Cau Massimo F, Ferraresso Francesca, Seadler Monica, Badior Katherine, Zhang Youjie, Ketelboeter Laura M, Rodriguez Geoffrey G, Chen Taylor, Ferraresso Matteo, Wietrzny Amanda, Robertson Madelaine, Haugen Amber, Cullis Pieter R, de Moya Marc, Dyer Mitchell, Kastrup Christian J
Michael Smith Laboratories, University of British Columbia, Vancouver, BC V6T 1Z4, Canada.
School of Biomedical Engineering, University of British Columbia, Vancouver, BC V6T 1Z4, Canada.
Mol Ther Methods Clin Dev. 2024 Apr 27;32(2):101258. doi: 10.1016/j.omtm.2024.101258. eCollection 2024 Jun 13.
Genetic manipulation of animal models is a fundamental research tool in biology and medicine but is challenging in large animals. In rodents, models can be readily developed by knocking out genes in embryonic stem cells or by knocking down genes through delivery of nucleic acids. Swine are a preferred animal model for studying the cardiovascular and immune systems, but there are limited strategies for genetic manipulation. Lipid nanoparticles (LNPs) efficiently deliver small interfering RNA (siRNA) to knock down circulating proteins, but swine are sensitive to LNP-induced complement activation-related pseudoallergy (CARPA). We hypothesized that appropriately administering optimized siRNA-LNPs could knock down circulating levels of plasminogen, a blood protein synthesized in the liver. siRNA-LNPs against plasminogen (siPLG) reduced plasma plasminogen protein and hepatic plasminogen mRNA levels to below 5% of baseline values. Functional assays showed that reducing plasminogen levels modulated systemic blood coagulation. Clinical signs of CARPA were not observed, and occasional mild and transient hepatotoxicity was present in siPLG-treated animals at 5 h post-infusion, which returned to baseline by 7 days. These findings advance siRNA-LNPs in swine models, enabling genetic engineering of blood and hepatic proteins, which can likely expand to proteins in other tissues in the future.
对动物模型进行基因操作是生物学和医学中的一项基础研究工具，但在大型动物中具有挑战性。在啮齿动物中，可以通过敲除胚胎干细胞中的基因或通过递送核酸来敲低基因，从而轻松建立模型。猪是研究心血管和免疫系统的首选动物模型，但基因操作的策略有限。脂质纳米颗粒（LNPs）能有效地递送小干扰RNA（siRNA）以敲低循环蛋白，但猪对LNP诱导的补体激活相关假过敏（CARPA）敏感。我们假设适当施用优化的siRNA-LNPs可以敲低纤溶酶原的循环水平，纤溶酶原是一种在肝脏中合成的血液蛋白。针对纤溶酶原的siRNA-LNPs（siPLG）将血浆纤溶酶原蛋白和肝脏纤溶酶原mRNA水平降低至基线值的5%以下。功能测定表明，降低纤溶酶原水平可调节全身血液凝固。未观察到CARPA的临床症状，在输注后5小时，siPLG处理的动物偶尔出现轻度和短暂的肝毒性，到第7天恢复到基线水平。这些发现推动了siRNA-LNPs在猪模型中的应用，实现了血液和肝脏蛋白的基因工程，未来可能会扩展到其他组织中的蛋白。
