Vaccine Production

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Udo Reichl - One of the best experts on this subject based on the ideXlab platform.

  • mathematical modeling as a tool to improve influenza Vaccine Production processes
    IFAC-PapersOnLine, 2018
    Co-Authors: Udo Reichl, Mandy Bachmann, Robert Dürr, Stefanie Duvigneau, Tanja Laske, Melanie Dostert, Achim Kienle
    Abstract:

    Abstract Cell culture-based Production of influenza Vaccines is emerging as a promising alternative to conventional Production in embryonated chicken eggs. Development and establishment of high-yield producer cell lines represents a major challenge to manufacture sufficient amounts of low-cost Vaccines. One possible option to optimize Vaccine Production is to manipulate the expression of host cell factors relevant for virus replication. Lentiviral transduction is a gene editing method that allows to modify the expression of single or multiple host cell genes. However, due to different copy numbers and integration sites of the gene constructs the expression level shows a large cell-to-cell variability within the cell population. In this study, we will investigate the impact of genetic modifications on virus yield with the help of a structured population balance model. Therein, cell-to-cell variability is represented in terms of distributed kinetic parameter sets obtained after bootstrapping for five cell lines overexpressing a single gene. Moreover, we evaluate four different strategies to predict distributed parameter sets for cell lines overexpressing multiple genes based on the parameter distributions of the underlying single gene modifications. Furthermore, we will apply the most suitable prediction strategy to find a combination of gene modifications that leads to the highest virus productivity.

  • Bioreactors for high cell density and continuous multi-stage cultivations: options for process intensification in cell culture-based viral Vaccine Production
    Applied Microbiology and Biotechnology, 2016
    Co-Authors: Freddy Tapia, Daniel Vázquez-ramírez, Felipe Tapia, Yvonne Genzel, Udo Reichl
    Abstract:

    With an increasing demand for efficacious, safe, and affordable Vaccines for human and animal use, process intensification in cell culture-based viral Vaccine Production demands advanced process strategies to overcome the limitations of conventional batch cultivations. However, the use of fed-batch, perfusion, or continuous modes to drive processes at high cell density (HCD) and overextended operating times has so far been little explored in large-scale viral Vaccine manufacturing. Also, possible reductions in cell-specific virus yields for HCD cultivations have been reported frequently. Taking into account that Vaccine Production is one of the most heavily regulated industries in the pharmaceutical sector with tough margins to meet, it is understandable that process intensification is being considered by both academia and industry as a next step toward more efficient viral Vaccine Production processes only recently. Compared to conventional batch processes, fed-batch and perfusion strategies could result in ten to a hundred times higher product yields. Both cultivation strategies can be implemented to achieve cell concentrations exceeding 10(7) cells/mL or even 10(8) cells/mL, while keeping low levels of metabolites that potentially inhibit cell growth and virus replication. The trend towards HCD processes is supported by development of GMP-compliant cultivation platforms, i.e., acoustic settlers, hollow fiber bioreactors, and hollow fiber-based perfusion systems including tangential flow filtration (TFF) or alternating tangential flow (ATF) technologies. In this review, these process modes are discussed in detail and compared with conventional batch processes based on productivity indicators such as space-time yield, cell concentration, and product titers. In addition, options for the Production of viral Vaccines in continuous multi-stage bioreactors such as two- and three-stage systems are addressed. While such systems have shown similar virus titers compared to batch cultivations, keeping high yields for extended Production times is still a challenge. Overall, we demonstrate that process intensification of cell culture-based viral Vaccine Production can be realized by the consequent application of fed-batch, perfusion, and continuous systems with a significant increase in productivity. The potential for even further improvements is high, considering recent developments in establishment of new (designer) cell lines, better characterization of host cell metabolism, advances in media design, and the use of mathematical models as a tool for process optimization and control.

