Department of Pharmaceutical Sciences, College of Pharmacy, Oregon State University, 2730 S Moody Avenue, Portland, Oregon 97201, United States.
Department of Biomedical Engineering, Oregon Health & Science University, 3303 SW Bond Avenue, Portland, Oregon 97239, United States.
ACS Nano. 2024 Sep 10;18(36):24842-24859. doi: 10.1021/acsnano.4c05088. Epub 2024 Aug 26.
mRNA therapeutics encapsulated in lipid nanoparticles (LNPs) offer promising avenues for treating various diseases. While mRNA vaccines anticipate immunogenicity, the associated reactogenicity of mRNA-loaded LNPs poses significant challenges, especially in protein replacement therapies requiring multiple administrations, leading to adverse effects and suboptimal therapeutic outcomes. Historically, research has primarily focused on the reactogenicity of mRNA cargo, leaving the role of LNPs understudied in this context. Adjuvanticity and pro-inflammatory characteristics of LNPs, originating at least in part from ionizable lipids, may induce inflammation, activate toll-like receptors (TLRs), and impact mRNA translation. Knowledge gaps remain in understanding LNP-induced TLR activation and its impact on induction of animal sickness behavior. We hypothesized that ionizable lipids in LNPs, structurally resembling lipid A from lipopolysaccharide, could activate TLR4 signaling via MyD88 and TRIF adaptors, thereby propagating LNP-associated reactogenicity. Our comprehensive investigation utilizing gene ablation studies and pharmacological receptor manipulation proves that TLR4 activation by LNPs triggers distinct physiologically meaningful responses in mice. We show that TLR4 and MyD88 are essential for reactogenic signal initiation, pro-inflammatory gene expression, and physiological outcomes like food intake and body weight─robust metrics of sickness behavior in mice. The application of the TLR4 inhibitor TAK-242 effectively reduces the reactogenicity associated with LNPs by mitigating TLR4-driven inflammatory responses. Our findings elucidate the critical role of the TLR4-MyD88 axis in LNP-induced reactogenicity, providing a mechanistic framework for developing safer mRNA therapeutics and offering a strategy to mitigate adverse effects through targeted inhibition of this pathway.
mRNA 疗法被包裹在脂质纳米颗粒 (LNPs) 中，为治疗各种疾病提供了有前途的途径。虽然 mRNA 疫苗可引发免疫原性，但负载 mRNA 的 LNPs 相关的反应原性带来了重大挑战，尤其是在需要多次给药的蛋白质替代疗法中，这会导致不良反应和不理想的治疗效果。从历史上看，研究主要集中在 mRNA 有效载荷的反应原性上，而 LNPs 在这种情况下的作用则研究不足。LNPs 的佐剂特性和促炎特性至少部分源自可离子化脂质，可能会引发炎症、激活 toll 样受体 (TLR)，并影响 mRNA 翻译。在理解 LNP 诱导的 TLR 激活及其对动物疾病行为诱导的影响方面，仍存在知识空白。我们假设 LNPs 中的可离子化脂质在结构上类似于脂多糖中的脂质 A，可通过 MyD88 和 TRIF 衔接子激活 TLR4 信号转导，从而引发 LNP 相关的反应原性。我们利用基因敲除研究和药理学受体操作进行了全面研究，证明 LNPs 激活 TLR4 会在小鼠中引发独特的具有生理意义的反应。我们表明 TLR4 和 MyD88 对于反应原信号的起始、促炎基因表达以及生理结果（如食物摄入和体重）是必需的，这些都是小鼠疾病行为的有力指标。TLR4 抑制剂 TAK-242 的应用通过减轻 TLR4 驱动的炎症反应，有效地降低了与 LNPs 相关的反应原性。我们的研究结果阐明了 TLR4-MyD88 轴在 LNP 诱导的反应原性中的关键作用，为开发更安全的 mRNA 疗法提供了机制框架，并为通过靶向抑制该途径减轻不良反应提供了策略。
