Partitioning and inactivation of viruses by the caprylic acid precipitation followed by a terminal pasteurization in the manufacturing process of horse immunoglobulins

Caprylic acid (octanoic acid), has been used for over 50 years as a stabilizer of human albumin during pasteurization. In addition caprylic acid is of great interest, by providing the advantage of purifying mammalian immunoglobulins and clearing viruses infectivity in a single step. Exploiting these two properties, we sequentially used the caprylic acid precipitation and the pasteurization to purify horse hyperimmune globulins used in the manufacturing of Sérocytol. To evaluate the effectiveness of the process for the removal/inactivation of viruses, spiking studies were carried out for each dedicated step. Bovine viral diarrhoea virus (BVDV), pseudorabies virus (PRV), encephalomyocarditis virus (EMCV) and minute virus of mice (MVM) were used for the virological validation. Our data show that the treatment with caprylic acid 5% (v/v) can effectively be used as well to purify or to ensure viral safety of immunoglobulins. Caprylic acid precipitation was very efficient in removing and/or inactivating enveloped viruses (PRV, BVDV) and moderately efficient against non-enveloped viruses (MVM, ECMV). However the combination with the pasteurization ensured an efficient protection against both enveloped and non-enveloped viruses. So that viruses surviving to the caprylic acid precipitation will be neutralized by pasteurization. Significant log reduction were achieved > or =9 log(10) for enveloped viruses and 4 log(10) for non-enveloped viruses, providing the evidence of a margin of viral safety achieved by our manufacturing process. Its a simple and non-expensive manufacturing process of immunoglobulins easily validated that we have adapted to a large production scale with a programmable operating system.     

Public vaccine manufacturing capacity in the Latin American and Caribbean region: Current status and perspectives

The vaccine global market is currently growing at a rate of 16.52%. Nowadays the vaccine manufacturing industry is limited in the sense that not all vaccine manufacturers have the capacity to execute all the steps necessary to produce a successful product. The biological variation inherent to vaccine manufacturing and the initial investment required to bring a vaccine to the market are some of the factors that discourage vaccine manufacturing initiatives.Given the current global context in vaccine innovation and production, and the increasing participation of vaccine manufacturers from developing countries in global markets, this paper aims to review vaccine manufacturing capacity in Latin American and Caribbean (LAC) countries with specific focus on trends in national or public sector manufacturing, presenting current challenges and future opportunities for the sector in meeting national and regional (LAC) needs.Despite the overall low vaccine manufacturing capacity reported within the LAC region within this paper, it is considered that the relatively high and concentrated capacity that exists within a number of countries, combined with political commitment of all countries within the Region, can provide the necessary platform for the continued development of capacity in vaccine development and manufacture within LAC.     

某私营木制家具制造企业2004 年与2010 年职业病危害因素检测 与评价报告比较和分析

目的:通过比较2个年份职业病危害因素检测与评价报告,评估职业病防治工作的效果。方法:选取两份有代表性的报告,比较和分析职业病危害防护设施的安装、个人防护用品发放与使用、警示标示以及各项检测结果的合格率。结果:该企业2010年各项调查指标相比2004年明显提高。结论:该企业各项职业病防治工作效果显著,劳动环境明显改善,但仍需注意噪声、木粉尘的危害。     

mRNA stability and antibody production in CHO cells: Improvement through gene optimization

The productivity of stably transfected cell lines is of critical importance for the manufacturing of therapeutic proteins. Various methods have been successfully implemented to increase the production output of mammalian cell cultures. Increasing evidence suggests that optimization of the gene coding sequences of an expression vector can improve specific cell line yield of the recombinant protein. Here we demonstrate that gene optimization substantially enhances antibody production in Chinese hamster ovary cells. When gene optimization was applied to the heavy and light chain genes of a therapeutic antibody, we observed increased antibody production in transient transfection. Elevated heavy chain mRNA level was associated with the increase of antibody production. Further analysis suggested that the increased antibody expression is attributable to enhanced mRNA stability resulting from gene optimization. Gene optimization also led to increased antibody production in stable clones.     

Cryopreservation timing is a critical process parameter in a thymic regulatory T-cell therapy manufacturing protocol

Regulatory T cells (Tregs) are a promising therapy for several immune-mediated conditions but manufacturing a homogeneous and consistent product, especially one that includes cryopreservation, has been challenging. Discarded pediatric thymuses are an excellent source of therapeutic Tregs with advantages including cell quantity, homogeneity and stability. Here we report systematic testing of activation reagents, cell culture media, restimulation timing and cryopreservation to develop a Good Manufacturing Practice (GMP)–compatible method to expand and cryopreserve Tregs. By comparing activation reagents, including soluble antibody tetramers, antibody-conjugated beads and artificial antigen-presenting cells (aAPCs) and different media, we found that the combination of Dynabeads Treg Xpander and ImmunoCult-XF medium preserved FOXP3 expression and suppressive function and resulted in expansion that was comparable with a single stimulation with aAPCs. Cryopreservation tests revealed a critical timing effect: only cells cryopreserved 1–3 days, but not >3 days, after restimulation maintained high viability and FOXP3 expression upon thawing. Restimulation timing was a less critical process parameter than the time between restimulation and cryopreservation. This systematic testing of key variables provides increased certainty regarding methods for in vitro expansion and cryopreservation of Tregs. The ability to cryopreserve expanded Tregs will have broad-ranging applications including enabling centralized manufacturing and long-term storage of cell products.     