  • impact of defective interfering particles on virus replication and antiviral host response in cell culture based influenza Vaccine Production
    Applied Microbiology and Biotechnology, 2014
    Co-Authors: Timo Frensing, Udo Reichl, Antje Pflugmacher, Mandy Bachmann, Britta Peschel
    Abstract:

    During the replication of influenza viruses, defective interfering particles (DIPs) can be generated. These are noninfectious deletion mutants that require coinfection with a wild-type virus but interfere with its helper virus replication. Consequently, coinfected cells mainly produce DIPs. Little is known about how such noninfectious virus particles affect the virus yield of cell culture-based influenza Vaccine Production. We compared infections of Madin-Darby canine kidney cells with two seed virus preparations of the influenza virus strain A/Puerto Rico/8/34 that contain different amounts of DIPs. A combination of conventional RT-PCR, RT-qPCR, and flow cytometry revealed that DI genomes indeed strongly accumulate in coinfected cells and impede the viral RNA synthesis. Additionally, cells infected at the higher DIP concentration showed a stronger antiviral response characterized by increased interferon-β expression and apoptosis induction. Furthermore, in the presence of DIPs, a significant fraction of cells did not show any productive accumulation of viral proteins at all. Together, these effects of DIPs significantly reduce the virus yield. Therefore, the accumulation of DIPs should be avoided during influenza Vaccine Production which can be achieved by quality controls of working seed viruses based on conventional RT-PCR. The strategy for the depletion of DIPs presented here can help to make cell culture-based Vaccine Production more reliable and robust.

  • Distributed modeling of human influenza A virus-host cell interactions during Vaccine Production.
    Biotechnology and bioengineering, 2013
    Co-Authors: Thomas Müller, Udo Reichl, J. Schulze-horsel, Robert Dürr, Britta Isken, Achim Kienle
    Abstract:

    This contribution is concerned with population balance modeling of virus–host cell interactions during Vaccine Production. Replication of human influenza A virus in cultures of adherent Madin–Darby canine kidney (MDCK) cells is considered as a model system. The progress of infection can be characterized by the intracellular amount of viral nucleoprotein (NP) which is measured via flow cytometry. This allows the differentiation of the host cell population and gives rise to a distributed modeling approach. For this purpose a degree of fluorescence is introduced as an internal coordinate which is linearly linked to the intracellular amount of NP. Experimental results for different human influenza A subtypes reveal characteristic dynamic phenomena of the cell distribution like transient multimodality and reversal of propagation direction. The presented population balance model provides a reasonable explanation for these dynamic phenomena by the explicit consideration of different states of infection of individual cells. Kinetic parameters are determined from experimental data. To translate the emerging infinite dimensional parameter estimation problem to a finite dimension the parameters are assumed to depend linearly on the internal coordinate. As a result, the model is able to reproduce all characteristic dynamic phenomena of the considered process for the two examined virus strains and allows deeper insight into the underlying kinetic processes. Thus, the model is an important contribution to the understanding of the intracellular virus replication and virus spreading in cell cultures and can serve as a stepping stone for optimization in Vaccine Production. Biotechnol. Bioeng. 2013; 110: 2252–2266. © 2013 Wiley Periodicals, Inc.

  • Trypsin promotes efficient influenza Vaccine Production in MDCK cells by interfering with the antiviral host response
    Applied Microbiology and Biotechnology, 2012
    Co-Authors: Claudius Seitz, Timo Frensing, Britta Isken, Björn Heynisch, Maria Rettkowski, Udo Reichl
    Abstract:

    Trypsin is commonly used in Madin–Darby canine kidney (MDCK) cell culture-based influenza Vaccine Production to facilitate virus infection by proteolytic activation of viral haemagglutinin, which enables multi-cycle replication. In this study, we were able to demonstrate that trypsin also interferes with pathogen defence mechanisms of host cells. In particular, a trypsin concentration of 5 BAEE U/mL (4.5 μg/mL porcine trypsin) used in Vaccine manufacturing strongly inhibited interferon (IFN) signalling by proteolytic degradation of secreted IFN. Consequently, absence of trypsin during infection resulted in a considerably stronger induction of IFN signalling and apoptosis, which significantly reduced virus yields. Under this condition, multi-cycle virus replication in MDCK cells was not prevented but clearly delayed. Therefore, incomplete infection can be ruled out as the reason for the lower virus titres. However, suppression of IFN signalling by overexpression of viral IFN antagonists (influenza virus PR8-NS1, rabies virus phosphoprotein) partially rescued virus titres in the absence of trypsin. In addition, virus yields could be almost restored by using the influenza strain A/WSN/33 in combination with fetal calf serum (FCS). For this strain, FCS enabled trypsin-independent fast propagation of virus infection, probably outrunning cellular defence mechanisms and apoptosis induction in the absence of trypsin. Overall, addition of trypsin provided optimal conditions for high yield Vaccine Production in MDCK cells by two means. On the one hand, proteolytic degradation of IFN keeps cellular defence at a low level. On the other hand, enhanced virus spreading enables viruses to replicate before the cellular response becomes fully activated.

