Maximizing mRNA vaccine production with Bayesian optimization

Messenger RNA (mRNA) vaccines are a new alternative to conventional vaccines with a prominent role in infectious disease control. These vaccines are produced in in vitro transcription (IVT) reactions, catalyzed by RNA polymerase in cascade reactions. To ensure an efficient and cost-effective manufacturing process, essential for a large-scale production and effective vaccine supply chain, the IVT reaction needs to be optimized. IVT is a complex reaction that contains a large number of variables that can affect its outcome. Traditional optimization methods rely on classic Design of Experiments methods, which are time-consuming and can present human bias or based on simplified assumptions. In this contribution, we propose the use of Machine Learning approaches to perform a data-driven optimization of an mRNA IVT reaction. A Bayesian optimization method and model interpretability techniques were used to automate experiment design, providing a feedback loop. IVT reaction conditions were found under 60 optimization runs that produced 12 g · L−1 in solely 2 h. The results obtained outperform published industry standards and data reported in literature in terms of both achievable reaction yield and reduction of production time. Furthermore, this shows the potential of Bayesian optimization as a cost-effective optimization tool within (bio)chemical applications

Purification of therapeutic & prophylactic mRNA by affinity chromatography

In vitro transcribed mRNA is an emerging therapeutic and prophylactic modality with the potential to transform medicine. The drug platform features exceptionally rapid development and versatility of manufacturing processes. Despite the prompt advancement of mRNA from trials to market, purification challenges remain. The cell-free synthesis of mRNA is responsible for the generation of product and process-related impurities, creating the potential for immunogenic effects and decreased translatability into the clinic. Affinity chromatography presents itself as an effective primary capture step for the isolation of functional transcripts from product and some process related impurities. Developing platform processes for the affinity purification of mRNA is hindered by the varying strand lengths of non-amplifying, self-amplifying, and trans-amplifying constructs, with disparities in capacity being observed. Ligand chemistries may contribute to non-specific binding events which remain challenging to characterise. Improved elution and wash conditions may be pursued through novel ligand chemistries, enhanced density and spacing. Regardless of the size or application of the product, the impurities generated by in vitro transcription represent a significant obstacle to the safe administration and long-term storage of mRNA. Affinity chromatography is a valuable tool in overcoming these challenges, with current commercially available products relying heavily on oligo deoxythymidine ligand chemistries. Whilst affinity chromatography is highly valuable in the purification of mRNA, the inability to separate key secondary structures such as double-stranded RNA means it remains to be seen if this technology will adopt the same position as protein A does in mAb manufacture

Single-dose administration and the influence of the timing of the booster dose on immunogenicity and efficacy of ChAdOx1 nCoV-19 (AZD1222) vaccine: a pooled analysis of four randomised trials.

