Hazard Characterization of Modified Vaccinia Virus Ankara Vector: What Are the Knowledge Gaps?

Okeke, Malachy I.; Okoli, Arinze S.; Diaz, Diana; Offor, Collins; Oludotun, Taiwo G.; Tryland, Morten; Bøhn, Thomas; Moens, Ugo

A robust hazard characterization of a virus vector will demonstrate that the vector is clonally pure and genetically stable at a passage level and titer that is equivalent to what is present in the production batch. The MVA vector is assumed to be homogenous following over 570 passages in CEF and subsequent three rounds of plaque purification [100] . The identical genome sequence of independent MVA strains from at least five independent laboratories is presented as evidence of MVA clonal purity and genome stability [42] . However, the next generation sequencing approach used in sequencing those genomes only reported the genome of the predominant MVA clone or variant, since it was not designed to determine if minor variants are present. A demonstration of the clonal purity of MVA will require deep sequencing of the stock or master seed virus (MSV) in tandem with sequencing of several MVA clones obtained from the stock following at least three rounds of plaque purification. While Dryvax ® vaccine has been shown to be polyclonal using genome wide sequencing designed to examine if variants are present [101] , isolation and genome wide mapping of variants within the MVA stock or MSV have never been reported. One study employing passage of MVA strains in immune-deficient mice has shown that the examined MVA strains with the exception of MVA-BN are polyclonal [56] . However, the observation from that study showing MVA-BN to be homogenous can only be confirmed if MVA-BN is subjected to deep sequencing in combination with genome sequencing of independent clones isolated from MVA-BN.