Yoshihiro Kawaoka - One of the best experts on this subject based on the ideXlab platform.

  • establishment of canine rna polymerase i driven reverse genetics for influenza a virus its application for h5n1 Vaccine Production
    Journal of Virology, 2008
    Co-Authors: Shin Murakami, Taisuke Horimoto, Shinya Yamada, Satoshi Kakugawa, Hideo Goto, Yoshihiro Kawaoka
    Abstract:

    In the event of a new influenza pandemic, Vaccines whose antigenicities match those of circulating strains must be rapidly produced. Here, we established an alternative reverse genetics system for influenza virus using the canine polymerase I (PolI) promoter sequence that works efficiently in the Madin-Darby canine kidney cell line, a cell line approved for human Vaccine Production. Using this system, we were able to generate H5N1 Vaccine seed viruses more efficiently than can be achieved with the current system that uses the human PolI promoter in African green monkey Vero cells, thus improving pandemic Vaccine Production.

  • An improved reverse genetics system for influenza A virus generation and its implications for Vaccine Production
    Proceedings of the National Academy of Sciences of the United States of America, 2005
    Co-Authors: Gabriele Neumann, Ken Fujii, Yoichiro Kino, Yoshihiro Kawaoka
    Abstract:

    The generation of Vaccines for highly pathogenic avian influenza viruses, including those of the H5N1 subtype, relies on reverse genetics, which allows the Production of influenza viruses from cloned cDNA. In the future, reverse genetics will likely be the method of choice for the generation of conventional influenza Vaccine strains because gene reassortment by more traditional methods is cumbersome. Established systems for the artificial generation of influenza A viruses require transfection of cells with the eight to 12 plasmids that provide the eight influenza viral RNAs as well as the polymerase and nucleoproteins of the virus. However, cell lines appropriate for human Vaccine Production (e.g., Vero cells) cannot be transfected with high efficiencies. To overcome these problems, we established a reverse genetics system in which the eight RNA polymerase I transcription cassettes for viral RNA synthesis are combined on one plasmid. Similarly, two cassettes encoding the hemagglutinin and neuraminidase segments and six cassettes encoding the remaining proteins were combined. We also combined three RNA polymerase II transcription cassettes for the expression of the polymerase subunits. By combining these cassettes, we reduced the number of plasmids required for virus generation significantly and produced influenza A virus in Vero cells with higher efficiency than with the traditional 12 plasmid system. This new system is thus suitable for influenza virus Vaccine Production and may be applicable to other reverse genetics systems that rely on the introduction of several plasmids into eukaryotic cells.

Wilfried A.m. Bakker - One of the best experts on this subject based on the ideXlab platform.

  • Scale-down of the inactivated polio Vaccine Production process
    Biotechnology and Bioengineering, 2013
    Co-Authors: Yvonne E. Thomassen, Aart G. Van 't Oever, Marian Vinke, Arjen Spiekstra, Leo A. Van Der Pol, René H. Wijffels, Wilfried A.m. Bakker
    Abstract:

    The anticipated increase in the demand for inactivated polio Vaccines resulting from the success in the polio eradication program requires an increase in Production capacity and cost price reduction of the current inactivated polio Vaccine Production processes. Improvement of existing Production processes is necessary as the initial process development has been done decades ago. An up-to-date lab-scale version encompassing the legacy inactivated polio Vaccine Production process was set-up. This lab-scale version should be representative of the large scale, meaning a scale-down model, to allow experiments for process optimization that can be readily applied. Initially the separate unit operations were scaled-down at setpoint. Subsequently, the unit operations were applied successively in a comparative manner to large-scale manufacturing. This allows the assessment of the effects of changes in one unit operation to the consecutive units at small-scale. Challenges in translating large-scale operations to lab-scale are discussed, and the concessions that needed to be made are described. The current scale-down model for cell and virus culture (2.3-L) presents a feasible model with its Production scale counterpart (750-L) when operated at setpoint. Also, the current scale-down models for the DSP unit operations clarification, concentration, size exclusion chromatography, ion exchange chromatography, and inactivation are in agreement with the manufacturing scale. The small-scale units can be used separately, as well as sequentially, to study variations and critical product quality attributes in the Production process. Finally, it is shown that the scale-down unit operations can be used consecutively to prepare trivalent Vaccine at lab-scale with comparable characteristics to the product produced at manufacturing scale.

