Circular mRNA against CleanCap linear mRNA vectors: comprehensive comparison, expression, active and passive...

Area of Science:

• Molecular Biology
• Vaccinology
• Biotechnology

Background:

• Messenger RNA (mRNA) technology has transformed vaccine development due to its universal, rapid, and scalable production.
• Two primary mRNA vector types, linear (cap-dependent) and circular (cap-independent), offer distinct advantages.
• A comprehensive comparative analysis of the most effective vectors of each type was lacking.

Purpose of the Study:

• To compare the expression efficiency, protective activity, and therapeutic activity of circular and linear mRNA vectors.
• To evaluate different configurations of linear and circular mRNA vectors for optimal performance.

Main Methods:

• Compared linear and circular mRNA vectors using luciferase reporter assays and target protein expression.
• Assessed protective activity via active immunization (immunogenicity and SARS-CoV-2 challenge) and passive immunization (antibody production and toxin challenge).
• Tested linear vectors with N1-methylpseudouridine or uridine and ARCA or CleanCap, and circular vectors with different IRES elements.

Main Results:

• Modified linear mRNA vectors demonstrated significantly higher expression levels both in vitro and in vivo compared to circular vectors.
• Active immunization studies showed comparable immunogenicity and protective activity between linear and circular vectors.
• Linear vectors provided significantly better protection in passive immunization models, correlating with higher in vivo protein expression.

Conclusions:

• While modified linear mRNA vectors exhibit superior expression, they did not outperform circular vectors in active immunization immunogenicity or protection.
• Linear mRNA vectors appear to be a better choice for passive immunization strategies that necessitate high target protein expression levels.