Scaling up the encapsulation process of the mRNA vaccine developed by Bio- Manguinhos against COVID-19

Introduction: Messenger RNA (mRNA) technology has been studied globally for over 30 years, but it was in 2021 that the first vaccine using this technology was approved.mRNA vaccines against COVID-19 played a crucial role in preventing severe infections from SARS-CoV-2, enabling a return to normalcy and helping to control the pandemic.For effective mRNA delivery, an appropriate system is essential, with lipid nanoparticles (LNPs) being the most studied and used system, ensuring efficient RNA delivery to target cells.During the pandemic, Bio-Manguinhos developed an mRNA vaccine using microfluidics for RNA encapsulation in LNPs.Initially, the vaccine was developed on a small scale using a microfluidic encapsulator, demonstrating promising results in vitro and in vivo, which led to the possibility of scale up the encapsulation process using a larger scale encapsulator.Objectives: This work aims to establish the appropriate parameters for scaling up the mRNA encapsulation process in LNPs of the COVID-19 vaccine developed by Bio-Manguinhos from small to medium scale. Methodology:The formulations were produced using a nanoencapsulation equipment suitable for volumes above 50 mL.The formulations were produced with the previously established proportions of 4 different lipids: an ionizable lipid, an auxiliary phospholipid, a cholesterol and a pegylated lipid.Different parameters of the production process were tested, such as: the Nitrogen:Phosphate ratio (N:P), the Total Flow Rate (TFR) and the Target Flow Rate Ratio (TFRR).After production, parameters such as average hydrodynamic size, polydispersity index (PDI), zeta potential and encapsulation efficiency.Results: Over 20 formulations were produced during these scale-up tests.Of these, 4 formulations exhibited an average hydrodynamic size between 80-100 nm, 16 formulations showed a PDI < 0.3, and all demonstrated encapsulation efficiency greater than 90%. Conclusion:The scale-up process to produce the mRNA vaccine developed by Bio-Manguinhos was successful, with promising results in critical quality parameters such as hydrodynamic size, polydispersity index, and encapsulation efficiency.The advancement in the LNP encapsulation process ensures the vaccine's efficacy and opens opportunities for future innovations in RNA-based vaccine development, with potential for treating other diseases.