E46 - Optimization of a mRNA Vaccine Candidate to Immunize Foals Against Rhodococcus equi

Abstract: Background- A vaccine against Rhodococcus equi (R. equi) is lacking because foals become infected after birth, when maternal antibodies inhibit responses to traditional vaccines. Vaccination with messenger RNA (mRNA) was recently demonstrated to circumvent maternal antibody inhibition and protect neonatal mice against influenza. We aim to develop a mRNA vaccine for foals targeting the virulence-associated protein A (VapA) of R. equi.

Hypothesis/Objectives- Expression and secretion of VapA by equine cells will vary significantly among 4 mRNA vaccine constructs encoding: 1) native VapA, including its transmembrane domain (TM); 2) a codon-optimized version of construct 1; 3) VapA without the TM domain and with an equine leader peptide; and, 4) a codon-optimized version of construct 3.

Animals- Equine bronchial fibroblasts (EBFBs) and equine bronchial epithelial cells (EBECs) cultured from healthy adult horses.

Methods- EBFBs and EBECs were transfected with each of the 4 mRNA constructs formulated in lipid nanoparticles and expression of VapA was evaluated by western immunoblot of cell lysates and supernatants using anti-VapA monoclonal antibody.

Results- Constructs lacking the TM domain (3 and 4) were expressed by EBFBs and EBECs. Construct 3 was more highly expressed than construct 4 in cells and supernatants. ELISA results for VapA expression in supernatants are pending.

Conclusions and clinical importance- Removing the TM domain improved secretion whereas codon optimization did not improve in vitro expression. Based on these results, we plan to evaluate the immunogenicity of construct 3 formulated in lipid nanoparticles by either nebulization or intramuscular injection to neonatal foals.