New approaches for improving the immunogenicity of modified vaccinia virus Ankara as a recombinant vaccine vector

Thesis

The CD8+ T cell response to a transgenic antigen expressed in Modified Vaccinia virus Ankara (MVA) can be augmented with enhanced transgene expression. This thesis presents different approaches to increase transgene expression in MVA. The transgene used was the sequence of the pb9 murine malaria epitope, fused to Renilla luciferase gene, and then fused to a secretion leader sequence (tPA) at the N terminus. The insertion of Internal Ribosomal Entry Site (IRES) upstream of the transgene, driven by the pB8 promoter and inserted at the B8R locus, or the modification of the pB8 promoter by nucleotide substitution both failed to enhance the transgene expression or immunogenicity. However, utilising endogenous promoters such as the pB8, pE3, and pF11 at their natural loci to drive the transgene resulted in lower transgene expression in vitro, but stronger immune responses to the pb9 epitope in BALB/c mice, as compared to the conventional p7.5 or mH5 promoters. Utilising these promoters at the TK locus (ectopic promoters) revealed similar patterns of transgene expression and immunogenicity, indicating that the improved immunogenicity is linked to promoters despite the low transgene expression measured in vitro. Therefore, the pF11 promoter was further studied to determine its activity in vivo and showed higher transgene expression, as compared to the mH5, which correlated with in vivo immunogenicity. Inactivation of the TK gene in MVAs with endogenous promoters showed reduced in vivo transgene expression and immunogenicity. Finally, in a related attempt to enhance MVA immunogenicity, deleting multiple immunomodulatory genes from the MVA genome did not affect MVA cellular immunogenicity in a range of vaccination regimens, testing either the pb9 epitope or the M. tuberculosis antigen 85A.

Authors: Alharbi, N

• Publication date: 2014
• Type of award: DPhil
• Level of award: Doctoral
• Awarding institution: Oxford University, UK
• Language: English
• Keywords: immunogenicity; Modified Vaccinia virus Ankara; poxvirus; transgene expression; vaccine; viral vector; promoter
• Subjects: Immunology; Viruses; Infectious diseases; Vaccinology