Modified Nucleic Acids for Low-Immunogenic Protein Expression
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Solution Overview
Problem
Existing methodologies for protein expression using heterologous DNA face challenges such as integration into host cell genomic DNA, leading to alterations and damage, and inefficient translation due to multiple processing steps, particularly in primary cells or modified cell lines.
Innovation Solution
Modified nucleosides, nucleotides, and nucleic acids with chemical modifications on the major groove face to disrupt binding with partners, reducing innate immune response and enhancing protein production efficiency.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If heterologous DNA is introduced into cells for protein expression, then protein production is achieved, but integration into host cell genomic DNA occurs causing alterations and damage
Solution Approach 1:
The patent uses modified nucleosides and nucleotides as intermediary substances that mediate between the introduced nucleic acid and the host cell's DNA repair and immune systems. These modifications act as a buffer that prevents direct harmful interactions while still allowing protein expression to proceed.
Solution Approach 2:
The patent applies chemical modifications to nucleosides and nucleotides (changing their chemical parameters) to alter how they interact with cellular systems. Specifically, modifications at positions 2 and 4 of pyrimidine bases change the binding affinity with major groove interacting partners, thereby reducing immunogenicity and genomic integration while maintaining translational efficiency.
2Productivity
If multiple processing steps are used for DNA to protein conversion, then complete expression pathway is achieved, but lag times occur before protein generation
Solution Approach 1:
The patent extracts the transcription step from the traditional DNA→RNA→protein pathway by directly introducing modified nucleic acids that can function as mRNA in the cytoplasm. This eliminates the need for nuclear transcription and reduces the number of processing steps, thereby reducing lag time while maintaining complete protein expression capability.
3Productivity
If unmodified nucleic acids are used for protein production, then standard translation pathway is maintained, but binding with major groove partners causes immunogenicity
Solution Approach 1:
The patent applies local chemical modifications at specific positions (2 and 4 of pyrimidine bases) rather than uniform modifications throughout the nucleic acid. This localized approach preserves essential translation functions while specifically targeting the regions that interact with major groove binding partners, thereby reducing immunogenicity without compromising translation efficiency.
Data Source
AI summary
The present disclosure provides modified nucleosides, nucleotides, and nucleic acids, and methods of using thereof.