshuai Zhang, Jing Dai, Man Qi, Feng Lan, Andrew S. Lee, Ran Zhang, Xujie Liu, Zhaoyang Chen · 发表于：Journal of Extracellular Vesicles · 年份：2025 · DOI：10.1002/jev2.70186 · 被引用次数：3 · 研究领域：Extracellular vesicles in disease、interferon and immune responses、Virus-based gene therapy research

Adeno-associated virus (AAV)-mediated gene therapies face critical clinical limitations, including immune-mediated neutralization by pre-existing antibodies and dose-dependent hepatotoxicity. Extracellular vesicle-encapsulated AAVs (EV-AAVs) offer a promising solution by shielding AAVs from antibody recognition, yet existing production methods remain inefficient and impractical for clinical application. Here, we developed a cellular nanoporation (CNP) platform that enables scalable, high-yield generation of EV-AAVs, achieving an approximately 11-fold increase in production efficiency compared with conventional methods. In LDLR-deficient murine models with pre-existing neutralizing antibodies (1:200), EV-AAV-LDLR at half the standard AAV dose robustly restored hepatic LDL receptor expression and attenuated atherosclerosis progression. Notably, EV-AAV exhibited superior immune evasion capabilities, maintaining 2.3-fold higher hepatic transduction efficiency than conventional AAV upon secondary dosing due to antibody shielding. Importantly, EV-AAV therapy markedly reduced hepatotoxicity, with serum AST/ALT levels comparable to saline-treated controls, thereby overcoming a critical safety barrier of high-dose AAV treatment. These results demonstrate CNP as a clinically translatable platform for scalable EV-AAV manufacturing, enabling effective multi-dose regimens while overcoming key immunological and toxicity barriers in liver-directed gene therapy for familial hypercholesterola...