BACKGROUND: The ChAdOx1 nCoV-19 (AZD1222) vaccine has been approved for emergency use by the UK regulatory authority, Medicines and Healthcare products Regulatory Agency, with a regimen of two standard doses given with an interval of 4-12 weeks. The planned roll-out in the UK will involve vaccinating people in high-risk categories with their first dose immediately, and delivering the second dose 12 weeks later. Here, we provide both a further prespecified pooled analysis of trials of ChAdOx1 nCoV-19 and exploratory analyses of the impact on immunogenicity and efficacy of extending the interval between priming and booster doses. In addition, we show the immunogenicity and protection afforded by the first dose, before a booster dose has been offered. METHODS: We present data from three single-blind randomised controlled trials-one phase 1/2 study in the UK (COV001), one phase 2/3 study in the UK (COV002), and a phase 3 study in Brazil (COV003)-and one double-blind phase 1/2 study in South Africa (COV005). As previously described, individuals 18 years and older were randomly assigned 1:1 to receive two standard doses of ChAdOx1 nCoV-19 (5 × 1010 viral particles) or a control vaccine or saline placebo. In the UK trial, a subset of participants received a lower dose (2·2 × 1010 viral particles) of the ChAdOx1 nCoV-19 for the first dose. The primary outcome was virologically confirmed symptomatic COVID-19 disease, defined as a nucleic acid amplification test (NAAT)-positive swab combined with at least one qualifying symptom (fever ≥37·8°C, cough, shortness of breath, or anosmia or ageusia) more than 14 days after the second dose. Secondary efficacy analyses included cases occuring at least 22 days after the first dose. Antibody responses measured by immunoassay and by pseudovirus neutralisation were exploratory outcomes. All cases of COVID-19 with a NAAT-positive swab were adjudicated for inclusion in the analysis by a masked independent endpoint review committee. The primary analysis included all participants who were SARS-CoV-2 N protein seronegative at baseline, had had at least 14 days of follow-up after the second dose, and had no evidence of previous SARS-CoV-2 infection from NAAT swabs. Safety was assessed in all participants who received at least one dose. The four trials are registered at ISRCTN89951424 (COV003) and ClinicalTrials.gov, NCT04324606 (COV001), NCT04400838 (COV002), and NCT04444674 (COV005). FINDINGS: Between April 23 and Dec 6, 2020, 24 422 participants were recruited and vaccinated across the four studies, of whom 17 178 were included in the primary analysis (8597 receiving ChAdOx1 nCoV-19 and 8581 receiving control vaccine). The data cutoff for these analyses was Dec 7, 2020. 332 NAAT-positive infections met the primary endpoint of symptomatic infection more than 14 days after the second dose. Overall vaccine efficacy more than 14 days after the second dose was 66·7% (95% CI 57·4-74·0), with 84 (1·0%) cases in the 8597 participants in the ChAdOx1 nCoV-19 group and 248 (2·9%) in the 8581 participants in the control group. There were no hospital admissions for COVID-19 in the ChAdOx1 nCoV-19 group after the initial 21-day exclusion period, and 15 in the control group. 108 (0·9%) of 12 282 participants in the ChAdOx1 nCoV-19 group and 127 (1·1%) of 11 962 participants in the control group had serious adverse events. There were seven deaths considered unrelated to vaccination (two in the ChAdOx1 nCov-19 group and five in the control group), including one COVID-19-related death in one participant in the control group. Exploratory analyses showed that vaccine efficacy after a single standard dose of vaccine from day 22 to day 90 after vaccination was 76·0% (59·3-85·9). Our modelling analysis indicated that protection did not wane during this initial 3-month period. Similarly, antibody levels were maintained during this period with minimal waning by day 90 (geometric mean ratio [GMR] 0·66 [95% CI 0·59-0·74]). In the participants who received two standard doses, after the second dose, efficacy was higher in those with a longer prime-boost interval (vaccine efficacy 81·3% [95% CI 60·3-91·2] at ≥12 weeks) than in those with a short interval (vaccine efficacy 55·1% [33·0-69·9] at <6 weeks). These observations are supported by immunogenicity data that showed binding antibody responses more than two-fold higher after an interval of 12 or more weeks compared with an interval of less than 6 weeks in those who were aged 18-55 years (GMR 2·32 [2·01-2·68]). INTERPRETATION: The results of this primary analysis of two doses of ChAdOx1 nCoV-19 were consistent with those seen in the interim analysis of the trials and confirm that the vaccine is efficacious, with results varying by dose interval in exploratory analyses. A 3-month dose interval might have advantages over a programme with a short dose interval for roll-out of a pandemic vaccine to protect the largest number of individuals in the population as early as possible when supplies are scarce, while also improving protection after receiving a second dose. FUNDING: UK Research and Innovation, National Institutes of Health Research (NIHR), The Coalition for Epidemic Preparedness Innovations, the Bill & Melinda Gates Foundation, the Lemann Foundation, Rede D'Or, the Brava and Telles Foundation, NIHR Oxford Biomedical Research Centre, Thames Valley and South Midland's NIHR Clinical Research Network, and AstraZeneca

Microfluidic devices towards personalized health and wellbeing

To make personalised medicine a reality, there is a significant need for cost‐effective methods that enable the rapid selection of optimal nutrient intake and/or disease treatment with a minimum of side effects. In this perspective, we briefly discuss the potential of merging the advances of microfluidic devices, organoid structures and advanced analytical methods to progress towards a personalised in vitro platform for health and wellbeing

