Since its beginnings in the 1970s, recombinant DNA technology has become integral in the production of pharmaceutically important proteins such as blood factors, hormones, growth factors, interferons, interleukins, and vaccines. Recombinant technology was developed in Escherichia coli, and because of its long history of use and ease of manipulation, the bacteria remains a common expression system. However, E. coli is limited in the types of proteins it can express, and production can be hampered by the formation of intracellular inclusion body aggregates which complicate the processes of protein recovery and purification. The non-pathogenic Corynebacterium glutamicum can accumulate amino acids extracellularly and has historically been used for the commercial production of amino acids glutamate and lysine. More recently, C. glutamicum has been developed as an expression system with the ability to secrete properly folded, functionally active proteins into the fermentation broth. Production in C. glutamicum has the same advantages of scalability and ease of culturing as in E. coli, but with more simplified recovery and purification processes…
Tag: <span>vaccines</span>
The impact of viruses—in geopolitical human health issues, in the production of vaccines and recombinant proteins, and in gene therapy and cancer treatments—highlights the need for a better understanding of the systems that are dependent upon them. A primary barrier to recognizing the full potential of these life-saving biomedical approaches is the scarcity of analytical methods capable of providing biologically relevant information without hindering the pace of development and production. ViroCyt® is a Colorado-based biotechnology company with one overriding focus: Enabling the rapid and specific quantification of viruses and virus-related particles. The ViroCyt Virus Counter® was designed to meet this objective.
From a regulatory standpoint, vaccine stability must be demonstrated, along with the prediction of stability during temperature excursions, before a vaccine can be approved for use in humans.
In this work, Abdala subunit vaccine thermostability was studied under thermal stress conditions (2–8°C [control], 25°C, 37°C, 45°C, and 60°C) for 15 days. Molecular integrity of the vaccine active pharmaceutical ingredient was monitored by SDS-PAGE, immunoblotting, RP-HPLC, mass spectrometry, and circular dichroism spectroscopy analysis. While functionality was monitored by immunogenicity assay, inhibition of binding between receptor-binding domain (RBD) and receptor, angiotensin converting enzyme 2 (ACE2), and RBD/ACE2 binding assay.
Results showed that no degradation, loss of disulfide bridges, nor modifications of secondary structure of the RBD molecule were detected at 25°C and 37°C. Moreover, high titers (1:48,853-1:427,849) of anti-RBD-specific mouse antibodies were detected with the ability to inhibit, to different degrees, the binding between RBD/ACE2.
In conclusion, the Abdala subunit vaccine is stable under thermal stress and storage conditions, which has an advantage over non-subunit vaccines previously approved or currently in development against COVID-19. The demonstrated high stability of this vaccine is a key factor in ensuring vaccine effectiveness, extending immunization coverage with fewer production runs, simplifying immunization logistics, and reducing cold chain-associated costs.
In the past 20 years, mammalian cell lines have been utilized to produce many viral veterinary vaccines. Cell lines such as baby hamster kidney (BHK)-21, Vero, and Madin Darby canine kidney (MDCK) are widely used because they help facilitate shorter manufacturing lead times and tighter process controls. As compared to other biotech products, viral vaccine manufacturing processes present some specific constraints linked to the cell substrates used. With the global veterinary vaccine market value predicted to be almost $7 billion per year by 2021[2], to remain competitively priced as well as profitable, bioprocess scientists are under pressure to develop methods for faster and more cost-efficient cell culture production. This has led to a shift from the use of expensive, two-dimensional T-flask and roller bottles to single-use, stirred tank bioreactors with microcarriers, or the adaptation of attachment-dependent cell lines such as BHK-21 for suspension culture. This requires time-consuming optimization and scale-up development experiments, which are real drawbacks. However, utilizing automated, single-use mini bioreactors as a scale-down model can enable more efficient use of time and optimization of media, feed, and culture conditions to de-risk upstream process development. In this article, single-use, mini bioreactors are evaluated to determine if they are geometrically comparable to benchtop bioreactors (both glass and single-use vessels) and pilot-scale, single-use bioreactors for effectively modelling mammalian cell culture at 2 L and 50 L scale…
Medicago manufactures influenza vaccine virus-like particles (VLPs) in an unusual production platform consisting of Nicotiana benthamiana plants. During the in vitro adventitious agent test (AAT) of certain Medicago B strain influenza vaccine VLP test samples, positive hemagglutination of guinea pig red blood cells was observed on day 14, but not on day 28. The positive result in the assay was surprising because the production process uses no animal-derived raw materials and contains a viral inactivation step. Plant-associated viruses would not be expected to infect the mammalian cell-based assay. No cytopathic effects or hemadsorption of red blood cells was observed in these AATs. The positive hemagglutination was observed at 2–8°C, but not at 36–38 °C, and only in a few of the six detector cell lines used in the assay. Because this is quite an unusual pattern of responses for an AAT, Medicago and the contract testing lab, Eurofins Lancaster Laboratories (ELLI) investigated the positive responses thoroughly for the presence of an adventitious agent or an alternative explanation not involving a viral contaminant. Investigation results indicated that the hemagglutinating activity associated with the vaccine test sample itself was responsible for the positive hemagglutination response. The positive hemagglutination on day 14 of these AATs was deemed an assay artifact, and preventive actions were taken to prevent recurrence of this type of false positive response…