The baculovirus expression vector system, which is based on infecting insect cells with recombinant Autographa californica nuclear polyhedrosis virus (AcNPV), is one of the most commonly used eukaryotic expression systems aimed at producing functionally active mammalian proteins. It offers advantages such as high-level protein expression and post-translational processing capabilities that are extremely important to the biological activity of certain proteins. This system utilizes a strong promoter of the very late gene, polyhedrin, to drive heterologous protein overexpression. Nevertheless, in order to generate milligram amounts of recombinant proteins, cell culture often needs to be scaled up to as much as 25 liters….
Tag: <span>protein expression</span>
Proteins and their promise for revolutionizing drug discovery have come virtually full circle in just a few decades. The advent of genetic engineering and the emergence of early recombinant proteins such as insulin and interferon dramatically boosted the perceived value of proteins in pharmaceutical research and of protein drugs in particular. Although the lights dimmed somewhat on the promise of therapeutic proteins in subsequent years, more recent times have seen a resurgence of interest in proteins, particularly monoclonal antibodies. Perhaps most telling has been the dawn of the post-genomic era, which has cast a bright spotlight on proteins, long respected as the work-horses of the cell, for their usefulness in exploring cell function, unraveling biochemical pathways, understanding disease, and for their massive value as novel drug targets…
The use of plants as protein expression hosts for human therapeutic proteins is emerging as a safe and cost-effective alternative to microbial and mammalian cell culture. Pharmaceutical protein production is typically carried out in microbes and mammalian cell culture because of their high production potential and/or ability to produce complex eukaryotic proteins. However, immense costs are typically required for production facilities to support their growth. To offset these costs, companies usually build and expand a production facility over several years. In fact, it has been predicted that the demand for high-value pharmaceuticals produced by cell culture will quickly surpass the ability of pharmaceutical companies to produce them…
The first use of mammalian cell culture for the production of vaccines dates back to polio vaccine development in the 1950s. The development of hybridoma technology in the 1970s further stimulated the use of mammalian cells for the production of monoclonal antibodies. Together with developments in genetic engineering, it therefore became possible to produce a wide range of recombinant proteins as well as to improve cell metabolism. Animal cells are now widely used in industrial processes to obtain complex glycoproteins with correct posttranslational modifications and biological activity for diagnostic and therapeutic applications. Animal cells are the main source for commercially available recombinant proteins such as tissue plasminogen activator (tPa), erythropoietin (EPO), DNAse, factor VIII, interferon-ß, and monoclonal antibodies…
At the onset of modern-day biotechnology, products typically fell into two distinct categories, the traditional high volume, low value products (e.g. beer and industrial enzymes) that had come to characterize the biotechnology industry, and low volume, high cost products. Recombinant proteins, the result of technological advances in molecular biology, have come to typify these latter products. Recombinant protein therapeutics have been hugely successful, potentially outstripping production capacity and continue to drive much of the biotechnology. Meanwhile, many recombinant proteins, those characterized as research tools and reagents, are governed by a price-volume relationship typical of industrial enzymes. In a competitive environment, they are fast becoming commodities — price sensitive, packaged as kits, coupled to instrumentation, and relying on heavy marketing and brand recognition. Ominously, the advantage protein therapeutics have enjoyed with patent protection and regulatory constraints on production is being threatened as patents expire and competition from generics increases…
More than 130 drug and vaccine approvals for 95 entities over the last 20 years have generated roughly $30 billion in revenue for the biotech industry. The vast majority of this revenue comes from 30 proteins that have manufacturing bottlenecks resulting from the complexities of consistent protein production. The lag times involved in constructing mammalian cell fermentation facilities keeps supply of immensely successful high-volume drugs like Enbrel, Rituxan, and Remicade well below estimated demand. In other cases, the complexities of peptide synthesis threaten the potential of soon-to-be-launched or recently approved drugs like Fuzeon. The Pharmaceutical Research and Manufacturers of America (PhRMA) has documented more than 371 new biotech drugs in development, supporting the view that demand for many biopharmaceuticals will continue to outstrip supply. That number does not include the multitude of biotech drugs still in research stages…
The Sf-9 insect cell/baculovirus expression system is one of the most commonly used protein expression systems. It is the preferred system for generating large amounts of protein in a short period of time, and it has been successfully used to express several hundreds of different proteins. A representative list of the different proteins made in our laboratory over the past decade with the Sf-9 insect cell/BEVS system is given in Table 1. These proteins are often used in drug screening studies and structure function analysis. Proteins intended for therapeutic purposes are not normally produced using this technology, although a few examples do exist. There is also an unexplored potential for the cells to be used for the production of recombinant viral vectors. Recent reports demonstrating the ability of baculoviruses to express proteins in mammalian cells, with mammalian promoters, indicate that BEVS technology might soon have a major role to play in the field of gene delivery…