Aspectos metodologicos do Projeto SBBrasil 2010 de interesse para inqueritos nacionais de saude

Abstract published in English and Portuguese English title: Relevant methodological issues from the SSBrasil 2010 Project for national health surveysThe SBBrasil 2010 Project (SBB10) was designed as a nationwide oral health epidemiological survey within a health surveillance strategy. This article discusses methodological aspects of the SBB10 Project that can potentially help expand and develop knowledge in the health field. This was a nationwide survey with stratified multi-stage cluster sampling. The sample domains were 27 State capitals and 150 rural municipalities (counties) from the country's five major geographic regions. The sampling units were census tracts and households for the State capitals and municipalities, census tracts, and households for the rural areas. Thirty census tracts were selected in the State capitals and 30 municipalities in the countryside. The precision considered the demographic domains grouped by density of the overall population and the internal variability of oral health indices. The study evaluated dental caries, periodontal disease, malocclusion, fluorosis, tooth loss, and dental trauma in five age groups (5, 12, 15-19, 35-44, and 65-74 years). = O Projeto SBBrasil 2010 (SBB10) foi concebido como um levantamento epidemiológico em saúde bucal, de base nacional, dentro da estratégia de vigilância em saúde. O objetivo deste artigo é apresentar aspectos da metodologia do SBB10 que possam contribuir para ampliar e desenvolver conhecimentos na área de saúde. Com relação ao plano amostral, trata-se de uma pesquisa por conglomerados e com múltiplos estágios. Capitais e municípios do interior das cinco regiões brasileiras compõem os domínios da amostra, cujas unidades amostrais foram, respectivamente, setor censitário e domicílio para as capitais, e município, setor censitário e domicílio para o interior. Nas capitais foram sorteados 30 setores e, no interior de cada região, 30 municípios. A precisão considerou os domínios agrupados segundo o grau de densidade no total da população e a variabilidade interna dos índices. Foram avaliadas as condições de cárie dentária, doença periodontal, oclusopatias, fluorose, traumatismo dentário e edentulismo em cinco grupos etários (5, 12, 15 a 19, 35 a 44 e 65 a 74 anos).Angelo Giuseppe Roncalli, Nilza Nunes da Silva, Antonio Carlos Nascimento, Cláudia Helena Soares de Morais Freitas, Elisete Casotti, Karen Glazer Peres, Lenildo de Moura, Marco A. Peres, Maria do Carmo Matias Freire, Maria Ilma de Souza Cortes, Mario Vianna Vettore, Moacir Paludetto Júnior, Nilcema Figueiredo, Paulo Sávio Angeiras de Goes, Rafaela da Silveira Pinto, Regina Auxiliadora de Amorim Marques, Samuel Jorge Moysés, Sandra Cristina Guimarães Bahia Reis, Paulo Capel Narva

Large heterogeneities in comet 67P as revealed by active pits from sinkhole collapse

Pits have been observed on many cometary nuclei mapped by spacecraft1,2,3,4. It has been argued that cometary pits are a signature of endogenic activity, rather than impact craters such as those on planetary and asteroid surfaces. Impact experiments5,6 and models7,8 cannot reproduce the shapes of most of the observed cometary pits, and the predicted collision rates imply that few of the pits are related to impacts8,9. Alternative mechanisms like explosive activity10 have been suggested, but the driving process remains unknown. Here we report that pits on comet 67P/Churyumov–Gerasimenko are active, and probably created by a sinkhole process, possibly accompanied by outbursts. We argue that after formation, pits expand slowly in diameter, owing to sublimation-driven retreat of the walls. Therefore, pits characterize how eroded the surface is: a fresh cometary surface will have a ragged structure with many pits, while an evolved surface will look smoother. The size and spatial distribution of pits imply that large heterogeneities exist in the physical, structural or compositional properties of the first few hundred metres below the current nucleus surface