Apply metabolic engineering to mammalian and insect cell hosts in order to increase yields and enhance product quality of biotherapeutics generated by these organisms

	Manipulate metabolic pathways of mammalian and insect cell hosts in order to alter glycosylation processing
	Inhibit the programmed cell death pathways of mammalian cells to increase viable cells in culture

	Insect cells can produce mammalian-like glycosylation patterns for the first time
	Glycoprotein therapeutics with increased immunoactivities are generated
	Viable cell densities and product yields are increased for mammalian cells in culture

	Potential to produce therapeutics for human use in insect cells and other eukaryotes
	Lower doses of monoclonal antibodies required for treating cancer and other disorder
	Lower pharmaceutical costs for health care patients receiving biotherapeutics as drugs

Abstract:

The majority of human biotherapeutics are secreted glycoproteins produced by mammalian cell culture. These glycoprotein products include monoclonal antibodies such as Rituxan, Hercepten, and Remicade, erythropoietin (EPO), tissue plasminogen activator (tPA), Factor VIII, Protein C, and many other proteins in clinical trials.  The market for monoclonal antibodies alone is estimated to grow 30% a year and reach sales of nearly $6.5 billion by 2004.  As glycoproteins, these biotherapeutics typically include oligosaccharide (carbohydrate) chains attached to the protein at specific amino acid residues.  The number, type, and location of the carbohydrate attachments on the protein can affect key properties of commercial biopharmaceuticals including clearance rate, immunogenicity, biological specific activity, solubility and stability against proteolysis.  Humans will typically accept only those biotherapeutics that have particular types of carbohydrate attachments and will often  reject glycoproteins that include non-mammalian oligosaccharide attachments.  As a result, mammalian cells Chinese Hamster Ovary (CHO), Baby Hamster Kidney (BHK), and Human Embryonic Kidney-293 (HEK-293) are used for the production of the vast majority of these glycoprotein therapeutics because of their capacity to generate glycoforms and perform other post-translational processing patterns that are accepted by human patients.  Unfortunately, production of biotherapeutics in mammalian cells can be expensive due to the need to grow these cells in costly cell culture environments and because mammalian cells often produce the proteins in low yields. Currently, there is a desire by biotechnology and pharmaceutical companies, researchers, consumers, and the government to lower the costs for producing high-value biopharmaceuticals.

Metabolic engineering is playing a central role in the development of more efficient methodologies for producing high-value biopharmaceuticals in mammalian, insect cells, and other eukaryotes.  Two different metabolic engineering strategies are being implemented to improve production of high quality biotherapeutics.  One strategy to increase the efficiency of producing expensive biotherapeutics is to enhance the capabilities and capacity of the mammalian cell hosts through metabolic engineering. Metabolic engineering of mammalian metabolism, cell cycle, apoptosis, and glycosylation can increase cell densities and yields and improve the quality of products from cell culture. Two specific examples of mammalian cell metabolic engineering will be presented:

1. Elimination of a glycosylation processing pathway to improve antibody product quality

2. Inhibition of cell death (apoptosis) pathways to increase monoclonal antibody product yields Glycosylation pathways and other post-translational processes can also be manipulated to obtain higher quality products from these mammalian cell hosts.

An alternative metabolic engineering strategy is to alter the physiology of non-mammalian hosts in order to allow these organisms to produce biotherapeutics accepted by humans. These alternative hosts currently lack the genetic and metabolic content for generating high-quality mammalian forms. However, these cells can often generate higher product yields than mammalian cells so metabolic pathways are being engineered so that these organisms can generate the more desirable higher value glycoproteins. The engineering of insect cells metabolic pathways to facilitate the production of mammalian-like glycoproteins is one example that will be presented. Increasing biotherapeutic production efficiencies in mammalian, insect cells, and other eucaryotes will lower the costs to develop valuable bio pharmaceuticals and ultimately lower health care costs for patients receiving these life-saving drugs.

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