T. Kovačič, H. Haas, L. Stotsky-Oterin, A. Štrancar, U. Bren, and D. Peer
Nature Reviews Chemistry, 2025
Abstract:
This study addresses the chemical complexity of RNA-LNP formulations, crucial for mRNA-based therapies but challenging in terms of quality, stability, and reproducibility. As formulations evolve to include multiple nucleic acids and targeting ligands, their complexity increases, requiring meticulous control in Chemistry, Manufacturing, and Controls (CMC). The intrinsic chemical reactivity between RNA and lipid components can compromise drug performance, with degradation reactions often undetected by standard assays. Orthogonal analytical tools like Reversed Phase-Ion Pair Liquid Chromatography (RP-IP-LC) and mass spectrometry are vital for detecting these degradants.
Sartorius BIA Separations PATfix LNP Switcher and CIM monolithic columns, play a crucial role in this study. The PATfix LNP Switcher is instrumental in accurately determining chemical impurities in RNA-LNP formulations, such as nucleic lipid adducts and fragmented RNA, using an improved chromatographic 2D method that allows direct deformulation without pre-treatment, ensuring no sample loss and yielding better results. Additionally, it facilitates the determination of encapsulation efficiency, size, size distribution, and total concentration. CIM OH columns are adept at selectively capturing particles, achieving resolution between encapsulated and free RNA, and enabling scale-up for LNP purification. CIM SDVB columns provide resolution between different sized RNA and hydrophobic RNA species, enabling quantification of RNA, its fragmentation, and RNA-lipid adducts. These products are essential in advancing the analytical techniques required for comprehensive characterization and successful development of novel RNA-LNP pharmaceuticals