  • Scale‐down of the inactivated polio Vaccine Production process
    Biotechnology and bioengineering, 2012
    Co-Authors: Yvonne E. Thomassen, Marian Vinke, Arjen Spiekstra, Leo A. Van Der Pol, René H. Wijffels, Aart G. Van ’t Oever, Wilfried A.m. Bakker
    Abstract:

    The anticipated increase in the demand for inactivated polio Vaccines resulting from the success in the polio eradication program requires an increase in Production capacity and cost price reduction of the current inactivated polio Vaccine Production processes. Improvement of existing Production processes is necessary as the initial process development has been done decades ago. An up-to-date lab-scale version encompassing the legacy inactivated polio Vaccine Production process was set-up. This lab-scale version should be representative of the large scale, meaning a scale-down model, to allow experiments for process optimization that can be readily applied. Initially the separate unit operations were scaled-down at setpoint. Subsequently, the unit operations were applied successively in a comparative manner to large-scale manufacturing. This allows the assessment of the effects of changes in one unit operation to the consecutive units at small-scale. Challenges in translating large-scale operations to lab-scale are discussed, and the concessions that needed to be made are described. The current scale-down model for cell and virus culture (2.3-L) presents a feasible model with its Production scale counterpart (750-L) when operated at setpoint. Also, the current scale-down models for the DSP unit operations clarification, concentration, size exclusion chromatography, ion exchange chromatography, and inactivation are in agreement with the manufacturing scale. The small-scale units can be used separately, as well as sequentially, to study variations and critical product quality attributes in the Production process. Finally, it is shown that the scale-down unit operations can be used consecutively to prepare trivalent Vaccine at lab-scale with comparable characteristics to the product produced at manufacturing scale.

  • Platform Technology for Viral Vaccine Production: Comparison Between Attached and Suspension Vero Cells
    Proceedings of the 21st Annual Meeting of the European Society for Animal Cell Technology (ESACT) Dublin Ireland June 7-10 2009, 2011
    Co-Authors: Yvonne E. Thomassen, Gerco Van Eikenhorst, Wilfried A.m. Bakker
    Abstract:

    Vero cells can be used as host for the Production of many different types of viruses. The standard Vero culturing process consists of growing Vero cells adherent to microcarriers. Due to surface limitation the cells will have to be transferred to new microcarriers regularly. This subculturing of the cells, especially at large scale, is a complicated technique. This drawback can be overcome by using single cell suspension cultures. Vero cells growing in single cell suspension (sVero) were compared to adherent growing Vero cells (aVero) for their capability of producing polio virus. Using flow cytometry it was determined that sVero cells contained the polio virus receptor and were able to produce polio virus. These results indicate that sVero cells may be an ideal candidate for a platform based viral Vaccine Production approach. The combination of sVero cells with the flexibility of disposable bioreactors completes their suitability for these purposes.

Yvonne E. Thomassen - One of the best experts on this subject based on the ideXlab platform.

  • Scale-down of the inactivated polio Vaccine Production process
    Biotechnology and Bioengineering, 2013
    Co-Authors: Yvonne E. Thomassen, Aart G. Van 't Oever, Marian Vinke, Arjen Spiekstra, Leo A. Van Der Pol, René H. Wijffels, Wilfried A.m. Bakker
    Abstract:

    The anticipated increase in the demand for inactivated polio Vaccines resulting from the success in the polio eradication program requires an increase in Production capacity and cost price reduction of the current inactivated polio Vaccine Production processes. Improvement of existing Production processes is necessary as the initial process development has been done decades ago. An up-to-date lab-scale version encompassing the legacy inactivated polio Vaccine Production process was set-up. This lab-scale version should be representative of the large scale, meaning a scale-down model, to allow experiments for process optimization that can be readily applied. Initially the separate unit operations were scaled-down at setpoint. Subsequently, the unit operations were applied successively in a comparative manner to large-scale manufacturing. This allows the assessment of the effects of changes in one unit operation to the consecutive units at small-scale. Challenges in translating large-scale operations to lab-scale are discussed, and the concessions that needed to be made are described. The current scale-down model for cell and virus culture (2.3-L) presents a feasible model with its Production scale counterpart (750-L) when operated at setpoint. Also, the current scale-down models for the DSP unit operations clarification, concentration, size exclusion chromatography, ion exchange chromatography, and inactivation are in agreement with the manufacturing scale. The small-scale units can be used separately, as well as sequentially, to study variations and critical product quality attributes in the Production process. Finally, it is shown that the scale-down unit operations can be used consecutively to prepare trivalent Vaccine at lab-scale with comparable characteristics to the product produced at manufacturing scale.

