(T1330-01-02) Formulation Development and Characterization of a Marburg Self-amplifying mRNA-Lipid Nanoparticle Vaccine with a Toll-Like Receptor 7 Agonist

Purpose: Synthetic self-amplifying mRNA (saRNA) vaccines have attracted increasing attention due to the rapid development potential and capacity for dose-sparing. saRNA vaccines need to be formulated as lipid nanoparticles (LNPs) for delivery to the cell and to prevent mRNA degradation. Toll-like Receptor (TLR) agonists, which boost the innate immune response, have been used as adjuvants in subunit vaccines; however, it is uncertain as to whether these TLR agonists will act as an adjuvant with the sa mRNA-LNP vaccines. To test this, we have performed formulation studies using the Marburg Virus Glycoprotein (MARV GP) saRNA, from Imperial College, as our model antigen.

Methods: The MARV GP saRNA has been encapsulated in ionizable LNPs formulation consists of ionizable lipids (DLin-MC3-DMA) DSPC, cholesterol and PEG-lipid using a standardized microfluidics approach. An optimized iLNP formulation for encapsulating sa RNA were used to incorporate the imidazoquinoline resiquimod (R848) (LogP = 1.55) or another form of TLR 7 telratolimod.

Results: the maximal % encapsulation efficiency of this small molecule TLR7 (R848) agonist was ~50%.  To address this issue, formulation development was subsequently performed using an alkylated imidazoquinoline, telratolimod, instead of R848, which led to a tangible increase in encapsulation ( >80% encapsulation). The critical quality attributes (CQAs) of the optimized telratolimod vaccine formulations were assessed using Dynamic Light Scattering (DLS), electrophoretic light scattering, RNA encapsulation efficiency, agonist encapsulation efficiency.  In addition, physico-chemical and morphological characterization was performed on the optimized vaccine formulation.

Conclusion: Overall, the data presented in this study provide a formulation development approach for the preparation of an saRNA-LNP with suitable TLR7 adjuvant for vaccine formulations to evaluate the immunogenicity and co-stimulation of the innate immune system.

(A) Encapsulation efficiency for TLR7 agonist forms in the optimized mRNA-LNP vaccine formulation. (B) Encapsulation efficiency for Marburg sa mRNA in LNPs incorporated with Telratolimod or Resiquimod