Work

N1‐methylpseudouridine modification level correlates with protein expression, immunogenicity, and stability of mRNA

Year: 2024

Type: article

Abstract: N1-methylpseudouridine modification level correlates with protein expression, immunogenicity, and stability of mRNADear Editor, Messenger RNA (mRNA) has undergone significant evolution, emerging as a robust platform for diverse therapeutic applications such as vaccines, protein replacement, and adoptive cell therapy across infectious, cancer, and immunological diseases.In vitro transcribed mRNA offers distinct advantages, including cytoplasmic transient expression, devoid of the risk of genomic integration.Nucleotide modifications, such as pseudouridine (Ψ), N1methylpseudouridine (m1Ψ), and 5-methylcytidine (m5C), play pivotal roles in mRNA immunogenicity, stability, and translational efficiency.Notably, standing out as a state-of-the-art modification, global m1Ψ modification has been performed in the two approved mRNA vaccines against severe acute respiratory syndrome coronavirus 2, mRNA-1273, and BNT162b2.The in vivo stability and translational duration of synthesized mRNA are partially restricted by its immunogenicity.Recognition of foreign RNA by toll-like receptor 3 (TLR3), TLR7, TLR8, and retinoic acid-inducible gene I (RIG-I)-like receptors triggers innate immunity, leading to RNA degradation.1,2 The therapeutic application of mRNA necessitates a delicate balance between immune activation and protein expression.For infectious disease vaccines, chemical modifications can minimize inflammatory responses while ensuring effective mRNA translation.Conversely, cancer vaccines require adequate innate immune stimulation for anti-tumor immunity.Thus far, most studies have investigated alternative m1Ψ modification individually or combined with others.However, it is unknown whether m1Ψ modification ratio in mRNA has an impact on the delicate balance between immunogenicity, stability, and translational efficiency.Therefore, we synthesized mRNA encoding enhanced green fluorescent protein (mEGFP) with different m1Ψ modification ratios (5%, 10%, 20%, 50%, 75%, and 100%) through in vitro transcription.HEK-293T cells were transfected with unmodified mEGFP or m1Ψ-modified m1ΨEGFP (m1ΨEGFP-5%, m1ΨEGFP-10%, m1ΨEGFP-

Source: MedComm

DOI: 10.1002/mco2.691

Authors: Shaoyi Chen, Zheng Liu, Jingsheng Cai, Haoran Li, Mantang Qiu

Institutions: Peking University, Peking University People's Hospital

Language: English

Topic: RNA and protein synthesis mechanisms

Subfield: Molecular Biology

Field: Biochemistry, Genetics and Molecular Biology

Domain: Life Sciences

SDG: Good health and well-being

Open Access status: gold

Estimated APC paid: $3,500