Mannich reaction-based combinatorial libraries identify antioxidant ionizable lipids for mRNA delivery with reduced...

Area of Science:

• Biotechnology
• Immunology
• Drug Delivery

Background:

• Lipid nanoparticles (LNPs) are crucial for messenger RNA (mRNA) delivery.
• The immunogenicity of current LNPs restricts therapeutic applications by limiting protein expression levels and durability.
• Developing novel ionizable lipids is essential to overcome these limitations.

Purpose of the Study:

• To synthesize and screen novel ionizable lipids for reduced immunogenicity in mRNA delivery systems.
• To identify an ionizable lipid that mitigates the inflammatory response associated with LNPs.
• To evaluate the efficacy of LNP formulations incorporating the novel lipid for enhanced protein expression and therapeutic gene editing.

Main Methods:

• Leveraged the Mannich reaction for the synthesis of ionizable lipids.
• Conducted in vitro and in vivo screening of six combinatorial lipid libraries.
• Formulated LNPs with the identified antioxidant ionizable lipid, C-a16, for mRNA delivery.
• Assessed LNP performance in mice using mRNA for gene editing (Cas9), protein expression (FGF21), and immune responses (tumor neoantigen, SARS-CoV-2 spike protein).

Main Results:

• Identified C-a16, an antioxidant ionizable lipid with reduced immunogenicity.
• LNPs with C-a16 mitigated intracellular reactive oxygen species generation, extending protein expression duration.
• Demonstrated significantly enhanced gene editing efficiency (2.8-fold) and protein expression (3.6-fold) compared to commercial LNPs.
• Observed stronger antigen-specific immune responses when delivering mRNA encoding tumor neoantigen or SARS-CoV-2 spike protein.

Conclusions:

• C-a16 is a promising antioxidant ionizable lipid for mRNA delivery.
• LNPs incorporating C-a16 show potential for improved therapeutic outcomes in gene editing and protein replacement therapies.
• Further investigation of C-a16 is warranted for advancing mRNA-based therapeutics.