Millisecond Rapid Mixing Stopped-Flow BioSAXS Study Comparing the mRNA and Ribonucleoprotein Loading Mechanism in BioNTech/Pfizer and Modern

The interaction kinetics of lipid nanoparticle (LNP) with its payloads such as proteins and mRNAs controls product stability, loading and release efficiency, as well as efficacy after administered in the biological system. We still have very limited understanding of interactions between charged LNPs and the gene-editing therapeutic candidates such as mRNAs and RNPs. The aim of the proposal is to investigate the structural dynamics of LNPs upon mRNAs and ribonucleoprotein (RNPs) loading on a millisecond time scale using the rapid mixing stopped-flow BioSAXS facility at the ID02 beamline. Steady state SAXS experiment has shown that the ionisable ALC-0315 lipid can form lyotropic liquid crystalline cubic and hexagonal mesophases which are responsive to pH. We will investigate how model mRNAs and RNPs drive nanostructural changes of cationic LNPs, including the BioNTech/Pfizer COVID-19 Vaccine LNPs (ALC-0315 and ALC-0159), and the Moderna COVID-19 Vaccine LNPs (SM-102 and PEG-DMG).

脂质纳米颗粒（lipid nanoparticle, LNP）与其有效载荷（如蛋白质、信使核糖核酸（mRNA））之间的相互作用动力学，决定了产品的稳定性、装载效率与释放效率，以及进入生物系统后的药效。目前学界对带电荷LNP与基因编辑治疗候选物（如mRNA及核糖核蛋白（RNPs））之间的相互作用仍缺乏深入认知。本研究计划旨在利用ID02光束线的快速混合停流生物小角X射线散射（BioSAXS）装置，在毫秒时间尺度下探究LNP在装载mRNA及核糖核蛋白（RNPs）后的结构动力学变化。稳态小角X射线散射（small-angle X-ray scattering, SAXS）实验已证实，可电离脂质ALC-0315可形成对pH具有响应性的溶致液晶立方相与六方介观相。我们将研究模型mRNA及RNPs如何驱动阳离子LNP的纳米结构变化，其中包括百奥泰（BioNTech）/辉瑞（Pfizer）COVID-19疫苗所用的LNP配方（包含ALC-0315与ALC-0159），以及莫德纳（Moderna）COVID-19疫苗所用的LNP配方（包含SM-102与PEG-DMG）。