Generation of Zika virus–specific T cells from seropositive and virus-naïve donors for potential use as an autologous or “off-the-shelf” immunotherapeutic

BackgroundZika virus (ZIKV) infection can cause severe birth defects in newborns with no effective currently available treatment. Adoptive transfer of virus-specific T cells has proven to be safe and effective for the prevention or treatment of many viral infections, and could represent a novel treatment approach for patients with ZIKV infection. However, extending this strategy to the ZIKV setting has been hampered by limited data on immunogenic T-cell antigens within ZIKV. Hence, we have generated ZIKV-specific T cells and characterized the cellular immune responses against ZIKV antigens.MethodsT-cell products were generated from peripheral blood of ZIKV-exposed donors, ZIKV-naive adult donors and umbilical cord blood by stimulation with pentadecamer (15mer) overlapping peptide libraries spanning four ZIKV polyproteins (C, M, E and NS1) using a Good Manufacturing Practice–compliant protocol.ResultsWe successfully generated T cells targeting ZIKV antigens with clinically relevant numbers. The ex vivo–expanded T cells comprised both CD4⁺ and CD8⁺ T cells that were able to produce Th1-polarized effector cytokines and kill ZIKV-infected HLA-matched monocytes, confirming functionality of this unique T-cell product as a potential “off-the-shelf” therapeutic. Epitope mapping using peptide arrays identified several novel HLA class I and class II–restricted epitopes within NS1 antigen, which is essential for viral replication and immune evasion.DiscussionOur findings demonstrate that it is feasible to generate potent ZIKV-specific T cells from a variety of cell sources including virus naïve donors for future clinical use in an “off-the-shelf” setting.     

Immobilized hematopoietic growth factors onto magnetic particles offer a scalable strategy for cell therapy manufacturing in suspension cultures

Hematopoietic therapies require high cell dosages and precise phenotype control for clinical success; scalable manufacturing processes therefore need to be economic and controllable, in particular with respect to culture medium and growth factor (GF) strategy. The aim of this work was to demonstrate the biological function, and integration within scalable systems, of a highly controllable immobilized growth factor (iGF) approach. GFs were biotinylated and attached to streptavidin coated magnetic particles. GF concentration during biotinylation, GF‐biotin ratio, and GF lysine content were shown to control iGF surface concentration and enable predictable co‐presentation of multiple GF on a single bead. Function was demonstrated for immobilized GMCSF, SCF, TPO and IL‐3 in GF dependent cell lines TF‐1 and M‐07e. Immobilized GMCSF (iGMCSF) was analyzed to show sustained activity over 8 days of culture, a 2–3 order of magnitude potency increase relative to soluble factor, and retained functionality under agitation in a micro‐scale stirred tank bioreactor. Further, short exposure to iGMCSF demonstrated prolonged growth response relative to soluble factor. This immobilization approach has the potential to reduce the manufacturing costs of scaled cell therapy products by reducing GF quantities and offers important process control opportunities through separation of GF treatments from the bulk media.     

Cell and gene therapy manufacturing capabilities in Australia and New Zealand

Cell and gene therapy products are rapidly being integrated into mainstream medicine. Developing global capability will facilitate broad access to these novel therapeutics. An initial step toward achieving this goal is to understand cell and gene therapy manufacturing capability in each region. We conducted an academic survey in 2018 to assess cell and gene therapy manufacturing capacity in Australia and New Zealand. We examined the following: the number and types of cell therapy manufacturing facilities; the number of projects, parallel processes and clinical trials; the types of products; and the manufacturing and quality staffing levels. It was found that Australia and New Zealand provide diverse facilities for cell therapy manufacturing, infrastructure and capability. Further investment and development will enable both countries to make important decisions to meet the growing need for cell and gene therapy and regenerative medicine in the region.     

Les statuettes féminines en ivoire des faciès gravettiens et post-gravettiens en Europe centrale et orientale : modes de fabrication et de représentation

During the Upper Palaeolithic, especially in Gravettian times, the hunter-gatherer societies had an economy closely linked with the exploitation of two local species in Eastern Europe: reindeer and woolly mammoth. The ivory objects are rich archives about their ways of life and their collective imagination, as in particular the ivory female statuettes show. These figurines, also called “Venus”, are one of the cultural characteristics of the Gravettian sites. To date, although they are more numerous in Eastern Europe, they were discovered, in a variable number, also in site of Central and Western Europe; today, we have no clue that this cultural tradition crossed the Pyrenees. The corpus of pieces from the Gravettian sites of the Russian Plain (dated between 25,000 and 21,000 B.P.) is the more informative about technological know-how of the Gravettian craftsmen. He consists in the leading material of this paper, completed with some data about Epigravettian and Magdalenian statuettes. Whereas the Gravettian figurines show figurative female representations, those of the later cultural facies are more stylized. In a technological point of view, there is a close link between the choice of blanks within the mammoth tusk and the morphology of the statuettes, whatever the period of time considered.     

The spatial correlation and interaction between manufacturing agglomeration and environmental pollution

Using statistical data from 285 cities in China, this paper studies the spatial correlation and interaction between manufacturing agglomeration and environmental pollution. Using a widely used spatial correlation index, bivariate Moran's I, we first estimate the spatial correlation between manufacturing agglomeration and environmental pollution. We show that there is significant spatial correlation between them, and distinct patterns of local spatial concentration are identified. Then, we use a spatial simultaneous equations (SSE) model to analyze the interaction between manufacturing agglomeration and environmental pollution. We show that manufacturing agglomeration aggravates environmental pollution, while environmental pollution restrains manufacturing agglomeration. In addition, manufacturing agglomeration and environmental pollution in any one city can be affected by manufacturing agglomeration and environmental pollution in surrounding cities through spatial spillover. Finally, we put forward specific suggestions based on the conclusions for more sustainable development.