Summary Background Use of oral live-attenuated polio vaccines (OPV), and injected inactivated polio vaccines (IPV) has almost achieved global eradication of wild polio viruses. To address the goals of achieving and maintaining global eradication and minimising the risk of outbreaks of vaccine-derived polioviruses, we tested novel monovalent oral type-2 poliovirus (OPV2) vaccine candidates that are genetically more stable than existing OPVs, with a lower risk of reversion to neurovirulence. Our study represents the first in-human testing of these two novel OPV2 candidates. We aimed to evaluate the safety and immunogenicity of these vaccines, the presence and extent of faecal shedding, and the neurovirulence of shed virus. Methods In this double-blind, single-centre phase 1 trial, we isolated participants in a purpose-built containment facility at the University of Antwerp Hospital (Antwerp, Belgium), to minimise the risk of environmental release of the novel OPV2 candidates. Participants, who were recruited by local advertising, were adults (aged 18–50 years) in good health who had previously been vaccinated with IPV, and who would not have any contact with immunosuppressed or unvaccinated people for the duration of faecal shedding at the end of the study. The first participant randomly chose an envelope containing the name of a vaccine candidate, and this determined their allocation; the next 14 participants to be enrolled in the study were sequentially allocated to this group and received the same vaccine. The subsequent 15 participants enrolled after this group were allocated to receive the other vaccine. Participants and the study staff were masked to vaccine groups until the end of the study period. Participants each received a single dose of one vaccine candidate (candidate 1, S2/cre5/S15domV/rec1/hifi3; or candidate 2, S2/S15domV/CpG40), and they were monitored for adverse events, immune responses, and faecal shedding of the vaccine virus for 28 days. Shed virus isolates were tested for the genetic stability of attenuation. The primary outcomes were the incidence and type of serious and severe adverse events, the proportion of participants showing viral shedding in their stools, the time to cessation of viral shedding, the cell culture infective dose of shed virus in virus-positive stools, and a combined index of the prevalence, duration, and quantity of viral shedding in all participants. This study is registered with EudraCT, number 2017-000908-21 and ClinicalTrials.gov , number NCT03430349 . Findings Between May 22 and Aug 22, 2017, 48 volunteers were screened, of whom 15 (31%) volunteers were excluded for reasons relating to the inclusion or exclusion criteria, three (6%) volunteers were not treated because of restrictions to the number of participants in each group, and 30 (63%) volunteers were sequentially allocated to groups (15 participants per group). Both no...

Summary The live-attenuated oral poliovirus vaccine (OPV or Sabin vaccine) replicates in gut-associated tissues, eliciting mucosa and systemic immunity. OPV protects from disease and limits poliovirus spread. Accordingly, vaccination with OPV is the primary strategy used to end the circulation of all polioviruses. However, the ability of OPV to regain replication fitness and establish new epidemics represents a significant risk of polio re-emergence should immunization cease. Here, we report the development of a poliovirus type 2 vaccine strain (nOPV2) that is genetically more stable and less likely to regain virulence than the original Sabin2 strain. We introduced modifications within at the 5′ untranslated region of the Sabin2 genome to stabilize attenuation determinants, 2C coding region to prevent recombination, and 3D polymerase to limit viral adaptability. Prior work established that nOPV2 is immunogenic in preclinical and clinical studies, and thus may enable complete poliovirus eradication.

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Efforts to increase cell growth and protein yields need to be complemented by the maintenance of the quality of the protein produced. Elevated oxygen pressure or rapid increases in oxygen content can cause oxidative stress within the cells, leading to oxidation of specific proteins and nucleotide sequences. In addition, transient or steady-state anoxic conditions can cause limitations in amino acid production and plasmid stability. Major pathways and mechanisms of oxidative damage to proteins expressed in bacteria are reviewed. Damage to nucleic acids involved in gene expression also is considered. The methodologies for identifying oxidative damage to macromolecules are improving but are not yet adequate for on-line feedback. This limits our ability to integrate information about these phenomena and the cellular responses into a quantitative model. Enough information is available, however, to consider changes in the time profile of dissolved oxygen as a cause for poor process performance.

uct characterization and release, production and purification processes that are scaleable and economical, and formulations enabling sufficient stability to guarantee an acceptable shelf-life. In this review, we highlight the many advances in these areas, and identify requirements for further research. In order to capture a comprehensive and current picture, we emphasize both the published and patent literature.We begin by discussing the common methods used for analysis of both purified and unpurified adenovirus (AdV) samples. These methods are critical to both process development and the release of GMP (good manufacturing practices) supplies to be used in clinical studies, and are referred to throughout the remainder of the review. Next, we discuss the options and rationale for vector design, and we describe the common complementing cell lines. Cultivation of AdV vectors including media selection, modes of operation (suspension vs. adherent, batch vs. fed-batch, and so on), and critical optimization and control parameters are then discussed, with an emphasis on current industrial practice. Adenovirus purification methodologies are addressed similarly, with a focus on the rational selection of unit operations for a scaleable process. Current industrial practices are compared, and critical issues such as the clearance of empty capsids and host cell DNA are reviewed in detail. Next, the development of remarkably stable liquid formulations is discussed, with an emphasis on excipient selection based on the mechanisms of inactivation. Finally, we briefly recap critical developments and highlight areas in need of additional research to support the next generation of adenovirus-based gene transfer products. 2Analytical methods for process and product characterization 2.1