Department of Chemical Engineering, Carnegie Mellon University, Pittsburgh, PA, United States of America.
Department of Chemical Engineering, Carnegie Mellon University, Pittsburgh, PA, United States of America; Department of Biomedical Engineering, Carnegie Mellon University, Pittsburgh, PA, United States of America.
J Control Release. 2022 May;345:819-831. doi: 10.1016/j.jconrel.2022.03.046. Epub 2022 Mar 26.
The broad clinical application of mRNA therapeutics has been hampered by a lack of delivery vehicles that induce protein expression in extrahepatic organs and tissues. Recently, it was shown that mRNA delivery to the spleen or lungs is possible upon the addition of a charged lipid to a standard four-component lipid nanoparticle formulation. This approach, while effective, further complicates an already complex drug formulation and has the potential to slow regulatory approval and adversely impact manufacturing processes. We were thus motivated to maintain a four-component nanoparticle system while achieving shifts in tropism. To that end, we replaced the standard helper lipid in lipidoid nanoparticles, DOPE, with one of eight alternatives. These lipids included the neutral lipids, DOPC, sphingomyelin, and ceramide; the anionic lipids, phosphatidylserine (PS), phosphatidylglycerol, and phosphatidic acid; and the cationic lipids, DOTAP and ethyl phosphatidylcholine. While neutral helper lipids maintained protein expression in the liver, anionic and cationic lipids shifted protein expression to the spleen and lungs, respectively. For example, replacing DOPE with DOTAP increased positive LNP surface charge at pH 7 by 5-fold and altered the ratio of liver to lung protein expression from 36:1 to 1:56. Similarly, replacing DOPE with PS reduced positive charge by half and altered the ratio of liver to spleen protein expression from 8:1 to 1:3. Effects were consistent across ionizable lipidoid chemistries. Regarding mechanism, nanoparticles formulated with neutral and anionic helper lipids best transfected epithelial and immune cells, respectively. Further, the lung-tropic effect of DOTAP was linked to reduced immune cell infiltration of the lungs compared to neutral or anionic lipids. Together, these data show that intravenous non-hepatocellular mRNA delivery is readily achievable while maintaining a four-component formulation with modified helper lipid chemistry.
mRNA 疗法的广泛临床应用受到缺乏能够在肝外器官和组织中诱导蛋白表达的递药载体的限制。最近，研究表明，在标准的四组分脂质纳米颗粒制剂中添加带电荷的脂质，可以实现 mRNA 向脾脏或肺部的递药。这种方法虽然有效，但进一步增加了原本复杂的药物制剂的复杂性，并有可能减缓监管审批过程并对制造工艺产生不利影响。因此，我们希望在保持四组分纳米颗粒系统的同时实现转导谱的改变。为此，我们用八种替代脂质中的一种替代脂质体纳米颗粒中的标准辅助脂质 DOPE。这些脂质包括中性脂质 DOPC、鞘磷脂和神经酰胺；阴离子脂质磷脂酰丝氨酸 (PS)、磷脂酰甘油和磷脂酸；以及阳离子脂质 DOTAP 和乙基磷脂酰胆碱。虽然中性辅助脂质维持了肝脏中的蛋白表达，但阴离子和阳离子脂质分别将蛋白表达转移到脾脏和肺部。例如，用 DOTAP 替代 DOPE 将 pH 7 时的阳性 LNP 表面电荷增加了 5 倍，并将肝脏与肺部蛋白表达的比值从 36:1 改变为 1:56。同样，用 PS 替代 DOPE 将正电荷减少一半，并将肝脏与脾脏蛋白表达的比值从 8:1 改变为 1:3。这些效应在可离子化的脂质体化学中是一致的。关于机制，用中性和阴离子辅助脂质制备的纳米颗粒分别最有效地转染上皮细胞和免疫细胞。此外，与中性或阴离子脂质相比，DOTAP 的肺部转导作用与肺部免疫细胞浸润减少有关。总之，这些数据表明，通过维持改良辅助脂质化学的四组分制剂，很容易实现非肝细胞内静脉内 mRNA 递药。
