Ghoshal Bartika, Chakraborty Debajyoti, Nag Manish, Varadarajan Raghavan, Jhunjhunwala Siddharth
Department of Bioengineering, Indian Institute of Science, Bengaluru 560012, India.
Molecular Biophysics Unit, Indian Institute of Science, Bengaluru 560012, India.
ACS Bio Med Chem Au. 2024 Nov 12;4(6):291-299. doi: 10.1021/acsbiomedchemau.4c00057. eCollection 2024 Dec 18.
Base modification and the use of lipid nanoparticles are thought to be essential for efficient in vivo delivery and expression of mRNA. However, for ex vivo immune cell engineering, the need for either of the two is unclear. Previous reports have suggested that nucleic acids may be efficiently delivered to immune cells ex vivo, through a nonendosomal delivery route, but the need for base modification has not been determined. Herein, we demonstrate that when a nonendosomal delivery method is used, unmodified mRNA performs equally well to the commonly used base-modified mRNA, including the methyl pseudouridine modification, in terms of protein expression and inflammatory response in cells. However, if an endosomal delivery route is used, then methyl pseudouridine modification is necessary for high expression and low inflammatory response, as demonstrated by others as well. Overall, we show that nonendosomal mRNA delivery renders nucleoside modifications nonessential and that unmodified mRNA combined with nonendosomal delivery route may be used for efficient ex vivo mRNA-based engineering of immune cells.
碱基修饰和脂质纳米颗粒的使用被认为是实现mRNA在体内高效递送和表达所必需的。然而，对于离体免疫细胞工程而言，这两者中任何一个的必要性尚不清楚。先前的报告表明，核酸或许可以通过非内体递送途径有效地离体递送至免疫细胞，但碱基修饰的必要性尚未确定。在此，我们证明，当使用非内体递送方法时，就细胞中的蛋白质表达和炎症反应而言，未修饰的mRNA与常用的碱基修饰mRNA（包括N1-甲基假尿苷修饰）表现相当。然而，正如其他人所证明的那样，如果使用内体递送途径，那么N1-甲基假尿苷修饰对于高表达和低炎症反应是必要的。总体而言，我们表明非内体mRNA递送使得核苷修饰不再必要，并且未修饰的mRNA与非内体递送途径相结合可用于基于mRNA的高效离体免疫细胞工程。