  • Scale‐down of the inactivated polio Vaccine Production process
    Biotechnology and bioengineering, 2012
    Co-Authors: Yvonne E. Thomassen, Marian Vinke, Arjen Spiekstra, Leo A. Van Der Pol, René H. Wijffels, Aart G. Van ’t Oever, Wilfried A.m. Bakker
    Abstract:

    The anticipated increase in the demand for inactivated polio Vaccines resulting from the success in the polio eradication program requires an increase in Production capacity and cost price reduction of the current inactivated polio Vaccine Production processes. Improvement of existing Production processes is necessary as the initial process development has been done decades ago. An up-to-date lab-scale version encompassing the legacy inactivated polio Vaccine Production process was set-up. This lab-scale version should be representative of the large scale, meaning a scale-down model, to allow experiments for process optimization that can be readily applied. Initially the separate unit operations were scaled-down at setpoint. Subsequently, the unit operations were applied successively in a comparative manner to large-scale manufacturing. This allows the assessment of the effects of changes in one unit operation to the consecutive units at small-scale. Challenges in translating large-scale operations to lab-scale are discussed, and the concessions that needed to be made are described. The current scale-down model for cell and virus culture (2.3-L) presents a feasible model with its Production scale counterpart (750-L) when operated at setpoint. Also, the current scale-down models for the DSP unit operations clarification, concentration, size exclusion chromatography, ion exchange chromatography, and inactivation are in agreement with the manufacturing scale. The small-scale units can be used separately, as well as sequentially, to study variations and critical product quality attributes in the Production process. Finally, it is shown that the scale-down unit operations can be used consecutively to prepare trivalent Vaccine at lab-scale with comparable characteristics to the product produced at manufacturing scale.

  • Platform Technology for Viral Vaccine Production: Comparison Between Attached and Suspension Vero Cells
    Proceedings of the 21st Annual Meeting of the European Society for Animal Cell Technology (ESACT) Dublin Ireland June 7-10 2009, 2011
    Co-Authors: Yvonne E. Thomassen, Gerco Van Eikenhorst, Wilfried A.m. Bakker
    Abstract:

    Vero cells can be used as host for the Production of many different types of viruses. The standard Vero culturing process consists of growing Vero cells adherent to microcarriers. Due to surface limitation the cells will have to be transferred to new microcarriers regularly. This subculturing of the cells, especially at large scale, is a complicated technique. This drawback can be overcome by using single cell suspension cultures. Vero cells growing in single cell suspension (sVero) were compared to adherent growing Vero cells (aVero) for their capability of producing polio virus. Using flow cytometry it was determined that sVero cells contained the polio virus receptor and were able to produce polio virus. These results indicate that sVero cells may be an ideal candidate for a platform based viral Vaccine Production approach. The combination of sVero cells with the flexibility of disposable bioreactors completes their suitability for these purposes.

Rainer Fischer - One of the best experts on this subject based on the ideXlab platform.

  • The Potential of Plant Virus Vectors for Vaccine Production
    Drugs in R & D, 2006
    Co-Authors: Vidadi Yusibov, Shailaja Rabindran, Ulrich Commandeur, Richard M. Twyman, Rainer Fischer
    Abstract:

    Plants viruses are versatile vectors that allow the rapid and convenient Production of recombinant proteins in plants. Compared with Production systems based on transgenic plants, viral vectors are easier to manipulate and recombinant proteins can be produced more quickly and in greater yields. Over the last few years, there has been much interest in the development of plant viruses as vectors for the Production of Vaccines, either as whole polypeptides or epitopes displayed on the surface of chimeric viral particles. Several viruses have been extensively developed for Vaccine Production, including tobacco mosaic virus, potato virus X and cowpea mosaic virus. Vaccine candidates have been produced against a range of human and animal diseases, and in many cases have shown immunogenic activity and protection in the face of disease challenge. In this review, we discuss the advantages of plant virus vectors, the development of different viruses as vector systems, and the immunological experiments that have demonstrated the principle of plant virus-derived Vaccines.