Development of a self-amplifying RNA vaccine against Rift Valley fever

Background: Rift Valley fever (RVF) is a mosquito-borne zoonosis caused by the Rift Valley fever virus (RVFV), which leads to high mortality in livestock and severe or fatal complications in humans. Despite its severity, no licensed RVF vaccine exists for humans, and licensed livestock vaccines have suboptimal safety and immunogenicity. This thesis aimed to design and preclinically evaluate self-amplifying RNA (saRNA) vaccines against RVF and transfer saRNA vaccine development capacity to a resource-limited setting (Uganda). Methods: Mutations enhancing plasma membrane expression and glycoprotein stability were introduced into a consensus RVFV medium (M) segment sequence (WT consensus) and evaluated for in vitro expression of glycoprotein n (Gn) and c (Gc) after cloning into the pcDNA3.1(+) vector. The WT consensus and the mutated construct with the highest plasma membrane Gn expression (Furin-T2A) were individually cloned into a Venezuelan equine encephalitis virus (VEEV) pDNA replicon, transcribed in vitro into saRNA and capped. The saRNA constructs were then assessed for immunogenicity in BALB/c mice after formulation in lipid nanoparticles (LNPs) in the UK and cationic lipids in Uganda. Results: LNP-formulated saRNA WT consensus and Furin-T2A candidate vaccines induced high levels of anti-Gn IgG after two vaccinations (mean = 522,000 ng/mL, SD = 424,736 ng/mL, p < 0.01) and (mean = 522,848, SD = 366,604, p < 0.01), respectively. However, only the WT consensus induced significant RVFV pseudovirus-neutralising activity (median IC50= 5089, IQR = 5500, p < 0.01) compared to the unvaccinated control group. In contrast, saRNA RVF vaccines formulated in cationic lipids induced variable and weak humoral and T-cell immune responses. Conclusion: This thesis demonstrated that an LNP-formulated saRNA RVF vaccine can induce robust humoral immune responses in mice and merits further development. It also demonstrated the transfer of saRNA vaccine development technology to Uganda, advancing local vaccine research and development capacity.